RESEARCH

IN PHYSIOPATHOLOGY
AS BASIS OF
GUIDED CHEMOTHERAPHY
With Special Application to Cancer
EMANUEL REVICI, M.D.
Scientific Director, Institute of Applied Biology,
New York, N.Y.
Chief of Dept. Of Oncology, Trafalgar Hospital,
New York, N.Y.

Published for the American Foundation for Cancer Research, Inc., by

D . V A N N O S T R A N D C O M P A N Y, I N C .
TORONTO

PRINCETON, NEW JERSEY

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COPYRIGHT 1961, BY
EMANUEL REVICI. M.D.
Published simultaneously in Canada by
D. VAN NOSTRAND COMPANY (Canada), LTD.

CONTENTS

FOREWORD

THEORY A ND FAC TS
EW OTHER PATHOLOGICAL CONDITIONS have aroused, as cancer has, the interest of so

many scientic disciplines. Problems related to cancer have become of continuously

increasing concern in virtually every eld of medicine. In some, such as pathology, they
are a major preoccupation. But in sciences other than medicine, cancer also has been
receiving increased attention. One of the most urgent activities of synthetic chemistry
today is the search for new compounds which might possibly be effective in the control of
cancer. Physical chemistry is trying to provide new explanations about the variety of
processes present in cancer. Even mathematical studies which recently have offered an
interesting application of quantum theory to carcinogenesis, have found new applications
in cancer.
With the rapid development of physical sciences, the medical research worker has
hoped that from them might come some contribution that could help him ultimately in his
difficult task. He also appears to have been anxious to take quick advantage of the
progress of other disciplines for another reason, hoping that, through employing their
ndings and methodology, medicine in general and cancer research in particular, could be
promptly changed from the empirical discipline it has been until now into a positive
science. He has brought as many applications of other disciplines as possible into his study
and this has led to a whole series of new methods of investigation through which
interesting new information has been obtained. Yet, most of these applications have been
tried chiey because they have been at the immediate disposal of the scientist rather than
because they have represented a missing link in the development of his own ideas.
The outcome has not been rewarding. Medical knowledge appears not to be
sufficiently advanced to successfully utilize the avalanche of new, highly specialized
information offered by the investigative methods derived from other disciplines. Basic
theoretical knowledge in medicine in general, and about cancer in particular, has not yet
reached the level necessary to relate and assimilate the new data. To a large extent, basic
concepts about pathogenic problems are not even formulated as yet. When the medical
scientist has tried to transform the new data into effective therapeutic procedures, he has
failed. And the failure has made more evident how much we need basic
physiopathological knowledge before we are able to take advantage of detailed data.

Meanwhile, normal development of cancer research has been hindered, sidetracked from its logical course. While thousands of scientists with almost unlimited funds
at their disposal are presently using the most advanced methods for the acquisition of
details, almost no attempts are being made to resolve basic problems, although the cancer
investigator is continuously obliged to realize the dearth of fundamental knowledge.
If we attempt to analyze this abnormal situation further, we can nd indications
that it may have its origin also in a distortion of the proper relationship between the two
factors that, together, make for progress in researchideas and experiment.
The experimental approach provides precise information about particular
phenomena under dened conditions. The analytical method tries to investigate reality by
recognizing the proper place of the various constituents of a whole, the parts being
identied as such by the experimental ndings. On the other hand, the conceptual
method not only provides an inkling of what the completed whole will eventually look like,
but also attempts to predict the properties and the relationship of the component parts.
In dealing with a highly rened and complicated subject, the analytical method by
itself appears inadequate. For example, in atomic physics, the results of experiments are
expressed by numbers giving the values of certain physical quantities that have been
measured. In order to complete the analysis, we must simultaneously determine the
numerical values of certain quantities dening the material bodies, the objects of the
experiments. This is prohibitive so far as canonical coordinates by Heisenbergs uncertainty principles are concerned. With experimental knowledge somewhat curtailed, theory
at present must attempt explanation.
In other areas as well, experiments present only limited numerical values pertaining
to some physical quantities. Were we able to measure all quantities, we could analytically
reconstruct the entire theme of the physical reality. However, when some quantities
cannot be simultaneously determined, this direct reconstruction is not possible and
experiments merely give an indirect approach to what we regard as reality.
If the inadequacy of the analytical approach by itself is evident in the highly positive
disciplines, such as in the physical sciences, it is even more so in biology. As Bohr and
others have intimated, the conditions of uncertainty seem to be much more pronounced
in biology than in physical science. The fact that experiments in biology give only
fragmentary and unrelated results is not surprising; the need for a synthetic theoretical
method in this eld is clear.
In medicine, which is applied biology, the need for the conceptual approach is
especially profound. lt is true that this approach, as the sole approach, has shown its
inherent weakness in the past. There was a time in the development of medicine when
available data were so scarce and unreliable, and the need for ideas to provide some sort
of guidance was so great, that the worker resorted to broad imagination, using it to
replace almost entirely any other form of investigation.
Largely as a reaction to the high proportion of speculations prevalent in the early

years, the experimental approach in medicine came to be emphasized. Claude Bernard,

who almost single-handedly was responsible for this, tried to give experimentation its
rightful role. However, in ensuing decades, the relationship between theory and
experimentation has been progressively distorted. An unrestrained exaggeration of the
role of the experiment, the erroneous view that pure facts represent the aim of research,
has led to an entirely unbalanced approach. Not only have almost any data obtained by
research been considered intrinsically interesting, but obtaining them has become the sole
purpose of much research. In scientic papers today, experimental data must be reported
as such; any allusion to theoretical meaning is considered undesirable. Generations of
scientists have been schooled to believe in the intrinsic value of the experiment. As they
have applied this belief to research in biology and as they have made unlimited use of new
methods taken from other disciplines with no ideological requirement for their use, we
have had more and more data and fewer ideas. Today, with great astonishment, some
scientists are at last beginning to recognize not only that data alone do not generate ideas,
but that science cannot progress without theory.
Ideas and experiments are integral parts of all scientic research. A balance
between them is needed to assure progress. It must be understood that the function of
experimentation is to guide our thinking, to help build up new concepts, and to prove
their accuracy in accordance with reality. Certainly, fundamental concepts must not be
mere speculations. They should be accepted only after conrmation through
experimentation. Experimentation is the necessary link between mental concept and
reality. To the attempts to consider any unresolved fundamental problems in biology, one
has to try to bring a rightful balance between conceptual views and experimentation.
The exaggerated importance attributed to experimentation in biological science, its
use even as a substitute for ideas, has led recently to a massive attempt to solve the
therapeutic problem of cancer by indiscriminate screening of chemical agents. Here,
empiricism has been brought to its culmination. After tests of tens of thousands of agents,
many workers are now beginning to realize that the results are almost worthless for
cancer therapy in humans, that seemingly promising agents have an effectiveness limited
to the conditions present in the actual animal experiments. By its impressive magnitude,
the failure of indiscriminate screening, of empiricism epitomized, has begun to impel
many workers to change their ideaas to what must be done if the cancer problem is to be
solved. A rst result of this change has been a new and, this time, unbiased evaluation of
just where we stand in our assault on the cancer problem. Every day more scientists are
making the evaluation in their reports to the medical profession and to the public with a
candidness which, only a few years ago, very few would have employed.

The Present State of the Cancer Problem

Surgery in cancer can be considered to have arrived now at or near its maximum efficiency.
Thanks to progress in operative techniques, and to advances in pre- and post-operative
care, ultraradical surgery is available today. The propensity of cancer to spread far from its
original site has made such surgery obligatory in many cases if there is to be an effort to
eliminate all malignant cells. Yet ultraradical surgery has not suiciently increased the cure
rate to justify horrifying mutilations, especially when the face is involved. With few
exceptions, surgical procedures do not prevent the patient from dying of cancer sooner or
later. The so-called ve-year-cure-rate represents, to say the least, an unrealistic appraisal.
Many authors consider that even the rate of ve-year survival is not improved by surgical
procedures, and the ultimate fate of these ve-year survivors, with few exceptions, is still
disastrous. Most of the cured cases still die from cancer.
Other recently discovered facts have increased skepticism about the value of
surgery in cancer. The polycentric origin of cancer, especially incases where the lesions are
far apartconsidered by some workers to be true even in malignant melanoma, for
instancewould greatly limit the value of surgery as a means of eliminating all cancerous
cells. It is recognized that to operate on a lymphoma is useless. Furthermore, it is known
today that cancer cells are present in the circulating blood. Surgical manipulation has been
found to induce a ow of these cells into the blood even from relatively small primary
tumors.
In view of all this, cancer cannot be considered to be a condition for which surgery
is a major hope. Surgery represents only an expedientto be tried so long as nothing
better can be offered. It is probable that in the future it will be reserved, in cancer
treatment, for the correction of mechanical complications, such as intestinal or other duct
occlusion.
Unfortunately, radiation has not been much more successful in its long range
results. In order to control cancer, it is necessary that radiation destroy all the cancer cells
present in the organism while producing minimal damage to normal tissue. It appears that
such high selectivity of action cannot be obtained. The lack of it may be implicit in the
nature of the effects achieved by radiation. A study of the biological effects of radiation,
which is to be presented later in this monograph, has shown that an important part of the
action of radiation is to induce changes in certain constituents of the body, principally fatty
acids. These changes are largely responsible for the favorable effects of radiation but they
also are largely responsible for the undesired effects. It is the nature of these changes
which limits qualitatively the capacity of radiation to inuence cancerous processes, and
makes it dubious that progress in technique can ever greatly improve the qualitatively
insuficient effectiveness of radiation. Clinical results to date provide conrmation of this
pessimistic view. The recent use of extremely high voltage radiation, of radioactive cobalt,
and of other radioactive particles has not greatly improved results over those obtained
with older forms of radiation twenty years ago, except for reducing some harmful

immediate skin and systemic effects. Now, as earlier, with few exceptions, the benets of
radiation are no more than temporary. Longlasting good effects still are limited to only a
few radio-sensitive tumors. The resort to isotopes, in which the scientic world has put so
much hope and millions of dollars, also has proved greatly disappointing.
Of the thousands of cases of various kinds of cancer in which isotope therapy has
been tried, only a very limited number of cancers of the thyroid have responded. Not only
because of its continuing failures, but because of its inherent qualitative inadequacy,
radiation does not appear, any more than surgery, to represent the solution for the
problem of cancer.
With surgery and radiation therapy incapable of resolving the problem, more and
more research workers have turned their efforts in other directions. The existence of some
cases of spontaneous remission has led many investigators to believe that immunological
procedures related to cancer would be able to resolve these problems. Unfortunately the
existing knowledge in this specic eld is too meager to permit more than some tentative
investigations, usually only repetitions of similar researches made many years ago with
limited success. Fruitful development of this approach would have to follow the normal
pathway, starting with more knowledge of the complex immunological processes
intervening in cancer.
An enormous amount of cancer research in recent years has been directed toward
chemotherapy. It is a fact that many agents and groups of agents have shown the capacity
to inuence tumor evolution. However, each has had limited usefulness. Results of
treatment have been characterized by inconsistency. Even in seemingly susceptible types
of cancers, results have been good in one case, poor in another and have varied even for
the same patient at different times. The inability to explain and remedy these variations
has discouraged many workers. Although it appears evident that the source of
discrepancies resides in the patients themselves, the general tendency among researchers
has been to try to resolve the problemby nding agents able to act independently of any
differences which exist between subjects.
In despair at the lack of progress in this approach, many workers today are using
the screening enterprise mentioned above as a kind of last resort. For this project, they
have renounced the scientic concept that pharmaco-dynamic activity must serve as the
basis on which an agent is to be tried in therapy. They have fastened into a purely empiric
approach. Now, all available chemical substancesand many others which will be
synthesized especially for the purposeare to be screened indiscriminately, for their
effects on animal tumors with no reason for this test other than that the agents are, or can
be made, available. We will not dwell here on the assumption that routine technique is
more likely than imaginative brainpower to resolve the problem of cancer. The results of
this screening to date have shown it to be an invalid procedure, as expected by most
critical workers. With tens of thousands of substances already tested, the busy screeners
are obliged to recognize that the approach itself is fundamentally erroneous. Experience
has proved that an agent can be wonderfully effective against one tumor and still be en-

tirely inactive in others. Of tens ofthousands of agents tested, less than a hundred have
shown effects on tumors in animals. None appears to have signicant value when applied
in humans. These results have emphasized again the importance of factors other than the
agent itself. One factor lies in the differences which exist between various tumors. Some of
the other factors include variations between species, between individuals of the same
species, between origins of tumors, between spontaneous and transplanted tumors, and
even variations in anyone individual at different times.
Faced with this situation, some workers have concluded that not one treatment but
at least hundreds of different treatments must be found in order to cope with the huge
variety of conditions.
Taking cognizance of these considerations, it has seemed to us that a more realistic
and logical approach is to try to understand the nature ofthe existing differences and to
attempt to make the treatment adequate on the basis of that understanding. It has been
this approach which has been followed in our research.
We have studied the problem of cancer for the last thirty years from an entirely
different vantage point than that used by other workers. Attention has been focused on
the physiopathological aspect of cancer, on the basic changes that occur in the different
patients, with the ultimate aim of understanding the part played by these changes in the
response of cancer to therapeutic attempts. This emphasis on the physiopathological
aspect of cancer has been made possible by applying a more general overall idea of the
nature of the disease.
This approach is based under various new concepts. They concern,
1) The role of the organization in the pathogenesis of the conditions.
2) A dualistic systematization of the manifestations related to normal and
abnormal physiology.
3) The predominant intervention of certain constituents such as lipoids and
chemical elements in the induction of the opposite manifestations.
4) The possibility to integrate the occurring processes into a system of defense
mechanism against the noxious inuence exerted by the environment.
Many general and special problems of physiopathology, some of them concerning cancer
and other conditions, have been analyzed in this framework.
The application of this approach to therapy has resulted from a logical
development of that approach. The recognition of the intervention of a variety of
pathogenic factors, not only differing from one subject to the other, but even changing in
the same subject during the evolution of the condition has emphasized the need for
individualized therapy. As opposed to the tendency to overcome the differences existing
between individual subjects through a standard therapy, the guided therapy utilizes the
knowledge of the occurring different pathogenic particularities in order to correct them. A
high degree of exibility in the treatment has appeared necessary.
As part of this approach to therapy, has appeared the need for more complete
knowledge of the existing differences and their interpretation in terms of the pathogen-

esis of the condition. The search for adequate analytical tests has thus represented the
rst task. The development of day-by-day analysis of the condition has been possible by
choosing relatively simple but reliable procedures. The information they offered was used
to determinethe nature of the agents able to correct with a certain specicity, the
encountered pathological conditions. These two parts the recognition of the existing
condition and the adequate agents, have concretized this approach.
These considerations explain also why the new developed guided therapy cannot
be understood and correctly applied without a sufficient knowledge of its
physiopathological and pharmacological basis. These same considerations have led us to
present the research concerning this approachas a block, instead of fragmented
communications. The form of a monograph has appeared consequently the best suited. In
a further effort to achieve a cohesive presentation, we have separated from the text most
of the technical and experimental data, and presented them as notes at the end of the
text.

ACKNOWLEDGEMENT
Progress in our research has been made possible only through the day by day
cooperation of different groups of co-workers who have contributed years of assiduous
work. Many of them are mentioned in the following pages where the research in which
they took part is presented. I am deeply indebted to them.
I wish to thank all those friends whose personal efforts or, who through their
organizations, have given their material and moral support to the continuation of our
research. Special thanks go to Mrs. Sherman Pratt for her tireless efforts on our behalf.
My sincere appreciation to those who have helped me in the preparation of this
book, Mr. L. Galton, Mrs. E. F. Taskier and Mrs. H. Kennedy for the editorial work; Mrs. E. F.
Taskier for the gures; and Mrs. P. Berger, Mrs. B. Doctors and Mrs. M. Prikasky for the
secretarial work.
I wish to gratefully acknowledge Miss Fanny I-Ioltzmanns devoted friendship and
help.

CHAPTER

t s A S I CC
] O N C E P T SO F
ORCANIZATION
nr\
rNrELLEcruALMEcHANTsM
I
usedby man to acquireknowredgehas
"t
led him to recognizethe existenceo[ relationshipsbetween the various
manifestationsencounteredin nature. He has employed abstractionand
integration to build up conceprualindividualitieswhich he identified as
separateentitiesin nature. Structuralcharacteristicsand dynamic properties have appearedto be the most suitablecriteria for defining theseentities. However,curiosityhas constantlyimpelledman to attemptto extend
his knowledgeby explainingand correlatingtheseentities,and an important means has been analysesbreaking them down into their component
parts.
Out of these analyseshas come recognitionof the fundamental importanceof organization.F^ot,as entitieshave been analyzedone after the
other, it has becomeclear that the seeminglyinfinite variety of them perceived by our sensesis in reality the result of the arrangementof a relatively small number of basic units, the molecules.Moreover, analysishas
shown that only a very small numberof chemicalelementsmake up even
the most complexmolecules;that combinationsof less than one hundred
elements,in differentproportionsand relationships,
account for tens of
thousandsof compoundsand many billionsof entities.And further analysis
hasrevealedthat elementsthemselvesrepresentdifferentdynamic arrangements of only n fsw-nscording to some hypotheses,only two-fundamental corpuscles.
Upon close analysis,nature, which appearsto be so greatly varied,
turnsout, in fact, to be basedupon only a very few fundamentalconstituents and it is the manner in which theseconstituentsare bound together,
which providesvariety.
theirorganizationinto a multitudeof combinations,
I

R E S E A R c Hr N p H y s t o p A T H o L o c y

The studyof organizationobviously,then,could furnish the most valuable information about nature.And it would not appearto be too much to
expect that, if nature's seerninglyinfinite variety stems from organization
of only a few constituents,then organizationitself might also be achieved
through a few, relatively simple fundamentalpatterns.If so, seekingout
suchpatterns-systematicanalysisof organizationcomparableto the efforts
to systematizeconstituents----could
be of primary importance to better
understandingof a host of problems.
Homotropy and Heterotropy in Nature
As man attemptedto recognizeordcr in the constantchangessecn in
nature,he noted certain patternsthat appearedto indicatedefinite"laws of
nature."Someof theselaws have been observedto operateunder such a
wide variety of circumstances
that they have come to be acceptedas
"fundamentalIaws."
ln 1824, Sadi Carnot formulatedone which is known as the Second
Law of Thermodynamics.Carnot observedthat, in a given system,work
involving the transfornrationof thermal into mechanicalenergy is only
accomplishedas heat drops from high to low temperature.In more general
terms,this meansthat work accomplished
in an isolatedsystemresultsin
progressively
eliminatingdifferencesin temperature.Clausiusrecognized
this as a fundamentalprinciple and postulatedthat the amount of energy
availablefor work alwaystendstoward a maximum. This condition, called
"maximum entropy," correspondsto uniformization of temperatureand
also to homogeneousdisorganization.At first, it appearedthat the principle was in conflict with the First Law of Thermodynamicswhich expresses
the rule of conservationof energy.However, Helmholtz soon was able to
demonstrateits validity by showingthat only the secondlaw could reconcile
the first with the impossibilityof perpetualmotion.
In a more philosophicalvein, we considered,in our research,that this
Second Law of Thermodynamicsin its broadest sense could define a
fundamentaltrend toward annihilation of any existing differencesin nature, through the triumph of total uniformity. SinceClausiusused the term
"entropy" in applying Carnot's original observationto closed mechanical
systems,it has seemedpreferableto avoid confusion by utilizing another
term for this general tendency toward uniformity in its broadest sense.
Therefore, we have chosen the term "homotropy."
Despitethe theoreticallyrapid trend in the directionof absoluteuniformity, or homotropy, no such final state has yet been achieved.It must
be concluded,therefore,that someotherfactoropposedto that trend exists.

BASIC CONCEPTS OF ORcANTZATION

Wehavechosenthe term "heterotropy" for this other factor, which tends

to maintain
or produceinequalityand thus to prcservethe order that is
evident
in nature.
In order to understandthe roles of these two opposing fundamental
tendencies
in the organizationof nature in as logical a fashion and with
asmucheaseas possible,it seemedadvisableto try to study their operationfustin one of the simplestand best known natural organizations,the
atom,passinglater on to higher and lower levels of organization.
TheAtom
Therole of the two opposingtendenciesin atom organizationbecomes
clearwhen we study the relationshipbetween the forces that form this
entiry.
Each atom consistsof a positivelycbargednucleussurroundedby
negatively
charged electronsin adequatenumber to balance the nuclear
charge.
Thee.ristence
of the atom dependsupon forcesacting betweennucleus
andelectrons.One group is of coulombiannature. These are the electrostaticforcesthat account for the attraction between oppositely charged
electrons
and nuclei, and for the repulsionbetweenelectronsbearing similarcharges.
If such forcesdid not exist, electronswould wander irregularly
andwouldnot be retained around the nucleus.
Yet, if electrostaticforces were unopposed,electronswould be drawn
closer
andcloser to the nucleusand would finally fall into it, therebybringtngaboutcompleteannihilationof all charges.The fact that electronsare
notabsorbedby the nucleusindicatesthe existenceof a second,opposing
force.
This secondforce is definedby quantum mechanicsand the quantum
rheoryof fields. euantum mechanicsascribesa seriesof discreteenergy
levelsto electrons within atoms. Radiation is emitted or absorbed only
whenelectronspassfrom one stationarylevel to another.The energylevels
ue relativelystable, and a state of minimum energyexists when each of
theelectronsof the atom is as close as it can be to the nucleus on the
groundlevel.
The energy levels correspond to the orbits describedby Bohr's theory
which,although not entirely accurate,affordsa good basisfor understandingatomic properties.Bohr envisagedthe electronsrevolving around the
nucleus
in definite orbits, each orbit moving continuouslyin these states,
theatomnot emitting radiation. This differsfrom thc older theory according to which the electronscan revolve around the nucleus on any orbit.
Suchcasualorbit motions would lead to loss of energy by radiation. The

R E S E A R c Hr N p H y s r o p A T H o L o c y

electrons would come closer and closer to the nucleus and would, as
already pointed out, finally be absorbedby it. The quantum theory of
fields accountsfor the absenceof radiation and for electronsremainingin
their particular orbits. However, the concept of stationary statesfails to
explainall the propertiesof the atom, particularlyits chemicalreactivity,by
virtue of which different atoms combine to form molecules.
According to anothertenetof the quantum theory, the Pauli Exclusion
Principle,an orbit cannot be occupiedby an indefinitenumber of electrons
but, at most, by two electronsthat spin in oppositedirections.The orbits
are arrangedin shells,each shell having a definite level of energy.A shell
is completewhen it containsthe maximum number of electronscompatible
with the Pauli Principle.Completeshellsconsistof.2, 8, 18, etc., electrons.
When an inner shell has its quota of elcctrons,additional electronsmust
occupy an outer shell. Consequently,
insteadof falling into the nucleus,
the electronsin their lowest energy stateswill continue to revolve at it
considerabledistancefrom the nucleus.
As already indicated,if there were only electrostaticforces, the electronswould have long sincefalleninto their nuclei,neutralizingall electric
charges.The universewould be in a state of maximum homotropy.No
strong atomic forces would exist and no chemical reactions would take
place.The intervcntionof quantumforcesavoidsthis. It is apparent,then,
that the organizationof the atom resultsfrom the operation of two types
of forces,electrostaticand quantum, the electrostaticservingto bring and
keep nucleusand clectronstogetherto constitutethe atom, the quantum
accountingfor a motion of electronswhich preventstheir total annihilation and the neutralizationof all electricalcharges.
Homotropic and Heterotropic Forces in the Atom
We rnay now attempt to considerclectrostaticand quantum forces in
the atom in terms of homotropicand heterotropictrends.Let us hypothesize
an atomic systemin which only electrostaticforces are active and compare
it with a real systemwhich also has active quantum forces.Whereasthe
fictitioussystemwill rapidlyevolvetowardsa stateof maximum homotropy,
with annihilation of all charges,this will not occur in the real system.
When the two systemshave reachedfinal statesof equilibrium, the homotropy of the imaginary systemwill be greaterthan that of the real system.
If the quantum forces that keep the electronsaway from the nucleus in
the real atom could be withdrawn, the electrostaticforces acting alone
would bring about a state of completeannihilation,thus making available
a certain amount of energy that had previously been preservedby the

B^sIc coNcEprs

oF oRcANrzATIoN

quantrrm forces. In this sense,it is apparent that the electrostaticattraction

betweennuclei and electronsis of a homotropiccharacter,while quantum
forces are heterotropic.
Fulfillment. ol Quantum and Electrostatic Forces
There are diverse coDsequencesfrom the operation of quantum and
electrostaticforces in the atom. The partial fulfillment of the electrostatic
forces keepsthe nucleus and electronstogetherin the atom, while quantum
forces ceaseto exist with the establishmentof complete electron shells,and
have, therefore,been called "saturation forces."
In the atoms of the noble gases,quantum and electrostaticforces are
simultaneouslyfulfilled. As a result, theseatoms are inert. They have no
physical or chemical activity and their entry into the formation of molecules
is explained by the intervention of van der waal's cohesion forces. In all
other atoms, the electrostatic forces are fulfilled when the number of
orbital electronscorrespondsto the nuclear charge. However, when this
occurs, the electron shells are incomplete and consequently unfulfilled
quantum forces :ue present. when the quantum forces are fulfilled, other
electrostaticforces appear.
under these circumstances,in order to complete its outer electron
shell,i.e., to fulfill the quantum forces,an atom may borrow or lose one or
more electrons.This is achievedwith a secondatom which, by the exchange, reduces or increasesits orbital electrons to fulfrll its quantum
forces and is left with a number of electronsconsistentwith a complete
outer shell. The fulfillment of quantum forces requires changesthat involve a displacementof electronsoutside the atom itself. This exchange
of electrons,properly called "electron transfer," fulfi"lls,to be sure, the
quantum forces of the atom. However, as a result of the transfer, the
relationshipbetweeneach nucleusand its orbital electronsis changed,resulting in covalentions. Those atoms that have gainedby the transfer and
have an excessof electronsnow have a negativecharge while those that
have lost electronshave a positive charge.As a result, new electrostatic
forces appearwhich, althoughconfinedto the atoms themselves,influence
their external behavior, as evidencedby the interaction between atoms.
An antagonisticrelationship can be conceivedbetween electrostatic
and quantum forces in the sensethat the fulfi.llmentof one usually leads
to appearanceof the other.
Electron transfer representsonly one mechanismfor fulf,lling the quan. We have unwillingly resorted to this too anthropomorphic term, the use of
which has to be excusedas didactic license.

.'

R E s E A R c HI N p H y s t o p A T H o L o c y

tum forces of the atom. Two atoms which do not have sufrcient electrons
in their external shells to complete the external shells of both can fulfill
their quantum forces by sharing some of their electrons.By achieving a
complete external shell for each atom, the sharing processsatisfiesthe
quantum forcesof both atoms.This methodof quantum fulfil.lmentthrough
the sharing of electrons also can lead to the appearanceof electrostatic
forces.If the two atoms are identical,the sharedelectronshave an intermediate position and, therefore,do not influence them. As a result, the
atoms have their quantum forces fulfilled without inducing new electrostatic forces. This is the so-called"homopolar bond." If, however, two
atoms are dissimilar energetically,their shared electrons will be located
closerto one atom than to the other, the distancebeing determinedby the
competitiveinfluenceexertedby the atoms upon the sharedelectrons.At
the same time, other electronswill be influencedby the bond, and, as a
result,their orbits will be alteredto someextent.Weakerelectrostaticforces
will result and the bond will be intermediarybetweenthe ionic and the
homopolar.Both kinds of fulfillment of quantum forces-one achievedby
transfer,the other by sharing-thus lead to the appearanceof new electrostatic forces in the ions or ionoids.
We must repeathere for emphasisthat the fulfillment of quantum forces
can take place through variousavenues,either by loss or gain of elcctrons,
or by sharingwhich can range from ionic to homopolar. The plurality of
possibilitiesfor fulfillment of quantum forces is very important, making
it necessaryto considerthe resultsof such fulfillment on a statisticalbasis.
The electrostaticforces act betweenchargedions of oppositesigns,or
betweenatoms bound by sharedelectrons.Through the balanccof thesc
electrostaticforces,bound atoms appearand correspondto ncutral formations, having their electrostaticforces fulfilled. However, it is only with
the intcrvcntionof suitablequantum forcesthat the bound atoms can form
a new entity, the molecule.
Quantum and Electrostatic Forces in Molecules
Alternate operation of electrostaticand quantum forces leads to the
organizationof atomsinto molecules.The quantum forcesin the molecules
interveneto permit organizationof theseentitiesso that the constituents
are maintainedat proper distancesand positions.The result is electrostatic neutrality.The appearanceof new quantum forces that maintain the
constituents,through their organizedmovement,at certain distancesand in
but also the stabilityof
certainpositions,insuresnot only the establishment
the new formations.Besidesvibrationalmovements,other more definite

B A S I C C O N C E P T SO F O R C A N I Z A T I O N

movementscan be recognizedin the new molecule.When two or more

atomsbecomeassociatedby sharcdelectronbonds,the sharedelectronsno
longer are confined to one atom but are displacedfrom their own orbits.
Under certain conditions,electronscan travel betweentwo or more atoms,
or even surround the moleculeas a whole. These movementswhich correspond to the interventionof quantum forcesgive stability to the molecule.
The fulfillment of intramolecularquantum forces will aflect molecules
in a similar way as fulfillment of atomic quantum forces aflectsatoms. By
a processsimilarto that governingmotion of electronsin atoms,motion of
entitiesthat enter into the structureof moleculesis also controlled.The
fulfillment of quantum forces is achievedin various ways. For example,
there may be localizationof the movementof electronsin the molecule.As
the result of relative immobilizationof theseelectrons.electrostaticforces
appear in the molecule as a whole.
The relativelyimmobilizedelectronscan be consideredas being related
to the moleculeas an entity, since they cannot definitely be attributed to
any of the constituentatoms. As a result,the moleculebecomeselectrostatically active.
The positionsof electronsand evenof atomsin moleculescan be understood easilyby consideringeventsat the molecularlevel in the sameway
we consideredthose at the atom level. The molecule whose electrostatic
forces are balancedis neutral. However, it has active quantum forces
which governthe positionand mobility of the constituents
and the relative
positionsof bound atomsor of certainelectronsin the entity. Fulfillment
of molecularquantum forces is realizedthrough changesin movement
of electronswhich lead to loss or gain of one or more electrons,protons,
ions or even groups of atoms. This leads to appearanceof electrostatic
forcesand the moleculebecomesan activeentity. In the molecule,as in
the atom, quantumforcescan be fulfilled in more ways than one-although
one may representa preferred situation. For this reason, activation of
molecules,through changesin mobility of molecularelectrons,has to be
consideredon a statisticalbasis.
The electrostaticcoulombiancharacterof an activatedmoleculeis the
resultof the changesin mobility of the electrons.Positiveor negativeareas
in the moleculedevelopaccordingto the abundanceor dearth of electrons,
at thesepositions.The new electronicarrangementsin a moleculecan be
seen as representinga preparatory step for the molecule to become an
activeentity in the same way that atoms are activatedand becomeions.
The molecule loses or gains one or more electrons,(or protons, ions or
groupsof atoms) and becomeselectrostatically
active,with positiveor nega-

REsEARcH rN pHysropATHoLocy

tive charge,dependingupon the nature of the lost or gained entity, and this
is the outcome of the fulfi.llmentof the molecularquantum forces.This is
illustrated by the following examples concerning the benzene molecule,
and the carboxyl and hydroxonium radicals,in which we shall limit ourselvesto changesproducedby quantum and electrostaticforces.
In the benzenemolecule,which is electrostaticallyneutral, the electrostatic positive and negativeforces of the constituentatoms are balanced,
However,all the electronsare not in fixed positions.The ,,' electronsof the
doublebondsmove around in the molecule.Becausethe moleculeis closed,
this movementis circular, thus accountingfor the stability of the molccule,
recognizedin part by the equal reactivityof all its carbon atoms which is
encounteredunder certain conditions and results in the Kekulian forms.
The fulfillment of the quantum forces accountsfor a kind of relative
fixation of these wandering,,,electronswhich is responsiblefor the other
structuresof the benzenemoleculedifferent from the Kekulian ones. It is
this localizationof electronswith the capacityto entcr into further reactions
which results in the activation of the molecule as seen in the resulting
Dewar structureswhich in turn accountsfor active centerssuch as ortho,
meta, and para positions.These excited molecules,electrostaticallyactive,
can readily take part in chemicalreactions.Study of the mobile zrelectrons
in many other moleculesallows us to undcrstandtheir role in providing
molecularstability, while their relative localizationfavors the appearancc
of electrostaticallyactive centersin the molecule and resulting reactivity.
Here again, localizationof electronsopens up many possible avenuesto
activation.
The carboxyl and hydroxonium ions represent typical examples of
another kind of activation. Inactive carboxyl occurs when the quantum
forcescausethe electronsto wander continouslybetweenthe two oxygens
of carboxyl.Becauseof this electroniccondition,the H atom seemsno
longer to be bound to either of the o atoms,but is situatedbetweenboth:
this form correspondsto the electrostaticatly
fulfilled condition. With fulfillment of quantum forces,the wanderingelectrontakes a more fixed position at one or the other oxygen.when this occurs.the H+ ion leavcsthe
carboxyl group, and the carboxyl acquiresa negativeelectrostaticequilibrium, leadingto further combiningactivity.This fulfillment of the quantum
forcesis responsiblenot only for the appearanceof an activatedgroup of
electrostaticcharacter,but also for the existenceof two structures.each
one with anotheractive oxygen.
A similar activation takes place when a molecule acquiresan ion, as
seen for the hydroxoniumion. Water can, under ccrtain circumstances,

BASrC CoNCEPTS OF oRcANTZATtON

bind a proton resultingfrom a hydrogenatom which has lost its electron.

This bond is achieved through a valency bridge, and can be regarded
as the fulfillrnent of molecularquantum forces.Different structurescan be
consideredas resultingfrom the fixation of the hydrogenbridge in different
positions in rclation to the tetrahedralconstitutionof thc oxygen atom.
They help to give this bridge bond its high resistance.
Bonding ol Molecules
Electrostaticforces in radicalsor activatedmoleculesmay be further
balancedwhen new bonds are realizedbetweenentitieswith oppositeelectrostatic forces. Bonding of moleculeshaving electrostaticallyexcited centers may or may not be of chemical nature which is considered to
correspondto changesin the structureof the molecules.More often, only
a physical bond between moleculestakes place, in which case there are
no changesin molecular structure.Both types of bonding result in fulfillment of the electrostaticforcesthrougha balancedneutralization,but bonding alone is not sufficientto establisha new entity. A new entity, with
structural and functional individuality, apparently is realized only when
quantum forces appear and establishdefinite relationshipsbetween the
bondedconstituents'
molecules,
placingthemin ccrtainpositionsand organizing their movements.The holistic concept emphasizesthe difterence
betweenmoleculesor radicalsbound only by the fulfillmentof their electrostatic forces of gencral coulombiancharacter,and the new entities resulting from the appearanceof specificquantum forces proper to them. Here
again,then, at the molecularas at the atomic level,progressin organization
is achievedby alternateoperationof electrostaticand quantum forces.
Polymolecular Formations ;
When an electrostatically
activemoleculeor radicalbinds an electron,
ion or even a small radical,the resultingentity is still considereda simple
molecule.The group resultingfrom the bonding of severalpolyatomic
radicalsis a complex molecule.Like simple molecules,complex molecules
also can group togetherand the bonding of sevcralleads to still more
complexformations,the macromolecules.
In turn, macromolecules
also can
be grouped and the bonding of two or more producespolymolecularformation.Thus, organizationprogresses
from simplemoleculesto polymolecular formations,first through the grouping togetherof similar entities.A
' A l l t h r e e t e r m s - m a c r o m o l e c u l e s ,p o l y m o l e c u l e s
and complex molecules-are
cbosenonly for didactic convenience.

l0

REsEARcH tN pHysropATHoLocy

new entity appearswhen one of thescgroups binds a respectivesecondary

part.
Micelles
A distinctivetype of new entity resultsfrom the bonding of molecular
formationswith simplerconstituents,such as ions or ionized molecules.To
this type of entity we have appliedthe name of "micelle." I polymolecules,
macromolecules,complex moleculesor even simple moleculescan form
the principal part of thesemicellar entities.
According to the above definition,micellesare entities formed by the
binding of molecules,as principal units, to ionized molecutesor ions, as
secondaryunits. The latter originally were consideredto be "impurities"
until Duclay showed their important role in establishingspecificentities.
According to our concept,micellesare producedwhen grouped molecules
and activeimpuritiesbecomebondedas the resultof reciprocalbalanceof
electrostaticforcesand the alternatcoperationof clcctrostaticand quantum
forces,seenfor atoms and molecules,appliesagain. Fulfillment of electrostaticforcesleadsto appearanceof quantum forces,this time proper to the
micelles.The quantum forcesmaintainthe micelleconstituentsin proper
positions and govern their movements,describedas vibratory for these
entities.The operation of the quantum forces, togetherwith fulfillment of
the electrostaticforces, accountsfor the stability of micelles.
The micellarquantumforcesalsocan be fulfilled, leadingto the appearance of unequal distributionsof micelle constituents.The micclle thcn
passesfrom relativelyneutral to an electrostaticallyactive form which can
enterinto further bonds,and it is primarilyin furtherbondagethat micelles
appear in a reticular aspect.
In an overall view of the developmentof organization,from atoms to
micelles,the relatively simple pattern of alternatingoperation o[ electrostaticand quantum forcescan be recognized.The regularityof the pattern
allowsus to considerit as fundamentalto the progressof organization.We
havc tried to go further and to recognizethe existenceof this samesimple
organizationalsystemfor formations below the atom and above the miceUes.We tcntativelyconceivedof subatomicformationsbeingorganizedin
the samemanner,i.e., by alternateoperationof electrostatic
and quantum
forces.We will not go into this study here as it would lead us too far from
r l t i s t o t h i s t y p e o f s t r u c t u r et h a t w e a p p l y t h e t e r m m i c e l l e , a s d i s t i n g u i s h e d
f r o m v a r i o u so t h e r m e a n i n g sf o u n d i n t h e l i t e r a t u r e .

BASIC CONCEPTS OF ORCANIZATION

l l

the subjectof this presentation.An outline of this subject is presentedin

Note l.
Organization ol Motion as Heterotropic Achievement
An interestingaspectof the concept presentedabove is the influence
of homotropic and heterotropicforcesupon the motion of particlcswithin
the organizationalframework. At-random mobility must be consideredto
be an attributeof entitiesfrce of any constraint,and thus corresponding
to
a homotropic state.Any changetoward constraint,leading to a degreeof
immobilization, must be considereda heterotropiceffect. The systematized
mobility producedby quantum forces,which apparsto prevent annihilation of opposingcharges,accountsfor the relative immobilization.While
mobility iself is an homotropicattribute,its systematization
is heterotropic.
The correlation of mobility with homotropy, and of fixation with
heterotropy, appears basic. The processof uniformization which corresponds to homotropy appearsto be possibleonly in the presenceof a
maximum of free mobility. The heterotropicsystematizationof movement
can be seenat variouslevelsof organization.Electronsin movementin the
atom differ from electronsin movementin the environmentthrough a systematizationof their mobility, as they are constrainedto follow definite
patterns. The relative fixation of certain electrons-for instance,shared
electrons-following the fulfillment of quantum forcesalso marks a further
heterotropicinfluence.This also appliesto radicalssuch as the carboxyl.
In the formation of - C =
..----o
: 9 the movemento[ electrons,in itself, is
homotropic,while the limitation of movementbetweenthe two oxygensis
an heterotropiceffect.With the electronfixed in one position,bound to only
one oxygen,a further step in immobilizationis achievedand representsa
heterotropicfactor. In more complex molecules,such as unsaturatedfatty
acids, for example,the tendencyof the electronsto wander is related to
homotropy, while their restrictionto the moleculeor even to certain areas
of the molecule representsan heterotropic cffect. This also applies to
micelles,where the water moleculesand impuritieshave a certain degree
of mobility. This mobility must be consideredto be a vestigeof the movement of free water molecules,with a high mobility consideredas a homopolar effectat this level.The retentionof water or other moleculesin the
micelle may be consideredas a heterotropiceffect.And their further fixation, as in the activatedmicelle,is a further heterotropiceffect.
Organization,which resultsfrom the alternateoperationof electrostatic

12

xEsEARcH rN pHysropATHoLocy

coulombianhomotropic and organizationalheterotropic tendencies,leads

to the realization not merely of stableconfigurationsbut, more significantly,
to entitiescapableof reactivity,and consequentlyable to respond actively
to the various changesof the environment.The fact that fulfillment of
quantum forces causesappearanceof new electrostaticforces, which will
be further neutralized,leads to the progressof organizationto ever-higher
levels.
At each step of organization,however, another characteristiccan be
recognized.It results,in part, from the fact that progresstoward higher
entitiesis accomplishedby an increasein complexity rather than only in
size.The increasein the positivecharge of nuclei in atoms, for instance,
bringsabout a parallelincreasein the number of surroundingelectrons,but
this goes on only up to a certain point. Actually, the size of the atoms is
limited by the size of the nucleuswhich, in turn, is limited by the quantum
forcesable to insurestabilityfor the nucleus.Nuclei becomeunstablewhen
they contain too many protons.
Levels, Entities and Constituent Parts
As alreadynoted, we definedan entity through its structural and functional individuality. we used the term "level" to indicate a conceptual
groupingof entitieshavingthe samebasicconstitution,suchas, respectively,
nuclei, atoms, molecules,micelles,etc.
We have used the term "part" to define an entity when it contributes
to the formation of anotherentity. Nuclei and electronsare parts that form
an atom. Molecules are parts when they are bound through electrostatic
and quantum forces to form micellar entities.In progressiveorganization.
eachnew entity thus is composedof partswhich are entitiesfrom the level
immediatelybelow, and the new entity itself servesas a part for the immediatelysuperiorentity. we call this relationship"hierarchic," one entity
being inferior to that which it forms and superior to those which have
formedit. So conceived,
eachnew organizational
entitycan be identifiednot
only through the nature of the parts forming it and the manner in which
they are bound, but also through its level in the hierarchic succession.
we have seenthat most of the entitiesare made up of dissimilarparts.
Analysis of what happenswhen an entity is formed has shown that the
processis complex.In order for an entity to act as a part in a higher level,
it must first pass through an activatedstage.Activation opens up many
opportunities,a plurality of possibleformations.So does another process.
an immediate consequenceof activation. Almost continuously, sevcral
5imilar entitiesare seento join togetherin a kind of "common grouping,"

B^SIC CONCEPTS OF ORGANIZ^TION

13

adding further to the multiple possibilitiesof new entities.The multiplicity

of possiblities
at eachstcpin organization
explainsthe exponentialincrease
in the number and complexityof the entities,resultingfrom the hierarchic
pattern of their formation.
According to the holistic approach,an entity exists only through its
own qualities.It must have characteristics
other than thoseof its constituents. It is the relationshipbetweenthe constituents,in the new entity,
largely resultingfrom the operation of quantum forces, that characterize
the entity.
Principal and Secondary Parts
As noted, cntities at progressivelevels of organizationare formed of
dissimilarparts.Theseparts do not haveequallyimportantroles.There is
a "principal" part which is characteristic
for a given level.l'here are "secondary" parts that are nonspecific
for the level,the sameonescan serveat
ditlerentlevels.The secondarypart for a hierarchicentity often is an entity
of a far lower level. (Fig. 1)

PrincipalParts
First [ntities

Secondany
Parts
o f t h e S e c o nEdn t i t y

S e c o nEdn t i t t e s
T h i r dE n t i t i e s
F o u r t hE n t i t y

f t h e T h i r dE n t i t y
f t h e F o u r t hE n i i t y

Ftc. | . The hierarchic organization. In the organization in general, the different

e n t i t i e s a p p e a r i n t e r r e l a t e da c c o r d i n gt o a c h a r a c t e r i s t i ch i e r a r c h i cp a t t e r n . E a c h e n tity is formed by a principal and a secondarypart. An entity is hierarchically "superior" to the cntities which form its principal part and "inferior" to rhose in the
formation of which principal part it enters.

Secondaryparts have important characteristicsin common. These become especiallyevident when lower level entitiesare examined.At levels
below the atom, secondarypartsfor all entitiesare electrons.It is of interest
to observethat in higher entitiesas well, such as moleculesor micelles,all
secondary parts have a negativeelectrical charge.

L4

xESEARcHIN pHyslopATHoLocy

Also characteristicof secondaryparts is their derivation directly from

the environmentin which the entitiesof which they are components,appear.
This is clearfor many crystals,wherewater moleculesrepresentthe added
secondarypart. This water of crystallizationis not free betweenthe ions
but bound to them. Water apparsindispensable
to crystal formation, since
its loss resultsin disintegration.Water can be consideredto play a secondary part. Similarly, in some crystalssuch as gold, some electronswander
betweenthe atoms while others are concentratedin certain regions. (Brillouin) These electronsrepresenta secondarypart in these crystalsas do
water moleculesin others. Water and electronscan be related to the environment from which they derive.
Derivation of the secondarypart from the environmentappearsto be
even clearerat the micellarlevel.In the caseof the gelatinousprecipitate
obtainedby coagulatinga couoidal solution,a part of the environmentin
which the gel is precipitatedentersinto the formation of the new micelle.
For example,the micelle of colloidal copper ferrocyanatecontainspotassium ferrocyanateas a secondarypart for each principal part of copper
ferrocyanate.Micelles of fcrric hydrate, obtained by the hydrolysis of a
boiling iron perchloride solution, provide another example. Besides the
Fe2o3, moleculesfrom the Fe6Clusolution used in the preparation enter
into the formation of this hydrosol.The role of the negativelychargedconstituentsbecomesapparentwhen a part of the cationof this secondarypart
is removedfrom the intermicellarfluid and the hydrosol still persists.It is
only whcn the chloridecontentbecomestoo low that coagulationresults.
The moleculesof potassiumferrocyanateor iron perchloride,once consideredto be impurities,must be looked upon as secondaryparts of these
micelleentities,Dcrived from the environment,they enter into the formation of the micelles,especially
throughtheir negativeelectrostatic
character.
I'his concept becomeso[ even greater importance when entities are
considcredin relation to the constantlychangingenvironment.The electrostaticbalancebetweenentity and environmentrealizedat any given time
cannot be consideredto be permanentbecauseof the changeswhich occur
in the environment as it travels toward ultimate total homotropy. As a
working hypothesis,
it can be assumedthat the relationshipbetweenentity
and environmentwould changeas the lattermovestowardtotal homotropy.
It can be assumed,too, that as hierarchicorganizationdevclopsin time,
secondaryparts from the environmentwould diffe.rfor differententities.
Changesin environmentwould providcevidencethat thcsesecondaryparts
are relatedmore closelyto the environmentas it existedin the past when

BASTC CONCEpTS OF ORCANTZATTON

15

these entities are assumedto have appeared.we will see below how important this is for the more complexentitiesof higher levels.
In the role played by secondaryparts in progressiveorganization,the
changesin their mobility are of specialsignificance.we have already seen
that in hierarchicentitiesthe secondaryparts are simpler units than the
principalparts,a factor which facilitatestheir mobility. The mobility can
be related to the fact that these secondaryparts are derived from the
environmentwhere they arc mobile, with their motion not systematized.
The interventionof quantumand quantum-likeforces,which help to create
new entities,can be seenas a kind of organizationof the relativemobility
of the secondaryparts. It is the systematization
of their movementwhich
preventscompleteannihilationof electrostaticforces present.The relationship between secondaryparts and environmentthus explains the
characterof the mobility encounteredthroughouthierarchicorganization.
It must be emphasizedhere that, becauseof the electrostaticnature of
the bond betrveenprincipal part and secondaryparts, a principal part is
capableof entering into the formation of more than one specifictype of
entity. similarly, fulfillmentof quantumforcescan lead to more than one
typc of structure,However, of the many possiblenew entities,or structures,only a few will fulfill the requirements
for developinga still higher
organization,i.e. will be capablcof acting as principalpart in a new cntity. some rcmain at their original level without progressing,
even after
being bound to other entities.Even many of thosewhich have somecapability for higher organizationcan go only one or two steps.Only a very
few will continueall the way up. [n other terms,only a few will be able
to utilizc new quantumforcesin order to realizencw entities.While various bondsand structuresoffer a largevarietyof possiblenew entities,it is
thc entity with a capacityto adcquatelyresistthe cffectsof the changing
environmentwhich will take part in progressiveorganization.
In considering the forces which intervene in progressivehierarchic
organization,one has to considerthe free cnergyavailablein the environment. The immenseamount of energyreceivedfrom the sun representsa
typc of heterotropicenergy which can intervenein organization.We will
see that this is easilyrecognizedfor higherentities.The ability of certain
entitiesto develop may be relatedto their peculiarability to utilize heterotropic forces, most of them of solar origin. The lesssuccessfuldisappearor
remain at lower levels.

16

xESEARcHIN PHYSIoPATHoLocY

The Organized Boundary

We have seen that an entity achievedthrough systematizationof the
movementsof its componentsacquiresa boundary betweenitself and the
environmentas a result of this restrictedmovement.The boundary does
much more than delimit the entity and constitutea barrier betweenit and
the environment.The electronsof the outermostshell form the boundary
of the atom, for example.It is through them that the atom realizesits relationship with the environment.Chemicalreactionis largely limited to this
boundary. In the atom, where the nucleusis the principal part, and the
electronsare the secondarypart, it is evidentthat it is the organizedmovement of the electronsthat providesthe boundary.The form and organization of electronicshells,and specificallyof the boundary shell of an atom,
are resultsof quantum forces.The environmentalnature of the secondary
parts and their buffering role make them of great importancein complex
boundary formation. Theoretically,hierarchicprogressmay be considered
to dependupon the developmentof secondaryparts which allow increasingly complex boundary formations. This explains the importance we
attachto the study of boundaryformation in higher entities.
To summarizethe above conceptof organization,the different entities
can be integratedinto a hierarchicorganizationalpattern which depends
upon alternate operation of the two fundamentalforccs, electrostaticof
coulombiannature, and quantum of organizationalnature. Entities can be
identifiedby the nature of the inferior entitiesthat act as constituentparts
and the relationshipbetweenconstituentsas principal and secondaryparts.
While the principal part is formed by an hierarchicallydevelopedentity,
the secondarypart is a secondentity from the environment.The incorporation of a part of the environmentinto a new entity correspondsto a systematizationof its motion. And it is through the organizedmotion that
appearsa boundary formation which marks the realizationof a new hierarchic entity.
This concept of organizationhas made it possibleto understandthe
relationshipof the seriesof entitiesthat composethe biological realm.

CHAPTER

BIOLOCICAL ENTITIES

\xr

VVrt" rHE coNcEpr of hierarchicorganization,it becomespossibleto

gain a new insight into, and understandingof, the biologicalrealm.
In the classicalview, just as simple substancesin nature are conceived
of as being formed by moleculesand atoms, biologicaUycomplex organisms are consideredto be composedof cells as fundamentalentities. In
arriving at complex organisms,however, it is granted that organization
has followed a definitepattern.At first glance,it is apparentthat cells are
grouped in morphologicallyregular ways to form tissues.Similarly, tissues
are groupd to form organs and thesein turn composethe organism.In
this classicalsystematization,
a complex individual would appear to be the
result of a grouping of cells, tissuesand organs,bound together in what
has been describedas an harmoniousmorphologicalrelationship.
In our study of organizationof the biologicalrealm, we have emphasized the individualization of both conceptualand material entities. In
some cases,entitieshave been simple to identify becausethey are easily
separablemorphologically.Where morphologicalseparationhas not been
immediately evident, other criteria-such as structural and functional
properties-have been used for identification.Besidesplaying its part in
organization,each entity has its own individuality,and consequentlycan be
recognizedholisticallyas a well-definedunity. Startingwith chromomeres,
the entity status is easily acceptedbecause,in addition to clear morphological and functional properties,there is a degree of independentindividuality. Following up, chromonemata,chromosomes,nuclei are other
entities.
The study of the organizationof biological entities has shown, however, that in all casesthere is a specificpatternof interrelationshipwhich is
more complex than the classicallyacceptedpattern. In the simplestmicrot1

18

R E S E A R c Hr N p H y s I o p A T H o L o c y

scopicallyidentifiedentities,it could be seenthat a seriesof chromomeres

are bound together through a specialfibrillar formation, which stainsdifferently from the chromomeres,
to producethe chromonemata(I,2), which
can be consideredholisticallyas a new entity. Two or four chromonematas
( 3 ), togetherwith the chromosomalsap,form the chromosomeas a new entiry. Simi.larly,severalchromosomestogetherwith another part-this time
representedby the nuclear sap and the proper nuclear membrane-form a
nucleus.In turn, the nucleus,plus protoplasmicformations,cytoplasmand
the cellular membrane,form the ccll. Even superficialanalysisindicatesa
common fundamentalpattern in the organizationalchangestaking place
for entities ranging from chromomeresto cells. Becauseof this pattern,
these entitics can be consideredto be "hierarchically" interrelated.One
entity is hierarchically"superior" to the entitieswhich form it and "inferior" to those it will itself help form. Two immediatelyinterrelatedentities thus are hierarchicallysuperior and inferior, respectively,just as in
organizationin other realms.(FiS. 1)
Analysisof the progressionof organizationfrom simple to more complex entitiespermitsus to recognizeother characteristics
of the fundamental
pattern.In order to form a hierarchicallysuperiorentity, severalsimilar entities first join to form a group. It is the group which then will bind
other constituentsto bring into cxistencea new, hierarchicallysuperior,biologicalentity. Thus, the chromomeresas a group join with a fibrillar formation to producechromonemata;
join with chromosomalsap
chromonemata
to form chromosomes.Groups of chromosomesplus nuclear sap form the
nuclei.It has appearedevidentthat the partswhich are bound to form each
hierarchicentity do not play equal roles. In each case, the principal part
is the one which is composedof similar entitiesacting as a group; the
other part is the secondary.Figure 2 offers a graphic representationof
hierarchicorganizationfrom chromomeresto cells.
In all entities,mentionedabove,thereis the samerelationshipbetween
principal and secondaryparts. The secondarysurroundsthe principal part.
The moqphological
relationshiphas helpedus to apply to thoseentitiesthe
hypothesisdiscussedpreviouslyconcerningthe mechanismthrough which
hierarchicprogressionhas taken placein nature.According to the hypothesis, severalsimilar entitieswould first associateand form a group. In a
secondstep, the goup would tend to maintain around it a small portion
of its immediateenvironment.From this portion o[ the environmentrvould
come the secondarypart for the next superior hierarchic entity. With a
boundary formation, separatingthis minute part from the rest of the environment,the new entity would be established.Such a processcould oc-

BroLocrcAL ENTtTIES

19

cur, although rarely, with a singlebiological entity serving as principal part.

Usually, severalentities grouped togetherwould be needed.This pattern
explainswhy the secondarypart can be conceivedof as a part of the environment retained around the principal part, and rvhy the establishmentof
a new and higher entity can be consideredto occur only when this secondary part is detachedfrom the rest of the environment and separated
from it through the interventionof a boundary formation,

Entities.

AddedSecondary

Format
ions
Chromomeres
Chromonemata

Fibrillar material
Chromosomial
sap

Chromosomes

lfucleus
CelI

Nuclearsap
Cytoplasma

Ftc.2. The hierarchic relationship in the organizationof morphological entities below

c e l l s . F o r e a c h e n t i t y i t s p r i n c i p a l p a r t i s r e c o g n i z e da s b e i n g m a d e b y a g r o u p i n g o f
entities hierarchically inferior to it. The secondarypart which correspondsto a kind
of environment for the principal part usually surrounds the principal part.

Having recognizedthis pattern of organizationfor lower entities, we

lr'ent on to determinewhetherit remainsthe same for higher entities.It
could be seenthat groups of cells,along with interstitialformationsand
fluids around them servingas secondarypart, createthe tissueas a new
hierarchicallysuperiorentity. Indeed,a proper boundaryformation morphologicallylimits and conceptuallydefinesthe new entity. The interstitial
fluids are separatedby a continuousendotheliumlimiting the lymphatic
spaccsas a systemclosedtoward the intercellularspaces.Under thesecircumstances,the lymphaticendotheliumservesas the correspondingboundary formation that limits the tissueentity. Severaltissuesgrouped together,
playing the role of the principal part, bind the lymph, as secondarypart,
to form the organ, as a new hierarchic entity. Lymphatic vesselsand
connective tissues represent the organ's boundary formations. Further-

; : : i '
. r ' :
I ]!a:; trr:4trr:itl'riitir{)

Nl

20

RESEARcH rN pHysropATHoLocy

more, organs grouped together, with blood as secondary part, form the
entity called the organism.(Fig. 3,)
Does the samepattern apply for entitieshierarchicallyinferior to chromomeres?For these lower entities, morphologicalinformation to define
the relationshipbetweenprincipal and secondaryparts is unavailablefor

Entitv

Added
Secondary
Formations

_ _ I n t e r s t i t i a lf o r m a t i o n s
Tissue--Lymphat
ic fornations
0rgans

0rganism

- -8lood f ormat
ions

Ftc. 3. The oganization abot'e the cells. The hierarchic pattern of the organization
in general is recognized also in the organization above the cells. Tissues, organs.
organism are seento representhierarchic entities,each one being made by a principal
p a r t f o r m e d b y t h e g r o u p i n go f e n t i t i e sh i e r a r c h i c a l l yi m m e d i a t e l yi n f e r i o r t o i t . T h e
principal part is bound to a secondarypart characteristicfor each entity. This secondary part would correspond to the proper environment in which the entities forming the principal part, have evolved.

the moment. Until electron microscopyand other means provide such

data,wc are obligcdto find other criteriato indicatewhich, in thesehierarchic entities,is principal and which secondarypart. We have considered
electricalcharacteristics
of the constituents
as criteria for identifyingtheir
role in the hierarchicorganizationof biologicalentitiesbelow chromomeres.
Biological Realm
Before going further in trying to analyze submorphologichierarchic
entities,one problem was to establishthe limits of the biological realm

BroLocrcAL ENTITtES

2l

itself. The questionwas,how far below the morphologicalformationscould

we go and still have entitieswhich can be regardedas biological.With the
progressof science,criteria previouslyusedto definelife have becomeoutmoded. A separationline betweenthe animate and inanimate no longer
cain be drawn. with the study of the propertiesof viruses, it appeared
impossibleto maintain the last vestigeof the old vitalisticconcept.According to most of the classicalcriteria, viruseswould representanimate entities sincethey are able to multiply and conservea strict identity. However,
they also form salts,are crystallized,broken down, and then reconstructed
in the same or another order. If the virusesare acceptedas ,,borderline"
entities,as proposed,then the concept of animate and inanimatecan no
longer be sustained.
Although the animateand inanimatecannotbe distinguishedin terms of
a specificproperty, we cannot totally overlook the fact that an important
group of entitiesappear quite different from others in nature. As animals
and plants are difterent from stones,even without any vitalistic concept,
we are obliged not only to recognizethe difference,but also to try to
establishwhere the differencelies. To conform to reality, we have applied
the term "biological" to a group of entities,but we have given the term a
new meaning.
Just as organic chemistryis considered-whether correctly or not-to
comprisecertain combinationsof carbon,we considerthe biological realm
to compriseentitieshierarchicallydevelopedfrom a specificchemicalradical. This basic radical includesnitrogen and carbon atoms bound together
to form the N-c-N-c group. According to this concept,starting from the
basic nitrogen-carbonformation, an entire seriesof entities has been developed through hierarchicorganization.Togetherthey form the realm to
which the term "biological" can be applied.
The N-c-N-c group, through combinationwith hydrogen,would result
in radicals with a strong alkaline property; that is, with strong positive
electricalcharacter.Someof theseN-C-N-C groupstake part in the formation of nitrogenousbases,pyrimidinesand purines,while some,by acting
as principalpart and bindingvariousamino acid radicalswould, with the
necessary
electrons,
build up a new group of entities,arginineand histidine
as alkalineamino acids.Thesecan be considered,
hierarchically,
to be the
immediatesuperiorsof the N{-N-C goup and thus to representthe first
biological molecules.The alkaline positivecharacterof these moleculesis
noteworthy.Following the hierarchicpatterndescribedabove,severalsuch
alkalineamino acidslinked togetherto a seriesof entitiesof the samelevel
(simple amino acids) will form new groupsthat are still electricallypositive:

22

nESEARcHrN PHYSToPATHoLocY

will act as principalpart

In a newstep,thesehistones
thealkalinehistones.
(Fie.a)
to produce
nuclearentities.
parts,can form not one
Principalparts,by bindingdifferentsecondary
but manynew and differententities.More than one hierarchicline can be
identified.An especially
importantline resultswhenhistonesbind one or
more entitiesof the nucleicacid group,to form nucleo-proteins.
Other
histonescan bind variousother secondary
partssuchas carbohydrates
or
lipids and, in so doing,form differentbiologicalentities.Someof these

Princtpal

Added
Secondarv
Format
ions

il-C-l{-CqrouD'

Alkaline-ami!o

Aminoacid group

Histones

aminoacids

Deoxycibo-, \
nutleoproteins

Deoxyribonucleic
acld

Genes

Nucleooroteins
and
o t h e ls^e c .e n t i t i e s

Ftc. 4. The organizalion ol the submorphological entities of the biological realm.

T h e s a m eh i e r a r c h i cg e n e r a lp a t t e r n .w i t h p r i n c i p a la n d s e c o n d a r yp a r t s i s r e c o g n i z e d .
S t a r t i n gw i t h t h e p o s i t i v cC N C N g r o u p , e a c h e n t i t y h a s i t s p r i n c i p a l p a r t m a d e b y t h e
grouping of immediately inferior entities bound to secondary parts, generally more
electronegative.

entitiescan continue their hierarchicdevelopment.Nucleo-proteinscan

have riboseor deoxyriboseand consequcntly
form more complexnucleoproteins.Through all hierarchicachievement,
it can be secnthat new lines
are formedwhen groupsof hierarchically
lower entities,actingas principal
organizedpart, bind diflerent secondaryparts, all more negativcthan the
principalpart. Theoretically,this would rcsult in a sericsof entitiesall at
the samelevel,i.e. entitieswith similar principalparts but havingdifferent
sccondaryparts.Somewould go on to developsuperiorhierarchicentities,
otherswould evolve no further.
Differententitiesof the the samelevelcan be groupedtogetherin various ways to form a variety of principalparts. Always the group must be
madeup of entitiesof the samelevel.This requirementhas been seento
can exist betweenthe
be generalat all hierarchiclevels.Sincedifferences

B t o L o c r c A L E N T T T T E S/

23

constituents
forming the principal part for a new entity, the predominance
of one or another constituentwill make entities at the same level differ.
This mechanismof differentiationthrough the constitutionof the principal
part has been extremelyimportant throughoutthe biological realm.
Nucleolus
The concept that plural groupingscan enter into the principal part of
the nucleusputs the role of the nucleolusin a new light. It was accepted
for a long time that the nucleolus representsonly the reserve material
necessaryfor metabolism of the nucleus. The strong positive electrical
characterof the nucleus,as recognizedthrough its rather alkaline reaction,
would give it roles more important than that of the other constituents.
According to a work hypothesiswhich we advance,successivenuclcolar
formationswould representthe principal parts of hierarchic organization
below the nucleuslevel. In chromonemata,chromosomeand nucleus,the
partscorrespondingto the nucleoluscan be recognized.These formations
are grouped together with genesto form the principal part of the chromonemata.Similarly, chromatineformationsrepresentingentitiesof the
samelevel as the respectivenucleolar formations,will form togcther the
groupscharacterizingthe principal part of chromosomes.In the nucleus,
the nucleolusis joining the other formationsto form is principal part.
Protop lasmatic F ormations
In the cell, a hierarchicallysuperiorentity, a similar condition also
appearsto persist.The protoplasmaticformationswith ribo-nucleicacid
can be conceivedas representingentitiesof a nuclearlevel, that is, a level
similar to that of the nucleus.Togetherwith the nucleusthey would form
the group correspondingto the cell. This kind of evolution of entities in
relatively separateparallel lines, with their further grouping together to
form principal parts for new entities,is part of the typical pattern of
organizationespeciallyevidentin the biologicalrealm.
Boundary Formalions
We have mentionedthat groupingsof severalentitieswould not be
sufficientto form a new entity so long as the secondaryenvironmentalpart
is not isolatedfrom the medium from which it originates.Consequently,
the new entity appearsonly when a distinctboundaryformation is formed.
Progressivehierarchic developmentis dependentupon the appearanceof
suchboundary formations.For the first biologicalentities,the radicals,the
boundary semsto be more an energeticproperty than a morphologically

24

nESEARcH rN PHYStoPATHoLooY

organizedformation. For the molecules,it can be consideredto consist of

molecular surface forces, recognizedas the van der Waals cohesionforces.
A similar but more apparent boundary formation can be found in higher
molecularcomplexes,especiallythe micelles.The molecular arrangementat
the surface of micelles separatesthem from their environment and consequently insurestheir identity. In the caseof morphologicallyidentifiableentities,of course,boundary formations can be easilyrecognized.Chromomeres
are welldefined and separatedfrom the chromosomal sap. The chromosomes,in turn, show a real membranejust as the nucleusand the ceil do.
The next higber entity, the tissue, is bounded by the endothelial cellular
layer, separatingthe interstitial formations from the lymphatic spaces.Usually the boundary of organs which have tissuesas principzrl and lymph as
secondaryparts is representedby organizedblood vessels.As far as the
organism is concerned,the mucous membranesand skin are boundary
formations.(Fig.5)
H i erarchi c I nterrelationship
Viewed as a heterotropiceffect, hierarchic organizationcan be consideredto be a method of conservingexistingentitiesas such, in spite of
changesoccurringin the environment.TeleologicaUyspeaking,by entering
into the formation of a new and superior entity tfuough the system of
hierarchicorganization,each entity, in fact, protectsis own individuality.
The hierarchic organizationmakes it possiblefor each entity to continue
to live in a medium which correspondsto its own environment.The constituentsof the secondarypart in the new entity are chosento correspond
to the environmentin which the principal part of the entity has existed.
The successive
secondaryparts, added during the hierarchicdevelopment,
act as multiple protectivebuffers for the first entities,thus insuring their
unaltcredconservationin spite of continuouschangesin the environment
brought about by increasinghomotropy.
PhylogeneticDevelopment
Hierarchic organization,when relatedto time, would appear to correspond to evolution. The conceptof ontogenesisreproducingphylogenesis,
appearsin a new light when analyzedin accordancewith hierarchic organization. The parallelism between actual hierarchic organization and
hierarchicphylogeneticand ontogeneticdevelopmentgreatly helps to increaseunderstandingof many principal problemsof biology.
In accordancewith the conceptof hierarchicorganization,when a new
level is realizedthrough the binding of entitiesfrom a lower level, as prin-

BroLocrcALENTrrrEs

25

cipal part, to differentsecondaryparts, severaloutcomesare possible.

Someof the new entitiesare unableto continueto existand disappear.
Of
the others,somearerelatively
wellbalanced
entitiesandconsequently
can

Enti ty

ChromomeresChromonemala/-Chromosomes.Nucleus- -

formation
Boundary

boundary
Chromomere
boundary
Chromonemata
membranE
Chromosomial
-- Nuclearmembrane
-.Cellularmembrane

- Lymphat
ium
ic endothel

C eIIs

Tissues-J-

0roans
- F-- I
\

B l o o dv e s s e l s
S k i na n dn u c o u s
menbranes

\
\
\

0rganism

Fto. J. The boundary lormation in the hierarchicorganizationof the biological realnr.

A proper boundory lormation delimits each entity, insuring lhus its individuality. It
is by separatingcach time the secondarypart from the rest of the environment that
the boundary formations have pcrmitted the progressivedevelopment of the hierarchic organization. The boundary formation governs the rclationship bctwecn the
entity and its environment.

persist.But with further changesin the environment,some of theseentities

disappear.Even among those that remain, some do not representfully
satisfactorysolutions and further hierarchic developmentis necessaryto
insure tbeir persistence.They must evolve further to correspondto the

26

REsEARcH rN pHysropATHoLocy

new unfavorableexternal conditions.This has led to the developmentof

the complex entities presenttoday in nature.
The multiple entity possibilitiesat each level have resultedin a wide
variety not only of entities which, with further evolution, could produce
higher complexes,but also of others which have ended their evolution at
Iower levels. So the existenceof many varied entities apDearsto be the
result of the existenceof multiple solutionsfor the same problems which
have been part of the mechanismof hierarchicevolution.
From this viewpoint,existingindependententitiescan be recognizedas
correspondingto the different levels of hierarchic organization.Viruses
can be consideredto be at the same level as genesor even the entities
immediatelybelow genes; the microbes at the same level as the nuclei:
monocellularorganismsat the level of cells, etc. Furthermore,in each
independentorganism, from simple to most complicated,the same progressionin organizationof successive
hierarchicentitiescan be seen,starting with the simplest biological entities and continuing until the level at
which the entity has actually stoppedits hierarchicevolution.
In a further step, having arrived at thc concept that the secondary
part of each entity correspondsto the kind of environmentin which the
entity found itself at the time of its phylogeneticappearance,we tried to
seewhat information about theseenvironmentscould be obtainedthrough
study of the secondaryparts.
The first biological entities,the alkaline amino acids, could have appearedin an atmosphererich in ammonia,water and methane.Experimentally,electricaldischargesthrough mixtures of these materialshave
induced the appearanceof amino acids.It appearsplausiblethat such an
environmentcould have existedaroundvolcanoesin times when the earth's
atmospherewas formed by ammonia. with steam formed by the heat of
the volcano, with methaneresultingfrom the interactionsof erupted metallic hydrocarbonswith watcr, and with lightning so frequent around
volcanoesin eruption, the necessaryconditionsfor synthesisof amino acids
may have been present.The simple amino acid moleculeswhich would
haveresultedcould haveconstituteda group neededfor hierarchicdevelopment. Out of a seriesof such amino acids,some could have bound the
group N-c-N-c which also could have been synthesizedunder the influenceexertedby radioactiveelementsor radiation.(Note I )
Additional information concerningthe constituentsof secondaryparts
which enter into the developmentof all the subnuclearentitiesis mcager.
Simple amino acids or urea are presentin the chromosomialand nuclear
sap which are practicallyfree from K and Na. We could thus consider

BfoLoorcAL ENTtTTES /

27

arumonium as the predominantcation for all hierarchicentities up to the

nucleus. This would accord with geologicaldata concerning a primitive
atmospherein which ammonia was predominantat the time when the first
biological entitieswould have appeared.we can then, tentatively,in view
of this cation common to all, classify the hierarchic entities below the
nucleusin what we will call "the nuclearcompartment."
In the same way, we analyzedthe compositionof cytoplasm,with the
thoughtthat it could provide informationabout the constitutionof a second
environmentin the evolution of biologicalentities.The principal cation of
cytoplasm is potassiumwhich also representsthe principal cation of the
earth'scrust. Curiously enough,potassiumand the other constituentscould
oe found in the same relative proportions in the earth's crust as in cytoplasm, (Note 2) a f.actwhich seemsto confirm the hypothesisthat mud,
humid earthcrust,represented
the environmentin which entitiesat nuclear
levels lived. The cytoplasmconservedthis constitution,with potassiumas
principal cation, when it was separatedfrom the environmentto become
the secondarypart which, with the nucleus,formed the cell as the next
superiorhierarchicentity. The cell by itself, representsa new compartment
with potassiumas principal cation.
It is only for the animal cell that the environment seems to have
changedagain.This time the new environmentwas the sea.when several
cells joined togetherto organizetissueas a new entity, they had to maintain their environment,now representedby the sea.The hierarchicentities
above the animal cell show sodium as principal cation in their secondary
parts, thus indicating that when they were organized the sea was their
environment.This characteristic
allows us to group togetheranimal hierarchic entitiesabove cells to form a new compartment,the metazoic,with
sodium as the principal cation.
With passagefrom marine to terrestriallife, air is found as the new
environment.Not integratedas a new secondarypart, without a separating
boundary formation, the part of the environmentkept in the respiratory
apparatusdoesnot enter however,into the formation of a new entity. Only
the presenceof air in the bonesof birds can be seenas such integration.
A certain fundamentalfurther development,in the same direction can
be seenin animalsas well as in humans,in the areaof sociallife. (Note 3)
Divisionof complexhierarchicorganization
into compartmentsappears
to be relativelysimple. There are changesin the principal cation from
compartmentto compartmentwhich correspondto similar fundamental
changesin the environmentthrough which the organismpassedduring its
phylogeneticevolutiol-f1e61 volcano to mud, to sea, to the surface of

28

REsEARcH rN pHysropATHoLocy

the earth. We tried similarly, to correlateother elementsin the periodic

chart with hierarchiccompartments.The resultswill be presentedin detail
later. For the moment, it can be stated that elementscorrelated to different environments are also found in the difterent compartments correspondingto these environments.For example, Mg like Na representan
elementof the metazoiccompartmentand of the sea where this compartment was phylogenetically
organized.Fe, Cr, Ni, Zn, Ca, Mn, Co, As, and
Se which fall into the cellular compartment, represent characteristic constituentsof the earth'scrust.
Constants
The conceptthat matterin generalis the expression
of the heterotropic
trend permits us to explain further some of its characteristicswhich have
been of great importance in biological development.Heterotropy can be
seenworking to maintain existingentitiesas long as possible,to conserve
their characteristicproperties in spite of changesin the environment. Heterotropy could result in unchangedvalues which would appear as constants of an entity and would indeed identify the entity. It is hierarchic
organizationwhich would tend to permit conservationof constants.
Each biological entity-just as any entity in nature----canultirnately
be defined by the series of characteristicproperties which it is able to
conserve.Constancyis the criterion which permits us to judge the importance of a property to an entity. The longer a property is kept constant
despiteenvironmentalchanges,the greaterits fundamentalimportanceto
the entity. The progressiveaddition of secondaryparts through hierarchic
organizationrepresentsan effectivemeans of preservingconstantsof the
lower entities. New properties, added with the formation of each new
hierarchicentity, representednew constants.This explainswhy considering
the constants,it can be seen that those incorporatedin the lower entities
are the best conserved.The higher a constantin the organizationalhierarchy,the lesswell-preserved
it will be.
The idea that constantscorrespondto the character of the environments through which the individual has phylogeneticallypassed,and that
they are conservedthrough hierarchicorganization,permits us to try to
extend our understandingof conditionspresent in past environments.
Cationsand even anions would representonly one type (apparentlythe
most important) of the constantsmaintainedthrough hierarchicalorganization. Other constants,correctly interpreted,would indicate in what
direction we must search for conditions which prevailed in the environ-

BroLocrcAL

ENTtTTES

29

mentwhen the respectivehierarchicentitiesthat make up a given organismwere establishedduring phylogeneticdevelopment.

As examples,let us considerthe conservationof salhity and temperatureas constants.Values for salinity of the metazoiccompartmentand of
theblood and valuesfor temperaturehavc been seento be constantscharacterizingspecies.Differencesbetweentheseconstantsin different species
showa succession
conforming with paleontologicaldata. In the interpretation given by Rene Quinton, constantswould indicate the times when
various speciesoriginally appearedin nature.
According to our concept of hierarchicorganization,these constants
may be interpretedotherwise.They would not indicate the moment when
the lowest entities of the respectivespecieswere formed, but rather the
time in the developmentof thesespecieswhen the hierarchicentitiescapable of conservingthe respectiveconstantsappeared.In other words, they
would not indicate the earliestmomentsof appearanceof the first entities
which later developedto form the respectivespecies,but would indicate
a relatively Iate moment in the creation of the metazoicentity which has
appearedable to conserve,as its own constant,this specificattribute of the
environment.In the caseof salinity,this would correspondto the constitution of the metazoicentity itself which has retainedthe compositionof the
early seain its intercellularfluids. As far as tempratureis concerned,the
entity which would appear able to conserveit has to be regardedas much
more complex and even to be related to the appearanceof systemsof
organswhich are sufficientlysensitiveto changesin temperatureand which
also possessthe means of insuring constancyfor temperature.
The conservationof different elementsin different compartmentsappears to be characteristic.
In order to maintainits constantsfor elements
the entity has to opposetheir uncontrolledcirculation.The role of hierarchic entitiesin conservingancestralconditionswould explain why an
entity would have to opposeparticularlythe penetrationof the constituents
which characterize
the succeeding
environments.
This has appearedevident
for the cations.The boundary formationswhich have to ptay the principal
role in the creation of each entity must also insure its identity by barring
uncontrolled penetrationof elementscharacterizingthe new environments.
Invasion by such elementswould correspondto an abnormal event which
must be corrected. If the invasion progressedbeyond a certain limit, it
would createa conditionincompatiblewith further existenceof the entity.

30

nESEARcH rN pHystopATHoLocy

|later Circulation
The passageof complex organizedanimals into the new environment
of terrestrial life brings to the fore the problem of the place of water in
hierarchic systematization.It appears to us an acceptable concept that
water doesnot circulatefreely in the organism.Its appearancein hierarchic
entitiescan be understoodif, as we did for the other constituents,we relate
water to its place in the environmentin which phylogeneticdevelopment
has taken place, In the first near-volcanoenvironments,which applied to
the subnuclearentities,water was relativelyscarce,which explainsthe high
concentrationof the constituentsin the nucleus.The mud of the earth's
crust is richer in water, which explainsthe difterencebetweenthe nucleus
and the cytoplasm, with the latter richer in water. The sea was the
environmentfor the metazoiccompartment,which explainsthe richnessof
water in this compartment.The so-called"internal sea" consequentlycan
be seen only in the metazoic compartment.With the passageinto the
terrestrialenvironmentwith its air medium,the water againbecomesscarce
and has to be conserved.The circulationof water betweencompartments
is governedby osmotic forces which are determinedby the original richness in water of the respectiveenvironments.The importance of water
circulation appearsevidentwhen an abnormalityin its distribution occurs.
Water arriving in a compartment-alone or with a cation-in an amount
abovethatcorresponding
tothe constantfor that compartment,isseparated
from the constituentsof the entity in order to reestablishthe characteristic
constant value. Such separationof abnormal amounts is accomplished
accordingto the compartment,through the appearanceof vacuoles,edema,
exudatesor diuresis.
Animals and Plants
The conceptof hierarchicorganizationin which each entity canconserve
its own environment allows us to consider in a new light various other
problems of living organisms.One concernsthe fundamentaldifferences
betweenanimalsand plants.Analysisof the constituentsof the metazoic
secondarypart providesa new criterion for distinguishingbetweenanimals
and plants and gives logical meaningto its distinction.Animals can be
characterized
as having sodium as the cation of thcir metazoiccompartment; from the cell level on, they have had the sea as their temporary or
even permanentenvironment.Plants,on the other hand, have potassium
as the principal cation for their metazoic compartment, indicating that,
from the cell level on, they have had the earth'scrust as their environment,

B r o L o c r c A LE N T r r r p s /

lt

passingthus directly from mud to air. By their actual attachmentto the

soil, plants continuethis relationshipto the mud. Their relative immobility
is in accord with continuation of the terrestrial-airenvironment in their
development.The mobility of animals, on thc contrary, can be seen to
have its origin in the fact that they have had the sea as thcir environment
at least for a period of time, r.e.,from the cell period until the appearance
of those animalswhich left the sea. We can interpretthe appearanceof
celluloseand lignin as part of the plant-sustainingmeans which would
bring to plants indispensableexternal protection against the hardnessof
the soil environment.Cellulose and lignin are not necessaryfor animals
which experienced
much of their evolutionin the sea.
Multiplication
The hierarchic organization of organisms,with the conservationof
successive
entities,puts the problem of cntity multiplicationin a new light.
According to the hierarchicorganizationconcept, the multiplication of a
complex entity means the reproductionof the entire seriesof hierarchic
entities forming it. In this process,the intervention of each hierarchic
entity appears highly individualized.And this applies not only for the
morphologicallyidentifiableentities,but even for the most primitive entities.The difterencebetweenthe role playedby the principal and secondary
parts becomescapital for theseprocesses.
While for each entity the principal part has to be built as such, the parts correspondingto the secondary
Parts are taken from the immediateenvironment,The quantitativedisproportion betweensome principal and secondaryparts makes the role of the
first diftcult to be recognized.The complex entity, through changesthat
are the reverseof thoseof ontogeneticand phylogenctichierarchicdevelopment, separatesthe successive
principalparts which characterizeit. With
the replication which takes place the division occurssuccessivclyfor these
hierarchic entities. In scissiparitythe division morphologicallyoccurs at
the cell level; in karyokinesis,it can be identifiedat the chromomerelevel
and certainlytakesplacemuch lower in the hierarchicentities.In replication in general,different constituentsavailableare adequatelychangedby
the respective principal part to form the necessarysecondary parts.
Through these changesthe same processesare reproducedwhich originally occurredwhen the entity had been phylogcnetically
organized.
individualization
With the
of the low hierarchic entities the problem
of replication is simplified.Once replicationoccurs,the sameprocesstakes
place successivelyfor the progressivelyhigher levels.Above chromomeres
this appears very clear in karyokinesis.

32

REsEARctf IN

PHYSIoPATHoLocY

After the chromomeresdivide, two or four chromonemataappear.The

processgoes on within the chromosome,nucleus and cell. In order to
protect its individuality each hierarchicentity is protectedduring its division. The chromosomialmembrane,the cellular cytoplasmand the cellular
membranecontinue to protect the respectiveentities as they divide. The
cell itself divides only when the two nuclei have had their protecting membranes rebuilt.
In the division and multiplication of a complex entity, the return to
entitiesas low as subnucleicparts indicatesthe relativeimportancefor the
characterizationof the complex entity and for the conservationof its particular properties of the parts added during the hierarchic development.
The entity must, in fact, rid itself of these added parts which, although
they have other importance,have a secondaryrole even in the processes
of multiplication.
It is interestingto note that a similar return to more primitive component entitiesalso occurs,althoughit is lesspronounced,when an entity,
tissue,organ or organismhas to fight a noxious intervention.The defense
is passedprogressivelyfrom the organ to the tissuesand from theseto the
cells.In effect,there is a renunciationof addedparts during thesemoments
of crisis.Even at the cell level, a similar processis seen.The added parts,
represented
formations,disappcar.The almostnonby the protoplasmatic
differentiatedcell fights the noxiousfactor at the lowest levelsof its organization.
Lile and Death
We have seenthat the term "biological realm" can be applied to hierarchic developmentstartingfrom the N-C-N-C radicals.We have employed
this term for didacticconveniencealthoughit is unrelatedto the commonly
acceptedconcept of life. The study of hierarchicorganizationalso led us
to considera conceptof life and death which, while retainingsome of the
common meaning, also accordswith the phenomenaof hierarchicorganization.
In the complex entity, each lower level entity lives and dies with
relative independence.An organ can be dead and yet have living cells
in it for a time. There are always dead cells to be found in living tissues
of the different hierarchicentiand organs.It is the relative independence
ties making up a complex entity that explainstheseseemingpeculiarities.
Our concept of life and death stemsfrom considerationof the nature
of hierarchic entities. We have seen that all matter in nature. from the
simplestto the most complex entity, is a result of heterotropy.The per-

B I O L O c T C A LE N T I T t E S

33

sistenceof constantsproper to each entity is distinctly opposed to homotropy. Life in its broadestsense,correspondsto the capacity of an entity
to maintain heterotropyby conservingits characteristicconstants.The life
of any entity aPpearsto be synonymouswith conservationof its constants.
An entity dies when it has permanently-that is, irremediably-lost its
capacityto conservethe constantswhich characterizeit. Death then representsexhaustionof heterotropyfor the specificentity.
The fact that, in essence,life appearsto be synonymouswith the conservation of constants and is heterotropic,relates it, and especiallyits
origin, to one of the important sourcesof heterotropicforce, solar energy.
A distinction has to be made betwcen heterotropyas one of the fundamental laws of nature and the meansby which it is exerted.Solar energy,
with all of its quantas,would greatly increasethe effectsof heterotropic
forces in nature. It would not createsuch forcesbut would simplify them
and extend their applications.The origin of matter and, as we have seen
above, of entities, biological or nonbiological,is in the final analysisthe
result of heterotropicforces.External conditionsqualitatively and quantitatively influenceoperation of heterotropicforces.
The sun's heterotropiccontributionshave to be consideredunder this
aspect.Throughthe quantasit disposes
of, solarenergyhasnot createdlife,
as conceivedabove, but by permitting more and more entities to appear,
has greatly facilitated their extension.Its effect, although certainly not
limited to any group of entities,seemsto be especiallyimportant to those
forming the biological realm. Similarly, the effect, of a special type of
energ.y,radiation,also must be considered.Radiation appearsto be related
to the elements,and will be discussedin a later chapter devoted to the
elements.
Sincelife itself is related to changesdirectly aimed at conservingconstants,in this broadestsenseit is no longer limited to the specificgroup of
cntitiesfound in the "biological" realm. Life has the samemeaningfor an
atom, crystal or micelle, as for a cell, organ or organism.It is for this
reason that knowledge of the mechanismthrough which constancy is
achievedbecomesof great importancein the study of all matter and especially of the biologicalrealm.
Maintenattce ol Constants
To study the mechanismused to maintain constants,we must define
exactlywhat constancymeans.According to Cannon'sprinciple of homeostasis,
constantshave beenconsidered
to correspondto a dynamicbalance
that results from the continuousoperation of two opposingfactors. And

34

R E S E A R c Hr N p H y s r o p A T H o L o c y

it was conceivedthat by actingconcomitantlyas coupled antagonists,these

factorsor groupsof factors insureconstants.The interventionof thesetwo
opposite factors becomesapparentonly if an exterior cause upsets their
balance.
However, our study of the processesthrough which dynamic balance
is maintainedhas permittedus to recognzea differentmechanismthan the
one which is commonly accepted.
A value consideredto be a constantfor an entity is not fixed or static.
It represents,rather, a statisticalvalue, the result of a seriesof dynamic
changeswhich must also be consideredin terms of time. Consequently,a
constanthas to be seennot only as the averagevalue of a seriesof organized changes,but also must be identifiedby the characteristicsof the variations. An averagevalue around which variationsoccur thus representsthe
first attribute of a constant. The second attribute is the existenceof a
rhythm in the variations,the third involvesintensityof variations.For
instance,when we say that human body temperatureis constant,we mean
that 37'C is averagevalue for oral temperature,and also that body temperature presentscharacteristicvariations having a 24-hour rhythm and
also that the occurringchangesconsistof variationsof a few tenthsof a
degreeabove and below the averagevalue.
The two antagonisticinterveningfactors do not operateconcomitantly
to maintain a constantvalue, but rather act alternately,each being predominantfor a period of time.The resultis not a continuouslysteadyvalue
for the constant,but an oscillatorymovementwith successive
passages
from one sideto the other of the averagevalue.This oscillatorymovement
appearsto be the generalrule throughoutnature, prevailingin everything
from the wavesin the smallestsubatomicparticlesto the pulsationof the
universe. The rhythm periods appear to correspond to environmental
rhythms.A rhythm relatedto the day, for instance,is seenfor temperature.
In other constantswe recognizea l2-hour rhythm which could correspond
to that of the oceantides.Other rhythms,with periodsrangingfrom two
hours to a few minutes are seen for severalchangesoccurring in blood.
Therc are also some in which the influcnceof thc moon is evident; for example,the hypophysis-ovarian
cycles;and for others,the influenceof the
seasonsis apparent.
Teleologicallyspeaking,balancerepresentsa very effectivemethod for
maintainingconstants.Any deviationin any direction as a result of an
externalinterventionwill be counteracted
by the opposingphaseof the oscillatory balance.This occursbecauseof the existenceof two phasesof thc

BIOLOCIC^L ENTITIES

35

oscillatorymovementitself. Such would not be the caseif there wcre fixed

valuesfor constants.
Related to the pattern of the organizationo[ matter in gcnera.t,this
oscillatorymovementcan be consideredto bc anotherinstancein which the
two oppositefundamentalforcesof heterotropyand homotropy, which are
basicto progressivehierarchicdevclopmentitself, also operate.This oscillatorybalancecan be relatedultimatelyto the alternatesuccessive
interventionof thc heterotropicand homotropictrendsin the organizationand the
manifestations
of entitiesexistingin nature.
Dualism
The concept of dynamic oscillatorybalanceis of great importance in
the study of biological phenomena.coupled facrors with opposite propertiescharacterizeall constantsand are involved both in the processes
throughwhich constantsare maintainedand in their manifestations
as well.
Recognition
of this dualismin all biologicalphenomenahas been of great
valuein the investigationof normal and abnormal physiology.
According to our concept,dualismresultsfrom the alternate.not concomitant,operationof two opposingfactors.And, as we have seen,these
factorsultimately can be related to the two fundamentalforces in nature,
homotropicand heterotropic.Thus, in a unifiedconcept,in every phenomenon in which dualism appars,one force will be homotropicthe other will
be heterotropic.Homotropy is relatedto fulfillment of clectrostaticforces,
andhasgeneralcoulombianelectriccharacter.Heterotropyis quantum-like
and organizational.Homotropy would keep entities simple. Hcterotropy
would lead to more organizedbonds and to more complex synthesis.In
everyphenomenonstudied,thesecharacteristics
of the two fundamental
forceshave permitteddualism to be recognizedand interpreted.The dualisticview has becomeour basicapproachfor all of the problemsrelatedto
matterand, more specifically,to biologicalentities.
The dualistic conceptof interveningforcesbrings an entirely new light
in any analysis in which a graphicalrepresentation
is different from a
straightline. From the curvesof spectralanalysisof constituentsto those
of complex phenomena,the existenceof oscillationsrevealsthe intervention of opposed forces and offers a valuable mean to study them. This
broad approach has a specialfield of applicationin biology.
Dualism can be further recognizedeasilyin the manifestations
of the
in
biologicalentities, their function and in the substances
composingthem.
In the case of the elements,such a dualismcan be relatedto atomic structure and the propertiesof the electronicshells,as will be seen below. In

36

nEsEARcH rN pHysIopATHoLocy

complex molecules, a simple form of dualism can be seen in acidity and

alkalinity, electrophily and nucleophily or, furthermore, in positive and
negativeelectrical characteristics.We will seelater, how important dualism
is for the different groups of constituentsand how, without this dualistic
concept, it would be di-fficultto understandthe roles of most of theseconstituentsin biologt.
In part, as a consequenceof the separationof the constituentsinto
two groups,dualism can be observedeasilyin the hierarchic organization
of higher entities. Dualism appearsin the relationship between primary
and secondaryparts, the fust having a more positive character than the
second.The study of cancermanifestationsunder this dualistic aspecthas
beenhighly rewarding and is the subjectof the following chapters.
In a more concretestep, the dualisticconcepthas provided new understandingof abnormality.
Normal and Abnormal
A normal entity can be conceivedof as one which is able to maintain
its constantswith their characteristicvalues, rhythms and intensitiesby
meansof the alternateoperationof homotropic and heterotropicforces.A
normal entity, thus, can be definedas one havingconstantswithin the limits
that statisticallycharacterizethis particular kind of entity. we can define
the abnormal entity as one in which a constant'scharacteristics-average
value,rhythm, intensity-are altered.It is alteration,without completeloss
of the characteristicsof constants,that differentiatesabnormality from
death.In death the constantsthemselvesare irremediablylost. This definition also distinguishesabnormal from physiologicalmanifestations.In the
physiologicalmanifestation,oscillatorymovementpersistsand only its intensity is influenced,usually becomingexaggerated.
As expectedfrom the dualistic concept, abnormal changescan take
place in either of two opposite directions and this is a significant fact of
abnormality.The two possibilitiesare inherent in the oscillatory balance
characterizingthe constant itself. It is the offbalance,resulting from the
predominanceof one of the coupledfactorsover its antagonist,
exaggerated
which leads to the abnormal. Persistentpredominanceof one factor abnormally affects,and even suppresses,
normal oscillatoryrhythm.
For each normal condition, then, two oppositeabnormalitiesare possible.By relating the abnormalcondition to one or more alteredconcepts,
and the alterationin each constantto one of the dual changespossible,a
new systematizedanalysisof the abnormal becomesfeasible.The large
number of constantswhich composeeach entity and which can become

BtoLOctc^L ENTTTTES /

37

abnormalhelp not only to explain the great varieg of abnormalitiesbut

also offer a meansof obtaininganalyticalpicturesof disease.
It is with this approachthat wc havetried to study pathologicalconditions. with specialemphasison cancer.This study is presentedin the following pages.

CHAPTER

CANCERAS AN ORGANIZED
CONDITION

il

.llN rue coNcEpr now most widely accepted,cancer is consideredto be

the result of abnormal changeswithin cells. Although it is admitted that the
diseasemay have different etiologies,it is the cell which is regarded as the
pathogenic entity. A group of specific changesin the cells is believed to
rcpresent the fundamental abnormality.
In today'sprevailingoutlook, differencesbetweentumors are attributed
to the multiple secondarycharacteristics
presentin the diseasedcells along
with a primary specificanomaly. The complex clinical manifestationsof
cancerare further explainedin termsof the relationshipbetweencancerous
cells, as pathogenicentities,and the whole organism. Clinical evolution,
from local innocuous process to lethal disease is related to anatomical
spread of cancerous cells from their original site. Abnormal metabolic
changcs seen in the organism are believed to result from the influence
exertcd by functional abnormalitiesof the cancerouscells. In a still narrower view, canceris consideredto be the result of abnormalityof a single
s-1-cificfunction of the cell-its gro\rnh. Qualitatively and quantitatively,
abnormalgrowth has becn consideredto be the capital factor in the pathogcncsisof the discase.(292)
In contrast to this classicalview, our studies have lc-d us to regard
ccnc!'r as somethingother than an abnormality limited to the cell alone.
As we have seen,the organismis a conrplcxhierarchicorganizationof
different biological entities.We soueht to detc'rminen'here cancer fits in
this complexorganization.
Can cancerrvith is manifc'stations
and is evolution be L*tter understoodif s1'stematized
in ;.rccordirnce
s'ith the hierarchical
organizationof the organism?Cm both nunifcst.rti(rnsilnd r'r'olution be
rclutednot alone to a cellular abnormaliry"
but rathc.rto :l pR)gressive
par38

CANCER ^S

^N

ORGANIZEDCONDITION

39

ticipation in the diseaseof the different hierarchiclevels of the organism?

we have found that suchparticipationcannotbe analyzedreadily in the
advancedcanceroussubject with so many and such varied manifestations
of the diseasealready present.Similarly, incipient caseswith a paucity of
clinical manifestationsare not ideal for the purpose.It was only by following the successiveappearanceof manifestationsduring the evolution of
cancerthat their relationshipto the level of hierarchicorganizationinvolved
could be clearly seen.
Identificationof the level involved at each point in the developmentof
cancerwas greatly facilitatedby conceptuallyseparatingthe clinical evolution of the diseaseinto a seriesof successivephasesand identifying the
changeswhich characterizethe passagefrom one phase to the next.
we have chosento call thesephasesprecanccrous,
noninvasive,invasive, painful, preterminal and terminal. we will briefly identify them and
their salient features here.
Precancerous Phase
In the precancerousphase, the diseaseis not clinically apparent.Yet
this phasehas beenrecognizedas pathogenicin experimentalcarcinogenesis
and its characteristicchangesalso have been identifiedin human subjects.
Morphologicalchanges-abnormalitiesin size and form--<an be observed
in the chromosomes.These changesare not identifiedas related to cancer
in human subjectsexceptwhere multiple centersof cancerizationare found
(as in the stomach,for instance). (Note 1) Thesechromosomalabnormalities can be considered to be precancerouslesions, since experimental
carcinogenesis
has indicatedthat thesechangespreccdethc appearance
of
cancerous
cells.In termsof hierarchicorganization,
then, the precancerous
phasecan be consideredto be limited to the subnuclearlevels.
Noninvasive Phase
In the noninvasivephase,also known as "cancer-in-situ,"abnormal
intra-epithelialcells are present.The abnormality involves two changes.
One, morphological,affectsthe nucleus;the other affectsarrangementof
the cells in the epithelium. The abnormal changesin the nucleus in this
phasehavebecn widely studiedin exfoliativccytology.(Note 2)
Abnormalityin this phaseappearsto be limited entirelyto the nucleus.
The cells continue to have an almost normally differentiatedcytoplasm,a
fact which originally led to the descriptionof this phaseas "cancer of differentiatedcells." Besidesthe nuclear changes,cells in this phase show,
histologically,an anarchicarrangementdifferent from the regular disposi-

40

REsEARcHrN pHysrop^THoLocy

tion which is one of the basic characteristicof the epithelium. Since the
regularrelationshipbetweenthe cells forming epitheliumcan be attributed
to dipolarity, the anarchicdispositionseenin this phaseof cancer can be
ascribedto loss of cellular dipolarity.
cancer-in-situ, in terms of hierarchic organization,would appear to
involve the level of the nuclei, and the noninvasiveness.
characteristicof
this phase,persistsas long as the "cancerous"abnormatityremainslimited
to this level, that is, as long as the cytoplasmof cells remains apparently
unaffected.
Invasive Phose
This phaseis characterizedby irregularproliferationof cells and penetration into neighboringtissues.To the anarchicarrangementsnoted in the
noninvasivephase, now has been added exaggeratedgrowth. And the
changeof a noninvasivecancer into an invasiveone can be consideredto
result solely from the addition of the new factor of abnormal growth. The
invadingcells will persistonly if, concurrently,there is a loss of the defense
mechanismof the invaded tissues,as will be seen later.
Studiesof invading cells have revealed,in this phase,an anomaly no
longer limited to the nucleusbut now encompassing
the cytoplasmas well.
Exfoliative cytology has shown an abnormal and rapidly disintegrating
cytoplasmand this has servedas an important diagnosticcriterion. From
the point of view of organization,it can be said that, with the participation
of the cytoplasm,the diseasehas progressedfrom the nuclear to the cellular level.
Painlul Phase
Pain is the principal clinical manifestationcharacterizingthe next phase
of the disease.As we shall explain in greaterdetail later, pain arisesfrom
changesin the pH of the intercellularfluid that bathessensorialnerve endings.For the moment, we can remark that biochemicalchangesnow occur
outsidethe cells, and, with the participationof interstitial formations,the
diseasehas progressedto the tissularlevel.
Preterminal and Terminal Phases
In the next stage,the preterminal,biochemicalchangesaffect the function of variousorganswhich may or may not in themselvescontain cancerous cells. while some changesin function may be seen even before this
preterminalphase,now, there is manifestimpairment.And, while the invasionof an organ by cancerousmassesis a factor precipitatingthe func-

c A N c E R A S A N

O R C A N I Z E DC O N D T T I O N

4 l

tionalchanges,
invasionis not indispensable.
Abnormatbiochemicalchanges
leadingto seriousfunctionalimpairmentsare seenin organsentirelyexempt
from tumor masses.
with further progress of cancer, metabolic functions that are systemicallyimportant becomeabnormal,Later, we will analyzr,in detail these
changeswhich affect the whole organismprofoundly.For the moment we
want only to note that, with thesechangescancerpassesfrom the clinically
preterminal to the terminal phose.
In the light of this systematization,cancer then appears to progress
clinically in organized fashion as it passesfrom the relatively innocuous
nuclearnoninvasivecancer-in-situto a lethal systemicdisease,the progress
being marked by the successiveparticipationof difterent hierarchic levels
of the organization. Table I sums this up.
T,fSLP

Organizational
Level
Subnuclear
Nuclear
Cellular
Tissular
Organic
Systemic

PhysiopathologicalChanges
Gene and chromosomeanomalies
Nuclearanomaliesand atypical
cellular arrangemenls
Atypical growth
Local pH changes
Organic metabolicchanges
Systemicmetabolicchanges

Ctinical phase
precancerous
Noninvasivecancer
Invasivecancer
Painful cancer
Preterminalcancer
Terminal cancer

By extrapolation,a similar progressiveparticipationof hierarchic entities can be conceivedof below morphologicallyrecognizablelevels.This

would permit us, as a working hypothesis,to attribute the pathogenesis
of
cancerto abnormalitiesin nucleo-proteinsor, even lower in the scale,to
abnormalitiesin histonesor alkaline amino acids.(Note 3)
This concept----ofprogressiveparticipation of successivehierarchic
levels in cancer-<ontrasts sharply with the view generally held today
which placesthe entire burden of anomalyon the cancerouscell itself. The
classicalconcepthas led to the currently prevailingaU-or-nothingapproach
in which therapeuticattemptsare directedto the cancerouscells as the only
avenuefor controlling the diseaseat any moment of its evolution. Under
our hierarchicconcept,therapeuticpossibilitiescan be extendedbeyond
the cancerouscell.
These considerationsraise the questionof the relative importance of
themultiplechangeswhichoccurin cancer.Subnuclear
and nuclearchanges
are of relatively little importanceas long as there is no progressof disease
beyondtheselevels.Corroborationfor this can be found in the great num-

42

RESEARcH rN pHysropATHoLocy

ber of casesin which cancer-in-situ

cellsare noted in an organ,yet clinical
cancerdoesnot follow. In our concept,the changeswhich occur at levels
abovethe nucleararc critical in the evolutionof the diseaseand, as such,
are the important pathogenicfactors. On the other hand, as we shall see
later,changesat higherlevclssimilar to thoseencounteredin cancermay
occur independentlyof cancer,and without a sequenceof changesat lower
levels.It is only when changesat higher levelsappearin proper sequence,
affectingalready abnormal entitiesof the lower levels,that clinical cancer
resultsand the malignancymovesrelentlesslyfrom the noninvasivecancerin-situ to the terminal phase.
This concept,then, focusesattentionon all changesoccurring at difterent hierarchiclevels of the organizationrather than on those in the cell
alone. It emphasizesthe importanceof the relative independencewhich
existsbetweenthe different hierarchiclevels,an independencewhich governstheir participationin the complexconditionwhich is cancer.
From the therapeuticstandpoint,then,it seemslogicalto supposethat,
if the progressiveparticipation of successivelevels can be interrupted,
many if not all of the noxiousmanifestationsand the courseof cancercan
be favorablyinfluenced.In view of this, it has beenessential,first, to obtain
more information about the cancermanifestationswhich are added as the
diseasetakes its hierarchicallyprogressivecourse and about the mechanisms that account for thesemanifestations.

CHAPTER

DUALISIU

'fhe
/ \\s wE HAVEotssERvED
ABOVE,
dualismprevailsin nature.
concept
of an oscillatorydynamic balance-the result of alternateopcration of opposedforccs-has beenof specialvalue in the study of most of the physiological phenomcna.Over the yearswe have alsoconstantlyobservedthat in
most physiopathological
manifestations,
dualisticpatternscan be recognized.
This dualisticpathogenicconccpthas helpcd to guide our study of disease.
In cancer, it has permitted bettcr understandingof many processesand
manifestations.It has also servedas a basis for our attemptsto influence
cancer and other conditionstherapeutically.
It was in the study of pain
that, initially, we found clear evidenceof pathogcnicdualism.

PAIN
Many years ago, during expcrimentswith an alcoholic cxtract of human placentaas a therapeuticagentin terminalcancercases,a curiouseffect was observed.In somepatientswith painful lesions,administrationof
the preparationresultedin a decreasein the intensityof pain and even in its
disappearance
within a few minutes,with rclicf usuallylastingfor hours. In
other cases,however,there was an oppositeeffect;pain increasedin intensity within a few minutesafter an injcction.In somesubjects,the exacerbation was so great and pain becamcso unbearablcthat the experimental
treatmenthad to be discontinuedquickly. In severalcasesin which the
preparationwas used in progressivelylarger doscs,another notcworthy effect was observed: after the first injections,pain decreasedand even disappearedfor severaldays, only to have a new pain arise as treatment
continued.This new pain becamemore intcnseaftcr each injcctionso that
the therapyhad to be discontinued.
Patientsclearly recognizedthe differpains.
encebetween
The new one frequentlyhad a burningcharacter.
43

44

REsEARcH rN pHysropATHoLocy

Thus, it became apparent that one substancecould increasepain in

some subjectsand alleviateit in others,and could even alter the nature of
the pain in the same subject. Pain, as demonstratedby antagonisticresponsesto a single agent, thus appearedto have a dual nature. Our immediateproblem was to investigatethis and its significance.
Physiological and Pathological Pain
In discussingthe sensationof pain,mostauthorshave found it necessary
to distinguishbetweendifferenttypesof pain. Somehave classifiedpain as:
l) spontaneous,
or 2) provoked,accordingto its modeof induction.others
have definedpain accordingto its site of origin and quality as: r) superficial or cutaneous,and 2) deep visceralor somatic.Superficialpain from
skin and mucousmembranesnear body orificeshas been describedas bright
or burning in quality, while deep visceral or somatic pain arising from
mesenchymalstructures,certain mucous membranesand viscerahas been
describedas diffuse and aching in quality. In many respects,attemptsto
define and classifypain in theseterms have servedto confuse rather than
to clarify the problem.
That differenttypesof painsdo exist is an observationbasedupon common experience.For example,when a stimulusof sufficientintensityis applied to the skin for an adequateperiod of time, a sensationof pain is
induced.This pain disappearsrapidly when the stimulusis removed.But
if the stimulushas been of such intensityand duration as to producetissue damage,an after-pain may recur spontaneouslysome time after the
stimulushas ceased.The originalpain servesas a warningthat the tissues
are endangered.The after-pain,however,cannot be consideredas a direct
effect of the applicationof an externalstimulus,but is rather a manifestation of true tissuedamage.
As a first step, two categoriesof pain-one induced in normal tissue
by external intervention,the other appearingas a pathologicalmanifestation of an existinglesion-were established.we called the first, which is a
normal sensorialsensation,"physiological"or "sensorial" pain. The second, a symptomof an abnormallocal condition, was called "pathological"
or "symptomatic"pain. This separationhelpedto eliminatediscrepancies
otherwiseencountered
in the studyof pain,discrepancies
which resultwhen
two entirelydifferentmanifestations,
one sensorialand the other symptomatic, are studied under the same heading and investigatedby the same
methods.
Pain may be induced in damagedtissuesby various stimuli not of
sufficientintensityto arousepain sensations
in normal tissues.This sensi-

DUALTSM

45

tivity constitutesan abnormal responseof tissuesthat have undergone

pathological
changes.Inflammatory,traumatic,circulatory,neoplasticor
otherpathologicalchangessimilarly may bring about either spontaneous
painor an abnormaldegreeof sensitivityof the inyolved tissuesto external
stimuli.
There are, therefore,two gcneraltypes of pain which are biologically
different.Thc first is a direct responseof normal tissuesto external stimuli
whichservesas a warning of danger.The organismreacts to this type of
painby seekingto run or to fight. The secondtype of pain arisesas a consequence
of tissuedamage or diseaseto which the body respondsby attemptingto put the injured area at rest. This secondtype of pain, whether
spontaneous
or provoked,superficialor deep,is biologicallydifterent from
the first pain experiencedfollowing the applicationof sufficientlyintense
externalstimuli to normal tissues.
For purposesof further study, it would appear to be advantageousto
distinguish
betweenphysiological
or sensorialpain which is the responseof
normaltissuesto noxiousexternalstimuli,and pathological
or symptomatic
painwhich is a manifestationof abnormaltissues.In the study of pain in
all its aspects,it is necessaryto keep this distinctionin mind. While various
investigativemethods have furnished data concerningphysiologicalpain,
the information thus obtainedis of very limited value when applied to the
problemof pathologicalpain. However,it is pathologicalpain which constitutesthe vital clinical problem, physiologicalpain being of concern in
me'dicine
primarily in the field of anaesthesia.
(Note I )
Dualism in PathologicalPain
It wasonly in pathologicalpain that a dual characterwas encountered.
For
onething, it was noted that in some patientswith chronic pain-associated
*ith tumors, arthritis or other conditions-the pain intensitywas not constant.In many of thesepatients,variationsin pain intensitycould be seen
to follow a pattern. Although the variations usually are referred to as
"spontaneous,"we could show that they were related to the time of day.
Furthermore,the variations were not the same for all patients. In one
group, pain was severein the morning and diminished toward evening,
whilein anothergroup, little or no pain was felt in the morning and exacerbationsoccurredin the evening.
The intake of food also had a dual influence.In some patients with
tumorsfar removedfrom the gastro-intestinal
tract, pain was increasedby
catingwhile in otherspain decreased.Patientsthemselvesoften recognized
this relationship and many whose pain was increasedwith the intake of

46

nEsEARcH rN pHyslopATHoLocy

food refusedto eat for fear of aggravatingtheir suffering,while thoseof the

other group wanted to eat wheneverpain was scverein order to reduceits
intensity.
These observationson the influenceof time of day and intake of food
pain
on
led to the study of the acid-basebalance of the body since this
balanceis known to be influencedby the same two factors-time of day
and food intake.
In a preliminary study we considereda specialaspectof the acid-base
balanceof the body, that is, the severalmechanismsinvolved and their
possibleinterventionin the changeof pain intensity.A study was made of
blood pH, titrimetric alkalinity,CO2 combiningpower, rclative chloride
distribution betweenerythrocytesand plasma, as well as urinary pH for
the indicationsthey furnish concerningthe acid-basebalance.The blood is
highly buffcred in order to avoid damage,through abnormal pH values,
to cells in generaland especiallyto those of the nervous system.Consequently, the variationsin the blood pH are as limited as possible.Alkaline
reserveand the chloridesrepartitionrepresentonly part systemsin the general acid-basebalance.Titrimetric alkalinity, correspondingto the sum of
ionized and nonionizcd constituents,furnish information of the broadest
scale of the acid-basebalance. Consequentlyit appears to be a higily
significantmeasurement.
Through the non-ionizedconstituents,it can vary
greatly without influencingblood pH. It reflectsthus otherwischidden
changesin the acid-basebalance.We could show that alone, among all the
variablefactorsof the blood acid-basebalance,total titrimetric alkalinity of
blood varies in parallel with the urinary pH. (Nore 2)
As an immediateresult of this research,it was possibleto utilize the
changesin the urinary pH as an indicationof the most importantvariations
occurringin the systemicacid-basebalance.This makes it possibleto use
changesin the urinary pH as an indicator of the relationship between
acid-basebalanceand variationsin pain intensity.
Pain and Acid-Base Balance Changes
When changesin pathologicalpain intcnsitywerc studiedin relationto
changesof the urinary pH, a correlationcould be establishedin the majority of cases.Two oppositekinds of relationshipwere observedwhen curves
of the variationsin pain intensitywerecomparedwith thoseof the urinary
pH.
with chronic pathological
Patientswho had experiencedpain associated
lesionsover prolongedperiodsof time were instructedto record carefully
the relativeintensityof their pain at regularintervals,such as every hour.

DUALTSM

47

No analgesicswcre administeredfor at least six hours before or during the

testperiod which was continued as long as possible,even for twenry-four
hours.Patientswere instructedto concentrateon a singlepainful area and
to estimate the degreeof pain intensity.They were told to consider an
averagedegreeof pain during each hour rather than momentarypeaks
during the observationperiod or the pain at the moment of recording. The
PTIN
Unbaroble
Very*vere
Severe
Moderole
Sl,9ht
None

pH
8.O
Unne
7.5
7.O
6.5
6.O
5.5
5.O

to

tz

'6
,.1,!

tB

22

24

Flc. 6. A pain pa!lern is recognizedby comparing the concomitant changes present

in the curves of pain intensity with those of the urinary pH, The parallel variations
of the two curves indicate an olkaline patter,t with the pain more intensive when thc
urine is more alkaline, as seen in a case of carcinoma of the coloq with painful abdominal mass.

degreewas recordedin relativeterms of no pain, slight, moderate,severe,

very severe and unbearable,or as figuresfrom 0 to 10.
Urine specimenswere obtainedeach hour immediately after the pain
intensityobservationswere recordedand the pH was determinedelectrometrically. Two curves-for the hourly variations in pain intensity and
for urine pH-were then plotted.
Two distinct types of correlationswere found. In the first, the two
curvesparalleled each other, the pain being more intensewhen the urine
pH was higher, and less severewhen the pH was lower. (Figs. 6 and 7)
Becausethe maximal pain of this type of correlationis associatedwith a
changetoward alkalinity, this was caUedan alkaline pattern of pain,

48

RESEARCH IN

PHYSIOPATHOLOGY

PAIN
Unbcorobh
Vcry Scvcn
Scwrc
Moduote
S,,9ht
l{ona
0H
Utina

6.2
6.O
58
5.6
5

5.2
5.
l'lour

Fto. 7. The alkaline paltern ol pain in which the concomitant variations in the curves
of pain intensity and urinary pH are parallel, seen iD a case of arthritis.

In the secondrype of correlation,the two curvesvaried inversely,pain

being most marked when the urine was most acid, and least so when the
urine was most alkaline.(figs. 8 and 9) This secondtype was called acid
pattern of pain becauseof its associationwith a changetoward acidity.
Consideringthe highly subjectivenatureof pain, the inconsistencies
which
occur are minor. Fig. l0 showsthesecurvesfollowedduring days.
The correlationbetweenthe pain intensityand urine pH curves arc
relativerather than absolute.The generallevel of urine pH apparently
m,A/
Ltibcorobfe
Very *ere
Scvcrc
Itlodatota
Sl,ght
Nona

pH
7.5
Uttna
7-O
6.5
6.O
5.5
5.O

Hour

Ftc. 8. An ocid pain patlern is seenin a caseof carcinoma of the prostate with meta.
s t a t i c b o n e l e s i o n s ,i n w h i c h t h e c o n c o m i t a n tv a r i a t i o n so f t h e c u r v e s o f p a i n i n t e n 5ity and urinary pH are divergent.

DU^LrsM

49

dependsupon other factors. Consequently,only the fluctuations of the

hydrogenion concentration,rather than the absolutelevels,are considered
in this relationshipto pain.
Changesin pain intensitywere found to have a similar dualisticcorrelationwith other factors as well as the acid-base!3l3nss-v/ith potassium
contentof blood serum, for example.Studieswere made to compare the
concomitantchangesin pain intensityand in potassiumcontent of blood
serurn.In severalcasesthe acid and alkaline patternsof thesepains were
determinedthrough the relationshipto urinary pH variations.At different
PAIN
U^Eorablc
Very fucn
Sevtre
Hoocrcle
Sl,ght
l{ona

_N_
Utlte

8.O
7.5
7.O
6.5
6.O
5.5
llou I

Ftc. 9. An acid pattern ol pain recognizedby the divergent variations of the curves
o f p a i n i n t e n s i t y a n d u r i n a r y p H i s s e e ni n a c a s eo f p h a n t o m l i m b .

moments,especiallywhen the pain was markedly different in intensity,

blood was obtained by finger puncture,and collectedin glass capillaries.
After clotting, the serum was immediatelyseparatedand the potassium
contentmeasuredusing a flame photometer.Curvesof the valuesof pain
intensityat these moments, establishedby the method previously rnentioned,and of the concomitantK+ contentwerecompared.Figs. I I and 12
showthe two curves in two casesin which, the two pain patterns-acid
andalkaline, were primarily recognized.The caseswith high serum potassiumvalues and with parallel changesbetweenthe two curves-pain intensityand K+ content-were seento correspondto alkalinepain pattern;
the other cases with less serum potassiumand opposite variations of the
two curyes were seento correspondto the acid pain pattern.
We extended the study of the pain pattern, as revealedby concomitant

50

nESEARcHrN pHysropATHoLocy

variationsof pain intensityandbody acid-base

balance,from cancercases,
in which a frank dualismhad beenseen,to other painful conditions.It
wasinteresting
pain can haveone or the
to note that,in manyconditions,
otherpatternbut thereare someconditionsin which only one patternis
consistentlyfound. Pain followingtraumaof any kind-the pain of post-

C o r c r n o m oo f R e c t u m
|9 4 3

+++ +
+ + +
c
++
o
o+

8.2
74
--

6'2
54
46
C o u r t e s yo f D r .M o r r oR o g n o n i

Fto. 10. The pain pattern can be recognizedalso through the characteristicopposite
variations of the curves of pain intensity and urinary pH, followed during successive
days-instead of hours-as seen in the above curves of a case of cancer of the rectum. (Courtesy of Dr. Rognoni)

operative and accidental wounds, burns and fractures, for example-always has an alkaline pattern. This is also true for the pain of gallbladder
colic. In other conditions,eitherpatternmay be presentand must be determined by analysis,For instance,the pain of neuritis and simple headachehas an acid pattern in somecases,alkalinein others.
In rheumatoidarthritis,an alkalinepain is almost constantlyfound.
ln osteoarthritis,the pain is of an acid type. In arthritic patientsin whom

DUALISM

5I

this relationshipdid not seemto hold, it was possibleto recognizenot only

the existenceof both rheumatoidand osteoarthritisbut also to note that the
pain pattern, as shown by test, was relatedto the more painful condition.
we utilized the diagnosisof the type of pain presentas an indication of
the nature of painful processes.We will seelater how this correlationhas
been confirmed by therapeutic trials.
lo
I

Pain
i n t e n ist y

6
4
2

(a)
K+ in serum
in mEq
.2
.0
1

H o u r s
Frc. I l. Pain pailern and potassium in blood serum. The comparison of the concomitant changesin the curves of pain intensity and those of the amount of potassium
in blood serum shows parallel variations in a case with an alkaline pattern.

Two types of pain associatedwith two different conditionspresent in

the same individual have been found to occur more frequently than expected,althougbusuallynot simultaneouslyactive.
In most patientswith two or more anatomicallyseparatedpainful foci,
parallelvariationsoccurredbetweenthe curvesof the differentpains. Only
in occasionalcaseswere the rwo painsfound to vary simultaneouslybut in
oppositefashion, Their oppositepatternswere well describedby patients
who observed"the two pains act as if they were part of a balance;when
one goesup, the other goesdown, and the opposite."In Figure 13, the pain
curye of lesion A is seento vary inverselywith the curve of urinary pH,
while the pain curve of lesion B is parallel with the urinary pH curve. Thus,

52

RESEARCH IN

PHYSIOPATHOLOGY

t0

Pain
Intensity

1
7
o

(b)
1.0

K+ tn serun 58

i n dq

16
54
32
3

Hours
F t o . 1 2 . D i v e r g e n t v a r i a t i o n sb e t w e e np a i n i n t e n s i t yc h a n g e sa n d t h o s e o f t h e b l o o d
serum potassium in a case of acid pain pattern.

PAIN LESION A
YaTt
Sevlrl
'.odarota
Sllghl

Nonr
MIN LESION8
Vr"t S.v.r.'l
Stvrrr
I
Hod.rlt
I
srbhf
{
I
ttolr

n!__UntlE ?.o
6.5
60
3.5

Frc. 13. Pains. Acid and alkaline pains can co-exist on different lesions,as seen in a
patient with multiple osseous metastasesfrom breast carcinoma. Lesion A, which
c o r r e s p o n d st o a n a c i d p a t t e r n . s h o w s d i v e r g e n t v a r i a t i o n s b e t w e e n t h e c u r v e o f
u r i n a r y p H a n d t h a t o f i t s p a i n i n t e n s i t y ,w h i l e f o r l e s i o n B . w i t h a n a l k a l i n e p a t t e r n ,
t h e v a r i a t i o n so f t h e c u r v e so f p a i n i n t e n s i t ya n d o f u r i n a r y p H a r e p a r a l l e l .

I _H..1"*_

:" .

_,

l,lil{Si"Ss\jilJ,1,$s-\i1,,i"r".+r,,"

DU^LrsM /

53

the pain of lesionA is of an acid patternwhile that of lesionB is of alkaline pattern.

Also interestingto note is the persistence
of the samepatternfor pains
associated
with chronic conditions.We have headachepatients,for example,in whom, during the 20 yearssincewe first determinedthe patternof
pain, there has been no changesof pattern. In others,on the contrary,
PAIN
UnDmmbla
Itry Scverc
Slvero
Modctolc
Slghl
lVorp
pH
ul tnQ

HrPOr

7.O
6 .o
5 0
4.O
8

9
t
Houl

F t c . 1 4 . T h e c h a n g e si n d u c e d i n t h e p a i n i n t e n s i t yb y t h e a d m i n i s t r a t i o no f a n a c i d i fying agent indicate the pattern present. Pain with an acid pauern is intensified foll o w i n go r a l a d m i n i s t r a t i o no f 1 . 5 c c . o f . a 5 o % s o l . o f p h o s p h o r i ca c i d . U r i n a r y p H
changesreflect the induced systemic acidification.

changes
occur rapidly. In a caseof sciatica.we have seenrapid and frequentchangesin pattern,especiallyin responseto therapeuticmeasures.
Worthy of being noted is the correlationfound betweenvariationsin
painintensityand changesin the acid-basebalanceof the body even in
casesin which nerves are directly involved in lesionsand in which a
mechanicalpathogenesis
usuallyhas been accepted.This would indicate
thatthe chemicalfactor mentionedabovehas a role in the pathogenesis
of
paineven in thesecases.
Acidilying and Alkalizing Agents
We next demonstratedthe cause-effect
correlationbetweenacid-base
changes
and variationsin the intensityof pain. Administrationof acidifying or alkalizing substancescould induce the same changesin pain
intensityas those causedby spontaneous
variationsin the acid-basebalanceof the body.
The relationshipbetweenacid-basebalancechangcsand pain intensity
was thus investigatedby administeringstrong acidifying and alkalizing

54

RESEARcH rN PHYsropATHoLocy

agentsto patientsafter the type of pain-urinarypH correlationhad been

determined.Administeredorally, strong acidifyingsubstances,
such as
phosphoricacid, ammonium chloride or mono ammonium phosphate,
increasedthe severityof pain with an acid pattern,(Fig. 14) and reduced
the severityof one with an alkalinepattern.(FiS. 15)At the sametime, it
causeda loweringof the urine pH.

PAIN
Severe

Poo
JHt

Moderote
Slight
None

pH
Urine

7.O
6.5
6,O
5.5

Hours
Flc. 15. Pain with an alkaline pattern is relieved by oral administration of two doses
of 1.5 cc phosphoric acid (50Vo sol.). Urinary pH changes reflect the systemic acidification.

Sodium bicarbonate or ammonium acetate in quantities that alkalinized the urine increasedintensityof pain with an alkaline (Fig. I6) ^nd
diminishedintensityof pain with an acid pattern.(Fig. 17)
These changes of the systemic acid-basebalance induced by administration of strong acidifying or alkalizing agents explains how similar
changes,when they occur spontaneously,affect pain intensity. The effect

DUALISM
PAIN
Unbrorcble
Very Scvere

lVoHCOr

Sevcre
Moderote
S1 , 9 h ,
NonG
pH
Uttt'

/ s s

80

7 0
6 0
5.O
9
Houl

F t c . 1 6 . P a i n w i t h a n o l k a l i n ep a l t e r n i s i n t c n s i f i e db y o r a l a d n r i n i s t r a t i o no f 5 g r a m s
of sodium bicarbonate.urinary pH changesreflect the systcmic alkalinization.

may be to increaseor decreasethe intensity,dependingupon the pattcrn

of the existing pain. It is intercstingto mention that similar changesof the
systemicacid-basebalance,spontaneousor induccd by the administration
of acidifying or alkalizing agents,do not influenceeither the thresholdor
intensity of physiologicalpain. we have often utilized this rcsponseto
alkalizing or acidifying substancesas a method of recognizingthe acid or
alkalinepattern of pain. (Fig. I8 bis)
Dualism in Local pH Measurements
Since it had been observedthat a definite correlation exists between
the variations in pain intensity of abnormal foci and changesin the gen-

fllA/
U^bP'oroble
Wy Severe
Sevcrc
Modercle

NoHCOs

Slighl
None
PH
Unne

8.O
7O
6.O
a n
I

a
Houl

F t c , 1 7 . P a i n w i t h a n a c i d p a t t e r n i s r e l i e v e dh y o r a l a d m i n i s t r a t i o no f 5 g r a m s o f
s o d i u mb i c a r b o n a t e .U r i n a r y p H c h a n g e sr e f l e c tt h e s y s t c m i ca t k a l i n i z a t i o n .

56

xESE^RcHrN pHysropATHoLocy

eral reaction of the body, it was desirableto ascertainwhat changeswere

taking place within the abnormalfoci themselvesat the same time.
Patientswith easily accessiblesuperficiallesions,especiallytumors, in
which painful areascould be well localized,were employedin theseexperiments. The pattern of pain, acid or alkaline, was first determinedin the
manner previously described.Local pH determinationswere then perCo of Utcrut
45
55 05

Hours 35

unbccroblc
vcry rcvtra
t cv0ra
modorolo
rlight
nona

Co of Prortolr
f5

25

35

45

1 . 5g m r . A m o n i u m

55

05

15

25

9m3A
. monium
occloll

ocr tot.

r.5eT
A m o n i u mo c c l o l c

Go of U l c r u r
fo
20 30 4 0

oo

oo ro
l

unbcoroblc O . 5g m t . A m o n i u mC h l o r i d r
varyr

Co of 8r.ort
20
30 40

50

l g m . A m o n i u mC h l o r l d r

taYcra
nodcroh
I light
nona

Co of Ulcrur

Co of Uteruc

30

YaTy scvarc
stvcrS

40

50

00

ro

to

? o r c 4 p 5 0

3O drope phorphoric ocid

off. sol.
I
I
r7

modorolc
rllght
nonc
Ftc. l8bis. The responseof the pain of a lesion to an agent permits to identify the
acid or alkaline pattern present.The effect of an acidifying and alkalizing agent correspondsto an increaseor a decreasein pain intensity, according to the pain pattern
present.First row, left side-acid pattern, right side-alkaline; second row, left sideacid, right side-alkaline; third row. right side-alkaline, left side-acid pattern.

DUALTSM

57

formed. Specialglasselectrodest were used for this purpose and determinations were carried out employing a sensitivepH meter. The tip of the
electrodewas placed on the surface of the area to be testedif ulcerated,or
was introduced into the tissue to be tested through a small incision. In
reality this givesa measurementof the pH of the local interstitialfluid.
Urine pH and local pH determinationswere then performed at difterent
times correspondingwith spontaneousvariationsin the pain intensityexperiencedby the patient. Simultaneously,the pH values of normal tissue
areas and, when possible,of nonpainful tumor areas were determined.
Similar studieswere carried out after administrationof strong acidifying
and alkalizingagents.
Many difficultieswere encounteredboth in the choice of suitableclinical subjectsand in techniques.Neoplasmsproved to be the simplesttype
of painful abnormalprocessto employ.The neoplasmhad to be locatedin
a readily accessibleregion so that the electrodescould be introducedinto
an ulceratedarea or through small incisions.The patient had to be able
to very accuratelylocalize the area of pain since considerabledifferences
in pH valueswere found to exist in difterentparts of the same lesion.The
pain had to be superficiallylocalizedbecauseaccuratedeterminationsin
the depths were not possible.Finally, the complete cooperation of the
patientwas essential.
The data obtained for a patient with an ulcerated,profusely draining
carcinomaof the breast is recordedin TnsLe II. Pain was most intense
T,rsl.e II
Onse.nvrtroNsIN A Crse wlrn Alx.,rllxe P,rlx P,rrrenx

Treatment (oral)

Phosphoricacid
S O V o . 2c c .
Sodium bicarbonate
5 grams

Pain Intensity pH Urine

pH Normal
Tissues

pH Tumor

None
Moderate
Very severe

5.4
6.2
7.1

7.3
7.4

7.6
8.1
8.5

Slight

5.5

7.3

7.9

Unbearable

7.8

1 A
I .t+

8.8

when the acid-basebalance of the blood, as reflected in the urine pH

changes,was relatively most alkaline, and was less intensewhen the balancwas more acid. The pH valuesof the painful areasof the tumor in
* S u p p l i e db y H a r t m a n a n d B r a u n , P a r i s .

58

R E S E A R c Ht N p H y s r o p A T H o L o c y

this case showed considerablelability under the influence of spontaneous

changesin the general acid-basebalanceof the body, reaching a high of
8.5. At this time, the pain was very severe.The pain becameunbearable
following oral administrationof 5 grams of sodium bicarbonate,and the
pH within the sametumor area reached8.8. The pH of the normal tissues
in this case,even after the administrationof strong alkalizingagents,never
exceeded7.4, while the tumor pH was neverbelow 7.6.
The findings in a casewith acid pain are recordedin TenlE III. This
T,rsr-EIII
OnseRv.rrroNs
rN A C,r,se
wrrH Acro prrN prrrrnN
a=_

pH TUMOR
Treatment(oral)

Pain
Intensity
None
Moderate
Severe

Phosphoricacid
5 0 V o, 1 . 5 c c .
Unbearable
Sodiumbicarbonate
5 grams
None

p H r pH Normal Painful
Urine
Tissues i area

7.0 l

7.4

i 66 . 3r

Nonpainful area

5.8
5.3

t.)

5.8

7.2
7.1
6.9

5.0

t.-)

5.5

6.8

6.6

7.2

l.+

patient had an extensivesarcomaof the face which was not ulceratedbut

involved the skin. Pain was most intensewhen the blood titrimetric alkalinity was relatively low and less severewhen the alkalinity was higher.
The pH values in the tissuesof a painful area of the tumor were always
more acid than the values found in another nonpainful area. The pH of
the nonpainful area was slightly below that within normal tissues.Spontaneouschangesin the acid-basebalance of the body brought about a
reductionof the pH of the painful tumor area to 5.8, at which time the
patient reported severepain. The nonpainful tumor area had a pH of 6.9
at the same time. Following administrationof L5 cc. of 5o% phosphoric
acid, pain becameunbearablcand the pH within the tumor tissuesof the
painful area fell at oncc to 5.5. At the same time, the nonpainful tumor
areapH was 6.8 and the normaltissuepH was 7.3.
Similar results were obtained in other casesand lcd to the following
conclusions:
The pH valuesof the interstitialfluid of painful lesionsstudied
in vivo differ from thoseof normal tissues,the hydrogenion concentration
being eithcr higher or lower than that of normal tissuesof the same individual. Thc pH of painful abnormal tissuesis much more labile and

DU^LrsM

59

extremelysensitiveto generalbody pH changes.Spontancousor induced

variations in the acid-basebalance of the body produce slight changesor
none at all in the pH of the interstitialfluid in normal tissues,but give rise
to more pronouncedchangesof the pH within painful pathologicaltissucs.
Through this research,we have thus been able to conncct pathological
pain to the pH of interstitial fluid of painful lesions.A further connection
could be made with the richnessof the interstitialfluids in potassium.In
the alkaline pattern of pain, more potassiumwas found in the interstitial
fluids. The presencein thesefluids of potassium-the cation of the cytoplasma-in higber amountsthan in normal conditionswas scen to induce
pain. The subcutaneousadministrationof potassiumcompoundswas secn
to be painful, while similar saltsof sodium werc well tolcrated.As a local
acidosiswas found in lesionshaving an acid pain pattern, a local alkalosis
and an increasein potassiumcontcnt appcarcdin those with an alkaline
pain pattern. The intensity of pain in this case was found to be proportionate to the degrec of abnormal deviation of thc local pH and to the
abnormalamountsof potassiumin the interstitialfluids. In addition to
variationsin the acid-basebalanceof the body, variationsin the amount
of serum potassiumappear able to alter pain intensityin caseswith an
alkalinepattern.This correlationis further explainedbelow by the place
of potassiumin the organization.
O.rido-red uction Potential
Differencesindicatingthe samedualismwere also found in other manifestations
in painfullesions.The oxido-reduction
potentill was measuredin
tumorswith pain. Patientschosenwere thosewith easily accessiblesuperficial tumors in wbich painful areas could be localized. Platinum needle
electrodeswere introducedin the painful areasthrough small incisionsor,
if the lesionswere ulceratedin the lesionsthemselves.
The measurements
of thepotentialpresentweremadeusinga BeckmanpH meter.As had been
thecasefor the pH, it appeared
importantthat thc patientbe ableto indicate
clearlythe painful areas.Becauseexactlocationof theseareasin the depths
was almost impossible,superficiallesionsusually were chosen for these
determinations.In general,in lesionswith an alkaline pattern, the measurementsshowedhigh values (such as from +100 to +350 millivolts)
whilein lesionswith pain of the acid pattern,the valueswere low (such as
-2 to -15 millivolts).

60

REsEARcHrN pHysropATHoLocy

Abnormal Substances
changes in the local pH, like changesin acid and alkaline patterns,
could be related to the appearanceand subsequentaccumulationof difterent substancesin the lesion. Increasedconcentrationof lactic acid was
found in the interstitialfluids of acid pattern lesions,while increasedconcentrationsof sodium and especiallypotassiumions were found in the
interstitialfluid of lesionswith an alkalinepattern.
Processesleading to a local acidosisare known and ascribed to the
well-known anoxybiotic metabolismof carbohydrates.They result from
lack of the "respiratory" oxybiotic phase of carbohydrate metabolism,
with the consequentconversionof pyruvic acid into lactic acid. only part
of the lactic acid is changedinto glycogenthrough the pasteur-Myerhoff
reaction,and an accumulationof lactic acid results.While the presenceof
lactic acid is well known, its presencehas not previously been related to
pain or other manifestations.
We could showthis correlationin somecases.
(Note 3 )
The scientificliteratureofreredno information concerningthe appearance of alkaline compounds.We were able to establishthat the presence
of sodium ions coincideswith another anomaly found in theselesions-a
high fixationof chloridesin the lesionsthemselves.
Valuesof 1600 mgr. of
cl., or even higher, per 100 gms. of wet tissuewere found in theselesions
insteadof about 400 mgr./ 100 gms. measuredin normal tissues.The local
alkalosisand the resultingalkaline pain pattern could thus be correlated
with an abnormal sodiumchloride metabolismat the tissularlevel in which
with chloride ions fixed by the cells, sodium ions remain free to combine
with carbonate anions and form alkaline compounds. If the abnormal
NaCl metabolismoccurs in the interstitial fluid, the subsequentalka.losis
inducespain. We will seelater how abnormal NaCl metabolismalso takes
place at other levelsof the organization.In thesecasesof alkaline pattern
of pain, the fact that abnormalamountsof sodium ions still enter the cells
will result in a loss by thesecells of potassiumwhich will accumulatein
the pericellularfluids,and form alkalinecompounds.
An immediateconclusionthat could be drawn from these studieswas
that there is a definitedualismin pathogenesis
of pain originatingin abnormal tissuesand that the two pain patternsevidencedin lesionswith acidosis
or alkalosisof the interstitialfluids indicatethat processesof two opposite
naturesgo on at the tissularlevel.

DU^LtsM

OTHER ACID AND ALKALINE

6l

SYMPTOMS

using the same method of investigationas in the study of pain, a relationship betweenvariationsin intensityof certain other symptomsand
variationsin urinary pH could be found. Acid and alkaline patternsof
itching could be recognized.The samepatternscould be found for vertigo
and impairedhearing.Among psychiatricmanifestations,
manicdepressive
statesshowedchangesthat could be relatedto acid-base
variations.
The correlationbetweendyspncaand acid-base
variationswas contrary
to what we expected.classically,dyspneais believedto be associated
only
with an increaseof acidityof blood. However,both acid and alkalinepatterns of dyspnea were observed,and could be related more directly to
changesat the tissuelevel.We will rcviewherethe pathogenesis
of itching,
vertigo,dyspnea,and other conditions,undcr this dualisticaspect.
Itching
The problem of itchingis interestingfrom other than the therapeuticpoint
of view, sincelittle is known regardingthe natureand causeof this condition. Various hypotheses
have been offeredbut have failed to explain its
pathogenesis.
our interestin itching originatedduring the study of pain
and led to an hypothesiswhich allowsus to approachthe problemfrom a
new angle. By analogy with pain, we separateditching into two types:
physiological,
as the responseof normal tissuesto externalstimuli; and
pathological.as a sensationarising within diseasedor damagedtissues.
Ticklingthe normalskin or evenstimulatingit throughheat.colcl,etc.,may
causa sensationof itchingand lead to scratching,Certainmucousmembranes,such as thoseof the nose,and the skin aroundnaturalorifices,arc
especially
sensitiveto such stimulation.This type of itching,as a response
of normal tissuesis phy,siological
or sensoriul.
Under pathologicalconditionshowever,the skin, mucousmembranes
andother formationsmay itch without externalstimulationor in response
to stimuli which ordinarilydo not producethis sensation.
The itching then
can be consideredas a manifestation
of diseasedor damagedtissuesand,
as such can be describedas pathological.Just as does pathologicalpain.
pathologicalitching representsa symptom rclated to abnormal changes
afreadypresent.(Note 4)
In spite of their relative independence,
the fact that similar fundamentalmechanismsare involved in the productionof itching and pain
explainscertain characteristics
they have in common. Like pathological

62

R E S E A R c Hr N p H y s I o p A T H o L o c y

pain, pathologicalitching varies in intensitywith the time of day. patients

with chronic pruritusare awareof this,some have more itching in the morning, others experienceexacerbationsat night. The same dualism is seen
with intake of food. For thesereasons,the relationshipbetweenchanges
in acid-basebalanceof the body, as reflectedin the urinary pH, and variations in intensityof itching was investigated.
Patientswith long standingpruritus associated
with a variety of chronic
skin conditionswere studied.They were askedto note over periods of six
to twelve hours the changesin itching intensity.Evaluation of the changes
was made by the patientsthemselves,using a seriesof qualificationssuch
as none, slight, moderate,severe,very severeand unbearable,or a scale
rangingfrom 0 to 10. They were instructedto considerthe averageinten-

lTCHING

Very Severe
Seyere
Moderote
Slight
None
PH
Urine

5.5

6.O
5.5
5.O
t

4
Hours

Ftc. 20. Thc alkaline puttern ol itching is recognized i n a c a s e o f s e n i l e v u l v a r

p r u r i t u s ,t h r o u g h p a r a l l e l v a r i a t i o n so f t h e c u r v e s o f t h e intensity of the itching and
of tbe urinary pH.

DUALTSM

63

sity of their itching for each hour, rather than to indicate the maximum
intensity at the exact time of recording. voided urine specimenswere
obtained at the end of each hour. The pH of the urine specimenswas determined electrometrically.A graph was plottcd to compare thc variations
in the subjectivedata furnishedby the paticntwith the concomitanthourly
changesin the pH of the urine. It was usuallynecessaryto repeat the test
severaltimes before the patient appearedable to satisfactorilyevaluatethe
changesin the intensity of the symptomfor hour-long periods rather than

ITCH IN G

VerySeyere
Severe
Moderote
Slight
IVone

fi_

Urine

7.O
6.5
6.O
5.5

Hour
FIc. 21. An acid patlern ol itching is recognizedin a caseof pruritus ani through the
divergencein the concomitant variations of the curves of itching intensity and urinary pH.

for just the moment of recording. (we also tried to judge the intensity of
itching through the frequency,intensityand duration of scratching,as
notedby an observer,but without success.)Fiftecn patientswere studied
and, becauseof the limited number, the resultsare presentedas merely
preliminary.

64

x E S E A R c Ht N p H y s l o p A T H o L o c y

In four cases,the curvesof itching and urinary pH did not show any
definitecorrelationeven after repeatedtests.of the remainingI I cases,7
showed a distinct parallelismof the two curves, Md in the other 4, an
inverserelationshipbetweencurveswas apparent.The graphs obtained in
two characteristiccasesare presentedhere. (Frgs. 20 and 2l )

Severe
lvloderole
Slf9ht

HsPOe

None

pH
Urine

7.O
6.5
6.O
5.5
5.O

Hours
F t c . 2 2 . A d m i n i s t r a t i o no f p h o s p h o r i ca c i d - 1 . 5 c c p h o s p h o r i ca c i d , s o l . 5 0 c Z - i n d u c e s t o g e t h e rw i t h a n a c i d i f i c a t i o no f t h e u r i n e , a n i n c r e a s ei n t h e i n t e n s i t vo f t h e
itching with an acid pattern.

There is a distinctparallelismbetweenthe two curves in Figure 20.

indicatingthat itchingwas more intensewhen the urine was relativelymore
alkaline,and slightor absentwhen the urine was more acid. We have consideredthis as an "alkaline pattern" of itching in accordanccwith the
designationfor pain, An inverserelationshipbetweenthe two curves is
seenin Figure 21. ln this case,itching was more intensewhen the urine

DU^LtsM

65

\\'asmore acid, and less severewhen the urine was more alkaline.This
representsan acid pattern.
The effect of a strong acidifying agent,phosphoricacid, in caseswith
acid and alkalineitching,is illustratedin Figures22 and 23. The intensity
of the alkalineitchingin the first casewas reducedby the acidifyingaction
of phosphoricacid, while the acid itching of the secondcasewas intensified. In Figure 24, the responseof a patient with alkaline itching to the
administrationof sodium bicarbonateis shown. The intensityof itching
wasgreatly increasedafter the alkalizingagent was given.

ITCHING
Seyere

IH'Po'

Moderote
Slight
None

pH
Urine

7.O
5.5
6.O
5.5
4
Hours

F l c . 2 3 . l t c h i n g w i t h a n a l k a l i n ep a t t e r n i s r e d u c e di n i n t e n s i t yb y t h e a d m i n i s t r a t i o n
o f a n a c i d i f y i n g a g e n t . I n t h e a b o v e e x a m p l e ,t w o d o s e s ,e a c h o f t . 5 c c p h o s p h o r i c
a c i d( 5 0 % ) , w e r e n e c e s s a r iyn o r d e r t o o b t a i n t h i s e f f e c t .

The fact that both pathologicalpain and pathologicalitching undergo

thesamechangesin intensityrelatedto the generalacid-basebalancewould
indicatethat a similar mechanismmay be involvedin the pathogenesis
of
bothsymptoms.It can be conceivedthat a slight local pH changeconfined
to the skin or mucousmembranecould act on the itching end organsand

66

R E S E A R c Hr N p H y s r o p A T H o L o c y

evokethe sensation
of itching.More intensepH wouldresultin pain.The
fact that itching,one of the principalsymptomsof dermatological
conditions,can be relatedto local acid and alkalinechangeswithin the skin
wouldpermit the integrationof skin pathologyin a more generalphysiopathological
mechanism.
The conceptof the interventionof two difterent

ITCHING

Seyere
lvloderote
Slight
None

pH
Urine

7.O
6.5
6,O
5.5
6
Hour

F r c . 2 4 . T h e a d m i n i s t r a t i o no f 4 g r a m s o f s o d i u m b i c a r b o n a t et o a s u b j e c tw i t h a n
alkaline patteril of. itching induces together with the alkalinization o f t h e u r i n e , e x acerbalion of the symptom.

one rcsultingin acid substancesand the other in alkaabnormalprocesses,

line compounds,reprcsentsa new approachto the study of many skin
conditions.The therapeuticapplicationof this concepthas producedinterestingresults.(See ChapterXIV)

DUALTSM

67

Vertigo
vertigo has been of specialinterest in our research.while it can be
inducedby various etiologicalfactors, a basic dualism in its pathogenesis
is evident.
Many patientsrvith vertigo have been found to experiencewide variations in the intensity of the symptom. Some have exacerbationsin the
morning,others in the evening.This observationsuggesteda possibledual
patternand we investigatedvertigo by the samemethod usedfor the study
of pathological pain.
In patients with vertigo, the intensitywas determinedat hourly intervalsby using either a scaleranging from 0 to 10, or a seriesof qualifica-

x
@
g

(u

5
U'
o
OJ
g
N
N

8.7

7.
H
!

7o
o 5.6
L

o
c
L

=
,'D

46

Noon

pm.

midnightatn.

Frc. 25. Shows the curves of the intensity of lerligo and of the urinary pH in a case
w i t h a n a l k a l i n e p a t l e r n , t h e v a r i a t i o n so f t h e t w o c u r v e s b e i n g p a r a l l e l . ( 8 . W e l t ,
A M A A r c b i v e s o f O t o l a r y n g o l o g y ,5 8 : 2 7 3 - 3 0 0 ,1 9 5 3 . )

68

REsEARcHrN pHysropATHoLocy

tions, such as none, slight, moderate,severe,very severeand unbearable.

At the same intervals,urine sampleswere collectedand their pH values
measured.Data were plotted in curveshaving time as common abscissa.
Here again, as for pain, two different correlationscould be observed.In
somecases,the two curves--{ne for intensityof vertigo, and the other for
urinary pH level-showed parallelvariations.(Fig. 25) The vertigobecame
more intensewhen the pH was high, and this was consideredto be vertigo

:\
v,

c
o
c
o

CD
L

(u

o
(_
ftt
E
L

Hours
Ftc. 26. Vertigo with an acid pottern. The symptom is more intensive when the
urinary pH values are lower. (B. Welt A M A A r c h i v e s o f O t o l a r y n g o l o g y ,5 8 : 2 7 3 300,1953.)

DUALtSM

69

of an alkaline pattern. In other cases-of an acid pattern-vertigo was

m o r ei n t e n s ew i t h l o w e r v a l u e so f u r i n a r y p H . ( F i g . 2 6 )
The acid-basepathogenesi.s
of vertigo was further confirmed by the
responseto acidifying and alkalizing agenrs.B. welt has widely investigatedvertigo by this method. He also has used the responseto therapcutic agentsas a criterion for the pattern present.The administrationof
[glenel, heptanol or unsaturatedfatty alcohols was seen to induce an
increasein the intensity of vertigo of acid pattern and a decreasein the

500cycles
--- 1000cycles
-2000 cycles
o2o
C)

'3a

o
o

,'=u4j)

an9^-to-tl-t2-/^;-Z-J

zt
Elq
L

Eto
L

I
n.n

t2

HouFs

t
e^1.

Fto. 27. The analysis of the concomitant changesin thc curvc of the intensity of thc
auditive acuity and of the urinary pH shows opposite variations iu a subject with
impairedhearing. The opposite variations arc especiallymanifest for some velucr2000cycles in this case. This rclationship corrcspondsto an acid pottern. (Courtesy
of Dr. B. Welt.)

70

*EsEARcH rN pHysropATHoLocy

alkaline type. Agents such as sodium thiosulfatehave an opposite eftect.

The great similarity in the responseof pain and vertigo to the same
agentshas establishedthe role of acid-basechangesin the pathogenesis
of
vertigo as well as the possibilityof influencingthese changesin order to
relievethe symptom.Thesestudieshave shown that, in spite of the variety
of etiologicalfactors which can induce vertigo, the condition can be considered,from the point of view of therapy,in terms of its dualistic pathogenesis.This has simplifiedthe therapeuticapproach,limiting it to a choice
betweenonly two groups of agents.We will see later how successfulthis
approachhas been in Welt's hands.

:b

.at

1to

{A

tO

&

.t-

r-.

a6

a-

F. f{.
In i t i a l
PostPlacebo
P o s tA c i d i f i c a t l o n

u,
q,

o
o
o,
C

.n
C'
-t

ttiohtear A.C.

\.

\
F r c . 2 8 . T h e a d m i n i s t r a t i o no f a n a c i d i f y i n g a g c n t - 2 g r a m s o f m o n o a m m o n i u m
phosphate-induces a marked increaseof the heoring acuity if. the abnormality presenl correspondsto an alkaline pattern. Only minimal or no changesare seen to occur
if a few drops of acetic acid are used as placebo. (Courtesy of Dr. B. Welt.)

7l

F. |l.
- - - lnltial
"'-" PostPlacebo
-Post Acidification

o
o,
D

(,
c,

o
c

ti
an
C'
J

L e f t e a rA . C .

Fto. 29. Alkaline pattern of hearing impairment correspondsto an increasedhearing

a c u i t yf o l l o w i n g t h e a d m i n i s t r a t i o no f 2 g r a m so f a m m o n i u m p h o s p h a t e .

.*i*r#d'ltW$l$*iliffik,s;l.ii$i,,i#:

72

RESEARCH IN

PHYSIOPATHOLOGY

o
A. l.

--- Initral
F o s tA c i d r f r c l i r o n

o
tf,
20

zf
?t
(y
cl
IJ

&
,t

o,

c ,o
rt
an
o

af

to

Left car
to

30. Alkalinc pattern of hcaring impairment.

DUALISM

73

Impaired Hearing
In studying thc various otologicalprobrems,B. welt and I noted
thatmany patientswith impaired hearingexperiencedvariationsin auditive
acuityat different times of the day, or even in conjunction with food intake.This led us to investigateimpairedhearingby the samemethod used
for pain. (Fis. 27) we studied the influenceof acidifying and alkalizing
agentsupon auditive acuity in casesof impaired hearing.
complete audiograms were obtained, employing differences of only
two decibelsbetweenmeasurements
and using all of the acceptedfrequenciesfor air and bone conduction.Audiograms were obtained in subjects
beforeand after administrationof acidifying agents,such as ammonium
fe

7fo

t6o

K.H.
- -- lnitial
' ..' ....
PostPlacebo
P o s tA c i d i f i c a t i c n
o
o
D
(J

c,
o
c
t^

o
o

'at-e'a

-'-

-./'/-./u"
'"
a-.

L e f t E a rA . C .

F r c . 3 t . T h e a d m i n i s t r a t i o n o f 2 g r a m s o f m o n o a m m o n i u m p h o s p h a t ei n d u c e s a
decreascin hearing acuity if the abnormal pattern is acid.

74

R E s E A R c Hr N p H y s r o p A r H o L o c y

chloride and ammonium monophosphate,and alkalizing agents such as

sodiumbicarbonate.In normal subjects,the audiogramsshowedlittle or no
change.In subjectswith impaired hearing,three types of responseswere
noted for any one agent.The audiogramwas either not changedat all or
an increaseor decreasein acuitywas seen.If a manifestincreasein acuity
2ooo

K .H .
----|nitial
- * - - P o s tP l a c e b o
-f65[
Acidification

U'

o,

-r/--t'

II

-'1

I
(u
o

t
"t""'-

o
U'
o

llight Ear8 . C .

ta

2ooo

4ooo

K.H.

at

o
o

--- lnitial
' - - - - PostPlacebo
-Post Acidification

(,

o
ct

C
v,
U'
C'
-J

L e f t E a r8 . C .
Ftc. 32. Acid pattern of hearingimpairment.

was obtained with one group of agents,an opposite effect was obtained
when an agentof the oppositegroup was administered.
changes of at least 6 decibelswere required before they were considered to be induced by an agent.when changesof such intensity were obtainedwith an agent,it was invariablytrue that oppositechangeswould be
obtainedwith an oppositeagent.It was also true that the same responses

DUALTSM

75

couldbe obtainedin the samepatient in repeatedtests.This appearsto be

highlysignificant,indicating that the responsewas, in fact, correlatedwith
changes
induced by the administeredagent,changessimilar to those seen
in thecaseof acid-basesymptoms.It was thus possibleto integratehearing
impairmentin the group of acid-basesymptoms and to recognize two
well-definedtypes, one correspondingto an acid pattern, the other to an
alkaline.Figures 28 through 32 illustrateseveralcasestaken from Welt's
observation.It must be noted that changesunder the influence of the
agentsare seenat almost all frequenciesin some casesbut at only certain
frequencies
in others.
It must be noted also that not all casesof impaired hearing could be
placedin one or the other category.while this could be done almost without excePtionfor young subjectsor for thosewith still evolvingconditions,
it could not be done for subjectswith old, fully evoluted impairment. It
aPpearsthat once the pathologicalprocesseshave arrived at a terminal
point-and an inactive scleroticscar is present-a responseto acidifying
or alkalizingagentsis no longer to be expected.

Manic-depressiveCondition
We have studiedthe relationshipbetweenintensityof manifestations
and
systemicacid-basebalanceof the body in another group of subjectswith
variousmental disorders.of all the casesstudied, the only condition in
whicha clear relationshipcould be shown was in manic-depressive
subjects.From patients presentingchangesduring the day, passing from
periodsof high excitation to calm, from deep depressionto calm, we obtainedhourly urine specimens.At the same time, observationsof their
mentalstate were made. The evaluationsof mental condition were made
by trained observersor membersof the family using a conventionalscale
whichpermitted translationinto graph. The pH of the urine sampleswas
measured
electrometrically.Curvesof the pH and of the mental conditions
wereplotted having the common time as abscissa.A striking correlation
betweenthe two curves was found in the first investigationsin manic patients.with the pH of the urine at higher values, the patient was calm,
whilethe more acid urine correspondedto periodsof intensiveagitation.
Figure33 showsan example.
This correlationalsocould be demonstrated
by administering
acidifying
andalkalizingsubstances
to manic patients.Acidificationthrough the administrationof phosphoric acid was followed by a manifest increasein
agitationwhile administrationof an alkalizingagent-sodium bicarbonate

76

REsEARcHrN pHysropATHoLocy

-was followed by a pcriod of calm. An oppositebut less evidentcorrelation was seen for severaldepressedpatients.we must mention that the
usual difficulty of judging accuratelythe degreeof depressionfrom one
hour to another can explain this lessercorrelation.Manic manifestations
were much more readilv evaluated.

o
(o
q)

o_

Io

(u

c 6o
L

Hours
Fro. 33. Tbe variations of the curve of the ogitotion of a lO-year-old boy are oppositc to that of the urinary pH, indicating an acid pattern.

DUALTSM

77

Dyspnea
Certain characteristicsof dyspnea suggestedthat this symptom might be
exploredby the srme meansemployedin studying pain associatedwith pathological tissue changes.A regular pattern is observed in certain types of
dyspnearelated to the time of day. Cardiac asthma or paroxysmal cardiac
dyspneaoccur mainly at night after patients have gone to sleep. However,
this is not due to the recumbentposition since patients may assumethis
position during the day without any ill effect. Harrison's "evening dyspnea"
(20) is characteristicallyabsent in the morning, but develops slowly during the course of the day, reaching a maximum in the evening.
Just as for pain, the degreeof intensity of dyspneawas studied in relation to hourly acid-basebalancechangesas indicated by changesin the pH
of the urine. Patientshaving dyspneaof prolongedduration as a result of
variouspathologicalconditions,with no treatmentof any kind for at least
six hours before or during the period of observation,were the subjectsof
this investigation.The degreeof intensityof the dyspneawas estimatedby
trainedobserverswho were in constantaftendance.Dyspneawas recorded
by the observersas absent,slight, moderate,severe,or very severe,estimationsbeing basedupon rate depth and evidentdifficulty in respiration.
At the time of these observations,hourly urine specimenswere obtained from patients with as little disturbanceas possible. No patient
showedevidenceof renal disease.The pH of the urine specimenwas determined potentiometrically.The curves showing hourly fluctuations in
the intensity of the dyspneaand the changesin the pH of the hourly urine
specimenswere then plotted and compared.Acidifying and alkalizingsubstanceswere administeredto patients during the course of some tests in
order to observe the influence of induced changesin the acid-basebalance
uponthe degreeof dyspnea.Phosphoricacid and sodium bicarbonatewere
usedfor this purpose.
Fourteen patients with different pathologicalconditions were studied.
In ten cases, a distinct correlation betweenthe intensity of the symptom
and acid-base variations was found.
Four patients had pulmonary edema associatedwith the symptom of
dyspnea.One patient had edemadue to congestiveheart failure; another
had pulmonary edema and lung metastases
from a carcinomaof the pancreas.Two other patients with cancer of the breast metastaticto the bone
andskin also had pulmonaryedema.In one of these,the pulmonaryedema
appearedto be a result of the accidentalintroduction of a fatty acid in oil
preparationinto the blood stream following an intramuscularinjection. In

78

n E S E A R C Hr N p H y s r o p A T H o L o c y

all four of the caseswith pulmonaryedema,the intensityof dyspnea

was
found to be increased
whenthe urinarypH showedchangestowardmore
alkalinevaluesand was diminished
whenthe pH changes
wereopposite.
In thesecases,the intensityof the dyspnea
wasrelievedfollowingadmin-

DYSPNEA
Seyere
Moderote
Stight
None

pH
Urine

6.5
6.O
5.5
5.O
4

Hour
Ftc. 34. The parallel variations between the curves of the intensity of dyspnea and
o f t h e v a l u e o f t h e u r i n a r y p H i n a c a s e o f p u l m o n a r y c o n g e s t i o nf o l l o w i n g a n
a c c i d e n t a li n t r a v e n o u si n j e c t i o n o f a f a t t y a c i d p r e p a r a t i o n ,i n d i c a t e s a n a l k a l i n e
patlern.

istrationof phosphoricacid, while sodiumbicarbonateincreasedthe dyspnea. By analogywith pain, we called this correlationan alkalinepattern.
(Fie.34)
Six casesshowedan oppositetype of correlationbetwecnthe intensity
of dyspneaand the acid-basechangesin the pH of the urine, All of these

DUALISM

79

caseshad mediastinalor pulmonary massesand failed to show signs of

pulmonaryedema. In these cases,the maximum degreeof dyspneawas
associatedwith a relatively more acid urine, and the dyspnea was less
intensewhen the urine was more alkaline.In thesecases,phosphoricacid
increased
the degreeof dyspnea,while converselysodium bicarbonatedec r e a s eidt . T h i s w o u l dc o r r e s p o n tdo a n a c i d p a t t e r no f d y s p n e a(.F i g s . 3 5 ,
36)
Thesefindings,althoughobtainedin only a limited number of patients,
stronglysuggesta similaritybetweenthe fundamentalorigin of both pain
anddyspnea.As in pain, the two patternsof dyspneawere associatedwith
a relative alkalosisand a relative acidosis.

Severe
Moderote
Slighl
None

pH
Urine

6.5
6.O
5.5
5.O
3

Hour
F r a . 3 5 , T h c o p p o s i t ec o n c o m i t a n tvariations betwcen the intensity of dyspnea and
urinary pH in a case of mcdiastinal metastasesof a hypernephroma indicatc an acid
paltefn,

80

REsEARcH tN pHysropATHoLoGy

certain differencesexist betweenthe investigations

of pain and dyspnea. In studyingpain, it was necessary
to dependentirelyupon the observations
of the patientas to the relativeintensityof the pain experienced
from hour to hour. In dyspnea,the patient'sown observations
werefound

oYSPNEA

NoHCOs

Seyerc

Moderotc
Slight
None
7.5

70
6.5
6.O

5.5
f

Hour
Flc. 36. The administration of sodium bicarbonatc increases the intensity of the
dyspneain a casc of pulmonary metastases
of cancer of the gall-bladder,indicatin3 an
alkaline pattcrn.

to be less reliable due to the emotional factors associatedwith dyspnea.

It was, however, possibleto depend upon trained observerswho could
estimateaccuratelyenough the degreeof dyspnea on the basis of rate,
depth and apparentdifficulty of respirarion.It was found that the greater
the experienceof the observer,the closer was the correlation betweenthe
curyesof intensity of dyspneaand the changesof the urinary pH values.

DUALtSM

8l

since it appearedevident that acid-basechangesplay a significantrole

in influencingthe degree of dyspneacxperiencedby the patient in two
oppositedirections,it was important to considerthis influencein relation
to the physiologyof respirationwhich is known not only to be affectedby,
but even dependenton acid-basechanges.
concerning the respiratorycenter, two different mechanismsof direct
chemicalstimulationhave beensuggested
as being responsiblefor dyspnea.
One is lack of oxygen, but considerableevidenceexists to indicate that
simpleanoxemiais not a true causeof dyspnea
. (zl, 22) on the other
hand,an acidosisof the respiratorycenter is known to causedyspnea,but
it has been establishedthat only a very profound generalacidosis,such as
that connectedwith diabetes,acid poisoningor emphysemacan create a
changein blood pH sufficient to affect the respiratory center. This is also
true for the chemoreceptorcenterslocatedin the carotid sinus, which are
still lesssensitiveto the generallack of oxygenor acidosisthan the respiratory center. while general acid-basechangessecm to play an important
role in dyspnea,thesechemicalchangeswould not appear to act directly
upon the respiratorycenter since the actual blood pH is not sufficiently
changed.
other factors-involving direct local, rather than systemicinfluencesmaybe considered.One suchdirect action would be relatedto reflex stimulation which is generally accepredas having an important role in the
controlof respirationand even dyspnea.This reflex control of respiration
is connected with nerve endings within the lung parenchyma which are
stimulatedwhen the walls of the alveoli are stretchedin the respiratory
phase.The stretch reflex inducesimpulses,carried by way of the vagus,
which acting upon the respiratorycenter, stop the inspiratory phase and
bring about expiration. It may be assumedthat the nerve endingswithin
theparenchymacan be stimulatednot only by mechanicalchangeswithin
theparenchymabut also by local chemicalchangestoo. Under abnormal
conditionssuch as pulmonary congestion,there may be a change in the
tissuereaction, as has been recognizedby a higher pH of the edema fluid
itself.Through this reflex mechanism,these pulmonary tissue reaction
changes
may producedyspnea.The generalacid-basefluctuationsinfluencingthe local alkalosiswould, as seenin pain, indirectlyinfluencethe degree
of dyspnea.such a mechanismmay accountfor dyspneawith an alkaline
patternfound in pulmonary congestion.
A simi.larlocal factor also could be seen for the acid pattern. It was
observedthat all patients with an acid pattern of dyspnea had tumors
locatedwithin the mediastinumor in the lung parenchymaitself. From

82

R E s E A R c Ht N p H y s r o p A T H o L o c y

the study of pain, we know that abnormal degreesof acidosiscan occur

in tumors.pH changestoward acidosiswithin a tumor tissuein the vicinity
of chemoreceptorsmay produce impulse dischargesin these centers especially sensitiveto changestoward acidosis.They may alter the character
of respirationand result in dyspnea.As in other conditions,these local
changesoccur even with reducedchangesin the systemicacid-basebalance.Through this mechanism,variationsin the intensity of dyspneacan
occur without the intensivechangein the generalacid-basebalancewhich
is considerednecessaryto affect directly the respiratory center itself.
The fact that relatively small acid-basebalance changesaffect the intensityof dyspneain two oppositedirectionscan thus be explainedby this
indirect influenceexertedupon a local process,Ieadingto alkalosisin abnormal conditions affectingthe pulmonary parenchymaand to acidosisin
lesionspresentin the neighborhoodof the chemoreceptivecenters.These
two mechanismsproposedas part of this working hypothesis,appear able
to explain the paradoxicalexpcrimentalfindingsin clinical studiesof this
symptom, where opposite responsesupon dyspneaare seen for the same
acidifying and alkalizingagents.
A more completestudy of dyspneaunder this aspectwill be published
separately.
Dualistic Patterns at Other Levels
Cellular Level
Cytologicalstudieshave revealedthat some characteristicsof the cell,
other than those typifying the cancerousanomaly itself, exhibit dual patterns.For the cellular level, they could ultimately be related to changesin
the aging processes.In tumoral foci having an acid pain pattern, cytological
characteristicsindicate a prolonged cellular youth. A round aspect of the
nucleus,with a fine texture of chromatin and well-separated
nucleolus,
and a basophylcytoplasmrepresentmajor characteristics
of cellular youth.
In lesionswith an alkalinepain pattern,the cells show rapid early aging.
The tendencyto lobulationof the nucleus,to separationof chromatinand
formation of clumps, to cytoplasmaticoxyphily and the appearanceof
azurophylgranulaecharacterizesuch aging.Rapid aging was seento lead
to prematurecellu.lardeath through piknosis and karyorrhexis.Opposite
cellular aging processescould be further related to differencesseen in evolution of tumors. Rapid aging of cells,associated
with alkalinepain patterns, results in necrotic tumors and ulceration of superficial lesions.
Frequently,it could be noted that a changefrom acid pattern to alkaline

DU^LtsM

g3

occursand is accompaniedby a melting away of massivetumors and their

replacementusually by ulceration.
TissularLevel
As seen above,pain, dyspnea,vertigo and itching can be considered
tissuelevel manifestationswhich show dualism. Additionally, the nasal
pH (Note 5/ deviateseithertoward acid or alkalinevaluesand thesedeviationscan be correlatedwith similar acid-base
changesat the tissularlevel.
Thesemeasurementsare useful as a diagnosticcriterion for tissue level
changes.
Organic Level
The same dualismobservedat the cellular and tissuclevcls also was
foundin signsand symptomsinvolvingthe organ level.Dual patternswere
foundfor dysfunctionsof variousorgans.Insomniaand somnolence,
diarrheaand constipation,oliguriaand polyuria,tachycardiaand bradycardia,
all representexamplesof dualism at the organic level.
The dualismevidentfor the lengthof persistence
of a wheal (Note 6)
inducedby intracutaneous
injectionof a salinesolutionwas also related
to the organ levelwith the skin considered
to be an organ.In normal subjects,the resorptionof the wheal is completedin about ls-zo minutes.
In onegroup the resorptiontime is short,evenreducedto a few minutes.In
othergroups, on the contrary, the resorption time is greatly prolonged,
thewheal sometimesbeing presentevcn after more than 90 minutes.
Svstemic
Level
Studiesof dualism at this level covered temperaturevariationsand
changes
in variousblood and urine values.
Temperalure
Two patternsof temperaturechangeswere found in cancer patients.
For oral temperature,37oc (98.6"F) was consideredas the reference
va.lue.
Temperaturesmeasuredseveraltimes during the day showedthat,
for many patients, the values were fixed either above or below this referenceline. Normal individualsexhibit daily variationsof temperature;the
curvenot only crossesthe referenceline but also shows broad changes.
In contrast,variationsusuallyare smallerin abnormalcases,and the curve
remains
on one or the othersideof the referenceline.Figures37, 38 and 39
showexamplesof such curves.The two patternshave been found to be

Flc" 37, The curve of oral temperutureof a casewith gcneralizedmctastaticmelanoma is pcrsistentlyabovc the 98.6'F (37'C) line, which corrcspondsto the averagevaluc of normal individuals.

Fre. 38. The curve of the oral tempcratureof a paticnt wilh gcneralizcdmctastases
of the breastshowsvaluesconstantlybelnx'lhe 9t.6"F averase
of adcnocarcinetma
line.

DUALTsM
r{i r1,..

R.
6-lo

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IT

* 1 "

rr .,r,a rrrk

I
/3

/2

/ 8 5

t,g

/{

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rl

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lt

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rt

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t

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I

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t

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Ftc. 39. Curve of the oral temperature of a patient witb bronchogenic cancer with
abdominal metastassshows values constantly helnw g8.6"F (37'C) avcrage line.

independentof type or site of origin of tumors. we will discussthe significaoce of tempefature patterns later.
.Syrtenrc Analyses:
Changesin urine or blood valueswere followed in hundredsof subjects
for long periods,even years. It appearedessentialto plot the data graphically. An average value, obtained from a significant number of normal
subjects,was riepresentedin graphs as a referenceline. In normal subjects,
as a general rule, relatively wide oscillations omur around the average
value line. By contrast, in diseasestates,there is a fixation of the curve on
one or the other side of the averagevalue, the curvesexhibiting only slight,
or even no, variations.Two opposingpatternsare thus evident in disease
sialesfor each type of analysisof blood and urine. The dualism indicates
once again the existencein systemicmetabolismof two kinds of abnormal
changeswith antagonistic characteristics.It is important to note that the
advancedcancerouscondition is characterizedby a marked dualistic pattcrn at the systemic level.

86

REsEARcHrN pHysropATHoLocy

Blood
In blood analyses,the concentrations
of potassiumand calcium, the
presenceof C reactiveproteins,the number of leucocytesand of circulating eosinophiles,(Nole 7) and the red cell sedimentationrate, have been
studied,usingclassicalmethods.They all show the samedualism.Figs. 40
to 43 show someof thesecurveswith the averagevaluesas referencelines.
with the intention of obtaining information concerning the amount of
potassiumpresent in cells, we investigatedthe content of this cation of
the red cells.(Note 8)
Urine
In urine studies, measurementswere made for pH, specific gravity,
surfacetension,oxido-reduction
potential,excretionof sodium,potassium,
calcium, chlorides, phosphates,sulfates, sulfhydryl, indoxyl, grucuronic
acid, peroxides, etc. Most studies were carried out by routine test techniques.For some analyses,however,conventionaltechniqueswere found
to be inadequateand new testsdevised.
For the urinary excretionof sulfhydryl,a new techniquewas devisedby
M. Bier and P. Teitelbaum in our laboratories.Using the warburg
micromanometer,
thc nitrogenliberatedfrom sodium azide in a buffered
solution in the presenceof free iodine was found to be directly related to
the amountof sulfhydrylpresent.The amountof nitrogenfreed at a determined momcnt-13 minutes-was most indicative.(Note 9)
For the information which we neededconcerningthe amount of calcium in urine a very simplemethodwas devised.(Note I0)
For measurementof urinary surface tension, we devised a new
technique.We used a capillary so calibratedas to give the surfacetension
in dyne/cm for a fluid with a specificgravity of 1,015. In this method,
severalarrestsor slowdownsof the descendingcolumn are noted and make
it possibleto obtain informationabout an extremelyimportant factor which
is usuallynot consideredin the measurement
of surfacetensionwith other
methods.It is known that urine is formed of differentconstituents.
some
of them with the tendencyto move toward the surfacewhile others tend
to move toward the bulk of the fluid. Changesin the distributionof these
constituentstake place. They induce changesin the value of the surface
tensionof the urine which occur even during the time measurements
are
taken. The descendingcolumn in a capillary will indicatethesechanges
which are important for precisemeasurement
of surfacetensionof urine.
(Note I I )

D U A L I S M

87

lz

!'
a1 a

Ftc. 40. Curve of the value of the K + in blood serunt in a case with periarteritis
nodosa. The values remain above 4.5 mEq, which represent the average value obtained from seriesof normals.

H
Y

5
c
o

a/,

F I c . 4 1 . C u r v e o f t h e v a l u e so f K + i n t h e b l o o d s e r u m o f a s u b j e c tw i t h c a r c i n o m a
of thc breast, liver metastasesand jaundice. The values remain almost constantly below the 4.5 mEq line.

,:{#:i;;!lil:tt;iiil:tiii'i1l,l1$.'*ii
{llii*ri'+

88

R E S E A R C H

I N

P H Y S I O P A T H O L O C Y

U'
O.J
t)
o

IJ
f

Days
F I o . 4 2 . T h e c u r v e o f t h c n u m b e r o f b l o o d l e u c o c y t e so f a c a s e o f b r e a s t a < J e n o carcinoma shows constantly values abovc 7.0O0,considercdas the average value for
normals.

9ao
C

7
(e
3n
(l,
(J

f,

(J

5
o,

-J

0ays
Ftc. 43. The curve of thc number of blood leucocytesof a casc of brcast carcinoma
showspcrsistentlyvalues below the averageline of 2.000.

DUALTSM

89

Two methods were employed to rneasurethe oxido-reduction potential

of urine. In one, the measurementwas made with a potentiometerusing a
platinum electrode,fust at the pH of the urine and once again after bringing the pH to 7 by adding the necessaryamount of NaoH or HCl. (Note
l2) rn the secondtechnique,difrerencesin potentialwere measuredthrough
the time necssaryto discolor a solution of toluidine blue in an acid
medium at l00oc. The value was determinedin seconds,the concentration of the dye having been chosen so that discoloration in 100 seconds
represented
an averagevalue for normal subjects.More rapid discoloration
indicated higher potential, while a low potential correspondedto slow or
evenno discoloration.(Note I3)
The presenceand relativeamountsof oxydizingsubstancesin the urine
were also investigatedby meansof two reactions:in one, through the passageof indoxyl into indigotine and indigorubineunder the influence of
sulfuric acid (Note 14); in the other, through liberation of iodine from
iodides in the acid medium.
It is interestingto mention at this moment, the variationsencountered
in the metabolismof nitrogen and the form under which it is excreted
through the kidney under normal and pathologicalconditions. we could
showthat in generalthe form of excretionis determinedby the amount of
wateralso excretedby the organism.High amountsof water will thus induce
the excretionof nitrogenprincipallyas ammonia,low amountsas uric acid.
This relationshipwas explainedby data found in comparativephysiology.
The availability of water in the environmentof variousanimalswas seento
determinethe form under which theseanimalsexcretenitrogen.The occurrenceof similnl conditionsconcerningthe excretionof water in pathological
states,furnishesan interestingexplanationfor the form under which it is excretedin thesepathologicalconditions.In Note l5 this problem is discussed
in more details.
Iu all tests requiring quantitative measurements,an important problem
arose.It appearedpractically impossibleto obtain 24 hour urines routinely
for periods of months in large groups of individuals in order to note the
continuouschangesin excretionof the differentsubstancesstudied.Measurements of various substanceseliminated in urine could be made under
thesecircumstancesonly by using isolatedsamples,The values so obtained
expressthe concentrationsof the substances
in the sampleand consequently
weredirectly related not only to the amount eliminatedby the kidney but
alsoto the amount of water excretedat the same time. The values varied
greatly from one specimento the next, according to the amount of water
excreted.Therefore thesedata could only be of relative usefulness,Since

90

xESE^RcH tN pHysropATHoLocy

specificgravity is also a direct functionof the amountof water excretedin

a urine sample,we related the concentrationof a substanceto the specific
gravity of the same sample.This ratio provided a new value which is independentof the amount of water in the sample and is more closely relatedto the amountof the othersubstances
which vary much lessin amount
than water. From the physiopathological
point of view, the data obtained
were seen to correspondto the degreeof active reabsorptionof a given
substance
by the kidney.While the ratio of the concentrationover specific
gravity would vary directly with the excretionof the substance,the inverse
ratio would rcpresentan index of retention,which increaseswith the retention of the substancein the body. Such indiceswere routinely applied for
the differentsubstancestestedin urine to obtain more reliable valuesthan
could be obtainedfrom the concentration
data alone.(Note t5)
Urinary Patterns
various urinalyseswere performedduring sufficientlengthsof timc on
a larger number of subjectsconsideredto be normal. For each test, an
averagevalue was thus obtained.This averagevalue servedas a referencc
line for the curvestraced with the data obtainedfrom the subjects.The
AcC u3i

Sagiibr,

| l ? o ? 2

?r
0" 2 1
.o 15
9

AI

ERS

Flc. 44. Curves of. diflerent anall'sesof daily urine samples of a normol 4O-yearo l d m a l e . F o r e a c h k i n d o f a n a l y s i s t, h e c o r r e s p o n d i n ga v e r a g cv a l u e w a s c a l c u t a t e d
from the measurementsobtained in more than one hundrcd normal subjects. The
curves are seen to pass from one to the other side of the lines correspondingto thc
r e s p e c t i v ea v e r a g ev a l u e s .A c e r t a i n c o r r e s p o n d e n c ies s e e n f o r t h e c h a n g e sw h i c h
take place at the same time in these curves. Parallel variations are seen between spccific gravity and chloride index, while opposite to thosc for the pH and surface
tension,

9l

D U A L I S M

study of thesecurves has permitted us to recognizeseveralcharacteristic

patterns.Under conditionsconsidered
normal,the curvcswere seento pass
from one side to the other on this averageline with relativelywide variations. When an abnormalityexisted,the variousurine analyseswere characterizedby a curve with only small variations,fixed on one or the other
side of the averageline. For each test, two such characteristicabnormal
palternsare encountered.Fig. 44 showsthe curvesof differenttestspassing
trl
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-'.
.-_..
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&n
9n
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t,

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pH

50
PH

6
I
6

Frc. 45. Curves of the valuesof the urinary pff followed during 50 days in five
s u b j c c t s .T h e c u r v e ( a ) c o r r e s p o n d st o a n o r m a l i n d i v i d u a l w h i l e t h e o t h e r f o u r
( b , c , d , e ) t o s u b j e c t sw i t b d i f f e r e n t c a n c e r o u sl e s i o n s .F o r t h e n o r m a l c a s e , t h e
values pass above and below the 6.2 value which is consideredas the average computed values obtained from normals, For the abnormal cases,the curves which show
l i t t l e v a r i a t i o n s a r e f i x e d a t o n e o r t h e o t h e r s i d e o f t h e a v e r a g el i n e , F o r c a s e ( b ) ,
an adenocarcinoma of the ovary' and case (c), a bronchogenic cancer, the curves
are fixed below the averageline. For case (d), with a bronchogeniccancer. and case
( e ) . a c a n c e r o f t h e b r e a s t w i t h g e n e r a l i z e dm e t a s t a s e st ,h e c u r v e s a r e a b o v e t h i s
averageline.

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D U A L I S M

/ e 3

July

June

?5

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Po?ient-Mr: Spe
Ftc. 47. Curve of the values of the urinary specific gravity in a case of cancer
of thc breast, with the values fixed above 1016 which is considered as an avcrage
value for the specificgravity as computed from a seriesof measurementsin normals.

from one side to the other of the respectiveaveragelines as corresponding

to a normalsubject.Figures45 to 5l showseveralexamplesof two opposite
patternsfor the various testsusedsuch as urinary pH, specificgravity, surface tension,chlorideindex and sulfhydrylindex.
F undatnental Ofi balances
When the curvesof variousanalyses
obtainedat the sametime for thc
samesubjectwere checked,an interesting
relationshipwas found.
In patientswith small localizedtumors, only some of the analyses
showedabnormalpatterns.However,with the evolutionof cancertoward
the terminalstage,abnormalpatternsbecameapparentfor more and more
analyses.In advancedcases,most of the analysesshowedabnormal patterns.

Moy

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F r c . 4 8 . T h e v a l u e so f t h e u r i n a r y s p e c i f i cg r a v i t y i n a c a s eo f c a n c e r o f t h e b r e a s t .
are the whole time below thc averatc value of 1016.

94

RESEARCH IN

PHYSIOPATHOLOCY

Ftc. 49. The urinary surloce tension remains constantly fixed abot,e the average
value of 68 for more than a year in a case of cancer of the urethra.

2oo
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F t c . 5 0 . T h e u r i n a r y s u l f h y d r y l i n d e x r e m a i n s b e l o w t h e a v e r a g ev a l u e o f
c a s co f c a n c e ro f t h e b r e a s t ,w i t h b o n e m e t a s t a s e s .

DUALISM

95

It was especia.lly
in advancedstagesthat definitegroupingsof patterns
with opposite characteristicscould be recognized.They correspondedto
two fundamentaloffbalanceswhich we have called "Type A" and ,.Type
D." 1"4" for anoxybiosis,"D" for dysoxybiosiswhich representthe principal manifestationsof oxygenmetabolismin a phaseof theseoffbalances.
)
The correspondencebetwecn the different abnormal patterns defining
the offbalanceA or D, is seenin Figures52 to 60. The abnormalpattern of

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Frc. 5 1 . T h e u r i n a r y s u l f h y d r y l i n d e x r e m a i n sa b o v e t h e a v e r a g ev a l u e o f r . 5 i n a
caso f h y p e r n e p h r o m aw i t h l u n g m e t a s t a s e s .

the low urinary specificgravity appearstogether with a pattern of high

valuesfor the pH, both corresponding
to the offbalancerype A. (Fig. 52)
A high urinary pH and low chlorideindex patterncorrespondto the offbalancetype A, as seenin Fig. 53. The analysesof a casewith low specific
gravity,high pH, low chlorideindex and high surfacetension,as present
in offbalancetype A, is shown in Fig. 54. An oppositecase,offbalancetype
D, with high specificgravity, low pH, high chloride index and low surface
tensionis shownin Fig. 55. A similarcaseof offbalancetype D is shownin
Fig. 56 with high urinary specificgravity,low pH, low surfacetensionand
low blood leucocytenumber.In Fig. 57, the low pH, high sulfhydrylindex
and low surfacetensionshow an offbalanceof the type D.
of the levelsmay explainwhy, in the samesubject,
The independence

e 6 /

RESEARCH IN

t94l
Moy
30

PHYSIOPATHOLOGY

Ju n e

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54
4.6
Polrenl- Mrs Von

Frc. 52. A correspondenceis seen between the patterns of. urinary specific grat,ity
and that of the urinary pH in a cancer of the colon correspondingto tbe type A offbalance.The changesstill present in the curves are opposite.

not all the analyticalpatternsobligatoryconcord at all times in the same

subject.Especiallywhen defensereactionsintervene,the offbalanceat one
levelcan be differentfrom that at otherlevels.Fig. 58 showssuchexamples.
Usually,as the diseaseprogresses,
many of thesedifferencesdisappear,the
manifestations-analyticaland clinical at different levels-entering in the
sametype of offbalance.Fig. 59 showssuch an example.

ZC
62
4 6

cl
I tiaar

2.5

F r c . 5 3 . I n a c a s e o f s a r c o m ao f t h e l e g w i t h l u n g m e t a s t a s etsh c c u r v e o f t h e a r i nury pH is fixed above the averageline (6.2) while the curve of chloride index Wl o w t h e a v e r a g el i n e o f 2 . 5 . T h i s r e l a t i o n s h i pindicates a type A of the offbalance.

D U A L I S M
Fab.n,t
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F t c . - 5 4 .T h e a n a l y s i so f a t e r m i n a l c a s eo f c a n c e ro f t h e b r e a s t ,s h o w i n g t h e
fundanrcntaloflbolance, low specific gravity, high pH, low chloride index and high surface tension, correspondingthus to the offbalance A.

The passageof a subjectfrom one offbalanceinto the oppositeone is seen

to occurduring the evolutionof the conditionmost often inducedby therapcuticattempts.Fig. 60 showssuchan example.
The two oppositeoffbalances,identifiedfirst through the urine analyses,
couldbe recognizedto exist for all the manifestations
taking place at the
diffc'rentlevelsof the organization.The manifestationsseenat theselevels
could thus be interpreted as correspondingto one of the two opposite
fundamental
offbalances,
type A and D. Table Iv showsthis coordination
of the different manifestationsaccordingto the two oppositeoffbalances.
on the basisof all thesedata, clinical manifestationsand analyses,the
dualisticpathogenicconceptappearedto be well establishedand the dual-

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60

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Ftc. 55. A terminal case of cancer of the breast shows all the analyses fixed in
opposite position to the case of Fig. 54, i.e., high specific gravity, low pH, high
chloride index and low surface tension, corresponding to an offbalance of the type D.

RESE^RcH IN

Leucocyt
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o

PHYSIOPATHOLOCY

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Ftc. 56. offbalance type D, shows high urine specific gravity, tow pH, low surface
tension and low blood leucocyte number.

66
64
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Ftc. 57. The 3 curves, PH, sulfhydryl index and surface tension show an offbalance
type D.

DUALtSM

gg

isticphysiopathological
mechanism
studiedin complexconditions
in gcneral
andin cancerin particular.
r5
5?

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F t c . 5 8 . T h e u r i n e a n a l y s e so f a p a t i e n tw i r h c a n c e ro f t h e o v a r y . s h o w i n g a d i s c o r d a n c e b e t w e e nt h e p a t t e r n sp r e s e n t .W h i l e a l l t h e a n a l y s e ss h o w p a t t e r n so f t h e
t y p e D , t h e c h l o r i d e i n d e x r e m a i n s t h r o u g h o u t t h e e n t i r e o b s c r v a t i o nc o n s t a n t l yo f
type A.

Place ol Dualism in Cancer Physiopathology

After recognizinga dual pattern in most manifcstationsof cancer,the
problem was to considerthe relationshipbetweenthis dualismand cancer
itself.
One critical observationwas that a pattern in a given patient may
change.During the evolution of the condition, seldom is the changefrom
abnormal to normal, but usually from one abnormal to the opposite abnormal pattern. This fact has a specialimportance as will be seen later. In
some cases,even the immediate,direct causcof such changcscould be
established.
The following case is illustrative.
Mr. S. L., a patientwith cancerof the prostateand metastaticdestruction of half of the sacrum,was in very severepain. Study of variationsin
pain intensity and in urinary pH, as well as the responseto acidifying and
alkalinizing substances,indicated that the pain was of a typical acid pattern.The patientstartedradiotherapyand pain decreased
with almostevery
treatment,until after a few sessionsit had disappearedcompletely.Out of
bed and feelingwell, he continuedthe treatment.Howevcr, at about the
nvelfth session,pain again appearedand thereafterincreasedwith each
treatment.After five more sessions,
he was back in bed and sufteringsuch
unbearablepain that radiotherapyhad to be discontinued.
At that time, a
new analysisof the patternof pain showedthat it had changedfrom thc
that this changemight be thc
originalacid to alkaline.It was hypothesized

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DUALISM

l0l

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d16
a

Ftc. 60. A condition can pass from one offbalanceto the opposite one. The passage
ol the analyses lrom the oflbalance lype D into the oflbalance trpe ,,1 is seen in a
caseof caocer of the lung. The analyseswhich first show high specific gravity, Iow
pH and high chloride index, pass subsequentlyto the opposite side, with low specific
gravity, high pH and low chloride index. This new oflbalance remains unchanged
for a long time as shown in the second part of the curve.

resultof irradiation. Study of other cases,after irradiation, has confirmed

the hypothesis.
Changesof pain from an acid to an alkalinepattern also were observed
patients
in
who had sufferedshock or seriouspyrogenic infections.viral
infection such as flu, or even smallpox vaccination,seemedto induce an
opposite change-from alkaline to acid. Changesobtained under the influence of therapeutic agents were frequently observed and will be discussedlater.
The dual pattern seen for most cancer manifestationswas integrated
into the concept of the diseaseas a complex organized condition that developsthrough progressiveparticipation of hierarchically superior levels of
the organization. From a clinical point of view, such participation correspondsto the successiveaddition of new manifestationsto a previously less
complexcondition. According to their levels,these"Added Factors" correspondto prolonged youth or rapid cellular aging,to acid or alkaline pain,
to blood and urinary analytical data fixed in one or another of two opposite
patterns.Dualism is the principal characteristicsharedin common by all
theseprogressively added manifestations.
In trying to understandthis dualism and its significance,we had to

102

*ESEARcH IN

PHYsroPATHoLocy

consideragainthe placeof dualismin the generalorganizationof nature.

The studyof reactivityin naturehas led to the recognitionthat a basic
dualismexists-that the forcesoperatingin naturecan be separatedinto
two oppositegroupsfor almostall the interveningfactors.Forceswhich
would tend to lead toward annihilationof differencesand producea state
of maximumentropyor homotropyor the antagonistic
forceswhich tend
TrsLe IV
M,tNtreSrrTIONS
kvel

Offbalance"A"

Offbalance"D"

Cellular Prolonged cellular youth

Rapid cellularaging
Less difterentiatedcells, especially More differentiated cells, espeof connectivetissues
cially of connective tissues
lncreased amount of connective Decreasedamount of connective
tissue
tissue

Tissular Increasedlymphatic tissue

Organic

Low
oxido.reduction poten
processes
Low chloride content
Local acidosis
Acid pattern symptoms
(Pain, dyspnea,itching,etc.)

Decreasedlymphatic tissue
High oxido-reduction potential
processes
High chloride content
Local alkalosis
Alkaline pattern symptorns
(Pain, dyspnea,itching, etc.)

Somnolence
Constipation
Polyuria
Exophtalmia
NasalpH below6.5
Slow absorbtionof skin wheal

Insomnia
Diarrhea
Oliguria
Enophtalmia
Nasal pH above6.5
Rapid absorbtionof skin wheal

Systemic Blood
Leucocytosis
Eosinophilia
High color index
Low potassiumcontent
Low R.C. sed.rate
No C reactiveprotein
Urine
Low specificgravity
High pH
High Cl, Na excretion
Low SH, Ca, phosphate,sulfate
excretion
High surfacetension
Death in coma

Blood
Leucopenia
Eosinopenia
Low color index
High potassiumcontent
High R.C. sed.rate
High C reactiveprotein
Urine
High specificgravity
Low pH
Low Cl, Na excretion
High SH, Ca, phosphate,sulfate
excretion
Low surfacetension
Presenceof oxidizing substances
Death whi.leconscious

DU^LrsM /

103

to maintainand increasedifferencesand lead to more complex organization

-and are thus cataloguedas ncgatively+ntropic,
ectropicor heterotropic
-would thus appear with each new added factor when a higher hierarchic entity is realized.For this reason,they will appearespeciallymanifest for the same added factors intervcningunder abnormal conditions.
Dualism consequentlyconcernsthe manifestationsprogressivelyadded.
The result is seenin the alterationof the oscillatorymovementwith alternatepredominanccof one and then the other of the antagonisticforcesfor
theseadded factors.This fact would explain the dualism seen especially
in abnormalities.It also explains the interventionof the dual patterns
mentionedabove.
Dualism becomeseven rnore important when it can be seento play a
capital role in the mechanismthrough which agcntsact upon abnormal
conditions.Originally,the dualisticactionsof agcntscould be ascertained
through the changesinduced in pain and various other patterns. It has
consequentlybeen possibleto classifythe effcct of various therapeutic
agentsaccording to their influenceupon these patterns and, through this
more readily measurableinfluence,on the fundamentaloffbalancesthemselves.
At the sametime, the study of the influenceexertedby variousagents
upon these patterns, which is presentedlater, has posed the problem of
preciselywhat effectsupon thesepatternsare producedby the substances
which are body constituents.And this led us to a third basic conceptdealing with the importantrole playedby body constituents
and, in particular,
by the lipids.

CHAPTER

TH E CONSTITUENTS

E-lr
.lL lltsnencHrc oRcANrzArroNand dualism have opened the way for

a
study of the body constituentsin an attempt to systematizethem and their
functioningin accordancewith thesetwo concepts.In this research,we
consideredbesidesthe constituents
separatedin groupsas lipids, proteins,
carbohydrates
and electrolytesalso the elementsas a sourceof interesting
information.
With most of the manifestations
integratedin dualistic patterns,rvc
plannedto test the differentconstituents
by noting their influenceon these
patterns.It was to be expectedthat some might have selectiveactivity at
certain levelsof organizationand the manifestationsrelatedto theselevels.
This selectivitydid becomeevidcntbut it also turned out that any agent,
in sufficientamount, cxertcd an effect at any level. The problem was to
selectthe manifestationwhich would respondmost readily to the greatest
number of agents.This would make comparisonsbetweenagentseasier
and serveas a practicalcriterionfor the start of classification
on this dual
basis.
The measurementof the influenceexertedby various agentsupon the
secondday wound crust pH (s.d.c.pH) proved to be particularlyrewarding and was employedin the first part of the investigation.
The s.d.c.pH
providedan indication not only of acidifyingand alka.lizingeffectsbut also
comparative values for these effects. Later, the influence exerted upon
many other manifestations
was studiedfor corroboration.Details of the
s.d.c.pH techniqueand thc resultsobtainedare in Note 1.
In studyingthe elements,we chose,first, simplecombinationsin which
they appear.Each anion was investigated
by studyingit as it occurredin
thc respective acid and in compounds in combinations with diffcrent
cations;each cation was studiedin its combinationswith differentanions.
104

THE CONSTITUENTS

105

ln this way, we obtained a seriesof data which enabled us to pinpoint the

influenceof each element.
TheElements
Using this method, we could determinethat elementssuch as Li, K,
Na,Fe, Ni, Zn, Hg, Bi, B, F, Cl, Br, I-in sufficientamount-produced in
the s.d.c. pH an acidifying effect. The opposite eftect-alkalization-was
seenfor Mg, Ca, Sr, Ba, Cu, Pb, S and Se. It must be emphasizedthat
someelements-such as K, Fe, Zn, Hg, Cl in the acidifying $oup and Ca,
Cu, S and Se in the other-produced an intense effect while others had
a weak though still clear, action. We must add that Ni and Cr showed a
relatively weak acidifying effect. This separationof elementson the basis
of acidifying or alkalizing effect agreed with almost all data available
about antagonismbetween elements-for example, the known antagonism
betweenK and Ca, Mg and Cu, and betweenMo, Zn and Cu.
As a secondstep,we relatedthe elements,through their influenceupon
the s.d.c.pH, to one of the two fundamentaloffbalances.
Those inducing
acidificationwere thus classifiedas an "inducing offbalancetype A," or
"anti offbalance type D" or "anti D," while those producing alkalization
wereca.lled"inducing D," or "anti offbalancetype A," or "anti A." Going
anotherstep, the acidifying elementswere consideredto have a tendency
towardincreasingheterotropy;the alkalizing,a tendencytoward increasing
homotropy.This led us to attribute to the first, the acidifying group, the
qualification"hetero" type, and to the second,thc "homo" type. We did
not give these designationsany other meaningthan that indicated above,
usingthem for didactic facility.
TheSeries
After classifying elementsinto hetero and homo groups, we studied
thesegroupings in terms of the place of the elementsin the periodic chart.
We could quickly see that when two or more elementsare part of the same
series,they also belong to the same group. For example, all elementsin
the I A and the VII A seriesare A inducing,or "hetero." The members
of the II A and VI A seriesare D inducing or homo. At this point, we
tentatively extended these hetero or homo characters to an entire series
after one or more elementsin it had been recognizedas such. The I A,
Il B, III A, V A, VII A seriesand the Fe subseriesof VIII were classified
ashertero or A inducing, while the II A, I B, VI A seriesand the Co subseriesof VIII were labelled homo or D inducing.
This hetero or homo grouping of the various series permitted us to

106 /

xESEARcHrN pHysropATHoLocy

make other correlations.We could seethat among all the seriesdesignated

as A in the periodic chart, those numberedoddly are hetero type, while
thosewith even numbersare homo type. Among the B series,the opposite
is true; those with odd numbersare homo type, while thosewith even are
hetero type. Extrapolating, we could classify all the series according to
this criterion. This view was confirmed by the hetero character seen in
Cr and especiallyMo, and the homo characterfor Mn.
We could go farther and correlatethe aboveclassification,made on the
basisof biologicalproperties,with the electronicconfigurationsof the elements. For the members of the A series,those with an odd number of
electronsin the valency shell were hetero type, while those with an even
number were homo type. Among the B series,this criterion did not hold
true. Wc found, howevcr,that a similarcorrelationexistedif consideration
was given not to the valencyshell alone but to the sum of the two external
shells,the valency and the shell beneathit. This accordswith the fact that
in the B serieselements,the two shellshave insufficientelectronsto frrlfill
the quantum numbers. we saw thus that those membersof the B series
with an odd number for thc sum of elcctronsof the two shellshave a homo
character,while those with an even number havc a hetero. This criterion
appliesto all membersof the B group, includingthose in the I B and II B
series,which have their full quantumquota of electronsin the shell beneath the valency shell. This samecriterion was used to classifythe three
subseriesof seriesVIII shown in the chart. The Fe subseriesand Ni subserieswere consideredhetero type, the Co homo type. This antagonism
was seento be in accordwith experimental
findings.
These considerationsalso permitted us to classifythe membersof the
Lanthanium and Actinium Series.Characteristically,all show nonfulfillment of three of their electronshells-the valencyand the two shellsbeneath.We established
for theseelementsa separateseriesdesignation,C.
Using thc sum of the electronsof the three shells,we separatedthe elementsof the C seriesinto hetero and homo categories.Here, the criterion
was the opposite of that used for the B scries.Thc members with odd
numbersof electronswere consideredhetero, those with even numbers
homo.
We will discusslater the biologicalsignificanceof this separationof
elementsinto A, B and C serieswith their respectiveone, two and three
unfulfllledshells.For the moment,we will only remark that if we consider
an even number of electronsas corresponding
to a kind of partial fulfillment of quantumforces,especiallyas comparedto an odd number, such
partial fulfi.llmentis seen,among all the A series,in those with even num-

rHE

CoNSTTTUENTS /

107

bers (II, IV, VI); amongall the B series,in thosewith odd numbers(I B,
III B, V B, VII B); and, amongthe C series,in thosewith alternatenumbers which can be consideredto correspondto even numbers. All these
series-with partial fulfillment for the sum of their shells-have a D inducing or anti A, character, and thus a homotropic tendency.
Periods
Coming back to the influenceexertedby the elementson test manifestations, we found that elementsof the same seriesshow some similar
propertieswhen acting at a certain level of organization,but some differencesappearwhen they are acting at difrerentlevels.Someof thesedifferences are important. Magnesium,calcium and strontium act similarly on
the s.d.c. pH, and againstconvulsionsas well. However, magnesiumhas
been found to induce somnolenceor even deep sleepin test animals,while
calcium immediately wakes them from magnesium-inducedsomnolence.
This type of antagonisticaction among membersof the sameseriessometimes apparsespeciallypronouncedbetweentwo consecutivemembersin
the series-for example,betweensodiumand potassium,magnesiumand
calcium,oxygenand sulfur, and sulfur and selenium.
Study of this "antagonism" has permitted us to recognizea specific
characteristic.When two elementsof the same seriesact upon the same
entity, one may substitutefor the other.Sodiummay replacepotassiumin
cells.Magnesiumand calcium.oxygenand sulfur,and sulfur and selenium
can replaceeachother in this kind of reciprocalactivity.There is no truly
antagonisticaction betweenthem. This explainsthc fact that two elements
of the same series,if in sufficientamount can have similar activity at a
given level-that of the tissular,for instance,where the changesof the
s.d.c. pH take place.
Further analysisof the activityof membersof the sameserieshas revealedanotherimportantcharacteristic
which has permittcdfurther classification. Differencesin activity of membersof the same seriescould be
related to the organizationalcompartmentsinvolved.This becameclear
when activity of members of the I A series was analyzed according to
whether these elementsform constituentsof the metazoic,nuclear or
subcellularcompartments.
Sodiumis the predominantcation of the metazoiccompartment,which consistsof the interstitialfluids,lymph and blood.
Potassiumis the principalcationof the cellularcompartment.Ammonium,
which corresponds in most of its propertiesto rubidium, representsthe
cation at the nuclearlevel.It could be seenthat the developmentof hier-

108

nEsEARcH rN PHYSToPATHoLocY

archic organizationhas involved elementswith progressivelysmaller atomic

weights.
Study of constituentsin compartmentsand in the environment has
further permitted us-as seen above-to correlate the metazoic compartment with the sea, the cellular compartmentwith the crust of the earth,
and the nuclear and subnuclearcompartmentswith the formations in
which their constituentswere found in the vicinity of volcanoes.
This correlation of the metazoiccompartment to the environmentof
the sea becameespeciallyinterestingwhen we could recognizein its constituentsnot only sodium,but, curiouslyenough,the other membersforming the sameperiod in the periodicchart. Chlorine, magnesiumand sulfur,
predominantin the sea and also found in the metazoiccompartment,are
in the same period in the chart as sodium.
We have thus tried to extend the conceptof a correlationbetweenthe
periods of the chart and the different compartmentsof hierarchic organization. Tentatively we correlated the second period of the chart to the
total organism as an entity. Oxygen, carbon and nitrogen-principal elements in air----enterinto direct contact with the organism as such. The
third period contains sodium, magnesium,sulfur and chlorine, which are
found in the sea and can be correlatedwith the metazoic compartment.
The fourth period containspotassium,calcium, iron, nickel, copper,selenium and bromine-all common to the earth's g1u5t-3nd, according to
our tentative systematization,correlated with the cellular compartment.
Following the same plan, we could relate the fifth period-<ontaining
rubidium, molybdenum,silver, tellurium and iodine-to the nuclear compartment.
As a possiblebasis for a working hypothesis,we could consider the
sixth period-with cesium,barium, gold, mercury, lead, bismuth-and the
lanthanium series to belong to a subnuclearor, rather, submorphologic
compartment.The seventhperiod includesthe radium and actinium series,
characterizedby radioactivity. This period could be related to the lowest
level of the biological organization,the primary one, probably even the
submolecularlevel. This would relate the interventionof radioactivityfrom cosmic rays and especiallyfrom the earth's radioactiveslsrnsnt5-ts
the beginningof the biological realm. Such radioactiveinterventioncould
have brought togetherC and N to form N-C-N-C, which we consideredin
our hypothesis to be the fust entity in the biological realm. This view
represents,at least,a new basisfor an interestingworking hypothesis.
Thus we have the conceptof hierarchiccompartmentsrelatedto changing environments.We also can correlate,further, the environmentsto the

THE CoNSTtruENrs

lo9

periods in the periodic chart to which their principal constituentsbelong.

It is difficult to accept as purely accidentalthe correlationof the changes
in environments with the progressivedisplacementof their constituents
toward periods in the chart with memberseach time having lower atomic
weighs. It is in this progressionthat we can see homotropy developing
toward its maximum, complete value. This view, which will be discussed
in more detail later, again relates evolution of the biological realm to
progress of homotropy, with the cnvironment representingthe concrete
realizationof homotropic evolution.
.
The entire chart can be consideredin terms of hetero and homo series
and of periods that correspondto hierarchiccompartments,as shown in
Tenle V.
For the moment, the possibility of relating an element, tfuough its
membershipin a series,to the hetero or homotropic trend and, through
its place in a period, to an organizationalhierarchiccompartment,helps
to explain many of the peculiaritiesseen in the biological distribution of
the elementsand especiallyin the role played by them at the "proper"
levels to which they belong.
As a generalrule, the presenceof an elementat the level to which it
belongsis directly correlatedto quantitativeoptimum valuescorresponding
to the constantsof the level and to the qualitativerole which it performs.
lts presenceat levelsother than its own must be interpretedin connection
with its activity at its own proper level. Increaseor decreasein the amount
o[ an elementhas a different meaningaccordingto the level at which the
variation occurs. If it occurs at the specificproper level to which the element belongs,it would indicate a direct quantitativeor qualitativechange
in the activity of the element.At other levels,this is not true. If the activity
of an element is qualitatively impaired at its own level, the amount of
the elementat the immediatesuperiorlevelwill increase.The increaseat the
superior level can be interpreted as taking place in order to keep at the
disposal of the impaired level a sufficient amount of the element for possible later use. On the other hand, an abnormally intensive activity of an
element at its own level will reduce the amount of it present at the level
immediately superior. The decreaseat the upper level can be interpreted
as a defensiveattempt to reduce the abnormal activity by limiting the sup-

plv.
The generalrule, which appearsto governthe variationsin distribution
of an elementwithin the organism,makesit important to know the proper
level of an element. Some exampleswill illustratewhat we mean, An increaseof copper is seen in the blood serum of cancer patients,althougb

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THE CoNSTrruENTs

lll

a manifest reduction in catalaseas well as in copper content is seen in

the tumor cells themselvesand in the liver cells. According to the view
presentedabove, these findings can be interpreted to reflect a primary
insufficiency
of copper at its specificlevel, that of the cell. copper is quantitativelydeficientat the levelof the cell not becauseof its low availability,
but becauseit cannot be utilized well enoughqualitativelyto form catalase.
The qualitativeimpairmentin copper'suse at its proper level would lead
to an increasedamount of copper in the immediatelysuperiorcompartment,that of the blood serum.
The knowledgethat copper belongsto the cellular levcl, becauseof
theperiod to which its belongs,could explainthis peculiarity.The organismdoesnot have too much copper althoughthe amount of it in the bloo<J
is incrcased.
Neitherdoesit have too little copperat the proper level.The
abnormalityresidesin a qualitativelyimpairedcapacityof abnormalcells
to utilizecopper.In tentativctherapeuticapplication,we haveto try neither
to increase
nor decrease
the amountof copperbut to obtainits proper utilizationby the abnormalcells.
This view appliesalsoto potassium.
An increaseof potassiumin blood
serumis seenin subjectswith type D offbalance.
with potassiumbelonging
to thecellularlevel,the primary abnormalityhas to be soughtat this level.
In fact such a primary anomaly of potassiummetabolismis seen in the
cellsfor, in oflbalanceD, the cellsare poor in potassium,possiblybecause
thecationmovesout of the cellsas the resultof beingdisplacedby sodium.
The increasein the amount of potassiumfound in the circulatingblood
thuscan be interpretedas secondary,designedto offer the cells a sufficient
amountof potassiumto be utilizedin attemptsto overcomcthis offbalance.
on the other hand, in abnormal offbalanceA, when quantitiesof potassiumare found presentin proliferatingcells,an abnormallyIow amount of
potassium
is found in blood (as low as 3.0 m Eq or less). As potassium
is stillexcretedthrough the kidney, this low blood potassiumis not to be
interpreted
as a quantitativedeficiencybut rather as a teleologicalresponse
to the abnormallyhigh utilizationat the cellularlevel.
A study of potassium,presentedunder this aspect,is thc subjcct of
Note2.
Thc relationshipbetweenelcmcnts,pcriodsand levelsof the organizationexplainsa curious distributionof elementsas seen in the following
e x p e r i m e nlt/.1 0 m o l a r s o l u t i o n sw i t h p H , o f d i b a s i cp h o s p h a t eosf l i t h tum,sodium, potassium,ammoniumand rubidium were prepared.Each
solution
was injectedintravenouslyinto mice, l/4 cc. per minute, until
theanimaldied. The organs,and especiallythe brains,werc immediately

ll2

RESEARcH rN PHYsroPATHoLooy

fixed in Bouin solution and studied histologically. For ammonium and

rubidium, vacuoleswere seen presentin cells and especiallyin nuclei; for
potassium,the vacuoles were present in the cytoplasm, while for sodium,
only a pericellular edema was noted. No such changeswere observed for
lithium. Considering the diiension of the atoms, an opposite occurrence
would have been expectedwith lithium penetratingmost into the cells and
rubidium the least. The fact that the heavier elements correspond to the
lower levels of the organization,accordingto the concept presentedabove,
explains the occurrence.
This could also be seenfor the distributionof seleniumand tellurium.
We could show that while seleniumaccumulatesin the cytoplasm,tellurium
-which is the next heavier elementof the VI series-is fixed preferentially
in the nuclei.
It must be recognized that many problems result from exaggeratedor
reduced amounts of elementsat compartmentswhere they do not belong
as characteristicconstituents.The therapeutic effort, until now, has been
to try to eliminale an excessor make up for a deficiency at any level. According to the view presentedabove, the main effort should be to try to
correct the anomaly in the metabolism of the element at its proper Ievel
for this will lead to correction at other levels. we will consider, later, some
examplesof such eftort.
The concept of dualism and of the place of elementsin hierarchic organization has opened a new way to study the influence exerted by these
elementsin abnormal conditions as will be presentedbelow.
The same type of analysis used for elements can be applied to the
other body constituents.we will start with those which we believe to be
the most important for the problem of the offbalances,the lipids,

CHAPTER

LIPIDS AND LIPOIDS

1r
pATHwAys,one theoreticaland the other experimental,
lLrvo DTFFERENT
have led us to considerlipids as possiblythe most important constituents
involved in thc dualisticpatternsof physiopathological
manifestations.The
study of all the constituentsof the organism--electrolytes,
proteins,carbohydrates and lipids-has shown that for each of them, a rough division
into two classeswith antagonisticreactivitycan be made accordingto the
positive or negativeelectrostaticcharacterof their polar groups-nucleophilic or electrophilicfor some,anionic or cationicfor others.However,
these fundamentaldifferenceswhich can explain thcir interventionin
processesin which dualism is apparent,do not representthe reason for
their rolc in the inductionof patterns.
The reactionsin which someof theseconstituents
take part are carried
out as rapid changeswhile others are complet.,ed
only slowly. It is these
slow rcactions,once accomplished,
which tend to be stablefor long periods
of time. Sinccsuchstabilityis characteristic
of clinicaland analyticalmanifestationswhich have dual patterns,it appearedlogical to considerthe
with slow reactivitywhich are relatedto thesemanifestations.
constituents
Becauseof their hydrosolubility,
and the rapidityof the reactionsin which
part,
proteins
take
most
they
electrolytes,
and evencarbohydrates
probably
play a lesserrole in theselong lastingprocesscs.
The lipids, on the contrary,seemto be especiallysuitedfor this role.
Many of the reactionsin which the lipids participateare slow. As we will
seelater,this is primarily becauseof their insolubilityin water.They form
in the organisma group "apart" from all the water solubleconstituents,
a fact which permits them to function through proper reactions largely
without continuous interferencefrom thc other constituents.For these
reasons,the lipids appearedto be the most likely of all constituentsto be
lt3

114 /

R E S E A R c HI N P H Y S t o P A T H o L o c Y

of major importance in physiopathologicalmanifestationswith long-lasting

patterns.The study of the lipids has substantiatedthis.
However, before discussingthese substancesand their properties, a
nosologicalproblem must be considered:What are lipids? How can they
be defined?
DEFINITION

OF LIPIDS

The literature fails to furnish an adequatedefinition for the group of

that show thosepropertieswhich biochemistryand experimental
substances
biology attributeto the lipids, A definitionon a chemicalbasis,such as one
which considerslipids to be fatty acids and fatty acid derivatives,appears
to be insufficient. It excludes substancessuch as those forming insaponifiable fractions which not only have properties attributed to lipids, but
continuouslyintervenein the processesrelated to them.
such as "greasiness"and solubility come nearer
Physicalcharacteristics
to the real situation without providing a satisfactorydefinition. Bloor's
definition (Note /J, widely acccptedtoday in spite of having been found
inadequate,has introduced-in addition to the important solubility characteristics-certain less acceptable criteria such as the origin of these
substances
and the direct relationshipto fatty acids.Without thesecriteria,
lipids would have to include the group of hydrocarbonswhich have the
same solubility property but usually are not encounteredin organisms.
However, with these criteria, Bloor's definition, besideslimiting the field
too much through the requirementfor a relationshipto fatty acids, excludes the entire important group of synthetic agents with similar properties.To be complete,a definition would have to include these artificial
substances.
Thus confronted by the need for a satisfactorygeneral definition, we
proposedone in 1940 (23) which has since been of great help to us in
all our research: a Iipoid is a polar-nonpolar substancein which the nonpolar part is predominant. It is thus lormed by one or more polar groups
bound to one or more nonpolar groups, the last being energeticallypredominant. In terms of intervening forces, this definition considers the
cohesionforces of the nonpolar part, and especiallythose related to its
surfaceand known as the constant"b" of van der waals forces, which in
lipoids are predominant upon the electrostaticforces of the polar part.
The definitionhas provided the key for the study of the multiple problems
in which these substancesappear to be involved. This definition appears
acceptablesinceit explainsall the known proprtiesof the lipids. Further-

LTPTDSAND

LTPOTDS

ll5

more,the study of the specificrelationshipbetweenthe forces involved

couldevenpredictnew properties,
as will be shownlater.
The distinctionbetweennaturaland syntheticsubstances
as a basisfor
a definifioahasbeenobsoletein biochemistryfor a long time. In our study
however,
it appearedto be didacticallyusefulto indicatewhetheror not
a substanceis encounterednaturally in the organism.Therefore,while
adheringto our generaldefinition,we haveemployedthe term "lipoids"
for the entiregroup of polar-nonpolarsubstances
with a predominance
of
thenonpoliupan, and have conservedthe term "lipids" to designatethe
naturallyoccurringmembers.With this separation,
we havealso avoided

Hydro
id
Borderl
ine
Lipoid

FN\\\
FI\\\\\\\\\

eot.rGroup

Group
Sl llon-Polar

Ftc.6lbis. Schematic representationof the predominant relationship of the polar

and nonpolar parts in hydroids, borderline substancesand lipoids.

a certain apprehensionfelt by many workers about incorporating indistinctly, in the same group of agents,substanceswith vastly different chemical constitutions,which until now have not been associatedwith lipids.
The fact that, beyond physicochemicalconstitution, biological properties
characterizingthe lipids are common to the entire group of lipoids, will
in time, we hope, help to reduce the importance of this separation betweenlipoids and lipids.
The structure of the lipoids-with a large variety of polar and nonpolar
Sroupsbut always with the same characteristicenergeticrelationshipbetweenthem-has led to a logical systematizationof thesesubstances,using
thenature of the polar and nonpolar goups as criteria.
CLASSIFICATION OF LIPOIDS

Lipoids may be subdividedaccordingto differentcriteria.

l. According to the polar group.
A. Lipoids classifiedaccording to the nature ol their polar group.
( -COOH)
1. Lipo-carboxylic acids
(-SH)
2. Lipo-thiols
(_sofi)
3. Lipo-sulfonic acids

l16

nESEARcH rN pHysropATHoLocy

4.
5.
6.
7.
8.
9.
10.

Lipeamines
Lipo-amides
Lipo-alcohols
Lipo-aldehydes
Lipo-ketones
Lipo-halogens
Lipo-metals
etc.

( -NH.)

-coNH,)
-oH)
-cHo)

(:CO)
(-Cl, etc.)
( -Na, etc.)

B. Lipoids classified according to the predominantelement ol the polar

group.
l. Lipo-sulfur compounds
a. Lipo'thiols
b. Lipo-sulfonicacids
c. Lipo-sulfides
d. Lipo-sulfoxides
e. Lipo-sulfones
f. Lipo-sulfites
etc.

(-SH)

( -so"H)
(:S)

(-so)
(:SO,)
(:SO")

2. Lipo-nitrogenderivatives
a.
b.
c.
d.
e.

Lipo-amines
Lipo-amides
Lipo-nitriles
Lipe.isocyanides
Lipo-nitro derivatives
etc.

(-NH,)
(-CONH,)
( -CN)
(-NC)
( -NO,)

C. Lipoids classified according to the energetic character ol their polar

8roup.
A. Lipoids with negativepolar groups
l. Lipoacids
a. Lipo-carboxylicacids
b. Lipo-thiols
c. Lipo-sulfonicacids,etc.
2. Lipo'atdehydes
B. Lipoids with positivepolar groups
3. Lipobases
a. Lipo-amines
b. Lipo,guanidines
c. Lipo-imines,etc.
4. Lipo-alcohols
ll. According to the nonpolar group.
A. Lipoids classifiedaccording to the structure ol their hydrocarbon chain.
l. Aliphatic
2. Alicyclic
3. Aromatic
4. Heterocyclic,etc,

L t P t D s A N D L r P o t D S

l 1 7

accordingto theircarhttnbond.s.
B. Lipoidscla.ssified
l. Saturated
2. Unsaturated
a. Ethenic(mono-,di-,poly-)
b. Ethynic
Someaspectsof this classification
requirediscussion.
The genericterm lipoacidhas beenemployedto describesimplelipoids
having polar groupswith acid functions.While the principallipoacidsare
the fatry acids,other membershavc other acid polar groups,such as SO3,
SH, NOr, etc. The significanccof this groupingtogethcrof lipoids with
negativepolar groupshas becomeevidentespeciallyin studyingthe similaritiesin the biologicaleflectsof thcsesubstanccs.
In certain aspectsof
our research,this correlationhas pcrmittedus to substituteone category
of lipoids (lipo-thiols or lipo-aldehydes) for another (lipo-carboxylic
acids), therebyavoidingcertain undesirableeffectsof the latter group of
substances.
Lipoids havingpolar groupsenergetically
oppositeto thoseof the acids
have been grouped togcther.O[ thcse,the memberswith a polar group
with alkalinefunctionshave becn classifiedas lipobascs.The tcrm baseis
generallyappliedto ionizablecompoundswhich influcncethe pH of solutions and combine readily with acids by losing an OH and gaining a
proton. Another group is formedby the lipoidicalcohols.Recentevidence
indicatesthat, in miiny circumstances,
the differencesin the reactionsof
alcoholsand common basesare quantitativeratherthan qualitative.Quite
often the reactionof an alcohol with an acid is analogousto the reaction
befweenan acid and sodium hydroxide,thc H' of the acid combiningwith
the OH- of the alcohol.The differcnces
betweenreactionsare considered
to be mattersof time-rate.Whereasthe reactionof the baseis almost instantaneous,
that of alcohol is a slow reactionand is less complete.This
behavior of alcoholic substancesis particularly clear when the hydroxyl
group of an alcohol is replacedby a halogento preparethe alkyl halides.
For instancc,accordingto Karrer (24): "This can be done by the action
halogcnacidson the alcohol:
of the concentrated
CH3OH + HCI -----> CHrCI + H2O
The reaction correspondssuperficiallyto the formation of a salt from an
acid and a base:
NaOH + HCI ------>NaCl + H:rO
There is, however,a differencebetweenthe two processes.
Basesand acids
are largelydissociated;when thcy come together,hydrogenions and hy-

ll8

,/

R E S E A R c HI N P H Y S t o P A T H o L o c Y

droxyl ions combine almost at once to give the very little electrostatically
dissociatedwater, so that the reactionwhich really occurs is:
Nal * OH- + H+ + Cl- -----> Na+ * Cl- + HrO
Ionic reactionsalwaysoccur instantaneously.
Reactionbetweenalcohol
and hydrogen halides is governed by other laws. Alcohol is only very
slightly ionized. For the removal of the hydroxyl group a certain time is
required.The reactionbetweenalcoholand acid with eliminationof water,
known as esterification.
is thereforea time reaction."No essentialdifference exists between the reaction of lipo-alcoholsor lipo-amines,for instance,with organicacids.
Theseconsiderationswould have been sufficientto allow lipobasesand
lipo-alcoholsto be grouped together.There are othcr considerationsas
well. Their common biologicalactivity and mutual interchangeability
also
justify
appcar to
grouping them together.Furthermore,the recognitionof
the existence
of a generalmutual antagonismbetwecnlipoacidson the one
hand and lipobasesand lipo--alcohols
on the other hand, chemically,physically and biologically,has proven of considerablevalue in explaininga
variety of experimentallyobservedfacts in many aspectsof our research.
Following this through, we have found it advantageousto define the
two groupsof lipoids by a more generalcharacter,the electricalaspectof
the polar part, negativcfor the lipoacidsand lipo-aldchydes
and positive
for the group of lipo-alcoholsand lipobases.The terms,"positive and negative lipoids,"servealso to emphasizethe natureof their antagonism.
The structureof thc nonpolargroup as it confersphysical,chemical
and biological propertieson the lipoids permits further subdivisions.Lipoids may be classifiedon the basisof thc a.liphatic,alicyclic.aromaticor
heterocycliccharactcrof the nonpolar group. While the negativelipids
are principally formed by fatty acids,the positive are made up principally
of sterols.The prcsenceor absenceof doublebondsdefiningsaturatedand
unsaturated
carbon chainshas becn one subjectof our study and considerablebiologicalimportancehas bcen found to be relatedto this character
as well as to the positionalrclationshipof the double bond to the polar
group and the polarity inducedby the doublebond.
The study of lipoids has shownthat, bcsidespropcrticscontributedby
the elementsand groups which compose thcm, they have additional
physico-chemical
and cven biologicalpropertieswhich are characteristic.
We have termedthcse"lipoidic propcrties"to indicatethat they are consideredto resultdirectlyfrom the particularconstitutionof the lipoids.

LIPIDS AND

LIPOIDS

II9

Physical Lipoidic Properties

Solubility
Solubility representsthe first and most important of thcselipoidic prop
erties.Characteristically,a lipoid has a greatersolubility in neutral solvents
than in water.This is explainedby the fact that the two constituentgroups,
polar and nonpolar,inducediffercntsolubilitypropertics,
to a frec movementbetwcen
As is well known, solubilitycorresponds
moleculesof the solventand the solute.(25) Solubilityis greaterwhen
the physicalpropertiesof the groupsformingthe solventand thoseforming
Lhesolutearc similar;it is impairedwhcn thcy are different.Consequently,
polar groupsin a solutewill tend to favor solubilityin solvcntswith polar
groups, such as water. At the same time, thcy will oppose solubility in
neutral solventsformed by nonpolargroups.On thc othcr hand, nonpolar
groupsin a substancewill favor solubilityin nonpolarneutralsolventsbut
will opposeit in polar solventssuch as watcr. Polar groups thus are hydrophilic and lipophobic,while nonpolarare lipophilicand hydrophobic.(26)
While the solubility characteristics
of substances
composedonly of
polar or nonpolargroupsare rcadily apparent,thc problcm is more complex when a substancecontainsboth polar and nonpolar groups. Since
groupswith antagonistic
such a compoundpossesses
solubilitytendencies,
"in
toto" will dcpendupon thc relationshipbctweenthe opits solubility
posing forces. For a borderline group of polar-nonpolarsubstanceswith
approximatelyequalforces,therc will be cqual solubilityin polar and nonpolar solvents.For other substances,
thc predonrinancc
of one or the other
group will determinesolubilitycharacteristics.
If thc electricalforces of
the polar group predominatc,the substanccwill be hydrosolublebut insolubleor only partly solublcin nonpolarsolvent.s.
If, on thc contrary,the
cohesion-i.e., the van der Waals forccs-of the nonpolar group predominate, the substancewill be solublein nonpolarsolventsand less,or even
not at all, solublein water. (Tnnn VI)
Polar and nonpolarforcescan be calculatedand thcir study can indicate the place of a substancein this systematization.
The importanceof
solubility for defining and systematizing
lipoids became apparent in a
physicomathematical
study of thescsubstances
carried out by Jean Mariani
in our laboratories.(Note 2) We havc defined as "hydroids" those substanceswith predominant polar groups which are morc soluble in polar
with no predominance
solventssuch as water.The "borderlinesubstances,"
of either group, show the same solubilityin polar and neutral solvents.

120 /

R E S E A R , C HI N

PHYSIOPATHOLOGY

TrsLe VI
CursstntcerroNoF Cueulc,rl Coupouxos
Composition

Predominance

Name

Polargroupsonly
Polar-nonpolar
groups

Nonpolar groups
only

Example
Water

Polar group
predominant

Hydroides

Glycerin

No predominance Borderline
substances

n-Propyl alcohol

Nonpolargroup
predominant

Oleic acid
n-Butyl alcohol

Lipoides

Paraffin

The "lipoids," in which the nonpolar groups predominate,are more soluble in neutral solventsthan in water.
As we have mentionedabove, from a practical point of view, a substancecould be judged to be a hydroid, borderlinesubstance,or lipoid by
consideringthe differencesin its solubility in water and in a nonpoliu solvent, such as petroleumether, which correspondsto a mixture of the first
aliphatic saturatedhydrocarbonsliquid at normal temperatureand pressure. ,{ polar-nonpolar substancemore soluble in water than in neutral
solvent is considered a hydroid; one equally soluble in both solvents is
classifiedas a borderline substance;w,hilea substancemore soluble in the
neutral solvent than in water is a lipoid.
Difterent polar groups such as COOH, OH, NH2, CO, SOz, SH, etc.,
enter into the constitutionof various lipoids. They differ considerablyin
their electrostaticforces. As a result, the forces of the nonpolar goups
required for predominance,if a lipoid is to be formed, also will difter. A
different nonpolar group thus is necessaryfor each different polar group.
For aliphatic molecules,it is principally the length of the chain which
determinescohesionforces and a differentnumber of carbons in the nonpolar group appearsto be necessary,dependingupon the polar group, in
order to form a lipoid. The study of homologousseries from this point of
view is interesting.
Sincethe value of the electrostaticforcesvariesgreatly from one polar
group to another, the first members of the various homologous series.
which are also lipoids, will differ from seriesto series,dependingupon

L T P T D S^ N D L t P o t D S

l2l

the nature of the polar group. The lengh of the carbon chain of the nonpolar group will thus indicate in what member of a series the lipoidic
characterappears.By comparingmathematicallythe value of the electrostatic forces of each polar group and the cohesionforces of the nonpolar
group in the respectiveseries,it is possibleto determinewhich member of
each homologousseriesof substanceswill first show the properties of the
lipoids. This a.lsocan be determinedexperimentally,as seen above, using
the solubility characteristicsof the lipoids. For the different membersof
the series,degreesof solubility in a polar solvent such as water, and in a
nonpolar solvent such as petroleum ether, were determined.The first member of an homologous series to be considereda lipoid was the one found
to be more soluble in the nonpolar than in the polar solvent.All members
with a large number of carbon atoms show lipoidic properties;those with
fewer carbon atoms lack those properties.
Thus, lipoidic properties first become manifest, among the carboxylic
acid series,in valeric acid,i.e., the five-carbonmember.The shortercarbon
chain membersare soluble to an equal or greaterdegreein water than in
petroleum ether, while those having a carbon chain longer than four show
a higher degreeof solubilityin the nonpolarsolventsthan in water. (Tnnr-E

vlr)
TesLe VII
Sor,usrLlrresor CrnsoxyLtc Aclo HovolocuEs

ility in

Substance'

Common Name

Methanoic acid
Ethanoic acid
Propanoic acid
Butanoic acid
Pentanoic acid
Hexanoic acid
Heptanoic acid
Octanoic acid

Formic acid
Acetic acid
Propanoic acid
Butyric acid
Valeric acid
Caproic acid
Enanthic acid
Caprylic acid

Polar
Solvent
( W a t e ra t 2 0 " )
I
2
3
4
5
6
7
8

Nonpolar
Solvent
( Petroleum
Ether)
insol.

cO

3 . 7( a t 1 6 " )
0.4
o.24

0 . 2 5( a t 1 0 0 " )

' Namesapproved
by International
Unionof Chemistry.
The sameis true for the trJkylalcohols.n-Propyl alcohol and the members below it are either miscible with both water and petroleum ether or
more soluble in water, indicatingthat the nonpolar forcesdo not predomi-

?.:.]l,:'j

\jii;

122

RESEARcH rN pHysropATHoLocy

natein their molecules.

Therefore,they are not lipoids.n-Butyl alcohol,
moresolublein neutralsolventthan in water,thus is the first lipoidic member of this homologousseries.However,this is not true for all its isomers.
The primary,secondaryand iso butanolare the first in their respective
seriesto possessthe solubility propertiescharacteristicof lipoids. In the
tertiaryalcoholseries,however,the four+arbonmember,the tert.-butanol,
doesnot showthe samesolubilityproperties.Tert.-butanolis misciblewith
water and neutralsolventand as such,is not a tipoid. For this tertiary
alcoholseries,it is the five-carbonmember,the tert.-amylalcohol,which
first showsthe solubilitypropertiesof a lipoid,being only 12.57osoluble
in waterandinfinitelysolublein petroleum
ether.Thus,of the four isomers
of butyl alcohol,three are lipoids,while one, tert.-butylalcohol,is not.
( Trn l n V III)

Trnle VIII
Sor-uslI-rrresoF THE Ar-xyr- Alconor-s
7o of solubilitvin

Substance'

Conrmon Name

Methanol
Ethanol
I -Propanol
2-Propanol
l-Butanol
2-Butanol
2-Methyl, 2-propanol
2-Methyl, I -propanol
l-Pentanol
2-Pentanol
3-Pentanol
2-Methyl, 2-butanol
2-Methyl, l-butanol
3-Methyl, 2-butanol
l-Hexanol
2-Hexanol
3-Hexanol
l-Heptanol
l-Octanol

Methyl alcohol
Ethyl alcohol
Propyl alcohol
Isopropyl alcohol
n-Butyl alcohol
sec.-Butylalcohol
tert.-Butyl alcohol
Isobutyl alcohol
n-Amyl alcohol
sec.act. Amyl alcohol
Diethyl carbinol
tert.-Amyl alcohol
n-act. Amyl alcohol
Isoamyl sec. alcohol
n-Hexyl alcohol
sec.-Hexyl alcohol
Ethyl propyl alcohol
n-Heptyl alcohol
n-Octyl alcohol

No. of
Carbon
Atoms

Polar
Solvent
(Water
at 20")

Nonpolar
Solvent
(Petroleum
Ether)

I
2

4
4
4
5
5
5
5
5
5
6
6
6
8

r Names approved by International Union of Chemistry.

7.9
t2.5
@

9.5
2.7
5.3
insol.
t2.5
insol.
sl. sol.
very sl. sol.
very sl. sol.
0.9
insol.
insol.

co
oO
co
oO
@

co

&

*
t

*
cc

r
co

LrPrDsAND LrPorDs

123

The same methods were used to recognizethe first Iipoidic membcrs

of various alkane derivativesstudied.-fnnle IX shows the first lipoid
membersof severalhomologousseries.
T,rgls IX
Flnsr Lrpololc MevneRs lN Vrnrous AlxeNr' Denrv,rrrve
Hovor-ocous SE'Rrr.s
S u b s t a n c eI

Conrnron Name

N{ethanethiol
Propanal
Propylcarbylamine
l-Butanol
2-Butanone
Butanamide
2-Methyl, 2-butanol
Pentanoicacid (n)
H e x y l a m i n e( n )
l, I Octandiol

Methyl mercaptan
Propionaldehyde
Propyl isocyanide
n-Butyl alcohol
Butyl ketone
Butylanride
tert.-Amyl alcohol
Valeric acid
Hexylamine
l. 8 Octandiol

Polar
Group
-SH
-CHO
-NC

-oH
:CO
-CONH.

-oH
-COOH
-NH'

-oH

i
]

No. of
Carbon Atoms
l
J
J
A

,
A
a

5
5
6
8

t N a m e sa p p r o v e db y I n t e r n a t i o n a l
Union of Chcmistrv.

Molecular l^oyer Formation

Other fundamentalcharacteristics
result from the different solubility
propertiesof the two parts, polar and nonpolar, forming a lipoid. Introducedin a diphasicmediumin which one phaseis watcr and the other oil
or even air, the polar group, with the tendencyto be solublcin water, will
penetratethe water. Since the nonpolar group, which is hydrophobic, is
predominant,not only will it not cnter the water but it will also prevent
theentire moleculefrom moving frecly in water.Conscquently,
the lipoid
moleculewill remain at the surfaceof the water with only its polar group
penetrating.
Becauseof this, the moleculewill assumean oricnted position
towardthe surfaceof water. If the secondphaseis a neutral solvent,the
lipoidmolecules,which will accumulateat the interphasewith the polar
group in water, will have the nonpolar group penetrating the ncutral
solvent.
In both cases,the moleculesform oricntedmolccurarlayerswhich, if
presentat the limit betweentwo phases,would appearas organizedformations.This property which appearsas a direct consequence
of the characteristic
constitutionof the lipoids has further consequcnces.
In such a
layer,the polar groups penetratingin water will influencethe properties
of its surface,and thus reduceits surfacetension.

124 /

REsEARcHrN pHystopATHoLocy

Through the coulombiancharacterof their electrostaticforces,the polar

groups will thus confer, accordingto their nature. a positive or negative
electricalcharacterto the layer.
In a mixture of water and oil, the presenceof a lipoid layer will lower
the intersurfacetension and will favor the breaking down of the phases,
facilitatingthe formation of an emulsion.The presenceof the same electrical chargeat the surfaceof theseresultingemulsiondroplets will act as
a repeUentforce betweenthem and increasethe stability of the emulsion.
(FiS.62) This is anotherimportantcharacteristic
of lipoids which results
from their peculiar constitution.

F t c . 6 2 . T h e p r e s e n c eo f t h e s a m e e l e c t r i c a lc h a r g e a t t h e s u r f a c eo f d r o p l e t so f a n
emulsion insures,througb the repellent forces, the stability of the emulsion.

Chemical Properties
Lipoids have two groups of chemicalpropertieswhich can be related to
their two principal parts, polar and nonpolar. The polar groups with their
electrostaticforces give to the lipoids one group of characteristicreactivities. A carboxylic lipoid will act like any other organic acid, while the
lipo-'alcoholswill act like other alcohols,a thiolipoid like a mercaptan,
and so on. A characteristicof chemical reactionsinduced by the polar
groupsis that, while they are occurringin a water medium, they are largely
limited to the site where the lipoid is localizeddue to the insolubility of

LtPtDS AND LtPOtDs

125

the entire molecule in water. Through this localization, the polar reactivity
of the lipoids becomeslargely a "surface reactivity." It is interestingthat
even minute amountsof lipids are able, through this localizationat separating surfaces,to induce important changes.
A second group of reactionstake place at the nonpolar group and
especiallyat the different formations presentin it, such as double bonds,
cycles, etc. The hydrophobic and lipophilic character of the nonpolar
groups confers a specialcharacteron thesereactions.Most of them take
place in nonionic nonpolar media. Many occur at the semipolar double
bonds with nucleophilicor electrophiliccarbonswhich appear to be especially suitablefor this reactivity. This would explain the fact thar nondissociatedmoleculesmay take part in thesereactions.Most of thesereactions
are relatively slow. This double reactivity,ionic through the polar group
and rather nonionic through the nonpolar group, makesthe study of these
Iipoids one of great interestand it will be discussedbelow in more detail.
Biological Properties
The biological properties of lipoids in general also can be related directly to their physiochemicalcharacteristicsand, thus, to their peculiar
constitution.
Lipidic System
The relative insolubility of the lipids in water and their solubility in
neutral solventshas permitted us to separatethesesubstancesas a group
from the other constituentsof organisms.For more than just didactic purposes,we considerlipids to constitutea separatesystemin the organism.
The part played by lipids in the organizationand the functioningof various entities supports this concept.For example,when a lipoid is introduced into the organism, it will be selectivelydissolved in, circulated
through, retainedby and metabolizedas part of the lipidic system.overtone's"Index of Repartition" of anestheticsin the organism can be seen
to be a direct corollary of the existenceof such a systemalthoughthe anestheticagent can be a lipoid or a nonpolar substance.
A great degreeof independenceof this system is morphologically evident as in adipous cells, when fats circulateas chylomicronsor when
they form oriented layers. We have seen above how the orientation of
lipoids at the surface of water results from the relationshipbetweenthe
solubilitiesof the two constituentgroups,polar and nonpolar. Along with

126 /

s,EsEARcH
rN pHysropATHoLocy

their insolubility in water, the orientation of lipoids has allowed them to

play a very important role in biology.
The very existence of biological entities appears to depend upon the
ability of lipids to build up boundary formations separating and thus assuring the individuality of biological entities.
Through peculiar, reciprocallyopposedorientations,two or more layers
of lipids can form a membrane with two polar faces which has the ability
to separatetwo aqueousmedia. In its simplestform, such a membrane
appears in mitochondria. (Note 3/ Similar boundary formations identify
nuclei and cells and appear in higher entities, as in the membranes and
intercellular cements of lymphatic and blood vessel endothelia. It is this
peculiar orientation which allows lipids to establishthe necessaryboundary
formations resulting in complex hierarchic organisms.The existenceof
biological entities, at least from the chromosome level up (and probably
even below that level), can be seento result directly from the intervention
of lipids as a separatesystem,particularlyin the formation of the dipolar
lipidic boundaries.
However, boundary formations which separatethe biological entities
would not have been efficient if they did not fulfill another capital role:
that of allowing selective passageof metabolites. A totally impermeable
membranewould isolate the respectiveentities and result in their death.
On the other hand, a totally permeablemembranewould have no usefulness.The boundaryformation has to act selectively,pcrmitting the passage
of some,but not all. substances.
But even this does not seemto be sufficient to insure an efficientboundary. Most important, such a membrane
must be able to alter its permeability,quantitativelyand qualitatively,according to variationsin circumstances.
Such capacityfor altering perrneability can be relatedto the presenceof the two groups of lipids, fafty acids
and sterols,with their antagonisticpropertiesrelating to permeability.
The fatty acids appear to induce permeabilityin the membranethey
form, especiallypermeabilityfor anions.The perpendicularposition to the
surfaceof water assumedby the nonpolar aliphatic groups when the fatty
acids form this boundary membraneappearsto be favorable for the passageof a substancethrough the membrane.The fatty acid moleculesthus
can be separated,permitting other moleculesto pass between them; that
is, to passthrough the membraneformed by the fatty acids.The negative
electricalcharacterof the polar groups of these fatty acids explainswhy
they representa kind of barrier to the free passageof cations.Thesecations
are attractedand retainedby the acid polar group. This would explain the
manifestchangesin permeabilityunder the influenceof calcium ion. The

LtPTDS AND

LTPOTDS

127

removalo[ calciumfrom cellularmembranes,

throughtrcatmentwith oxalates, increasespermeability,while trcatnrentwith calcium salts reduces
permeability.The bivalentcalcium ion, when it binds the polar groups
of two adjacentfatty acid moleculcsin the membrane,prcventsthe passage
of other moleculesbetweenthcse parts o[ thc membrane,a fact which
explainsthe manifestdecrease
in permeabilityinducedby this cation.
The other group of lipids, the sterols,have an effect on permcability
opposite to that of fatty acids. This can be related in part to the bond
rvhich these sterols make with the fatty acids. Consequently,they block
any passagethrough the part of the membraneformed by fatty acids.The
impermeabilityis due, to someextcnt, to a peculiarityof the layersformed
by thesesterolsthemselves,
The polycyclicmolcculesof sterolsdo not take
the same perpendicularposition toward the surfaceof water as fatty acids
d o . ( 2 7 ) ( F i S . 6 J l S i n c es t e r o lm o l e c u l eass s u m ea p o s i t i o na l m o s tp a r a l l e l
(a)

-ll+llllll
ooooooooc'

Fattyacidorientedlayr

ratgr

(b)

o
/z///,///,.

rater

oooooooo

Sterol orientedlayer

l.-

polar group
rrcn-polargroup

Ftc. 63. Schenraticuspcct ol orientcd interluce lu1'ers.The perpendicularposition to

t h e s u r f a c eo f w a t e r o f f a t t y a c i d m o l e c u l e s( a ) f a v o r s t h e p a s s a g eo f o t h e r m o l e c u l e s
t h r o u g h t h e s e p a r a t i n gm e m b r a n e st h e y f o r m . O p p o s i t e l y t, h e a l m o s t p a r a l l e l p o s i t i o n
t o t h e s u r f a c eo f w a t e r o f t h e p o l y c y c l i cm o l e c u l c so f s t c r o l s ( b ) p r e v e n t st h e p a s s a g e o f o t h e r m o l e c u l e st h r o u g h t h e m e m b r a n et h e y f o r m ,

to this surface,the layer which they form exhibits no permeabilityproperties.It evenopposesany passage
throughit. It seemsthat fatty acidsand
sterolsmake separate"spots" in the cellularmembranesso that, through
their quantitativerelationships,
they confer differentdegreesof permeability to differentregionsof the mcmbrane.The changesin permeability
which resultfrom the antagonistic
interventionof the two groupsof lipids
seemto play an important role in norrnaland abnormal physiology.
With fatty acids inducing permeability,and sterols opposing it, the
fundamentalcharacter of their biological relationshipcan be recognized.
It would seemthat part of the function of sterolsis to opposethe activity
of fatty acids.Conceptually,sterolswould appear,in this specificactivity,
to be "anti-fatty" acid agents,with a capacityto control the activity of fatty
acids rather than be active by themselves.
Partly for this reasonas well as
for greater general understanding,it is necessaryfirst to investigatefatty
acid activity.
'EllElt;tlf

[Ln[it8.AIf

128 /

nESEARcHrN pHystop^THoLocy

FATTY ACIDS
Besides their constructive role in cstablishingboundary formations,
fatty acids appear to serve various othcr purposesin the organism.They
can be used as caloric metabolites,and they play an active functional role
in a biological change. while all fatty acids may exhibit these three
activities---<aloric,constructiveand functional-there are important individual difterences.with the carboxyl as common polar group, the differencesbetween the various fatty acids can be related to the nonpolar
groups.we will discussthis aspectof fatty acids, emphasizingonly what
can be consideredto be new contributionsto understandingthe biological
role of the substances.
Rancidity
The srudyof changeswhich take placein virro, on lipids and especially
on fatty acids after they have been separatedfrom the organisms,led us
to consider a possible parallelism between them and the changes which
take placein the organism.We tried thus to utilize especiallythe knowledge
furnishedby the study of the chemicaldeteriorationof natural fats generallyknown as rancidity(28), to betterunderstandand alsoto systematize
many of the processes
occurringin vivo.
Three typesof rancidityare describcd.In onc-hydrolytic rancidityfats are separatedin free fatty acids and glycerol (or mono- or di-glycerides), through thc intervcntion of lipolytic enzymes. These are often
producedby molds (Penicillium,Aspergillus,ctc.) or by microbesrich in
suchlipolyticenzymesor cven thc lipasepresentin the tissuesfrom which
the lipids are obtained.]-he characreristic
of rhis typc of rancidityis the
interventionof enzymesand the appearance
of frce fatty acidsas a result.
In a secondtyp. of rancidity,also occurringunder the interventionof
enzymes,an oxidativeprocessis involved.Thc characteristic
of this typc
of rancidityis that it affectsaJmost,if not cxclusively,saturatedfatty acids,
convertingthem into methyl-ketones
by a bcta oxidation process.This
"perfume rancidity" called so becauscof the odor of the methyl-ketones
with seven,nine or cleven carbonswhich rcsult-takcs place apparently
through the interventionof a peroxidaseprescntin certain molds (such
as penicillium glaucum) , one of its characteristicsis that it occurs
e s p c c i a l loyn s a t u r a t e fda t t y a c i d sw i t h a l o w n u m b c ro f c a r b o n s( 8 t o l 2 ) .
The third type of ranciditygroupstogethcrthe oxidativechangeswhich
take placeat the unsaturatednonpolargroup of the lipids. As they result
from the interventionof double bonds, the reactionsdiffer accordinsto

L T P T D SA N D L t P O t D s

Lzg

the energeticcenterpresent.In one which occursat room temperatureonly

for the conjugatedfatty acids,such as eleostearicacid, and at 100"c only
to some extent for oleic, linoleic and linolenic acid, the oxidation leads to
thc appearance
of peroxides.(29) In anotherform of this oxidation,taking
place for oleic. linoleic and linolenicfatty acids at room temperatureor
below 50"c, hydroperoxidesresult, as it has been shown by Farmer and
corvorkers,
first for rubber (30) and afterfor fats (31). Another important
fact seen in rancidity changesis that the atmosphcricoxidation of polyethenoidfatty acidscan resultin a displacement
of the double bondswith
the appearance
of conjugatedisomcrs.(32)
The study of natural rancidityhas represented
the basicguide for our
study and systematization
of the processes
encounteredin normal and abnormal physiology. we searchedand found this similarity not only in
generaloutlines,but also for most of their details.By referringto the processesfound in rancidity, we were able to identify, besidesenzymaticlipolysis and enzymaticKnoop beta oxidation,known to occur in the organism,
also the interventionof hydroperoxides,peroxidesand the conjugationof
double bonds. Not only the processesthemselvesbut a.lsothe conditions
under which they take place and their inter-relationship
have been found
p
a
r
a
l
l
e
l
to
i n v i v o t h o s ew h i c h c a n b e s e e ni n v i t r o .
We rvill seeall along in the studyof fatty acidshow far the biological
interventionof this parallelismgoes.
Caloric Metabolism
Although all the fatty acidsarc ulrimatelyusedby differentorganisms
as caloric metabolites,the saturatedand monocthenicmembersare most
importantfrom this point of view. Among the saturatedand monoethenic
fatty acids,the memberswith long chainsappcar to be thosc which are
kept in reservefor caloric purposes.we could show that the principal form
of caloric desmolysis,
the Knoop beta oxidarion,takes place directly, almost exclusively,on memberswith relativelyshort chains,tlat is, with a
maximumof l0 or 72 carbons.
While fatty acids with short chainstake part directly in thesecaloric
metabolicchanges,thosewith longercarbonchainsmust undergopreliminary changesbefore entering into caloric metabolism.A desaturation,
changinga saturatedfatty acid into a monoethenic,appearsto be a first
stepin caloricmetabolismof the long chain members.The monoethenoids
thus can be seento be intermediaryforms bctwcenthe saturatedreserve
and the short-chain,easilymetabolized
fatty acids.
The double bond in these monoethenicacids would rhus appear to

130

RESEARcH IN

PHYSIoPATHoLocY

have two uses: one, to reducethe melting point below body temperature
and thus permit easymobilization,and two, to induce changeswhich lead
to the breaking up of the long moleculeinto two shorter ones which can
be metabolizedthrough the Knoop oxidation.
All the data indicate that this fission would not take place at the double
bond but through a more complex process.A first change consists of
oxygen fixation at the carbon near the double bond. This leads to the appearanceof a hydroperoxidegroup. It is only in a subsequentstep that
the moleculebreaks at a place betweenthis carbon near the double bond
and the double bond itself, resulting in the appearan of short chains
which have an even number of carbonscapableof being directly metabolized through the beta oxidation.(Note 4)
The positionof the doublebond in the naturally occurringmonoethenic
fany acids, separatingalmost always a group of nine carbons toward the
carboxyl or the mcthyl end, (Note 5/ acquiresa special significancefor
the breakingdown of the moleculesfor caloric purposes.
The desaturationof the saturatedfatty acids,which would representa
first step toward allowing them to participate in metabolic caloric changes,
would usually takc place in the liver, apparentlythrough the same processesby which polyunsaturatedfatty acidsare partially saturated.(Note 6)
An interestingpart of the caloric metabolismof the saturatedand
monoethenicfatty acids, which will be shown below, is their combination
with glycerol to form triglycerides.
Constitutional Role
Although saturatedand monoethenoidfatty acids enter into the formation of boundary membranes,the di-, tri- and tetraenicmembersseem to
havea particularlyimportant role in the constructivefunction of fatty acids.
Some of them entcr directly into the formation of the membrane;some
forrn complex lipoids such as lecithinewith the glycerophosphoricradical
and nitrogen containing bases.As a rule, these last representa lipoidic
substratewhich would act as a neutral natural solvent present in membranes,and as such, intervenein the realizationof a diphasic medium at
the level of the boundary formation.This medium would largely insure
the orientationof the fatty acidsat the separationsurfaceand the formation of permeablelipidic layers,
FunctionalRole
The third role of fatty acids is as functional agentstaking part in certain reactions.This activity appearsto be strongly related to two factors:

LTPIDSAND LTPOTDS /

l3l

the presenceof an uncombinedcarboxyl $oup and the energeticintervention of the double bonds of the nonpolar part of the polyunsaturated
members.
Free fatty acids appear to be functionally active while the combined
ones usually are inactive.The activity is relatedonly partially to the direct
capacity of the carboxyl to realize new combinations.It resultsfrom the
induction exertedby the carboxyl upon the nonpolar group. The so-called
free fatty acids of the organism are probably bound in a labile form to
proteins, but this bond will not influencethe induction effect exerted upon
the nonpolar group. The intensive positive carbon of the carboxyl, together with the zig-zagdispositionof the fatty acid molecule,causesthe
inductive effect to charge the successive
carbons of the chain. They will
thus show alternativesigns.The even carbonsshow a negativecharacter,
while the odd ones are positive.The fact that oxygencombineswith positive carbonsexplainsnot only why, as in Knoop oxidation,this bond occurs
at C3, which is strongly positive,but also explainsthe so.calledalternate
oxidation (33) where the other following odd carbonsare binding oxygens.
Through the influenceexerted by the carboxyl, the double bond shows a
specialactivity which has been worth studying.
Double Bonds
There has been some tendencyto regardthe double bond as a weak,
easily broken point of the molecule.Actually, it emergesas an important
center of activity. With its capaciry to become a semipolar center, and
consequently,to bind or lose radicals,the double bond is an energetic
center in the molecule.Is importantcharacteristic
is the ability to effect
such changeswithout altcringthe chain of the moleculeitself. Since this
type of reactionis reversibleand can be repeatedfor the same molecule,
the double bond appearsto representa functional entity. Becausethe reaction principally involvesnonmetallicelements,the unsaturatcdfatty acid
takesan activepart in the metabolismin which theseelementsappear.
The study of rancidity has helpedus, by analogy,to systematizeoxidaas they take placein vivo. In addition to Knoop beta, several
tion processes
other g'pes of oxidation could be recognizedin which double bonds intervene more directly. The double bond, with its semipolarcharacter,influences nearby carbons, rendering them highly reactive. In one form of
oxidation, a molecularoxygen is bound to a nearby carbon to produce a
formation,as was shownto occur in vitro by Farmer. (31)
hydroperoxide
this oxygenfixationbecomesreversible.
When,under certaincircumstances,
the fatty acid will liberate the oxygen. It appearshighly probable that in

132 /

R E s E A R C Hr N p H y s r o p A T H o L o c y

such a process,the oxygen is libcratedas a free radical, the entire process

thus corresponding
to an activationof oxygen.The changeof a molecular
oxygcn into a frec radical would representthe physiologicalrole of unsaturatcdfatty acidsin oxidationprocesses.
The presenceof two double bondsin non-parallelposition,contmon to
most of the naturallyoccurringpolyunsaturatcd
fatty acids,is even morc
important; the two double bonds exert a parlicularly strong influcnce on
the specialcarbonwhich is in thc intermediarypositionbetweenthem. Becauseof the alternateinduction producedby the strongJypositive carbon
of the carboxyl,the carbonsof the chainhave alternatecharacters,
positive
and negative.When an intermediarycarbonalso has a strongpositivecharacter, it appcarsto be cspeciallyable to fix oxygen.This strong.lypositive
intermediarycarbon,occurringin naturalpolyunsaturated
fatty acidswith
more than two double bonds, may be the rcason for the important role
playedby theseacids whcn they act as essentialfatty acids in the organism. (Note 7)
The study of rancidity has further shown that, while the in vitro
oxidationof an unsaturated
fatty acid under mild conditionssuch as room
temperatureleads to the appearanceof hydroperoxidcs,oxidation at a
higher temperaturewill result in anothcrfixation of oxygen,this time at
the doublebond itself.Epoxidcsor peroxidcswill appearaccordingto the
ionic or molecularcharactcr of the oxygen. This extremely important
processalso occurs in rancidity under the inl'luenceof an enzyme. It is
highly probablc that a similar proccsstakes placc in vivo in those pathologica.l
conditionsin which pcroxidesappearin the urine.Radiation,certain
inflammations(especiallythosc due to streptococci),administrationof
seleniumpreparations
or of highlypolyunsaturated
fatty acidsare followed
by the appearanceof theseoxidizingsubstances
in urine. As mentioned
above, when these substancesappear, there also arc increasesin indoxyl
and glucuronicacid, which can be considered,up to a certain point, to
resultfrom abnormallyintensiveoxidationtakingplaccon tryptophaneand
glucose.(See below.) while acrivationof oxygenis a physiologicalprocess,peroxidesappearunder abnormalconditions.

ABNORN{AL FATTY ACIDS

T h e s t u d yo f t h e r e l a t i o n s h i bp e t r v c c na b n o r m a cl o n d i t i o n sa n d l i p i d s h a s
progrcssively
led us to considerthe existence
of clualitative
changesin these
l i p i d s ,b e s i d e st h e c l u a n t i t a t i voen e s .T h e c x i s t c n c eo f a b n o r m a ln r e t a b o l i c
processes,
and espcciallythe fact that such abnormalitiesare often of long

L T P t D SA N D L t P o t D s

133

duration,could hardly be attributedto variationsin the quantityof the interveninglipids alone. More probably they would result from changesin
the nature of the interveninglipids themselves.
We have investigated
this
aspectof the fatty acids presentunder abnormal conditions.
As a guide for the direction to be followed in theseinvestigations,we
used the information furnishedby the study of rancidity. We believedrancidity would be ableto indicatebroadly the natureof the qualitativechanges
which the lipids may undergounder abnormalconditions.Conceptually,the
abnormal can be consideredto result from a loss of the capacity of the
organism to sufficiently control occurring processesand keep them in
the frame of the constantswhich characterizethe entity. Due to this lack of
effectivecontrol, the in vivo occurringchangesunder abnormal conditions
would closelyapproachthose which take place in vitro where such a control doesnot exist,Theseconsiderations
led us to searchfor changessimilar
to those seenin rancidity,or occurringin vivo in lipids, under abnormal
conditions.
As mentioned above, in rancidity a first group of changesconcerns
the polar group. Someof them result in the appearanceof free fatty acids,
others correspondto changesin the carboxylsthemselves,while still others
are representedby processes
of oxidationwhich occur in the chain near the
polar group. A secondgroup of rancidity changesconcernsthe nonpolar
group and especiallythe energeticcenterspresentin it, the double bonds.
The study of this last group of changesled us to consider,the changes
appearingin vitro under the direct influenceof heat and oxygen.As part of
thesechanges,we consideredof specialimportancethe conjugationof the
double bonds seen to occur in vitro as a step in the oxidation of polyunsaturatedfatty acids.This conjugationcorrespondsto a characteristicdisplacementin the moleculeof two or more of the double bonds present,so
as to result in parallel reciprocalpositions.
While in the simple bond two tetrahedriccarbons are bound through
their peaks,in the double bond they are bound by one edge, and in the
triple bond by a surface.In the conjugatedformation, the common edges
of two doublebonds,being separatedby one simple bond, are consequently
parallel.The planesin which the electronsof thesedouble bondsare moving
for eachdouble bond and which are perpendicularto that of the bond itself,
becomeparallel. Through the resultingreciprocalinduction their energetic
value is enhanced.
We have studied systematicallythe different qualitative abnormalities
concerning the fatty acids, guided mainly by the information obtained
throughthe study of rancidity.

.^

134 /

nESEARcHrN pHysropATHoLocy

Methods ol Investigation Used

Followingthis line,we firstinvestigated
the forms underwhich the lipids
in gcneralare presentin the organism.We utilizedthe differences
in solubility betweenthesedifferentforms, separatingthem into free lipids, lipids
kept in a labile bond with other constituents,
as in cenapse,lipids bound
throughtheir polar group as in fats,or in the still strongerform as lipids in
combinationsso firm that thcy cannot be separatedexcept through saponification.The methoddevisedfor this study and someexamplesare in Note
8A. This researchshowedthat under abnormalconditions,very important
variationsoccur in the amountsof the difterent forms. This study pointed
out that the free lipids are greatlyresponsible
for the importantnranifestations in which lipids appearas activeagents.
In the study concerningthe abnormal metabolismof the carboxyl and
nearby carbons, we investigatedthe appearanceof fatty aldehydesor
ketonesin blood, urine and in the cells.
Onc of the major problemsencountered
was the appearance
in vivo o[
conjugatedfatty acids,as abnormalfatty acids.In order to ascertaintheir
presenceand to measuretheir amounts,we had utilized three differcnt
methodsof investigation:spectralanalysisin ultra-violetand in the first
portion of the visible spectrum(Note 8Bl; the study of the place of thc
doublebond in the fatty acidsmoleculethroughthe fissionof thesemoleculesand the analysesof the resultingfractions.(SeeNote I, Chapter 101
More recentlywe have tried the vapor fractionationmethod (gas chromatography) (Note 8C).
The first, and especiallythe secondmethod,gave us valuabledata permitting us to recognizethe interventionof conjugatedfatty acids in ab'l'hese
normal conditions.
studiesrevealedthe appearanceof conjugated
fatty acids,especiallyas trienes,the increaseof their amountwith the progressof the conditionsand especiallythe fact that deathoccurswhen their
concentration
in the bodieshasreacheda criticalvalue.This has markedthe
importanceof thesesubstances
in physiopathology.
The fact that gaschromatographydid not revealthe presence
of conjugatedfatty acidsappearsdue
to the conditionsunder which the methodactuallyworks.
Later, we will frequcntlyreturn to the variousproblemsrelatedto interventionof conjugatedfatty acids.We could thus directly correlatethe
interventionof theseabnormalfatty acidswith the pathogenesis
of the manifestationsof many conditionssucb as trauma,shock,adrenalectomy,
and
especiallywith the noxiousmanifestations
following irradiation.(Chapter
I0)

LIPIDS AND

LIPOIDS

,/

I35

In abnormal metabolic changes,an important factor is the intervention of abnormal fatty acids in the metabolismof chloride ions, producing
an especiallystrongfixation of the chlorideion to the carbonsat the double
bonds. The conjugateddouble bonds in a fatty acid moleculeappearto be
especially suitable for this since an abnormal, irreversible fixation of
chlorides occurs in two steps. First, the halogen is fixed at the extreme
carbons of conjugatedformationswith a displacementof the double bond
in the intermediaryposition. In the secondphase,the fixation takes place
in the intermediarycarbons,too. (Nole 8D)
Functionally,fatty acids induce activationof oxygenas a norrnal process, but the appearanceof peroxidesor irreversiblefixation of ctrloride
ions is an abnormal event.
It is the abnormal fixation of chloridesby the conjugatedfatty acids
which leads to a more complex group of processcsinvolving sodium
chloride metabolism.with the chloride ion fixed. the sodium ion of sodium
chlorideremainsfree to enterinto othercombinations,
especially
with a carbonateion, producingstronglyalkalinecompounds.This processexplains
the appearanceof local alkalosisas a result of thc intcrvention of conjugated abnormalfatty acids,corresponding
to the chloride phaseof ',D."
The division of fatty acids into four groups-l ) saturatedand monounsaturated,2) di-, tri- and possiblyalso tetra-unsaturated,
3) tetra- and
higher polyunsaturated,and 4) conjugated---{orresponds
schematicallyto
the four principal roles---caloric,organizationa.l,functional and pathogenic
-which fatty acids play in the organism. These roles are seen to be
dictated both by the differentstructuresof the fatty acids and the different
substancesto which they are preferentiallybound. The fate of a fatty acid
in the organism seemsto be greatly influenced by its bond to other substances.As already noted, we have called these other substances"antifattv acids."
THE

ANTI.FATTY

ACIDS

Gl;'cerol and Glyc'erophosphoricAcid

It is classically acceptedthat the intestinal absorption and circulation
of fatty acids is made through bonding to varioussubstances.
The analysis
of this absorption shows, however,that different fatty acids have preferential bonds. For saturatedand monothenicfatty acids,the bond is principally with glycerol. Although mono- and di-glyceridescan be identified
in the cells of the intestinalmucosa,these fatty acids leave the intestine
as triglycerides,forming the largest part of the chylomicrons.They are

135

nESEARcH rN PHYStoPATHoLocY

also found in reserye in adipous cells as triglycerides. The di-, tri- and
even tetraenic fatty acids usually enter the circulation as phospholipids,
that is, in direct combination with glyccrophosphoric ions. The polyunsaturated acids are bound to sterols when they enter the blood, circulate
and are stored. While the structures of the various fatty acids determine
their difterent roles in the organism, it is the anti-fatty-acid constituents
to which they are bound which enhancethcseroles.The study of the antifatty acids has shown that these substancescan even dictate, by themselves,difterentfates for the variousfatty acids they bind.
The combination of glycerol with any fatty acid seemsto establisha
caloric metabolic character. This is true for the very different fatty acids
found in plants and animalsas triglycerides.Even the ricinoleic triglyceride,
if fresh, is used as comestibleoil, castor oil. The same is true of the oil
of triglyceridesof polyunsaturated
fatty acids found in marine animal oils.
In the seeds,all the triglyceridesof fatty acids,even the conjugatedones
such as eleostearicand parinaric,representenergeticsources.It seemsthat
it is their combination with glycerol which has given all these fatty acids
value as caloric metabolites.The same is true for the bond to glycerophosphoric ion.
Combination with glycerophosphoricacid endows various fatty acids
with the abi.lityto participatein the constructionof membranes.The bond
to sterols,on the contrary, inducesan ultimate functional activity provided
the fatty acid iself is so constitutedas to be able to fulfill this function.
The influenceexcrtedby anti-fattyacidscan be understoodin terms of
the changesthey induce in the activity of the fatty acids.Sincethe activity
of the last is largely relatedto their presenceas free substances,
it is principally through their combinationwith fatty acids that the anti-fatty acids
intervene.By inactivatingthose free fatty acids which form a membrane
and insure its permeability,an anti-fatty-acidagcnt can causethe membrane to changeits permeabilityand evento becomecompletelyimpermeable. Similarly, an anti-fatty acid, by combining with a polyunsaturated
fatty acid, can reduce or even suppressits functional activity. It is to be
notedthat,by both changingpermeabilityand suppressing
functionalactivity, the anti-fatty acids exert their influenceultimately by altering oxygen
metabolism.From this point of view, metabolismbecomespredominantly
anoxybioticin contrast to normal oxybiotic metabolism.For glucose.for
instance,suppression
of the oxidativephasearrestsmetabolismat pyruvic
acid which passesinto lactic acid. The appearanceof acid substancesas a
biological effect of the action of anti-fafty acids results,in fact. from the

r.rPrDs^ND LrPotDS

137

rc'ductionof the fatty acid's activity, aflectingoxidative processesdirectly,

or indirectly through reductionof membraneFrcrmeability.
while one group of anti-fatty acidscan be directly relatedto hydroids,
and especiallyto glycerolor to glycerolbound to phosphoricacid as in the
ion, a secondgroup is represented
by lipoids, principally
-elycerophosphoric
formed by derivativesof a characteristicring system,the cyclopentanophenanthrene.As anti-fatty acid lipoids, thesc compounds,the steroids,
were of special intcrest in lipoid research.only some aspectsof the biologicalactivity of steroids-mainly, thosewhich reprcsentnew views in the
studyof thesesubstances-willbe discussed
here.

Steroids
A fundamentalrole of thesesubstances
in biology is determinedby the
'l'his
fact that they are polycyclic.
leadsus to considerthe role of the ring
iself in reactivity, as shown by a study of the steroidsin oppositionto the
fany acids.In thc fatty acids.the bondsbetwecncarbonsas presentin the
aliphatic chain, insure a high reciprocalmobility between these carbons.
As a result, the entirealiphaticchain is highly llexible.on the other hand,
ri-uidityis characteristicfor all the rings,and is increascdby the polycycling
of the nrolcculcs.The constitucnts
of the moleculesare kept in fixed reciprocalpositions.while, in the fatty acids,the flexibility of the chain permits
the energeticcentersto take different relative positionsamong themselves
towardother nrolecule-s,
the rigidity o[ thc p<llycyclicmoleculcsmaintains
theenergeticcentersof the cycle,or thoseattachedto it, in the samerelative position. This fundamentalcharacteristic
of the cyclic moleculesappearsto be an important factor in determiningthe biological role of the
variousagentswhich have such cyclesin their molecules.
In the case of steroids,this attributcacquiresspecialimportance.An
understanding
of the differentbrologicalactiviticsof steroidscan be obtainedby an ana.lysisof the forces resultingfrom this characteristiccomposition.Besides the energeticcentersor formations attachcd to it, two
energeticcenters appear as part of the steroid nucleus itself. One is at
Cr and the other center is rcpresentedby the cyclopentanicgroup. The
factthat these centersare maintaincdin fixed relativeposition through the
rigidityof this polycyclicnucleushas resultedin an importantpropertyof
thenucleus itself which becomestranslatedinto a dipolarity of the molecule.The study of thesetwo energeticcentershas advancedour knowledge
of the role of steroids.
The study of the polar groupsbound to C,, of the polycycleskeletonof

138

nESEARcH rN pHysropATHoLocy

steroidshas permitted us to recognizethe conditions which induce stronger

activity for these polar groups, conditions which are usually fulfilled in the
naturally occurringmembers.It could thus be seenthat the reactivityof an
oxygen bound to Cs is increasedif another double bond present in the
cycle is parallel to the double bond through which the oxygen is bound to
Cr.A doublebond betweenC1 and Cr,,as shownin Figure 64 (a), fulfills

(a)

(b)

(L'

F l c . 6 4 . I n f l u e n c ee x e r t e du p o n t h e o x y g e n b o n d a t C ' b y t h e p o s i t i o n o f t h e d o u h l e
bond in the cycles I and 2 of the cyclopentanephenanthrene
molecule. A parallelism
b e t w e e nt h e d o u b l e b o n d o f o x y g e na n d t h a t p r e s e n tb e t w e e nC , a n d C , i n c r e a s e st h e
e n e r g e t i cc h a r a c t e ro f t h e c a r b o n y l ( a ) . A s i m i l a r i n f l u e n c eb u t l e s s a c t i v e , i s e x e r t e d b y t h e d o u b l e b o n d b e t w c e nG a n d C , . A d o u b l e b o n d a d d e d b e t w e e nC , a n d
C ' ( c ) i n c r e a s e st h e a c t i v i t y . S t i l l m o r e a c t i v i t y w o u l d r e s u l t f r o m a t b i r d d o u b l e
b o n d a d d e d b e t w e e nC c a n d C , ( d ) .

such a condition.A similar influenceis exertedindirectly by a double bond

betweenC6 zrndCz (b) which, through induction,will influencethe parallel
Cr and C5 bond and further the double bond of the oxygen.This explains
the influenceexertedby the double bond presentbetweenC, and C, (c).
as in the synthetic,prednisolone.
Further enhancement
of reactivitywould
be obtainedwith a third doublebond addedbetweenC6 and C7. The parallelism bctwecn three double bonds (d ) would produce an increased
reactivity.
For the hydroxyl, a similar enhancedreactivity is induced by double
bonding of the carbon to which the hydroxyl is attachedwith a double
bond for the Cs - Ca or C, - C., as shown in Figure 65 (a and b). A
similar condition is fulfillcd if a doublc bond is prescntin the moleculc
parallelto any of thesebonds,as seenin Figure 65 (c) where the double

r*

,\/-t-r*oAA
(a)

rff

-k

.*

Ho,/W

*o&
(b)

(c)

6)

F t c . 6 5 . T h e i n f l u c n c ee x e r t e du p o n t h e h y d r o x y l b o n d a t C . b y a d o u b l e b o n d i n
t h e c y c l e I a n d 2 o f t h e p h e n a n t h r e n ei s i n c r e a s e di f t h e d o u b l e b o n d i s a d j a c e n to r
parallelto the bonds of C., bearingthe hydroxyl.

LTPtDSAND LrPorDS

139

bond is betweenC5 and Co. An enhancedreactivityof thesecompounds

would be obtainedwith one double bond betweenC3 and C. and another
betweenCr and C,, (d ).
We will come back to this important intervention of the double bonds
in cyclic molecules.
The energeticproperty of the cyclopentanegroup apparsto be correlated with its odd number of carbons.The a.lternatesuccessionof carbons
with positive and negativecharactersresultingfrom the induction effect
causestwo carbonsof this cycle to have the sarnesign. This "twin formation" inducesa specialmolccular reactivity related to the pentanic cycle
of the steroidmolecule.(Note 9)
The special reactivity seen for c3 of thc cyclopentanophcnanthrene
molecule can be explained through a hypothesiscovering the origin of
these substances.
Although the origin of a cholesterolmoleculethrough a
cyclizationof squalene(35) appearsplausible,this seemsless probable
for the corticoids.We havetried to connecttheir origin to arachidonicacid.
Severalconsiderationssuch as the high levelsof arachidonicacid and
corticoids in the adrcnals,and the reduction of the former when an important amount of the latter is excreted(Note I0), seem to establisha
correlation between these substances.According to our hypothesis,the
steroidswith a two carbon chain at Cr;, as seenpresentin the corticoids
and luteoids,would result from a cyclization of the arachidonicmolecule.
(Note 11l This would explain the specialreactivityof C., which woutd
correspond to ce of the arachidonicacid bound in this molecule by a
double bond.
A study of the different steroidsunder this energeticaspecthas permitted us to understandtheir physiologicpropcrties.
with the c3 having a hydroxyl or an oxygen as polar group in almost
all the steroids,the variety of the biologicalproperrieswould be relatedto
the different conditionsat the other extremity of the molecule,principally
at Crz, which result from the specia.lenergeticconditionsprevalentat this
regionof the molecule.The simpleststeroidsare thosehavinga polar group
representedby an OH or O fixcd at C17.Such naturally occurring steroids
have propertiesrelated to secondarysex characteristics.We will discuss
them briefly here.
Sex flormones
This group of steroidshas two polar groups, one at C.qand one at C17.
The energeticcenter at C3 can have negativeor positive polar characters,
accordingto the presenceof oxygen or hydroxyl. The energeticcenter at

140 /

R E S E A R c Hr N p H y s r o p A T H o L o c y

Crz also can have an oxygen or hydroxyl group and thus be negative or
positive. An important factor for the properties of the substanceis the
relationship between the two polar groups in the same molecule. It is apparent that the polarity of the molecule will vary according to what polar
groups are present at C3 and C17.[n a very simplified concept, which we
consider only partially accurate, we have tried to associate female and
male hormonal charact,eristicswith this polarity.
In the simple steroid molecules,a folliculinoid or estrogenicbiological
property seems to be conferred if the two polar groups-at Cs and Crz
-are formed by hydroxyls. The molecule appears to have a dipositive
polarity. It seemsto be important that the two hydroxyls be kept in the
relatively fixed reciprocal positions---+orrespondingto C3 and C,7-as
part of the solid skeleton of the steroids.Estrogenic properties are present
in various steroids that fulfill this condition. Furtherrnore, substancesfar
removed from the steroids have folliculinoid properties if they have this
relationship between the two hydroxyls. As shown in Figure 66 diethylstilbestrol,which has its two hydroxyls maintained in a fixed relative position similar to that of steroid estrogens, also shows potent estrogenic
activity.
OH

Estradiol

D i e t hIvs ti l b e s t r o l

Fro. 66. The lolliculinoid activity appears to be related to the existence of ru,o
hydroxyls kept at lhe same relative position. as it appearsin estradiol and in diethylstilbestrol.

ln the same way, we tried to correlate testoid activity with the presencc

of positive-negative
polarity; that is, with two polar centersenergetically
different, one corresponding to an oxygen and the other to a hydroxyl,
maintained in the same fixed relative position. The importance of this
relative steric position of the two polar groups for testoid activity becomes
evidentwhen it is found in substances
other than testosterone,
the principal
male hormone,with an oxygenat C3 and a hydroxyl at Cri. Testoid activity is presentin androsterone,which has an oxygenand an hydroxyl maintained in the samereciprocalpositions,althoughhere the oxygen is at C17

LIPTDS AND

LtPotDS

l4l

and the hydroxyl at C3, the reverseof testosterone.In both substances,

testosteroneand androsterone,the two conditions for testoid activity, positive-negative polarity and the same relative position between the polar
groups, are fulfi.lled.(Fis. 67)The differenceswhich exist betweenthese
substancesin their specifichormonal activity can be explained through the
difterent influence exerted upon the two polar groups in these substances
by the rest of the molecules.

Testosterone

Androsterone

Ftc. 67. The testoid activity seemsto be related to the presenceof a hydroxyt and a
carbonyl in the same fixed relative position which is insured by tbe rigidity of the
steroid molecule. The same positional reciprocal relationship is seen to exist between
these two polar groups in testosteroneand androsterone.

The testoid activity secn for cortisone, hydrocortisone and other hormones also can be explained by the presencein these moleculesof oxygen
and hydroxyl at C3 and C17,and maintenanceof the fixed position between
thesetwo polar groups.
Conceprually,the antagonismbetweenestrogenicand testoid biological
activitiescan be consideredto be ultimately related to the differencesin
polarity, which in one form or anothercan be found in other factors differing for the sexes.we will men[ion here only that a similar differencebetween male and femalecharacteris seenin the sexualchromosomes,where
the female characteris related to the XX chromosome,and the male to
an X and a Y chromosome.As we will see below, a relationshipexists
betweenlipids in generaland sex.
Besidesthe sex hormones,fatty acids appear to be connectedwith male
sex characteristics,
while femalecharacteristics
are relatedto anotherg.oup
of steroids,the sterols.
Sterols
Characteristic of the structure of this group of steroids is the presencc
of a hydroxyl at Cs and a long chain bond at C,z. Through the hydroxyl,

:-ttr I

t.t

t;

142 /

xESEARcH rN pHysropATHoLocy

the center at Cg has a nucleophilic character. This is reinforced by the

presenceof a double bond botweenC5 and C6 which, by paralleling the
bond betweenC3 and C1, increasesits ionic character and consequently
the reactivity of the hydroxyl bond to C3. Through this hydroxyl, sterols
combine in general with substanceshaving a negative polar group to form
esters.
Besidesthe capacityto combine with fatty acids in general,one of the
most important characteristicsof the principal sterol of animals, cholesterol, is its selectiveaffinity for certainfatty acid membcrs,the polyunsaturated.We tried to explain the specificityof this bond through an interesting
processwhich could be called "steric coupling."
In this process,two molecules,usually lipoids, are kept together not
only by the combination of their polar groups but also through a bond
betweentheir nonpolar parts. The two moleculesare reciprocallyattracted
through the multiple forces presentin the nonpolar groups. Some are related to attached centers, while some, such as those correspondingto
cohesionforces,are relatedto the rings themselves.An important factor is
the rigidiry of the sterol moleculewhich permits another molecule,if it is
flexible,to make the steric coupling.The rigid skelctonnot only kecpsthe
energeticcentersof one moleculein a fixed positionbut permits the flexible
aliphatic chain to cover over the polycyclic molecule and thus bring the
energeticcenters of one molecule in contact with those of the other.
Through this, steric coupling completesthe bonding of the polar groups.
The greaterthe concordanccbetwcencnergcticcentcrsin both molecules,
the more perfect the coupling is, for the more complete is the reciprocal
neutralizationof the energeticcentersof the two molecules.Stericcoupling
explains why, of all the fatfy acids presentin the organism, cholesterol
seemsto prefer to bind thosewith polyunsaturatedchains.It is thesefatty
acidswhich have severalenergeticcentersin thc nonpolar group as representedby double bonds. The long chains of these fatty acid molecules,
havinga certaindegreeof flexibility,will then completcthe stericcoupling.
(Note I2 )
Stericcoupling, in addition to its generalimportancein biology, where
it representsa kind of molecular reactivity,seemsto cxplain the antagonistic influence exercisedby different constituents,especiallythe sterols
and polyunsaturatedfatty acids.Through stericcoupling.cholesterolcould
influencethe activity of thesefatty acids more dircctly related to the nonpolar group. It has to be emphasized,however, that the neutralization
resultingfrom steric coupling is not irreversible,On thc contrary, through
the interventionof various factors,such as the breakingdown of the bond

LTPTDS AND

LTPOTDS

143

betweenthe polar groups,the two coupledmoleculescan regain their independence.This would explain the relativelability of the combinationsberween fatty acids and sterols. The antagonism between fatty acids and
sterols is an important aspectof biological dualism which will be discussed
in more detail later when these substancesare studied in terms of their
influence at the difterent levels of organization.

Steroids with a Two-Carbon Chain

Among the most important steroids are those having a two-carbon chain
fixed at C17.
Two groups, the Iuteoids and corticoids, appear directly related to
allopregnanehydrocarbon,the steroidpolycyclewith a two-carbonlateral
chain fixed at Cr;. As we have alreadyseenin the hypothesisconcerning
the origin of the steroids(Note 11l, this hydrocarboncould have been directly derived from arachidonicacid, thc two-carbonlateral chain correspondingto the tail chain of this acid, a tail which remainsafter cvclization.
The Luteoids
The prototype of the luteoids is progesterone.Two polar groups C : O
are present,one at C3 of the polycycleand the other at C:roof thc tail chain.
A paralleldouble bond betweenCa and C; completesthe formula. Energetically,progesteronepresentsa first center at C3 which appearsstrongly
nucleophilicfor two reasons:first, becauseit correspondsto the potent
electronegative
Ca and second,becauseit is reinforcedby a double bond
presentbetweenC1 and C5,and whichis henceparallelwith the doublebond
of the carbonyl.The second : O is attachedto the Cs,yof thc tail chain.
This alsoappearsreinforced,the doublebond of this carbonylbeingparatlel
to the bond betweenthe Crr and C17,which in the cyclopentane,
accordhypothesis
of
ing to the
twin carbons,binds two negativechargedcarbons.
Through its constitutionprogcsterone
is also a lipoid, thc complexhydrocarbon group being predominant over the polar groups. With its polar
nucleophilicccnters,progesterone
has the fundamentalcharacterof acid
luteoid activitycorresponds
lipoids. Progesterone's
to the presenceof two
relativelystrong neutrophiliccenterskept in the characteristicpositions,
one at C3 and the other at C2s.
We can see that any disturbancein this energeticpicture, any change
from the dinucleophilicat any center, decreasesthe luteoid propertiesof
the substance.With more profound changes,the luteoid activity is even
(Note 13)
suppressed.

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nESEARcHtN pHystopATHoLocy

Corticoids
The corticoidsrepresentthe group of hormonesupon which the attention of scientistsrecentlyhas beenintensivelyfocusedbecauseof their new
therapeutic applications.
chemically, they appear to be the same as luteoids,derivativesof the
s:rmeparent hydrocarbon, allopregnane.Structurally, all these adrenocorticoid hormoneshave: a) a c3 bindingan o group; b) a double bond betweencr and cc in the first cycle;c) a two-carbontail chain with an o
attachedin ketone form to Czui d) an oH as primary alcohol presentat
c2t. This structure,common to all corticoids,seemsto be responsible
for
the principal propertiesof thesesubstances.
Corticoidshave been separated
into subgroupsbasedupon the presenceof aftachedgroups oH or _--o at
Crr or OH at Ctz. The presenceor absenceof attachedradicalsat C,,
appearsto be most important. Corticoidswithout attachedradicals at the
C1l have a major influenceon the metabolismof electrolytes.The second
group of corticoids,having the radical, are known as neoglucogeniccorticoids,the nameindicatingtheirprincipalbiologicalcharacteristics.
Energetically,the corticoids present a nucleophilic center at c3,
reinforcedby the presenceof the double bond in the cycle betweenCr and
cr. The doublebond is parallelto the doublebond of the carboxyl,and thus
inductivelyincreasesthe ionic characterof the latter.
A secondenergeticgroup of the tail chain appearsin toto as a strong
tripolarcenterwith a nucleophiliccenterat C.:,of this chain and an electrophilic center at c31. (Note 14) To this basic pattern is added, in the
neoglucogenic
corticoid,a separateenergeticcenter at C11.which can be
either clectrophilic,formed by a hydroxyl,or nucleophilic,formed by an
oxygen.
Corticoidsappeil, in general,to act as positivelipoids.(Note t5)
Becauseof their importancein relation to anti-fatty acid activity, we
will discussfirst the neoglucogeniccorticoids,the members with a polar
group also at C11.Accordingto our hypothesis,
thesesteroidshave a special biological activity, a role in the processof synthesisin the organism.
The part of the molecule between C11 and Crr constitutesan energetic
formation with a peculiar property. It representsa kind of energeticmold
or template,in which each carbon has its specificenergeticcharacter.Different radicalswould be attractedby the energeticcentersof this template
formation accordingto their own energeticnature.Kept in their respective
positions,they would be induced to bind together in order to fornr new
substances.In this manner this template formation would promote new

L T P T D SA N D L T P O T D S /

145

syntheses.[n difterent corticoids the constitution of the Crr : Czr formation

will differ and this will determine which substanceis to be synthesizedby
the respectivemold or templateformation.(Note I6)
Using the template hypothesis, we studied an entire series of body
constituentsforming the "gluco group." Glucose,galactose,glucosamine
and galactosamine,with their respectiveacids, as well as ascorbic acid,
:ue rmong thesessubstances.According to our hypothesis,thes neoglucogenic corticoids would have the important role of producing, possibly
along with other mechanisms,the entire seriesof "gluco" constituents.The
existence of difterent template formations would result in a variety of
synthesizedconstituents.
The intervention of the template formation in synthesis can occur
again and again without aftecting the molecule of the corticoid as such. It
is interesting to note here a structural curiosity which could be interpreted
as being related to template activity. In this template,the group of successivec,r, c,r, c13 and crz ?re part of the rigid skeletonof the cyclic
molecule,while C:o and Czr are forming the lateral chain attachedto C17.
This can be regarded as conferring a certain propr mobility to this lateral
chain as related to the polycycle.It is conceivablethat this lateral chain
would becomea closedformation when synthesistakes place.A movement
of the chain at Crz would permit the mold to open and thus liberate the
synthesizedmolecule. It is interesting to note here the importance of the
structure of the template for the constitution of the substancessynthesized.
Besidesthe polar group at c17, that at Crr is also important for neogluco
genic activity since it insuresa six-carbonchain in the synthesizedmolecules. A hydroxyl or carboxyl at the Co of the synthesizedsubstancewill
appar, according to the nature of the polar group at Cn of the steroid.
The respectivecharactersand positionsof Czr and C12 will permit thc
appearanceof a cycle formed by five carbons and an oxygen, characteristic
for the pyranic form of newly synthesized
substances.
An interesting confirmation of the template hypothesis was obtained
when glucosamine which, according to the hypothesis,is synthesizedby
the cortisonemolecule,was found to inducein patientsmany of the clinical
changeswhich are obtainedby treatmentwith cortisone.We will consider
these resultslater in our discussionof therapy.The capital role played by
glucosamine,galactosamineand the respectiveuronic acids in the constitution of the connective tissue representsthe "missing link" for the
explanation of the relationship betweencortisone,the other neoglucogenic
corticoids, and this tissue. Some part of the therapeutic eftect obtained

146 /

R E s E A R c Hr N p H y s r o p A T H o L o c y

with theseneoglucogeniccorticoidsin diseasesof the connectivetissuehas

to be attributed to the intervention of the amino sugars.
In the study of anti-fatty acid activity, glycerol and glycerophosphoric
ion werefound to control the absorptionand circulationof saturatedmono-,
di-, or tri-unsaturatedfree fatty acids.The sterolsappearto counterbalance
the normal polyunsaturatedmemberswhile adrenal corticoids, and especially the neoglucogeniccorticoids, counteract the toxicity of fatty acids in
general and of the abnormal conjugated members in particular. Research
done in our laboratoriesby E. F. Taskier indicatesthat the adrenalsintervenein the defensemechanismagainstfatty acids,and especiallyagainst
the conjugated members which appear to be related to abnormal conditions. (Note 17)
The part of our researchconcernedwith the role of lipoids in normal
and abnormal physiologyhas been almost entirely guided by the concept
of an antagonismbetween the two groups, one with a positive and the
other with a negativepolar character.This specificaspecthas led us to
study, together with the fatty acids and the anti-fatty acids, other substancesrelated to this antagonism.In the group of Iipoacidsor acidic lip
oids, as obtained from tissues,organs or organisms,we rccognizedthe
group of porphyrinic acids,relatedto various hemespresentin the organisms. In the group of anti-fatty acids obtained from the same sources,
differentconstituentsform the insaponifiablefractions.
As related to this dualistic aspectwe have studicd another group of
substances,
which appearto act in the organismagainstthe anti-fatty acids
themselves.These other substanceswould representa kind of biological
brake to counteract an exaggeratedintervcntion of anti-fatty acid constituents,
we have made a specialinvestigationof two substancesof this group,
glucuronic and sulfuric acid anions, which charactcristicallyseem to opposecertain anti-fatty acid substances.
Thesesubstances
appearas a result
of an exaggerated
oxidationof normalmetabolites.
Under abnormalconditions, the oxygen resultingfrom the interventionof peroxidemay be fixed
to carbohydrateseven bcfore they have undergonethe preliminary fermentativetransformationsseenin normal metabolism.with the aldehyde
group bound to phosphoricacid, the oxidationtakesplaceat c6, the second
most reactivecarbon h the molecule.This direct oxidation would represent, accordingto our view, one of the sourcesof glucuronic acid. Similarly the sulfuric anion would result from the oxidation of sulfur present
in the organism.They correspondto the oxygenphascof offba.lance
D.

LIPIDS AND

LIPOIDS

147

Glucuronic and Sulfuric Anions

urine specimensthat contain abnormal oxidizing substancesshow significant amounts of glucuronic and sulfuric acid compounds.(Nore Ig)
The analysisof the conditions under which these two substancesexert
anti-toxic activity permits a better understandingof their role in general
biology. A certain parailelism exists, and has always been emphasized,
between a detoxifying and an etminating function exerted by these two
radicals. Not only do sulfo- and glucurono-derivativesappear in the urine,
but it often has been noted that glucuronic acid intervenes when large
amounts of certain substances,such as menthol and phenol, are present
and there are insufficient sulfuric acid radicals to insure detoxification and
elimination. when mineral sulfatesare administered,the proportion of
sulfederivatives increases.
This parellelism appearsespeciallyinterestingwhen we recognizethat
sulfuric acid representsthe end result of the oxidation of sulfur introduced
into the organism in combinations in which it is a bivalent negative element. Only a smaller amount of sulfur is introduced as a hexavalentpositive element: that is, as sulfate. Sulfur is introduced mostly in bivalent
negative form, as in methionine,cystine, etc. Both sulfuric and glucuronic
acid result from oxidative processes,acting in one case upon the thiol
group and in the other upon glucose.
The relationship botweensulfuric and glucuronic acid goes still further.
It has been noted that glucuronic acid appearswhen enough sulfuric radicals necessaryfor detoxifying action are not available. However, this is
not entirely true since one processdoes not duplicate the other. Qualitative
di.fferencesintervene. (Note I9)
The significanceof glucuronic acid in the defense mechanism seems
clearer when we recognizethat, with but few exceptions such as benzoic
acid, all the substanceswith which glucuronicacid combinesare lipids or
lipoids having one or more positive polar groups. The combination with
glucuronic acid takes place through these positive polar groups. (Note 20)
In our view, glucuronic acid like sulfuric acid, has a specific role in the
defense of the organism and this seemsto be directed especially against
lipids or lipoids with a positive polar group. Bound by glucuronic acids,
the latter are eliminated as excrementalsubstances.Glucuronic acid thus
would act against many anti-fatty acid agents.We can conceiveof sulfuric
and glucuronic acids as mears by which organisms are protected against
an exaggerated activity of anti-fatty acid agents. Along the same lines,
when lipoids with positive polar groups are predominant and able to act

148 /

nEsEARcH tN pHysropATHoLocy

in an exaggeratedmanner to oppose the fatry acids physically and chemically, thc same means can be utilized to reduce this exaggeration.Thus, the
intervention of glucuronic acid as a result of an abnormal oxidation of
glucooe induced by fatty acids appears to be biologically sound. This is
also true for the sulfuric radical.
The importanceof thesesubstancesdoesnot resideonly in the fact that
the organism can easily produce them in larger quantities than fatty acids,
The fac-tthat they combine to form excrementalsubstancesis important
too, for in this way, they help in materially eliminating the anti-fatty acid
substancesfrom the organism. This would not take place if only a combination with fatty acids were possible,since the esters of fatty acids are
usually rotained in the organism and, under certain circumstances,can
again liberate their constituents.The intervention of glucuronic acid and
sulfuric acid appearsto be more effeotivethan that of the fatty acids which
have their own activity and are more toxic in exaggeratedamounts.This
appears to be especially true in the case of glucuronic acid becausethe
amount of glucose available is practically unlimited as compared with
other metabolites.Glucuronic and sulfuric acid would thus intervene in
the biological antagonism botween fatty acids and anti-fatty acid substances,inactivating and eliminating agontsfrom the last group, especialty
when in excess.Teleologically speaking,their intervention appears to be
still more interestingsince the body has, as part of its defensemechanism,
a tendencyto manufactureanti-fatty acids in excess.The interventionof
agentsother than fatty acids would prevent a vicious circle and permit an
excessof anti,fatty acidsto be removedby excretion.
FATTY ACIDS VS ANTI.FATTY

ACIDS

This study of the relationship betweenfatty acids and anti-fatty acids has
been guided by the dualistic concept. It must be recognized,however, that
the direct activity of thesesubstances
could be largely reducedto that of
one 8roup, the fatty acids. The action of anti-fatty acids is largely indirect.
They control and thus limit the activity of the fatty acids. It is within this
framework that the different anti-fatty acids selectivelyinfluencedifferent
specificfunctions of the free fatty acids.
The lipids and asscciated constituents with their multiple activities
create for each entity a balance responsiblefor many of the manifestations
of the entity. Variations in manifestationscan be attributed in large part to
qualitative and quantitative variations in the intervening lipids. A sys-

:,,;'r)

LTPTDSAND

LTPOTDS

t49

tematizationof these variations would help us understandmany of the

processesencounteredin normal and abnormal physiology.
As we have mentioned before, the balance between two antagonistic
forces, especially for normal conditions, is not static. Instead, there is
alternating predominance of the forces, which results in an oscillatory
movement. Several groups of such coupled forces, each group with its
proper rhythm, are at work. Operating simultaneously,they make for a
series of very complicated variations. Yet analysis is possible since each
of the variations follows a dualistic pattern. The variations, as they occur
at different levels and with different intensities,have been identified
through various tests. In a secondstep they have been tentativelycorrelated with changesin lipids. And next, didactically,lipid changeshave
been related to various etiologic factors, some intrinsic and some extrinsic.
^Se.r
The influence exerted by the sex of the organism upon l-ipidicbalance
was brought to our attention by a curious effect seen when cholesterolwas
administeredin an ether-oil solution to rats. only the females showed
paraplegiaand ulcerationsof the hind legs.While castrationor administration of sex hormonesdid not alter this response,it was influencedby the
administrationof two groups of lipids. The insaponifiablefraction of human placenta,for instance,was seen to induce a high sensitivityto this
preparation of cholesterol even for males, while the acid lipidic fraction
of placentapreventedparaplegiain females.(Note 2l)
Similarly,the fact that in femalesalone,adipouscells appearedquickly
in the skin of the ear after the application of sulfur mustard could be
related to the interventionof the insaponifiablefractions. (Note 22)
Starting with these observations,it could be seen that, in generar,a
higher proportion of positive lipids exists in females than in males. This
could be shown by direct analysesand by analysesof manifestationsrelated to such lipids. while many differencesare to be seen in various
manifestationsbetween females and males, only some could be related
to the direct or indirect interventionof sex hormones.In such instances.
castrationwith or without the administrationof sex hormoneswas able to
change,and even to reverse,the differencesin manifestationsseenbetween
sexes.However, in instancesin which thesemeasureswere without effect,
the differencescould be relatedto rhe interventionof lipids.

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x E S E A R c Hr N P H Y s r o p A T H o L o o y

Age
The changesin lipidic balancerelated to agehave been made the object
of an extensivestudy which also sought to determine the role of lipids in
aglng processes.A general predominanceof positive lipids, more manilest
in the cellular and tissue levels than in the blood, was seen in youth. This
would be expected in view of the special motabolic influence exerted by
this group of lipids. The anoxybiotic charaoter of metabolism induced by
sterolsresultsin the intervention of dehydrogenases
which lead to an abundance of hydrogen ions. This, in turn, leads to a predomhance of the kind
of syntheseswhich favor anabolism. Growth thus could be related to the
predominanceof lipids with positive polar groups, especiallysterols.
Agng processes,on the conEary, could be related to a predominance
of lipids with negativo polar groups, especiallyfatty acids. This predominance could be found especiallyat the cellular level, as seen in cultures of
tetrahymena.(Note 23) ln complex organismsor in rats (Note 24) in which
an increasein the proportion of fatty acids at the cellular level is present,
an opposite change occurs at the systemic and even at the organic level.
There is an excessof cholesterol,this time limited to the higher levels, as
revealedthrough analysesof the blood, for instance.Changesin the blood
vesselsare related in part to this excessof sterols at the systemic level.
Many manifestationshave confirmed such an offbalance with sterol predominanceat higher levels.For example,we found the urine surfacetension
abnormally high in old age. (Note 25l Similarly, skin wheal absorption
in old people requires more than 90 minutes for completion as against
approximately 20 minutes in middle-agedaduls. A predominanceof fatty
acids at lower levels and of sterols at higher levels would thus characterizs
the changesin lipidic balancerelatedto old age.(Fig. 68)
O ther Physiological F actors
The study of the role of lipids in various physiological functions was
made indirectly for the most part, using the tests previously mentioned
which were interpreted in terms of dualistic patterns. These were related to
the general offbalances A and D and, through them, attributed ultimately
to a predominanceof sterols or fatty acids.
Sleep in itself, without relation to night or day, was found to induce
a marked change,comparableto a type A offbalancewith predominance
of sterols. Subjects with pain of an acid pattern often correlate the appearanceof pain with sleep, the pain occurring uniformly at the moment
they wake up. In these cases,the urine shows a low specific gravity with

L T P T D SA N D

LTPOTDS

l5l

a high pH and a high surface tension,correspondingto an A type offbalance.As we will see, in subjectswith an intensiveA type offbalance,
nocturnal polyuria and pollakiuria occur.
sexual intercoursein males was seento induce,in analyses,transitory
changessimilar to an offbalanceof type D, correspondingto a predominance of fatty acids.In femalesthe changecorrespondsto a transitory offbalanceof type A, manifestedby changesat the systemiclevel. Muscular
exercisewas seen to induce, in a first phase during the exerciseitself,

r00

too

60

/"

40

20

0 1 0 20 l0 10 50 60

minutes
S u b j e c t sb e t w e e n
15 and35 years

l020lo{o5060?oo

minutes
S u b j e c t sb e t w e e n
65 and87 years

F I c . 6 8 . T h e d i s a p p e a r a n ctei m e f o r t h e w h e a l i n d u c e d b y t h e i n t r a d e r m i c i n j e c t i o n
of 0.2 cc saline, varies with the age. In old age, the wheal oftcn persistsfor more
than 90 minutes.

changescomparableto an offbalancetype D. This phaseis followed by a

much longer phase of type A, indicating sterol predominance.Intensive
rnentalexerciseproducesa markedchangesimilar to offbalanceA, with all
urinary testsshowingthe patternsfound with predominanccof sterols.
The responsesattributed to influencesexerted by external lactors could
be integrated in tle same dualistic mechanism,All the data indicate the
manifest influenceexerted by the time of day. Two marked changesare
seen,one around four o'clock in the morning and the other usually around
eight or nine o'clock in the evening.The morning changecorrespondsto a
predominanceof sterols,the eveningto predominanceof fatty acids.These
changes together with the clinical manilestations related to time of day

152 /

REsEARcHrN pHysropATHoLocy

appear in a new light when interpreted not as being the direct results of
time changesbut rather of patterns of diurnal activity and nocturnal rest.
This explainswhy in rats and mice, which are nocturnal animals,most of
the analysesshow variations related to the time of day opposite to those
in humans. other variations could be recognizedmore strongly related to
time of day. variations with a 24-hour rhythm could be seen,for instance,
for urinary surfacetension in mice. But, when rats and mice were maintainedfor a length of time under artificialcondirions,with light during the
night and dark during the day, the animals changedtheir habits, becoming
active during the day and sleepingduring the night. Afrer a certain time,
most of their analytical patterns such as urinary pH, blood leucocytes,
eosinophilesetc. changed,acquiringthe type of variation seen in humans.
Urinary surface tension remained unchangedfor a long time. (Note 8
chapter IV) Even more interestingwere other changeswhich could be related to changes in externa.ltemperature.The urinary surface tension
measuredin rats in the morning for long periods of time showed variations relatedto changesin the temperatureof the environment.(Note 26)
(Fis.69)
The importanceof temperatureled to its more detailed investigation.
Variations in lipidic balancehave been found to para-llelvariations in body
temperature.The blood of normal individualsis richer in sterolsthan the
blood of those with hypothermia.Furthermore,in moments of high temperature,more sterolsare found than in momentsof low or normal temperatures.An increaseof fatty acidsoccursin conditionswith hypothermia.
These changeswere confirmed also by the correlation between blood content in lipids and temperaturein different abnormal conditions. In shock
with hypothermia the blood is rich in fatty acids, while in infections with
fever.it is rich in sterols.
The role of.temperaturewas also investigatedby studyingthe influence
upon the lipidic balance by externally applied heat or cold. Characteristic
variationscould be seenin human analysesunder the influenceof hot and
cold days, and of local applicationsof heat and cold. Manifestationscorrespondingto predominanceof lipids with positive characterwere induced
by heat, while others correspondingto predominanceof Iipids with negative characterwere inducedby cold. Variationswere seenin animalskepn
in an incubator or in a refrigerator. ( N ote 27) w e will seebelow, by studying their influence at different levels, the importance of these variations
produced by temperature.
The influence exerted by barometic change.scould be seen in changes
in total blood potassium,the two curves being paraUel.Similar changes

LTPTDSAND

LTPOtDS

153

could be observedrelated to the atmospherichumidity. Other tests as well,

such as urinary pH, calcium excretion,etc., show a similar relationship but
to a much lesserdegree.(Note 28) The influence exerted by the environment could explain the changesseenfrom one day to the other in various
analyses.(Note 28)

t2
t,
q,
L

5t

.\t ao
(-

c,
o- 32
E
o,

6'

v,

(u

.U
z

v,

(a

s5
vt

(u

(t
E
(u

6t

l!

./,
q

0aYs

Ftc. 69. The curves of the average values of surface tension of two groups of 20
malc rats cach, and two groups of female rats each, show parallel changes with thc
inverse curve of the temperatureof the environment.

ths influence exerted by changesin seasonswas studied. An increase

in fatty acids in winter and of sterols in summer could be noted. These
increa-sesalso could be largely related to the seasonalvariations in temperature. Very hot days were marked by analysesindicating intervention of
lipids with a positive character. The influence exerted by the seasonswas
sen even in the responsesof organismsto pathological conditions such as
t.:t,t:i

154 /

nESEARcHtN pHysropATHoLocy

tumors; variations in character and growth of experimental cancer could

be noted. (Note 29) The relationship of many viral infectious diseasesto
seasonalchangeswhich has been noted in many epidemiologicalstudies
coufd be rclated to the changesin lipids. (Note 30)
Efiect ol Antagonistic Lipids a Difierent Levels
A more complete study of lipids under the dualistic concept was made
by considering their activity at different levels of hierarchic organization.
This researchwas greatly facilitated by the degree of individuality which
different biological levels exhibit when they are part of the hierarchic organizationof complex organisms.It was also aided by the availability of
lower organismsin nature which correspondto various hierarchic levels.
Through this double approach,the information obtained showed the importance of the relationshipbetweenthe levels of the complex organism
and the influence exerted by lipoids. If high doses of the agents are applied, the inJluenceis exertedupon all the levels.A preferentialinfluence
is exertedupon a singlelevel if reduceddosesare used.when medium size
dosesare administered,to the preferentialeffectupon one level a reactional
responseat other levelsis added.This resultsoften in concomitantopposite
effectat theselevels.
Eflects ol Lipoids on Viruses
The antagonisticeffectsof positiveand negativelipids were evident in
the sfudy of their action upon viruses.Generally,agentswith a positive
polar group appcaredto create favorableconditionsfor the development
of viruses,while those with a negativepolar group had an oppositeeffect.
This influence,which was first seenin phagesin vitro, becamestill more
evident in viral infections.
subcutaneousadministrationof positive lipids, such as sterols or insaponifiablefractionsof organs,inducedgreaterlocal receptivityto viruses.
In experimentswith smallpox virus in rabbits,for instance,virus inoculation of the skin induced an exaggeratedresponsein those areas where
positivelipids previouslyhad been injectedsubcutaneously
comparedwith
the responsein other previouslyuntreatedareas.In less sensitivespecies
such as mice and rats, positive lipid injectionsinduced abnormally high
local receptivity to virus inoculation. Intracerebral injection of sterols
followed by subcutaneousinoculationwith smallpox virus invariably produced nervous system localizationof the virus. Intraperitonealadministration of sterolsin very high dosesin mice prior to smallpox inoculation
produced a great degree of central nervous system localization. Intra-

L T P T D SA N D

LlPOlDs

155

cerebralvirus inoculation,after subcutaneous

administrationof high doses
of anti-fatty acids, brought death earlier in test animals than in controls
given intracerebralvirus alone.
A sniking opposite effect was noted for lipids with a negativepolar
character. In rabbits, subcutaneousinjection of a polyunsaturatedfatty
acid set up a local skin area refractory to smallpox virus inoculation,although inoculation was positive in other areas of the body. Death also
occurredlater,followingintracerebral
inoculationwith a neurotropicvirus
in test animals given subcutaneousor intraperitonealinjections of fatty
acids, than in controls.This partially protectiveeffect was oppositeto the
increased receptivity seen in animals injected subcutaneouslyor intraperitoneallywith insaponifiablefractionsand intracerebrallywith the same
virus, wheredeath appearedearlier than in controls.
The antagonisticeffectsof the two groups of tipids for viral infection
appearedinterestingfrom severalpoints of view. The cffects were local,
at the cellular level, where virusesthemselvesact. Subcutaneousinjection
of lipids inducedmanifestchangcsin responsetoward the virus in the skin
at the site of injection, and little or no changeat all elsewhere.we have
utilized this fact, as we will seebelow, to obtain information regardingthc
level at rvhich various agents act. A change induced in receptivity to
viruses,limited to the skin at the site of injection, woutclindicate activity
of the agentat the cellular level. Tests basedupon the skin responseto
smallpoxvirus infection have shown that, among the lipids with a negative
polar character,a maximum of influenceis exertedby the insaponifiable
fraction of organs of exodermicorigin from speciessensitiveto the virus.
The insaponifiablefractionsof rabbit skin and rabbit brain were the most
activeof the lipids tested.Among the fatty acids,the prevcntiveeffectwas
seen to increasewith the degreeof desaturation.It was almost entirely
absent in saturatedfatty acids, notably present in polyunsaturatedfatty
acids.
The increaseand decreasein receptivityof the skin to smallpox virus
following injection of lipids also furnishedinformation about the roles of
the polar and nonpolar parts of lipids in this specificactivity. An opposite
effect was seen between two groups of substanceshaving the same nonpolar group but differing in their polar groups.While the polyunsaturated
fatty acids of saffower oil, for instance,greatly reduced receptivity, the
same polyunsaturatedmembershaving alcoholsas polar groups increased
receptivity. The polar group--negative or positive-appears to be the
factor inducingthe oppositeeffect.
The role of the nonpolar group was studied by comparing saturated

156

REsEARcH rN pHystopATHoLooy

and unsaturatedacids and alcohols. Almost no activity was seen for the
saturated.The unsaturatedmemberswere active in general,with activity in
any direction increasing with the degree of desaturation of the nonpolar
group. Thus, it appears that the nonpolar group determines whether a
substanceis active or inactive, but the nature of the activity-that is, increasingor decreasingreceptivity-is determinedby the polar group.
The influence exerted by agents with a positive charact,er upon viral
infection would explain the seasonal changes in clinical manifestations
which are especially interesting for the paralytic form of poliomyelitis.
we could show experimentally that when mice, after being inoculated
subcutaneously with smallpox vaccine virus, are kept in an incubator at
37"c, all develop cerebral involvement, while such involvement appears
in only a small proportion of other animals kept at room temperature, and
does not appear at all in those kept in a cool place. As we could also show
that one of the effects of exposure of an animal to a higher temperature
is an increase in the body of the amount of free lipids with positive character, this could explain the increasein the virus sensitivity of cells in the
central nervous sys0emwhich are especiallysensitiveto these lipids. This
relationship would also explain the increasedincidence of paralytic polio
casesduring hot weather.
The presence of greater amount of lipids with positive character in
youth helps also to explain the frequency and intensiry of viral infections
in children.(Note 31)
The study of the effects of temperature and lipids upon viruses has
shown that those effects are not limited to the host but also are exerted
upon the viruses. (Note 32) The influence of heat and cold upon virus
activity was studied in bacteriophages,where effects for virus and host
could be separated.The direct influenceupon the virus appearedrelatively
small and secondaryto the changeswhich appear in the host itself. Bacteriophage,separatedfrom microbes by filtration and kept in an incubator at
temperatures 2-3 degrees C higher or lower than controls, showed no
change in virulence. This was true as long as microbes were not present.
Microbes kept at higber temperaturewere more sensitiveto phages;when
kept at lower temperature, they were less sensitive. This influence went
so far as to change a sensitivestrain to a refractory one, and vice-versa.
The fact that microbesgrown at higher temperaturesfavor the develop
ment of bacteriophagewhile those grown at lower temperatue hinder it
could be correlatedwith the changein the richnessof lipids in the microbes
themselves.Similar results were obtained when microbes were grown for
a time in media containing fatty acids or insaponifiablefractions and were

l;r:iS$r
i

LIPTDSAND LrPolDS

157

then removed and exposed to phages.These experiments(Note 3Jl indicate the direct role played by the lipids of the hosts in the activity of
phages,and would explain the influenceexerted by temperature.Through
the change in the lipids of the hosr, the virus changestoo, becoming more
active if grown in microbes at a higher temperature and less aggressiveif
passedthrough microbes kept at lower temperature.
Efrects ol Lipids on Miuobes
The antagonisticeffectsof the two groups of lipids upon microbes were
investigated. As an example, we will mention here the characteristic
changesin Bac. anthracis treated with polyunsaturatedfatty acids and insaponifiablefraction preparations.(Fig. 70) We investigatedthe microbes
for their morphological, tinctorial, cultural and virulence characteristics.
with the fatty acids added to media, changeswhich can be consideredto
be mutationalwere induced,leadingto tiny Gram negativemicrobesgrowing on agar as transparentsmall colonies.The changes,however, were
reversible.Usually several passagesin normal media were sufficient to
produce reversal.First small and separate,then larger and more confluent
Gram positive granules were seen to appear in the microbes which, themselves, also became progressivelyplumper. Ultimately, all the characteri51is5,-rnsrphological, tinctorial and cultural---of the normal microbes
reappeared
. (Fig. 7l )
Microbes showed opposite changes when treated with insaponifiable
fractions,(Fig. 70) losing their bacillusform. Abnormally intensiveGram

6
(a)

treated.
with
sterols

i=L
\-

(b)

(c)

t?:ttio.:ilh

controI

Fro.70. lnfluence ol lipids upon microbes. Schematic drawing of the changes in.
duced in Bacillus Anthracis by the influence exerted by the two groups of lipids.
Treatcd with sterols (a) as in tbe unsaponifiable fraction of placenta, the microbes
change into cocci irregularly shaped and intensely retaining the gram stain. Treated
with fatty acids (c) from cod liver oil, the bacilli change into very tiny gram negative microbes. (b) shows untreated microbes.

..,;

t.t;.'i,*i;;',;,.

."t.

""#.;-14

158

nEsEARcH rN pHysropATHoLocy

=/--'
- -

a{a*

e&

-{.'
\

Fto' 71. Lipids and microbes. Drawing of the progressivepassagetoward normal

bacilli of the tiny gram negative microbes obtained through the treatment of Bac.
Anthracis with fatty acids. The passagetakes place usually in successivesteps. Thc
gram positive formations appear first as fine granules;they later become clumps ancl
finally give the microbes their normal aspect.

positive cocci appeared. They grew on agar as very thick creamy white
colonies. These changeswere seen to persist for a long time and seldom
were sPontaneouslyreversed.Treatment with fatty acids induced reversal
although inconsistently. we attempted to correlate the differences in
changesinduced by different lipids to the different levels of the microbe
at which they work. The changeto cocci can be regarded as corresponding
to an influenceexerted upon the membraneand the change to Gram positive to an influence upon differentiated formations present in the body.
( 3 1 3)
Effects of Lipids on Protozoa
The effects of lipids upon monocellular organisms,especiallytetrahymena pyriformis, were studied and an effort made to relate the nature
of the main changesinduced in these protozoa to changesobserved at the
cellular level of complex organisms. An initial effect was noted on the
polarity in protozoa which seemed to be oppositely influenced by long
chain polyunsaturatedfatty acids and sterols.Lipids with a positive character were seen to induce a change in the form of protozoa causing them
to become almost round, a change considered to correspond to reduced
polarity. Lipids with a negativecharacterhad an oppositeeftect;the tetrahymenabecameabnormallyelongated.
The administration of higher amounts of polyunsaturated fatty acids
was seento induce immediate changeslocalized at the anterior pole of the
organism, changeswhich ultimately lead to the breakdown of the membrane particularly at this point. This effect parallels in intensity the degree
of desaturationof the fatty acids. other changeswere seen in growth rate
and survivxfrime and, thus,in the agingprocess.(Nole 3a) (Fig.7a)
At the same time, resistanceto heat was seen to increaseas the result
of treatment with negative tipids, while it decreasedafter treatment with

LIPIDS AND

ControI

LTPOIDS

r59

with
Treated
fatty acids

Ftc. 74. In a direct action of fatty acidr on tetrahynlcna, a pas:,agcof fluicl occurs
at the surface with a break of the membrane especiallymanifest at the anterior polc
( a ) , c o n t r o l u n r r e a t e d( b ) , ( 1 2 0 0 x ) .

the positive sterols.(Note JSl The sameinfluenceupon thc aging processes,

as manifestedin a prolongation of the lifc-span, was noted for polyunsaturated fatty acids with a long chain and evcn for some mcmbcrs of the
saturated seriesbut with a shorter chain.
Effects ot' Lipids on Complex Organisnu
Morphological changes-The same level separation was uscd in thc
study of the effectsof lipids on complcx organisms.Acting at chromosomal
levels, lipids led to the appearanceof monstrosities.various lipids, especially insaponi{iablefractions of organs, rvere injectcd into larvae of flies.
While an immediate change in the cells of the larvac could be traccd to
the subnuclear level as secn in chromosornes,monstrositiesrvere seen to
be induced in the resulting flies. A similar eJTectbccame evident when
lipids were injectedinto hens' eggsbeforeor during incubation.Especially
with cholesteml but also with insaponifiablefractions of organs, a high
proportion of chickens were hatched with spasticparaplegia.
The sarneproblem is being studied, in collaboration with P. Fluss, in
Drosophila melanogaster, grown for many generations in nedia to which
an entire scriesof difierent lipids from one or the othcr group are addcd.
This study is in progressand the resultswill be published later.
We have seen that the antagonisticeffects induccd by the two groups
of lipids could be related ultimately to oppositechangesin the fundamental
biological proces$ of aging. This appeared clear for lower morphologic*l
levcls of organization and was especially evident for cells. While anti-fatty

160 /

R E S E A R c Hr N p H y s r o p A T H o L o c y

acids induce changes which can be regarded as corresponding to prolonged

youth, the polyunsaturatedfatty acids induce rapid aging with pyknosis and
karyorrhexis and death of the cells as old entities. This could be seen
clearlyin tumors,in which cellswith youthful characterlead to non-necrotic
tumoral ma.sses,
while cells that age rapidly produce necrosisin the tumors
followed by ulceration il the tumor is superficial.
The eftect of conjugated fatty acids was somewhatmore complex, indicating an abnormality in the induced processes.Their sdminisfixtion was
followed by the appe:uance of cytoplasmic and even nuclear vacuoles,
correspondingultimately to an anomaly of water metabolism.
The effect of lipids upon adipous cells appearedto be of special interest. The anti-fatty acids, especially the sterols, when injected subcutaneously in animals, induced a characteristicprocessin the adipous cells near
the injection site. These cells became very enlarged and higlrJy irregular,
with their content changed into an emulsion only slightly stained with
sudan. The fatty acids, on the contrary, imparted to adipous cells an abnormal resistanceto destruction. They remained persistently unchanged
even in the midst of very active processeswhich usually cause them to
disappear.Unchangedadipouscells were found encircled by the hvading
cancerousceus,deep within tumors in animalstreatedwith fatty acids.
On Pain-From the start of this research,the opposing effects of the
two groups of lipids upon pain has been most impressive. For the fatty
acids the degree of saturation is important. The saturated members of the
fatty acid seriesand even oleic acid are entirely without effect. Linoleic
and linolenicacidsshow a slight influence,while the polyunsaturatedmembers show a marked effect. Administration of highly unsaturatedfatty
acids and of acid lipidic fractions of certain organs,such as placenta,liver,
spleenor blood, uniformly decreasedpain of an acid pattern and increased
pain of an alkaline pattern. These oppositeeffectshave, from the beginning
of our study, contradictedthe idea that this influenceupon pain was the
result of the direct action of tlese agentsupon the nervous system.Furthermore, the opposite effects exerted upon the same pain by the other group
of lipids have confirmed the hypothesisthat the acrion takes place at the
level of the painful lesion, where the difterencesbetweentre two pains was
found to correspondto two oppositeacid-baseoffbalances.
In the study of the effectsof lipids on the pH of the secondday wound
crust,madein collaborationwith CarlosHuesca,we have demonstratedthat
lipids influence pain through changes induced on the acid-base balance
presentat the tissuelevel. The poeitivelipids constantlylowered this pH
while the negative lipids elevated it. (Note I Chapter V) Even more im-

L T P T D SA N D L T P O t D S /

151

portant than this temporary pH effect in establishingthe mechanism of

lipid action in pain was the change in the actual pattern of an existing
pain after administration of these agents. Polyunsaturatedfatty acids in
sufficient amount were found to convert an acid-pain pattern to an alkaline pattern, while sterols changed an alkaline pattern into an acid one.
We will return to this important fact later.
A pathogenic role for lipids becomesevident too, when pain can be
induced through the administration of lipids in previously painlesslesions.
Such lesions treated with large amounts of lipidic preparations became
painful. An alkaline pattern of pain was seen to appear after fatty acid
administration, while an acid pattern followed use of the insaponifiable
fraction. (Note 36)
At the tissuelevel,lipids also affectsuchacid-basesymptomsas vertigo,
itching, dyspnea,tremor, and even mental diseases.
In theseconditionsthe
same antagonismbetweenthe two groups of lipids-and the same opposite
effects upon the acid and the alkaline pattern---can be noted along with
the same possibility for changing the pattern to the opposite type if big
doses of lipids are administered.
wound Healing-The same manifest antagonism between the two
groups of lipids was also noted in their influence upon the evolution of
wounds. Changesin the sloughingand healing processwere followed by
measuring the size of wounds (Note 37) as well as by serial histological
examinations.The lipids with a negative polar group were seen to retard
the evolution of the processesby prolongrngthe first catabolic phase. positive lipids generally had an opposite effect. However, here too it could be
observedthat sterolshave relatively little effect on the healing of connective
tissue, but manifestlyfavor proliferation of the epithelia. This was especially evident in the changesin scar formation of the skin of treated animals. In rabbis treated with cholesterol,the epithelial scars were found to
have 8-10 layers insteadof the 2-3 characteristicfor the rest of the skin
and for the scars in control animals.
Regenerati6n-11 collaboration with E. F. Taskier we studied the
eftect of lipids upon the regenerationof liver in rats, after the resectionof
almost 3/q of. this organ. The rate of regenerationcould be estimated by
correlating it to the time of appearanceof fatty droplets filling up almost
all the cells. as a first step in the regenerativeprocess.In very young animals, this changein fatty liver cells was seento take place even within the
first 24 hours after resection.The change was progressivelydelayed as the
age of operated animals increased.In old animals the change in fatty liver
cells appearedonly after the fourth day.

162 /

REsEARcH rN pHyslopATHoLooy

The administration of lipids had a marked effect on appearance time

of fatty cells. Sterols induced precociousappearancein old animals. From
this point of view, sterol-treated animals appeared to react as young individuals, with fatty cells evident even on the secondday. The fatty acids
and acidJipidic fractions of organs showed an opposite effect, delaying the
time of apparance of the fatty cells. Young animals treated with polyunsaturated or conjugated fatty acids showed no fatty droplets in the liver
cells for as long as three to four days. with higher doses of the same
agents,the fatty infiltration did not occur at all.
It is interesting to note a parallelism between fatty infiltration of liver
cells and the richness of adrenals in sudanophil subsrances.An almost
complete lipid depletion of the adrenals was seen after high doses of fatty
acids and coincided with a total lack of fatty cells in liver regeneration.
(Note 38)
organic Level-Effects of the two antagonistic groups of lipids at the
organic level have been studied in terms of manifestationsclearly associated with various organs. we will review these effects briefly here, with
more details to come when the therapeuticuso of lipoids is discussed.
lnlgslings-The influence of lipids upon intestinal function is marked
by the same antagonismbetween the two groups of agents.oral administration of large amountsof fatty acids,especiallyhigher unsaturatedsuch
as obtaincd from cod liver oil, was usually fotlowed by diarrhea. Diarrhea
also occurred after parenteral administration of these substancesin large
arnounts. It was interesting to note that parenteral administration of the
acid lipidic fraction of placenta, blood or even organs had a marked influence upon the colon and rectum in particular. High doses produced
tenesmuswith a mucous or even sanguinolentsecretion.This localization
of the effectsof the lipidic fraction appearedto be especiallyinteresting
from a therapeutic point of view, as will be seen later. The oral or parenteraladministrationof the oppositegroup of lipids, sterolsand insaponifiable fractions, has an opposite effect, a constipating one, which we will
discusslater together with its therapeuticaspects.
Kidney-The manifcst opposite effects exerted by the two groups of
antagonisticlipids upon diuresis raise the question of where these effects
take place. While a systemiceffect can be recognized,a more direct intervention upon the kidney also must be considered.The addition of the acid
lipidic fraction of organs,and especiallythose obtainedfrom pork kidney,
to the perfusion fluid in a dog kidney preparation produces a manifest
decreaseof excretedurine. The administrationof insaponifiablofraction

LtPrDs AND LTPOTDS /

163

has a marked diuretic eftect which we will discuss below with its therapeutic aspect.
Nervous system-rnteresting eftects by the two groups of antagonistic
lipids upon many manifestationsof the central nervous system have been
noted.
Convulsions-Administration of sterols and insaponifiablefractions of
many organs such as placenta, liver, butter, eggs, etc., in large amounts
induces convulsionsin rats. Convulsionsalso were noted in humans when
huge dosesof these agentswere administered.But even in relatively small
amounts,theselipid agentssensitizedanimalsto the administration of other
c-onvulsantagents.In rats or mice receiving such lipids, thiamine chloride
induced convulsionsin doseswithout effect in controls. (Note 39)
An opposite effect was observed for lipids with negative character.
Saturatedfatty acids showedno influenceon thiamine-inducedconvulsions.
Such convulsions were prevented by the administration of nonsaturated
members.The effect was related to the degreeof desaturation of the fatty
acids. with increasesof the iodine number, the necessaryeffective doses
of these fatty acids becameprogressivelysmaller. While hundreds of milligrams of mono- and diethenicacids were necessaryfor each 100 gram of
body weight, the anti-convulsanteffect was obtained with only a few milliSams of clupanodonicacids,and with still lessof the nonenicacid, bixine.
The study of the pathogenesisof convulsionsalso covered the influence
exerted by these lipids of the adrenal corticoids. The administrationof
mineralocorticoids,especiallydesoxycorticosterol,
even in small doses,to
who
subjects
had receivedany one of the lipids with a positive polar group,
such as cholest,erolor insaponifiablefraction of placenta,liver or kidney,
was followed almost invariably by convulsions.we will presentmore details on this effect later in the discussionof syntheticsubstances.For the
moment we want only to note the relationship between mineralocorticoids
and lipids with positive character in the pathogenesisof convulsions.The
concomitant intervention of the two factors-an oftbalance induced by
lipids with positive character, and action of mineralcorticoids-seems to
provide new light on the pathogenic problem of epilepsy and convulsions
in general.
Coma-The role of cortical hormonesin the pathogenesisof convulsions was confirmed by the opposite effect produced by neoglucogenic
corticoids. We will see later that the administration of cortisone to subjects
receiving higher alcohols such as heptanol, octanol or octandiol in large
doses,induced a subcomatosecondition at first which progressivelychanged
into coma. (Note a0) Oppositepropertiesof the mineral and neoglucogenic

164 /

REsEARcH tN pHysropATHoLocy

corticoids,which made Seyleseparalethem accordingto their "phlogistic"

and "antiphlogistic" activity, would explain the two opposite manifestations inducing convulsionsand coma, produced in individuals previously
treated with the same anti-fatty acid agents.
On Cardiac Rhythm-The influence exerted by the two groups of lipoids upon the cardiac rhythm was studied under the same dualisl"icaspect.
The effects observed can easily be interpreted considering the role of the
differentiation of the cardiac cells for their part in the cardiac physiology.
The role of a cell in cardiac physiology is a direct function of its own
automatismwhich can ultimately be related to its degreeof differentiation.
The fact that the two groups of lipids acr antagonisticallyupon this cellular differentiation, the acid lipids exaggeratingit and the insaponifiable
fraction of sterols reducing it, has explained some of the effects induced
by these agents upon normal and abnormal cardiac rhythm. (Note 4l)
on oestral cycles-The action of the two groups of lipids at the organic level was also studied in the rat ovarian cycle. Daily, and even twice
a day, vaginal smears were made in animals treated with these agents.
when large amounts were administered,both groups suppressedthe cycle.
with smaller doses,only the lipids with positive polar groups, especiauy
sterols,produced this effect.
Systemic Level-Blood has appeared especially suitable for in vitro
and in vivo studiesof the effectsof the two groups of lipids at the systemic
level. The effects on different blood constituentswere analyzed and led to
very conclusive results. We will outline here the principal points of this
study.
Under the influence of anti-fatty acids, the erythrocytes become more
turgescent increasein volume, show a strong refringency of their crown
in dark field examination, and remain isolated from one another. The
sedimentationrate, if previously high, is reduced by treating blood in vitro
with insaponifiable fractions of organs. oxygen appears to be retained
longer in treated red cells than in controls.
The fatty acids have an opposite effect. Under their influerrce,the red
cells becomecrenelatedand develop a tendencyto form sludges.The sedimentation rate is increased.The color of the treated blood is dark and,
even after oxidation, rapidly darkens again. In vivo, lipids wittr a positive
charaoter induce leucocytosis,tlose with a negative character leucopenia.
This last effect is seen even in vitro. In Note 42, the influence exerted by
lipids upon the blood is presentedwith more details. (36)
On Temperature-The administrationof sufficient amoun{s of positive
lipids induces a frank elevation of temperature,while hypothermia follows

LrPrDs AND LtPorDs

165

thc administrationof negativelipids. The relationship betweentemperature

and lipids, however, is not so simple since changesin external lemperature
influencethe balance betweenthe antagonisticlipids. For example, animals
kept in incubators at a temperature of 35oC show an increase in lipids
with a positive character.Animals kept in a cool place, such as a refrigerator, show an increasein lipids with a negativepolar group. The organisms
are able to combat the increaseof lipids with negativecharacter by means
of the normal defensemechanism,but are less capable of dealing with an
increaseof lipids of pcitive character.Therefore, while a high proportion
of animals kept in refrigerators adapt themselvesto the new conditions,
those in incubators die in a few days.
on systemic Patterns-The influence exerted by lipids upon various
other systemic manifestationswhich are reflected in abnormal patterns in
urine analyseshas been studied. In genora.l,the fatty acids induce patterns
corresponding to the offbalance of type D, while the sterols induce patterns
of the type A offbalance.Here again we must emphasizethat any lipid, if
administered in large quantity, influencesa.ll analytical values. A certain
specificity, however, is noted since, in relatively small doses,lipids induce
changesonly in certain values. Becauseof the inherent technical problems
conceming the patterns, only a few analysescould be followed accurately
over the period of time necessaryfor a clear recognition of changes in
small laboratory animals. It is for this reason that most of our studies in
this area were made on humans where pattern changes could be easily
identffied and followed over long periods.
It is to be emphasizedthat, under these conditions, the influenceof
lipids is exerted especially upon zrlreadyexisting abnormal patterns, increasingor decreasingtheir deviationsfrom the normal, or changingthe
patterns entirely. Abnormal patternswere induced through huge amounts
of lipids, which very seldomwere administeredto patients.Tnelp X shows
schematically the analytical changesinduced by the two groups of lipids
upon various urine and blood analyses,expressedas patterns corresponding to abnormalconditions,as well as upon the manifestations
presentat
other levels.
we will discuss these effects in more detail when describins the
pharmacodynamicpropertiesof lipids and lipoids.
Mechanismol the Lipidic Biological Activity
The analysisof the changesinduced by lipids has emphasizedcertain
characlerswhich appearof capital importancefor the understandingof the
biological interventionof thesesubstances.
In one kind of activity a lipid

rirt';:l;

l.';.rr

166

RESEARCH IN

PHYSIOPATHOLOGY

acts throughits Iipoidic properties.From the data concerningits distribution

in the organismit can be seenthat, due to its solubility characters,a lipid
introducedin an organismwill be selectivelyretainedby the existinglipidic
system.When such interventionthrough its lipoidic propertiestakes place,
the nonspecificcharacterof the activity of the lipid is prevalent.A second
kind of activity results from the bond realized through the charge ol the
polar groups.The positiveor negativecharacterof thesepolar groupsdeterTrsle X
LEVEL

EFFECTS OF STEROLS

EFFECTS OF F^TTY ACIDS

Cells

Prolongsyouth character
Increases
potassiumcontent
Decreasessodium content
Reducesmembranepermeability
Reducescellularoxidation
Reduceschloridecontent

Inducesrapid aging
Decreasespotassiumcontent
Increases
sodiumcontent
I ncreasesmembranepermeability
I ncreasescellular oxidation
Increaseschloride content

Tnissaes

Lowers pH of lesions
Lowers chloride content of
lesions
Lowers water content of lesions

RaisespH of lesions
Raiseschloride content of
lesions
Raiseswatercontentof Iesions

Organs

Inducessomnolence
Inducesdiuresis
Inducesconstipation
lnducestachycardia

Inducesinsomnia
Inducesoliguria
I n d u c e sd i a r r h e a
lnducesbradvcardia

Systemic Induceshyperthermia
Induceshypertension
Blood
Increases
RC volume
DecreasesRC sed.rate
Increasespersistenceof
oxygen fixation
Determinespersistence
of
RC isolation
Determineshyperleucocytosis
Determineseosinophilia
Decreaseskalemia

Urine Induceswater excretion

Inducessulfhydrylretention
Inducescalciumexcretion
lnduceschlorideexcretion
Inducessodiumexcretion
Inducesphosphateretention
Inducesretentionof surface
active substances

Induceshypothermia
Induceshypotension
DecreasesRC volume
IncreasesRC sed. rate
Decreasespersistenceof
oxySenfixation
Determinesformation of sludge
Determinesleucopenia
Determineseosinopenia
kalemia
Increases
Induceswater retention
I nducessulfhydryl excretion
lnducescalciumretention
I nduceschloride retention
Inducessodiumretention
Inducesphosphateexcretion
Inducesexcretionof surface
activesubstances

LIPIDS AND LIPOTDS /

167

minesthus the natureof this secondkind of activity.A third kind of activity

resultsfrom the chemical constitutionof the polar group, which will induce
selectivecombinationsand consequentlywill have a more specificinfluence.
A fourth group of changesare inducedby the activity which takes place at
the nonpolar group of the lipid and more specificallyat the energeticlormations presentin it. They will have a still higher characterof specificity.
with this systematizationof the activity of the lipids, a further systematic analysisof the influenceexerted by the lipids appears possible.
Through its selectivedistribution, the administrationof a substance
having lipoidic propertieswill influencethoseentitieswhich have lipids in
an active form in their constitution.The influence exerted will thus be
proPortional to the richnessof the entity in theseactive lipids. This fact
explains why the administrationof a lipid or lipoid affectsselectivelythe
abnormalentitiesrich in free lipids and to a much lesserdegree,the normal
ones. It is this selectivedistributionwhich will further limit the activity of
the lipoid to the lipidic systemand most manifestlyto the abnormalentities.
In the frame of this limitation, this activity resultsfrom the chargeof the
polar group. Similar effectsare thus obtained for all the different lipoids
which have the same electric positiveor negativecharacterof their polar
group. This explainswhy one can usedifferentagentsfrom the samegroup
and still obtain similar results.Agentschemicallyso differentas fatty acids,
mercaptans,persulfides,aldehydesor epichlorohydrine,have similar activity becausethey all have negativepolar groups.The characteristicof the
effectsresultingfrom the electricalcharacterof the polar groups,is that they
are common for the groupshavingthe samesign and diametricallyopposite
for the agentswith a positiveor a negativepolar group.
This effect was clearly seen in fatty acids in which the negativeciuboxylic polar group was changedinto the positive primary alcohol. The
biological effects of the new substancewere opposite.
It is in the third kind of activity that the chemicalnature of the lipoids
intervene.Certain effectsresultingfrom the bond of an amino polar group
will thus be differentfrom that of the alcohols,althoughboth act as positive
energeticcentersand as such have exertedother common effects.The same
is true for the carboxyland thiol groups.
Still more specificappear the effectsresultingfrom the interventionof
the energetic factors present in the nonpolar group, such as the double
bonds,and the energeticformationsthey realizesuch as conjugated,or two
double bonds separatedby a methyleniccarbon.
The various mechanismsinvolved have explainedfurther the different
kinds of biologicaleftectswhich result.The action of the lipid by meansof

168 /

nEsEARcH rN pHysropATHoLocy

the lipoidic effect will thus influencegeneral,nonspecificmanifestations,

suchas thoseconcerningthe permeabilityof membranes.only secondarily,
will thesechangesin membraneperrneabilityinfluencethe different metabolic processes
which the membranegoverns.
In the secondgroup of changes,relatedto the interventionof the polar
groups,the antagonisticeffectsinducedwere seento concern processesresulting from membranepermeability.It is only in a third change that a
more specificaction upon the differentmetabolicprocesseshas to be considered.These are concernedwith an interventionupon metabolitesor the
agentsgoverningthem. The characterof this last lipidic interventionis its
specificinfluenceexertedupon a definitemetabolicsystem.
we tried to interpretthe influenceexerredby a lipid or lipoid according
to the above systematization.
The recent developmentof the biochemical
methodsof investigationhas put into limelight many biochemicalprocesses
by consideringthem as isolatedmetabolicentities.Most of them were seen
to result from the interventionof enzymesupon more or less specificsubstrata. One of the principal objectivesof the actual pharmacodynamic
studiesis to correlateas directly as possible,biological eftectsof diftcrent
agentsto specific metabolic processes,most of them correspondingto a
change in an enzymatic process.This approach, while very interesting,
would not take into considerationthe important role played by the nonspecificactivity of lipids and lipoids. These nonspccificinfluencesthrough
changesin the lipidic systeminducedifferentchangesin different metabolic
processes.A nonspecificchangein membranepenneability will affect many
enzymaticprocesses.It explainsthe existenceof similar influencesexerted
upon theseprocessescommon to agentswhich have nothing more in common than their lipoidic propertiesand the presenceof a positiveor a negative characterof their polar group.It is this characterwhich binds an eftect
to the nonspecificintervention.This so-systematized
analysishas thus permitted to separatethe biologicalactivity of the lipids and lipoids, the more
specificfrom the lesserinfluences,and correlateeach one to a proper or
commoncharacterof the agent.This view has amply simplifiedthe study of
the pharmacodynamic
interventionof thesesubstances.
OTHER CONSTITUENTS
In addition to the chemical elements and lipids, other constituents have
been studiedfrom the dualisticpoint of view. Although the other constituents havc receivedless emphasis.interestinginformationhas been obtained.

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LrPrDs AND LlPorDs

169

Amino Acids
Amino acids have been separatedinto groups basedupon their effects
at different levels. The first group includes the simple amino acids. In
these members the portions of the molecule which are added to [he amphoteric amino acid group, are usually electrically neutral. The amino acids
polymerizedthrough the anaphotericgroup serve as building materialsfor
the bigger protein molecules. They have appeared to be inert without
effects upon the different levels. Beyond these simple amino acids, are
two groups, energetically active, which have a second energetic center
with a negative or positive character in their molecules. while the amino
acid group servesto make thesesubstancespars of higher proteins through
the same bonds of amino acid groups as the simple members, it is the
other energeticcenter, with acid or alkaline character, which confers upon
these amino acids a positive or negativecharacter.
We studiedeffects,at difterent levels,of arginine,lysine and histidine,
which are members of the group with alkaline centers; of glutamic and
aspartic acids which have acid centers; and of methionine with a thiolic
center. Like for the lipids, the last two groups have shown similar propertias,but oppositeto thoseof the memberswith dkaline centers.The nature of their intervention appearedevident through the interesting opposite
effectsexertedupon microbes.Cultures of B. subtilis in broth containing
membersof one or the other of the antagonisticgroups show characteristic
changes.Unlike controls in which the long chains of microbes remain isolated, the microbes were seen to be kept together in media with atkaline
amino acids, forming a consistent gelatinous mass separated from the
medium. In broth with acidic or thiolic amino acids, the microbes remained separatedor formed very small aggregates.This appearedinteresting when we considered the positive character present in a.lkaline amino
acids, as related to the heterotropic,constructivetrend, while the negative,
as in the acid and thiolic members,is related to the opposite trend, We saw
further the sameantagonism betweenthe influenceexerted by histonesand
nucleic acids, the first paralleling the alkaline amino acid groups and the
second the opposite group. The more manifest effect of the ribonucleic
acids could be seen to take place at higher levels of the organization and
possibly explains the more direct action upon the genes.
We investigated the eftect of the two groups of amino acids at the
tissuelevel upon pain. Arginine,lysineand histidinedisplayedan analgesic
effect upon alkaline pain, while glutamic acid and methionine had this
effect upon acid pain. The effect could be related more to the basic tend-

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170

xESEARcH rN pHysropATHoLocy

ency of thesesubstancesto act through metabolicchanges,than to a direct

influence upon the acid-basesystemicbalance. The first group acts as
heterotropicagentsand the secondas homotropic, as mentionedabove.
Abnormal Amino Acids
Our researchled us to severaltentativesto defineabnormal amino acids
and the proteins they form. one concernedtheir rotatory capacity. The
naturallyoccurringamino acidsare all levorotatory.However,the organism
constantlyhas enzymesableto attackdextrorotatoryamino acid membersas
if it would have to be prepared to encounter and destroy them. Such
dextrorotatorymemberscan be conceivedto appearon a statisticalbasisas
the result of the resonanceprocessseen to occur in all the synthesisin
nature. The interventionof specificenzymesagainstthem would have the
aim to control their existenceand especiallyto prevent their interventionin
further evolution. In a work hypothesisconcerningthe cancerousprocess,
we consideredthat their persistenceand especiallytheir participation in
forming hierarchic entities would correspondto the specific abnormality
characterizingthis condition.
In another work hypothesiswhich concernsalso cancerousprocesses
and which we will discusslater, abnorma.lproteinsare thought to appearas
a resultof the bond of a carbamicradical (295) to the amino acid group.
The resultingcyclic formation having the characteristicNCNC group in
it, would correspondto abnormal amino acids which would representthe
primary characteristicformation of the cancerouscondition. (Seechapter
II,Note I)
Carbohydrates-Glucose acts as an anti-fatty acid agent, possibly becauseof the glyceryl compoundsresultingfrom its metabolism.We have
studiedit in opposition to the respectiveacids-gluconic, glucuronic and
saccharic.Glucosehas an analgesiceffect,although limited, upon pain of
an alkaline pattern, and an oppositeeffect upon pain of an acid pattern.
The acid group has an oppositeeffect upon pain. This could be correlated with the changestoward acidosisseen in the local pH of the lesions.
A manifest changetoward acidosiswas seenunder the influence of glucose
in the second day wound crust pH. We have noted previously the role
playedby glucuronic acid as an agentwith anti-positive-lipidactivity. We
believethat it is largely through this mechanismthat it favorably influences
acid pain, having an indirect action similar to that of fatty acids.

CHAPTER

DEFENSE
rTr\
REcocNtrtoN THAT multiple factors are responsiblefor abnormal
I
"t
conditions,
and that these factors can be systematizedaccording to the
concepts presented above, throws new light on a specific aspect of the
relationship between the different entities and the environment when this
tends to alter their characteristicorganization.This responseis concretized
as the defenseagainstthe noxious.The analysisof this defensehas been
facilitatedby emphasizingthe relativeindependence
of the endtiesforming
the complex hierarchic organism,the dualistic patterns of response,and
the critical role of the lipids as well as of proteins. Abnormal processesin
an organism's defense system may be better understood when they are
compared to those correspondingto normal physiologicalprocesses.For
this reason,we start with this last aspect.
The direct intervention of a noxious agent upon a biological entity can
be characterizedby its tendencyto induce heterogenization,
through an alterationof the entity'sconstituents
or the relationship
betweenthem.This, in
turn, affectsone or more of the constantsthat characterizn,the entity. The
ensuing defense response is directed ultimately at restoring the altered
constantsto their normal values.
Involved in a first stage of defenseare those very factors which normally maintain the constants,the factors which induce the oscillatory
dynamic balance. As a first response, they become exaggerated. Such
exaggeration,which takes place successively
for the opposite phases,resolves many slight noxious interventionswithout clinical manifestations.
Through a damping movement, the exaggeratedoscillationssoon return
to normal. If the normal constantsare reestablished,
the phenomenoncan
be consideredto be a physiologicalresponse.
But if the alterationsinducedby the noxiousagentpersist,an abnormal

L7r

172 /

RESEARcHIN pHystopATHoLocy

condition results. Indeed, in this casethe exaggerationof oscillatory movement can be so great that an abnormaliry may result even from this exaggeratedattempt of the entity to reestablishnormalcy. In fact, offbalances
are induced by just such changeswhich often represent,by themselvesas
will be secnlater, one of the major immediatefactors inducing the abnormalcy. As long as an abnormal condition is not resolved,the biological
entity will try to utilize new meansin order to reestablishthe normal balance. If the constantsdisturbed by the noxious intervention are fundamental, or if the changesresultingfrom the defensemechanismitself are
too great, death of the entity will result.
As expected,responseswill differ accordingto the level to which an
affectedentity belongs.However, despitethe many differencesrelated to
levels, a common and relatively simple pattern can be recognizedwhen
manifestationsoccurring at different levels as tlre result of the noxious
intervention are compared and referred to the basic pathological concepts
already noted.
Most of the information about this simple pattern was originally obtained by srudyingresponsesat the systemiclevel. Blood was parricularly
suitablebecauseof its availability,its multiple constantsand manifestcapacity to conserve them, and particularly because of the facility with
which noxious agentscould be inducedto act upon it.
The interventionof a noxious agent able to change the energeticbalance of blood sets in motion immediatelya group of successiveprocesses
which may or may not be clinically apparent,dependingupon their intensity.They have been describedas hemoclasiaby widal and hemo-shock
by many authon. Although widely investiga,ted,
the mechanismdid not
appear clcar. From our studies, we have arrived at certain conclusions
which we will briefly prasenthere.
Diphasic Phenomenon
As a noxiousfactor, we usedan intravenousinjectionof killed microbes
or of a colloidal suspensionof a metal. within a few minutes, a group of
changesoccurred.They were revealedthrough a seriesof analysesmade at
very short intervals.(Note /) The changeswere found to affect most of
the blood constituents,The most characteristicchangein our opinion is a
leucopeniawhich especiallyaffects the granulocytes.With it, there is a
lowering of serum antitryptic power; a decreaseof serum albumin; appearance of degradatedproteins,esteraseand amylase;increaseof free fatty
acids; and a lowering of coagulabilitywith reduced clot retraction.Clinically, these changesare accompaniedby hypothermia and hypotension.

DEFENSE

173

Together they representwhat we will call the "negative phase" of the immediate response.
This group of changesrepresents,in fact, only the first part of a diphasic phenomenon.The negativephase is usually followed by a second
and opposite one which we call the "positive phase" of this immcdiate
response.It results from the tendencyof the body to correct, and even
over{orrect, the changesoccurring in the fint phase. After hypothermia
and hypotension,hyperthermiaand slight hypertensionfollow. At the same
time, thc number of granulocytesincreases,as does the antitryptic power
of serum and its albumin contcnt. The serum appears richcr in frec
stcrols. Blood coagulabilityand clot retractionalso increase.After moving
rapidly to a peak, all thesevaluesreturn slowly to normal. The existence

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l'{inutes
Frc;, 75. Diphusic responsein the delense.The intravenousinjections to a normal individual of a foreign material such as of a suspensionof killed microbes or of a colloidal
metal induces a typical responsewhich correspondsto the hemoshock. A diphasic
curve seen in most of the analysescharacterizesthe occurring variations. The curve
presentedcorrespondsto the total number of the blood leucocytes.A parallel diphasic
curve is seen for other blood analysessuch as clot retraction, albumin content of the
s e r u m , a n d a n t i t r y p t i c v a l u e s o f t h e s e r u m . S i m i l a r d i p h a s i cc u r v e s , b u t o p p o s i t e i n
sens,are seen for blood coagulation time, amount of arnylase and esterasein the
\ e r u m . a m o u n t o f K , i n t h e s e r u m , a n d f o r t h e a m o u n t s o f p r o t e o s e sa n d p e p t o n e s

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174 /

xEsEARcHrN pHysropATHoLocy

of two phases can be recognized in all the changes occurring in hemos h o c k(. F i g . 7 5 )
In trying to correlatethe multiple changestaking place, it is the lysis
of leucocytes,especiallygranulocytes,which can be considcredo[ primary
importancein the developmentof hemo-shock.This is evident from the
relationship between granulocytopeniaand the intensity of the diphasic
phenomenon.The administrationof morphine or other opium derivatives
to an individual,prior to the applicationof the noxious factor, will reduce
or suppressthe granulocytopeniatogether with all the manifestations.
(Note 2/ Intensive physical exerciseconcomitantwith the application of
the noxious factor will increasethe granulocytopeniaparallel with all the
manifestationsof hemo-shock.(Note 3)
According to our hypothesis,lysis leads to liberation of proteolytic
enzymeswhich may be present as such or may be present in precursor
form in the leucocytes.And it is the interventionof theseenzymeswhich
reducesthe antitryptic power of the blood and, by digestingblood constituents,Iowers the amount of albumin presentin the serum, and induces
a parallel increaseof productsof protcin hydrolysis.The increasein anrylase as well as in esterasepresentin blood is related to the other hydrolytic enzymesliberated in this phase,and is also probably correlatedwith
leucolysis.The esterascacts hydrolytically upon the neutral fats present
and this would explain, at least in part, the liberation of free fatty acids
seenin this phase.In the changescorrespondingto the first phase,digestive
effect of theseenzymesupon the blood constituentscan be recognizedas
being one of the most important interveningfactors.
We confirmedthe correlationbetwcenthesechangesand leucolysisnot
only through thcir parallel variations, as mentioned above, but also
through in-vitro experiments.Lysis of leucocytesresultedin liberation of
hydrolytic enzymcs.An exudatc rich in granulocyteswas obtained by
injectingsterilebroth, or an aleuronsuspension,
into the pleura of rabbits.
To this exudate,removed through pleural puncture, a small amount of a
colloidalsilver-proteinpreparation(Collargol0.lvo) was added and the
preparationmaintainedat 38oC. This was seento induce the appearance
of vacuolesin the leucocytes,following the phagocytosisof silver grains.
The vacuoleswere observedto grow rapidly to huge dimensionsfollowed
(FiS. 76)
by burstingof the leucocytes.
Analysisof the pleuralfluid treatedin this mannerhas shown the same
changeas those seenin the first phaseof hemo-shock:loweringof antitryptic power with a decreasein albumin content, increasein products
formed by partial digestionof proteins,appearanceof amylaseand ester-

DEFENSE

l7S

ase, and an increaseof free fatty acids. There were also the same nuclear
"shadows" as encounteredin large amounts in the circulating blood at this
phase. The increaseof the potassiumcontent of serum seen in this phase,
and the increase found also in the supernatantpart following centrifugation of the exudate to which Collargol had been added, representsa further
confirmation of the role of leucolysisin this first phase.These data enabled
us to consider that the mechanismthrough which the blood tries to combat the intervention of a noxious agent corresponds,in the first phase,
primarily to a lysis of granulocytesfollowed by hydrolytic digestion.

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FIc. 76. Drawing of the changesinducedby a colloidul su.spension
ol silver proteinate
upon leucocyles.The leucocyteswerc obtained by injecting broth intrapleurally to
rabbits. Silver proteinate was added to thc suspensionof leucocytesand thc changes
observed in a microscope heated chamber maintained at 38'C. The phagocytosisof
the silver proteinate leads first to the appearanceof this substanceas intracellular
granules, followed by the formation of vacuoles.As these grow to a huge size the
cells burst. The nucleus remains as nuclear shadow.

The second phase, which would correspond to efforts to correct the

exaggeratedeftectsof the first digestivephase,involveslargely a mobilization of reservesof those blood constituentswhich were altered during the
first digestivephase.The spleenpours a part of its storedblood into the circulation. The richnessof spleen in reticuloendothelialcells explains the
liberation of sterolswhich is seenduring this secondphase.This is recogrluzr,dby the fact that, at this time, the spleenefferent blood is richer in
sterols than the afterent blood. Other constituentscome from intercellular
and lymphatic spaces.This mobilization,characterizingthe second phase
of hemoshock, appearsto be achievedin large part mechanically,through
a direct intervention of the vegetativenervous system inducing the contraction of the smooth muscularfibers, as seen during chill, which marks
the beginningof this secondphase.Fever which follows, is in part, due to
the sterols liberated largely by the reticuloendothelialsystem.

176 /

nESEARcHrN pHysropATHoLocy

If this hemeshock, in spite of its frequently violent clinical manilestations, resolves the effects of the noxious intervention upon the blood, it
can be considered to be, to a certain degree, a physiological response.It
amounts to an exaggoration of the oscillatory mechanism through which
the characteristicconstantsof the blood are maintained. By employing the
hydrolytic enzymesstored h the leucocytes,the blood tries to resolve the
influence exercised by the noxious factor, digesting and thus breaking
down either the factor itself or the results of its direct intervention. Acting
upon blood constituents,the noxious agent often inducesthe appearanceof
micelles bigger than those normally circulating. The fatty acids liberated
by hydrolytic enzymes would insure, in the first place, a higher boundary
permeability, thus permitting the passagethrough the capillaries of substancesotherwisebarred. At the sametime the fatty acids bind the antigen
in a lipidic complex.
In the second phase, the organism tries to repair damagescaused by
the exaggerateddigestive processor by the intervention of fatty acids. If
the organism is able to resolve through a successfuldiphasic reaction the
changesinduced by the noxious agent,it returns to normal.
Prolonged Hemoshock
Inability of the organism to resolvethe noxious interventionthrough
the mechanisminvolved in the diphasic phenomenonleads to abnormal
prolongationof one phaseor another.If it is unableto destroythe noxious
factor in the first phase or to mobilize the repair process in the second
and thus correct the damage induced by the first phase, the organism remains in a prolonged first phase of hemo-shock.If the second phase is
quantitativelyor especiallyqualitativelyinadequate,the organism remains
in a prolongedsecondphase,continuingto try to resolvethe offbalanceby
a quantitativelygreater mobilization of the otherwisequalitatively inadequate weaponswhich are at its disposal.It is the predominantintervention
of the lipids which characterizestheseextendedphases.We wish to note
again that the fatty acids intervenein the prolongedfirst phasewhile antifatty acid agents,especiallysterols,are active in the second.
The adrenalsplay a particularly important role in the immediate and
prolonged defense process.In the first phase, the increasedamount of
fatty acids with four or morc double bonds found in blood and in the
organismin generalappearsto conrefrom the adrenals,which are usually
extremelyrich in thesesubstances.
In an exaggeratedlyintensiveprolonged
first phase,we found small reddish adrenalspracticallydevoid of fatty
acids.This occurrence,togetherwith the concurrentincreasein fatty acids

DEFENSE

177

in the blood, relates thesechangesin fatty acid content of the blood largely
to a Iiberation of the adrenal fatty acids into the circulation. Another important factor for the prolonged first phase appears in the intervention of
lymphocytes able to induce a lysis of compounds of very high fatty acids
such as presenteven in waxes.(Note 4) A lymphopeniacorrespondsto the
prolonged first phase. In the elevation of the amount of anti-fatty acid
agents in blood, characteristic of the prolonged second phase of the diphasic phenomenon, the adrenals seem to intervene again providing a
portion of the increasedcirculating sterols. The exaggeratedmanufacture
of sterols can be attributed to the reticuloendothelial system in general.
cranulocytosis and lymphocytosis occur in this prolonged second phase.
The interventionof sterols, which are relatively simple steroids,can explain the clinical manifestationssuch as fever, which charactcrizethe prolonged second phase, since fever can be induced by the administration of
Iarge amounts of sterols.
we can separate, from the point of view of its manifestations, the
immediatediphasichemo-shockphenomenonwith a short evolution, from
the more prolonged forms. while the former, if not too exaggerated,would
correspondto a physiologicalphenomenon,t}tc latter is always abnormal.
In the former, the principal interventionis that of hydrolytic enzymes;in
the latter, lipids play the most predominant role. pathogenically,each
phase of the diphasic phenomenon,if unable by itself to resolvethe immediateproblem, will be followed by a correspondinglipidic predominance.
The result may be either one of the two phascs,with fatty acids or sterols
predominant.We call this entire response"the antiheterogeneous
reaction"
of the defense,separatingits diphasic manifesrationsinto immediate hydrolytic and prolonged lipidic stages.
A rui heterogeneous Reaction
Although, in the prolongedlipidic stage,a certain specificiryfor particular antigens can be recognized,the antiheterogeneousresponse in
generalrepresentsrather a nonspecificeffort of the organismto resolvethe
problemscausedby the presencof any heterogeneous
factors as such.
Before going further, we want to cmphasizesomc important characteristics of this antiheterogeneousresponse related to organization. The
catabolic processespresent in the first phasc appear to result in part from
the direct hydrolytic process and in part from the biological intervention
of the products of hydrolysis, especiallyfatty acids. The hydrolytic enzymatic processis homotropic in nature by delinition, as it breaks down
difterent constituents,liberatinggroupswith anionic and cationiccharacter.

178

n EsEARcH rN pHysropATHoLocy

Enhancing the catabolic character of the first phase processesare the

anionic groups which appear to have a predominant role. The second
phase, a reparative one, is anabolic and therefore, heterotropic in character.
The study of the antiheterogeneous
responseemphasizesanother fundamentalcharacteristicof the processes.
A basic differenceexistsbetween
the direct effectsinduced by the interveningagentand thoseresultingfrom
the defenseprocessesthemselves.A direct effect of a noxious intervention
correspondsto heterogenizationof the constituents.Some changes will
appear through this heterogenizationitself, others through the defense
processes
which representthe responseof the organismto the heterogenization. While the first correspondsto a direct action, the last is catalogued
as antigenic,its manifestationsbeing grouped as defenseprocesses.The
same substancecan have a direct action and an antigenic one revealed
principally through the manifestationswhich it induces. A direct action
can be noted instantaneouslyif the changesinduced are sufficientlyintensive.The antigen effectsalways require a certain time before manifestationsappear. This time can range from a few minutes for the first
enzymaticresponseto hours or days for the prolongedresponse.
An important feature of the prolongedstageis that it persistsas long
as the noxious agent is present.This is evident in cases in which the
noxious agent can be suppressedthrough external intervention.For example, with suppressionof microbial activity by antibiotics, the corresponding clinical condition disappears.Of more interest is the effect of
antimicrobialand antitoxic immune sera. Administration of a specific
scrum, if it can neutralizethe noxious agent,producesa curative effect at
this stage.In a short time, symptomsdisappearand the organism reverts
to normal. Although nonspecific,the prolongedantiheterogeneous
reaction
shows such a straight correlation with the presenceof the antigen as to
makeus designatethis stageas primaryor toxic. In this stage,the organism
reactswith clinical manifestationsof diseaseif the antigen is capable of
inducingsufficientnoxious changes;if not, there are no clinical manifestations. Persistencein the organismof an antigenbeyond the rapid diphasic
phenomenonindicates,in general,an incapacityof the organismto achieve
its disposalsuccessfully.The need for more complex meansto combat the
antigenbecomesimperative.
with or withoutclinicalmanifestations-thatis, evenwithout a primary
toxic stageof the disease-as long as the antigcnhas not been fully neutralized, the organism will still try to resolve its intervention and return
to normal, resortingto other means.It will produce antibodieswith a cer-

DEFENSE

179

tain degreeof specificity toward the antigen. Two kinds of antibodies will
be manufactured and will differ in their fundamental characteristics,the
time of their appearance,and their role in the defenseprocesses.
CoagulantAntibodies
The first group of antibodies have a characteristicproperty. Together
with the antigen, on which they fix with a degree of specificity, they form
highly energeticcomplexes.This is manifest in a marked tendency to bind
together such complexesas well as constituentsof the blood and form huge
aggregates.When such antibodies are produced for, and act against, a
specific microbe, agglutination results. Conglutination, prccipitation and
flocculation occur when similar antibodiesact against other antigens.Due
to their tendency to establishantigen antibody complexesresulting in huge
formations, these antibodiesare generallygrouped as coagulant antibodies.
Although the coagulation characteristic is not demonstrable in vitro for
all antibodies in this group, we use the term "coagulant antibodies" for
didactic purposes.
The huge complex formation resultingfrom the binding of coagulant
antibodieswith an antigencan appear as a precipitate,agglutinatedmicrobe
or conglutinated red cells. Once established,this formation representsa
new heterogeneousentity of much larger dimensions than the antigen
alone. As srrch,it becomesby itself a new noxious agent for the organism
which consequentlyreacts against it. The organism utilizes the same processesagainst this noxious antigen-coagulantantibody complex as it uses
for any heterogeneousagent, with the same immediate diphasic or prolonged mechanism.
Teleologically,the formation of coagulantantibodiescan be interpreted
as an attempt of the organism to defend itself anew against the an,tigen.
The antigen, this time fixed through these antibodies in a new and more
noxious formation, will once again incito the nonspecific defense mechanism. First there will be the antiheterogeneousresponsewith its diphasic
phenomenonand, if once more this is not effective,a prolonged new lipidic
intervention will follow. If the quantity of heterogeneousformations is
gteat, the first phase of the diphasic phenomenoncan be so severe as to
cause death in a few minutes. If less severe,this first phase is followed by
the second,with chills and high temperature,As in all the antiheterogeneous reactions, the organism tries to combat the presenceof the noxious
factor-in this case, the flocculate produced by the antigen-antibodybond
-attempting to digest it through hydrolytic enzymes or to neutralize it
through constituents brought in during the second phase of the diphasic

180

RESEARcH rN pHysropATHoLoGy

phenomenon. If it fails, the abnormal prolonged form of this response

follows with characteristiclipidic liberation.
In terms of biological meaning, the formation of coagulant antibodies
representsa new chance for the organism to resume tho fight against antigens by using the same fundamental means, the antiheterogeneousreaction. However, since the new agent, the antigen-antibody complex, is
much more noxious than the antigen alone, the intensity of the response
will be much stronger and the chancesof disposing of the antigen will be
greater.
AIIergic Reaction
The genericterm "allergic" is reservedfor the entire group of processes
in which a coagulant antibody takes part. The reaction now against the
antigen-antibody complex is a typical antiheterogeneousresponse. A
fundamental difference exists, however, between the antiheterogeneous
reaction in the primary toxic stageand the reaction which occurs when the
heterogeneousfactor is a complex antigen-coagulantantibody usually with
more noxious character than the antigen alone. It is the nature of the antigen-antibody,the result of the bond of the coagulantand the antigen,which
gives it the allergic character. This faot explains why, although we can
possessspecificimmune sorum able to neutralizean antigen alone, this will
not influencethe allergic response.Already bound to a coagulant antibody,
the antigencannot be bound again and consequentlyneutralizrd by another
antibody. The specificneutralizing serum will have no effect upon the antigen-coagulant-antibodycomplex already formed and consequently will
have no effect upon the procssesinduced by this complex. The immune
serum does not influence the allergic manifestations which represent tbe
response to antigen-coagulant-antibodycomplexes. This would theoretically explain the favorable effects of a specific serum upon a condition
which is in the toxic primary stage,where the antigen intervenesas such,
and the lack of such favorable eftectsin the allergic stage,where the antigen is representingon.ly a part of a complex new noxious formation.
This mechanism would also explain why t}le same immune serum, although without curative effect upon the allergic stage of a condition, will
have preventivo activity. Before the onset of the allergic stage-that is,
before the coagulant antibodieshave appeared-the active immune serum
will bind and neutralizethe antigensstill free in the organism.Under these
conditions, when coagulant antibodies appear, the antigens are no longer
availableto be botrnd by them to form the noxious antigen+oagulant-antibody complexes.Without curative action, immune serum is effective as a

DEPENSE

l8l

preventive only when administeredprior to appearanceof coagulant antibodies.

An important factor for the allergic responseis the time of liberation
of coagulant antibodies. Generally, a period of 6 to 8 days is required.
Under specialcircumstances,as in casesin which the organism has manufactured the same antibodies in the past, the time necessaryfor their appe:rranceis reduced even to 4 days. In other cases,for certain antigensor
for older subjects,the time may be as long as | 4 days or even longer. For
certain antigens,or under specialcircumstances,the body appearsunable
to make coagulantantibodiesat all. In that case,no allergic manifestations
appear.
It must be emphasizedthat antibodieswill be liberatedeven if the antigen is no longer present.The presenceof antibodiesalone does not give
rise to any reaction and their appearancewill pass without any manifestations. However, they can persistunder certaincircumstancesfor months or
years and become a potential source of abnormality. At any time if the
same antigen becomes present in the body, the coagulant antibody will
form the allergic bond with it. The body will then react against the newly
formed complex with an antiheterogeneous
response.If this occurs in the
blood or central nervous system,it can appear as an immediate violent reaction which corresponds to anaphylactic shock. It is the intensive first
phaseof the diphasicphenomenonwhich kills in anaphylacticshock. such
shock can be easily producedin animalsas passiveanaphylaxyby making
coagulantantibodiesand antigensavailableconcomitantlyin the blood.
Li pi do- p rot ei c A rui bodies
Analysisof allergicantibodiesindicatesthat they have two constituents,
lipids and proteins.Electrophoreticanalysesreveal that they are displaced
mostly as beta globulins. Experiments show that such lipidoproteic antibodies lose their activity if broken into their constituents,neither the lipid
fraction nor the protein alone being able to bind the antigen.
The study of the lipido-proteic antibodies,brought us back to consider
the role of the lipids in the immediateor prolonged first phase.Moet, if not
all the natural antigenshave lipids and lipoproteinsin their structure.As
we have seenabove,someof the fatty acidsinduce defenseresponses.
The
administration of fatty acids is followed by a leucopenia-especially a
lymphopenia; administration of sterols,by a hyperleucocytosis.Some fatty
acids,suchas thoseobtainedfrom the tuberclebacilli,inducecharacteristic
lesions such as giant cells. There are both naturally present lipoproteins
and those resulting from the bond of the body's freed fatty acids to rhe

182 i

nEsEARcH lN

pHystopATHoLocy

antigen,which seemto act as specificantigens,inducing the appearanceof

coagulant,allergic antibodies.Experiments,which we will discussbelow,
have shown that, while the specificityof the antibodiesis highly related to
the protein fraction, the allergic or immune characterof the resulting responseis due to the fact that lipido-proteinsare involved in these processes.The injection of the procluctresultingfrom the action in vitro of the
acid lipidic constituentsof an organism upon proteins of another species
acts as an antigen inducing the early appearanceof coagulantantibodies.
The repeatedinjectionsof the product obtainedthrough the action in vitro
of foreign lipoacid fractionsof variousorigins upon different body proteins
also inducesallergic response.Just as we have connectedthe appearance
of the first diphasicphenomenonto hydrolytic enzymesand the prolonged
form of the antiheterogeneous
defensemechanismto the intervention of
lipids, we relate the allergicbody defenseto the interventionof lipidoproteic formations.The allergicstageof defensethus could be considered
to be a lipido-proteicdefenseresponseagainstlipido-proteic antigens.
N eutralizing A ntibodies
The unsuccessfulfight of the organismagainstan antigen through the
diphasic,lipidic or allergic responsesoften can evolve further, making use
of a more effective measurewhich correspondsto another kind of antibody, different from the coagulanttype. The characteristicof this second
type is that it forms, specificallywith the antigenagainstwhich it is manufactured,a new kind of bond, an antigen-antibody-complex,
this time entirely nonnoxious to the organism. This complex is energetically so
balancedas to corrcspondto the constantsof respectivelevelsof the body
whereit occurs.Through this new bond, the antigenis biologicallyneutralized in the sensethat the resulting antigen-antibody-complex
is entirely
harmless.
This type of neutralizingor immune antibody usually appearson or
after thc l5th day followingthe momentwhen the organismhas startedto
organizeits defenseagainstthe antigen.It can occur whether the antigen
is still prcsent or not and whether it is free or bound to coagulant antibodies.It reprcsentsthe best meansthrough which the organism opposes
the influenceexerciscdby an antigen.The appearanceof the neutralizing
antibodiescorrespondsto the last stage,the immune one, in the defense
mcchanism.If the antigenhas produceda clinical condition,the neutralizationof the antigenby the new antibodyresultsin the slow disappearance
of morbid manifestationsand a progressivereturn to normal. With or without prior clinical manifcstations,
the presenceof the neutralizingantibody

DEFENSE

183

in the organism provides a potential weapon to prevent the same antigen

from again causing trouble. This has led us to identify this part of the
defensereaction as the "immune stage" of the defensemechanism.
The action of these neutralizing antibodieshas been demonstratedbeyond doubt through passive immunity. Their administration confers protection againstthe antigen. This action is limited to the antigen so long as
it is not bound by another antibody. Clinicatly, neutralizing antibodies
have a curative value if the antigen is present, inducing the primary-toxic
response.They have, also a preventive effect upon the allergic form of the
disease if they are administered before the appearanceof the coagulant
antibodies.
lltu6alizing antibodies are globulinic in nature. They are displaced in
electrophoretic analyses as gamma globulins. Isolated zrs pure globulins,
they do not lose their activity. This would differentiate them from the
coagulant antibodies which, as previously noted, are lipideproteinic in
nature.
The defenseresourcesof organismsagainst antigensthus can be didactically separated into four fundamentally distinct groups: enzymatic
hydrolytic, lipidic, lipoproteinicand proteinic.They correspondto distinct
stagesfrom the point of view of reactionsinduced and biological meaning.
The first representsa primary, direct, immediateresponsecharacterizedby
a rapid nonspecific digestive process and followed by the exaggerated
mobilization of repair processes.If the immediate responseis inadequate,a
second stageas a prolonged lipidic defensefollows. Although it has a certain degreeof specificity,this last responseis still directed againsta heterogeneousconstitution of the agent as such. If unsuccessfulin inactivating
the agent, all these responsesare followed by another defense stage in
which action is taken against the antigen through more specific coagulant
lipido-proteinic antibodies and through the antiheterogeneousreaction to
the resulting complex. with the last stage, which is characterizedby the
intervention of proteinic neutralizingprotectiveimmune antibodies,the fight
against the antigen is usually concluded successfully.Table XI, below,
summarizes this syst,ematizationof the defenseresponse.
Defense and Hierarchic Levels
The above changes represent, in a schematic manner, what happens
when a noxious agent acts directly or indirectly upon blood. The same
basic patterns of defense,the substancesused and the processesinvolved
can be found at various hierarchic levels.It is easy to seethat, with such a

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DEFENSE

185

Jomplex mechanism occurring in different individuals and againsga great

variety of antigensr gest variations in manifestations will be evidenced.
When an antigen enters the organism and is not fully neutralized by
the intervention of immune antibodies.the mechanismof defenseis set in
motion. This can be limited to a group of entities, to one level, or can
aftect more levels and entities. According to the nature of the antigen and
the capacity of the di.fferententities to respond, the different procss will
proceed all the way to the stage of protective immunity or it can stop at
any stage.These factors, nature of the antigen, levels and entities involved
and degreeof responsefor each of them, determine the pathogeniccharacteristics of the resulting condition. The manifestationsfor a stage and a
grouP of entities can be so exclusiveas to produce a characteristicclinical
disease.The ability to respond through only a part of the defensemechanism dependson the nature of the antigen and on the conditions existing in
the different entities affected.The clinical manifqstationsfurnish important
information concerningthe defenseprocesseswhich are occurring.
C Iinical Manil estations
The first stageof the defensereaction,if highly intensive,can be manifested clinically as superacuteshock. This occurs minutes after the
intervention of the noxious factor. If the second phase of the diphasic
phenomenonis intensive,a chill with high temperaturewill appear.A state
of shock occursif fatty acids remain predominant,as in the prolongedfirst
phase. A feverish condition correspondsto a predominant anti-fatty acid
intervention in the prolongedsecondphase.while the first phase of the
condition appearsimmediately after the intervention of the antigen,the apparance and persistenceof the second phase, which corresponds to the
prolongedlipidic, dependsupon the nature and especiallythe amount of
antigen presentin the organism.Incubation time may vary from minutes to
severaldays. This relativelyshort and variable incubation time represents
an important characteristicwhich enablesus to recognizethis stage.Another characteristic of this stage is the disappearanceof symptoms after
administrationof neutralizingimmune antibodiesspecificfor the antigen.
If the antigenby itself has no toxic effectsupon the organism,its presence will not induce important manifestations.The direct response,enzymatic or prolonged lipidic, will be so limited as to have either minimal
clinical manifestationsor none at all.
we have seen that a.llergiccoagulantantibodiesusually appear after
the 6th day following penetrationof the antigeninto the organism.If the
antigen is still presnt, the appearanceof the antibodieswill induce an

186

REsEARcH rN pHysropATHoLocy

allergic condition. In caseswhere the antigen already has produced toxic

manifestations,the appearanceof the allergic stage will be marked either
by new symptoms or increasedintensity of existing ones. With a nontoxic
antigen, the presenceof which was not revealedpreviously by any clinical
manifestations,the appearanceof the allergic complex will coincide with
the appearanceof symptoms and, thus, with the appearanceof the clinical
condition. The period before the appearanceof symptoms correspondsto
the time before the appearanceof theseallergic antibodies.This incubation
time, as for all allergic manifestations,will be 6 or more days, which corresponds to the time necessaryfor the appearanceof the coagulant antibodies. An obligatory incubation time of 6 days or greater thus is an
indication that the processhas an allergic pathogenesis.
If the organism has previously manufactured similar coagulant antibodies againstthe same antigen, this incubation period could be shortened
to 5 or even 4 days. Against certain antigens,some organismsneed a longer
time to produce coagulant antibodies and the incubation time may run as
Iong as severalweeks.
Organisation and Delense
Going beyond the defensereaction in its general aspect, that is, independent of the place in the organizationwhere the characteristicprocesses
occur, we considered it in relation to hierarchic organization of entities.
conceptually, it can be acceptedthat each hierarchicentity, having a certain degreeof biological independence,
will have its own problems to resolve when it faces an antigen. Each will react for itself and. for this reason
alone, there will be differencesin the defensemechanismat different levels.
It can also be acceptedthat becauseof differencesin the means at their
dispoeal,the various entities will show individual peculiaritiesin their
resPonses.
Although some information is missing,our systematizationof the defense reaction according to the manifestations at various organizational
levels,stands.For each stage,manifestations
having common basic characteristicscan be identified at different levels such as cellular, tissular, organic, and systemic.At the cellular level, the first phase of the diphasic
phenomenoncorrespondsto a manifest increasein membrane permeability
and intracellularhydrolytic processes.
The changesseen in the first phase
of shock can be interpreted as resulting in part from such processes.we
will note here only that, for cells,the first phaseis characterizedby vacuolization of the cytoplasm and even of the nuclei similar to the vacuolization
seen in the leucocytesin the presenceof a colloidal metal. In superacute

DEFENSE

187

shock, which correspondsto the first part of the diphasicphenomenon,we

observed such vacuoles in central nervous system, liver and pulmonary
alveolarcells.(SeeShock,Chapter9) The sameprocessat the tissularlevel
causeslytic changesand, if this lysis acts upon vessels,producespetechiae.
For the systemic level, the first phase of the diphasic phenomenon is
marked by the changesoccurring in blood. The leucocytes,rich in hydrolytic enzymes,have a pronounced lytic tendency. The liberated enzymes
act upon the blood constituents and can impart to this stage the acute
dramatic aspectoften seen in clinical hemo-shock.
The prolonged lipidic phaseof the antiheterogeneous
reaction will have
different manifestationsaccording to the level affected. These differences
will correspond closely to the antagonisticinfluence exercised by the two
groups of lipids, sterols and fatty acids. we have discussedpreviously the
interventionof theselipids at differentlevels.We will mention briefly here
their role in the difterent phasesof the defensemechanrsm.
we have seen that, in general, the changesat the nuclear level correspond to prolonged youth if they are produced by the predominant intervention of sterols and to a rapid aglng with karyorrhexis and pyknosis
when produced by fatty acids. Similar manifestationscan be recognizedin
this phaseof the defensemechanismat the cellular level for the cytoplasm
and protoplasm formations, with aging signs and necrosisinduced by fatty
acids, and predominanceof youthful characteristicsinduced by sterols.For
the tissular level, intervention of fatty acids induces local alkalosis and
edema, while the sterols induce a local acidosisand fibroblasticreaction.
Lysis of vesselswith hemorrhagesoccurs in processesin which fatty acids
predominate.The predominanceof sterolsleads to a marked tendencyof
the vascularendotheliumto proliferateand this, in turn, can lead to vascular obliterationand ischemicinfarcts if the vesselsare terminal. At the organic level, the prolonged lipidic responseis more manifest than at lower
levels as the result of impaired specific function of the organ. Dualism in
clinical manifestationsis evident; oliguria or polyuria, diarrhea or constipation, insomnia or somnolenceare examplesof organic impairments seen
as clinical manifestationsof this stage of the defense mechanism. At the
systemic level, dual manifestationsare even more pronounced. Hypothermia, hypotension,cold perspiration,enophthalmia and dark-colored
blood are related to predominance of fatty acids, opposite manifestations
to predominance of sterols. Although these prolonged manifestations,part
lipidic response,can occur concomiof the nonspecificantiheterogeneous
tantly at the various levels of the organization, usually they affect one or

188 /

nESEARcHrN pHysropATHoLocy

severallevels. The manifestationscorrespondingto the one level or several

levelswill predominate.
The allergic stage shows the same clinical manifestationscommon to
the antiheterogeneousresponse with its enzymatic or prolonged lipidic
The fundamental difterencein the allergic stageis the obligatory
Processes.
incubation perid of 6 or more days. once the alleryic complex i5 lsarized,
the manifestations
are the sameas thoseproducedby highly active antigens
inducing a direct antiheterogeneousreaction.
The qualitative difterencesin the capacity of the hierarchic entities to
combat various noxious agentscan explain the diflerencesin the manifestations of allergic Processestaking place at the cellular or tissular levels as
comparedto those in the blood, which is at the systemiclevel. At any level,
the mobilization of lytic enzymesable to break down the alleryic antigenantibody complex can be so intense as to bring rapid death of the entity
or can be slow and prolonged. However, at the systemic level, as in the
circulating blood, the producs resulting from exaggeratedlysis are more
rapidly and completelydisposedof than in cytoplasmor interstitialfluids.
In the latter, they will be presentfor a long time and their noxious in-fluence
will persist.If the lytic products appear in moderate amounts in blood, the
organism may be able to disposeof them without any clinical manifestations.
For thesereasons,the presenceof antigensin the btood when coagulant
antibodies start to apPear will not induce serious manifestationsand will
even prevent them. As coagulant antibodiesappear gradually in the blood,
only small amounts of antigen-antibodycomplex will be produced at any
one time. Although highly noxious in large amounts, the complexescan be
resolved through lytic processesif formed gradua-lly,and consquently will
not provoke clinical manifestations.
At the tissular and cellular levels,a similar progressiveappearanceof
antigen-coagulant-antibodycomplex cannot be resolved in the same way.
The lytic reactions which break down this complex cannot occur with the
efficiencynoted in the blood. The complexesand tro products of lysis will
progressivelyaccumulate and the consequentmanifestationswill become
more and more intensive. It is for this reason that longJasting allergic
manifestationscolTespondto seriouslocal conditions.Even if the antigenantibody complexesare produced at a moderate rate, when antibodies appear and the antigen is present, they will induce little or no systemic
manifestations.On the contrary, seriousallergic manifestationswill arise
if the complexesare formed at the cellular,tissularor even organic levels.
Becausethe defenseProcesscsat theselevels cannot resolve them at the

DEPENSE

189

sams 1319as they appetr, as defenseprocessesin the blood can do, severe
local manifestations result. This may lead to necrosis and even rejection cif
alteredcells or tissues.These representthe very important difterenceswhich
exist between allergic processeswhich occur in the blood and those which
occur at the difterent levels following the appearanceof allergic antibodies
while the antigen is still present.
The fact that there will be no reaction when small amounts of the
allergic complex are progressivelyformed, as in casesin which antigensare
presentin the blood at the moment of appearanceof the allergic antibodi$,
is confirmed indirectly by the possibitity of prevonting severe systemic
manifestationsthrough skeptophylacticor desensitizationprocedures.The
introduction of very small amounts of antigen thus produces only small
amounts of complex at any one time, avoiding clinical manifestations.With
progressivedoses however, the antigens will fix circulating antibodies in
srrfficient proportion to prevent the formation of important amounts of the
same complexes after further administration of the antigen. The presence
of the secondphase of the diphasic phenomenon,with the exaggerationof
constituentsantagonisticto those presentin the first phase,will also act to
Prevent the occurrenceof an intensivefirst phasewhen the antigen appears
anew.
The situation changesentirely when antibodiesappear and the antigen
no longer is present. They can then accumulate in the blood in large
amounts. Thereafter, sudden appearanceof the antigen in sufficient quantity will form a large amount of the allergic complex and the subsequent
reaction can be so violent as to kill the subject.This occurs in anaphylactic
shock. When the antigenis limited to other levels,important local changes
can be induced.
The neutralizing immune antibodies, if manifestations already exist,
will prevent new ones from appearingand this will permit healing prooesses
to take place without further interference. The antibodies will prevent
manifsstationsat the respectivelevel if the antigen appearsagain.
Affinity ol Antigens
In the defense procsses,another factor intervenesto produce difierencesbetween responsesat difterent levels-the special affinity of antigens
for various cells, tissuesor organs.This affinity will determine not only the
levelbut also the individual entitieswhere manifestationswill occur. It has
to be emphasizedthat the independenceof the levelsor of groups of entities
in an organism goes so far as to allow the defenseprocessesto progressto
difierent stages.While defenseprocessesat the tissular level, for instance,

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190

REsEARcH rN pHysropATHoLocy

cannot go beyond the stage of prolonged lipidic response,those at the organic or systemic level can arrive at the allergic stage.we will see below
the importance of this unequal responseof the difterent levels.
The unequal capacity of different tissuesto manufacture allergic antibodies could be postulatedto explain the propensity for local allergic conditions. The ectodermic system appears especially inclined to allergic
responses,as seen for the skin. We tried to relate this to the natural richnessof these organs in sterols. This would explain the fact that the brain,
which is richest in sterols,seemsto show the earliest allergic manifestations,
which could be interpreted as resultingfrom early or more constant appearance of coagulantantibodies.
Besidesthese difrerencesin the responsesof various entities, an important factor intervenes in the induction of localized allergic manifestations. It corresponds to unequal affinity of the antigen itself for various
entities.This would localize the antigen in cells, tissuesor organs so that
when coagulatedantibodiesdo appear, the noxious allergic complex will
be formed locally in the sameentities.It seemsthat this localizationof the
antigen,such as upon nerves,kidney,lung, etc., is more important than the
capacity to produce antibodies in determining predilection of pathological
pr@essesfor specificcells, tissuesor organs.
one of the most interestingaspectsof the defensemechanismis the
relationshipbetween successivesteps.we could show, generally,that an
intensiveresponsein one step representsa favorable condition for appearanceof an intensiveresponsein the next step.It is a known fact that manufacture of immune antibodies is influenced by an inflammatory process.
This is the reason for the customaryinjection of tapioca, for instance,in
horsesduring their immunization for the production of therapeuticsera.
We could show that injectionsof lipids, lipid acids or insaponifiablefraction of placenta,or of organsof animalsof the same speciesfor instance,
manifestly hasten the appearanceof the next step in the defense against
the microbe.
It seemsclear that under the influenceof the lipids used,the agglutinins
appear in blood earlier and their amount increasesmore rapidly than in
the control animals.
Antigenic Factors
The interventionof different mechanismsin the defensehas led to the
suppositionthat each one would be induced by relatively specificfactors
presenteitherin the antigenitselfor appearingduring the defenseprocesses.

DEFENSE

l9l

An analysisof this aspectof the problcmof the defenschas broughtfurther

interesting
information.
The interventionof the first mechanism,that of hydrolytic enzymes
acting througha processsimilar to digestion,would have as aim to break
down the antigenitself as well as the groupsresultingfrom the bond between antigenand body constituents,
especiallyproteins.By analogywith
the processof digestion,the factorpresentin the body which would induce
this responsewould correspondto abnormallylow numberof micellcs.The
low numberof micellespresentis revealedby a cryoscopicindex near zero.
The digestivedefensemechanismwould thus intend to lower this cryoscopic index back to its normal valuesor even bclow them.
The secondmechanism,
that of the lipidic intervention,would havetwo
aims.One, to act againstfree lipidseitherprcsentin the antigenor resulting
from the hydrolytic action upon fats, and sccond,to bind hydrosoluble
constitucntsinto complexeswith a lower hydrosolubility,
and consequently
rvith lower diffusioncapacitythroughthe aqueousmedia of the organism.
This concernsthe antigenas well as the productsrcsultingfrom the lytic
intervention.The bond would take place throughthe activepolar part of
the lipid molecules.
The third mechanismis characterized
by the intervcntionof the allergic
antibodieswith thc aim of binding the antigenin higher complcxes.The
lipido-proteicantibodieswill opposea lipido-protcicfraction presentin
the antigenitselfor resultingfrom the bond bctweenlipids liberatedin the
second mechanismand proteinsof the antigenor of the body. The coagulant effcctwould resultfrom the bond throughthe polar and nonpolar
groups of the lipido-proteicantibodiesand those of the lipido-proteic
antigenicfactors.
For the fourth mechanism,characterized
by the protcctiveantibodies,
the antigenicfactor would be represented
by the proteicconstituents
of the
antigen,which leadsto an antireplication
in the specificantibodies.
It shouldbe notedthat in the complexdefensemechanismthe resultsof
the interventionof a defenseprocessrepresentantigenicfactorsfor the next
step. The presenceof productsof the enzymaticdigestionleadsto the interventionof the lipidic phase,largelyaimed to immobilizeand inactivate
them; the bond betweenlipids and antigenleadsto the appearanceof the
allergic lipido-proteicantibodies.Possibly,the occurring lipido-proteic
complexeswould intervene,facilitatingthe appearanceof the protective
antibodies.The idea that successive
antigenicfactors would induce the
of differentstepsin the defensemechanism,has led to a series
appearance
of studieswith the aim to obtain desiredreactionsthroueh the use of such

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192 /

n E s E A R c Hr N p H y s r o p A T H o L o c y

antigenicfactors. we will describehere very briefly severalsuch applications which were interestingalso becauseof the practicalresultsobtained.
Hydrolysis Products
we tried thus to utilize the productsresultingfrom the breakingdown
of body constituentsor of other materialsin order to induce through their
administration,the appearanccof the seconddefensemechanism.Applying
the dualistic concept, we separatedthus in the products of hydrolysis of
different materials,those with an acid characterfrom the group with basic
and alcoholic characters.Various materials were thus hydrolyzed using
KoH, NaoH or ammonia.The solublepart, separated,
was treatedwith an
acid and a precipitateobtained.After washingit, this precipitatewas red.issolvedby a.lkalizing
to a pH still below neutrality.This has represented
the
"acid fraction." Besidesacid lipids, this fraction containsalso acid protein
groups and even humic acids.
The part which remainedinsolubleafter treatmentwith KoH (separated from the soluble part) was treated with an acid. The part which
becamesolublewas then separated,
reprecipitated
by alkali, and partially
redissolvedby bringingthe pH, throughacidification,near 7. This representsthe "alkaline fraction." With differentdegreesof chemicalhydrolysis,
various fractions-more or less broken dsu/n-sps obtained for both the
acid and alkaline fractions.The degreeof this "digestion"has appeared
highlyimportant.The amountof the productsobtaineddecreases
for an insufficienthydrolysisas well as for a too highly pushedhydrolysis.
Accordingto the mechanisnrmentionedaboveit was expectedthat these
fractions,resultingfrom the breakingdown of body constituentsor of the
antigen and correspondingto the effect of the first enzymatic defense
mechanism,
would inducethe secondstepof the defensemechanism.This
would correspondin part to the interventionof the properdin systemand
of the lipidic defense.It has as characteristic
the fact that it would appear
only within a certaintime. The followingexperimentillustratesthis clearly.
The "acid fractions" of human blood, hydrolyzedby KoH was obtained
and then injectedintraperitoneallyto mice. At different intervalsfollowing
this injectionthe mice were inoculatedwith 3,000,000microbesof a fresh
c u l t u r eo f B a c .p r o t e u sI.n c o n t r o l st h i s i n o c u l a t i o n
w o u l d r e s u l ti n a I O O V o
lethalinfection.No protectionwas seento appearin the l6 hoursfollowing
the injectionof the "acid fraction."At thc l6th hour, J,2,,were protected.
This protectionincreasedwith time to be completeafter 22 hours, when all
the animals survived. This protection was still present after a few days.
The inoculationof thc nontreatedblood in the same proportion was

DEFENSE

le3

seenincapableof conferringthe samedegreeof defense,a fact which indicatesthe importanceof the breakingdown processin this ,.24 hours" defense.These resultsare similar to those obtainedby I. A. parfentjev with
malucidin,a product of hydrolysisof yeast.
Another applicationof the sameconcept was in the use of the lipidoproteiccomplexes.
In a group of researchstudies,we utirizedthe products resultingfrom
the bond betweenan antigenand a lipid, with the intent to obtain a lipidoproteic antigen and through it. a lipido-proteic defense response.Often
the mixture of the antigenwith the lipidic preparationsappearedsufficient.
Fatty acids, such as oleic, linoleic, arachidonicor eleostearic,acting
directly upon the killed typhoid microbeswere usually seento enhancethe
production of agglutinins and of specific immune antibodies. The same
effect was produced by lipoacids of the same spciesas the test animal.
Lipoacids of guinea pigs were especiallyactive in promoting the apparance
of agglutinins but less potent in inducing the appearanceof immune antibodies. The lipoacid fraction of bacteria such as B. subtilis, coli, diphtheria, acting in vitro upon typhoid killed microbes,led to the appearance
of antibodies against typhoid microbes but produced almoet no antibod.ies
againstthosemicrobesfrom which the fatty acidswere obtained.The lipo
acid fraction of tuberclebacitli bound to killed typhoid microbeswas seen
to induce agglutininsbut seemedto roduce and even prevent the appearance of immune antibodies.The same influencewas seen with the lipids
obtained frorn tlre seedsof Bixa orellana but was less accentuatedfor the
lipids from fish and squid. Butanol and especiallyheptanol were seen to
retard the appearanceof all antibodies,allergic and immune.
Allergic Precipitates
The injection of killed typhoid microbes agglutinated by a specific
serum was followed by rapid production of immune antisera. The serum
of rabbits injected with these mixtures prevents a lethal condition induced
in mice by intraperitonealiniectionof living microbesin much smallerdoses
than serum obtained with untreated microbes.
on the other hand, the injection of the same killed ryphoid microbes,
mixed together with a flocculate obtained, for instance, from egg protein,
and an antiegg precipitant-guinea pig serum-produces a much lcss
rapid appearance of antityphoid immune antibodies than injection of
microbes alone.
Another form of lipido-proteiccomplex, utilized as agent with the aim
to induce not a lipido-proteic responsebut a higher one in the defense

194

nEsEARcH tN

pHyslopATHoLooy

process,was that of allergic precipitates.Through a blender, we obtained

from rat and mousetumorshomogenates
in which it was no longerpossible
to seecells.After centrifugationthe supernatantfluid was separated,and
usedas antigen.Part of it was inoculatedto guineapigs,twice at 3-day intervals.The amount of appearingprecipitineswas determinedperiodically
and the animal bled when the serum had a sufficientlyhigh titre. Using the
sameantigenand the obtainedsera properlydiluted,flocculateswere obtaincd.The precipitateseparated
was injectedto animalshavingthe tumor
grafted.In a high proportion 6f g35s5-in more than Tovo in some experiments-the tumorsstartedto showchanges24 hoursfollowingthe injection
of the precipitate,to ulcerateor disappearin the subsequent
days. Similar
research,usingpooledhuman tumors,is in progress.
Intermediary Lysatesand Antigens
of interestwas a specialuseof the intermediarylysatesin order to obtain changesin the antigens,which would facilitatethe defensemechanism.
Microbes,tissuesor other products,serving as antigenswere injccted,
mixed with intermediarylysatesfrom blood or other sources.This was seen
to resultin a morc specificsecondday defenseresponse.
Mice injectedwith
such a mixture of blood intermediaryacid fraction plus killed microbes
showedresistanceto the inoculation,24 hours later, of the same living
microbesin dosesotherwiselethal.The protectionobtainedhas a marked
degreeof specificity.
In experimentsnow in course,we utilize blendedtumors mixed with
the intermediaryacid lysatefraction,to inducea defensein animalshaving
the sametumor grafted.
Thc discussionaboveconcernswhat could be called the immunological
part of the defensereaction.It has to be coupledwith many other processes
or phenomcnawhich can be systematized
as endocrine,vegetative,central
nervousor even psychologicalresponses.
Someof them could be indirectly
related to the interventionof lipids, and possibly involved through them
in the immunologicalresponses.
This concept of inrmunologicaldefense,even under its incompleteaspct has helpedus to understanda numberof importantpathogenicproblems, includingtwo which have becn of particularinterestto us: infectious
discaseand cancer.Our study of the infectiousdiseasesunder this aspect
w a s r e p o r t e di n a p r e l i m i n a r yc o m m u n i c a t i oinn 1 9 1 9 . ( 3 7 ) I n 1 9 4 2 ,t h i s
part of the researchwas presentedat the Congressof Medicine in Mexico
and publishedin the journal "Pasteur."(38)

DEFENSE

r95

INFECTIOUSDISEASES
Toxic and Allergic Conditions
In infectiousdiseasethe antigenis a micro-organismwhich may bc a virus,
microbe, protozoa,mycct, etc., or even a product elaboratedby a microorganism.Thc responscof an organism to the presenceof an infectious
antigentendsto follow the same successive
stagespreviouslyoutlincd. If
the meansat the immediatedisposalof the organismare qualitativclyand
quantitativelysufficientto neutralizethe antigen,the entire processwi.ll be
resolvedasymptomatically.
othcrwise, the first stage of the defensercaction, the primary toxic diphasicphcnomenon.will bc set into motion.
According to the qualitativeeffectiveness
o[ this response,manifestations
will vary from simplesubclinicerJ
changesto clinicalreactions.If thc second
phaseof the diphasicresponsecannottake place.a prolongedform of the
first phasewill result.It correspondsto shock,which is encounteredonly
in very severeinfections.The rapidly lethalconditionresuttingfrom transfusion of massivelyinfectcdblood is an examplc.
The secondphasebringschill and fever.If thc secondphaseresponse
is qualitativelyinsufficicnt,the prolongedform ensucs,bringing fever,the
usualmanifestationof many infcctiousdiscases.
Thc fever persistsas long
as the nonneutralizedantigcnis present.In this stageof the defcnsereaction againsta micro-organism
or its toxins,the symptoms,althoughresultingfrom thc responscof the organism,are still directly rclated to the
presenceof the antigen in sufficientquantity.The quantity necessaryto
induccthe clinicalmanifestations
can be reachcdwithin a short time after
the penetrationof the antigeninto the organism.The toxic reactionthus
can appear in a few hours. Consequently,
there is no specificobligatory
incubationtime. The manifestations
will disappcarwhen the amount of
antigenis decreased
sufficiently.For somemicrobes,antibioticshave such
action,resuitingin a decreasein the amount of the antigenprcsent,and
consequently
in the disappearance
of the clinicalmanifestations.
A similar
decreasein the amount of the free antigenprcsentcan be obtainedby its
neutralization
throughspecificimmunescra,if available.Conseqently,
such
serahave curativeeffectsin infectiousdiseases
characterized
by a primary
toxic pathogenesis.
Allcrgic antibodieswill appearafter an obligatoryincubationperiod of
6 or more days. If the antigenis still prcsent.it may be destroycdby the
new defensiveantiheterogeneous
responses
mobilizedagainstthe resulting
allergiccomplex.ln this case,the appearance
of the allcrgicantibodiesre-

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REsEARcH rN pHysropATHoLocy

sttlts in a kind of clinical crisis which can lead to the cessationof the disease. However, if this eftect does not occur, the appearanceof allergic
antibodieswill cause an increasein symptornsor in their gravity.
In cases asymptomatic prior to the appearanceof the allergic antibodiesbecauseof low direct toxicity or insufficient quantity of the antigen,
the diseasewill become cl.inically apparent only with the appearanceof the
allergic manifestations.The clinical condition thus will have an obligatory
incubation of 6 or more days, since this representsthe time necessaryfor
the coagulant antibodies to be produced. Since the manifestationsin such
casesare due to the allergic complex and not to the direct action of antigen,
they will be nonexistentor minimal during the incubation time. Due to the
allergic complex, the condition will not respond to specific immune sera
able to neutralize the antigen but ineffective against the allergic complex.
Specific immune sera are not curative for these infectious conditions of
alleryic pathogenesis.As already noted, only when administered before
allergic antibodies have appeared,during their incubation period, do these
sera have a marked preventive effect.
Thus the pathogenesisof an infectious diseasecan be toxic or allergic
in nature. The two pathogenicmechanismscan be identified easily tfuough
incubation time of major clinical manifestations. An infectious disease
which appears shortly after the entrance of the antigen without an obligatory incubation has to be corcidered, according to our concept, to be ol
toxic pathogenesiswhile one which appears alter an incubation time obligatory greater than 5 or 6 days has to be considered allergic.
Applying this concept, we have separatedthe clinical infectious diseases
into two groups, toxic and allergic, using incubation time as the criterion.
We wish to note here the great similarity in the incubation time for the
diseasesin each group. Most of the allergic group have an obligatory incubation time ranging from 6 to 14 days, which coincides with the usual
time neededfor the appearanceof the allergic antibodies.The incubation
time is independent of the fundamental nature of the etiologioal agentvirus, microbe, protozoa, etc.---or of the nature of their products+xo.
toxins, endotoxins, etc. This indicates that the principal factor in the
incubation time is the allergic pathogenic mechanism itself.
Based upon the criterion of obligatory incubation time, the following
diseaseswith brief incubation time have been consideredas having a toxic
pathogenic mechanism: diphtheria, botulism, anthrax (Bac. anthracis),
meningococcalinfections, cholera, some streptococcalinfections, dysentery
(especiallyShiga Kruse bac.), plague, scarletfever, pneumonia,etc. In the
allergic group, with an obligatory incubation time above 6 days, we find:

DTFENSE

l9'7

typhoid, typhus, tetanus,prertussis.

rabies,mcaslcs,poliomyelitis,glanders,
etc. (TnnlE XII) In both groups.thereare variedetiologicalagents.Thus,
in the allergicgroup, for example,the antigensinclude a microbe with an
exotoxin (tetanus) with an endotoxin (typhoid), a rickcttsia (typhus),
and a virus (rabies).
'f,rsLe

Xll

lNrrctrous Dtselsus
Incubation
Low
Dyphtheria
Anthrax
Botulism
GaseousCangrene
Plague
Erysipelas
Dysentery
MeningococcicInf.
Cholera
Pneumococcus
Inf.

Ohligatoryubove 6 duys
Typhoid
-f
etanus
Pcrtussis
Glander
Tularenria
Leprosis
Typhus
Rabies
i\'teasles
Munrps
Poliomyelitis
Smallpox
Chickenpox
Recurrentfever

The concept of toxic and allcrgic pathogenesisfor these diseasesis

impressivelyconfirmed when we consider the cfTectsof specific immune
seraupon their evolution.The specificserademonstratecurative properties
for all diseasesin the first group with brief incubationtime, consideredin
our conceptbecauseof this incubatingtime as toxic. Not one of the conditions of the secondgroup, consideredas allergicon the basisof their
incubationtime alone,can bc curedby immunescra.Still more impressive
is the fact that, in spiteof the lack of curativeeffect,the sameserahave a
marked preventiveeffect upon the sameallcrgic conditionsif administered
beforethe onsetof the symptoms,that is, duringthe incubationperiod.This
confiffnsour explanationthat the therapeuticinefficiencyof the sera in the
secondgroup is due to the allergicpathogenesis
of the diseaseand not to a
lack of active antibodies.Moreovcr, the sarnesera have a curative action
upon infectionswith brief incubationperiods induced experimentallyin
animalswith the same agent.Thc concepthas been confirmed in most of
the infectiousdiseasesand we will discusssomeof thesediseasesbriefly.
Before discussingthis aspectof infectiousdiseasesin more detail, we
want to mentionanotheroccurrencewhich can bc interpretedalso through

198

p , E s E A R c Hr N p H y s r o p A T H o L o c y

the concept of allergic conditions. It concerns a kind of recurrence of

symptomsseenoften around the 7th day after the beginningof the clinical
condition in infectiousdiseaseswhich, by themselves,have allergic pathogenesissuch as typhoid, mumps,measles,pertussis,etc.
While in thesecasesthe condition itself can be consideredan allergic
manifestationagainstthe infectiousagent as antigen,a 7th day exacerbation in the course of the clinical condition can be interpretedas a second
allergicreaction.This time a new antigenhas to be considered.This appears
to occur with the first allergic manifestation,the new reaction appearing
7 days later. The complex antigen-coagulant-antibodies
responsiblefor the
clinical manifestationsof the allergic condition could representthis secondary antigen. Besides the antiheterogeneous
reaction-+nrymatic and
lipidic-which determine the symptoms of the condition, this complex
inducesthe appearanceof a new group of coagulantantibodiesagainstit.
Around the 7th day after the beginning of the clinical condition when
thesenew coagulantantibodiesagainstthis secondaryantigen appear,they
induce the exacerbationseen.
The existenceof these secondarya.llergicreactionstoward secondary
antigens,often themselvesof allergicnature, explainsmany of the tardive
manifestations
seenin the courseof infectiousconditions.
Pneumococcic P neumonia
This disease,which appearsafter a very short incubation pcriod, has
the characteristicsof a primary or toxic condition, with chill marking the
beginningof the clinical manifestations.Antipneumococcicimmune sera,
correspondingto thc type or even to the subgroupof the etiologic microbe
are curativewhen administered
in time and in adequatedoses.In the natural evolution of the disease,a crisis usually appearson the Sth day. This
correspondsto a marked aggravationof symptomswhich can be so intense
as to lead to death.The term "crisis" indicatesthis characteristicexaggeration of the manifestations.The coincidencebetween appearanceof the
crisis around the 8th day and the moment when allcrgic manifestations
gcnerallyoccur has suggestedan allergicnature for this crisis.
usually, the courseof the diseasechangessuddenlyafter the crisis.
Most of the marked manifestationsdisappearin a short time. This allergic
crisis, with an initial increasein the severityof the condition, is not seen
in patientswho have receivedspecificserumwhich has actedto preventthe
crisis as allcrgic reaction.In pneumoniathe crisis has its beneficialeffect,
a fact which accordswith the conceptthat allergic interventionprovides a
new opportunity to resolvc the interventionof the noxious agent. As we

r::l::.
'';."";..
,

DEFENSE,

,/

199

have seen,the allergicstagerepresentsa second,more complex method of

combattingantigens.In the caseof pneumococcic
pneumonia,this allergic
defenseeftort is often successful,The evolution of the diseascis stopped.
Diphtheria
We considereddiphtheria,becauseof is characteristicshort incubation
time, to be a typical primary-toxic discase.This is also confirmcd by the
curative effectof its immune serum.However,the diseasehas a manifestation in which we recognizean allergic nature: diphtheria paralysis.with
a usual obligatoryincubationperiod of about g days, and always of morc
than 6 days, diphtheriaparalysisis a typical allcrgic condition. once present, it resistsdiphtheria immune serum,yet it can be efficientlyprevented
by the same senrm. In animals such as guinea pigs and hamsten, such
paralysiscan be induced and its allergiccharacterclearty recognized.The
heatingof toxin at 56"C reducesits direct toxicity without impairing its
antigenicproperties.Administrationof even huge amountsof heatedtoxin
does not produce any immediatetoxic effect. yet the heated toxin, evcn
though it has lost its toxic effect,inducesparalysis,
classically, this para.lysishas been related to a hypothcrical thermostabilefractionof the toxin, with a high incubationtimc. This view does
not agree with thc resultsof our cxperimcnts.When different amounts of
the same heatedtoxin are injectedin guineapigs of the same sex, age and
weight, the incubation time for paralysis,although always above 6 days,
changes,becomingparadoxicallylonger when higher dosesare used. with
great amountsof the heatedtoxin, correspondingto 20,000 lethal dosesof
the nonheated toxin, incubation time becomesas long as 14-17 days in
contrastto 8-9 days for relativelysmall amounts.TesLr XIII showsthis
relationship.
T,rsle XIII
Changes
in theincubation
timeof paralysis
incluced
by diftcrentamounts
of heateddiphtheria
toxin
Arnoutrt used

.5 LD
I L D
5 L D
20 LD
IOO LD
1000 LD
5000 LD
20000 LD
'Onein [our animals
didnotshowparalysis.

Incubation ti,tleurcrage ol 4 anintals

8.33 daysr

8.25
8,00
8.25
9 . 75
I 1.00
13.75
t5.75

200

nEsEARcH rN PHysropATHoLocy

If the paralysis were induced by direct action of a thermoatabilefraction of the toxin, then higher doses of the fraction should reduce, or at
least, not increase the incubation time. This paradoxical fact can be explained simply through the mechanismof allergic pathogenesis.It is a fact
common to immunological reactionsthat an organism has gteater difficulty
in manufac{uring any antibody when very large amounts of antigen are
Presentthan when smaller amountsare involved. This difficulty is translated
into a longer time necessaryfor the appearanceof the antibodies.As seen
in our experiments,in the caseof an a.llergicreaction, this difficulty in the
manufacture of coagulant antibodies would result in a longer incubation
time.
The localization of the auergic manifestation as paralysis can be explained in part through the affnity of toxin as antigen for nerves and in
part through the participation of the nerves in the allergic reaction. The
levels at which the diphtheria toxin acts seem to be tissular, organic and
systemic,with preferencefor the adrenals,inducing characteristicsuprarenalitis. When coagulant antibodies appear, no manifest systemic allergic
reaction will occur with the antigen still present in the blood. The allergy
will be manifest,however,at the lower tissuelevel and especiallyin the
nearby nerves. Antigen must be presentin the nerve at the moment of
appearanceof the coagulant antibodies if paralysis is to occur. This can
be demonstratedby using sensitizingand triggeringinjectionsof toxin in
animals.
we sensitizedguinea pigs to heated and unheatedtoxin by injecting
relatively small amounts intravenously.On the sixth or seventhday, another small quantity of the same toxin was injected, this time near the
sciatic nerve. The total amount of toxin was far below the lethal dose. Two
or three days later, paralysisdevelopedin the injeaed timb in a high proportion of animals while no such paralysiscould be observedin animals
injected only intravenouslyor with the same total amount of toxin at once
in the limb. In other experiments,the daily injection of small amounts of
toxin, whetherheatedor nonheated,near the sciaticnerve, induced paralysis although the total quantity of toxin was much lower than that which
ordinarily would induce para.lysis
in any similar animal. paralysisappeared
in thesecasesafter an incubation period of about 12 days. The animals
sensitizedby one or more injectionsof heatedtoxin respondedto the nonheated as triggering injections and vice versa, indicating that antigenic
propertieswere responsiblcfor rhe paralytic allergic manifestation.
In humans,anti-diphtheriaserum,eftectiveagainsttoxic manifestations.
had no effect upon paralysisonce it had appearedbut is very effective in

i tr',riilliiiiLiliill

DEFENSE

201

preventingit. The s,unewas true in animal experiments.Injected 24 hours

before the appearanceof paralysis.the serum had a consistent preventive
action. This fact confirms again the allergic pathogenesis
of the paralysis.
In anotherexperiment,we showedthat administrationof cortisone,with
its anti-allergic action, also reducesthe incidenceof paralysis without having the same effect upon the direct toxic action. It can be noted, too, that
among small laboratory animals,diphtheria paralysiscan be induced readily
in guineapigs,lessreadily in hamsters,and not at all in adult rats and mice.
In addition to the sensitivityof guineapigs to diphtheria toxin, this can be
related to the great capacity of theseanimals in general to produce allergic
antibodiesand thus to be subjectto anaphylacticreaction.Basedon these
considerations,we can classify diphtheria paralysisas a typical localized
allergicreaction.
Typhoid
Typhoid, as seen in humans, is an allergic condition with an incubation period obligatory longer than 6 days. However, in experimentalanimals the samemicrobe inducesa condition with a short incubationperid,
a fact which indicatesa primary toxic pathogenesis.
This differencecan
explain the striking difterence in results with immune sera. The literature
emphasizesgreat efficacy in experimentalanimals for various immune sera
prepared againstthis microbe and its endotoxin (Chantemesse,Besredka,
Kitasato, wassermann,etc.), but no efficacyin the human disease.
In our experiments,
whena suflicientamountof microbes,aliveor dead,
was injectedat once, a primary-toxiccondition was induced in guineapigs.
The incubation period was brief. with the same microbe, alive or dead, we
were able to obtain the allergic form in guinea pigs with repeated,daily
injections of small amounts.The ailergic form similar to that seen in humrns was induced. After about 12 days, temperaturestafled to rise and
usually remained high for more than two weeks, even with the dead
microbe if the injectionswere continued.lf living microbeswere used,the
condition continuedeven without new injections.We even obtainedpositive
hemocultureat this time. Under similar conditions,the same allergic form
of typhoid also was induced in rabbits although much less consistently
than in guinea pigs. An antityphoid serum obtained from rabbits showed
activity against the toxic form of infection induced in guinea pigs. It was
entirely ineffective againstthe allergic form when that was already present,
although the total amount of microbes injected over a period of many days
was smaller than was used to induce the toxic form. Injected before the

202 /

R E S E A R c Hr N p H y s l o p A T H o L o c y

8th day, the same serum prevented the appearanceof the allergic form of
the experimental disease.
Tetanus
In the light of our concept, we studied tetanus pathogenesisin an effort
to explain the classicallyemphasizedseparationbetweenthe so<alled small
and big animal disease.(39) In mice, tetanushas a short incubation period
and is manifestedby localized contractions,while the diseaseof so-called
"big animals" starts with trismus after an incubation period of more than
6 days. Based upon incubation times, we consideredtetanus to be the toxic
form in small animals, the allergic form in large animals.
we could, in fact, induce a condition in mice manifested by trismus
and epistotonus,and having an incubation period longer than 8 days, by
daily repeatedintravenousinjectionsof small amountsof toxin. The special
affinity of this toxin for the nervous system,and the strong bond between
nervous tissue and toxin, limited the response to the antitetanic scrum
even of the primary form. If injected in time, however, the antitetanic
serum controlled this primary form. The same serum appearsto be highly
effective in the prevention of the condition in animals which have been
prepared for the allergic form, provided it is administered before the allergic manifestations have appeared. The same senrm is totally inaaive
once the allergic condition is present.
Rabies
Under all circumstances,rabies needsan incubation time of more than
5 days. From our point of view, therefore, it has to be considered an allergic condition. when the rabies virus was passedrepeatedly through the
brains of rabbits, incubation time became continuously shorter and ultimately was fixed at 6 or even 5 days. Classically,this progressivelyshortened incubationtime is interpretedas being due to progressivelyincreased
virulence of the virus after these passages.In the tight of our concept of
the pathogenesis
of infectiousdisease,a reductionof incubationtime is the
result of increasedvirulence onJy in casesof primary direct toxic pathogenesis.In the allergiccondition,which has an entirely differentpathogenic
mechanismwith the incubation period related to the time necessaryfor
the body to producecoagulantantibodies,a shorterincubationwould correspond to a different change.It resultsfrom a greaterfacility of the organism
for manufacturingantibodiesagainstthe infectious agent. In the case of
rabies, a short incubation period of 5 days for a "fixed virus" would mean
that the organism is able to manufacture allergic antibodies more easily,

DEFENSE

703

and consequentlyearlier. Apparently,antibodyproduction is more difficult

for the "streetvirus" which has a longer incubationtime. The fixed virus
consequentlyappears to be not only a brain-adaptedvirus, but also a
weaker antigen,againstwhich the body and especiallythe nervoussystem
is more easilyable to manufactureallergicantibodies.
The possibility of using the fixed virus as a vaccine in an individual
already infected with street virus can be explained by differencesin the
relationshipbetweenthe organismand the two viruses.Usually the street
virus has a much longer incubation time, indicating that the body needs
much more time to manufacture the allergic antibodies.The same as the
animal is able to make allergic antibodiesin a short time against the fixed
virus, the vaccinatedindividualwill be able to manufacturemore rapidly
also the protectivcneutralizingantibodicsagainstthe same changeclfixed
virus. The neutralizingantibodieswill thus appearearlier and act against
the "street virus" before the organismin generaland the nervoussystemin
particular has made allergic antibodiesagainstit.
It is possible,however,that an additionalfactor may intervenein this
case. In vaccine, we use an allergic complex as it is present in the nervous
system. This corresponds to a further step in the general process of immunity and its presencecould shortenstill more the time necessaryfor
the appearanceof protectiveneutralizingantibodies.
The conceptof rabies,clinical and experimental,as an allergic condition has recent-lyreceivedconfirmationthrough the results obtained with
a specific antirabiesserum (Koprowski). With no curative capacity, this
serum is able to prevent the diseaseif it is injectedbefore the appearance
of the clinical condition,even if onJyshortly before.It helps in caseswhere
no more time is left for active immunity to be establishedby the body itself
as a responseto the vaccine.This passiveimmunity is consequentlyindicated for the case in which vaccinationstarts late. The serum, with no
curative effect once the clinical condition has started, has a preventive
eflect, a fact which accordswith the conceptof rabiesas a condition with
allergic pathogenesis.
Syphilis and Tuberculosis
Syphilitic chancre has the characteristics
of an allergic condition. The
first lesion,often a small blisteron a noninduratedbase,showsa minimal
reaction in spite of its richnessin treponemas.It is only after an incubation
of about 9 days that the intensivereaction appears,with the characteristic induration. Becauseof this, tle chancre can be consideredto be a
specificallergic manifestation.The positive lutein reaction also corresponds

204

nEsEARcH rN pHysropATHoLocy

to an allergic response.The appearanceof secondarymanifestations also

can be indicative of allergic pathogenesis,but with another antigen than
the treponema involved.
Severalpossible antigens have to be considered.One would consist of
constituentsof the microbe itself againstwhich the body needsalmost one
month to manufacturespecificcoagulantantibodies.Another antigen would
correspondto constituentsof the body itself becoming heterogeneousunder
the influenceof the treponema.A lipido-proteinicantigenseemsplausible.
Complement fixation, flocculation, and other diagnostic tests for syphilis
use antigenswhich are not directly obtained from the microbes but usually
correspondto lipido-proteic fractions of organs,such as heart. In the origlnal reaction of Wassermann,the antigen was an extract from organs rich
in treponema,such as the liver of stillborn infants with heredosyphilis.
This would favor the hypothesisthat a secondaryantigen is involved and
that it has its origin in the body constituents heterogenizedthrough the
influenceof the spirochetae.A similar antigenwould accountfor the pathc.
genesisof the secondarymanifestations.By extending this concept, the
tertiary lesionsand parasyphiliticmanifestationscan also be seento be of
similar allergic pathogenesis,with other newly formed antigensinvolved.
Secondaryantigens would conceivably develop in tuberculosisas well.
White the primary tuberculouschancrecan be seen as an allergic manifestationhaving the lipido-protein of the microbe as antigen, the cavern
formation can be attributedto a secondaryantigen.
It appears highly probable that it is these "secondary" antigens,together with the inability of the organismto manufactureefficient immune
antibodies,that keep the defensein the allergic stageand impart to both
tuberculosisand syphilis not only their chronic character but also their
clinical gravity. The allergic pathogenesis
explainsalso the inefficiencyof
all the tentativesto obtain sera againsttheseconditions.
Streptococcal I nl ections
Erysipelasand many other streptococcicinfectionsappear as primary
toxic diseaseswith a short incubation.Active immune sera in sufficient
amountsinjectedin time give good results.Often a marked changein the
symptomsis seen toward the Sth day, a fact which could be considered
to indicate passageinto the allergic phase.Many other manifestationsof
streptococcal infections, such as those seen in rheumatic fever, can be
consideredallergic.
The glomerulonephritiswhich appearsas a complicationof scarletfever
or of pharyngealstreptococcicinfection is especiallyinteresting.While a

,;
:.

";*:
, " .si-"":^:,,, ;- .1: ;.
'
'-\;ij
uigfff;
-:i. * s

: . ,, :is ,'

DEFENsE

205

change in gencral symptomsin theseinfectionsis seen toward the ltth

'l-he
day, this complicationusuallyappearstoward the 24th day.
intervention of a secondaryantigenresultingfrom the bond betweenlipids and thc
renal tissuescan be hypothecatcdin the light of studics concerning immunologica.ldefense processesagainst tissues,which we present in the
following pages.
We do not want to leave the problem of infectiousdiseaseswithout a
few more words about the use of lipids in the defensemechanismagainst
microbes.The fact that lipids liberated in the first defenseresponsesare
bound to microbes and intervene in this complex form to promote the
appearance
of higherdefenseprocesses
has led us to use similar bonds in
order to stimu.latethis defense.We have seenabove how lipids other than
thoseofferedby the infectedorganismcan be used.The injectionof killed
microbes treated with lipoacid from hetcrogeneous
sources,such as from
speciesnaturallyrefractoryto the microbe.hasenhancedthe defensemechanism. Microbestreatedwith lipoacidsof the tuberclebacilli or of Bixa
orellana were seento induce a strongspccifica.llergicresponse.
Interestingresultswcre obtaincdthroughthe usc of insaponifiable
fractions bound to the microbes.The fractionsobtainedfrom refractoryspecies
appearedto be most effectivein enhancingthe defensemechanismin general. The insaponifiablefractionsobtainedfrom the entire body of rats,
animals refractory to most infections,gave the best resultsfor most of the
infectionsstudied.
in addition to using killed microbestreated
In these investigations,
with lipids in vitro, we employedanothermethod to treat the microbes.
Lipoids were added to the media in which the organismswere grown.
Somc of the lipoids were seento increase,and othersto decre:u;e,microbial
virulence.Killed and usedas vaccinesin casesof resistantinfections,these
lipoid-treatedmicrobeswere seen to induce more eflectiveimmunization.
Experimentsin progressindicatethe possibilityof using such microbes
-and even virusesso treated-to obtain long-lastingimmunity. Microbes
with very reduced virulence are used as live vaccines.Their capacity to
induce effectivedefenseresponsesin a short time also his led to their use
as "late" vaccines,i.e,, vaccineswhich can be administeredduring the
incubationtime of an infection.As thesestudiesare still in progressan
evaluationof the resultsis not yet possible.
An interestingaspect of the influenccexerted by lipids upon microorganismsis their use in producingqualitativechangesin antibiotics.Prcliminary researchshowsthat the addition of lipids of the microbesagainst

I iil

.,.iii

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n E S E A R c Hr N p H y s r o p A T H o L o o y

which more active antibioticsare sought seemsto alter the antibiotics

that they have a higher degreeof specificityagainstthesemicrobes.
IMMUNOLOGICAL DEFENSE AGAINST
CELLS AND TISSUES
Heterogenizationof the Transplants
It is known that the introductionin a normal subjectof cells or tissuesfrom
an animal of another speciesor even a transplantfrom thc same spccies
will induce the appearanceof defenseprocesses.These differ with the degree of heterogeneityof the transplant.Experimentally,we can vary this
degreeof heterogeneityof transplantedcells and study the different responsesin the frame of the normal and abnormaldefensemechanism.
For the highly heterogeneous
transplant,such as cells or tissue of a
strangespecies,a primary responseoccurs, with liberation of hydrolytic
enzymes and lipids. If sufficiently strong, this response will destroy and
eliminate the transplant. If the transplant is moderately heterogeneous.
such as one from an individual of the same species,the primary reaction
is milder so that the transplantedtissuesurvivesthis attack. It will. however, be killed and rejected with the appearanceof the second defense
stage,i.e., that of the allergic reaction.The damageto the transplantcan
be attributedto the antiheterogeneous
reaction,which this time appearsto
be directedtoward the product resultingfrom the bond betweenthe transplant and tissueallergic antibodies.
For a still less hetcrogeneous
transplant,such as one from young animalsof the samespecies,
the two defenseresponses
arc nrild. However,the
transplantis often destroyedthrough a later interventionof immune antibodies. This is seen to occur after some months for organs or for cell
transplantssuch as bone marrow cells for the treatmentof severeradiation
damage.In these cases,the defensemechanismwhich intervenesmonths
after grafting can be correlatedto the immune stage.An autograft,which
is a perfectly homologoustransplant,usually will survive. The fate of a
transplantthus appearsto be determinedby its heterogeneity.
This heterogeneity, howcver, does not result only from the differenceswhich exist
betweendonor and receiver.Even an autotransplantcan be heterogenized
by surgicalmanipulation,heat or other treatment,or by changingits organizationalrelationshipto other entities.to such a degreeas to be destroyed
by an immune, allergicor even a primary defenseresponse.
The heterogeneityof the transplant-intrinsic or induced by the appli-

DEFENSE

2O7

cation of external agents-represents only one factor which determines

the nature of the defense processes.Another factor corresponds to the
changesin the antigen or constituentsinduced by the intervention of prifl&rJ, allergic or even protective irnmunological reactions. The study of
the defense reaction against the organism's own tissues or cells heterogenized by previous immunological responsesis of special interest, in view
of the role of such heterogenizedentities in a more complex defensemechanism. The organism often heterogenizesits own entities through the agents
used in the defense against foreign entities. primary, allergic and even
immune reactions induce various degreesof heterogenizationof the organism's own constituentsat various levels of the organization.
Through the intervention of hydrolytic enzymes,lipids, allergic antibodiesor even neutralizingantibodies,differentchangesin an organism's
orvn entitiescan be induced.From these,the heterogenization
of body entities by lipids was studiedin particular.The heterogeneous
effectof lipoacids
could be shown in many experiments,
as in the following: Suspensions
of
cells of differentorgansof guineapigs,in a concentrationof I gram of cells
to l0 cc.of saline,were prepared.At the sametime lipoacidsuspensions
in
saline were obtained starting from ZVo solutionsof different lipoacids in
alcohol. Four weekly administrations
to guinea pigs of the separatecell
suspensions
or of the lipoacid suspensions
were not followed in most of the
animalsby any seriousmanifestations.
A heterogenization
of the cells was
obtainedthroughthe actionof the lipoacidsuspension
upon the cetls.While
one singJeinjection of the so-treatedcells showed no noxious manifestations, consecutiveinjectionsat weekly intervalswere seento induce,in less
than a month, importantchangesgenerallyconcerningthe respectiveorgans
from which the cellsderived.
The lipoacid-cellcomplexactsas an antigen,with the type of cell determining the organ where the abnormal changeswill occur, and the lipid
determiningthe characterof the occurring reaction.Depending upon the
lipoacid, the effect will vary from minimal tissular lesions all the way to
changesleadingto death.
massivedegenerative
The degreeof heterogeneityof the lipoacid appearsto be one factor
which determinesthe stageof defenseinduced.Oleic and linoleic acids,and
the lipoacidsfrom human placenta,cow liver or total body of guinea pigs
had a slightereffect in inducing organ lesionsthan the lipoacids obtained
from Bixa orellanaand especiallyfrom the tuberclebacilli which led to
seriousdamagein the respectiveorgans.Tuberculinacting upon the cells
had the sameeffect as lipids obtainedfrom tuberclebacilli.
Through variations in the nature of the autogenousfactors-hydrolytic

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R E s E A R c Hr N p H y s r o p A r H o L o c y

enzymes,lipids, allergic antibodiesor even immune antibodies-a graduated seriesof changesin an organism'sown entities can be induced. of
the factors which intervene in the heterogenizationof such entities, we
have studiedthe lipids in particular. An antigenicrole for lipoacidscould
be shown in many experiments.For example,suspensionsof red cells or
cellsof different tissuesof guineapigs in a concentrationof I gram of cells
to l0 cc. of saline were prepared,At the same time lipoacid preparations
wereobtainedin the followingmanner.5 cc. of a Z% solutionin alcoholof
difterentlipoacidsor mixturesof lipoacidswere added to ll0 cc. of water
and the preparationboiled under low pressureuntil reducedto 100 cc.
The cell suspensions
were administeredto guineapigs in four injections
at weekly intervalswith no seriousmanifestations.
The same was done for
the lipoacidsabove. A preparationwas obtainedthrough the action of the
lipoacidsupon the cell suspensionin the following manner. 5 cc. of the
colloidallipoacid aqueoussuspension
were addedto 5 cc. of the suspension
of cells of different tissues.To 5 cc. of red cells, only I cc. o[ the lipidic
suspensionwas added. The mixture in each case was incubated for two
hours at 37"C and centrifuged.The cellular residues,separatedfrom the
supernatantfluid, were resuspendedin saline zr.ndkept frozen. While one
injection of the so-treatedcells showed no noxious manifestations.consecutiveinjectionsat weekly intervalswere seen to induce, in less than a
month, manifest changesin the respectiveorgans. with the red cells, a
marked anemia was induced. oleic and linoleic acids, and the lipoacids
from human placenta,cow liver or total body of guinea pigs had only a
slight effect in inducing organ lesions.The lipoacids obtained from Bixa
orellana and especiallyfrom the tuberclebacilli led to seriousdamage in
the respectiveorgans and resultedin death usually in less than 3 weeks.
Tuberculin in thesecaseshad the same effect as lipids obtained from tuberclebacilli.
The degreeof heterogeneityof the lipoacid appearsto be the factor
which determinesthe stageof defenseinduced,The lipoacid-cellcomplex
acts as an antigen.with the type of cell determiningthe organ where the
abnormal changesoccurs, and the lipid determiningthe character of the
occurringreaction.Dependingupon the lipoacid. the effect will vary from
minimal tissularlesionsall the way to massivedegenerative
changesleading
to death.
The intervention of a bond betweencells and lipids apparsevident
when acid lipid preparationsare injected repeatedlyat week.lyintervalsin
the sameorgan. Lesionsare obtainedwhich are similar to but lessintensive
than thoseproducedby the cell-lipoacidcomplexes.

DEFENSE

20c)

It was highly interestingto note the differences

in lcsionsdependingon
the origin of the cells injected.Zones of necroeis.often with subacute
cellular degenerationand even with inflammatoryprocesses.were induced
by not too heterogeneous
fatty acids.Acute glomerulonephritis.
Iiver degeneration,pneumonia,enteritis or encephalitisresulted from repeated
injectionsof cel.lsfrom kidney, Iiver. lungs, intestinesand brain treated in
vitro with bixin, lipoacidsof tuberclebacilli,lipoacidsof fish or even fatty
acid mixtures of cod liver oil.
Against tissuetransplants,injectionsof the host with lipoids with negative character-such as fatty acids.mixture of lipoacidsof differentorigins,
and lipids with SH or SeH as polar groups-have exaggeratedall phasesof
defenseprocesses.
skin transplantsbetweensiblings,which usuallygive a
high percentageof acceptedgrafts,were rejectedcompletelyafter treatment
with some of theseagents.The degreeof heterogeneityof the agentsap
peared particularly interesting.with lipoacids of the same species,very
higl doseswere requiredto induceonly minimal changcs.on the other
hand, preparationsof lipoacidsof fuh, mollusk, molds and microbesproduced marked effects.Transplants treated with these preparationswere
rejected or absorbedafter eight or more days. Seldom was an immediate
rejection seen.
Even more interestingwere the resultsobtainedby direct action of the
lipids upon transplants,achievinga bond betweenthem. For theseexperiments, the agentswere usedin oil solutionsas weUas in salinesuspensions.
Transplants were dipped into diflerent preparations.Even autografts if
treated with lipoacidsof the same speciesoften werc rejected.This took
place even after more than three weeks.when more heterogeneous
lipoacids were used, such as those obtained from other species,autografts
were rejectedas completelyas transplantsof the same species,i.e., around
the eighth day. This also occurred with relatively heterogeneousagents,
such as the lipoids of microbes,especiallythose of the tubercle bacilli.
with still more heterogeneousagents,such as lipoids with SH or seH
polar groups, the treated transplantswere rejectedthrough an immediate
direct inflammatoryreaction.
The influenceexerted by injectionsof the opposite group of ripoids
with positive character was in the opposite direction. The percentageof
acceptedtransplantswas increased.
By dipping skin transplantsof animalsof the samespeciesin preparations of the insaponifiablefractionsof the species,the percentageof persistentgraftswas highly increased.ln someexperimentsall the transplants
between siblingswere positive.Even betweendifferent strains of mice,

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R E s E A R c Hr N P H Y s T o P A T H o L o c Y

such positive results were obtained. The treatment of transplants with

butanol alone was not effective.Adding butanol to the preparationof insaponifiablcfractions,however,enhancedthe effectof the latter.
The most interesting results were obtained by cross-treatment-in
which the transplant was treated with the insaponifiablefraction of the
strain of the host and the host with the insaponifiablefraction of the donor.
An unusual number of positive grafts were obtained between strains of
mice and, in exceptionalcases,even betweenspecieswhen a mixture of
the two preparationsof insaponifiablefractionsof the donor and host was
usedfor the treatmentof both transplantand host.
Even more interestingresultswere obtainedwhen, in addition to these
treatmentsof transplantand host, anothertreatment-that of the "bed" of
the transplsnf-"rin5added.The wound receivingthe transplantwas soaked
with the mixture of insaponifiablefractions. Often aJter the graft, treatment with the fractions was continued through small injections into the
bed of the transplant.Injections into the transplant itself, if possible,increasedthe number of positiveresults.
Before pursuing further the study of these interestingproblems, an
analysisof another aspectof the responseto hcterogenizedmatcrial has
appearednecessary.It concernsthe interventionof different levels of the
organizationin the defcnsemechanism.This was seen to vary according
to the degreeof heterogeneity
of the transplant.The defenseprocessesthus
can be limited to the heterogenized
entity or to entitiesof the samelevel.
or they can extcnd far into the hierarchic organization.With a highly
heterogenizedmaterial, a broad hierarchic reaction occurs with several
superiorlevels intervening.In thesecasesprimary enzymaticor prolonged
processes
involve the tissular,organicand systemiclevels.With lessheterogcnization,the ensuingprimary reaction is not strong enough to destroy
and eliminatethe heterogenized
entitiesand an allergicreactiontakesplace.
This involvcsother levelssuch as tissularand even organic.With still less
intensiveheterogenization,the defenseremains localized at the affected
level itself and is weaker.With the defenseinefiicientin its primary or allergic stages,a protectivestagebecomesnecessaryin order to take care of
the heterogenized
entities.
The analysisof many conditionsindicatesthc importanceof the different factorsfor the developmentof the clinical manifestations.
Seventh Day Manilestations in Trauma
we have studied these defensereactions for trauma, the degree of
intensityof the trauma indicatingthe extentof the exogenousheterogeniza-

DEFENSE

2lI

tion. Very intensivetrauma can produce a lethal superacuteshock which

corresponds,as we shall see below, to a generalizedprimary response.A
less intensive trauma may induce a tissular necrosis with consecutive
sloughingas a localizedprimary defense.A still less intensivetrauma may
induce an allergic tissular response.The importanceof these changesfor
clinical manifestationsis such that it appearsnecessaryto emphasizethem.
After surgery,for instance,a slight temperatureelevationis often observed
betrveenthe 7th and 9th day, This has to be interpreted as an allergic
reaction. When intensiveenough, this allergic responsewith the ensuing
l}tic action passesfrom thc tissular level to the higher level of the blood
vesscls.Along with inflammation and pain, local hemorrhagesoften appear. Severehemorrhagesoccur at this time after various traumatic incidents. The most disturbing complicationsfor plastic surgcry of the nose,
for instancc,are the sevcre "7th day hcmorrhages."The fact that they
start at this critical moment indicatesthcir allergicpathogenesis.
The study
of theseallergicchangeshas shown that they occur in the evolution of all
traumatic lesions. They can occur and remain clinically inapparent and
uneventful,as seenin the followingexperiment.
In groupsof rats of the samesexand agekept under similarconditions,
parallel skin incisions 3 cm. long at t/z cm. intervals were made. The
lesions were excised at differcnt times and chloride content determined.
Fig. 78 shows the curvc of averagevalucsof the total chloride content
of theseskin woundsin goups of six animalsfor each day. It can be seen
that intcnsivelocal chloride retentionoccurswith the first <lefense
reaction,
with valuesas much as four times greaterthan thoseof normal tissues.On
the third day, chloride content falls. It goes below normal tissue values
after thc fifth day during the hcarlingprocess.However, in an othcrwise
regular curve, thcre is a distinct temporaryincreasein chloride content on
the 8th day. Its occurrenceat this time, when coagulantantibodiesappear,
indicates its allergic nature.
The same allergic pathogenesis
explainsthe exacerbationof symptoms
seen about the 7th day in many conditions.In patientswho have suffered
a myocardialinfarct, for example,recurrenceof pain is often scenthe 7th8th day aftcr the infarct.
Part of the effectsof chemical,physicaland hormonal agentscould be
interprcted in terms of influenceexerted upon thc different factors which
intervenein the defenscmechanism.Someagentssuch as opium derivatives
were seen to affcct the liberation of hydrolytic enzymeswhile others interfere with the manufactureof allergicor immune antibodies.The influence
exerted by radiation upon the defensemechanismcan be related to its

2r2 /

RESEARCH IN

PHYSIOPATHOLOGY

o
f
!
j

g
o

E
:

|J

Flc 7E. Wound chloride ctrrve. The curve of the amount of the chlorides present
in skin wounds in rats correspondsto the average value obtained in 6 rats for each
figure. A first phase, with high values correspondsto tbc offbalance D. This is followed by a second phase characterizedby an offbalanceA. A variation in the curve
correspondingto the 8th day is constantly seen as corresponding to an allergic reaction. The values representmgs. of chlorine per 100 gr. of weight of the wet malerial.

effect upon granulocytesand lymphocyteswhile neoglucogeniccofticoids

affect the connective tissueand lymphocytes.
Researchin all these directions is still in progressand the results will
be communicatedin further publications.
For the time being,they havebrought more information and suggestions
of researchin the specialcaseof the immunologicalproblem of cancer.
IMMUNOLOGICAL

PROBLEMS IN CANCER

We usedthe data obtainedfrom the analysisof the defenseprocessagainst

cells and tissuesin the study of the immunologicalproblem of cancer.The
different canceroushierarchicentities,as defined previously may be con-

l}.;gii:r,+.#t
;,*4dliiljiliaq$;$,f,j##i;.;issi

DEFENSE

213

sideredto correspond,up to a certain point, to heterogeneous

entities-the
grafted tumors to transplantsand the spontaneoustumors to heterogenized
entities of the individual.
We tried, in a first seriesof experiments,to follow the interventionof
the differentmechanismsof defenseon graftedtumors in animals,employing transplantsof various degreesof heterogcneity.Different types of tumors were used. Highly heterogeneous
tumors obtainedfrom speciesother
than the host would not grow when transplanted.The death of the transplant. even its rejection if mechanicallypossible.occurs in a short time.
The necrosisof the transplantand the relativelywide inflammatoryprocess
that developsaround it immediatelyafter the graft indicatethe intervention
of the first stageof the defensereaction from the cellular to the organic
level.
with a secondgroup of tumors, usuallyfrom the same speciesor even
moderatelyheterogeneous,
the grafts take and the tumors grow for a time.
often, around the 8th to the l5th day. the transplantstarts to show pro
found changes.The changesaffectthe entire tumor which involutesrapidly
and is often expclled.That this is due to allergicreactioncould be shownby
the following experiment.Fragmentsof the samekind of tumor which had
been obtained from different animals were transplantedat 2 to 3 day intervals in the same host. In spite of the different agesof the transplants,
the death and rejection of all occurred at short intervalsand in the same
manner, indicatingthe interventionof a mechanismtaking place in the
host and relatively independentof the evolution of the transplant itself.
Such a mechanismwould be the interventionof allergic antibodies.
In a third group of grafts of low heterogeneity,
the tumors continue to
evolve for an even longer period of time, and it is only in a few animals
that these tumors are entirely rejccted.This change,which consistsof
cytolysisof the tumor, takes a certain time to be completed,which indicatesthat it probably resultsfrom the interventionof protectiveantibodies.
The first and the seconddefensemechanismsappearedto be inadequateto
conquerthe tumor and it was the third stage,with formationof protective
antibodies,which apparentlywas able to accomplishit. This mechanismis
confirmedby the fact that later grafts are negativefrom the time they are
transplanted,through cytolytic changesin the tumor and not through an
intensiveinflammatory processas seen in the primary reaction. The immunologicalnature of the defensein thesecasescould be seenalso through
the passiveimmunity which could be inducedin other animals with the
serumof the host.

214 i

xESEARcHtN PHYsropATHoLooy

Grafts in Humans
The grafting of cancerouscells in normal humans usually leads to the
appearanceof a growing tumor which (315) after a period of days, almost always sufters the same fate as a moderately heterogeneoustransplant: death followed by resorption or expulsion. The time when this processo@urs indicatesthe allergic natureof the defensemechanism.It is highly
probable that, without the intervention of this efficient allergic defense,the
cancerousprocess would have continued to evolve.
Such continued growth occurs if grafts are made in subjects already
having their own cancerousprocess.This would indicate that the allergic
defensemechanism against the graft is no longer operating in these cases.
The fact that a canceroussubject acceptsa new tumor graft while the
normal one rejects it indicates that defenseprocessesare different for the
normal and this canceroussubject.The inability of thesesubjectsto reject
a grafted tumor through an alleryic responseappears to be the major immunological difference between the normal and the cancerous subject. Still
more important is the fact that in cancer patients,the anomaly would correspond to a lossof the capacity to reject the grafted tumor which the normal
subjectseemsto have.
In trying to determine the nature of the immunological anomaly in the
evolution of a spontaneouscancr in patients, we have to relate it to this
loss of the defenseprocessesas seen above.
In studying this occurrencein general,a loss of defenseagainst:rn antigen can be conceived to occur for any of the three different mechanisms
involved in defense:primary, allergic or protective.In caseswhen this takes
place, the loss of the protective stagewill take place first. The allergic defense will be affected next and finally, the primary response.This explains
why the inability of an individual to achieveone stageof the defenseleaves
the defenseresting in the immediately previous stage.The inability to manufacture protective globulinic antibodies,for instance, will leave an individual in the allergic stagewhich, in the developmentof defense,preccdes
manufactureof the immune antibodies.This results in a potential allergic
condition if the antigen is absent or an actual allergic condition if the antigen is present.We have seenthat this occurs in most of the chronic infectious conditions.Similarly,with the inability of an organismto manufacture
coagulantantibodies,the defenseremains in the previous defensestage,the
primary lipidic one.
Before going further, we have to discussa factor believed by many
authon to be involved also in the defensemechanismagainstcancer. A few

..,:::: :,,::.

.)

DEFENSE

215

yearsago, the defensemechanismin generalhad beenrelatedto the properdin system.Howcver,whenconsidered

in termsof the systematization
of thc
defenseprocesses,properdin has to be regardedas a nonspecificdirect antinoxiousreaction.It appearsto be involvedin the antiheterogeneous
defense
response,appearingafter the enzymatichydrolytic attack and at the bcginning of the prolongedlipidic intervention.
The decrease
in the properdincontentof thc blood of subjectswith cancer has causedvarious authors to try to explain through it the differences
in the reaction of cancerousand normal subjectstoward a new transplant.
The analysisof the conditionsunder which this occurs,however,has shown
us that the anomaly does not residein the antiheterogeneous
processesof
defense,which are the same againstantigen and allergic complex, but in
the allergic reaction itself. From the immunologicalpoint of view, the differencebetweena normal subjectand one with invasivecancer residesin
the loss of capacity to induce the secondtype of defense,the allergic,toward the canceroustissue. Correspondingto an inability to manufacture
coagulantantibodies,this deficiencywould explain the lack of respective
antiheterogeneous
reaction toward the antigen-coagulant-antibodies
complex and consequentlythe low blood content of properdin seen in these
cases.
Failure of the allergic defensemechanismspecificallyagainst cancer
entitiesneed not mean generalfailure of allergic defense.The failure may
be limited to inability to manufactureallergic antibodiesagainsta specific
antigen.We have seen,especiallyfor the infcctiousdiseases,that primary
and allergicprocessescan occur with great intensityand still not be qualitativelyefficient.The agent,the microbe,for instance,can still rcmain present despiteeven violent ailergic reactions.The mere presenccof defense
processes
does not implicitly mean successful
defense;they may be qualitatively insufficient.
In cancer,if the allergicdefcnseis insufficicnt,
two eventualities
havc to
be considered:either the organismin generalcannot passinto the allergic
stageof defenseand thereforeis unableto manufactureallergic antibodies,
or this responseis only qualitativelyinsufficient.In thc latter casc,the general and even local reactionscould be quite intensivebut still be ineffective.
This seemsto occur only in certain forms of cancer such as those with a
high inflammatoryprocess;for instance,in the inflammatoryform of breast
carcinoma.As this cancer starts and evolvesas an acute mastitis,very intensive defensiveprocesses,apparentlyonly of the primary stage,occur.
But they are unable to check the diseasewhich usually cvolveseven more
rapidly in these cases.This is also true for other cancerswhere fever is

216

nESEARcHrN pHyslopATHoLocy

present,indicatinga prolongedprimary, toxic stage.The lack of local reaction seenat the site of the growing transplantin the canceroussubject at
the time when the normal individual kills or rejects the transplant points
to the fact that the anomaly residesin qualitative inability to manufacture
allergicantibodies.
The next problem was to investigate the reason for the failure of
allergic defense against the tumors. we could show that the cancerous subject has not lost the capacity in general to manufacture coagulant antibodies.Even subjectswith very widely spread cancer were able
to respondwith a local skin allergicreactionto a secondinjection of an antigen (proteins from mollusks) made more than ten days after a first preparatory one. (Note 7) Their inability to fight transplantedcancer cells
through a similar allergic reaction indicatesthat the loss of this capacity
is not generalbut relatively specifictoward the cancerouscells. The lack
of an intensive inflammatory process, as well as the existence of high
amountsof lipids in the canceroustissue,also would indicateindirectly an
inability of the canceroussubjectto resolvethe existing immunological
problem of fighting cancer through an allergic reaction. The presenceof
large amountsof lipids indicatesthat thc defensemechanismhas been
arrested in the stage of pronounced lipidic predominance. Abnormal
amountsof lipids thus could representan indirect meansof recognizingthe
failure of an allergic responseto cancerousentities.
The next problem was to try to determinewhere in the organization
the failure occurs.The different levelsof the organizationare independent
to a certaindegreeand passageof an abnormalityfrom one level to another
induceshierarchicprogressionof the condition. This has posed the problem of the progressiveloss at the differentlevelsof the natural capacityto
defend againstcancer. Recently many investigatorshave shown that cancer cells passinto the lymphatic systemand into the generalcirculation in
a much higher proportion than had been suspectedbefore. Malignant cells
in the circulation are destroyed,however,by the defcnsemeanswhich are
not lost at this level.The samepatientthus may still have an activelygrowing cancerat the tissularlevel,indicatingthat this defenseprocess,although
successfulfor the higher levelsof the organization,does not interveneat
this lower level.
The hierarchicprogressionof cancercan be seenas a proefessiveloss
of the immunologicaldefensecapacity.while the organism conservesthe
capacityto fight at a higher level, a lower hierarchicentity no longer opposesthe cancerouscondition. It is not the absenceof cancerouscells in
blood or organs which explainsthe lack of an explosivespreadof the dis-

DEFENSE

217

ease,but the presenceof efficientdefensemeansat theselevelswhich keeps

a cancer still localized.
Metastases
The relativeindependence
in the lossof the defensecapacityof different
entities would explain one of the most baffing problems of cancer-why
certain cancerstend to metastasize
to certain organsor tissues.Somecancers show bone metastases,
othersspreadto many organs,while still others
spreadonly to certain specificorgans.This can be explainedby a loss of
the defensecapacityat the organ level. While some organslose,others still
maintain their allergicdefensecapacity.The circulatingcancer cells will
inducemultiplc metastases
in the first but will not be able to take hold in
the latter.
A similarmechanismcan alsoexplainthe persistence
of inactivecancer
cells for years after an operation.The defensemechanism,while it is not
able to aftect the cells and destroy them, is still sufficientlyactive at the
tissular level to prevent the condition from progressingat this level. The
cancr cells will start to invade this level only when the tissular level is
unable to defend itself further through an allergic responseagainstthe invading cells. By losing its allergic responsecapacity,the tissular level will
even exaggeratethe correspondinglower primary stageof defense,that is,
the prolongedlipidic phasewith the consequentchangeswhich this brings
on. Among them would be the appearanceof pain.
Under thesecircumstances,
the generalimmunologicalcondition favorable to the hierarchically progressivedevelopmcntof cancer has to be
regardedas the loss of the capacityof the different levels for an effective
allergicresponsetoward cancerousentities.The immunologicalproblemof
cancer consequentlyappearsin a speciallight, different from all the other
known conditionswherean unsuccessful
immunologicalresponseis present.
In the other conditions,the problem of the inability to conquer an antigen
is one involving the incapacityto mobilizeor developan eftectiveimmunological response.In cancer,the body appearsto have lost a previouslyexisting capacitythat was presentbeforethe diseaseappeared,In other diseases,
the immunologica.lproblem is to create a new and favorable condition in
the fight againstan antigen,by developingmeanswhich do not exist in the
normal individual. In cancer. the immunologicalproblem would be to
prevent the loss of a property possessed
by normal subjectsor. if already
Iost.to find somemeansto regainit.
Theseconsiderations
and the study of the differentfactorsinvolvedin
the developmentof the progressive
defensestagehas led us again to the

218

RESEARcH rN pHysropATHoLocy

role of the lipids in these processes.The appearanceof a stage in the

defense mechanism seems to be strongly related to the frrlfillrnent of
the qualitative requirementfor the previous stage.Deficiency of essential
factors in one stage representsan impediment for the next stage. In the
case of cancer, failure of the specffic allergic phase thus could be traced
to a qualitativelyinadequateprecedinglipidic phase.A level is unable to
surmountan allergicdefensebecausethe lipids which can mobilize arequalitatively inadequate.Even abnormal richnessin lipids thus could be interpreted as resulting from their qualitative inadequacy.This very excess
indicatestheir importance.
Imntunological Therapeutic Approach
The fact that effectivedefenseresourcesare present at a highcr level
does not mean that they inevitablywill act at a lower level.
This view of the abnormal immunological processesin cancer has
pointed the way for some new therapeuticapproaches.From the therapcutic point of view, the problem becomesone of how to induce the body
to regain, at the necessarylevel, the lost immunologicalcapacity,and
througha specificallcrgicdefense,to combatthe cancerousentities.Furthermore, sincethis specificallergicdefensecapacityis lost independentlyby the
various levels of organization,the immediate problem would be how it
could be recoveredfor the particular level where the loss occurs. The existenceof adequatedefensecapacityat a higher level, such as the systemic,
docs not provide a solution for cancer presentat lower levels such as the
tissular,becauseof the independence
which exists betweenlevels.only
the manufactureof coagulantor immune antibodiesagainstcancer at the
proper levcl would put the individualin a sufficientlyactivedefensephase
to enablehim to resolvethe condition at that level.
The problem of immunity at the proper level thus appearsto be critical
for any immunologicalattack againstcancer.It is evident in other conditions as well and has inspired the use of local vaccination in localized
infcctiousconditions.( 307, 308)
The study of immunity againstviruseshas permitted us to recognize
the importanceof immunity at differentlevels.Virus infection is a typical
cellularcondition,the virus multiplyingonly within a cell. Theoretically,an
immunity at all levels can be induced for viruses.According to the view
discussedabove,however,a systemicimmunity with circulating antibodies
will not insure a cellular defense.It would interveneonly when the virus
is passingthrough the systemiclevel and its activity would last only during
the time when circulating antibodiesare present. During this time, the

DEFENSE

219

virus will be prevented from reaching the cells. Once tbe circulating antibodies iue no longer present, the cells cease to be protected. For an efficient defenseagainst viruses an immunity within the cell appears thus to
be indispensable.The use of dead virus vaccine will induce only systemic
immunity, which can be recognizedthrough the circulating antibodies. It
is unlikely that the killed virus enters the cells. It does not aftect them,
and consequentlydoes not induce cellular immunity. Even a mild cellular
infection with living virus will give the necessarylongJasting cellular imrnunity. This would explain the need for living and not killed vaccinesfor
viral infections, as first postulatedby Pasteur.
A similar level immunity can explain the differencesseen between the
immunity resulting from the use of microbial vaccines and that produced
by natural disease.Typhoid infection gives lifelong immunity; the vaccination, only relative and temporary immunity. An explanation can be found
in the fact that, in the disease,along with the septicemia,manifestchanges
occur in organs and tissues.Spleenand lymphatic tissuesare highly affected
in typhoid and it is possible that the developmentof the defense at their
level would explain the lifelong immunity that follows the natural infection.
rn cancer, the problem would be to induce not a systemic defense,
which is still present for invasive cancer, but an effective tissue or even
cellular defense.Immunologicaltreatmentof cancer would have to make
tissular and possibly cellular levels regain their capacity to defend themselves through efficient allergic responses.The immunological prevention
of cancer would lie not in the creation of this defenseor in increasingit
quantitativelybut enhancingit qualitatively.A successfulallergic defensc
at this level apparently would have a preventive and even curative effect.
The use of lipids in the induction of the defensemechanismagainsttissues
has an interestingapplicationin cancer.A systemictreatment with lipids
or lipoids can change the defenseresponseso that it can be effective at
a specific level where it is otherwise inadequate.For invasive cancer, the
lipid activity must be induced at the cell level. The active lipoids for this
purpose are those with a high affinity for the cancerouscell.
As abnormal cells in general show similar capacity to bind the lipoids
administered,this general affinity becomesa handicap if abnormal entities
other than cancerouscells are present. These considerationshave led us
to attempt to use methodswhich will insure the activity of lipids at the ccll
level.
In one of these methods,the chosenlipoids are brought directly into
contact with cancerouscells through local injections into the tumors. SingJe
injections produce only limited changes in tumors. Local injections re-

22O /

REsEARcH rN pHysropATHoLocy

peatedso as to insure the presenceof the lipoids once, and then again 15
days later, are required to induce an eftective response.The lipoids or
lipids are so chosen that, when bound to body constituentsthey will induce allergic or immune defenseresponses.The acid lipids of tubercle
bacilli, bixine or guinea pigs are especiallyprone to induce allergic reactions,while the lipids of microbes-such as coli, typhoid or diphtheria
-produce immune responses.
In another method, lipids chosenwere bound to cancerousentities in
vitro. Cancerouscells were obtainedand treatedin vitro with lipids under
whoseinfluencethe body is able to manufactureallergic or immune antibodies. colloidal suspensionsof the lipids or lipoids were prepared as
mentionedabove,mixed with suspensions
of cancerouscells, kept at 37.C
for a few hours, then separatedfrom the non-fixedlipids and injectedinto
patients. In order to obtain good results it was necessaryto inject this
material at least twice, at an interval longer than two weeks, in order to
insurean allergicreactionagainstthc cell-lipidpreparation.While a single
injection produced good resultsonly in a very small number of cases,repeated injections were manifestly more effective.When canceroustumor
cells could be obtained through biopsy from the patient, we used thenr
for the in vitro treatmentwith lipoacids.when biopsy material was not
available,we used cancerouscells of similar origin as the tumor of the
subject,preferablypooled.
The condition for successof these methods has appearedto be the
presenceof the cell-lipidcomplex at the moment of appearanceof antibodies. This is assuredonly by the repetition of the injection. Another
interestingaspectof the immunologicalproblem in cancer, related to loss
of the natural defensemechanism,is the loss by cancerentitiesof their
capacity to utilize certain elements known to intervene in the defense
mechanism.The role of magnesiumin the properdin system, copper in
cytochromeoxidase,of calcium in generaldefense,suggestsa correlation
betweentheir deficientutilizationin cancerand the loss of the defense.We
will discussthis problembelow,after reviewingthe pharmacological
aspect
of theseelements.

CHAPTER

T H E C O R R E L A T I O Nt s E T W E E N
T H E B A S I CC O N C E P T S
rTrl
rs A cLEARinterrelationship
l[
betweenthe four basicconceptspre"r^u
viouslydiscussed
which permitsus to considerthem togetherand to establish a unified viewpoint.For all four can be scento representdifferent parts
of the same fundamentalproblem in biology: the manner in which an
entity resolvesenergeticdifferencesbetweenitself and the environment.
we have seen that, in the framework of fundamentallaws governing
nature, matter can be consideredto correspondto isletsof heterotropyopposingthe homotropictrend of evolution.Conservationof an existingentity
appe:us to be the principal meansby which heterotropycan be achieved.
And heterotropyis fulfilled, specifically,through maintenanceof the constantsof entitiesas valuesdifferentfrom thoseof the environment.
H i erarchic Organization
The continuoustendencyof natureto progresstoward maximum homotropy has made the conservationof existing entities a persistentlyacute
problem. The problem posed by the progressivelychanging environment
cannot be solvedthroughchangeswithin entitiesthemselves.
Any "adaptation" of the entity itselfwould affectits constantsand, consequently,
would
be contrary to the fundamentalpurposeof heterotropy.Nature has resolved
the problemin an entirelydifferentway. Sincethe entity itself must remain
unchanged,and yet the influenceof the environmentmust be conteracted,
nature has made use of hierarchic organization.Secondaryparts, reproducing the immediateenvironment,are joined to existingentities.often
surroundingthem and acting as buffersagainstenvironmentalinfluences.
Through theseadded secondaryparts,hierarchicentitiesare organizedso

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..'iJ**u.Ab$.s;:i.il;:.,'
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222

xESEARcH rN pHysropATHoLocy

that they reproducethe characteristicsof the environmentpresent at the

time of their formation. Through this means successivelyrepeatedmany
times, an entity can be kept unchanged,in a medium similar to the original
one, despitecontinuingchangesin the environment.
Hierarchic organizationthus representsthe main mechanismthrough
which the heterotropic achievements,representedby entities, counteract
the influence exerted by the homotropic force. Conceptually,hierarchic
organizationcan be seento representa form of defensedevelopedin time
by entitiesagainsta specificfactor, progressivehomotropic changesin the
environment.The successivesteps of the hierarchic organization respectively the hierarchicentities,reproducein short, the evolution of the relationship which has been developedbetweenthe entity and the changing
world. Hierarchic organizationcondensesthe phylogeneticevolution of
this specificpart of the dcfenscbetweenthe entity and the changingenvironment.Through this view, we can integrateorganizationin the general
dcfense,the hierarchicorganizationbeing part of the mechanismused
againstprogressinghomotropy.
Constituents
In the same manner, we can further integrate into the same defensc
mechanismthc various constituentswhich form the secondaryparts of the
hierarchicentities.We have thus tried to correlatethesc constituentsmore
directly to the successive
environmentsin which entitiesevolved.We have
seenabove, how this appliesto elementswhich are common to, and predominantin, both the entitiesand the environmentswhich correspondto
the media in which theseentitiesevolved.Through the correlationbetween
elementswhich enter into hierarchic organizationat various levels, and
their positionsin the periodic chart, the successivephylogeneticpassage
from one environmentto anotherhas a specificmeaning.In media formed
by elementswith lower atomic weight,the influenceexertedby progressive
homotropy is less manifest.The changesin the elementsas body constituents can bc thus also integratedin the samedefensemechanism.
Besidcsthe elements,other constituentscan be similarly integratedinto
the defenseagainstthe changesof the environment.In the immediatedefense processagainstnoxious agents,we have seen the successiveintervention of different constituents---{nzymes,lipids, lipido-proteins and
proteins,in that order. The high degreeof individuality and independence
of the entities in the hierarchicorganizationhas permitted us to conceive
of theseconstituentsas participatingwith a certain independencefor each
cntity. The presenceof all these constituentsin each higher biological

T H E C O R R P L A T I O NB E T W E E N T H E B A S I C C O N C E P T S

223

cntity, which is part of the complex organism,suggeststhat theseconstituents entered into the formation of these entities as the result of thcir
interventionin the defensemechanism.Thus, it can be conceivedthat. in
its phylogeneticdevelopment,each entity has passedthrough a succession
of defense phases in which specific groups of constituents-nzymatic,
lipidic, lipido-proteicand proteic-have been predominant.In actual organization,while all higher entitiescontain fundamentallythe sameconstituents, different substancesare predominant at different levels. This can
be explainedby the predominanceof a particular defensemechanismat a
particularlevel.According to this view, this defenseis principally in the first
stage,that is, of enzymaticnature, for most endodermicformations. It is
in the prolongedlipidic stagefor ectodermicformations,lipido-proteicfor
the reticuloendothelialsystem,and proteinic for cells. Through thesecorrelations,constituentscan be more completelyintegratedin defcnse.
The kind of special defensedevelopedfor the different levels of organization, through predominant specific constituents, has not been
followed by a total discard of the other constituents,which do not have
such roles. Instead,the latter have been retainedin the entities in smaller
amounts and in inactive forms. This confers upon the entity the capacity
to mobilize these constituentsand use them when the need to respond
to an acute emergencyarises.Pre-fermentsand cven ferments in mitochondria; fatty acids and anti-fatty acids bound as esters;lipido-proteins
and proteins in various combinations-all these are inactive constituents
which can be changed easily into active agents. when fighting a new
noxious intervention,an entity will resort to liberating or activating these
constituentskept in reserve.Each entity and level of organizationdoes
this indepcndentlyof other entitiesand levels,yet constituentsactivatedat
one level can act at other levels, too. The successor failure of defense
especiallyin its first stages,dependsnot only on the intrinsic value of thc
constituentsavailable, but also on the capacity of the affiicted entity to
utilize these means by activating them. Although activation processesbecome strikingly evident in abnormal conditions similar processessecm to
be important even in the maintenanceof existingentities.
Dualism, as we have seen, characterizesboth normal and abnormal
physiology.That which is considered"normal" is the result of an altcrnating intervention of two groups of opposite constituents,producing an
oscillatory movement and a dynamic balance. The dualism seen in abnormalities,when one or the other opposedfactor is persistentlypredominant, is related to hierarchicorganizationand the defensemechanism.
Dualism results from the intervention of two fundamental forces in

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nESEARcHrN pHystopArHoLoGy

nature-homotropy and heterotropy. Even the simplest analysesmake

evidenteither the homo- or the heterotropiccharacterfor many manifestations and processes.
For instance.an ulcerationor an enzymatichydrolysis
of a protein has to be interpretedas an homotropiceffect while a growing
tumoral mass or the synthesisof a protein can be seen as an heterotropic
one. For other manifestations,this characterappearsless immediatelyevident and it is through further analysisthat it can be recognized.Dualism,
like the other concepts,thus can be integratedin the defenseof entities
against an environment progressivelychanging toward maximum homorropy.
We have used this conceptual fundamental view in studying many
problemsin biology. It has aided us to formulate helpful working hypotheses.Despite its shortcomings,when applied to particular situations,
this basic concept has served as a guide in correlating specific problems
with the fundamentallaws governingnature. [t has also engenderedhelpful new interpretationsof availabledata. Through the relationshipof the
four conceptsdiscussedabove and the fundamentaldefensemechanism,
we have been able to analyze many problems without reverting to empiricism. Certain of these problems. to whose better understandingthis
approach appears to have contributed, are discussedin the pages that
follow.

CHAPTER

S H O ( ]K
r
ll* ro,r, oF THE pRocRESs
rcalizedin the lust decade,shock remains
one of the most challengingproblemsin medicine.That lipids have a
critical role in shock pathogenesis
seemsclear from a long-term study
which beganwith an investigation
of the activity of fatty acids in the induction of the abnormallydark color of blood secn in shock.The results
of this study will be discussedhcre not only becauseof the intrinsicinterest of the problem of shock itself but a.lsobecauseshock often represents
the terminal phaseof cancer as it does of many other diseases.In this
presentation,we will try to remain as much as possiblewithin the framework of our direct contributionto an understanding
of shock. A portion
o f t h e s er e s e a r c h ewsa s p u b l i s h e di n 1 9 4 3 .( 4 0 )
In studyingthe very complex phenomenonof shock, one has to consider a seriesof well-definedproblems.Shock has been related not only
to a large number of causesbut also to a seriesof vcry varied clinical
manifestations.
An initial problem was to determinewhetherthere is any
common relationshipbetweenthe different types-between the shock, for
instance,which kills a subject within a few minutes after a severesudden
trauma, and the shock that kills in days through profound systemicmetabolic impairments.What is common to, and what is different between
them, from the point of view of pathogenesis?
What constituentsintervene
and how, in shock?Theseand many other problemshave beenapproached
systematically.
T-vpesol Shock-As a starting point, we attenrptedto classify the
types of shockand found an interestingrelationshipaccordingto the time
that is, the intervalbetweenapplicationof thc noxious
of their appearance,
Three typescould be identifiedwith
stimulusand onsetof manifestations.
this criterion.
225

226 /

R E s E A R c Hr N p H y s r o p A T H o L o G y

There is an immediate type of shock which appears within a ferv

minutes after the application of the noxious agent. It is induced experimentally in animals by intravenousinjection of a noxious substance,by
scaldingthe animal in hot water, or by strong mechanicaltrauma. It has
predominantcentral nervous system manifestations,including exophthalmia and paralysis of the posterior limbs, followed by clonic convulsive
movements,and usuaUy is terminated by death. A similar superacute
type of shock is occasionallyseen in humans following transfusionsof
blood with an incompatible group. It also may be seen following very
severetrauma. In the case of bullet wounds, for example, large calibre
bullets may bring rapid death. Neither immediate hemorrhage nor any
organ impairment is sufficient, in itself, to account for the speed of death
in many of these cases.However, it can be explained by the rapid and
intensive participation of the central nervous system in this superacutc
type of shock. Sometimessuch shock is not lethal in animals or humans
and is followed by a period of prostrationand ultimate but slow recovery.
We called this type of shock the "superacute."
In a second type of shock, more frequently encounteredin humans,
the manifestationsappear after a certain period of time. Such shock often
is seen after direct transfusions,when the rate of injection has been too
rapid or when the syringeand tubeshave not been well coated with oil or
paraffin,or when thcre has been a subgroupincompatibilitybetweendonor
and receiver. The patient usually experiencesa severe chill within 30
minutes, The chill is succeededby a rise in temperature which usually
Iasts 15 to 60 minutes or more. The patient next experiencesdiaphoresis,
after which the episodeusually is concluded.In some casesthe symptomatology is different. At about the same time-30 minuts5-after transfusion, for instance, hypotension with hypothermia, cold and clammy
perspiration,and intensivedyspneaare noted. In thcse casesdeath can
follow in a short time. The same reaction is sometimesseen to occur,
usuallyalso in about 30 minutes,after the releaseof a tourniquet.We have
employedthe term "acute shock" to describethis secondtype characterized by its appcaranceat approximately30 minutes after the noxious intervention.
A third form, the "state ol shock," is considerablyslower in onset and
persistsmuch longer.Characterized
by hypotension,impairmentof circulation, cold and clammy perspirationand markedenophthalmia,it may lead
to death after scveral days during which the condition progressivelyincreasesin severity.It can, however,also end in recovery,This is the form

sHocK

227

most often encounteredin clinical medicine, in cancer and many tenninal

conditions.
The next problem was: could a common pathogenicmechanismbe
recognizeddespite the greatly varied manifestationsof these three forms
of shock?
Shock Mechanism
We saw one primary correlation between the three clinical types of
shock in the fact that sometimesone type is followed by another. Superacute shock, if not lethal, may be followed by acute shock which, in turn,
can change into a state of shock.
But it was the chemical analysisof blood, organs and entire bodies of
animals killed by any of the three types of shock which indicated the possibility of a mechanism common to all three. A low antitryptic power of
the blood, and the presenceof substancesresulting from protein hydrolysis
were found to characterlznall 3 types of shock. Additionally, ut increase
in the amount of free fatty acids, and the presenceof abnormal members,
occurred h all three types.
Fatty acids were studied from the point of view of the reciprocal position of their double bonds, through the oxidative fission method mentioned
previously. The appearanceof oxalic acid following oxidative fission indicates the presence of conjugated double bonds. The oxalic acid index
obtained indicates the proportion of these conjugated double bonds. In
normal rats, this oxalic acid index usually is zero in the total amount of
fatty acids; in normal mice, values below I are seen. In all animals in
shock, even in cases of superacuteshock followed by sudden death, the
oxalic acid index is invariably much higher. Furthermore, the death of an
animal in acute shock or state of shock appearsto be related to the presence of a critical oxalic acid index, indicating a concentration of abnormal
fatty acids incompatible with life. Whether it appears in a relatively short
time as in acute shock, or many days after the noxious intervention as in
the state of shock,the oxalic acid index found in dying animals is between
14 and 17. Such high values are not found in superacuteshock but the
oxalic acid still is markedly increased.Thus, the presence of hydrolytic
processestogether with abnormal fatty acids appears to be a common
pathogenic factor for the difterent forms of shock.
Pathological Clanges
The three types of shock-because of the presencein all of hydrolytic
processesand abnormal fatty acids-<ould be related to the first phase of

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228

R E S E A R c Ht N p H y s t o p A T r { o L o c y

the immcdiatediphzsicdefensephenomenonor its prolongedform. The

next problemwas to determinewhat othcr factorsmight influencethe developmentof diflering manifestationsso as to make shock appear in three
forms.
The study of pathologicalchangescharacterizingeach of these forms
was undertaken.We found cellular vacuolationa characteristiclesion in
animals in superacuteshock. Vacuoles are present in the parenchymal
cellsof the liver, to a lesserextentin the alveolarcellsof the lung, and to
a still lesserextent in kidney ceUs.Of special interest was the fact that
thesevacuolesare often seen in the cytoplasmand even in the nuclei of
brain cells. These findings explain the predominanccof the neurological
symptomsin this form. [n a publicationin 1943,we describedthis vacuolation as a characteristicof the superacuteshock. The fact that the characteristicpathologicalchangeencounteredin superacuteshock is the presence
of vacuolesin different cells suggeststhat this form of shock occurs principally at thc cellularlevel.
ln the acute type of shock, which usually appearshalf an hour after
noxiousintervention,there may be someevidenceof cellularvacuolization,
but the principal changesare at the tissularlevel. The changesare largely
localizedin the immediate areasdamagedby the noxious agent and are
manifestedby vascularand interstitialpathologysuch as marked edemaor
capillary hemorrhage.Splanchnicvasodilatationand pctechiaeat the surlace of pleura or peritoneumappear when the noxious agent acts indirectly in the blood or is applieddirectly to it through intravenousinjection.
The degreeof generalizedvasculardamagecorrespondsto the degree of
direct participation of the blood. We have discussedpreviously, in the
chapteron defense,the changesoccurringin the blood which characterize
hemoshock.Thc characteristicleucolysis,which is followed by hydrolytic
digestion,explainsthe high degreeof breakdownof blood constituentsand
vessels
observedin this kind of shock.While the participationof the cellular level-and especially of the central nervous system----characterizes
supracuteshock, particiPationof the tissularlevel leadsto the acute form.
We considerpathologicallycharacteristicof the stateof shock-in addition to the changesseenin blood, such as hemoconcentration,
dark color,
tendencyto form sludges,etc.-two other specificmanifestations;
milliar
lesionsin the gastric mucosaleadingto hemorrhageand ulceration,and a
manifestfluid accumulationin the first portion of the small intestine.Since
the variouschangesin the stateof shock affect the blood and two organs,
the stomachand duodenum,they can be consideredto involve the organic
and systemiclevels.

This analysishas permitted us to continue to u.";;;rt*r;,r:t:

that all three forms of shock stem from the samefundamentalmechanism
-the appearancof abnormal fatty acids as part of the first phaseof the
diphasic defense reaction. The differencesin manifestations between the
formsof shock are due to the level at which the mechanismoperates,cellular for superacuteshock, tissular for acute, and organic and systemic for
the state of shock.
The study of a special condition, hemoglobinuria "a frigore," or
paroxystic hemoglobinuria, has helped us to understand the time factor
in shock. In this condition, immersion of the hand in ice water, for instance,induceshemoglobinuriaand violent chill about half an hour later.
We have been able to demonstratethat in the developmentof such a
manifestation,two or often even three hemoshocksoccur, each one characterizedby a diphasic phenomenon.The first shock appears within ten
minutesafter immersion of the hand in icy watcr. Usually the first sensation and chill are very slight and while a reduced hemoglobinemiais present, hemoglobinuria is almost nil. It is the second hemoshock, appearing
approximately 30 minutes later, which is usually very intensive with manifesthemoglobinuria.The third shock,which appearsabout two hours after
immersion in ice water, is usually clinically inapparent and is revealed
only by blood analysis.
The study of this condition has indicatedthat in the appearanceof the
three episodesof hemoglobinuria,besidesthe changesin the red cells under
the influenceof cold. which are characteristicfor the condition as seen in
test, the importantfactor is the leucolysisoccurthe Donath-Landsteiner
ring as part of the hemoshock.The subsequenthemolysisleads to free
hemoglobin in the blood which, if in sufficient amount, passesinto the
urine. The changesinduced in leucolysiswill determinethe degreeof consequent hemolysis. The suppressionof leucolysisby administration of
morphine or other opium derivativesprevents any manifestation,while
physicalexerciseundertakenfollowing the immersion of the hand in icy
water induces,in addition to a very intensiveleucolysis,exceptionallyintensive clinical manifestations, The time when the three hemoshocks
appearalso marks the time when the threc forms of pathogenicshocksuperacute,acute and state of shock-are seen.The interventionof three
different noxious heterogenizedconstituentsappearsplausible.(Note l)
Fatty Acids and Sodium Chloride in Shock
We noted that in all three types of shock, abnormal fatty acids can be
found. A studv of the role of thesefatty acids permitted us to further un-

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RESEARcH rN pHysropATHoLocy

derstandthe mechanisminvolved in thesethree types of shock. Since these

samefatty acids have been seento figure in abnormal metabolismof sodium
chloride, the next logical step was to investigate the correlation between
the latter and shock. Following this line, efforts were made to see if the
difterencesbetween NaCl metabolismat different levels of organization
would help explain the peculiaritiesof the different types of shock.
As we have noted, when abnormal fatty acids impair sodium chloride
metabolism,two processesoccur. First, there is abnormal fixation of chloride ions by abnormal fatty acids; then, sodium ions, freed following this
chloride fixation, become bound to carbonateions, resulting in alkaline
substances.The pathological nature of chloride fixation results principally
from the fact that the binding taking place at the conjugateddouble bonds
is abnormally strong. Occurring in two steps,with a displacementof the
double bond in the first, the bond betweenthe conjugatedfatty acids and
chloride ions appear nonreversible.(Note 8, Chapter 6)
The great inequality in the ability of chloridesand sodium ions to pass
through membranescan serveto separate,anatomically,the fixed chlorides
from the free remainingcations.When this occurs, two distinct processes
can be recognized,one involving the binding of chloride ions by abnormal
fatty acids,the other involvingthe binding of carbonateions by sodium ions
and the resultingappearanceof alkaline compounds.In the cells, the two
processestake place separately,the sodium alkaline compound inducing
the appearanceof vacuoles.In tissues,the chloride fixation takes place
predominantlyin the cell, while the binding of carbonate occurs in the
interstitialspaces.This leadsto a localizedintercellularalkalosiswith consequentedema.
The same mechanism is involved in the changes associatedwith the
state of shock, except that theseprocessesoccur at the systemiclevel. It
is the part played by the sodium chloride of the blood in normal physiology,
especiallyin the processof digestion,which explainsthe abnormal changes
seen as characteristicof the pathologicalmanifestationsin the state of
shock.
Normally, chloride ions are excretedinto the stomach,where they are
bound to hydrogento form hydrochloricacid. An almost equal amount of
sodium ions, bound to carbonateions, is eliminatedin a secondstep into
the intestinesvia the pancreaticand intestinalsecretions.The chloride and
sodium ions are later liberated to form sodium chloride which is entirely
reabsorbedin the distal portion of the intestinaltract, the colon. The sodium and chloride ions are not simultaneouslysecretedin the digestive
tract. The interval betweenthe excretionof chloride ions into the stomach

sHocK

231

andof sodium ions into the intestinesa@ountsfor the physiological"alkalinetide" associatedwith digestion.
when chloride ions are pathologicallyfixed to abnormal fatty acids in
theblood, they can no longer be dissociatedand secretedby the stomach
in the form of hydrochloric acid. Instead,they remain bound to the fatty
acidsand accumulatein this form within the gastricmucosa.The multiple
rnilliar gastric mucosal ulcerations in the state of shock results from the
interventionof these abnormal fatty acids brought into the mucous membraneby the chloride ions to which they are bound. The ulcerationsare
causedby the catabolicaction of fatty acids.Thus, the first phaseof abnormal sodium chloride metabolismleadsto the characteristicmultiple gastric
ulcerations.
The secondphaseis relatedto the metabolismof sodium. The sodium
ions are secretedas carbonatesby the pancreasand intestinal mucosa in
the fint part of the small intestine.In the state of shock, becausethey do
notencounterthe chloridesnormally coming from the stomach,they remain
as carbonates.As sodium carbonateis accumulatedin the first portion of
the small intestine,a local alkalosisoccurs, leading in turn to an important
loca.lretentionof water. It shouldbe noted that this is a very differentsituationfrom achlorhydriaor hypochlorhydria
in which,while the chlorideions
arenot secretedinto the stomach,no excesses
of sodium ions appearin the
blood or in the intestines,and consequentlyno local alkalosisor fluid
accumulationoccurs.
The differencebetweenthe systemicand tissueprocessesin shock lies
in the localizationof the abnormal sodium chloride metabolism.In tissue
anomaly,the separationof sodium chloride takes place between the cells
andthe pericellularstructures.At the systcmiclevcl, it occurs betweenthe
stomachand intestines,
with the blood servingas intermediary.This mechanismexplainsthe larger amountsof water which distendthe upper parts
of the intestine,as observedin autopsiesof animalswhich have died in this
form of shock.
The close similarity betweenthe abnormal processesthat take place in
sodiumchloride metabolismat the tissueand systemiclevelsprovidesthe
basisfor another working hypothesisconcerningthe mechanismin superacuteshock. We have seenthat the productionof vacuolesin cells characterizesthis latter form of shock. The unequal cellular permeability for
chloridesand sodium in their dissociatedform is known. Chloride ions can
circulatemuch more easily betweencells and the pericellularspacesthan
can sodium ions. An initial effect of the intervention of abnormal fatty
acidsin cellularpathologyis the fixationof chlorides.At the sametime, an

232

RESEARcHrN pHysropATHoLocy

increasedpermeability in membranesoccurs. This would permit more

sodium ions to passthrough cell membranesand to accumulateintracellularly, inducinga liberationof potassium,the cellular cation. As the chloride
ions are bound to fatty acids in the cells, the sodium ions in the cetls liberate potassium and join it to form alkaline compounds. Isolated in vacuoles, thesecompoundsalso accumulatewater.
Thus, we have a concept of single pathogenesisfor all three forms of
shock basedupon abnormal sodium chloride and water metabolism,with
the abnormalitytaking place at differentlevelsof the organization,cellular
for superacuteshock,tissularfor the acute form, and systemicfor the state
of shock. The displacementof potassiumby sodium in cellular physiology
contributesto the increasein serumpotassiumfound in all forms of shock.
Water Metabolism
The localized retention of water, prompted by the alkaline sodium
compoundswhich result from abnormal sodium chloride metabolism,occurs in the cells. tissuesor intestinesin the different types of shock. Many
of the differencesin manifestationsbetweenthe three forms of shock can
be explainedin terms of localizationof this abnormal water metabolism.
The sensitivityof the cells of the nervoussystemto intracellularchanges
explainsthe predominanceand severityof the nervous system manifestations in superacuteshock, Abnormal tissuewater metabolismexplains not
only the predominantlylocal characterof the manifestationsseen in acute
shock,but also the hemoconcentration
valuesin thesecases.As often seen
in burns, important amountsof water are driven out of the blood into the
damagedtissues.
The abnormal water metabolismhowever,appearsto be the principal
manifestationin the state of shock. Upper intestinal water accumulation,
rather than a general unlocalizedfluid loss, can be demonstratedin the
pathogenesisof this form of shock. In opposition to the local lesion with a
high retentionof water, the generalsubcutaneoustissuessustaina loss of
water rather than an accumulationduring shock. This would not occur if
therewere a generalincreasedpermeabilityof all capillaries,allowing water
to pilss freely. The role of water accumulationin the first portions of the
intestinedue to the abnormal loss of systemicwater was demonstratedin
animal experiments.When the small intestinal tract had previously been
removed, and a state of shock was later induced by trauma, no hemoconcentrationoccurred.
It is the participationof one or anotherof the three principal levels of
tissular or systemic-which explains why the
the organization----cellular,

sHocK

233

same pathogenicprocess,abnormal sodium chloride and consequentabnormal water metabolism, produces such different manifestations in the
various types of shock. It must not bc forgotten howcver, that in the last
analysis,the abnormalitiesin sodium chloride and water metabolismresult
from the interventionof abnormal fatty acids.Fatty acid intervention,together with the abnormal sodium chloride and water metabolism confirm
the unitary pathogenesisof the three forms of shock.
Other Changes
Other changesassociatedwith shock also can be related to the influence
exercisedby abnormal fatty acids. The appearanceof rouleaux of red cells
may be easily explained by fatty acid intervention. It is the replacementof
the nonpolarity normally present at the surface of the red cells by a dipolarity which results in the formation of the rouleaux. This can be induced
by fatty acids in vitro. Sludge formation would represent a still more advanced step in this same processand would apparto result from a polypolarity at the surfaceof the red cells.Sludgeformationshave been induced
in vitro by fatty acids added in larger amounts to plasma. (Note 2) They
contribute to the circulatory impairment considered to be an important
factor in the tissular respiratory troubles seen in shock.
We have noted that the richnessin free fatty acids interferes with the
ability of the red cells to keep oxygen fixed, a fact which would impair its
transport. This, together with hemoconcentrationand circulatory impairment, has been found to account for the black color of the blood in shock.
(Note 3/ The clinical manifestationsare characteristicof offbalanceD.
Experimentally Induced Shock
The hypothesisthat the three types of shock are caused by the intervention of the same factol-3lnospal fatty acids-has been further confirmed experimentally. The cellular changes that characterlze superacute
shock can be induced by the rapid introduction into the blood stream of
c-ven minislsl amounts of fatty acids in preparationsin which they are
bound to plasmaconstituents.
Pooled heparinizedplasma of mice was treated by stirring it in a nitrogen atmospherefor one hour with a preparation of conjugated trienic fatty
acids. The nonbound fatty acids were separatedthrough short centrifugation. The plasma was injected intravenouslyin mice. For control, plasma
treated under the sane conditions with stearicacid was used. While control animals did not show any apparent discomfort, the mice injected with
the plasma treated with conjugated fatty acids died immediately, in most

234

xEsEARcH rN PHYsToPATHoLooY

caseseven during the injection itself. With such preparations,superacute

shock was induced in what we consider a direct way, the sudden death
contrastingwith the casesof hemoshockwhere death occurs usually after
an interval of a few minutes.This characteristicof direct immediatedeath
is consistentwith the pathogenicrole of fatty acids in superacuteshock.
The tissuechangesthat character'tzs,
the acute type of shock also may
be induced by local administrationof abnormal fatty acids with the condition that sufficientamounts are used. (lt'lote4) The systemicchangesthat
typify the state of shock can also be produced by prolonged absorption
of fatty acids, as when they are repeatedlyintroduced intraperitoneally.
(Note 5 )
The relationshipbetweenshock and lipids can be further seen in the
antagonisticeffect exercisedupon shock induced with standardizedtrauma
by two groups of lipids with positive and negative characters.We have
utilized the Noble-Collipp drum on a large scale to induce shock in rats.
In some groups of animals shock induction was constant;in other goups
under the sameconditions,shockcould be inducedonly in some animals.
Nevertheless,
it was still possibleto recognizeopposite effectsinduced by
the administrationof the two groupsof lipids. In some animalseven ap
parently little influencedby the trauma, the injection of a mixture of conjugated fatty acids immediatelyafter trauma brought death within a short
time. In no other animals, traumatizedunder the same conditions, have
we seendeath occurring within the sameshort interval of time. This also
appliesto animalsinjcctedbeforetrauma.In thesecases,the animalsdied
evenduring the trauma,that is, in the drum. (Note 6)
Conversely,the administrationof sterols,especiallypreparationsof the
insaponffiablefraction of human placenta,before induction of trauma prevcnted lethal shock alnrost without exccption,whereasunder the same
conditions the same trauma produced death in a high propoftion of the
controls.Evcn when injectedimmediatelyafter trauma,this sterolpreparation preventedthe developmentof lethal shock in a high proportion of
cases.(Notc 7)
The different forms of shock, althoughresultingfrom the same fundamental abnormal process, appear to respond differently to therapeutic
agents-again becauseof the localizationof the abnormal processesat
differentlevels.Adrenalin and relatcdcompounds,when administeredin
time, are able to control superacuteshock, but they are almost entirely
without influcnce upon the other forms. While acute shock can be influencedby the administrationof a largeamountof sterolsand butanol, superacute shock is unaffected,possiblyalso becauseof the slow absorption of

sHocK

235

the sterols.None of theseagentsis of significancein the treatmentof the

stateof shockwhich is only mildly influencedby butanoland ccrtaincortical
hormones such as hydrocortisone,especiallywhen introduced directly in
the circulation.
In order to act upon the fatty acids and sodium which produce the
abnormal water metabolism,we have utilized glycerophosphoricacid administeredin large amountsparenterally.Diluted with saline,it was usually
injectedintravenously.The good resultsobtainedare discussedlater.
The use of heptanoland of polyunsaturated
alcoholshas also led to
interestingresults.It was howeverwith preparations
havingsevcralof these
agents,working at differentIevelsof the organization,
that the bcst results
were obtained.
The measurement
of the chlorideindex and of the surfacetensionof
the urine haverepresented
valuablemeansto judgethc changesoccurringin
shock,in their clinical evolutionand especiallyin the action of the agcnts
in relationshipto the occurring recovery.
The study of shock has contributedto the knowledgeof the therapeutic
problemsof cancer and other conditions.The causeof death, when a predominance of fatty acids occurs as a systemicmanifestation,corresponds
to the state of shock.The possibilityof successfully
influcncingthis form
of shock would furnish a valuabletool for the treatmentof all severemanifestationsrelated to predominanceof fatty acids.

O
C H A P T EI R
RADIATION

"ln
lL Hp sruoy

oF rHE efiects produced by irradiation upon all biological

entities has resulted in the accumulationof a large amount of data. Being
unsystematized,
this information has helped but little to resolvethe many
physiological and therapeutic problems connected with radiation. New
light on theseproblemscan be providedby relatingthem to the basic concepts we have been discussing.
Other factors also have led us to study the problem of radiation. Its
widespreaduse for the treatment of cancerouslesions with indisputable
successin many cases,and the fact that some of radiation'seffectsappear
to be quite similar to those induced by the administration of different lipids,
led us to investigatethe mechanism through which radiation works and,
especially,the possiblerelationshipbetweenradiation and lipids. We will
discusshere briefly, someof the resultsof this investigation.
Irradiation ol Lipids
We began by trying to determine the eftects of radiation upon normal
lipids in vitro. As always, we tried to guide the researchby theorctical considerations.
Investigationof in vivo and in vitro effectsof radiationupon
proteins in general showed that histones,protaminesand alkaline amino
acids are most sensitive.These constituentsof complex protein molecules
have positive electricalcharacter.This relationshipbetween sensitivityto
radiationand positiveelectricalcharacterprovideda clue as to whereto look
in fatty acid moleculesfor changcsinducedby radiation.Severalpositive
centersare presentin the energeticstructureof fatty acid molecules.One
is representedby the carbon of the carboxyl. Its positive characteris due
to its bond to two oxygen atoms. This would explain the exaggeratedion-

izationwhichtakesplaceat the levelof this carboxylgroup.

RADr^rtoN

237

other positive centers also can be recognized.we have mentioned

previously that the positive character of carbon propagates through the
chain in an induction effect that causesalternate odd carbons to be positively charged, although the strength of the positive characrcr decreases
rapidly with distance from the carboxyl. Since a double bond geatly enhances the energetic character of the carbons linked by it, induction will
result in a center in which a more intensivepositive carbon is present.
Study of the reactions that take place at the double bond in a fatty acid
molecule confirms this view, since an electrophiliccharacterpredominates

g
o

v,
0

I
6
t
.6
L

fa

I rradiated
fbnirradiated

i l aY e

Length

(np)

Fto. 79. Irradialion ond conjugation in vilro. Spectral analysesin ultraviolet of samp l e s o f c o m m e r c i a l l y a v a i l a b l el i n o l e i c a c i d ( w i t h s m a l l a m o u n t s o f l i n o l e n i c a c i d
present) irradiated with gamma rays from 80 mgr of platinum filtered radium/10 cc,
f o r 6 d a y s a t r o o m t e m p e r a t u r eF. o r t h e a n a l y s e sd, i l u t i o n so f 0 . 0 0 2 9 bi n a l c o h o l ,w i t h
alcohol as reference, were used. The absorption spectra of the irradiated (-)
linoleic acid compared with the nonirradiated ( . . . . ) shows the appearanceof conjugated trienes recognizedthrough the characteristicpeaks.

238 /

n E s E A R c Hr N p H y s l o p A T H o L o c y

at this point. when treated with sodamine,carbons forming double bonds

combine selectivelywith it, indicating the positive electrostaticcharacter
of this formation. Consequently,we thought that the effect of irradiation
would be most likely to occur here. This has been confirmed experimentally. We could demonstratethat radiations causechangesespeciallyin the
reciprocal position of the double bonds in the molecule.
The results of this researchwere originally presentedbefore the Fifth
InternationalCongressof Radiologyin London in July 1951. We will timit
ourselveshere to a short resumeof the proceduresand findings:
Irradiation in Vitro
a) Radiation of polyunsaturatedfatty acidsin vitro inducesa conjugation of their double bonds, which increasesquantitativelywith the intensity
of the radiation.This has beenshownby spectralanalysisand by the oxidative fission method. (Note // Samplesof commercially available linoleic
acid which, through analysis,have beenfound to contain variable amounts
of linolenic acid, or cottonseedoil were treatedwith radiationsof different
sources,such as radium in platinum needlesfor gamma radiation,in monel
metal for beta radiation,thorium X for alpha radiation and X rays. Table
XIV showsthe resultsof analysisof the oxalic index. Figs. 79 and 80 show
the direct spectralanalysisof the samplesbefore and after irradiation as
well as the result of their chenricalconjugation.
Comparison of direct spectral analysesshows the appearanceof an
important amount of conjugatedtrienesin the irradiatedsamples.Analysis
after chemical conjugation of irradiated and control samples shows a
greateramount of trienesin the irradiatedsampleindicating that a process
of desaturationa.lsohas taken place through irradiation.
A dirc.ctrelationshipwas observedbetweenconjugationand amount of
radiation. (Tnalr XV) The quantity of conjugatedisomers,determined
by spectralanalysesand measuredby the oxalic index was seento increase
as radiationwas increasedeither by prolongingexposuretime or increasing
the amountof radium used.
b) Irradiation of fatty acids appearsto induce the appearanceof conjugated trienes. when a mixture of polyunsaturatedfatty acids, such as
thosefound in cod liver oil, was exposedto a radiation sourceconsistingof
platinum filtered radium, the changeswere limited to the appearanceof
conjugatedtrienes.Conjugateddieneswere seenin only some experiments
and then only in small amounts.The presenceof conjugated members was
recognizedthrough their characteristicabsorptionpeaksin ultra-violetanal-

-l
,tl

RADIATION

I
,t-1

30

E
an

c
o
F

"-*

"l
-l

uo']

t
I
I

'ol
'ol

too

rl

I
I
I
I
I
I
I
I
I
I
I
I

oo1

tn

t-'\

,
t
,
t
,
I
I

20
20)

.9

239

-aa

c
.9

t
a

t
t
t

320

300 ?30

240

220

200

Wove Length (my)

Frc. 80. Irradiation and desaturalion. Spectral analyses in ultraviolet (0.002% in
cthyl alcohol with alcohol as reference) of untreated sample (-)
and of the
radiated sample (. . . . ) both after alkaline isomerization.They show that the irradiation has induced also an increase in the amount of trienes present, which indicates
that a desaturation also occurred.

ysis. When the same mixture of fatty acids was treated by the usual
chemical methods ernployedto produce conjugation,i.e,, with potassium
hydroxide in ethylene glycol or glycerol ( 4l ), the spectral analysis
showed that the preparation contained fatty acids having between 2 and 6

240

RESEARCH TN PHYSIOPATHOLOGY

T,rsle XIV

Fatty Acid

Type of
Radiation

Sourceof Radiation

Exposure
Time
(Days)

Oxalic
Acid
mg/E
Fatty
Acid

6
8
l5
4

0
8.3
9.9
13.3
9.85

7
27

4.3
6.8

53

12.4

5
8

0
5.2
8.0

Linoleicacid
tt

tt

tt
t

tt
r

tt

gamma
gamma
gamma
beta
alpha
x-ray

tt

x-ray

50 mg Radium
120 mg Radium
120 mg Radium
25 mg Radium
in Monel metal
150 uc Thorium X
5000r daily--deep
therapymachine
5000r daily--{eep
therapymachine

Cottonseedoil
t
tl

t
tt

gamma
gamma

80 mg Radium
80 mg Radium

doublebonds.Figure 8l showsthe curvesof spectralanalysisfor such an

experimentin which 3 cc. of a cod liver oil fatty acid preparation were
treated for six days with 100 milligrams of radium filtered through platinum. In curyo "a," of the untreatedsample,it can be seenthat there is no
absorption due to the presenceof conjugatedmembers.Curve "b," for the
irradiated fraction, shows the typical conjugated rrienes, while Fig. 82
showsthe result of the chemicalconjugationof the nonirradiatedpreparation with membershaving from 2 to 6 double bonds.
Trst-e XV
Errecrs or Inn,rnrrrloxUpoNrHe Qurxrlry oF Oxrlrc Acro
PneseNrArten Oxto,rtrveFrssroN

Fatty Acid

Linoleic Acid
l0 cc.

l0 cc.

Exposure
time:
Days

Oxalic
Acid
mg g
Fatty
Acids

0
4
6
8

0
4.5
6.2
9.9

tt

15
20

r3.3
t4.4

1 5 0m g R a
,,

6
15

8.2
16.3

Sourceof
Radiation
0
1 2 0m g R a

RADIATION

241

IO

20

30
c
o

10

(a)

tn
.n
E

50

v,
E
.tt
t-

60
ia

70

I rradiated
Noninradlated

80

s
28o
260
i t a v e L e n g t h

2,to
(D!)

220

Frc. tl. Spectral analysis (0.002Vo in alcohol) of a mixture of. latty acids lrom cod
with 100 mg platinum
l i v e r o i l , u n t r e a t e d( . . . . ) ( a ) a n d i r r a d i a t e d( b ) ( - )
filtered radium/3 cc, at room temperature for 6 days. The analysis shows that the
conjugation which takcs place leads to the appearanceonly of conjugated trienes,
in spite of tbe presence of di-, tri-, tetra-, penta- and hexaenic unsaturated fatty
acids as shown by the absorption spectrum of the same mixture after chemical conj u g a t i o n w i t h p o t a s s i u mh y d r o x i d e i n e t h y l e n eg l y c o l . ( c ) a s s e e n i n F i g . 8 2 .

c) The changesinduced in fatty acids are essentiallythe same regardless of radioactive source. Thus the effect upon a linoleic acid preparation
containingsomelinolenicacid was the samewith alpha particlesof Thorium
X, beta rays from radium in monel metal, gamma radiation from platinum

242

xESEARcH IN

PHYSIoPATHoLocY

by a 400 kw. therapyunit. Figures

fi.ltered
radium,and X-rays generated
83 and 84 and TableXIV showtheseresults.
Irradiotion in vivo
d) Irradiation of the body of normal animals killed by decapitation,
producessmall amountsof conjugatedfatty acids.
under ether anaesthesia,
In early experiments,different organs obtained from slaughterhouseswere
irradiated and fatty acids examined. In general,even after intensive radia-

30

40

50
C

.9 -^

.3bu

c
o

Fzo
)
80
90

400

380

36U.

340

320
300
28c
WoveLength(mp)

260

240

220

F r c . 8 2 . C u r v e o f s p e c t r a l a n a l y s i sof the cod liver oil fatty acids after chemical

conjugation, shows the presenceof d i - , t r i - , t e t r a - , p e n t a - , a n d h e x a e n i c m e m b e r s .

tion, correspondingto 4,000 r. in one treatment,the oxalic index of fatty

acidswas neverfound to be above l, correspondingto I mg. of oxalic aciJ
obtainedfrom one gram of fatty acids.
e) On the other hand, the amount of conjugated fatty acids in the
bodies of living animals receiving radiation increases significantly. The
following experimentis illustrative.Eighty rats of the same sex, age and
weight(about 180 gms.), separated
into severalgroups,weregiven 1500r..
delivered by a therapy unit with no fi.lter.Four control animals were killed

243

RADIATION

l0

20

30

/r'\'.

f .,

I '

c
o

.a

40

att

/,'

l t

\'.\

\'..i
t v

t
t
,

ah
t

E
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)
,
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I

C'

I
t
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r-

60
t

}Q

m
I

l i
I r

80

l i
I t

t^'

Ir r a d i a t e d
llonirradiated

i
90

---.-'2
oo

2qo

U a v e l e n g t h ( m p )
Fro. 83. Absorption spectra (O.OO2Voin ethyl alcohol/ethyl alcohol) in ultraviolet
o f c o m m e r c i a l l ya v a i l a b l el i n o l e i c a c i d ( w i t h s m a l l a m o u n t o f l i n o l e n i c a c i d ) n o n with beta particles from 25 mg monel metal
irradiated ( . . . . ) and irradiated (-;
filtered radium/10 cc at 37'C for four bours. Some conjugation occurs in thc control
when kept in the incubator,

raiff$.r,4,{a{tfr
{iffi#(ffi'}*$itl's i#t1'

244

nESEARcH rN pHysropATHoLocy

beforeexposure.Groups of four treatedanimalswere sacrificedperiodically, startingimmediatelyafter irradiation,at 2, 6 and 24 hours aftcr irradiation,and eachday thereafteruntil all animalsdied or had beenkilled.
During this time, nontreatedcontrol animals,kept under the same
conditions,were also sacrificed.The quantity of conjugatedfatty acids

c
o

e
c
C'
L

>e

f a v e l e n g t h ( z r A )
F r c . E 4 . A b s o r p t i o n s p e c t r u m o f t h e s a m e m a t e r i a l a s i n F i g . E3, non-irradiated
( . . . . ) a n d i r r a d i a t c dw i t h o l p h a p a r t i c l e sf r o m 1 5 0 m c T h o r i u m X / 1 0 c c a t r o o m
t e m p e r a t u r ef o r 7 d a Y s . ( - )

RADr^rtoN

245

found in the entire body of each individualanimal at rhe time of death

was determinedby meansof the oxalic acid index method.
The oxalic acid index for fatty acids in the bodiesof untreatedcontrol
animalswas usually zero.occasionallythere was a variation from 0 but it
was always below 0.6. Irradiated animals killed within the first two days
showedan irregular increasein conjugatedfatty acids, with oxalic acid
indexvaluesof between0.6 and 5.1. Threedaysafter irradiation,the oxalic
index was above 3 in all the dead or sacrificedanimals. Thc index rosc

*
r

xt
X

r r r
l

,
x

8
r.ft
:

x
c

;
.!
x
o

I
E

(t
L

Ftc. E5. Futty acid conjugution indttted hy-irradiation in vir'<t.Changes in the oxalic index of total fatty acids of rats irradiated with a letlrcl dose (1500r). Sacrificed
at different intervals, the oxalic index of their fatty acid shows progressivelyincreasi n g v a l u e s ,T h e a n i m a l sd i e w h e n t h e i n d e x h a s a r r i v e d a t a c r i t i c a l v a l u e b e t w c e n l 4
and 17.

246

npsEARcH tN pHysropATHoLocy

even in animalswhich showedno visiblei.[l eftectsat the time they were

killed.
The index increasedcontinuouslywith the passageof time until the
animalsdied. By the fifth day, it was above6 for all animalsand, by the
seventhday, with few exceptions,it was around 10. After the twelfth day,
it had risen above 12 rn most of the animals.In all animalswhich were

oigl&
I ndex

7 days 12,2

!o
I

1.4
4.L
3.t
c
o { o
6

o
c
6

Eao
la

Ncrnal

3 hrs.

3,2

7o

Eo

Joo

"o ,.r. Llf,itr.,(rA'"

22o

F t c . E 6 . S p e c t r a l a n a l y s i s ( . 0 1 i n e t h y l a l c o h o )l o f t h e t o t a l f a t t y a c i d s o b t a i n e d
from tbe body of rats irradiated with 1500r shows changes more manifcst around
270 mp. The oxalic index of the preparation is indicated and shows a parallel increase
with the changesin the curve.

RADtATTON

247

sacrificedafter the l3th day or which had died at any time, the index
showedvaluesbetweenl4 and 17. (Note 2/ Figure 85 showsthescresults
in the group of rats describedin this experiment.These changeswere observedwhen the same procedurewas repeatedin other goups of animals.
These experimentsclearly indicated that the quantity of conjugatedfatty
acids progressivelyincreasesin the days following the exposurein animals
lo

[\.*'

,"-

'iiradtated

o
an
o
e
-q

,O

E
(!
L

'a

Eo

Jto

tto

.t?o

frl"ue
tenqffir1l

,r<

Frc. 87. Spectral analysis (.01 in ethyl alcohol) of the fatty acids of the lotal body
of a mouse irradiated with l50Or, shows an increasein fatty acids with the absorption
correspondingto 270 mp., as compared with the control.

treated with one lethal dose of X-ray. Death occurred when the amount of
conjugatedfatty acid reacheda critical level equivalentto an oxalic acid
index value betweenl4 and 17. The spectralanalysisof fatty acidsof animals treatedwith radiation showedchangescorrespondingto the presence
of conjugatedisomers.These appearin the samplesof fatty acids obtained
from the entire body of theseanimals.(Figs. 86 and 87l Still more evident
werethe conjugatedtrienesin the fatty acidsof organs.Figures 88, 89 and
90 show the differencein such analysesas comparedto correspondingun.

248

xESEARcH rN pHysropATHoLocy

treatedcontrols.The presence
of conjugated
trienesappearsclearlyin the
characteristic
peaks.
The concept of a critical value for the oxalic index is supportedby
other studiesin which the same value is found in animalsdying after
adrenalectomyor after thermal, chemicalor traumatic statesof shock.
Even in animalsdying in superacuteshock,within 3 to 5 minutesafter
beingseverelyscaldedin hot water,the level of conjugatedfatty acidsis
higherthan in controls.

2c)-+-

0 . 0 1 2t n e t h y l a l c o h o l / e i h yal l c o h o l
Imariiated

,o

(o

c
o
q
vl
c

E6o
|9
t

Da

70

9o

,.o

3@

2to

waritnothr#Ft

22o

F r c . 8 8 . S p e c t r a la n a l y s e s( . 0 1 i n e t h y l a l c o h o l ) o f t h e f a t t y a c i d s o f t h e k i d n e y o f a
normal rat and of a rat irradiated 6 days previously with 1500r. The peaks characteristic for conjugated trienes are seen.

R^DrArtoN

249

when the irradiated

dosewasnot a lethalone,that is, below600 r. in
our experiments,
the oxalic index increasedat fint but decreased
after
about two to three weeks.It neverreachedthe critical value of. 14-17.
( F i g.9 l ,N o te3 )
Local Efiecls
f) We completedthe studiesof the eftectsof radiationupon fatty acids
in vivo by considering
them at the localtissuelevel.The first requirement
20

0 . 0 1 2i n e t h y l a l c o h o l / e t h yall c o h o l

- lmadiated

Jo

t
a
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"?.r. 1.61fr
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220

Frc. 89. The spcctral analyses (.01 in ethyl alcohol) of the fatty acids of the liver
of a normal rat and of a rat irradiated 6 days previously with 1500r shows the appearaoceof the cbaracteristicpeaks of conjugated trienes,

xEsEARcH rN pHysropATHoLocy

250 /

was to establisha radiation procedurewhich would producea standardized lesion.When radiationswereapplieddirectlyto the skin of animals,
the individualdifferencesin responsewere quite marked.Thesecould be
explainedin part on the basisof ageand particularlyof sex,the difference

,o

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,l r r adiated

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F t c . 9 0 . S h o w s t h e s p e c t r a la n a l y s e s( 0 . 1 i n e t h y l a l c o h o l ) o f t h e f a t t y a c i d s o f t h e
lung of a normal rat and of a rat irradiated with 150016 days previously. Pcaks correspondingto coDjugaledtrienes are present.

betweenthe skin of male and female rats being manifest.However, there

were also pronouncedindividualdifferencesin animalsof the samesex, age
and weight living under the sameconditions,so that even when the experimental procedurewas carefully controlled,the same arnount of radiation
causedreactionsthat varied widely from slight erythemato ulceration.

RADIATION

,/

251

The problem of variability was satisfactorilyresolvedby radiating abnormal tissues,such as thoseof a wound, insteadof normal tissues.Standardizedlesionswere first producedand then irradiated.we usedthe following technique:an area of the skin on the back of male ratsweighingaround
200 grams was epilatedand, under ether anaesthesia,
a 2 cm.long incision

a
c)
E
q,

(o

P
L
r

'

x
T

Ccntrol

'H.r.

,
2
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Ftc. 91. Fatry acid coniugation and irradiotion in yiv'o. The changes in the oxalic
i n d e x o f t o t a l f a t t y a c i d s o f r a t s s u b m i t t e dt o s u b l e t h a li r r a d i a t i o n ( 6 0 0 r . ) . O n l y a
temporary increasein the oxalic index of the fatty acids of the animats is seen, the
a m o u n l s n o t r e a c h i n gt h e c r i t i c a l v a l u e s .

was made, penetratingthe skin and subcu'raneous

tissuesdown to the
dorsal aponeurosis.A needlecontainingradium was then placed between
the lips of the cutaneouswound. A thread passedthrough the hole at one
end of the needlewas used to fix it to the skin. Two retainingsutureswere
also used to maintain the radium needlebetweenthe lips of the wound,
The needlewas left in place for the desiredlength of time and then easily

252

R E s E A R C Hr N p H y s r o p A T H o L o c y

removedwith the help of the thread passedthrough the hole of the needle.
The retaining sutureswere also removed and the wound left open and undressed.
The length of time that the needle was left in place varied with the
amount of radium, the nature of the filtering metal, and the radiation burn
desired.we found that l0 mg. of platinum filteredradium had to be left
in place for about 90 houn in order to produce a standardizedulceration
that would last about four weeksbefore healing.The same effect was obtained when 25 mg. of monel metal filtered radium was kept betweenthe
lips of the wound for only two hours. when monel metal needleswere
used for only one hour, too great difterencesappearedbetween the ulcerations obtainedand the time necessaryfor their healing.A two-hour exposure causedan ulceration which usually required 4 to 5 weeks to heal in
control animals.lf the needlewas left in place for 3 hours, the ulcerations
were quite uniform but they required over two months to heal and more
than half of the wounds never healed.Failure to heal and more extensive
necrosisresultedfor periodsof exposurebeyond 3 hours.
Therefore,we utilized l0 mg. of radium in platinum for 90 hours in
one group of experiments,and 25 mg. of radium in monel metal for 2
hours in another group, in order to produce standardizcdulcerationsthat
would generally heal spontaneouslyafter 4 to 5 weeks.This technique has
been used in several hundred animals for various experiments. The fatty
acids of thesestandardizedradiation lesionswere studied.
Days after irradiation, the ulceratedlesionswere removed along with
about one cm. of surroundingtissueand submittedto analyses.It was always necessaryto use as many as 5 or 6 lesionsto obtain the quantity of
fatty acids needed for an oxalic index determination.The lesions werc
found abnormally rich in conjugatedfatty acids. Commonly, indices as
high as 4o-and in exceptionalcasesas high as 65-were found (Trnle
XVI) in comparisonwith 0 or 0.3 for normal skin with its subcutaneous
tissues.
Lipids and Radiation Burns
S) T'1t" apparanceof conjugatedfatty acids as an effect of radiation
has posed the problem of the role of theseabnormal fatty acids as intermediary agentsin the biological changesinduced by radiation. In trying
to solve it, we compared the eftectsobtained by administrationof conjugated fatty acids with those of radiation at different levelsof organization.
This study was facilitatedby consideringthe changeswhich take place in

R A D I A T I ( ) N

2s3

Trnle XVI
Ox,rlrc lNoex on Frrry Actos or Reorrrrox Bunxs
ElapsedTime

NormalSkin
Non-treated
wound

Woundwith 25 mg. radiumin

monelmetalfor 2 hours

24 hours
48 hours
7 2 hours
6 days
2 hours
24 hours
4 8 hours
4 days
I week
')

weeks

3 weeks

Average
0.1
2.2
3.9
2.3
1.8
t.7
6.1
l3.9
l9.1
31.0
46.0
49.4

the cellular cytoplasm and nuclei as induced by various substancesdesignated as radiomimeticagents.

It could be seenthat apparentlyall agentswhich induce radiomimetic
effectsare lipoids with negativepolar groups.The effectsof higher polyunsaturatedfatty acids, and especiallythe conjugatedisomers,appar to be
the same as thoseof known radiomimeticagents.The similarity between
the effectsof these fatty acids and those of radiation makes it logicd to
consider that at least some of the radiation-inducedchangesresult from
the interventionof theseabnorma.lfatty acids,
We have seenthat the changesinducedby fatty acids upon cell metabolism are in large part due to an increasein cell membranepermeability.
A similar change of cell membrane permeabi.litycould be recognized
among the eftectsof radiation. Following radiation, it could be seenthat
sodium of the interstitialfluids penetratesinto the cells more readily. This
was observedwhen radioactivesodium was used. (42) The cellular vacuolization seen to follow radiation, especiallyhigher doses, representsa
corollary of the abnormal penetrationof sodium into the cells which partty
resultsfrom the increasein membranepermeability,
h) At the tissularlcvel,the influenceexercisedby radiationupon pain
was seento $eatly resemblethat inducedby administrationof fatty acids.
Radiationefficientlyrelievespain that has an acid patternbut it may increasepain of an alkaline pattern.Furthermore,pain which appearsfollowing radiationhas an alkalinepatternand consequently
is increasedby
further radiation, or administrationof unsaturatedfatty acids. (Note 4)
i) At the tissularlevel,it could be seenthat the area of ulcerationof
the standard lesions obtained throueh irradiation of skin wounds was

254

REsEARcH rN pHyslopATHoLocy

increasedby the administrationof polyunssluratedfatty acids in general.

In somecasesthe ulcerationdoubled in size as comparedto controls. The
administrationof fatty acids also markedly delayedwound healing.when
the quantity of fatty acids administeredwas great enough,the wounds did
not heal at all. Six daily subcutaneous
injections,each of I cc. of a l\vo
oily solutionof cod liver oil fatty acids,preventedhealing.(FiS.92) The
area of ulceration was even greaterwhen only rA c*. of. a ljVo solution
of the conjugatedfatty acid isomers,obtained through an in-vitro conjugation of the preparation of cod liver oil fatty acids, was administered
under the sameconditions.This showedthat conjugatedfatty acid isomers
had a more manifest effect upon radiation wounds than the unconjugated
acidsobtained from the samesource.
j) we followed effectsof intensiveradio and radium therapy in humans
at organic levels. In caseswith radiation-inducedproctitis, mucositis, or
epidermitis,the changesobservedwere seen to correspondto the pattern
encounteredwith fatty acid predominance,
and especiallyto the pattern
induced by abnormal fatty acids.The appearanceof oxidizing substances
in the urine is frequently observedin patients with radiation burns after
extensiveX-ray therapy.They were almostconsistentlyseenin those cases
in which radiationlesionswere produced.The administrationof fatty acids.
and particularly of conjugatedfatty acids, to these patients increasedthe
intensityof the lesions.
k) Systemic changes induced by intensive radiotherapy were also
analyzed.Here again, the changesfollowed the pattern observed when
there is a predominanceof fatty acids,particularlyof abnormalfatty acids.
The appearanceof oxidizing substancesin the urine was frequently noted
after intensiveX-ray therapyand, as mentionedpreviously,was consistently
observedin those casesin which a radiation lesion was produced. Other
systemiceffectsof intensiveradiotherapywere seento include an increase
in urinary cxcretion of surfaceactive substances,
an increaseof potassium
in serum,a retentionof chloridesand water, and particularly,an increase
in the sulfhydryl index indicating an exaggeratedexcretion of this group.
Thesechangcsfollowing intensiveradiation are, as previouslynoted, similar to thoseseenwhen a predominantinterventionof fatty acids occurs.
Certain of these changesappear to have prognostic importance for
radiationtherapy.For example,in severalcaseswith very low urinary surface tension,high retentionof chloridesand absenceof urinary peroxides.
The continuationof irradiationled to death. (Note 4/ This is consistent
with the findingsin animalsthat lethal effectsof inadiation are directly re-

BADIATION

255

,,.,
,;,]]:,,i.

u
C

Ftc. 92. Lipids anct radiatian *'ounds. Radiation rvounds 5 weeks aftcr exposure to
l0 rngr. radium in platinum-for 96 hours. (a) Untrcated conrols: (b) treatcd tlnily
with I cc of a cod liver oil fatty acids l0{?. solution; {c) treated daily with 0.5 cc of
a 109* rolution of unsaponifiable lipid fraction cxtracted from human placcnta. The
lrcatmenl with fatty acids results in larger lesionsthan in controls, with no tendency
to hesl. Thc treatmcnt with the unsaponifiablc fraction lcads to a hcaling of the lesion
in rround threc wocks.

256

*ESEARCHrN pHysropATHoLocy

lated to the appearanceof large amountsof surfaceactivesubstancesin the

urine.
ROLE OF ANTI.FATTY.ACIDS
The study of the biological effectsof radiation also has revealedan
important role for anti-fatty-acid agents. The intervention of these substancesin the physiopathologicalprocessesthat occur in the organism
under the influenceof radiation can be consideredto be reactional.They
correspondto a responseof the organismto changesinduced by radiation
upon the constituentssuch as fatty acids.
This antagonismis clearly shown in experimentswith animals. The
administrationof sterol preparationsnot only reduccsthe size of ulceration
in standardskin radiation lesionsbut also significantlyimproves the rate
of healing. Insaponifiablefraction preparationsfrom human placenta, beef
liver, spleen or blood, as well as from butter, produce such favorable
effects.(Fie. 92)
If sterols are administered24 hours after the radium is inserted (or
later), the influence upon the dimensionsof the ulcer that develops is
reducedand is further reducedwith increaseddclay. In some of the animals treated with I cc. of a 5% oily solution of the insaponifiablefraction
of human placenta in sesameoil for seven days a week, healing with a
normal scar was complete within two weeks. Controls, treated with the
oil vehicle only, required an averageof more than four weeks to heal.
similar eftects were obtained with the administrationof I cc. of. a 7vo
solutionof butanol in salinetwice a day, beginningwith the day of radium
application.
The useof small amountsof radiationhas,in general,a differenteffect
to that of intensiveradiationfrom this point of view.This can be attributed
to the reactiveinterventionof anti-fatty acids.An exaggeratedscar-forming
effect,prolongedfibroblasticreaction,exaggerated
connectivetissueformation, and vascularsclerosisand thrombosisresultingfrom endothelialproliferation are all part of this long-term responseto moderate amounts of
radiation.The same effectsare producedby anti-fatty acid preparations.
All the manifestationsare opposite to those obtained with high doses of
radiationor fatty acids.
From a clinical point of view. the administrationof the insaponifiable
fractionpreparationhad a beneficialradiationeffect.Even in lesionsthat
had persisted
for years,the pain was observedto disappearafter a few days
with t.i.d. dosesas low as I cc. of a 59t solution of the insaponifiable

R^DrArroN /

257

fraction of placenta in oil. In severalcases,chronic lesions three to five

years old healedin only a few months of treatment.
l) The oppositeclinical responseto high or low doseswas frequently
observedin the systemicchangesin patientsreceivingX-ray therapy. While
high dosesled to a manifest lowering of the surfacetension of urine and
an increaseof the sulfhydryl index, togetherwith the other changes@rresponding to offbalanceD for small doses of radiation, certain opposite
effectsrelated to a predominanceof sterolswere noted. Of particular interestwas the absenceof oxidizing substances
in the urine, and the changes
in urinary surfacetension. In all casestreated,a first reactionto radiation
was a higher sulfhydryl index and low surfacetension correspondingto a
fatty acid predominance.When small or moderate amounts of radiation
were used, this reaction was very slight and rapidly disappearedin favor
of a secondchangecorrespondingto a predominanceof sterols with high
urinary surface tension, for instance. It is interestingto note at this point
that this secondaryresponsehas been observedespeciallyin those patients
for whom radiation also has had a limited therapeuticeffect. As we will
see, the dualistic interpretationof data furnished by urinary analysis in
patientsundergoingradiationtherapycan be usedto guide this therapy.
Role ol Adrenols
m) The study of the systemicsecondaryanti-fatty acid responseto
radiation has led to an evaluationof the interventionof the different antifatty acid agentsand to the role of the adrenals.It is known that adrenal
hormoneshave a peculiar effect upon the lymphatic system.They induce
a shrinkageof the thymus, spleen,and lymph nodes, along with blood
lymphopenia.Sincesimilar effectsare producedby irradiation,the problem
of the part played by an interventionof these adrenal hormones in the
radiation responseis of interest.
When the adrenalswere removed,shrinkageof the thymus and spleen
and lymphopeniastill occurred after radiation,but was markedly reduced
as compa.redto nonadrenalectomized
irradiated controls. Since shielding
of the adrenalsduring irradiationdoesnot alter the effectupon the lymphocytes and lymphatic organs,the role of the adrenalsappearsto be an indirect one. Adrenal hormonal secretionappearsthus to be a responseto
the systemicchangesinduced by inadiation. Adrcnalectomywould eliminate this secretionand thereby diminish the degreeof lymphopenia and
the involution of lymphatic organs.However, the secretiondoes not result
directly as an effect of irradiation of the adrenalssince shieldingdoes not
influenceit. Another factor seemsto intcrvenc to stimulate adrenal hor-

258

RBsEARcH rN pHysropATHoLocy

monal secretion. The difterences between the effects seen in adrenalectomized and nonadrenalectomizedinadiated animals correspondsthus to
the adrenal responseto the systemicchanges.
In experimentson rats, we have shown that polyunsaturated,and especially conjugated,fatty acids inducechangesin the number of lymphocytes
a short time after their administration,and that this is followed by involution of the thymus, spleenand lymph nodes.This seemsto occur through
the intervention of the adrenals since it takes place to a greatly reduced
degreewhen the sameamountsof fatty acidsare administeredto adrenalectomized animals.The abnormal fatty acids seem to influencethe adrenals
and their responseelicits lymphopeniaand involution of lymphatic organs.
However, this indirect action through the adrenal glands is only part of
the story. Large dosesof the samefatty acidswill directly induce a certain
amount of lymphopenia and involution of the lymphopoeticorgans since
these changesalso occur when these fatty acids are administered in large
dosesto adrenalectomized
animals.
n) All of this researchindicatesthat two of the mechanismsthrough
which radiation acts upon the organisminvolve changesin lipids. In one,
the action is directly through fatty acids; in the other, as a responseto
thesefatty acids, anti-fatty-acidagentsintervene.The role of the adrenals
appearsto be still more interestingconsideringthe nature of the fatty acids
produced by radiation. As seen above, the conjugatedtrienes appear almost specificallyas a result of irradiation of mixturesof fatty acids.It was
also seenthat the corticoidsintervenespecificallyagainsttheseconjugated
fatty acids.This correlationseemsto representthe link betweenradiation,
conjugatedfatty acids and the adrenalresponse.(Ch. 6, Note l7)
Direct Action ol Radiation
o) In spite of the importanceof fatty acids and anti-fatty acids, they
representonly one part of the mechanismthrough which radiation acts.
The direct and indirect action of radiationon other constituentsalso must
be considered.Thc influence exercisedupon these constituentscan be
largely relatedto variouschanges.There is a quantitativerelationship,for
instance,betweeninductionof mutationsand the direct impactof radiation
on proteins.Changesin fatty acids also are the result of such a direct impact. It appearedinterestingto ascertainhow much and which of the pathological changesthat follow irradiation are due to the direct impact upon
lipids and how much to the impacton the other constituents.
The three kinds of biologicalactivity of radiation-through other constituents,through changed fatty acids. and reactional through anti-fatt)'

R^DrArroN /

259

acids---couldbe studied at different levels of the organization. We note

here a few of the resultsof thesestudies.
Below the cellular level, the influenceof lipids seen$to decrease,causing the direct eftect of the radiation on other constituentsto appear predominant.For nucleo-proteinsand below them, only this last eftect seems
to occur, the changes induced apparently affecting histoncs and alkaline
amino acids. The close mathematicalrelationshipbetweenthe amount of
radiation and mutation would seemto indicatethat, even at the genelevel,
only the effect upon constituentshas to be considered.
The introduction of anti-fatty acids into the medium in which tetrahymenaor suspendedcells (as from ascitestumors) were irradiated,served
to distinguishthe direct effectsfrom those induced through fatty acids. In
the presenceof anti-fattyacids,vacuolizationand even changesin the nuclei
seenin the irradiatedcontrols are prevented.The fact that these changes,
which characterizethe radiomimetic effects,were reduccd by anti-fatty
acids, indicatesthe role of fatty acid changesin the pathogenesis
of these
effects.Among other agentstested,the insaponifiablefraction of organs,
and especiallyof placenta, appearedto be most effective in preventing
radiomimeticeffects.The high alcoholsor glycerolalso showedsuch influence,but to a lesserextent, In complex organisms,the diflerenceof
the effect of radiation on fatty acids and on other constituentsis increased
at the higher levels. At the systemiclevel, this effect is almost limited to
the fatty acids.
The introductionof polyunsaturated
fatty acids to the medium greatly
increasethe toxic effectsof radiation,as comparedto controls exposed
to radiation or fatty acids a.lone.The proportion of mutations was not
changed,however.
The ultimate effect of radiation at difterent levels dependsupon the
relationship between three factors: changed fatty acids, other changed
constituents,and the intcrventionof adrena.ls.
The effect of the adrenalsis
progressively
more manifestat the higher levelsof the organization.At
lower levels,the direct interventionof the abnormallipids becomesmore
important than the adrenalresponse,the latter being less able to act at
theselevels.At the cellularlevel,the influenceof lipids is still predominant.
At the tissularlevel, the direct lipid effect is still striking, while the influenceof the adrenalresponseis limited to the connectivetissue.Although
(as part of the adrenalresponse)
the effectupon the lymphaticconstituents
is important at the organic level, the steroid responsebecomesmore important at the systemiclevel.
p) On the other hand, it appearspossibleto vary the amount of the

260

xESEARcH rN pHysropATHoLoGy

lipidic effect by changing the nature of the radiation. The use of more
penetrating rays or of different corpuscleshas to be investigatedin terms
of the relationship betweeninfluenceupon fatty acids and the effect on
other constituents.It could be seenthat, in correspclnding
dosages,the less
penetratingradiations had a greater influence upon fatty acids than the
more penetrating.The fact explainsthe reductionof radiationburns directly
relatedto the interventionof fatty acids.Similarly,in a systemicprocedure,
such as teleradiotherapy,
the effecton other constituentsis reduc'edas compared with the direct influenceexertedupon the fatty acids.
It is possiblethat radiationsusing neutronswould induce an increascd
direct impact on other constituentswithout a correspondinglyincreased
effectupon the systemicfatty acids,The skin effect,which is minimal with
theseradiations,would indicatelittle interventionof fatty acids.
The unequalpart playedby lipids at the different levelscan be utilized
to obtain variationsin radiation eftects.If effectsupon the lowest levels of
the hierarchicorganization,such as upon histonesand basic amino acids,
are desired,radiation could very well be the tool to be chosen,becauseof
the small amountsof lipids prcsent at these levels. If the influencecould
be limited to such action, radiation could be consideredideal for such
therapeuticeftects. Unfortunately, this is not possible even when very
penetratingradiation is used, and the eftect of radiation upon lipids still
constitutesone of the principal factors which must be consideredwhen
radiationis usedas a therapeuticweapon.
Thus radiation is not the ideal meansfor affectingthe subchromosomal
level, in spite of the fact that it may, througlr its effect upon proteins,have
a profound influencebelow this level. Its ability to causea conjugationof
fatty acidsrepresentsthe seriousobstaclcto its use. In view of this. the
effectof radiation upon lipids actuallycan be consideredas an undesirable
epiphenomenonwheneverthe purposeof the therapy is to achievea local
effect at the lowcr levels. Frequently, the changeswhich require discontinuationof radiationtherapycan be recognizedto correspondto abnormal
local or systemicmetabolismproducedby the abnormalfatty acids.
It must, however,be recognizedthat the appearanceof abnormal fatty
acids has some advantageseven upon protein effects,since indirectly they
can make local tissuesmore sensitiveto radiation.We have previously
noted that abnormalfatty acids causcchangcsin the tissueand cellular
metabolismwhich lead to local alkalosis.This local pH changemay have
favorableresultsby acting upon the amphotericproteinsand by increasing
the positivelychargedmemberswhich apparentlyare the only ones sensi-

R^Dt^rtoN /

261

tive to radiation. Indirectly, the interventionof the abnormal fatty acids

will thus increasethe sensitivityof tissuesto radiation.
Before going further, we wanted to emphasizean aspect of the offbalanceD for which the studyof shockand radiationbroughtan important
contribution.A separationcan be made betweentwo phasesof offbalance
D. one in which oxygenis principallyfixed and anotherin which chlorine
is fixed. The first phase,"oxygen,"has as characteristic
the appearance
of
peroxidesin the urine, and clinicallyhas little noxiousmanifestation.
The
othcr phase,"chlorides,"with fixationof this ion leadsthe seriousmanifestationsas seen,for instance,in shock.For this reason,in radiationthe
disappearance
of urinary peroxideswith persistingoffba.lance
D, as seenin
the other patterns.will indicatea passage
"oxygen"
phase
from
into phase
"chlorides,"which correspondsto the appearanccof a seriouscondition.
(See also Note 4. Chapter l0)
Radio-Therapy
The above considerationsappear inrportant in the radiotherapy of
tumors. The tissular and systemicchangesrelated to the intervention of
fatty acids,especiallywhen thesechangesare sufficientlyintense,in themselvescan act upon tumors. However, when abnormallyintense,they can
constitutea seriouslimitationfor continuationof radiation.The manifestations that result from the pathogeniceflect of abnormal fatty acids, if intense, can prevent the use of large doses of radiation which would
otherwise be necessaryto influence a tumor through a direct effect upon
the lower levelsof the biologicalorganization,histones,nucleo-proteinsand
even genes.Consequently,the appearanceof abnormal fatty acids, which
represent an important factor in the biological effect of radiation, can be
consideredas a favorable effect when we seek to bring about systemic
changesand influencepain and metabolism,particularly at higher levels.
At the sametime, they can also representa principal obstacleto the more
effectiveuse of this same therapeuticagent when one wants to obtain an
effect at lower levels.
As for the effectsobtainedthrough the influenceexertedby fatty acids,
they can be decreasedby changingthe antagonisticrelationshipbetween
the abnormallipids and the defensemechanismof the adrenals.With small
amounts of radiation separatedby long intervals,the interventionof the
adrenals,as long as they function normally,can overcomethe effect of the
fatty acids. With higher dosesapplied more often, the fate of the irradiatcd individual dependsupon whatever antagonisticfactor predominates.
With high dosesor with a relativeadrenalinsufficiency,the direct effect of

262

xESEARcH rN pHysropATHoLocy

the abnormal fatty acids can becomepredominant.In that case,the type

D offbalancewill be more pronounced.It is in such offbalancethat subjects
die from too intensiveradiation.Thesefactors can be of major significance
in the intervention against accidental radiation as well as in guiding the
therapeuticuse of radiation.
Becauseof the interventionof abnormal fatty acids,systemicradiation
does not seemto be the best procedureunlessa very intensesystemiceffect
is sought. If this effect is desired,it can be obtained through a method
other than radiation.Furthermore,as we have noted above,the conjugating
effect of radiation upon fatty acids is almost entirely limited to the production of trienes and dienes.The biologicaleffectsof such conjugatedfatty
acidsare more apparentat the tissuelevel and aboveit. The energeticvalue
of conjugatedtrienesand dienesseemsto be too meagerto permit them to
act intensivelyat levelslower than the cellsor nuclei.
In order to have a manifestfatty acid effect, it appearsnecessaryto
have an adequateapplicationof radiation. since an exaggeratedsystemic
action of the abnormal fatty acidsmay even induce lethal eftects,radiation
does not appear to be the therapeuticmethod of choice for an influence
exercisedthrough fatty acids.Radiation,however,is more compatibleeven
with a desiredlocalizedeffect through the limitation of the field in which
the changesin fatty acids are induced.In this case,fatty acids may intervene with a lower systemicinfluence.This accountsfor the analgesicaction
of radiation which probably is relatedto an effect exercisedby local fatty
acids. Even here, however.the appearanceof an alkaline pattern of pain
can lead to undesiredchanges.In this case,radiationwill increasethe intensity of pain. TNs fact reducesthe indicationsfor use of radiation even
at the tissuelevel.
Biologically-Guided Radio-Therapy
The knowledgeof the important roles played by abnormal fatty acids
and anti-fatty acids in the biologicaleftectsof radiation has suggesteda
biologicalguide for radiotherapy.Urinalysis,by reflectingvarious systemic
changes,can serveas a valuableindicationof manifestationsand processes
presentin subjectsundergoingradiation. The persistenceof a pattern related to predominanceof fatty acids indicatesthat the patient has passed
into an imbalancethat can only be increasedby further irradiation and, if
it becomessufficientlyintense.may even prove to be lethal, causing the
patient to die with symptomsof severeshock. [n contrast,a pattern correspondingto the predominanceof sterolscould be consideredas being consistentwith a preponderantreactive response,which would indicate that

R^DIATION

263

higher amountsof radiationcould be usedwithout dangcr.From a practical

point of view, the information given by the urinary surface tension has
appearedvery valuable.The moment when, and the amount of, irradiation
to be given can be determinedby these analyses.A low surface tension
would contraindicateadministrationof radiation while high values would
indicatethat radiation should be increased.
The administrationof lipids or lipid-like substanceswould representa
method of controlling undesirableprocessesand a-llowingmore effective
useof radiation. If the reactiveinterventionof adrenalsappearstoo strong,
a lipoid with negativecharacter could be added to counteract this and.
consequently,could increasethe desirableeffect of the radiation. Subjects
receivingfatty acidsor sulfur preparationsalong with radiation have shown
intensivelocal effectswith very small dosesof radiation. Epidermitis and
mucositiswere seenin such patientseven with dosesas low as 600 r. The
sameintensiveeffectcould be seenin the tumors.The useof lipoids appears
indicatedwhen an intensiveeftect through fatty acids is sought,as in lymphatic tumors. On the other hand, if the effectof fatry acids is higher than
can be accepted,and representsa handicapfor the desiredeffect on proteins, then adrenal hormonesor other anti-fatty acids must be added. By
reducing the effect of abnormal fatty acids, it becomespossibleto obtain
a more intense impact on proteins and, at the same time, to avoid the
otherwiseinherent undesiredside effects.The choice of the anti-fatty-acid
agent must be guided by the level at which the effectsof the abnormal
fatty acids would make themselvesfelt. While corticoids act especially
upon systemic and organic levels, sterols and other positive lipoids act
upon the lower levels.Butanol and similar agentsare effectiveupon local
changes,such as pain.
The guidanceof radiationtherapy,as an exampleof how this new view
may be used to improve therapeuticapproaches,will be discussedlater.

C H A P T EI RI
P R O B L E M SI N C A N C E R

NT

I \ ew lNsrcHrs rNTo many pathologicalproblems-and those of cancer

in particular'-are oftered by the conceptswe have been discussing.Let us
take, for instance,the problem of just what cancer is. Classically,one is
entitled to speak of a condition as cancerous when cells with cancerous
character are present in the body. whether, on the one hand, only cancerin-situ cells arc identified or, on the other hand, the patient is dying and
has almost no organ or function left untouched-the condition is considered cancerous. Yet, so long as the concept of cancer is associatedimplicitly with the concept of malignancy, to consider clinically healthy
individuals to be cancerous only because of the presence of cells with
cancerousnuclear characters,when most of them will never show further
developmentof the disease,is entirely confusing.It is essentialto separate
the two concepts,the presenceof cancerousentities and actual malignancy.
The fact that the hierarchic levcls of the organism participate in the
various manifestationsof cancer puts the problem in its true light. A cancerouscondition does not implicitly mean malignancywhen it involves only
the prcsenceof an entity with cancerouscharacter. Other attributes must
bo considered.In the hierarchic progressionof cancer, malignancy begins
to be manifestedwhen the cellular level participatesand inducesinvasive
canoer. With malignancy an attribute of only some of its phases,cancer
can be seen to embrace many changes,beginning with those of the lowest
hierarchicentitiesand terminatingwith the systemiclethal condition.
The plurality of phasesof cancer,with the broad variationsin time and
other factors which de0erminethe passagefrom one phase to the other,
logically raises several immediate questions.
In view of the multiple phases,one cannot speak of pathogenesisof
of the different phases.Concancerin general,but rather of pathogenesis
264

PROBLEMS tN

CANCER

265

sequently,even postulatingthe existenceof somespecificoriginal cancerous

change,such a change would not, alone, induce the entire diseaseand
determine the passagethrough successivephases. Different pathogenic
factors must be consideredto intervenein order to have cancer passfrom
one phase into another. The evolution of the cancerouscondition has to
be relatedto thesedifferentfactors,someof them possiblymore important
than the original specificchange.The passageof a cancr from the noninvasiveto invasivephase,or from tissueto systemic,is surely more important than the appearanceof a low level cancerousentity. An original
change at a lower level appears,in fact, to be of very little importance, not
only becauseof its ubiquity but also becauseit is not implicitly related to
malignancy.From this point of view, then, cancerno longer can be defined
as some singlespecificchangein a cell, nucleus,chromosome,geneor other
biological entity.
has to be conceivedof in an entirely new way, in terms
Carcinogenesis
of plural factors and their relative values.Acceptingthe phasesabove invasive cancer as the only ones which correspondto clinical malignancy,
they have to be regardedas the end result of a seriesof cancerouschanges
developedat progressivelevels,with the interventionof many factors, not
just one.
Diagnostic Tests
This view puts the problem of diagnosisof cancer in a new light. The
recognitionof a cancerouscondition by itself, althoughimportant, has little
clinical meaning.The presenceof "cells with cancerouscharacteristics"in
the prostate of almost all men 40 years of age and older, and in the
thyroid, lung and stomach in a high proportion of the population, has
fai.ledto produce a general feeling of despaironly becausesuch findings
are commonplace.While they still mean cancer,they do not implicitly indicate malignantdisease.It appearsvery clear that a diagnosisof cancer
is incomplete without immediate qualificationas to its phase. We can no
longer speakof cancerwith any degreeof practicalmeaningunlesswe add
a descriptive adjective-noninvasive, invasive, tissular, organic or systemic.
And the searchfor a test to detectcancerwill have no meaningas long
as we have not definedin advancethe informationwe want. A test, biochemicalor immunological,which indicatesthe existenceeven of a specific
anomaly in the noninvasivephase or before, while interesting,wiU have
little significancesincethis anomalyexistsin so many subjectsand for most
doesnot go beyond the noninvasivephase.The test will not indicatemalig-

266

xesEARcH rN PHYsToPATHoLocY

nant cancer in the clinically frightening sense.On the other hand, the
processeswhich are addedto a noninvasivephaseform of cancr and turn
it into the invasive,tissue,organicor systemicphase,have no characterof
specificity. Similar growth changes,or the appearanceof lipidic predominance, which representadded factors are seen in many other conditions.
By using them for diagnostic purposeswe will not recognizn,the cancerous
condition but only nonspecificinterveningfactors. While these factors are
responsiblefor the changes,malignancydevelopsonly when, and because,
thesefactors operateon already abnormalentities,i.e., cancerousentities.
This explainsthe nonspecificityof many proposedtestsand the misleading
positive results obtained in conditions such as pregnancy where one of
theseadded factors, (active growth processes)is always present.
A test for cancer, to have clinical value, would have to indicate two
things: one, the specificearly changewhich is widely distributedbut representsthe cssentialcondition for the potential developmentof malignancy;
and, two, the concomitantprcsenceand concomitantoperationof the nonspecific factors which can cause the actual developmentof malignancy.
This kind of diagnostictest undoubtedlywill come from further systematized study of biochemicalchangesinducedby the simultaneousaction of
the two groups of factors.
Immunological studies reprcsentan approach of value for diagnosis.
The different phasesof cancer can be interpreted,in the final analysis,to
correspondlargely to the interventionof the defensemechanismat different stagesat the different levels.As mentionedabove, a changein a phase
resultsalso from a changein the defensestageat the respectivelevel. We
have seenthat the immunologicalaspectof cancer cannot be understood
without acceptinga relative independenceof the levels in their different
stagesof defense.This view explains somc seeminglyparadoxicaloccurrences.
Cancerouscells are frequently found circulating in the blood yet this
does not indicate generalizedcancer. While the organism defends itself
successfullyat thc systemiclevel againstcancerouscells, the cancer can
still progressat the lower level of the tissues.The loss at this Iow level of
an effectivedefense,principally primary or allergic,which is still persistent
at the systemiclevel, explainswhy the cancerouscells invade the tissues.
A test indicating the presenceor absenceof any immunologicalreaction
would consequentlyhave value only when related to hierarchic levels. It
must furnish indicationsonly of what is happeningat a specificlevel. The
nature of the immunologicalreaction in cancer is also different from the
reactionin other conditions.Defensecapacity-natural defensecapacity-

PROBLEMstN cANCER

267

at differentlevels is lost as the respectivelcvel participatesin the disease.

This is in distinction to the immunologicalpr(xessesin other conditions
in which the normal individual lacks specificimmune bodies.An immunologicaltest for cancerwould have to revealthe lossof a previouslyexisting
defensemechanismrather than the appearanceof an immunological response.This loss can be revealedin different ways. In one, the response
to a cancerousantigen is investigated,and its lack would indicate the
existenceof a cancerouscondition at this level.
In a study now in progress,we utilize pooled human tumoral tissuesas
antigen,and try to see if an allergicreactioncan be induced with it, in two
administrations,
sensitizingand trigger. Two intradermic injections at the
samesite are made 12 days apart. They induce an allergic reaction in
normal individuals, but are without eftect in patients with active malignancy.If the eftect is negative,a third injection is given l5 days later. A
similartest is made for the conjunctiva,with sensitizingand trigger instilIationsof a similar antigen.No allergic reactionindicatesa positive result,
whilea reaction is consideredto be normal.
Another test which we are studyingis basedon the same lack of efficientdefensemechanismat the tissuelevel. Such a lack of defensewould
permit a cancerousantigen to be present without the body offering a
sufrcientlyeffectivedefenseagainstit. The presenceof such an antigen in
the tissuesis revealedby inducing an allergic reaction, through the administrationof specificcoagulantantibodies.Scra of guinea pigs injected
with pooled human tumors and having a high precipitin content are used
in intradermicinjectionsor in eye instillation.An immediatereaction indicatesa positive result. As control, we use normal guinea pig sera. The
studiesare now in progressand the diagnosticvalue of thesetests will be
reportedin a later publication.
Circulating Cancer Cells and Surgery
These immunologicalconsiderationshave appearedimportant in consideringa problem related to the use of surgeryin cancer. The existence
of a veritable flow of cancerouscells in the general circulation, largely
inducedby the manipulationsinherent in operative procedures,has producedgrave doubts as to the value of the measurestaken by surgeonsto
preventlocal spread of cancerouscells through the surgical act itself.
Paradoxically,however, these precautionshave been followed by good
clinicalresults.Analysis from the point of view of the defensemechanism
involvedcan clearly explain this situation.In the invasivephase,the systemicdefensemechanismis stiU adequateto insure destructionof cancer-

268

RESE^RcH tN pHystop^THoLocy

ous cells which get into the blood. This is not true at the level of the
interstitial formations, that is, at the tissular level, where such defense
means are failing. The real danger during surgeryconsequentlyis not so
much the presenceof cancer cells in the blood, since the blood can still
take care of them, but the spreadof thesecells at the tissular level where
the defensecapacityhas beenlost. and where a cancerouscell consequently
has every chancenot only to remain alive but also to grow.
The independenceof the defensemechanismat different levels also
must be takeninto accountin explainingthe differences
in the eventswhich
follow the appearanceof a spontaneous
tumor and those which occur after
cxperimentaltumor transplantation.

EXPERIMENTAL

CARCINOGENESIS

The problem of carcinogenesis

apparsin a new light when cancer is consideredunder the conceptspresentedabove.Classically,the experimental
induction of cancer is judged successfulonly if the result is a tumor in
the invasivephase,that is, with abnormal cells invading normal surrounding tissues.This is consideredto correspondto a fundamental specific
changewhich transformsthe normal cells into cancerousones. The entire
diseaseis held to stem from the relationshipbetweenthese abnormal cells
and the organism. (29O, 291, 303) To these simple views of the abnormality, we have proposedanotherone.
In our view cancerrcpresents
a hierarchicallyorganizedcondition.Its
invasiveform is only one phasein a long sericsof changeswhich transforms
successive
hierarchicentiticsinto cancerousentities.Carcinogenesis,
thus.
is not simply a changeof a normal cell into a cancerousone but a step by
step progressivehierarchicdevelopment.A cell is cancerousonly if it has
a cancerousnucleusjust as a nucleusis cancerousonly if it is formed by
cancerouschromosomeswhich, in turn, are cancerousif they have cancerous genes.With the same reasoning,it is possibleto go far down in the
organization,below gcneseven to nucleo-proteinsor still lower to histones
or even alkalineamino acids,to find that the first changes,which can be
consideredto be specificfor cancer,take place at the bottom of the organization of the biological realm. In other words, a cell becomescancerous
after specificcancerouschangeshave occurred in all the hierarchicallyinferior entitiesthat composeit. Thus. a successfulexperimentallyinduced
cancer.i.e., one that is alreadyin the invasivephase,meansthat changes
would have affectedthe entire seriesof hierarchicentities,including the
cells,whoseparticipationresultsin the invasivecharacter.Seenunder this

i,ffi#l:*

PRoBLEMS rN c^NcER

269

aspect,carcinogenesis
no longer can be acccptedas a simple processoccurring in the cells, but must be regardedas a successionof organizcd
processes.
This becomesstill more interestingwhen it is realizedthat changesin
the constituentsat the lowest levels of the organizationcan occur on a
statisticalbasis,that is, independentlyof the direct interventionof external
agents.As thesechangeshave to be developedfor many successivehierarchic entities, it takes a certain time for them to be realized. This would
explain why most cancers appear after a certain age. Cells with cancerous
nuclei, i.e., ln the noninvasivephase,frequentlyare present,in older people, in many organs without producingclinical manifestations.Conceptually, in order for an agent to be considered a successfulcarcinogen, it
must act upon these noninvasive entities to such an extent as to change
them into invasiveones. It can thus act upon entities which have already
progressed,by themselves,far enough in the hierarchicdevelopmentof a
cancerous processand have arrived at the noninvasive phase without any
manifestation.The excessivelength of time necessary,even for the most
active agents,to induce invasive cancer would suggest,however, that more
than a simple passagefrom an already existing noninvasive cancer into
an invasiveone is involved. A plurality of changesmust be induced,some
or all at levels below the cell.
We are inclined to favor this last hypothesiswhich obliges us to consider that a carcinogeninduceschangesat differentlevelsof the organization. It is supportedby a seriesof facts. In addition to having the capacity
to induce invasive tumors, carcinogenic agents also induce precancerous
lesions which correspondto cancerousentities below the invasive phase.
Cells with abnormal nuclei or with only abnormal chromosomesare almost
constantly seen in induced carcinogenesis.
Even agents which produce a
high proportion of invasive cancer consistentlyinduce such changes at
Iower levels as well. For carcinogenswhich induce a low proportion of
invasive cancers,the effects often appear to stop at Iower levels. Such
activity at subnuclearlevels of the organizationis seen in the capacity of
most of the carcinogensto induce mutationsand monstrosities.
In the conceptof hierarchicorganization,mutations are consideredto
result from changestaking place at the gene level, with lower levels left
unaffected.Monstrositiesresult from changesat the chromosomelevel.
with mutationsand monstrositieshas led us
Comparisonof carcinogenesis
to consider that cancerouschangesbegin at levels much below those involved in mutations and monstrosities,possiblyat the nucleo-proteinlevel
or. even below. The complex cancerouscondition to which the invasive

270 /

x z s E A R c Hr N p H y s r o p A T H o L o c y

form correspondscan thus be seento be the result of a seriesof anomalies

which have taken place at differentlevelsbelow the tissular.Carcinogenesis
at the invasivephaseis conditionedby the existenceof changesat all the
lower hierarchiclevels.while they can appearas the result of the development of the organization, conceivably these changes can be hastened or
even inducedby the carcinogens.
The concept of multiple changesin carcinogenesishas caused us to
searchfor multiple factors in carcinogensthemselves.The possibilitythat
such factors might be found was suggestedby the existenceof so+alled
co-carcinogenicagents.These are substances
without carcinogenicactivity
of their own but capableof inducingsuch activity in casesin which some
carcinogensare administeredin dosestoo small to induce invasivecancers
by themselves.This peculiar intervention of co-carcinogenscan be explainedthrough the multiple factors in carcinogenesis.
It can be conceivedthat the factorspresentin a carcinogendo not have
equal activity. The differencesappear evident when the carcinogenis administeredin very sma.llamounts.While some factors still have sufficient
potency in these small amounts to accomplishtheir part in the complex
processof carcinogenesis,
others are quantitativelyinsufficientand do not
induce changes.The total eftect is an incompleteset of changes.Under
thesecircumstancesthe addition of a co-carcinogencan replacethe action
of the quantitativelyinadequatefactors,and consequentlycomplete the
plural action necessaryto produce an invasivecancer. Becauseany one
co-carcinogencan replaceonly certain factors, co-carcinogenactivity has
a certain specificity.
With the hypothesisof multiple actions in the same carcinogen,the
next step was to try to recognizethem. A study, identifyingdifferent active
energeticcenters in the structuresof carcinogens,has substantiatedthe
hypothesis.
We attempted,as a first step to systematizethe analysisof such energetic centersin carcinogens.A short resumeof this study is presentedhere.
Energetic Factors
A well known and generallyacceptedconcept tries to correlate carcinogenicactivity with the presenceof one energeticfactor. identifiedas a
"condensationof electrons,"at certain regionsof a moleculeand revealed
by the physicomathematicalapproach offered by Pulman and Dawdel.
were started
Studiesof the role of electrondistributionin carcinogenesis
by Otto Schmidt (43), which showedthat an electrondensityexceeding
0.44e/* in the meso region of the moleculeappearsnecessaryto confer

P R O B L E M Sr N C A N C E R

271

carcinogenicproperties.This conceptwas partially modified and amplified by Pulman,Dawdeland their co-workers(44) who have shown,by
quantumanalysisof variouscarcinogens,
that the densityof the a electrons
is increasedin certain preferredregionsof the molecules,the K regions.
They showedthat, whenelectrondensities
exceed1.292eat theseregions,
the substances
have carcinogenic
properties.Figure 95 showssuch a K
region.

o)

i-=
\z

\,2

CHs

9:l0 Dimethyl
| :2:7:8 Dibenzanthracene
Flc. 95. The regions K in carcinogenicmolecules.

From our point of view, a tentativelyinterestingaspctof this condensation of rr electronslies in two facts: the presencein some carcinogen
moleculesof more than one such K region, and the presenceof different
valuesfor theseK regionsin differentmolecules.It would be the presence
of more than one K region in the same molecule which would result in
interventionin more than one processand thus contributeto plural activity.
Further analyses,however,suggestedthat the condensationof z electrons in K regions would represent only one of the factors that would
induce activity in theseagents.We have identifiedanother energeticfactor
in the presenceof two atoms having the same electrical sign and being
bound togetherwithin the molecule.
Twin Formation
We have considered the existenceand importance of these "twin formations" as indicationsof energeticactivity in the course of studies on
electronicmoleculararrangements.In a molecule,an alternationof successiveatomsresultsin part from the alternatingpolarity of theseatomswithin
a molecule and in part from the opposite charactersconferred upon the

272

R E S E A R c Hr N p H y s r o p A T H o L o c y

two carbon atoms when they form acetic acid, an important precursor in
biologicalsyntheses.It is through alternatepolarity that an induction effect
of an energeticcenter in thc molecule propagatesitself along the chain.
The presenceof any energeticcenter in the moleculerepresentedby polar
groupsor a lateral chain, for instance,will enhancethis alternalepolarity.
When one or more such inductiveetTectsare propagatedthrough the chain,
two adjacentatoms may be found to possessthe same electrical sign for
their chargeor ionoid character.The twin formation which results representsa center of increasedmolccular reactivity. This reactivity can be so
intense as to lead to breaking down of the molecule, something which
occurs often in inorganic substances.
This has led Pauling to believe that
this condition,called "adjacentchargeru|e," cannot exist.
"Pauling has pointed out that the murual potential enerry of two electrical chargesof the samesign is so high that a canonical structurehaving
net residualchargesof the same sign on any adjacent atoms would have
too high an energy level to contribute appreciably to the real molecular
structure." So notes William A. Waters in "Physical Aspects of Organic
C h e m i s t r y . "( 4 5 )
The form suggestedfor nitrogenperoxide (N.On), (Fig. 96l would
appear to be impossiblebecauseof the high energy developedat the two
positive nitrogens.

NitrogenPeroxide
Frc. 96. The existencc of nitrogen peroxide molecule is prevented by the high energy developed at the two adjacent positive nitrogens.

However, the forces that exist in most of the organic molecules are
much weaker, so that the resulting"twin formations," although energetically potent. arc not strong enough to induce the breaking down of the
molecule.Consequently,they would exist and rePresentimportant energetic
centers.

P R O B L E M Sr N C A N C E R

273

we have studied a number of carcinogenicagents,seekingtwin formations. Analysisof the ionoid characterof the carbonsof the methylcholanthrene molecule revealsthe presenceof twin formations which could be
localizedat various points of the molecule.Figure 97 shows the energetic
aspectof methylcholanthrene
and the ionoid characterof its carbons.It is
the presenceof the cyclopentanegroup in the molecule that induces the
samesign in two adjacentcarbons.The presenceof the methyl group would
determine the electrical character of C2e and consequentlythe srccession
of alternate signs. On the other hand, the double bonds will determine the
probable localization of these twin formations in the molecule at the K
formation itself, that is, at C5 and C6.

Hsc

1)

20-Me
t hyI choI anthrene
Ftc. 97. The energeticaspectof methylcholanthrene.with twin formations.

Twin formations can be found in many carcinogens.It must be emphasized,however,that unequal energeticvaluescan be recognizedeasily
for difterent twin formations and would explain differencesin their activity, a fact which would confer possible plural properties upon this group
of qualitativelysimilar energeticformations.
Another aspect of the relationship between these formations and carcinogenesisappearsto be even more interesting.While no twin formations
can be found in severalagents,the formations are presentin the substances
resultingfrom metabolism of these agentsin the body. The relationship of
twin formation to carcinogenicactivity can be suspectedwhen such changes
appear simultaneouslywith carcinogenicity.
For example,no twin formation occurs in 2-naphthylamine,(Fis. 98)
whosedirect carcinogenicityis questioned.but such a formation appearsin
heterocyclic3:4:5:6 dibenzcarbazole,
one of its intermediates(46), which
is known for its carcinogenicproperties.(Fig. 98bis) This is also true for

274

nESEARcH rN pHysropATHoLooy

NHz

(a)

2-Napht
ylamine
Ftc. 98. No twin formations exist in 2-naphthylamine.

aminofluorcne,which is also relatedto 2-naphthylamine.(Fig. 99) The

existenceof a twin positive carbon group or a twin negativein the same
molecule can further explain the diversity of the tumors produced by this
carcinogenand its acetyl derivative,which has the s:uneenergeticpicture.
( 4 7 , 4 9 , 4 9 ,5 0 , 5 1 , 5 2 )

(b)

3:4:5:6 Dibenzcarbazole
F r c . g E b i s .A t w i n f o r m a t i o n a p p e a r si n t h e i n t e r m e d i a t e3 : 4 : 5 : 6 d i b e n z c a r b a z o l e .

Twin carbons can be correlated with the degree of carcinogenicity of

the sulfur isosters(53) in each of which a thiophenenucleusreplacesthe
This also appliesto the
benzoneringof 9:10 dimethyll:2 benzanthracene.
:rzo compounds with twin formation at the level of the azo bond. Figure
100 showsthe presenceof a twin nitrogen at the level of the azo bond, due
to the influence exerted by the symmetric rings.

PROBLEMS IN

CANCER

275

NHz

Z-Aninofluorene
F l c . 9 9 . A t w i n c a r b o n g r o u p i s p r e s e n ti n a m i n o f l u o r e n e .

Furthermore,it is the relationshipof twin formation to carcinogenicity

which indicatesthe need for consideringthe metabolismof various carcinogensin the organism.
Dimethylamino-azobenzene,
butter yellow, which has a twin formation and is an active carcinogen,can becomestill more active through
the metabolicchangcsoccurringin the body which lead to productswith
trvin carbons.The sanrc2:2'-azonaphthalene,
with a twin formation, becomes more active becauseof its transformationinto amines passing
throughhydrasecontpounds.2:2'-diamino-l: l'dinaphthyl, with twin car(-t

(-)

cHg

N: l'l

cHg
4

Dimethylamino
azobenzene

F l c . 1 0 0 . A t w i n f o r m a t i o n i s p r e s e n ti n 4 - d i m e t h y l a m i n o - a z o b e n z eantet h e l e v e l o f
the azo bond.

bon formation, is more active than the precursor,2:2 Azonaphthalene.

( 5 4 ) , ( F i s s .l 0 l a n d 1 0 2 )
It is possiblethat benzidinerearrangements
of the hydrazo dcrivative
determinetwin formation and thus explain its carcinogeneity.
The similarity in kinds of tumors produced by the derivativesof 4(Fig. 103), and the aminofluorene
aminostilbene
derivatives(55), makes
us think that twin formations can appear in this case through changes
occurringin the organism.

ti.,v:.Si:"Sffi&Sl

276

n E S E A R c Ht N p H y s r o p A T H o L o c y

<\
N

(+)

(+)

2:2' Azonaphthalene
Frc. l0l . 2:2' Azotaphtbalenehas only a slight activity.

Some artificial estrogensof high potency (56) diethylstilbestroland

triphenylethylenic
acid, ( 57), (Figs. 104 and 105) are known to have carcinogenicactivity. Whilc a twin carbon is present in both, such a formation is assumedto appearmore activein the latter,as the result of metabolic changesin the body.

2:2'
l: I'

Di a mni o
dinaohthyl

F r c . 1 0 2 . T h e p a s s a g eo f . 2 : 2 ' a z o n a p h t b a l e n ci n t o t h e a c t i v e 2 : 2 ' - d i a m i n o l : l ' - d i n a p h t h y l r e s u l t s i n t h e a p p e a r a n c eo f a n a c t i v e t w i n c a r b o n f o r m a t i o n d u e t o t h e

influencc excrted by the amino-group.

An interestingaspectis furnishedby urethaneand other estersof carbamic acid.Figure 106 showsthat no twin formationscan be seendirectly
or through a changein the molecule.This accordswith these substances'
lack of capacity,noted by many authors,to induce cancerouslesionsor
e v e nt u m o r s .( 5 8 , 5 9 , 6 0 ) O n ( 6 1 ) r e l a t e sl e s i o n sp r o d u c e db y c a r b a m i c

PROBLEMS tN

CANCER

277

acid esten to chronic inflammations,noting their regressionwhen treatment is discontinued,(Note l1

From analysesof the substancesable to induce invasivecancers,it can
be observed that many present a twin carbon or nitrogen formation, usually activatedby the inductionexertedby a polar group or by double bonds.
Some of these substancesoriginally without twin formation become carcinogensonly when changesoccur in the body leadingto the appearance
of a twin formation.

/C",,,

\.r. ,*,

CH: CH

4-DimethyI aminosti I bene

FIc. 103. 4-Aminostilbenederivative.

It must be emphasized,however, that according to the concept of

plural factors in carcinogenesis,
twin formation does not appear to be an
obligatorycondition for carcinogenicactivity; other factors can produce
suchactivity.
It is interestingto note that in most carcinogens,especiallyin the hydrocarbons,the twin formation is electrophobicdue to its richnessin electrons.For the present,we wish to stressonly that in substances
considered
to be activelycarcinogenic,i.e., capableof inducing invasivecancer, twin

lott

D i e t h y l si ltb e s t r o l
F r c . 1 0 4 . A t w i n f o r m a t i o n e x i s l si n d i e t h v l s t i l b e s t r o l .

278 /

x E S E A R c Hr N p H y s r o p A T H o L o c y

Triphenyl-ethylene
F r c . 1 0 5 . T h e p o s i t i o no f t h e t w i n f o r m a t i o n i n t r i p h e n y l - e t h y l e n e .

formation appearsto be an added factor which insurescomplex activity.

Intervcntionof groups of two energeticcenterswith the same character,
in carcinogens,places in a speciallight a group of agents which, under
particularcircumstances,
inducetumors.One group with alkylatingactiviry.
is formed by the nitrogenmustards,diepoxides,polyethyleneaminesand
dimethanesulfonoxyalkanes.
One of the physicochemicalcharacteristics
of this group is the presenceof two electrophiliccentersnear enough to
eachother to permit joint action. Still more important seemsto be the fact
that, through changesin all thesesubstances,
new formations may appear
which energeticallycould be ultimately consideredsimilar to twin formations. Through this character,their activity could also be parallel to that
mentionedabove.
in the carcinogens
encountered

NHe-C0-OCzHs
Urethan
F r c . 1 0 6 . U r e t h a n h a s n o t w i n f o r m a t i o n a n d a p p a r e n t l y - a c c o r d i n gt o m a n y a u t h o r s
- n o d i r e c t c a r c i n o g e n i ca c t i v i t y .

Nitrogen Mustud Derivatives

of this group are
and betterstudiedsubstances
The most representative
by the 2-halocthylamine
thc nitrogenmustard derivativcs,characterized
(Fig.
group
107) attachedto a radicalwhich can be aliphaticor aromatic.
It seemsthat it is through hydrolysisthat the compound becomesbiolog-

PRoBLEMSrN CANCER

279

ically active,and Haddow has shown that activity is presentonly if hydrolysis is sufficientlyhigh. (62) The inequalityof hydrolysisin different
memberscan be related to the influenccexertedby the radical bound to
the nitrogen.It seemsthat the presenceof a strongercnergcticcenter,as
it appears in positively or negativelycharged atoms bound to the cyclic
radical, reduces the dissociationof the chloroethyl group. Generally.
nucleophilicgloups would retard the dissociation.
Sufficientevidenceexists
to show that biologicalactivity follows the elimination of the chloride ion
and the appearanceof a carbonium ion as a reactive intermediate.A
further passageinto the ethyleneimonium
ion, consideredmore stableand

R *a"'
'cH2

cHzcl
c+z ct

NitrogenMustard
F l c . 1 0 7 . T h e n i t r o g e nm u s t a r dd e r i v a t i v e s .

thereforelessreactive,seemsto completethe transformation.Figure 108

showsthesechanges.
The haloalkyl side chains in the moleculeappar indispensable
for
biologicalactivity. (63, 64,65, 66) Thcy lead to the immediateappearanceof two positiveelectrostaticenergeticcenters.This does not represent
a minimalcondition,accordingto Landingand co-workers.( 67, 68) These
investigators
have shown that in nitrogenmustards,cytotoxicityincreases
with the number of haloalkylside chains.It is to be noted that a double
electrophiliccenter is found not only in the two original haloalkyl side
chains,but also in the later product,the ethyleneimonium
ion. In view of
the more frequentappearanceof this ion also for other agents,the analysis
of the relationshipof this group to twin formation appearsinteresting.
group, while a negativechargecan be seenat
In the ethyleneimonium
the nitrogen,a positivechargeappearsto be presentbetweenthe two CH,,,
providing a certain polarity. With two carbons positively charged and in
a relativelyfixed position,this group is similar energetically
to a positivc
tn'in carbon group. A two-step change,with the imonium group in the
first, and a carbonium in the second,can explain, as we shall see below.
the strangebiological activity of the nitrogen mustardswhich have a carcinogenicactivityonly throughchangeswhich take place in the organism.

280

xEsEARcH rN pHysropATHoLocy

zCHz-CHz-CL

C H-sN <\ - C H z - C H z - C L
I

nCHz

*ct-

CHr-r'r.(CH.

\r*,

- c H z- c t

+Hz0

J
-.CHz-CHr-oH
cHs- N< - C H . - C H r - c L

, L H r - C H ro H
/

CHs- u "(-Cn.

\l ' C H r

+ Ct-

+ HrO
/.'CHz
C H s- N . _ _^ . .

- CHu - OH r+l

-CHr -CHr - 0H F)

Dielectrophilic

'Ihc
Fto. 108.
c h a n g e sc l c c u r r i n gi n t h e n i t r o g e nm u s t a r d l e a d s t o e t h y l e n e i m m o n i u m
i n w h i c h a n e n e r g e t i ca s p e c ts i m i l a r t o r h a t o f a t w i n f o r m a t i o n i s p r e s e n t .

This agreesstrongly with the nature of the more recently studiedrelatively

activecarcinogens,the ethyleneimines,
wheresimilar centersare seen.(Flg.
i,09) The biologicaleffectof the ethyleneimine
group has been considered
to be relatedto a reactiveintermediatc.
Generally, if sufficient influence is exerted by another center in the
molecule,the imine group becomesactive.This centercan be a nitro group
as in 2:4 dinitrophenyl-ethyleneimine,
or other ethyleneiminegroups as

P R O B L E M Sr N C A N C E R

2gl

in methyleneimine
l:3:5 triazine,(Fig. l,r0l Through the influenceexerted
by these centers,the ethyleneiminegroup can have its carbons charged
sufficientlyto bccome a dielectrophilicformation. The possibilityof a reactive intermediate and a more stable electrophilic form thus appears
common to the two groups,mustardsand ethyleneimines.

;>. a=-<-;
\z
N

H,cArr,
Triethylenimine
2-4-6-Triazine
Flo. 109. Ethyleneimines are active carcinogens,probably related to their energctic
aspectwith a formation energeticallysimilar to the twin formation.

Eporide Cucinogens
A similar condition is also found in the epoxide carcinogens.carcinogenic activity has been recognizedfor substanceshaving two epoxide centers in close proximity in the molecule. The epoxide center by itself can
leadto a formationsimilar to that of carboniumion, as seenin Figure 108
and thus to the same formation found in mustards and ethyleneimines.
The analogy goes still further. The energeticcenter appears insrrfficientto
accomplishbiologicalchangeswithout an inductiveactivation.In the caseof
epoxides, this is usually brought about by another similar epoxide group
in the samemolecule.
As no carcinogenicactivity has been found in substanceswith only one
epoxidecenter or with two epoxidecentersfar apart, the inductive centers
seemto be of primary importance.The two energeticcentersforming the
epoxide g'oup, similar to those of the ethyleneimines,do not alone appear

282

,/

xEsEARcH rN pHysropATHoLocy

sufficiently reactive to induce important changes.Only when enhanced by

reciprocal induction is their reactivity adequate to induce either the appearanceof a reactive intermediateor a sufficientcharge in the ethylene
carbonsto produce biologrcalactivity. Thesechangescan be measuredby
the reaction with thiosulfate ion and consequentlycan be related to the
reciprocalpositionsof the two epoxidecenters.
The biologrcalactivity of dimethanesulfonoxyalkanes
can also be related to a similar energeticformation. Such a formation appearswhen the

NOz

i)
N

H,cAcH,
2 : 4 D i n i tr o p h e n y l
Ethylenimine
F I c . l l 0 . T h r o u g h t h e i n f l u e n c ee x e r t e db y t h e n i t r o g r o u p u p o n t h e e t h y l e n e i m i n e ,
t h e i m i n e g r o u p o f 2 : 4 d i n i t r o - p h e n y l - e t h y l e n e i m i nbee, c o m e sd i a l e c r r o p h y l i c .

moleculeis metabolized,with the differencethat the two CHr in this instance seem to come originally from other chains. (Fig. I I I ) For the
it is possiblethat a similar processoccurs during the
methylolamides,
changesthat take placein the organism.
Some corroboration can be found in the fact that two forms can be
observedin theselast groupsof carcinogens.One is electrostaticallyactive;
that is, it has a certain ionic character.The secondhas a dual electrophilic
activity which can be relatedto a twin formation with nrolecularreactivity.
Thus, twin formation,with its specialreactivity,appearscommon in
many carcinogenicagents.To be biologicallyactive,the twin formation has
to be sufficientlystrong and this is insured by an induction effect exerted

PRoBLEMS

rN

CANCER

283

by other formationsin the molecule,such as double bonds in parallel position or polar groups.A twin formation as energeticcenter in the molecule
would exert a molecular field effcct. It would thus representa center of
molecularreactivity which has to be consideredas such in the analysisof
plural activity.

Ar.N{

- . C H z - C H z 0 S0 2 ' A r

RCH2+
t

\cHr c H 2- o s o 2 .A r

-0502' Ar
R C H 2 ++

Sulfonoxyalkane

-0302'

Ar

ReactiveIntermediate

F I c . t l l . C h a n g e so c c u r r i n g i n s u l f o n o x y a l k a n c sl e a d i n g t o t w o a c t i v e C H r c e n t e r s .

SynjugatedFormations
The study of variouscarcinogenshas permittedus to recognizeand
relate to complexcarcinogenicactivityanothercnergeticinfluenceexerted
by two or more doublebondswhen prcsentin a parallelreciprocalposition
in cyclic molecules.This led us to the conceptof "synjugatedformations"
with 2, 3, 4 or more suchparalleldoublebonds.
In studyingmethylcholanthrene,
one o[ the most potent of the known
carcinogenicagenLs,the curve of its absorptionin ultraviolet light was
considered.This curve is shown in Figure | 12. The place and form of
the peakscould be interpretedin a peculiarway when conjugateddouble
bond formationswere considered.In thc curve of methylcholanthrene,
we
portions
that corrcspondto an inverseof the curvesobcould recognizr
tained from variousconjugatedpolyenes.Furthermore,the curve obtained
through the spectralanalysisof methylcholanthrene
can be consideredto
have high similaritiesto the inverscof the curveof a mixtureof conjugated
polyenes.Figure I l3 showsthe spectralanalysisof conjugatedcod liver oil
f a t t y a c i d s ,w h i l eF i g . l l 4 s h o w st h e i n v e r s ec u r v eo f m i x t u r eo f c o n j u g a t e d
fatty acidsof cod liver oil in which conjugateddi-, tri-, tetra-,penta-and
hexaenesare identified.Figure I l5 shows the comparisonbetweenthe
and the inverseof the peaksof the mixture.
curve of methylcholanthrene
We were thus led to consider the conceptualinterpretationof these
curvesin termsof the specialrelationshipthat existsbetweendoublebonds
in the same molecule.In the classicalconcept,two double bonds are consideredconjugatedif two of their carbonsare joined by a singlebond. ln
the zig-zag representationof aliphatic molecules,the conjugated double

r,iti{

284

RESEARcH rN PHYSToPATHoLocY

bondsfnlfill this condition.(Frg. 1I6a) Applying this relationshipto cyclic

what wasconsideredto correspondto conjugation,accordingto
molecules,
this criterion,did not showpropertiessimilarto conjugatedaliphaticmembers.(Frg.II6b) This madeus consider,as the conditionfor the properties
presentin conjugated
formations,anothercharacter:the reciprocalparallelismbetweendoublebondspresentas they appearin the aliphaticmole-

c
o

I
c
L

Y a v eL e n g t h( n 1 )

Fto. I 12. An interpretation ol the spectral analysesol methylcholanthrene. Curve (a)

shows the spcctral analysisin ultra-violet of methylcholanthrenc.

cule. Two or more double bonds in a cyclic molecule would thus realize a
similar kind of energeticformation when parallel, and would do so independentlyof the number of the singlebondspresentin-between.(FiS. 1I6c)
For didactic purposes,we have applied the term "synjugated" to energetic
formations resulting from parallel double bonds separatedby more than
one single bond.

- , , r - : r . $ : : , :':+--"1ar.i:i:
{1$.
:1 .i"r: i..;itri. ,

I riilL

P R O B L f : M S I N

C A N C E R

285

c
cl

; $
tn

E
o
C
.E
L

*60
br

mpp))
f i a v e L eennqqt thh ( m
F t c . I 1 3 . T h e c u r v e s h o w st h e s p e c t r a a
l n a l y s i so f r h e m i x r u r e o f c o n j u g a t e df a t t y
a c i d sw i t h m e m b e r s h a v i n g f r o m 2 t o 6 d o u b l e b o n d s . a s o b t a i n e d b y t r e a t i n g c o d
liver oil fatty acids with KOH.

Thus, in the methylcholanthrene

molecule,there exist formationscomposedof two, threeand four paralleldoublebonds(Fig. I 17), which we call
di-, tri-, and tetraenicsynjugatedformations.It is logicarto assumethat
they are important in determining the energeticaspect of this molecule

286

R E S E A R c Hr N p H y s r o p A T H o L o c y

when the relationshipof its spectral analysisto the curve corresponding

to the inverseof conjugateddi-, tri- and tetraenescan be recognized.Frorn
the point of view of its relationshipto the plurality of factors determining
the carcinogenicityof a substance,the presenccof parallel double bonds,

c
ct
o
tt

E
tt
c
(!
L

a.a

tlave Lenoth (-r)

F t c . I 1 4 . T h i s c u r v e i s t h e i n v e r s eo f t h e c u r v e o f F i g . l l 3 .

and the synjugatedformationswhich they constitute,is interesting.Theoretically,eachone of thcsesynjugatedfornrationswould by itself represent

a reactivepossibility.Althoughqualitativelysimilar,they would showmanifest quantitativedifferences.It must be noted that, while they are not presthc'yare in most active,realizingdi-, tri-, tetra- and
cnt in all carcinogens.

PRoBLEMS rN CANCER

287

evenpenta-synjugated
formations.According to the concpt of plural activity in carcinogenesis,
synjugation,while not indispensablefor carcinogeneticactivity,would representone of the factorsthat can make it possible.
Togetherwith the condensationof the electronsin the K regionsand
".
the presenceof polar groups, the twin and synjugatedformations would

?to
o
(-

-c
c

,'
,

.I'

l\
, l

Eta

|
I

-c

269

I
L -

60

l ' .

J \

c
o

'

it
234

, ' I

'izo
c
(g
L
}Q

Joo

280

250

llaveLenoth('r) for Methylcnolanthrene

F r c . l l 5 . D i r e c t c o m p a r i s o n b e t w e n t h e c u r v e o f t h e s p e c t r a l a n a l y s i so f m e t h y l c h o l a n t h r e n ea n d t h e j n v e r s eo f t h e p e a k s c h a r a c r e r i s t i fco r t h e different conjugated
f a t t y a c i d sa s s e e ni n F i g s . l 1 3 a n d l 1 4 .

288

REsEARcH tN pHysropATHoLocy

confer high plural activity upon the molecules of active carcinogens.An

energeticspectrumof a carcinogencan be establishedin which thesefactors
can be presentedsystematically.
F i g u r e l l 8 s h o w s a s p e c t r u mf o r 9 : 1 0 D i m e t h y l I : 2 : l : 8 D i b e n z anthracene.
In the light of this analysis,it appearslogical to conceive that the carcinogenicity of a chemical compound is a result of many factors, and that
the great differencesin carcinogenicpropertiesof various agentsis the re-

a
Frc. l16. Conjugation and synjugation. ln the aliphatic chain (a) the presence of
single bonds between the double bonds induce the parallel position of double bonds.
It is this parallelism, which through thc reciprocal influence cxertcd, induccs thc
energctic characteristicsof the conjugated formations. Io the benzene molecule (b)
whcre thc double bonds, although separated by single bonds, arc not paraltel, thc
lack of this parallelism explains the lack of the properties charactcristic to thc conjugated formation. Tbe parallelism when present in cyclic molecules (c) realizcs thc
"synjugated" formations.

sult of difterences in their energetic spectra. The di.fterencesare consequently qualitative as well as quantitative. From this viewpoint, it is
possiblethat the great carcinogenicactivity recognizedfor some substances
would correspond to the presencein them at once of a great number of
energeticfactors.
The study of the correlationbetweenthe presenceof various energetic
centers and carcinogenesishas been facilitated by relating carcinogenic
changesto levels of organization.Taking place at different levels, the induced processes
can be seento correspondto an entire seriesof manifestations which, while present also in invasive cancer, often can be recognized

PRoBLEMSrN cANcER

289

Tetrasynjugat
ed

p;
HS
/

.r'

Trisynjugat
ed/----/-'- J

/'

Eisynjugat
ed

ion in 20-Methylcholanthrene
Synjugat
F I c . l l 7 . T h e p a r a l l e l p o s i t i o n o f t h e e x i s t i n gd o u b l e b o n d s i n m e t h y l c h o l a n t h r e n e
c o r r e s p o n d st o a b i - , t r i - , a n d t e t r a s y n j u g a t i o n .

Tl l n ai
.a--

\
-\

cHs

\ i
il-l
r ll
\ / l

{
I
f-\ I

,
t

.
/

t
l -

lt /

'l r l r

-'\
t

T w o K R e g i o n s+ .
T r oT r i n F o r m a t i o n s
[tneTrisynjugated
Bond
Tro Tetrasynjugated
Bonds

L'{r H :
/

Tetra

-V

Tetra

9 : 1 0D i m e t h y ll : 2 : 7 : 8 - D i b e n z a n t h r a c e n e
F r o . l l 8 . ' f h e e n c r g e t i cp i c t u r e o f 9 : 1 0 d i m e t h y l . l : 2 : 7 : 8 d i b e n z a n t h r a c e nseh, o w s
t h e p r e s e n c eo f t w o K r e g i o n s ,t w o t w i n n e g a t i v ef o r m a t i o n s ,o n e t r i s y n j u g a t e db o n d
and two tetrasynjugatcdbonds.

29O /

n E s E A R c Hr N p H y s r o p A T H o L o c y

in casesin which an invasivecanceris not induced.Followingthis view, it

can be expectedthat carcinogenesis
is the summationof a wholeseriesof
actionsinducedin the organism,someexogenous
and othersendogenous.
Consideration
of thepluralityof the factorswhichintervenein chemical
carcinogenesis
and makeit a complexprocessleadsus to considerviruses
in the etiologyand pathogenesis
of cancerin a similarlight.
VIRUSES AND CANCER
More than fifty years ago, Borrel presentedhis hypothesisof viral origin
of cancer basedprimarily on analogies,Sincethen, although an enormous
amount of material on this subjecthas accumulated,much of it has been
contradictory and it has appearedto be impossible to arrive at any clearcut conceptof the role of virusesin the pathogenesis
of tumors. However,
it seemedthat an attempt to correlatemost of the data with information
furnishedby the study of chemicalcarcinogenesis
in the light of the concept
of cancer as a complex hierarchicdiseasemight be of some value for an
initial simplificationof the problem.(293, 312) (Note 2)
Some theoreticalconsiderationshave helped in systematizingthe data
and in indicatingthe probablelimits of viral interventionin carcinogenesis.
Just as with chemicalcarcinogens,it could be assumedthat virus intervention may bring to bear multiple factors.An analysisof the processeswhich
occur under viral influenceindicatesthat this hypothesisis plausible.
Even more than chemical carcinogens,virusesare able to act only at
certain levels of organization,Their intracytoplasmaticand often intranuclear developmentconditionsthe interventionof theseagentsat theselevels.
Fundamentaldifferencesin carcinogeniceffectcould be expectedif vinrses
are able to influencethe subnuclearlevels,or the nucleus,or, on the other
hand if their activity is limited to the cytoplasm.This view has permitted
us to understandthe striking differencein influence exerted by various
viruseswhich, although recognizedby any worker in the field, has not been
the subjectof any specialconsideration.
Two Types ol Carcinogenic Eflects
The differencc lies in the time neededfor a virus to produce carcinogenic effects.Inoculation of fowls with purified Rous sarcoma virus, for
instance,has been seen to produce a clear<ut, immediate eftect. Changes
have been recognizcdwithin 48 hours at the site of inoculation.They take
place in the nuclei of fibrocytesand consist of swelling, appearanceof a
more distinctnuclearmembrane,clearednucleoplasm,marginationof chro-

PRoBLEMS

tN

CANCER

2gl

matin and one or more enlargednucleoli,In as few as onc or two days,

cytoplasmicchangesare also cvident.Thcre is manifestbasophiliawith
swellingof the cell which becomesgreatly enlarged.Concomitantwith
thesechanges,the abnormal cells invade the fibrillar tissue.The tumor
whichdevelopshasthe characterof the classicalspindlecell Rous sarcoma.
Thus a canceroustumor in the invasivestage,with typical nuclear and
cytoplasmiccell characters,is induccdin only a ferv days at the site of
inoculation.This is charactcristicof one type of viral carcinogenesis,
the
extremelyactiveone. Integratedin the conceptof complexcarcinogenesis
presentedabove, it would mean that the cntirc seriesof changes-from
thoseat the lowestlevel which determinethe cancerouscharacterto the
cytoplasmicchangeswhich producethe prolifcrative,invasives6ngs1-ha5
been achievedby the virus in this short time. ln fact, this tumor grows
rapidly, is palpable even at the f,fth day and fatal in rwo to three weeks.
Almost diametrica.lly
opposedto this type of carcinogcnesis
are tumors
which representanothertype of virusintervcntion,
suchascertainmammary
cancersin mice. viruses that producesuch tumors can be obtainedfrom
variousorgans,even from thoseof animalswithout apparenttumors.They
inducethe appearance
of tumorsbut only undervery characteristic
circumstances.Preferablyintroducedin the first days of life-subcutaneously,
intraperitoneallyor even orally-they will producetheir effect only aftcr
many monthsor even afterone or two years,as tumorsof a specificorgan,
such as the mammary gJand,for example.However,such tumors appear
almostonly in femaleswho havehad onc or more prcgnancies.
In this case,
the virus acts only upon highly differentiated
cells and acts independently
of the site of inoculation.The extremelylong period without manifestation,
the fact that the virus can be found to someextentin variousorganswhich
show no change,and the specificlocalizationin a highly differentiated
organ such as the mammarygJand,would indicatethat the carcinogenic
interventionof the virus is highly relatedto a specificcharacterof thesecells,
their parriculardifferentiation.This would place virus interventionat the
cytoplasmiclevel where differentiationoccurs.
Under this interpretation,the length of time necessaryfor tumor apparancewould be relatedto the time neededfor a natural evolutionof
the mammary cells to the point where they arc sufficientlydifferentiated.
It appearsprobablethat this lengthof time corresponds
to that neededby
abnormal hierarchicentitiesof the mammarycells to have arrived,independentlyof the virus, at a state correspondingto that of precanceror
noninvasivecancer.Interventionof the virus at the cytoplasmiclevcl would
then transform the relativelvadvancedbut still noninvasivecancer cells

292

xEsEARcH rN pHysropATHoLocy

into invasivecancer cells. Viruses would act, in this case, as cytoplasmic

carcinogens.
These two types representthe extremesof carcinogenesisin which
virusesplay a role. They help to rnterpretmany of the other data furnished
by experiments.For didactic purposes,we sha.llregard as "broad-scale"
viruses those which act from very low to high levels of the organism, and
as "cytoplasmatic"thosewhich act at the higher cellular level only.
some of the rapidly acting broad-scaleviruses will induce evolving
tumors in a much shorter time than any known chemical or physical carcinogen,a fact that can be interpretedas meaning that these viruses are
more capable of inducing not only the cytoplasmaticcarcinogenicchanges
but also the entire scale of preparatory changesleading to invasive cancer.
Viruses differ from the usua.lactive chemicalcarcinogensin their special
capacity to induce changeseasily at the cytoplasmiclevel where they are
particularly capable of multiplying and acting. This would contrast with
most chemicalagentswhich generallyhave low carcinogenicactivity at the
ceUularlevel. Chemicaland virus carcinogenic
activity would complement
each other. This is in accord with experimentsof Russianscientists,which
have shown that culturesof cells treatedwith methylcholanthrene
in vitro
becomehighly carcinogenicwhen inoculatedin animals if a cancerousvims
is also added.
This view of the activity of cytoplasmaticvirusesat the higher cellular
level, as contrastedwith many chemicalcarcinogensusually more active
at lower levels, appearsalso to be in agreementwith the experimentsof
Rous and Kidd (69, 70) which demonstratedthe capacity of coal tar
extractsto localize the Shope papilloma virus. The high cytotropic character of this virus is well known. It would easily act upon cells already
transformedfrom normai into noninvasiveform by chemicalagentswhich
are active at the lower levels. These chemical agentsthus "localize" the
vira-lactivity. This is in accord with the ability of chemical agentsin the
Rous and Kidd experimentsto increasethe percentageof invading carcinomas as compared to thc papillomaspresent. According to the view
presentedabove,the papillomaas a benigntumor would representchanges
similar to those seenin cancerbut limited exclusivelyto the higher levels,
without cancerousentitiesat the lowcr levels.The addition of an agent
with a broad-scaleof carcinogenicactivity, that is, acting also at the lower
levels,suchas the chemicalagent,would give the resultinglesionthe entire
canceroussca-le,
that is, the characterof nralignancy.
The integrationof viral carcinogenesis
in the concept of cancer as a
complexcondition and recognitionof the two extremetypes of viral car-

PRoBLEMS lN

cANcER

293

cinogenicactivity permits us to understandthc reservc of most authors

overthe viral etiologyof cancer.For instance,many have refusedto accept
as a carcinogenicfactor the virus shown by Bittner ( 7l ) to be presentin
maternalmilk and to influencethe appearanccof mammary carcinoma in
mice.The refusalis basedupon comparisonof this virus with that of the
first typ seen in fowl tumors. With the systematizationpresentedabove
and the concept of broad-scaleand cytoplasmaticcarcinogenicviruses,
this reason is not vaiid. Furthermore,a carcinogenicvirus should not be
consideredto be the indispensable
factor able to induce proliferativecancer
in animals which usually have a virus cancer.This would explain why, in
certainbreastcarcinomasin mice, a viral agentcould never be found. (72)
The specificcapacity of a virus to act upon a dilrerentiatedcytoplasm
explainsthe fact that a virus may be widely distributedamongorgansbut
does not induce tumors except in spccial cells. previous prepzuatory
changesseemnecessaryfor the cytoplasmaticvirus to intervene.This is in
accordwith a low incidenceof tumors in certain strainsof mice despitean
abundantpresenceof the "milk factor" virus. ( 73)
These facts shed a new light on the entire problem of the relationship
between viruses and tumors. Viruses can multiply in organismswithout
inducingcancer.The virus of mammarycarcinomain mice can be transmitted to femalesthrough spermatozoaand can be found in large amounts
throughoutthe organism.The virus is presenta long time before any cancerouslesionsare seenand is presentin organsthat will neverhave tumors
and even in animals that never develop cancer.The developmentof this
virus,like all viruses,takesplace in the cell cytoplasmwhich doesnot necessarilymean the induction of carcinogenesis
as long as other factors are
not present.No tumors appearas long as the cell has not undergonethe
prior changesrequired if the cytoplasmaticcarcinogeniceffect is to take
place.without the previouschanges,the virus will not influencethe cell
any more than many other noncarcinogenic
viruses.It is only in the presenceof an advancedcellularchangethat the virus will produccan invasive
cancer.
Plural Activiry
The capacityof a broad-scalevirus to inducean invasivetumor in a
short time through plural activity at different levels has been related to
its richnessin lipids. Its analysismakes us suspectthe presenceof several parts in the virus, each one able to act at a diflerent level, as in the
caseof activechemicalcarcinogens.
A similar plural influencecan be seen

294 /

B , E s E A R c Hr N p H y s r o p A T H o L o c y

exertedby virusesother than those with carcinogenicactivity. The study

of bacterialviruseshas shown the existenceof such plural parts. (Note I )
Luria (74) has shown, after irradiating a bacteriophagewith ultraviolet Iight, that if lytic activity can no longer be obtainedby the intervention of a singleone of thcseparticlcs,it can be inducedwith two or more
of them. They act as though severalparts, which usually are presentin the
virus but which were unequally inactivatedby the irradiation, would be
necessaryin order to induce the processof lysis. This agreeswith the experimentsof Debruck and Hershey (75), which have shown that new
typesof viruseswith new propertiescan be obtainedwhen units of the same
phagestrain or related strainsare mixcd together.The new propertiesare
combinations
of thoseof the mixedunirs.(298)
Similarchangesin thc plural constitutionof the viruseswould explain
other peculiaritiesobservedin bacterialphages.Bacteria can carry phages
for generations
beforeany lytic activityoccurs.The lysogenicstrains(76)
of bacteriaare examples.It is possiblethat a virus may undergotemporary
changesunder certain circumstances;this would explain the frequent impossibilityof finding a virus immediatelyafter it infects a bacterium, ild
the "disappearance"of somevirusesin animalsimmediatelyafter infection.
Since, in both cases,the virus is found later, a change which makes it
unable to act and therebybe dctectedis plausible.The "masked" virus
would be one with only some of its plural propcrties present.The possibility of recoveringa lost property was demonstratedin the Berry-Dietrich
phenomenon,when a heat-inactivatedmyxoma virus recoveredits lethal
capacityif inoculatedalong with a libroma virus.
This conceptof plural activity finds further applicationin the explanation of many phenomenaobservedin virusesin generaland in variations
in carcinogenesis.
The "self-sterilizationof the neuro-infections"described
by Levaditi has to be regardedrather as partial inactivationof the viruses
especiallyif the virusescan be reactivated.This occurrencemust be separated from caseswhere a total destructionof the virus can be supposedto
have taken place.
The lethal infection induced in mice injected intraperitoneally with
salivary gland virusesof certain strains is an example of the latter. The
presenceof thc inclusionbodiesin liver and other organs,and the total
inabilityto produce the diseasein other mice (77) can be interpretedas
a signof a destructionof thc virusesin the organism.The inclusionbodies
can be interpretedas resultingfrom an agglutinationof the virusesthemselvesas shown by Nicolau in herpes.(78) In other cases,such as protracted herpesinfectionin rabbits (79), or vaccinalinfection in rabbits

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(80), only partial inactivation can be consideredto occur since electrophoresis,repeatedpassage,or even dilution restorespathogenicity,The
restorativefactor can be of varied nature.Casesin which pathogenicityis
restoredby a nonviral agent-activation of the virus of swine influenzain
the presenceof Hemophilusinfluenza(81), for example-are most revealing.
Virus and the Host
This conceptof plural activity cxplainsthe rclation betweentumorgenesisand destructioninduced by viruses.Often "neoplastic" infection
and "destructive"infectionare inducedby the samevirus. (82)
The herpes virus thus induces necrotic lesions in the chick embryo
whenintroducedin early stages,but if the cmbryo is more developed,the
samevirus producesproliferativechanges.(83) The myxomavirus induces
more proliferative lesions if attentuatedthan does the unchangedvirus.
(84) Under specialcircumstances,
such as in oldcr animals,shecp pox
virus inducespapilloma insteadof pustular infection (85). It must be
remarkedthat thesedifferentresultsare not limited to viruses;they occur
with radiationor even with other infectiousagents.(86) Bartonellabacilliformis,which inducesoften-lcthaloroya fevcr, seemsto be the causeof
"verruga peruviana," a fibroangiomatoustumor often seen in subjects
recoveringfrom the acute disease.
The differencesin activity of thc same virus appear to be related to
the age of the host. Generally,youth of the host increasesthe virus' capacityfor acting at more levels.The virus can produce lethal destructive
disease
in young animalsbut only a neoplasticrcsponsein adults,as seen
for the fibroma virus in rabbits.Furthermorc,the neoplasticresponsealso
occursin young animalsbut only if a small amountof virus is inoculated,
or if an attenuatedvirus, such as a long-storedone, is used. (87) This is
clearin the caseof the Rous sarcomaand other chickcntumors.
When injected into very young animals. Rous sarcoma and other
chickentumor virusesproduce a hemorrhagiclesion (88) but they will
inducetumors in adult animals.The dcstructivccffect can bc repeated
with repeatedpassages
of the virus in very young animalsbut in adults
eachpassageproducesthe ncoplasticresponsc.This is also true for some
strainsof lymphomatosisvirus (89) which induce tumor formation in
adultsand necrotizingprocesses
in young animalsor embryos.It is also
true for the virus of neurolymphomatosis
(90), and of gliomas. (91)
Theseviruses,althoughselectivefor thc nervoussystem,induceinflammatory or neoplasticlesionsaccordingto the age of the infcctedanimal.

;1,,,,i1#ff$ii#ffiffi,($

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In a generalway, it has beenpostulatedthat for viruses,as for bacteria,

the young animal represcntsa favorableterrain, while a certain resistance
is encounteredin the adult. waters and Bywaters (92) have shown that
the filtrable agentisolatedby Prickett and Belding (93) is not transmitted
spontaneouslyif the animal is older than 40 days. It is transmittedthrough
the eggs, although months elapse before there are manifestations.(94)
The problem cannot bc limitcd to the host, since,by passingthe virus
through young or old animalssomeof its propertiescan be changed.Gross
(95) has shown that a cell-freeextract obtainedfrom leukemicmice of the
AK strain, would induce the condition in c3H mice, provided the inoculation is given within a few hours after birth. The resultspublishedby Gross
and the inability of other authors ro reproducethem (96) could be explained by differencesin the virus strainsused. (97)
This was shown in the experimentof F. Duran-Reynals(98), in
which the Rous sarcoma virus undergoeschangesduring passagein the
adult chicken, which make it adaptableto another species,namely ducks.
The virus growing in young chicks seemedunable to induce the disease
when inoculatedin ducklingsor in older animals.Tumors obtained even
through cell suspensions,some of them very large tumors, could not be
transmittedfor more than one or two generations.However, filtrates of
tumors from older chickens,when injectedinto ducklingsno more than a
few days old, induced tumors which easily could be passedto young as
well as adult ducks. This changein the virus was strictly conditioned by
the age of the chicken; it occurred only if the animal was between three
and ten monthsold. If the animalwas more than 19-20 monthsold, injection of the filtrateswas alwaysunsuccessful,
and injection of the cells only
rarely induced tumors.
These changesin the virus arc explainedby mutation. Among various
resonanceforms which appear on a purely statisticalbasis, one different
from those previouslypredominantfinds favorableconditionsfor its development---+onditions that are not favorable for the predominant forms.
These experimentshave permittedus to further correlatethe intervention
of viruseswith the influenceexcrtedby severalchemicalfactors upon the
complextumor pathogenesis.
Virus and Lipoids
In studying the influenceexcrted by lipoids upon viral activity. we
could show that the presenceof free fatty acids, especiallypolyunsaturated,inducedchangesoppositeto thoseinducedby the anti-fatty acids.
In rabbits, administrationof various preparationsof fatty acids, espe-

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cially polyunsaturated,induced an unusual degreeof resistance.Animals

previouslygiven subcutaneousinjectionsof fatty acid preparationsshowed
a reduced generalresponseto chicken pox inoculation as compared with
controls, and practically no responsein the skin at the site of the fatty
acids injection. on the other hand, adminisrrationof insaponifiablelipid
fractions obtainedfrom tissuesof receptivespecieswas followed by manifest responseslocalized in the zone of injection, even in speciesotherwise
refractiveto viral infection. It is under this specialinfluenceof lipoids that
we have further investigatedthe interventionof the virusesin carcinogenesis.
cells vary in their content of lipids. we could see that richnessin
sterolsof a group of cellsincreascs
their receptivityro, and favorsthe development of, viruses in general,while richnessin fatty acids, especially
polyunsaturated,has an opposite effect. The local increasein a tissue's
richnessin sterolsmakes it more susceptiblcto the localizationand development of a virus, as is shown in thc followingexperimcnt.
In rabbits, intracutaneousor subcutaneous
injectionsof a colloidal
suspensionof cholesterol were made on epilated skin at several sites.
Twenty-fourhours later, the animal was injectedintravenouslywith suspension of smallpox vaccine.Characteristiclcsions were observedto develop at the sitesof the cholesterolinjections.
The generaleffectof sterolsupon receptivityto viruses,noted in many
experimentsin animals, was also recognizedin humans. The following
observationappearsinteresting.Mrs. D. R. had alwaysappearedrefractory
to smallpox vaccines.Until the age of 40, repeatedinoculationsproduced
constantlynegativeresponses.
She was treatedat that timc with a cholesterol preparationfor precordialpain, receivingdaily 2-3 cc. of a 2.5vo
solutionof cholesterolintramuscularly.
After threeweeksof this treatment,
shewas obligedto go abroad and it was nccessaryfor hcr to have the routine smallpoxvaccination.For the first time in her life, a characteristic
positiveresult was obtained.
The relationshipbetweensterolsand viruses,which would explainthe
affinityof most virusesfor the nervoussystcmand skin, sinccboth are of
exodermicorigin and particularlyrich in free sterols,would also explain
why young cellssimilarlyricher in sterolsare more susceptible
to viruses,
and the facility with which almostall virusesdevelopin embryos,such as
in chicken embryos.
Changesin richnessin lipids wereobservedundernaturalcircumstances
other than thoserelatedto age.Thus seasonal
changescould be noted,the
cold seasonleading to an increaseof fatty acids while the summer season

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brought an increaseof sterols,This would help to explain the seasonal

changesusua.llyobservedin naturally occurring viral infections.(99)
The epidemiologyof poliomyelitismay be related to the organism's
richness in sterols in the summer, particularly on hotter days. Seasonal
changeswere noted in naturally occurring tumors in which a viral etiology is seen. A certain resistanceappears in the fall and increasesin the
winter in the caseof leukosesand possiblyin other natural viral tumors.
( 100) This would explain the manifest seasonalchangesobservedby us
in the transplantedWalker tumor in rats or in grafted tumors in mice in
general,and in the induction of tumors through carcinogens.Similarly, the
induction of teratomas in testes through local admini5jlsllsn of zinc
chloride was noted to be influencedby the seasons.( l0l )
The influenccexertedby sterolswould explain the fact that virusesable
to act only at a higher level, as in the cytoplasm,tend to developin animal
cells abnormallyrich in sterols.It is highly probably that once it has penetrated, a virus will develop within a cell only under favorable conditions,
and these are hsured by the presenceof sterols. The virus will persist,
interferinglittle with the fate of the cell until other changesoccru at lower
levels.These othcr changestake many months or even years to be completed,and only then would the influenceof the virus be apparentthrough
its activation of the noninvasiveabnormal cell. Activation can occur regardlessof seasonalchangesin sterol richness.It seemssuperfluousto note
that this relationshipholds more for cytoplasmaticviruses than for those
with broad-scaleactivity.The latter are also more active in young animals.
changes in age of the host and other circumstancescan modify the
characterof viral carcinogenesis,
leading to rapid or very slow develop
ment, or even to completclack of rcsponse.This was often noted for Rous
sarcoma.In young animals,small amountsinducedrapidly growing tumors
with multiple hemorrhagicmetastases
rich in fi.ltrablevirus. In adult animals, despitethe largc amount of virus necessary,tumors took months to
appear,seldom metastasized,
and could be transmittedwith difficulty, or
not at all, by f,ltratesor evenby cells. (102)
The relationshipbetweenviral carcinogenesis
and lipids has been the
basisfor a group of experimcntsin which we tried to influencethe carcino.
genic activity of a virus by administrationof stcrols. Experimentsstill in
progress,using sterolsobtained from chicken embryos, seem to indicate
that lipids can strongly changeviral carcinogenicactivity. In general,they
inducean increascdresponseto viruses.
Many other peculiaritiesof the relationshipbetweenvirusesand carcinogenesishave been analyzedin terms of intervention of lipids as an

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intermediaryfactor. The capacityof a virus to induce tumors in different

organs-as seenfor leukemictissuecell-freeextractsin mice,which induce
peculiar salivarygland tumors ( 103), or tumors in the adrenalgland or in
the subcutaneoustissues (104)---can be explainedby certain peculiar
affinitiesof the virusesfor differentiatedtissues,possiblyrelatedto certain
specificlipids found in thesetissues.A similar affinity for the salivarygland
is seenfor rabies virus. There are also the affinitiesshown by the neuroand dermatotropicviruses,and by the neoplasticvirusesfor mammary
piland,lymphatictissue,ncuroglia,etc. while affinityfor the adrenalgland
could be related to its richnessin sterols,afffrnityto others sites could be
relatedto other lipids.
The plurality of localizationsof virusesalso can be relatedto affinities
of mutatedagentsfor differentcells.The experiments
of Gross(105), which
show the possibilityof separatingout of the samefiltrate the agentsresponsible for salivary gland tumors and for leukemic changes,would indicate
that a changein the virus must be also considered.However, the change
can be interpretedas a mutation and can be relatedto the influenceexerted
by lipids upon the virus. Treatmentof virus with lipids has shown the possibility of inducing changesin its behavior.Data showing the influence
upon tissularreceptivityof such changcswill appearin future publications.
with this concept of the role of virusesin the pathogenesis
of cancer,
it seemspossibleto explain other peculiaritiesthat have led to confusion
in this field.
It has been noted that viruses act as factors determining the change
to a cancerousentity which, once induced,can continueto developwithout
need of further interventionof the virus. This posesthe problem of the
relationshipbetweenviral carcinogenesis
and developmentof the virus in
the tumor itself. Even in a tumor, the multiplicationof the virus has to be
separatedfrom that of the $owth of the tumor. Although often interrelated,
they must be consideredas two different processes.The growth and even
direct transmissibilityof the tumor can continue,independentof the presence of the agentthat originallyinducedit. when tumors have been induced by a chemicalcarcinogen,they can be transmittedin continuous
generationsover many years, producing large tumors each time, a fact
which would precludeany possibledirect interventionof the agentin these
later tumon. Similarly,tumors once inducedby virus can be further transmitted by cells,the virus no longer being apparentin the tumors. A tumor
inducedby a virus often servesas a medium for the multiplication of the
virus.However,even while the tumor can continucto grow, it can become
an adversemedium for the further multiplicationof the virus.

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This explainsthe peculiar facr that tumors induced by a virus can be

rich in or can lack an appreciableamount of virus, as often seen in a
s h o p e p a p i l l o m a( 1 0 6 ) , o r e v e n i n t u m o r s i n f o w l ( 1 0 7 ) , w h i c h p a s s
through periods when transmissionthrough cell-freefiltrates becomesimpossible,while transmission
throughthe transplantof tumor cells still continues, The virus multiplication capacitycan vary not only with the host
but with the virus itself, thus explainingthe changesnoted above.
The resultsof the interestingstudiesof Bryan, Galman and Maloney
(108) who have investigated
the relationshipbetweenrichnessin virus of
an induced Rous sarcomaand the percentageof positive resultsin induction easily can be interpretedunder this view. The chancesof inducing
tumors increasein casesin which the host is also a favorable medium for
the multiplication of the virus, and vice versa. This would explain why
theseauthorsfound little or no activevirus in casesin which the injected
material producedless than 5oo/o positiveresults,but casesoriginated by
a material that induced a large proportion of positive resultswere rich in
active virus. The capacity to multiply after inoculation in the host itself
thus increasesthe ability of the virus to act as a carcinogenicagent, which
seemslogical. The relative independence
of the two processes-the multiplicationof the virus and the inductionof [sp615-sppeared clearerin the
casesmentioned above, where the virus developsin the entire body of
mice, even in successive
generations,
without inducingtumors.
The presenceof virusesin the organism,even without inducingtumors.
helps to explain the rathcr puzzlingexperimentin which a transmission
through filtrates,consideredcharacteristic
for viruses,was seen to occur
for tumors inducedby chemicalcarcinogens.
Carrel has claimed to have
transmittedthrough filtrate passages
tumors induced by arsenic,tar preparationsand even indoles.Thesetumorswere of the Rous type obtainedin
fowl. More recently,similar tumors transmittedthrough filtrates were obs e r v e db y M c l n t o s ha n d S e l b i e( 1 0 9 ) , M a i s i n ,H a d d o na n d H a a g e n( l l 0 ) ,
and oberling and Guerin ( I I I ) after injection of methylcholanthrene.
especiallyin fowl, The considerationspresentedabove furnish a logical
explanationfor theseobservations.
A first factorto consideris the presence,
in animalsregardedas normal.
of a virus able to interveneto produce a neoplasticeftect under spe.cial
circumstances.
we have noted that such a virus can be presentwithout inducingtumors,Fowl appearto be especiallysusceptible
to viruses(l l2),
statisticsshowingthat viral lymphomatosesare responsiblefor SoVo of the
malignancies
in chickens.Even while some speciesdisplay an inborn reto viral infection,othersare higily receptive,as seenfor the viruses
sistance

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of sarcomas(l13) and lymphomatosis.

( I 14) viruseshavebeenfound in
as matryas l0%o and even20Vo of.chickens,accordingto somereports.

(2e4)

The number of animalswith virusesand no tumors must bc considered

still higher when presenceof viruses is revealed by antibodies.DuranReynals, in collaborationwith the East Lansing Agricultural Experiment
station (115), has shown that, while not one of 23 chickenskept isolated
and free of lymphomatosisshowedantibodiesin rhe blood, hundrcdsof
chickenstaken at random did have the antibodies.I his facr makesit highly
probablethat the presenceof thc virusesin the chickensused for carcinogenic studieswas independentof the adnrinistrationof the chemical carcinogen. Furthermore, the role of chemical agents acting alone as
carcinogenscan be discountedbccausctheir elfectsdiffer widely. In the
seriesof Mclntosh, the tumors appcarcdfar from the site of injection which
is very unusualfor methylcholanthrene.
Furthermore,thc agentsuscd by
Carrel,excepttar extracts,gencrallyhavelittlc or no carcinogenicactivity.
The two hypotheses----one,
that chemicalcarcinogensalone can induce
tiltrabletumors;the second,that this is only coincidenceand the tumor is
entirelyof viral origin----canbe rcconciledunder the conceptof plural intervention.Thus the chemicalcarcinogenwould induceonly part of the proccss; the remainder,at highcr levels,would result from viral intervention.
The changethat occurs in thc cells through the influenceof the chemical
carcinogencould also favor the changein the virus, making it not only
more activebut alsoo[ neoplasticcharacter.'l'he conccptof plural changes
neededto induce active carcinogenesis
permits us not only to integratethe
interventiono[ virusesin the conceptof carcinogencsis
in general,as presentedabove,but also to considerthis intervcntionin relation to other
[actors.
Hormones can play a part; they are neededto induce the degreeof
differentiation that is a condition for viral co-carcinogenicintervention.
Inoculated intraperitoncally,the Bittner milk factor, although active, will
rarely induce mammary tumors in virgin femalesalthoughthe virus can be
proved to be in the body. The hormonalchangesrelatedto pregnancyand
lactation influencethe mammary gland and cause a differentiation.Since
this diftercntiation representsa condition for viral neoptasticactivity in
these cases,hormonal interventioncan be integratedas an added factor
importantfor the viral carcinogenicactivity.Thc hormonal factor would
appearto be an indirect co-carcinogen,and it is under this aspectthat its
role in carcinogenesis
has to be studied.
The concept of plural co-carcinogenicinterventionpermits us not only

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to relatethe diftercntpathogenicfactorsinvolvedin carcinogenesis;

in addition, by relating these factors to various levels of organization,it allows
us to obtain new insight into the geneticfactor.

GENETICS AND CARCINOGENESIS

The geneticfactor in carcinogenesis
can be understoodin terms of phylogenetichierarchicdevelopment.Such developmentresultsfrom a seriesof
progressivechangeswhich lead to successive
hierarchiclevels of organization. As we have seen,for each level of the organizationa seriesof different solutionsare availablewhen a new hierarchicentity is to be realized.
This results first from the fact that various numbers of entities take part
in the constitutionof the principal parts. Since different constituentscan
form the secondaryparts of these new hierarchicentities,the number of
solutions is increased.The resulting solutions can be considered on a
statisticalbasis. Many of these new entities will die immediately; others
will subsistas such; still otherswill progess. Their fate resultslargely from
their interrelationshipand the conditions present in the environment in
which they find themselves.
The striking similarity to the resonanceprocessstudied in the lowest
lovelsof organization,such as atoms or molecules,has led us to consider
that changesat higher levels are of the same fundamentalnature. of all
the resonanceforms that occur at each of the levcls, there are some that,
on a statisticalbasis,persistand develop.Thesepersistingresonance
forms
make up the normal organism. The favored resonanceforms are determined by heredity and also by environmentalconditions.while the resonanceforms apPearon a statisticalbasis,the environmentalcondition can
vary and new resonanceforms will mark the intervention of external
factors. As a normal entity is composedof the pcrsistentresonantforms,
abnormalityoccurswhen suchan entity persists.The characteristics
of any
individual are provided by the resonanceforms which have developed
phylogeneticallyand also ontogenetically.These predominant forms are
"isotropic."For didacticpurposes,we callcdthe others"allotropic."
Allotropic ResonanceF orms
It must be acceptedthat, originally,it was the interventionof allotropic
resonanceforms which permittedthe appearanceof new forms able to respondwell to the environmental
changes.The phylogenetic
developmentof
different phylae, species,strains and even individuals, can be seen as
resultingfrom suchdifferentsolutionsfor the sameproblems.when, horv-

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ever, an allotmpic resonanceform appears,during ontogeneticdevelop_

ment, it resultsin an anomaly, At the level of genesor chromosomes,
it
producesa mutation or monstrosity.At a still lower level, such allotropic
resonanceforms may rcsult, not in monstrositiesor mutations, but in
cancerousentities. The concept of cancer as a hierarchically organized
diseaseaccordswith this view of allotropic resonanceforms. A first cancerous entity would, therefore, develop when an allotropic resonanceform
occurs at a low level of the organization.Undcr favorable cond.itions,the
allotropic entity would develop hierarchically,passingon through the different levelsof the organization,and realizingallotropic chromosomes,nuclei. cells and tissues.
In order to have an invasivecancer, it would thus be necessarythat
an entire successionof favorableconditionsbe presentinsuring the development of a continuousline of hierarchiccancerousentities.Thesefavorable conditionscan occur spontaneouslyat each level, and both cancerous
and normal entitiesmay have many allotropicforms.Carcinogenesis
would
correspond to the creation o[ these favorable conditions, and, as seen
above, it can take place at the diffcrent levels of the organization.The
result is a hierarchic successionof the persistingforms of the allotropic
seriesof respectivecancerousentities.
The long time usually necessaryfor a cancerouscondition to appear
accordswith this mechanism.The interventionof any external factor consideredcapableof inducing, by itself, the developmentof cancerousallotropic forms, has to be regardedas favoring the conditions necessaryfor
the persistenceand developmentof the successionof allotropic cancerous
cntities.(304)
This would also explain the relationshipbetween hereditary factors
and carcinogenesis.
Just as the individualhas the capacityto realizethe
successiveallotropic forms, so a strain or even a speciescan inherit the
tcndencyto developsuch allotropicforms.This explainsthe persistence
of
mutationforms. It appliesto the devclopmento[ strainswith high or low
incidenceof spontaneoustumors. It would also explain the vast differences
among different speciesand strainsin their responseto carcinogens.
The conceptof cancer as correspondingto a seriesof allotropic resonanceforms at the successive
hierarchiclevelsis of importancein terms of
the intervention of external factors. Such environmentalinfluencescan
establishconditions favorable for the developmentof ailotropic forms at
successivelevels, permitting the progressionof the allotropic line. The
inequalityof their action at differentlevelsaccountsfor the big differences

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x E S E A R C Hr N p H y s t o p A T H o L o c y

seen betweenthe various carcinogens.It is under this specialaspectthat

we saw above thc important problem of induced carcinogenesis.
The relationshipbetween carcinogenesisand the defensemechanism
can be understoodin terms of the differencesin defensecapacitiesof allotropic and normal entities.From the study of cancerousand normal subjects, it appearsthat the latter have the capacityfor defenseresponses.It
apPearshighly possiblethat the characteristicof the "normal" resonance
forms residesin their ability to resolvenoxious interventions.The allotropic forms lack this ability. The lossof the defensemechanismat various
levels,which is characteristicof cancer,can be regarded,up to a certain
point, as being the result of the interventionof allotropic resonancforms,
which would appear to be fundamentallyinadequateto oppose the hierarchic progressionof cancer.Incapableof respondingwith the full defense
mechanism when confronted by continuous noxious interventions,cancerousentitiesuse the simpler, primitive defenseforms, and especiallythe
lipidic prolongcd form. This lipidic predominancerepresentsthe principat
factor in producingthe actual manifestationsof cancer,with their dualistic
nature.
The interventionof noxious factors in carcinogenesisis well known.
Traumaand microbialviral infectionsin particularare suchfactors.Co-carcinogenssuch as croton oil, and somesolventssuch as benzeneand toluene.
can be considerednoxiousfactors. (295)
This view of a plurality of factors interveningtogether to realize the
hierarchicallycomplex conditionof cancerhas anothervalue. It helps to
reconcilevariouscxplanations
of the pathogcnesis
of cancer,eachattributed
to a different etiologicalfactor, and each basedon incontestableevidence.
Accordingto our view, with the possibleexceptionof the broad-scalevirus.
which leadsto the appearance
of cancercells in a couple of days. in all
other casesa number of factorsof differentnature intervene.To resonance
changes,would be added chemical,viral, metabolic,hormonal or defense
influences,at the sameor at differentlevels,in order to provide the necessary circumstancesfor cancerdevelopment,The fact that, regardlessof the
nature of agentsused to induce them, cancers,once induced, differ very
little or not at all makesplausiblethe hypothesisof plural exogenousfactors
actingto bring about the necessaryfavorableconditions.
The above presentation-a resumeof our 1sss21sh-muststill be consideredto be a working hypothesis.
Other aspectsof the cancer problem have been analyzedanew in the
light of the conceptof pathogenesis
discussed
above.

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Lipids and Cucinogenesis

As we have seenabove,the recognitionof the fact that a seriesof successivechangestake place in carcinogenesis
has invalidatedthe concept
that in order to considcra substanceactivein this field, it has to induce
the entire seriesof changesby itself.includingthe passageinto the phase
of invasivecancer.An agentcan be consideredactiveif it inducesonly a
part of the successive
scriesof changcs.Didactically,we can thus consider
changesconcerningthe subnuclear,nuclear,cellular and metazoiclevels.
We have investigated
the interventicln
of lipids in carcinogenesis
from this
specificpoint of vicw.
The coordinationof datafrom variousobservations
and experiments
has
indicatedthat some of the lipids would act especiailyat the cellular and
tissuelevels.
Statisticaldata have indicatcda greaterproportion of cancer of the
cervix in non-Jewishwomen as comparedto Jewish women. This was
tentativelyrelated to the circumcisionof the respectivemales and this
correlationwas confirmedby statisticaldata concerningother groups of
populationpracticingcircumcision,such as the Moslems.The probable
role of smegmawas seenin experiments
in animalsmadeby differentworkers. It was reportedthat in mice, smegma,sterilizedor non-sterilized,
introducedin the vaginaof mice followedby the sutureof the vagina,would
inducepapilomatousand cancerousIesionsof the cervix,
Statisticalstudies(324) showeda similar correlationbetweencancer
of the prostateand circumcision,with a lower proportion of cancer in
circumcisedindividualsthan in those not circumcised.Entirely different
resultswere obtained by other workers. Studiesmade by the group of
Memorial Hospitalin Cleveland,concerningcanccr-in-situ,
showedno differencesbetweenJewishand non-Jewishwomen.Similarly,severalworkers
have reportedthat the prostatesin individualsovc'r 40 ycars of age, who
diedof conditionsotherthan canccr,havepresentin high proportioncancerin-situcells.No correlationwith circumcision
could bc found in thesecases,
the same proportionbeing found in all the ethnicgroupsexamined.(325)
We tried to interpretthesetotally discordantconclusionsof the two
groupsof statistics
and found thc explanationin thc conceptof plural interIn the lirst group of statisticsin the casesof both
ventionof carcinogens.
femalesand males,the cancerousprocesses
considercdwere those in the
invasivecellular stage,while in the secondgroup of statistics,the noninvasivecancerphasewas considered.It appearsthus quite clear that the
interventionof the circumcisionand respectively
of the smegmaexistsbut

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has to be placedat a precisepoint in the progressive

changesin the evolution of the cancerouscondition,at the passagefrom noninvasiveinto invasivecancer.without any influenceupon the appearancc
of the cancer-insitu,the smegmawould act manifestlyby changingthe proportionof active
cancerpresent.Its influenceappearsthusto be exertedat the cellularlevel,
where the occurring changesresult in the passageof the noninvasiveinto
the invasivecancer,and at the tissuelcvcl wherethe lossof the capacityto
defend itself againstthe cancercells permits their invasion.This led us
to the hypothesisthat the cancerousprocessin the cervix and prostate,
evolvesindependently
of circumcisionuntil the cancer-in-situ
step,but will
make the next step toward invasivecancerpredominantlyunder the influenceof the smegmaof noncircumcisedmales.Without such intervention
at this stage,sucha changemay occur but in an impressively
lower proportion. The invasivecancerof the cervix is thus almostentirelynon-existent
in virgin nuns. This consideration
and the richnessof smegmain positive
lipids has permittedus to go farther and considerthe problem of the role
playedby thesclipids in carcinogenesis.
We have tried for a long time, but with litle success,
to inducecancer
in animalsthroughthe administration
of unsaponifiablc
fractionsalone.The
several positive cases obtained in mice with repeated injections of the
unsaponifiable
fractionsof placenta,chickencmbryos,eggsor butter,have
not appearedsufficientlyconsistentto warrantany conclusions.
In our attemptto investigatcan interventionof the positivelipids for
the specificchange,corresponding
to the passageof the noninvasiveto the
invasivecancer, we carried out the following experiment.we selected
femalemice with at leasttwo previouspregnancies.
After a third pregnancy,
when in lactation,the mammaryglandwas injectcdwith small amountsof
fractions.The resultsseemto indicate
the abovementionedunsaponifiable
a higherproportionof mammarycarcinomain thcscmice than in controls.
Similarly,we have tried to influencecellsof the cervix in mice. through
the introductionof the samedifferentpreparationsof unsaponifiable
fractions in the vagina of ex-breedermice, followed by suture of the vagina.
The first resultshave shown a high proportionof malignanttumors. The
has led us
conceptof the interventionof abnormallipids in carcinogenesis
to utilize in similar expcriments,insteadof the above preparations,unto be heterogenized
by beingheatedabove
fractionsconsidered
saponifiable
which
are in progressseem to
320"C. At the moment,theseexperiments
indicatethe existenceof such an influence.
It is of even greaterinterestto note the influenceexertedby the unfractionsupon animalsthat had receivedurethane,according
saponifiable

PRoBLEMS tN C^NcER

3O7

to the experimentsof Berenblum.By combiningthree factors, urethane

for the first changesin the amino-acids,
severalpregnancies
for the changes
until the noninvasivephaseand unsaponifiable
fractionsespeciallyheterogenized for the passageinto the invasivephase,a high proportion of invasive mammary and ccrvical canccrssecrnsto bc induced in the first
experiments.
Of importanceappearsthe factor that a certainlapseof time
is necessarybetween each two factors which are applied in the above
mentionedsuccession.
This concordswith the conceptof plural successive
changesin carcinogenesis
discussedabove.
CarcinogenicActivity ol Urethane
The interestingresearchof Berenblumhas brought an importantcontribution not only for the largelydebatedrole of urethaneas carcinogen,
but also for the problem of carcinogenesis
in general.The fact that croton
oil, applied to the skin, inducesthe appearanceof malignanttumors in
animalspreviouslyfed with urethane,concordslargely with the conceptof
plural changestaking place in carcinogenesis.
The ana.lysis
of the influence
exertedby carbamicacid upon amino-acidswould place the interventionof
tbis agentat the first membersof the biologicalrealm. It can thus be seen
that the bond betweenthe amino-acidgroup and the carboxyl and amine
Sroupsof carbamicacid occur in a way similar to that which occursbetween
two amino-acidswith the big differencethat in the first caseit would result
in tbe appearanceof the CNCN formation. (Fig. 201) As mentioned
above,this CNCN formation representsthe group which characterizesthe
first biologicalentity. The placeof this CNCN group, not at the end of the
moleculeopposedto the carboxyl as in the alkalineamino-acids,but as correspondingto the bond which rcsultsin polymers,representsthe anomaly,
which accordingto the work hypothesis
we advance,would correspondto
-[he
the first cancerousentity.
fact that thc specific activity of urethane
takes place at the lowest levelsof organization,explainsthe necessitythat
a certaintime separatesits interventionfrom that of croton oil, which would
act only at the higher levels, probably inducing the passagefrom noninvasive to invasive phase. This time is necessaryfor the first cancerous
changesto build up the seriesof canceroushierarchicentitiessince the
cocarcinogen,
croton oil, would act only in thosemore evolvedcancerous
entities.In experimentsin progress,the passageof the urethane-induced
noninvasive
cancerousentitiesinto invasivecancer,is successfully
obtained
by treatmentwith preparationsof unsaponifiable
fractionsof placentaor
eggs.

,,,giw'*ffi,

308

R E S F - A R c Hr N p H y s r o p A T H o L o o y

INTERVENTION OF PSYCHOLOGICAL FACTORS

IN CANCER PATHOGENESIS
The role of psychologicalfactors in the pathogenesisof cancer, although
still obscure,has beenof increasinginterestin recentyears.Various theoretical considerations(i.e. the relationshipbetweenthe known eftect of
emotionsupon hormonal and biochemicalbalancesand the possibleeftect,
in turn, of the latter upon neoplasrn^s),
as well as a number of clinical reports and experimentaland statisticalstudies,point toward psychological
influencein cancerpathogenesis,
but little is actuallyknown in this area.
In order to explore this matter further, a researchprogram has been
carriedon in our Instirutesince 1952 by Dr. L. LeShan.This study has
includcd the evaluationof projectivepersonalitytests given to over 300
c a n c e rp a t i e n t s i;n t c r v i e r vos f 2 t o 8 h o u r s e a c h w i t h o v e r 1 5 0 p a t i e n t s :
and extensivecxplorarorypsychorhcrapy
(of from 60 to 400 hours) for 25
patients.Control groups werc also included in sach category.For patients
undergoingpsychotherapy,regular comparisonswere made between the
personalitypicturcand variousbiochemicalactivitiesreflectedin blood and
urine analyses.
A "back-and-forth"methodbetweenthe threetechniquesof personality
evaluationhas bccn employcd.Hypotheses
formulatedfrom data obtained
with one techniquehavebeencvaluated,refinedand clarifiedby data from
the others.whcn a hypothesiswas consistently
supportedby all three approaches,an attempt was made to formulate it in tenns permitting it to be
subjectedto critical test by experimentalor statisticaltechnique.
As an examplc,an hypothesiswas dcvelopedthat the cancer patient,
more often than chancewou.ldallow, had lost a major emotionalrelationship, and had beenunableto find a satisfactorysubstitute,sometime before
the first apparentsympromof cancer.This hypothesisappearedto be validated by data from all three techniques.It was then formulated in terms
by which it could be tested.If the hypothesiswere true, then certain social
groups which, a priori. had known higher rates of such lossesshould also
have a highcr cancermortality rate. Thus, for example,if marital status
were taken as the only variable,then, after age was cancelledout, we
shouldexpectthe highestcancermortalityrate in the "widowed," the next
in the "divorced,"the next in the "married" and the next in the "single."
Publisheddata, such as censusmaterial,could be used to exptorethe accuracy of this prediction.various predictionsof this type-all based on
the hypoths5i5-ws1smade.When testedagainstpublishedstatisticaldata,
a l l w e r e d e m o n s t r a t etdo b e v a l i d . ( l l 6 , I l 7 )

P R O B L E M Sr N C ^ N C E R

309

At this point of the research,one general.cmotionalpattern has been

found in over 50o/cof the 300 studiedcancerpatientsand in approximately
lOVo of the equatedcontrols:An early life historywith much self-doubting
and someanxietyover relatingto others;the establishment
of one personal
relationshipthat affordeda high dcgreeof satisfaction,
meaningand validity to the individualand providedhim with a "raison d'tre"; and the loss
of this relationship,followcd by inabilityto find a substitute,and a period
of intense(if often concealcd)dcpression.
This has beenelaboratedupon,
and casehistoriespresentedin variouspublications.(118, l19, lzo, lzl,

t22\
In summingup this researchin a recentpapr (123), the following
conclusions
couldbe reachedby LcShan:
l. There seernsto bc a correlationbctweenthe existenceof neoplasticdiseaseand the persistence
of certaintypesof psychological
situations.
2. The most consistentlyrepnrted.relevantpsychologicalstate has
beenthe lossof a major cmotionalrelationship.Often the psychic
state resulting from this loss could be traced to a period shortly
beforethe first notedsymptomsof cancer.
3. There appearsto be somerelationshipbetweenpersonalityorganization and the evolutionof the cancerouscondition.
4. There may be some relationshipbetweenpersonalityorganization
and the type or location of a cancer.
It would seemas if future researchin this area,to bc as usefulas possible,must focusupon the chemicophysiological
changeswhich resultfrom
variationsin psychic statesin general.It is highly probable that these
changesare mediatedthrough the endocrinesystem.Through the linking
of psychic stateswith hormonal changes,we may be able not only to
integratepsychologicalfactors with the many other factors influencingdevelopmentof cancer,but also to relatcthcm to certainlevelsof organization. It may be possibleto establishthe relationshipof psychologicalfactors
to other influenceswhich favor or even inducepassageof cancerfrom one
phase to another. This may prmit a more complete understandingof
cancer and help in finding new points at which some therapeuticvalue
might be expectedfrom psychological
intervenrion.
The relationshipbetweenthe adrenalsand psychicstateson the one
hand,and betweenthe adrenalsand the lymphaticsystemon the other hand.
couldexplainwhy the inlluenceof a psychologically
unresolvedproblem is
most evident,among all cancerousconditions.in lymphomas.LeShanhas
been able, by analyzinga sizablenumber of these lymphoma cases,to

310

./

nEsEARcH rN pHysropATHoLocy

recognizemore clearly and more consistentlythan in other conditionsthe

existenceof a patternof psychological
changesoccurringprior to clinical
illness.Recurrences
of symptoms,or periodsof exacerbation,
also could
be connectedto eventswhich had deep repercussions
upon the psychologicalstate.
Researchalong theselines, seekinginformationon psychicfactors in
the pathogenic
mechanism
of cancer,is beingpursuedactivelyby LeShan
andhisco-workers
at our Institute.

C H A P T ELR2
PHARN,IACODYNAIUIC]
ACTIVITY
TT

we have investigated
l[* ou* REsEARCH,
the pharmacodynamicactivity
of a seriesof substances
in terms of thc physiopathological
conceptsdiscussedabove: the level of organizationat which thcy act, the dualistic
natureof their activity,their relationshipto body constituents,
especiallyto
lipids, and the changesthcy induce in the dcfensemechanism.We will
limit ourselvesin this presentation
only to thoscof the substances
investigated which are of therapeuticinterest.
The dualistic concept has pcrmitted didactic separationof agentsinto
two groupswith antagonisticproperties.In the last analysis,this separation
could be related to the two fundamentaltendenciesin nature, homotropy
or heterotropy.For inorganic agents,this criterion appearsto be directly
relatedto the elementspresent.However, for organic agents,especiallyfor
lipids and lipid-like substanccs,
this simplecritcrionis lessvalid. Another,
the positive and negativeelectricalcharacterof their active polar groups
could be used. It must be emphasized,however,that some of these polar
goups show either positivc or negative properties depcnding upon the
medium in which they work. An alcohol,for cxample,can act as an acid
under special circumstances,in which case it forms metal alcoholates;or
it can act as an alcoholforming esterswith acids.For this reason,in conwe havc limited the sphereof changesstudiedto
sideringthesesubstances,
thosecorrespondingto the conditionspresentin biologicalentities.
Since, as seen above, substanceswith negativc polar groups actively
while those with positivepolar groups conintervenein various processes,
trol tbe activity of the first, we will start this presentationwith the former.
3ll

312

xESE^RCHrN pHysropA't-HoLoGy

AGENTS WITH ACTIVE NEGATIVE POLAR GROUPS

Among agents which have negative polar groups, a further division
can be made accordingto the natureof thesegroups.Under biologicalconditions,the most important of negativepolar groups is the carboxyl. Losing
a proton when dissociated,the carboxyl confers acidic property to tbe
moleculeinto whose structure it enters.We will consider,first, such carboxylic acids. Among such agents,the individual difterencesseen in biological activitieshave to be related to their nonpolar parts.
In the frame of our research,with organic acids, we were especially
interestedin those with a predominanceof the nonpolar group; that is,
those having lipoidic proprtics.They are, principally, fatty acids, which,
accordingto the presenceof one or more double or triple bonds in the
nonpolar part, can be separatedinto saturatedand unsaturated.
F"t$

Acids

SaturatedFatty Acids
The lipoidic character in this homologousseries starts with the five
carbon valeric acid, although caproic acid is the first lipoidic member
found in animals.The role of the saturatedfatty acids in organizationis
largely related to the activity, as caloric metabolites,of the memberswith
even carbon numbers.These saturatedfatty acids are absorbed,circulated
and stored as triglycerides,and it is also this bond to glycerol which ap
parentlyfavorstheir biologicalrole as caloricmetabolites.
As previously noted, only the memberswith a relatively short chain,
lcss than 12 carbons, can be directly metabolizedthrough Knoop beta
oxidation.Longer chain saturatedfatty acids undergo a breakdown before
further metabolic degradation.Members with l6 and more carbons also
play a limited constitutional role, usually together with an unsaturated
member, when bound to glycerophosphoricacid to form phospholipids.
Even when frec-that is, whcn their polar group is not neutralized-they
do not have a manifestfunctional role and, consequently,show no important pharmacodynamicactivity.
The higher fatty acids exert no dcmonstrableinfluenceupon microbes
nor upon viruses such as bacteriophages.
No influencehas been observed
upon monocellularorganisms,cells or tissuesin vitro. No changeswere
seenin the respirationof liver slicesor of Sa 180 ascitescancerouscells in
the Warburgmicrorespironreter.
Similarly,no influencecan be seenon red
blood cells nor on leucocytestreated in vitro. No changesin the chloride

P H A R M A C o D Y N A M T CA C r r v t T Y

3 1 3

contentof tumorsor woundsare inducedby administrationof theseacids.

The fact that no changccould be induccdin the secondday wound crust
pH explainsthe lack of any effectupon pain.
Even largc doses,such as 20 cc. of a l0% solutionof theseacids in
sesameoil, do not changethc intensityof pain of alkalineor acid pattern
in humans.No influencehas been noted upon tumor evolution, even with
the direct techniqueof dipping tumor transplantsin the agent, repeated
for successive
transplants.Existingedemaor ulcerationis not influenced.
At the organic level, no changescan be seen in the function of various
organs or upon liver regenerationrate. These fatty acids do not influence
convulsionsinduced by thiamine in rats even when administeredin very
large doses,such as 5 cc. or more of l}o/o solutionsfor 100 gr. of body
weight. The same lack of influenceis seen on systemicmetabolicmanifestations,as recognizedthrough variousanalyses.Except for palmitic acid,
which showsan adrenaldefenseindex of 12, the index for the other members of this seriesis below 6, indicatingonly a certain participationof the
adrenalsin the defensemechanismagainsttheseacids.
No changescan be found in the number or characterof leucocytesin
animalstreatedfor a few days with theseacids.No changein the sodium
or potassiumcontent of the blood or in urinary analysesis induced by
thesesubstances
even in subjectswhoseanalysesrevealabnormalpatterns.
No action upon body temperatureand no influenceupon the evolutionof
the defensemechanismcan be observed.
It is interestinghowever,to note the effectof the sodiumsaltsof these
acids,especiallyon in vitro lysisof differentcells.(Note I )
UnsaturatedF atty Acids
Of the unsaturatedfatty acids,oleic acid is the most widely encountered
in nature.As previouslynoted,ntonoethenoids
have a group of 9 carbons
either toward the carboxyl or mcthyl end of the molecule.with the double
bond betweencs and cro in a moleculeof 18 carbons,oleic acid appears
to satisfyboth tendencies,which could accountfor the ubiquity of this acid.
oleic acid circulates,is depositedas reserve,and is used as caloric
metabolite especially when bound to glycerol. In considerably lesser
amounts,when bound to glycerophosphoric
acid, it takes part as a phospholipid in the formation of the lipidic systemof the organism.It is only
slightlyactive in oxidation processes.
For this reason,large dosesare necessary to influenceabnormal processcsat differentlcvels. Even then, only
limited changesare induced.In in vitro studies,oleic acid producesno
changesin bacteriophages.
A certain influenceis noted for receptivityof

;,lr1[arr,
Fil..:'S

314

,/

xESEARcHlN

pHysropATHoLocy

dermotropeviruses.Injected subcutaneously
in rabbits, oleic acid induces
in the skin, at the site of injection a zone of low receptivity for smallpox
virus. Oleic acid has a reducedeffect upon microbes,causingthe appearance of some gram negativeindividual forms of Bac. subtilis, for instance.
Mixed directly with blood, oleic acid induceshemolysis.when plasmais
treatedwith this acid, and then mixed with red cells.the influenceexerted
upon red cells becauseof the small amount of acid fixed upon plasma, is
reduced. Although olcic acid may influence the pH of the second day
wound crust, causingan increasedlocal alkalosis(Fig. II9), its influence
0leic actd

F t c . l l 9 . T h e s e c o n dd a y w o u n d c r u s t p H f o r o l e i c a c i d
s h o w st h e c o n s t a n tp r e s e n c eo f a c h a n g et o w a r d m o r e a l k a l i n ev a l u e s .

upon pain is almostnil. Little or no changeis scen in standardizedradiation woundsin animalstreatedwith the acid. A limited influencecan be
observedon tumor grorvthby using thc techniqueof dipping transplants
gencrations.Usc of a 1094.solutionof oleic acid in tricap
for succcssive
gcncrations,
impairsgrowth of
rylin bcforegrafting,rcpeatcdfor successivc
after thc sixth or sevcnthgenerationin
Ehrlich mouse adenocarcinonra
some experiments,even later in othcrs.Negativepassagetakes place betweenthe cighth and tenth graft. Under the samecondition,very little or
no changesare noted in other tumors,and no changesare seenin tumors
in animalstreatedwith this acid even thoughchangeshave been reported
of cells of differentorgans,treated
by some authors, (124) Suspcnsions
of oleic acid and injectedin animalsof
in vitro with colloidalsuspcnsion

P H A R M A C O D Y N A M T CA C T T V T T Y

315

the same specics,induced no changcswith a single injection.Repeated

injectionsaftcr 3 weeksinducedlcsionsin the respcctivcorgans.
At the organic level, some effectscan be obtaincd by using high doses
of oleic acid. For example,to preventconvulsionsinduced by thiamine
chloride,dosesas high as severalhundredmilligramspcr 100 gr. of body
weight are required and even then tbis eflect is not constantlyseen.Systemic changes,rccognizedthrough blood and urine analyses,are almost
ni-I,even with large dosesof oleic acid. The compoundhowever,does prolong liver regenerationtimc.
'fhe
Among the diethenoics,we studied linolcic acid.
caloric activity
of the uncombinedfatty acid is rcduccdwhile constitutionaland functional
activitiesare increased.Linoleic and linolcnic acid appear to be organizational rather than functional fatty acids becauseof their preferredbond
to glycerophosphoric
acid. They are absorbedfrom the intcstines,mainly
'I'he
as phospholipids.No effectsupon phage can be seen.
refractivity to
smallpoxvirus inducedon rabbit skin is more manifcstthan for oleic acid.
However, the effect upon microbes,such as subtilis,is less evident than
for oleic acid,Crenelatedred cells,with increased
tendcncyto conglutinate
and increasedsedimentation
ratc, are found altcr linoleictreatmentof the
blood in vitro. To avoid hemolysis,the acid is not added directly to the
blood but to the plasmawhich is then reunitedwith thc cclls. Crenelated
cells also appear in vivo in rats injectcd intraperitoncallywith large
amountsof this acid, such as l0 cc. of IO9h in oil. A definiteshift toward
alkalosisis found in the secondday wound crust pH, which explainsthe
pain of an alkalinepattern
influenceseenupon pain. Linoleicacid increascs
pain of an acid pattern,though only slightly.Only in relaand decreases
t i v e l y l a r g ed o s c s( 2 - 4 0 0 m g . / 1 0 0 g r . a n i m a l ) , d o c s i t p r c v e n tt h e c o n vulsionsinducedby thiamine.
In lesionsinduced by radiation,the administrationof smarlldosesof
this fatty acid often producesa favorablehcaling effect.This effect can be
related to the growth-stimulating
action of essentialfatty acids in small
just
quantities.The effectis
the rcverseof the unfavorableinfluenceon healing observedwhen largerdoscsare administered.
An effectupon tumors in
be
achieved
only
through
animals can
repeatedtreatment of successive
generations
transplantsor through the treatmentof successive
of the host.
The effect of repeatedinjectionsof tissuecells treated in vitro with a suspensionof linoleic acid was more manifestthan that obtainedwith oleic
acid. The effect upon the systemiclevel, althoughmore manifestthan for
oleicacid, still is limited,even undcr abnormalconditions.Blood and urinary analysesare only slightly and briefly changedtoward the D type of

,-t.f

316

R E s E , ^ R c l rr N p H y s r o p A T u o L o c y

offbalanceeven when large amountsarc administered.Blood eosinophiles

are decreasedand potassiumcontentincreased,both only slightly. Body
temperatureis slightly depressed.
Preparations
especiallyrich in tricthcnoidlinolenit'oc'idwere used and
no differencesbetween their bioloeical activitv and that of linoleic acid
could be noted.
We obtained preparationscspeciiilly rich in arachidonic rcid f.rom
salmon oi.l. The caloric contributionof this acid can be consideredalmost
nil comparedto its functional role. It is absorbed,circulated and stored,
principally as esterifyingsterols.Although this acid is presentin the body
in relativelysmall amounts,it reprcscntsmore than 25Vo of.the fatty acids
of the adrenals.In view of the highly functionalrole of the adrenals,it is
logicalto supposethat arachidonicacid'sabundancein the glands is not
merely coincidental.A libcrationof this acid, togetherwith other higher
polyunsaturatedfatty acids from the adrenalsapparsto take place during
the first phaseof the diphasicdefensephenomenon.
In the first minutesfollowing a noxiousinterventioninducingshock,a depletionof fatty acids in
the adrena.ls
occurs,coincidingwith increasedamountsin blood.
Besidesthcir role in the dcfensemechanism,the adrenal fatty acids
seem to intervene in normal physiology.Successiveliberations seem to
occur, alternatingwith liberationsof sterols,as weli as corticosterones.
Together,theseproducc the diphasicoscillationswhich characterizethe physiologic dynamic balance.
Preparationsrich in arachidonicacid secm to act at di-fferentlevels.
No manifestinfluenceupon phagesis observed.The influenceupon smallpox virusesand upon microbesis similar to that of the linoleic preparations.
The in vitro and in vivo effectsupon red blood cells, such as crenelation,
conglutinationand increasedsedimentationrate, are more manifest than
for linoleic acid. Leucopeniaalso is seen.The effcctsupon the pH of the
second day wound crust and upon pain iue, however, the same as for
linoleic acid. The intensity of acid pain is reduced while that of alkaline
pain is increased.In dosesmuch smallerthan for linoleic acid, arachidonic
acid preparationsare able to preventconvulsionsinducedby thiamine. But
they do not seem to changethe evolution of radiation lesions.The influence upon tumors is very similar to that of linolcic acid, and only limited
changes are observed after treatment of successivegenerations,either
through the dipping of transplantsor through treatment of successive
of this fatty acid were
hosts.Organ cells treatedin vitro with suspcnsion
organs,if injectedtwice at 3 weeks
seento inducechangesin the respective
interval.Systemicinfluenceis not manifesteven under abnormalconditions.

PHARMACoDYNAMTC ACrlvrTY

317

Blood and urine analysesare slightly and temporarilychangedtoward the

panern of fatty acid predominancc.
Continuing thesc studies,wc have investigatedpolyunsaturated
latty
acids with more than four double bonds, particularlyclupanodonicacids
from cod liver and sardineoils. Most of the studieswere made by using
the fractionswhich when brominatedarc solublein acetoneat a low tempera t u r e ,i . e . ,a r o u n d - 6 3 " C , T h e d i l T e r e nf tr a c t i o n so b t a i n e dw e r e i d e n t i f i e d
t h r o u g hi o d i n cn u m b e r ,n c u t r a l i z a t i ovn: r l u ca n d s p c c t r aal n a l y s i sa f t e rc o n j u g a t i o nt h r o u g ht r c a t m e n w
t ith KOH
All the biologicaleffectsupon viruses,microbcs,cells, etc., are more
accentuated
than for linoleicor evenarachidonicacid.Changesin red cells
and leucocytcsare much more apparcnt.At this point we must emphasize
the preferenceof thesefatty acids for red blood cells over plasma both in
vitro and in vivo treatmcnts.(Note 2)Thcy have pircatereffectsupon pain
than do linoleicand arachidonicacids,reducingpain of an acid patternand
exacerbatingpain of an a.lkalinepattern. 'fhe local pH, as determinedby
secondday wound crust measurements,
also is shifted toward increased
alkalinity.Convulsionsinducedby thiamineare influencedby much smaller
dosesthan those required with othcr unsaturatedfatty acids. For some
preparationswith iodinc indicesaround 350, dosesas low as 35 mg./100
gr. of body wcight arc sufficientto preventconvulsions.Changesin the
evolutionof tumors are more striking with thesepreparations,especially
when the transplantsof succcssivc
gcncrationsarc dipped in the preparat i o n . W i t h t h i s t e c h n i q u en, e s a t i v er c s u l t sa r e o b t a i n e de v e n a t t h e t h i r d
transplantfor Ehrlich mammary adenocarcinoma.
An obvious effect is
noted on radiation-induced
lesions,with ulcerationincreasedand healing
slowed.The influcnccupon cholcsterollevelsin the blood and upon hypertension is greater than for arachidonicacid. Thc effectsupon systemic
changes,obscrvedthroughanalyses,
are tcmporaryand no greaterthan for
linoleic and arachidonicacids.
Acid Lipidic Fractions
Bearing in mind the role of acid lipidic constituentsin the physiology
of variousbiologicalentities,preparationscontainingthesefractionswere
obtained.Tissues,organs,organisms,and organicproductswere saponified
and acid fractionssoluble in ether were isolated.we called them "acid
lipidic fractions," or "lipoacidic fractions." Their analysesrevealed,in addition to variousfatty acids,othcr substances
with lipidic and acid character.Some were identifiedas porphvrinicacids.
Significantdifferencesrelatedto the sourcesof thesepreparationscould

rfll:'-.
Rli

.ji

$ffi
;,**?#S1.'i,.

318

;''

RESEARcH IN

PHysIopATHoLocy

be recognizedin biologicaleffectsat differentlevels.Preparationsobtained

from intestine,for instance,had no obviouseffectson any of the manilestations; there were no systemicor organicchanges,no effectsupon pain, red
cells or leucocytesin vitro, and no influenceon tissue respiration.Treatment of successivetransplantsshowedno apparenteffects,even after ten
passagegenerations.No effectsupon organsthrough repeatedinjectionsof
cellstreatedin vivo by thesepreparationswere seen.

:6
a
c
,a

fn
aa

lave Lengthfnp.;

F l c . 1 2 0 . S p e c t r a la n a l y s i so f r a t l i v e r f a t t y a c i d sa f t e r c h e m i c a li s o m e r i z a t i o ns h o w s
t h e p r e s e n c eo f f a t t y a c i d s w i t h 3 a n d 4 d o u b l e b o n d s . ( 0 . 0 0 2 % i n e t h y l a l c o h o l )

On the other hand, other prcparationsfrom placenta,liver, blood, etc.

did show activity upon all manifestations,
including pain. They induced
negativeresultson tumor growth after transplantdipping for just two or
three gcnerations.
The differcncesin activity could bc relatcd to the richnessof these
preparationsin polyunsaturated
fatty acids.It could be shown that it was
not the total numberof doublebondspresent,as determinedby the iodine
number,that was significantbut rather the relativeabundanceof higher

;illl:

PHARMACODYN^MIC ^CTIVITY

3I9

c
o
g
c
L
ira

lave Lemth6n*;
F t o . l 2 l . S p e c t r a la n a l y s i so f t h e i n t e s t i n a lf a t t y a c i d so f r a t s a f t e r c h e m i c a l i s o m e r i z a t i o n s h o w s m i n i m a l a m o u n t so f m e m b e r sw i t h 3 . 4 o r m o r e d o u b l e b o n d s .( 0 . 0 0 2 V o
in ethyl alcohol)

unsaturatedmembers, as rccognizedby special analysis after adequate

c h e m i c acl o n j u g a t i o nF. i g u r e s1 2 0 , l 2 l a n d 1 2 2 s h o w s u c ha n a l y s e s .
Abnormal Fatty Acids
Becauseof the role of abnormal fatty acids in the pathogenesis
of the
offbalancetypc D, particularlyrelatedto radiation,a study was made of
preparationsof acid lipids obtainedfrom abnormaltissues,organsand organisms.In an initial group of researches,
animalsthat had died of radiation sickness,shock, acute infections or after adrenalectomywere used.
Guinea pigs infectedwith B. anthracisand mice infectedwith strep hemolyticus were used as sourcesof abnormal lipids in a large number of experiments.Acid lipids obtainedfrom autolysatesof tissueswere employed.
We also used the fatty acids of cod and sardineoil, which may be consideredto correspondnot to natural but to alteredfatty acids sincethey were
obtainedafter autolysisof cod liver and whole sardinebodies.
After having determinedthat conjugationof double bonds is the basis

.ffi{r',
ffii,j

. $qi- jj![t

320

RESEARCH IN

PHYSIOPATHOLOCY

"l
;

*1
I

i*t
t

I rol
I

I
6u

fb

8c

f a v e L e n g t h( n p )
Frc. 122. Spectral analysisof the fatty acids of the entire bo<ly of rats, after chemical isomerization shows the presenceof di- and triethenic members, and littlc of
m e m b e r sw i t h m o r e d o u b l e b o n d s . ( 0 . 0 0 2 V oi n e t h y l a l c o h o l )

of abnormality in pathogenicfatty acids, conjugatedisomers of different

fatty acicisw'erepreparedand studied.Eleostearicacid obtainedfrom tung
oil was used extensivelyin animal and clinical research.Parinaric acid
was obtained from nuts of parinarium laurinum and was used to a lesser
degrce.various conjugatedfatty acid isomers,recognizedthrough spectral
analysisand oxalic indices, were obtained in a higher proportion from

ffi,-N

PH^RMAcoDYNAMTcAcrrvrTY

321

different lipoacidrcpreparationsby using a modificationof classicalmcthr.. (Note 3) Nlany mixtures of naturally occurring fafty
ods of conjugar..c
acids found in sapcnifiablefractionsu'cre conjugatedby the same method.
Conjugated di, tri, tetra, penta-and hexaenic fatty acids werc further
separatedfrom tl::sc itii:turesand studied,The unexpectcdrelationshipof
t h e s ec o n j u g a t c Jf , , l : ' , ' a c i d tso o x y g e nr v a so f i n t e r e s tW
. hilethe treatment
of linoleic acid at 37"C with oxygcn lcads ro progressiveincreasein the
amountof peroxidespresent,this doesnot occur for the conjugatedisomer
(FiS. 123) apparentlybecauseof repeateddestructionsof the peroxides
formed.
In general,the effectsproduced by conjugatedfatty acids at difterent
levels of the organizationare more intensiveand persistlonger than those

t2

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Frc. 123. Curves of peroxides of samples of linoleic acid (a) and its conjugated
isomer (b) induced tbrough the passageof oxygen ( 100 ml per minute for 30 cc of
sample) at 37'C. While peroxides are progressivelyincreasing i n t h e l i n o l e i c a c i d
prcparation,they do not change in the conjugated isomer,

322

n E S E , A R c Hr N p H y s r o p A T H o L o c y

of the nonconjugatedisomers.It is interestingto mention here the in-fluence

exertedby thesepreparationsupon viruses.Although there are no changes
for bacteriophages,
a marked influenceis seenin vivo upon receptivity of
the organism to viruses.Subcutaneousinjection of conjugatedfatty acids
in rabbits establishesa zone of refractivity toward inoculationof smallpox
vaccine in the skin, which is greater and more persistentthan that observedfor nonconjugatedisomers.The effect upon microbesalso is clear.
Gram negativestrainswith manifestmorphologicalchangeswere obtained
from B. anthracis,and persistedas such for 6-15 passages
beforethe old
morphologicaland tinctorial charactersreturned.The effect upon red cells
and leucocytesin vitro is more apparentthan for correspondingnonconjugated isomers.This is also true for the influenceupon the respiration in
vitro of tissuesor ascitescells.
The difference in the effects of conjugated and nonconjugatedfatty
acids is particularlystriking in certain manifestations.
For example,pain
with an acid patternis more easilychangedto alkalineby treatmentwith
conjugatedfatty acids than with nonconjugatedisomers.Once the alkaline
pattern appears,it persistsfor a long time. A manifesteffect is seen upon
lesionsinducedby radiation.Standardized
radiatedwoundsin rats, which
heal in about four weeksin control animalsand require six to eight weeks
to heal when treatedwith nonconjugated
fatty acid preparations,fail to
heal at all when treatedwith correspondingdosesof conjugatedisomersof
the samefatty acids.
The isomers of fatty acids also differ in their effect upon animals in
shock. When anaesthetizcd
animalsare scaldedby immersionup to the
level of the xiphoid in water at 90oC, immediatefatal shock occursif the
immersionlastsfor more than four scconds.With a three secondscalding
the animalsdie after severalhours.The administrationof fatty acid preparations markedly reducesthe survival time, and this effect is more manifest
for conjugatcdmembers,especiallyfor eleostearicacid. In general,animals
in shockinducedby any means,suchas by the Noble-Collipdrum, show a
specialsensitivitytoward conjugatcdfatty acid preparations.
The effectof conjugatedfatty acids at the systemiclevel, as recognized
by blood and urine analyses,is in the same direction as that for the nonconjugatedbut is more manifest.
There are significantdifferences
betweenthe effectsof conjugatedand
nonconjugatedfatty acidsupon the evolutionof transplantedand spontaneous tumors. In a small proportion of mic, a mixture of nonconjugated
fatty acidspreparedfrom cod livcr oil or sardineoil preventsthe appearance of methylcholanthrene-induced
tumors, The conjugated fatty acids

PHARMACODYNAMIC ACTTVITY

//

323

obtainedthroughthe treatmentof thesepreparationsshow this preventivc

effectin a large proportion of mice.Theseexperimentsarc interestingfrom
severalstandpointsand are presentedin somedetailin Note 4.
Conjugatedfatty acids producean incrciuc in the chloridecontentof
woundsin animals.Values are 407o highcr than in untreatedlcsions.The
same increaseof chloridesoccurs in tumors. When Dba mice with adenocarcinoma were treatcd for ten days with conjugatedfatty acid preparations, and tumors were removedand chloridecontentdetermined,values
were up to l80o/ohigherthan for controlswith untreatedtumors.(Note 5)
An interestingeffectwas noted in two rat tumors.For years,passages
of
Guerin'srat tumor and of sarcomainducedin our laboratoryby the injection of benzpyrene
have showna peculiarcharactcr.Althoughthey grow to
large size,the tumors have no necroticzones.After treatmentwith conjugated fatty acids,large necroticzonesappeared,leadingto ulcerationand
death.The appearance
of thesezoncsof necrosiscorresponds
to changesin
the fundamentalcharacterof the tumor. Transplantsof fragmcntsof thesc
treatedtumors,althoughtakenfrom nonulcerated
rcgions.or youngtumors,
resultedin tumors showingearly central ulceration.This charactcrpcr'l'he
sistedwithout further treatmentin succceding
gcncrations.
ulcerating
tumors appearto be a mutant of the originaland the nrutativechangecan
be relatcd to interventionof conjugatedfatty acids.This was confirmedby
the fact that similar changeswere constantlyobtainedin the same tumors
with thc sameagens. A similar but lessmanifestand less constanteffect
is obtainedwith preparationsof cod liver oil fatty acids administercdrepeatedlyin largeamounts.The samceffectwas obtainedwith the injection
of the fatty acidsdirectly in thc tumor itself.
The secondday wound crust pH showsmarkedchangcstoward alkalinity under the influenceof conjugatedfatty acids.The eflect upon regeneration time of liver is also manifest;cells full of fatty dropletsdo not
appear at all or appear much later than in untreatedcontrols. In rats
weighing200 grams,with a largeenoughdose,such as 2 cc. of l0% solufatty acidsobtainedfrom cod liver oil and repeated
tion of polyconjugated
for two days,the adrenalsshow completedepletionof fats. They become
small and red in color and contain no sudanophilmaterial.The liver rcgenerates
with smallcellswith compactcytoplasmand almostentirelybare
of fatty droplets.
The effectof conjugatedfatty acidsupon the lymphaticsystemis manifest.A marked involutionof thymus,spleenand lymphaticgland follows
particularlyof a mixthe injectionof conjugatedfatty acid preparations,
tureof conjugatedcod liver oil fatty acids.The effectupon tumorsis irregu-

324

xESEARcH

IN

PHYSToPATHoLoGY

lar. In some animals a marked retardationoccurs,in others no effect can

be observed.In convulsions,no greatereffect is seen than that produced
a color darkerthan when
by nonconjugated
isomers.Blood in vitro assumes
other fatty acidsare used.Eosinophilcs
arc markcdly rcducedafter administrationof conjugatedfatty acid preparations.Important changesin organs
were obtained with repeatedinjectionsof the respectivecells treated in
vitro with suspensions
of theseacids.Changesin the analyticalvaluesof
urine, however,are not greatly differentfrom those obtained with nonconjugatedisomcrs.With sufficientamountsof conjugatedfatty acids,a frank
hypothermiais obtained.
Bixine
In the samegroup of fatty acidswe can place bixine, a member of the
polyterpene
family with 9 conjugated
doublebondswhich we obtainthrough
saponificationof the seedsof Bixa orellana and have studied widely. The
changesproduced in microbesare similar to those with other polyconjugated fatty acids. Changesin connectivetissuesin animals appar to be
particularly interesting.The first reaction to subcutaneousinjection of an
oily solutionof I Vo bixtne in rats or mice is an inflamntatoryprocess,with
the injected material dividing into hundredsof tiny droplers.Some, however, melt away and the unabsorbedinjectedmaterial again forms one or
two big drops. The wall containingthe drops is very thin and transparent
and appearsto be made up of very few conxectivecells which have extremely long fibrils, rcpresentingthc highestdcgreeof differentiationof
thesecells.
The effect upon pain is similar to that of conjugatedfatty acids. The
secondday wound crust pH showsa manifestchangetoward alkalosis.The
effectupon radiationwoundsis the sameas for conjugatedtrienes.In animals injccted with convulsantdosesof thiamine, only a few milligrams of
the bixine preparationare requiredto preventconvulsions.Of all the fatty
acid preparationsused, bixine apparsto be the most efficientin its anticonvulsivantaction.The iodinenumberof 430, found in our preparations.
confirms,thus, in this case too. the correlationseenbetweenanticonvulsivant effectand richnessin double bondsof the fatty acids.
The distributionof bixine followingadministrationis interesting.Chromatographicstudyof blood constituents
after hydrolysisshowsthat almost
all the bixine is in the red cells,with minimal quantitiesin plasma.Lesions
such as wounds or tumors, after the administrationof bixine, become particularly rich in this substancein comparisonto normal tissues.Changesin
evolution of tumors also are manifest.The administration of this agent

P H A R M A C O D Y N ^ M I C ]^ I ' T I V I T Y

325

often leadsto rapid necrosisald cdenra.In aninralsand huurans,we saw

massivetumors becomeulceratcdin a fcw days after use of only a few
milligramsof the substance.
The ulceratingcffectof conjugatedfatty acids
u p o n t u m o r sr e a c h e si t s m a x i m u n rw i t h b i x i n e .l n a n i m a lt u m o r sw h i c h ,i n
s u c c e s s i vt er a n s p l a n t sh,a d n c v c rs h o w ns p o n t a n e o uusl c e r a t i o n st h, e i n j e c t i o n i n t o t h e h o s t o f o n l y a f e w m i l l i g r a m so f b i x i n e i n o i l y s o l u t i o n
produced,in addition to ulceration,a changein the tumor which can be
c o n s i d e r e dt o c o r r e s p o n dt o i l m u t a t i o n .F u r t h e r t r a n s p l a n t cs o n s i s t e n t l y
developedulceratingtumors and the ulcerativecharacterpersistedin succccdinggcnerations.
Massivedegencrating
changeswereobtainedin organs
after repeatedsubcutaneousinjcctionsof suspcnsionsof cclls obtained
frorn theseorgansand treatedin vitro with bixine.
The changcstoward oflbalanceD induccdin varioussystcmicpatterns
in humans by bixine are sinrilar to those producedby conjugatedfatty
acids.However,once induccd,thcscchangesare very persistent,
and often
remain uninfluenced
by anti-fattyacids.It is this characteristic
of resistance
t o f u r t h e rc h a n g e sw h i c h r c p r e s e n tas c e r t a i nh a n d i c a pf o r t h e r a p e u t iuc s e
of this agent.
Fudittg Response
I n c o n t r a s t o t h c r e l a t i v e l yp e r s i s t e ncth a n g e si n d u c c db y b i x i n e ,t h c
s t r i k i n gc h a r a c t e or f t h e e f f c c t su p o n p a i n o r s y s t e m i cn r a n i f e s t a t i o no sb tained with fatty acid preparations.and especiallywith the conjugated
i s o m e r s ,i s t h e i r s h o r t d u r a t i o n .F u r t h e r m o r c a, t t h e b e g i n n i n go f t h e i r
administration,theseagents,evcn in relativelysmall doses,exert intense
eflects,but such eflectscannot bc obtainedlatcr without continuouslyincreasingthe amountused.Aftcr a certaintime. cvcn largedoseshave very
little effect.
An explanationof the fadingcharacterof the resultsobtainedwith these
and many other agentscan be found in thc fact that the organismdefends
itself againstany factor able to inlluenceits balancc.In the caseof conjugatedfatty acids,this defenseseemsto bc providedmainly by the adrenal
glandsE
. . F . T a s k i c rh a ss t u d i e dt h i s a s p e c ot f a d r c n a d
l c f e n s ci n o u r l a b oratory and this researchis presentedin Note 17, Chapter 6. An adrenalectomizedanimal is usually less rcsistanttoward the administrationof
animal.This drop in resistmany agentsthan a normal or sham-operated
ance is expressedas an adrenaldefenseindex, as the ratio betweenthe
minimal lethal dose for the normal and for the adrenalectomized
animal.
preparations.
this index is betweentwo and three,and
For most fatty acid
greater
conjugated
for
the
fatty acids.Fatty acidswith three conbecomes

326

R E S E A R c Hr N P H Y S T o P A T H o L o c Y

jugated doublc bonds, however,have an adrenal defenseindex of 120.

Eleostearicacid is 120 times lesstoxic for sham-operatedcontrols than for
adrenalectomizedanimals, indicating a specific adrenal defense against
these acids. Progressivelyincreasedintervention of the adrenals would
explain the fading effect mentionedabove.
Alpha Hydroxy Fatty Acids
Alpha hydroxy fatty acids were obtained by fixing an OH at the carbon
adjacentto the carboxyl. Some of these acids exist in nature-in significantamountsin the brain and skin, and in vcry small amountsin the
kidneys. In our research, they were originally prepared from natural
sources,such as animal brain and skin. Most of the studieshowever,were
made with syntheticalpha-hydroxy-fatty
acids.For experimentalpurposes
in animals and humans,we principally used pure syntheticalpha hydroxy
fatty acids.Mixtures of thesemembersobtained through the treatment of
acid lipidic preparationsalso were cmployed.
In animalsand humans,alpha hydroxy fatty acids induce lesssystemic,
organic, tissue and cellular changesthan do thc correspondinguntreated
acids. However, we would like to mention a striking exception: the responseof lymphosarcoma
6C3HED in mice to the administrationof alpha
hydroxy-caprylic
acid. Althoughthis tunlor uniformly kills control animals
within 10-12 days, it disappearsin ovcr 8O7oof animalstreatedwith alpha
hydroxy caprillic acid, even if treatmentis institutedlate, that is, when the
tumor has alreadygrown to I cm. in diameter,a size usually rcached 2 or
3 days beforedeath.In the few animalsin which the tumor does not disappear,its growth is so sloweddown that survival time is extendedto three
or more weeks.(Note 6)
Other alpha hydroxy fatty acidscloseto caprylic acid, such as alphahydroxy-caproicand capric acids,show no influenceupon evolution of this
tumor. We could not obtain similar effectswith any of the other saturated
alpha hydroxy fatty acids that have chains with 4-20 carbons, nor with
alpha hydroxy-olcicor linolcic acids.Nor did alpha hydroxy-caprylicacid
or any of its homologuesappear to have any influenceupon other transplanted tumors in mice, the Walker tumor in rats or, spontaneousmammary tumors in mice.
Other Fatty Acids
The fading eftect secnwith naturally occurringfatty acids is so great a
handicap for therapeutic use of these substancesthat we searched for
fatty acids which the organismdoes not normally encounter
heterogeneous

PHARMACoDYNAMIC ACTtVTTY

327

and against which it would not be prepared to defend itself. This brought
us to the study of fatty acids having different nonpolar groups than those
of the normal and abnormal constituents.while most of these were prepared syntheticallyin the laboratory, we utilized on a substantiallylarge
scale two natural fatty acids which exist in plants and are sufficientlyheterogeneous,ricinoleic and crotonic acids.
Ricinoleic acid has a double bond between9 and l0 and a hydroxyl at
carbon 12 insteadof the seconddoublebond found in linoleicand linolenic
acids. As a result of the induction effect propagatedfrom the carboxyl
through the chain, the crr is a positive carbon. The positive characteris
enhancedby the adjaccntdouble bond betweence and c1e, and by the
hydroxyl bound to c12.C,, thusis very stronglypositive.we relatedthe intenselocal alkalosiswith consequent
watcr cxcretioncorresponding
to the
a l k a l i n ew a t e r yd i a r r h e at o t h c c f f e c to f r i c i n o l c i ca c i d l i b c r a t c di n t h e i n testine,and have consideredit to correspondto a local organicoffbalance
similar to that induccd in tissuesby unsaturatcdfatty acids.This would
explainthe intensivelaxativecflectof castoroil.
We uti.lizedricinoleic acid parenterallywith the aim of obtaining a
similar effcct in abnormalcellularand tissuelesions.The oily solutionof
ricinoleicacid has low toxicity when administeredparcnterally.However,
no manifesteffcct upon tumors or at differentlevelsof organizationwas
obtained.Crotonicacid did not showthe expectedinfluenceat theselevels.
Other hcterogeneous
agents,polyhydroxy fatty acids, were studied.
They were preparedby adding one or morc OH groupsto the nonpolar
groups at the double bonds of unsaturatedfatty acids.9, l0 dihydroxy
and 9, lO, 12, l3 tetrahydroxyfatty acidswere no differentthan thc corresponding unsaturatedfatty acids in their effects upon pain or systemic
analysesin humans,or upon tumor growth in animalsand humans.
Peroxide Fatty Acids
Peroxidesand epoxidesof fatty acidswcre preparedand studied.They
showedmore manifesteflectsupon virusesand bactcriain vitro and in vivo
than the other fatty acids. Investigationsof the effectsof these fatty acids
upon higher levels havc been limited until now. It seemsthat the effects
upon systemicand organic manifestationsare somewhatdifferent than
those obtained with use of the fatty acids from which the peroxidesand
epoxideswere derived. Influenceupon pain and upon tumors was grcater
for the correspondingunsaturatedfatty acids. This research---cspecially
with polyepoxidefatty acids-is still in progress.

328

R E S E A R C H

I N

P H Y S T O P A T H O L O G Y

Halogenic Compounds ol Fatty Acids

The study of changesin lesionswhereabnormalfatty acidsare present
has shown the importanccof fixation of chloride ions in thesesubstances.
Consideringits place in the periodicchart, chlorine is an elementwith heterotropic character.consequently,it could be conceivedof as being antagonisticto fatty acids,counterbalancing
their homotropiccharacter.This
was confirmed by pharmocologrcalstudy of fatty acids to which chloride
ions were added at the double bonds. We were particularly interestedin
conjugatedfatty acids in which effectsof treatmentwith chlorine could bc
followed through spectralanalysis.
when a mixture of conjugated fatty acids from cod liver oir was
treated with chlorine, the peaks in spectral analysis progressivelydisap
peared.(Fie. I2a) This did not lead,as expected,
to increasedtoxicity.Thus
far, in early trials, the differentpreparations,from 9-10 dichloro-stearic
acid to the polychlorinatedmixture of fatty acids from cod liver oil, have
not shown effects greatly different from those of conjugated fatty acids
upon tumor evolution,systemicanalyses,and pain. No differencehas been
noted betweenthe derivativesand their correspondingfatty acids in animal

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h a y e L e n o t h( b N

F r c . 1 2 4 . I n f l u e n c ee x e r t e db y c h l o r i n e8 , a su p o n t h e c o n j u g a l e df a t t y a c i d s .A p a r a l l e l
d e c r e a s ei n t h e a m o u n t o f a l l t h e m e m b e r si s s e e n .( 0 . 0 0 2 % i n e t h y l a l c o h o l )

P H A R M A C o D Y N A M T c^ C r l v l r Y

3 2 9

cxperimentsor in rescarchin hunrans.Experiencewith these products,

howcver. indicatesthat they may inducc milliar gastriculcerationswhich
u'e considerto rcsult from the inllucnceexertedby the fatty acids upon
the gastricmucousmembranewherethey are broughtby the chlorideions
to which they arc strongiybound. Researchin this directionconfirmsthe
part which thesefatty acids,solidly bound to chloride ions, take in the
pathogenesis
of the stateof shock.
An over-allanalysisof the pharmacological
activity of the fatty acids
mentionedabove showsa similarity in fundamentaleffectsobtainedwith
most of thesepreparations.
This can be interpretedas resultingprincipally
f r o m t h e f a c t t h a t a l l h a v ei n c o m m o n ,t h e i r l i p i d i cc h a r a c t ear n d t h e s a m e
polar groupwith acid properties-the carboxyl.As a result,thesesubstances
are fixed in the samepositionin abnormalentities,a fact which seemsto
representthe most importantfactorin their pharmacodynamic
activity.The
further biologicaldifferenccs
secnbetweenthe inlluenceexertedby the various membersstudiedwould be rclatedto a secondaryeffectof thesesubstancesresuJting
from the interventionof the nonpolargroups.This finding
-that the fundamentalpharmacological
activityof fatty acidsis connected
rvith the sitc of activity which is detcrmrnedby the lipoidic characterand
the polar group presentwhilc' the proper pharmacological
activity is due
to the interventionof the nonpolar group-has bcen of capital importance
not only for undcrstandingthc activity of thesesubstances
but also for
determiningthe path of our further research.Becauseof this, we investig a t e d ,i n a s e c o n ds t e p ,l i p o i d sw i t h o t h c r n e g a t i v ep o l a r g r o u p s .

LIPOIDS WITII OTHER NEGATIVE POLAR GROUPS

Lipoaldehydes.'
With a carbonylas polar group, the lipoidic properties
appearwith propanalin the homologoussericsof the aliphaticaldehydes.
with the carbonyllessdissociated
than the carboxyl,the lipoaldehydes
rep
resentnegotivelipoidsableto act for a longertime than the respective
acids.
We were especiallyinterestedin threegroupsof aldehydes.In one, with a
nonsaturatedshort nonpolar chain, we searcheda conjugatedformation
betweenthe double bond of thc oxygenof the carbonyl and the double
bond of the nonpolar chain. Another goup of the lipoaldehydescorrespondedto long chain fatty acids,while the third was formed b1'saturated
s h o r tc h a i n a l d e h y d ew
s i t h a n o d d n u m b e ro f c a r b o n s F
. rom the energetic
point of view, there were the first and especiallythe third groups which
appearedas the most interesting.ln the last group the opposite influ-

330

nESEARcHrN pHysropATHoLocy

ence exerted by the carbonyl and methyl groups upon the intennediary
carbonsof the chain givesthe entire moleculean especiallyhigh reactivity.
This oppositeinfluenceis seenat its maximum in propanal,where cL suffers the influenceencrgeticauyoppositeof the carbonyl and methyl group.
The fact that, due to its relativelyhigh solubility in water, propanalwhich is a lipoid--{an be administeredin aqueoussolutionsand still act
upon the lipidic system-makes of it an especiallyinterestingagent.
we have investigatedthesegroups of lipoaldehydesfrom the point of
view of their influenceexertedupon the two offbalances.In the group with
unsaturatedshort chains, we studicd acrolein and crotonic and malcic
aldehydewithout seeingany specialeffect upon the other levels than the
cellular one, where a vacuolationwas obtained.Furthermore their toxicity
has represented
an handicap.More interestinghas appearcdthe group of
the saturatedshort moleculeswith odd number of carbons. while with
heptanalwe havc obtainedbesidesan influenceupon pain, also a manifest
inhibitoryeffect upon thc growth of expcrimentaltumors, it was propion
aldehydewhich has shown the most interestingcffectsupon pain.
This was secn for the group of aldehydeswith aliphatic saturated
chain such as propionit' and heptylic aldehl,tlesor with cyclic, as .rclicy'lic
aldehydes.Inadequatedoscs-f rom ,urocc. to 2 cc. of the l0% solutionof
propionicaldehyde,or of the I % solutionof the heptylicor salicylicaldchydes-a manifcstinlluencewas obtainedupon the systemiccondition as
well as upon pain. Patientsin offbalanceA with pain and generaldiscomfort, wcre seento have a decrease
of the symptomsaftcr the administration
of propionicaldehyde.The eflectupon tumorswas reducedand propionic
aldehydedid not changethe cvolutionof the tumorsin spiteof the marked
improvementof thc gencralconditionand even of the ccssationof pain.
Lipoids n'ith Thiol Groups
Mercaptans: According to thc systematizationof lipoids presented
above, a thiol group, acting as a polar group, will form a lipoid when
prepondcrantaliphaticor cyclic nonpolargroup.
bound to an cnergetically
In thc homologousrncrcaptanseries,evcn thc lowestmembersare lipoids
forcesof the thiol group.
becauscof the weak electrostatic
Although methyl mercaptanis a lipoid accordingto our classification,
this substanceis too volatilc to be used.Therefore,the first low membcr
was ethyl mercaptan.
of this homologoussericsto be investigated
The effectsof ethyl mcrcaptanupon microbeswere more limited than
those seen for fatty acids. To determinethe effectsat the different levels

PHAR,M^CODYN^MIC ACTIVITY

331

of the organization,ethyl mercaptanhad to be administeredparenterally.

As for all other membersof this scries,we utilizedethyl mercaptanin 5 or
lOVo oily solution in cottonseedoil. In acute toxicity tests,the lethal dose
was found to be 145 mgs./100grams of body weight for mice, and 153
mgs./100 gramsfor rats. The immediateeffectsupon nuclei were similar
to, but lessintensethan thoseof conjugatedfatty acid preparationsleading
to caryorrhexisor pycnosisin abnormal cells. No abnormal mitosis was
seen in organswith high mitotic activity.such as the intestinalmucosaor
bone marrow, although appreciablechangeswere observedin mitosis in
animal tumors. A secondaryeffect,an exaggerationof aging processes,

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F l c . 1 2 5 . S e c o n dd a y w o u n d c r u s t p H s h o w sa c o n s t a n tc h a n g et o w a r d m o r e a l k a l i n c
v a l u e s f o r a l l t h e s u b s t a n c e sh a v i n g s u l f u r i n t h e i r p o l a r g r o u p .

was first recognized,severaldays after administrationin the changesin

granulocytesat the site of injection.The averagenumber of nuclear lobes
of the leucocyteswas often very high, even above seven.This was also
true for leucocytesin circulating blood when the product was injected
intraperitoneallyor even subcutaneouslyin rats. The immediateeffect of
such injectionswas a prolongedleucopenia,especiallya lymphopenia.
The chloride content of tumors and wounds was especiallyincreased
through treatment with mercaptans.At the tissular lcvel, local pH of the
second day wound crust was increased,an effect characteristicfor all
l i p o i d sw i t h s u l f h y d r yal s t h ep o l a rg r o u p .( f i 9 . ; , 2 5 ) T h i sc x p l a i n st h e d i r c c t
effectsof ethyl mercaptanupon pain and othcr alkaline or acid symptoms.
Theseeffectsare qualitativelysimilar to thoseseenfor the polyunsaturated
fatty acids but much slower to appear.While placentaacid lipid prepara-

332

nESEARcHIN PHYStoPATHoLocY

tions, for instance,produce an effect upon pain-an increasein intensity

for an alkaline pattern and a decreasefor an acid pattern----venwithin
5 to 8 minutes after parenteraladministration,the effect of mercaptansis
reducedand appean after half an hour, or later.
The eftect upon tumors in animals was especiallymanifest upon a rat
sarcomaoriginally induced by benzpyrenein our laboratoriesand passed
through successivetransplantsover a period of many years. Throughout
this period,this tumor showedl00Vo positivetakes,characteristicallygrowing to huge sizr, at times as large as the rest of the animal, but without
ever showing either spontaneousregressionor necrotic areas.When ethyl
mercaptanwas injected subcutaneously
in daily doscsof t/z cc. of a SVc
solution in cottonseedoil into animals with these tumors, interesting
changeswere observed.If the tumors were alreadylarge, above 6 cm. in
diameterfor instance,only a few regresscd(5 /20 for tumors of 6 cm. in
diameter).In tumors that did not regress,large necroticareasdeveloped
and were followedby ulceration.Most becameinfected,leadingto death.
Tumors smaller than 6 cm. in diameter frequently regressedrapidly and
then disappeared(between9/20 and 20/20 in difterentexperiments).No
such striking results were observedin any of the other tumor strains in
rats or mice treated with ethyl mercaptan,although in several,growth arrest occurredor necrotic zonesappeared.Ethyl mercaptaninjected in the
tumors themselvesinduced the same necrotic changesin most animals.
( F i s .1 2 6 )
The effect upon lymphaticorganswas manifest.Spleen,thymus and
lymph nodes were involuted in animalstreated for a few days with mercaptans.The eftectupon convulsionswas irregular.Even with small doses,
convulsionswere preventedin some casesbut, in general, the effect was
lessconstantthan with fatty acids.Eosinophilesdecreasedrapidly in animalsor humanstreatedwith this substance.
All urinary analyticaldata were
influencedby administrationof mercaptan,with changestoward the patterns of type D offbalance.
Becauseof the very disagreeableodor of ethyl mercaptan,we were
obligedto discontinueits useso that effectsof this agentupon some analyses,suchas surfacetension,sulfhydryland calciumexcretion.could not be
studied in humans.
Other Mercaptans:Superiorhomologuesof the mercaptanserieswere
used.They were divided into three groups.The first includedpropyl, butyl,
amyl and hexyl mercaptans;the second,heptyl and allyl mercaptans;the
third, memberswith more than l0 carbons.The first group producedmuch
the sameeffects,which tendedto diminish as the number of carbonsin-

P tt A R N{AC ()t)Y N A N{| ('

AC T I V I TY

1 1 1
-t .1 .1

creascd.For examplc.cflectswerc considerablyrcducedfor hexyl mcrcilptan as comparedwith ethyl mercaptan.Thc secondgroup-thc heptyl and
'I
allyl mercaptans-procJucedmorc intensivceffccts. his was espcciallytrue
for allyl mercaptan.Membersof the third group-rvith longercarbon chains
such as dodecyland hexadccylmcrcaptans-producedcflccts so slight that
ttrey seemedalmost nonexistent,exceptupon pH of the secondday wountl
crust, which showed valucs far abovc the rangc of thc controls, just as
with other membersof this homologousscrics.
Yesks

Control

ooo
Start treatoent

Treated
rl
ethyl
ercaptan

Treatedrith
ethyl
mcaptan

OOOI
Start treatnenl
U l s e r arto n

e O - .
i;.

F t c . l l 6 . l n f l u c n c e c x c r t e d h 1 ' c t h r ' l n r c r c a p t i u l u p ( ) n l \ : r r c o n l i r r n d t r e c t lb l , h e n z pyrene. For biggcr lumor\ jt induccs constltntly ln ulccration, while for rmall tun)ori,
thcir disappearance.

With the idea of usingthiol as a polar group and havingrnothcr ccntcr

in the molecule &s a sccondirrycenter, we studiecla seriesof othcr sub(the B A L preparation) oftcn used
stances.One was dimercapto-propanol
for heavy metal poisoning. [t proved to be conrplctcly rvithout cffect on
pain, tumor grorvth and systcmicchanges.It had lcss activity than the
highermercaPtans,
*'hich theniselves
rvcrclessactivethan prlyunsaturated
and abnormalfatty acids.
The difficultiesencountercdin adnrinisleringmcrcaplans,nrainly relatcd to offensiveodor of lou'er mcnrbcrsand inactivityof thc lessobnoxiousmembers,lcd to a scarchfclrothcr chemicalagentsthat rnightbc activc
without being evil smelling.Extensivestudy n'asmade of variousprcpara-

334

RESEARcHrN pHysropATHoLocy

tions that appearedto have a bivalent sulfur bond at the nonpolar group.
We investigatedcolloidal sulfur which, if introduced in the organism,
seemedto undergo changessimrlar to those of bivalent sulfur. We found,
thus, that the sulfur absorbedaftcr being administcredin colloidal form,
was almost entirely eliminatedafter oxidation in the form o[ sulfates.In
animals,no pharmacodynamicinfluenceswere observed.OnIy the pH of
the secondday wound crust was increasedwhen thesepreparationswcre
administeredparenterally in suspensionor were given orally mixed rvith
food, in proportions up to 4%. There was no evident change in tumor
evolution in animalsor humans.
Hydro-Persulfides
Another sulfur compound,so-called"sulfurizedoil," in which sulfur
and fatty acidsappearto form a hydropersulfide,
(Note 7) was tested.This
hydropersulfidepreparation,althoughit exhibitedno influenceupon viruses
in vitro, did inducea good degreeof local resistanceof the skin to smallpox
infection.The effectsupon microbeswere reduced.There was little direct
influenceexerted upon cells. The preparationswith 0.5 to lVo srtfur
bound to cottonseedoil were well toleratedlocallv when administeredintramuscularlyor intraperitoneally.
The effect of parentera.land oral hydropcrsulfrdeupon pain was slow
to appear,in contrast to the effect of fatty acids and even of mcrcaptans.
However, it persistedfor a long time. Pain of the acid pattern was eased;
pain of alkaline pattern was exacerbated.The influenceupon the second
day wound crust pH was marked.The local pH increasedto valueseven
higherthan 7.80. In radiationlesions,the dimensionof ulcerationincreased
and healingwas retardedor even prevented.In some tumors in animals,
the rate of groMh was slorved.This latter effect was not uniform in the
different types of tumors tested and even in the same type of tumor in
different groups o[ animals. systemic changesalso varied. Doses correspondingto 5 mgr. of sulfur were not toxic for 30 grn. mice in a single injection.Nor were l5 mgr. dosesin 200 gr. rats. chronic toxicity studies
showedthat 0.2 mgr. daily injectionsin mice and 5 mgr. injectionsin rats
for as long as three monthsdid not inducepathologicalchanges.High doses
suchas I to 2 cc. of a lVo preparationinjectedseveraltimes a day in humanswas almost uniformly followed by a rise in temperature,usually after
a few davs.

PH^RM^CODYN^MIC

ACTIVTTY

335

Other Compoundswith a Thiol Group

The resultsobtained with hydropersulfidepreparationsled us to seek
other compoundswith sulfur bound to fatty acids insteadof triglycerides
as in those mentioned above.
Sulfur compounds were prepared from various conjugated fatty acids
such as conjugatedlinoleic acid and eleostearicacids and from mixtures of
conjugatedfatty acidsobtainedfrom cod liver oil, fish oil, human placenta,
blood and various organs.while active in smalleramounts,they were not
qualitatively difterent from hydropersulfideprcparations obtained from
cottonseedoil, producing the same pharmacologicaleffectsin most tests,
especiallyupon pain, systemicmanifestationand evolution of cxperimental
tumors.
The fact that sulfur bound to the nonpolargroup,as in hydropersulfides,
produced lcss manifest results in animals and humans than mercaptans,
which have a thiol group as a polar group, led to the study of other substancesin which thiol radicalsinsteadof sulfur were added in similar positions,and consequently
wereconsideredto act as secondaryenergeticgroups.
preparations,
A seriesof
in which one or more thiol groupswere fixed at the
double bonded carbons in various conjugatedor nonconjugatedpolyunsaturatedfatty acids,were obtained.Thesesubstances
differ fundamentally
from the fatty acids mentionedabovein which sulfur atoms were fixed not
at the carbons bound by double bonds but at the carbon adjacent to
the double bond. 9-10, dithiostearicacid, 9, 10, 12, 13, tetrathiostearic
acid, as well as polyunsaturatedand conjugatedfatty acids with thiol
groupsfixed at their double bonds, were obtained.In general,they showed
no marked biologicaleffectson animals,no influenceupon pain or systemic
patternssimilxl to those observedfor the other lipidic productswith bivalent sulfur.
The hydropersulfidegoup was addedto soaps.Sodium and ammonium
soaps were obtained through saponificationof the triglyceridesof fatty
acids on which sulfur was already fixed. Effects at the different levels of
organizationwere markedly reduced.There was no influenceupon pain,
organic or systemicmanifcstations.However, a striking eftect was noted
on many microbes.Growth of Bac. anthraciswas preventedin someexperiments,even with dilutions of. | /2,000,000. For staphylococcusaureusand
streptococcushemolyticus, a similar effect was obtained with dilutions
higber than l/200,000. In animals,even oral administrationin drinking
water in a dilution of 1/500 and, in someexprimentseven l/1000, con-

336

p . E S E A R c Hr N p H y s t o p A T H o L o c y

trolled infectioncausedby thesemicrobes.The antibiotic activity appeared

to be similar to that of penicillin.(Note 8)
7'etrahydronaphthalene
Persulfides(SulfurizedTetraline)
we utilized the known marked tendencyof tetrahydronaphthalene
ro
fix oxygen with the resultingexplosiveperoxides,to fix sulfur in similar
combinations.Persulfidesof this substancewere thus obtained and their
pharmacologicalactivitystudied.white only a limited effectwas noted upon
viruses and microbes, the influence upon tetrahymena pyriformis approached that seen with active polyunsaturatedfatty acids. The producr
with 5 gm. sulfur fixed for 100 gramsof tetrahydronaphthalene,
showeda
relativelylow toxicity in normal animals,T5 mgr./100 gr. in mice and
125 mgr./100 gr. in rats beingwcll toleratedin intraperitoneal
administration. The influenceupon wound healingin animals,upon pain, and upon
the systemicanalysesin humans,was similar to that seenfor the mercaptans.Although the immediateeffecton pain was limited, prolongedadministration was effective.The analyticalchangesof the urinary surfacetension
and blood potassiumwere the most manifest.The toxicity was highly in,
c r e a s e df o r a n i m a l sw i t h a s c i t e st u m o r s ( S a 1 8 0 , E h r l i c h a n d K r e b s ) .
However,the influenceexertedupon transplantedand spontaneoustumors
in animalswas one of the most favorableones, compared to the effect of
other testedagents.
Thiosullates
We investigatedthiosulfates with the intention of studying agents
which, in addition to a manifestreductioneffect,would act througb bivalent sulfur liberated in the body. The elimination of part of the sulfur of
thiosulfatesas mercapturicacid has led to the suppositionthat the bivalent
sulfur ion, separatedfrom the thiosulfateion, would act through combinationssimilar to those found in the metabolismof thiolipoids.It is for
this indirect action that although hydrosoluble,without any direct connection to lipoids, we investigatedthe biological activity of thiosulfate
together with and under the same specific aspect as the lipoidic sulfur
compoundsconsideredabove.
There was a very limited effect upon microbesand viruses,no effect
upon phages,and no changein the receptivityof rabbits to smallpox virus.
With either oral or parenteraladministration,the immediate effect upon
pain was manifest.Oral administrationof 1/z cc. of a lOVo solution of
sodium thiosulfate in water, or intramuscularinjection of. l5 cc. of the
4o/o solution,was usually followed by a definite effect upon pain in less

PH^RMACODYNAMTC^CTlvtTY

33'1

than | 0 minutes.As this was found to be oppositefor the two patternsof

pain, sodiumthiosulfatewas usedcven for diagnosisof pain pattern.Pain
of an alkaline pattern increasedwhile a decreaseoccurred in pain of an
acid pattern. The second day wound crust pH increasedmanifestly with
the use of this substance.
Cellular and nuclear changesfollowing administrationof thiosulfate
preparationswere similar to those produced by mercaptans.The effects
upon the lymphaticsystemand on liver regcneration,
however,wcre minimal. Convulsionsinduced by thiamine were controlled well by thiosulfate
in dosesof 120 mgr. per 100 gram of body weight. Injectedsimultaneously
with administration
of thiamine,thiosulfatepreventedconvulsionsin a high
proportionof animals(17/20).
The effect upon radiation lesionswas lcss manifest.An increasein
wound size and prolongedulcerationoccurredonly with use of relatively
large amounts of sodium thiosulfatcdaily. Dosesabove 4O mg./100 grn.
of body weight were needed to obtain thesc efTccts.The hcaling of a
simplewound was retardedonly with largc doses,around50 mg./100 gm.
of body weight.On the other hand, when very small doseswere administered, such as 5 mg./100 grn., the healing effect was cnhanced.Effects
upon tumorswere lessmanifestin animals.Slightand inconsistent
changes
were seenin graftedtumors. Very often in the sameexperimentalgroup,
tumors disappearedin some animalswhile in others the pgowthrate was
only slowedor remainedunchangcd.The erratic resultson tumors in animals producedby thiosulfateswcre similar to thoseseenfor many of the
sulfur preparations,and appearedas characteristicfor this group. Repeated
injectionsof thiosulfatesin tumors were seento induce the disappearance
of the tumor if growth was slow cnough to permit such injections for
severalweeks.At the systemiclevel,the most markedeffect,other than that
on sulfhydryl index, was on surface tension which usually dropped with
the administrationof a sufficientlylarge amount.
Most of the researchon thiosulfatewas done with sodium salts.In a
few cases,very high dosage,such as 6-10 grams daily, produced moonfaceand slightleg edema,apparcntlyrelatcdto sodiumretention.This disappearedwith cessationof the medication.
Changing the cation of the thiosulfatc from sodium to magnesium
appearedto increase,sometimesmarkedly,the resultsobtainedin our experiments.Potassiumthiosulfate seemedto be more effective,especially
againstpain. Its use however, has been limited by disadvantages.When
administeredparenterally,it causesconsiderablelocal pain at the site of
injection as most potassiumsalts do. lf administeredto patients having

338

xESEARcH IN pHYSropATHoLocy

pain or other symptoms of an alkaline pattern or systemic manilestations

correspondingto type D, a more markedincreasein intensityof symptoms
occursthan for the sodium or magnesiumsalts.
We also investigatedsodium tetrathionate.Except for lower dosagerequirements,no other advantageswere found in its use. Its relative instability is a handicap.
Alpha-T hio-F atty Acids
In other studies,we tried to introducesulfur into the fatty acid molecule,this time changingthe polar group itself. With the sulfhydryl replacing
the hydroxyl of the carboxyl group, a bivalent negative sulfur was introduced, thus realizing a thionic group. (R-COSH)
We prepared severalmembersof this thionic acid series corresponding
to various saturated,polyunsaturatedand even conjugatedfatty acids. we
studied in particular the efiect of hexylthionic acid, correspondingto
caproic acid. The resultsobservedwcre essentiallythe same as those seen
with the other bivalent sulfur containinglipoids mentioned previously.In
addition to influencingpain and systemicchanges,hexylthionic acid produced some interestingeffects upon cxperimental tumors, reducing the
growth of a few of them. However, there were no important differences
from the effectsof the other sulfolipoids.
Another entire seriesof productswas prepared by introducing a thiol
goup at the Cz, or alpha position, of various fatty acids, with the intention
of creating a more complex polar group similar to that present in alpha hydroxy or alpha amino compounds.Alpha-thio-fatty acids were thus obtained for the entire homologousseriesof saturated,and for many of the
nonsaturated,fatty acids. Some membersof this seriesof alpha-thio-fatty
acids,such as caproic,caprylic and myristic, were studiedextensivelyboth
in animals and humans.From the biologicalpoint of view, however,they
showedno manifestdifferencesover the thiolipoidspreviouslydiscussed.
Thioglycolic Series
All these researcheswith limited biological results brought us to the
study of lipoids in which the thiol representsa polar group but in which a
secondarypolar center is present in the molecule. Many such synthetic
thiolipoidswere preparedin our laboratorywith the hope that they would
prove biologically more effective and would have alkylating activity as
well. Two seriesappearedto be interesting,since they were being active
particularly at lower levels oi organization.This led us to utilize them also
on a larger scale in clinical work, While consistentresults were obtained

PH^RM^CODYN^MtC

^CTIVITY

339

on pain and systemicchanges,the influenceupon animal tumors was erratic

and no different from that of other preparationswith thiol groups or the
sulfur compounds mentioned above. There were marked effects in some
animals with tumors; in others with the same tumor treated identically,
there were no effects at all. In humans the eftectson pain, tissular, organic
and systemiclevels were similar to those of many other sulfolipoids.
Starting with these substances,derivativeswere prepared. One group
comprisedderivativeswith a specialcharacter.In order to have only one
active polar group, one of the two polar groups had to be blocked. For
thioglycolic acid which we studied,either the thiol or the carboryl group
could be blocked,leavingthe uncombinedradical as the active polar group.
Since we were interestedin substanceshaving the thiol group as active
polar radical, the carboxyl group was blocked by replacing its hydrogen
with a methyl group. Methylthioglycolatehas been thoroughly studied in
our laboratory. Its pharmocologicalactivity is similar to that of the other
thiol preparationsmentionedabove.
other thioglycolateesterswith ethyl, propyl or butyl insteadof methyl.
were preparedand studiedbut showedno advantageover the methyl ester.
we tried to obtain the allyl esterin order to have a more potent secondary
centerbut we were unableto synthesizeit.
In the same group of agentswe studied another substance,beta mercaptopropanoicacid, having a thiol and a carboxyl as polar groups.uscd
uncombined,it could be seenthat here again,as with thioglycolicacid, it
is the carboxyl that acts as active polar group while the thiol acts as a
secondaryenergeticcenter at the nonpolar group. This acid is very toxic
in animals,producingas a peculiareffect,manifestmuscularspasms.The
compound also produced abnormal muscular rigidity. seen immediately
after death. In nontoxic doses,it showeda marked influenceupon tumor
growth. Many tumors disappeared;in many others,a reduction in size occurred. Impressiveresultswere obtained in spontaneousmammary carcinoma in mice where a fairly high proportionof tumors disappeared(2g/
40). Repeatedinjectionsinto theseanimalsgavegood resultsif the gowth
was slow enoughto pcrmit treatmentfor a periodof at leasta month. The
preparation,however,showedtoxic effcctsin animalswith tumors, producing weight loss similar to that producedby the thioglycolicseries.To
change the polar group and have the thiol act as such, we blocked the
carboxyl with a methyl in some experimentsand with an allyl group in
others. But the influenceupon tumors in animals was reducedthroueh
thesechanges.

340

nEsEARcH rN pHysropATHoLocy

Relationship to Sullur Metabolism

The study of lipoids with sulfur posedthe problem of the relationship
betweentheir structure and biological activity. we mention this here because it not only explains the influenceexerted by these agentsbut also
becauseit indicatesthe manner in which this researchhas had to be developed. We have seenthat, accordingto the systematization
of the elements,
sulfur representsa nonmeta.lanti A element,active especiallyat the metazoic level.It is its actionas an isolatedelementthat appearsinteresting,in
additionto the metabolicchangeswhich it inducesin the organismwhen
presentin a negativelipoid.
It seemsthat the organismgenerallymetabolizesbivalent negativesulfur by changingit ultimatelyto the hexavalentform. Ferguson and du
Vigneaudhave studiedthe metabolismof methionineand cysteinewhich
are the principalsourcesof sulfur in the organism.(125, 126) While the
evidencewe have on this subject is too limited to provide more than a
working hypothesis,it would indicate that other compounds with lipidic
.h313s191-1hs thiol-containing lipids-h3vs more important biological
activity.
The metabolismof thesecompoundsvaries among individuals,especially thosewith abnormalconditions.The study of the excrerionof sulfhydryls through the urine, expressedas the sulfhydryl index, has servedus as
a guide in studying the metabolismof sulfur up to a point. Thc high excretion of the thiol group seemsto be related not only to low oxidation
but to an abnormal form, as mentionedabove, since with the exaggerated
excretion,the thiol level in the blood is reduced.This form is to be consideredas an excrementalone. This seemsto be confirmedby the fact that
the administration
of bivalentsulfur,evenin a largeamount,is not followed
consistentlyby its eliminationin the form of thiols in the urine. In some
subjects,an impressivelyhigh proportion of the thiols administeredappearsin the urine, and this is true even for relativelysmall dosesof thiols
or of bivalentsulfur as in thiosulfates.
In other subjects,on the contrary,
even when larger amountsare administered,
the increasein eliminationis
minimal or does not occur at all. The abnormalityin sulfur metabolism,
which appearsto be a limited capacityto oxidize it to the hexavalentpositive form, also means an exaggerated
interventionof the thiol group as
such in the economyof the organism.This occurs along with symptoms
and signs, previously noted, correspondingto an exaggeratedoxidative
interventionof fatty acids.in which processes
the thiols probablytake part.
We tried to study the capacityof the organismto fully oxidize thiolic

PHARMACODYNAMIC
ACTtvrrY

341

sulphur by following the responseto the administrationof a known amount

of sulfur in bivalentnegativeform. Thc changeof the sulfhydrylindex of
urine would serve as a tolerancetest for thiol metabolism.
After injectionsof 80 mg. of sodium thiosulfate,the difterencesin the
capacity of various organismsto metabolizeit could be seen and related
to pathological conditions. This concept of thiol metabolismcan be the
basisfor understandingan abnormal form of thiolic sulfur which may be
involved in the pathogenesisof abnormal conditions.Substancescontaining a thiol group,suchas methionine,cysteineand particularlyglutathione,
are presentin sizableamounts in the organism,but it is not this form of
thiol that intervenesin the abnormalmetabolism.A large amount of the
normal form of thiol is presentin the blood of subjectswith a low urinary
sulfhydrylexcretion.when another form, the abnormalone, intervenes,
it is
excretedin the urine. The organismeliminatesthis "abnormal" compound
with sulfhydryls.It seemsquite probablethat this abnormal thiol compound is in a lipidic form since the sulfhydryl-containingcompound is
readily extractedby ether from the urine. Its affinity for the lipidic system
would explain the influence exerted upon fatty acids and the oxidative
processes
occurring in thc lipidic system.The thiolipoidsintervenecatalytically in the oxidation of the fatty acids,as seenin experimentsin vitro.
Thus, the thiol group in lipoids containingbivalent sulfur rather than
metabolizedsulfate would increasecatabolic metabolism. Although the
thiol in this abnormal form is largely eliminatedby the urine, apparently
as a defensemechanismagainstits pathologicalactivity, someof it is probably retained in the cellular lipids where it continuesits activity. Circulation of sulfur in thiolic lipidic form, with consequentimpairment of its
changefrom the bivalent negativesulfur into the hexavalentpositivesulfur,
would thus appearas the fundamentalsourceof the participationof sulfur
in the abnormal pattern. The influenceexertedby administrationof thiosulfatesupon the sulfhydryl index can serve as an indication for these
specific changes.
Sulfur is an anti-A elementand it is activeas such in all the forms in
which it existsin the organism,althoughthe intensityof its action varies
at differenthierarchiclevels.The activityof thiol as an anti-A factor can
be relatedto the influenceexertedby carcinogensor other agentsupon the
biological activity of this radical, especiallywhen it is taking part in the
formation of enzymes.This relationshipexplainsthe resultsobtainedby
repeatedinjections of organ or tumor cells treated in vitro with agents
having a thiol as polar group. The heterogenizationinduced leads to the
of severechangesresultingfrom the allergicreaction,
appearance

342

xESEARcH rN pHysropATHoLocy

It is interestingto note that sulfur has an anti-A tendencyeven in the

hexavalentpositiveform in which it appearsas sulfate.The sulfateion has
a capacityin the organismto inactivatelipoids of a positivecharacter.The
sulfate becomesbound to such substances,
thereby tending also to facilitate their excretion in urine. Many of these substancesare eliminated in
combinationwith the sulfuric radicalin the forms called "sulfo<onjugated."
In view of this, the effectof sulfateions appearsto parallel that of the
fatty acids.Both opposesubstances
having a positive polar character;that
is, both are biologically antagonisticto anti-fatty acids. In the sensethat
they opposeantifattyacids,both the bivalentnegativeand hexavalentpositive sulfur havc anti-A activity, the first directly and the last indirectly.
The characteristic influence exerted by sulfur in its different forms is
basedon its action as an agent inducing changestoward increasedhomotropy, by acting at levels above the cellular. This is a typical example of
the relationship between an element'sactivity and its place in the periodic
chart. Sulfur is a member of the serieswith homotropic action; it belongs
to the period which correspondsto the metazoic compartment and thus
acts above the cellular level.

Selenium Lipoids
The influence exerted by bivalent sulfur upon oxidation processesin
which fatty acids participate,servedas a guide for further research.Seeking agentsthat would act at a still lower levcl of the organization,even
below the cclls,we consideredother substances
which also affectoxidation
processes.Theoretically,at least, it appearedpossibleto induce changes
at a compartmentbelow the metazoic,at which sulfhydryl-containingcompoundsact.
We have discussedpreviouslythe systematization
of the biologicalactivity of elements,their fundamentalanti-A and anti-D influence,their distribution among the various levels of the organism, all related to their
atomic structureand their place in the periodic chart.
All of this led us to investigateseleniumwhich is the nonmetal element next to sulfur in the sixth seriesof Mendeleeffsperiodic table-the
serieswith an anti-A characterin which oxygenis the first member.According to its period, seleniumbelongsto the cellular compartment.
The first problem was the nature of the compound in which it would
be active.We wcre particularly interestedin using bivalent negativeselenium becauseof the activity of bivalent negativesulfur. However, we did
investigatethe selenicand seleniousacids.Theseacidsor their sodium salts

PHARMACODYN^MIC

^CTIVITY

343

have limited effect upon viruses and microbes. An interesting effect is

seen in Tetrahymenapyriformis,where a manifestcellular vacuolization
is induced.The influenceupon tumors, pain, organic and systemiclevelsis
less manifest and toxic effects are great. Thereforc, we prcpared lipoid
compoundswith a predominantnonpolar group and with a negativebivalent selenium.We utilized, on a larger scale,hexyl and heptyl diselenides
synthesizedin our laboratoriesby M. Bier,
Hexyl and Heptyl Diselenides
Studieshave shown that hexyl and heptyl diselenidesare lower in acute
and chronic toxicity than selenicand seleniousacidsand their sodium salts.
In wounds and tumors, these selenium preparationsinduce a relatively
limited fixation of chlorides,the increaseabove controls b"ing only about
l6Vo. In only a very few casescould any direct effect upon pain be observed within a few hours. However, the long range effectsafter several
daysof administration,seemedto be superiorto thoseof various fatty acid
preparations.This was true both for the decreasein intensityof pain with
an acid pattern and the increaseof pain with an alkaline pattern. The
eftectspersistedfor many days. In animal tumor experiments,there were
relativelyslight changesin growth or survival time,
No manifest influence was seen at the organic level. with relatively
large amountsof theseagents,an involutionof the lymphaticsystemwas
obtained. Thymus, lymph glands and spleen were markedly reduced in
sizein animalsdying after acutetoxicity tests,and adrenalswere small and
appearedto be depletedof their sudanophiliccontent. In rats, a frank
lymphopeniafollowed administrationof largesdosesof thescpreparations,
and eosinophilopenia
also was uniformly seen.Changcsin urine analyses
also were obtainedwith high doses.
It is noteworthythat administration
of disclenide
to a subjectwith a type
A pattern inducesthe appcaranceof oxidizingsubstanccs
in the urine, as
first
one of the
changes.
Effects at the cellular level are seen even with microgram dosages.
Vacuolizationoccursin the cellularcytoplasm.It is interestingto note that,
despitethe cellular vacuolization,pericellularedemaoccurs.The fact that
theseselcniumcompoundsare active in smail dosesmay be an indication
that they act entirely at the cellular and not the metazoiclevel. This effect
at the cellular level is confirmedby the fact that almost constantlythe administrationof seleniumif in sufficientamount is followed by a manifest
increasein serum potassiumvalues,and a decreasein the amount in rcd

344

RESEARcHIN PHYSIoPATHoLocY

cells.This changein serum potassiumis apparentbefore any other change,

and is generallyobtainedwith relativelyvery low dosesof selenium.
The effectsupon cells of another lipoidic selenium preparation, with
seleniumthis time as the polar group, warrant mention. The preparation,
synthesizedin our laboratory, is hexylselenoicacid in which the hydroxyl
of the carboxyl group has been replaced by a SeH radical. A manifest
eftect is produced by this agent in animals with ascitestumors. Intraperitoneal injection leads almost constantly to the disappearanceof such
turnors,even if the compound is used after ascitesis already present. We
usedthis product to bind seleniumin vitro to cancerouscells as will be seen
below.
Tetrahydronapht halene Perseleni de
The fact that in the first phasesof the defensemechanismthe organism
usesfatty acids acting largely through their productsof oxidation, has directedus-as we have seenabove-to searchin the therapeuticapproach
for agentshaving as pharmacodynamic
activity an interventionof peroxides.
parallel
peroxides,
a
further
we investigatedsimilar prodIn
step,
to lipidic
ucts in which, insteadof peroxides,persulfideswere present,sulfur being
the elementimmediatelyabove oxygenin the Vlth seriesof elements.We
studied thus the persulfides,among which the tetrahydronaphthalene
persulfide has been an interestingcompound. Its activity was explained, according to the biological systematization
of the elementsin which oxygen
correspondsto the organismlevel, while sulfur representsa metazoic element.Following the sameline, we searchedsimilar compoundsfor selenium
-3n slsrnentstill higher in the VIth series,which correspondsto the cellular level.We thus preparedand studiedperselenides
by boundingselenium
to tetralinein the sameway as was done for oxygen and sulfur.
The eftectsof the perselenideson microbes or animals were similar
to thoseof the other seleniumpreparationdiscussedabove. Tetraline perselenideshowedlow toxicity in animals,rA cc. of the lOVo solution of the
product obtained having 25 mg. seleniumVo, was not toxic in intraperitoneal injections in mice. Administeredorally in humans, in doses from
1/ru-2cc. of the solutioncontaining25mg. of seleniumper I cc., repeated
even severaltimes a day, did not show toxic effects.The influenceexerted
on pain and systemicchangestook some time to appar as with the other
seleniumpreparations.The influenceexertedupon the growth of experimental tumors in animals was more manifestthan for the other selenium
of naphpreparation.Similar resultsrvereobtainedwith the perselenides
thaleneand other aromatichvdrocarbons.

PHARMAcoDYNAMTC
ACrrvtTY

345

An investigationof the influenceexerted by the immediately heavier

member of this VIth series,tellurium, is in progress.
The foregoingdata on lipoids with negativecharacterindicatethat their
activity generallyis related to changesin processesin which ultimately an
interventionof oxygen takes place. This brings us to the first member of
the sixth series,oxygen,a nonmetalwith D inducing biological activity.
we studiedthe eftectsof ionic oxygen,usingcompoundswhich liberate
oxygen readily. These includedhydrogenperoxideas well as peracidsand
their salts, such as perchloric,perboric,persurfuricand periodic.
The changesinduced by thesesubstancesupon microbes,viruses and
cells are similar to those obtainedwith polyunsaturatedfatty acids and all
are cataloguedas radiomimetic.This fact tends to confirm the importance
of oxidation changesin the pharmacodynamyof fatty acids. The effects
upon pain and at organic and systemiclevelsalso were similar to those of
polyunsaturatedfatty acids. It is interestingto note in the same frame of
activity the appearanceof oxidizing substancesin the urine following the
oral administrationof theseagentsin higher doses.
we investigatedthe effectsof turpentineoil which is known to induce
the appearanceof peroxidein vitro. Highly oxidizedthrough treatmentwith
oxygen or especiallybound to sulfur, turpentineoil has shown interesting
pharmacologicalactivity. An old therapeuticdevice was parenteral administrationof turpentineoil to stimulatethe defensemechanismin cases
of septicemia.However, we saw no such stimulating effect in the fight
againstcancerouscells.The influenceexertedupon the cellular level of the
organizationwas quite reduced.The action of atomic oxygen appearsto
be difterent from that of the molecular.as we will seelater.

ALKYLATING

AGENTS

We investigated,as compounds with negative character, certain alkylating agents,choosing from the large number availablethose which also
showed lipidic properties.We were especiallyinterestedin two members,
sulfur mustardand epichlorohydrine.
Sulfur mustardcontains,along with
one active polar chloroethyl group, a secondrepresentedby the bivalent
sulfur polar group. It has the effect at different levels of organizationof
producing an offbalancewith predominanceof the acid lipids. We will
discussbriefly here some of the experimentsin which this influenceupon
the body lipids has been observed.

346 /

x E s E A R c Hr N P H Y s r o P A T H o L o o y

Su$ur Mustard
In studying sulfur mustard,we were first interestedin its effectsupon
body lipids and, through them, upon the lipidic systemof organisms.In inducing sulfur mustard'scharacteristicskin lesions,an interestingrelationship wasobserved.Pure sulfur mustardwas appliedto mechanicallyepilated
skin of rats. If a suffficientamount-2 to 3 drops-was used and spread on
one squarecm., the animal died. However,the time of death varied. If the
lesionshoweda massivenecrosis,followed by deep ulceration,similar to a
burn of the third degree,the animal died in about three weeks. However,
if the lesion was only erythematous,similar to a burn of the first degree,
the animal died in only 3 to 4 days. It seemedas if the lesion itself intervened secondarilyin the pathogenesisof changesleading to death. Sick
but still living cells appearto have an activity which is highly detrimental.
The abnormal cells apparentlyproduce substanceswhich are responsiblefor
rapid death of the animal. In widely necrotic tissues,these abnormal but
still living cells are limited in number; in an erythematous lesion, they
form the lesion itself. This correlation of toxicity with local Iesion was confirmed by the fact that excision of the lesion itself, il performed in time,
preventeddeath in some animals.The administrationof ferrous sulfate to
rats having sulfur mustard applied to their skin was seen to induce the
erythematousform of the lesions,with death in 3-4 days.
The similarity between the influence exerted by mustard burns and
caloric burns with a sufficientlyextensivefirst degreeburn producing more
rapid death than a third degreeburn, was of interest.
Analysis of the body of an animsl killed by a mustard burn reveals
abnormal amounts of unsaturatedfatty acids and reduced amounts of
sterols. In some cases,where death occurred after more than three weeks,
body sterols were found to be almost completelylacking. In these cases,
almost no insaponifiablefraction could be found. The lesion itself, especially an erythematousand edematousone, was very rich in unsaturated
fatty acids.Histologicalstudy of theseskin lesionsrevealedchangessimilar
to those obtained through the intradermic injection of concentratedsolutions of body acid lipidic fractions.The study of these lesionsfurther revealedthat the lesionsthemselveswere separatedfrom the organism by a
barrier of adipouscells,the resultof an exaggerationin number of the cells
of the subdermicfatty layer.
We studied these important changesfrom severalpoints of view. We
could show that an exaggerationof the adipouslayer underneaththe skin
occurs wben lipoids with negativecharacter,such as polyunsaturatedfatty

PHARMACODYNAMTC ACTMTY

347

acids,thiolipoids, etc. act upon the skin. Thus, this subcutaneousadipous

formation appearsto be a defenseweapon,designedto keep such lipoids
from passinginto the organism. The defenseappearsto be unequal for
males and females,as shown in the following study.
In collaborationwith the late Prof. R, Leroux of the Faculty of Medicine in Paris, we studiedhistologicalchangesin the ears of rats after local
applicationto the skin of a small amount of pure sulfur mustard.Normally
there are no adipouscells in the pavillion itself except at its base. Twenty
minutes after the application of sulfur mustard on the skin of the ear, 2
or 3 layersof adipouscells were seenin the connectivetissuebetweenskin
and cartilage.(Chapter 6, Note 22 )
curiously enough, the rapid appearanceof adipous ceils 20 minutes
after application occurred in femalesand not in males. The spaying of
femalesor castrationof males did not changethis responseeven after a
lapseof months.The administrationof male sexhormonesto female ratsspayed or not---{r of fcmale sex hormones to males----castratedor not
-also produced no change.It was only by the administrationto males,
ovcr a period of days, of a sufficiently large amount of the insaponifiable
fraction obtainedfrom the bodies of rats that this rapid responsewas induced. The administrationto femalesof the acid lipid fraction obtained
from rat bodies was seen to prevent the rapid adipous response.This differencein responsebetwecnmalesand females,can be relatedto the differencesin the amounts of membersof the two groups of lipids ordinarily
found in malesand females,as mentionedabove.
we studiedsulfur mustardfrom the point of view of pharmacological
activity.Dosesof 100 mcgr./100 gr. of body weight 1of a O.7Vosolution
of sulfur mustardin oil) were nontoxic in rats and micc. Except for an intensivelocal reactionat the injection site, no important immediatechanges
were obtainedin humanswhen I to 3 cc, of the 0.2% solution was injected
intramuscularly.The influenceupon pain-a decreasein the intensity of
acid pattern and an increasefor the alkaline pattern-was only temporary.
The influenceon tumor evolution was not sufficientto warrant clinical use
of this agent, especiallyin view of the persistentand intensivesystemic
changestoward an offbalance of the type D which appeared after a few
days. Through its anti-A action, which is the most intensiveof all agents
tested,sulfur mustard remains one of the most interestingsubstancesfor
experimental studies, especiallyfor the eftects exerted upon the anti-fatty
acids.

348

xESEARcH rN pHysropATHoLocy

Epichlorohydrine
The importance of the relationship betweenthe energeticcenters present in alkylating agentsand their ability to produce type D offbalance, led
us to study a group of these substanceswhich, at once, have both short
molecules and two polar groups in close proximity. The desire to have
such an agent with lipoidic property as well led us to study epichlorohydrine which correspondsto propane and has an epoxy group binding C2
and C3, while cr binds a chlorine. Solublein neutral solvents,epichlorohydrine becomes soluble in water only after hydrolysis. Its biological
activity differs from other chlorohydrinessuch as chloropropanediolor
trichloropropane,both of which can be consideredto be closely related to
the substanceproducedby hydrolysisof epichlorohydrine.The acute toxic
dose of epichlorohydrinewas found to be 6 mgr./30 gr. for mice and 25
mgr./100gr. for rats by intraperitoneal
administration;22and 35 mg./100
for mice and rats by subcutaneous
injection.In testsfor chronic toxicity, it
was apparentthat dosesof 5 and L5 mgr. injecteddaily were well tolerated
respectivelyby rats and mice. with higherdoses,the animalsbecamerapidly
emaciatedbeforedying. Used orally in drinking warer,a solutionof l/3000
was well toleratedby rats and mice even for months.With the use of solutions of l/2000, only a few animalsdid not lose weight,while a solution of
l/1000 invariablyinducedwcight loss.
There were no effectsobservedupon microbesor bacteriophage.
It appearsthat epichlorohydrine,actingbelow the morphologicallevels,
induceschangessimilar to thoseseenfor other alkylating agents.However,
it is not upon the desoxyribo-nucleic
acidspresentthat an important action
is seen but in the lipidic system at these lower levels. Epichlorohydrine
seemsto act also at other levels.The influenceupon pain-an increasefor
alkaline pattern, a decreasefor acid-was more noticeablethan for sulfur
compounds.Delaycd eftects,however,were more obvious than immediate
ones. The influence upon wound healing was similar to that of polyunsaturatedfatty acids. Cancerouscells, such as those from mouse ascites,
were destroyedin vitro by a 0.5 solutionof epichlorohydrine.The effect in
vivo upon sarcoma 180 or Ehrlich ascitestumors was most interesting.
Administeredby subcutaneous
or intramuscularinjection,epichlorohydrine
had no effecton the tumor even in dosesas high as 2,5 mgr. daily. However,
when administeredin drinking water in a 1/1000 solution,it prevented
the developmentof ascitesin 19/20 animals.But the toxicity was too high.
A L/2000 solution,usedas drinking water,controlledthe condition in morc
than 50o/aof the animals, while a I /3000 solution showed favorable re-

P H A R M A C o D Y N A M T CA C ' r l v t r Y

349

sults in only a few animals. Under the same conditions,there was no

apparent effect upon solid tumors in mice, even those induced by subcutaneousinjectionof ascitestumor cells.
In humans,all effectsupon the tumors were interestingand wilr be
discussedbelow. The influenceupon systemicpatterns was relatively reducedexceptfor a markedeffectupon the eliminationof calciumin urine,
obtainedeven with small doseswhich producedno other changes.Repeated
injectionsof organ cells treated in vitro with epichlorohydrinewere able
to induce severedegenerative
changesin the respectiveorgans.Experiments
with tumor cells treated and administeredin the same manner are stitl in
progress.
THE ELEMENTS
We have discussedpreviouslythe method used to classifythe elementsin
accordancewith their predominantbiologicalintcrvention.The place of
the elementsin the periodic chart, which establishes
the relationshipbetween their structureservedas a further basisfor this systematization.
The
capacity to induce changestowards an offbalanceof the same type was
found to be :r common property for clementsin the same series in the
periodic table. The series could be separatedinto two groups, one Ht
(from heterotropic)inducingan A and the other Hm (from homotropic)
inducinga D offbalance.Elementssystematizndas different periods in the
chart have been found to have predominantactivity in various compartments or groups of levels which form the hierarchicorganizationof complex organisms.we have related each element to a compartment (or
sometimeseven to a level), which didactica.llyis called the compartment
(or the level) of the element.We have tried to utilize this svstematization
in the study of the pharmacologicalactivity of elements.
From the beginning,severalbasicfacts about biologicalactivity became
apparent. Often the elements,used as such, do not induce the changes
which characterizetheir physiologicalactivity. Basicallyelementsact in
normal physiologythrough specificcompoundssuited to the compartment
or level to which they belong.In general,knowledgeof the level of an element permits us to identify also the proper compoundand its activity. The
interventionof an elementat levelsof the organizationother than its own
can be understoodonly in terms of the relationshipbetween the element
with its characteristic
compoundthe proper leveland the compoundspresent at other levels.
Two factorsare fundamentalin determiningthe activityof an element:
a) its availabilityand b) the possibilityto enter into its proper combination.

350

xEsE^RcH rN pHysropATHoLocy

The nature of any abnormality in activity of an element can be recngoizrl

only by relatingit to thesetwo factors.The amount of the elementavailable
and the capacity of entities of the level to which the element belonp to
manufacturethe proper compound,ultimately governsthe amount of the
elementat its own and at the other levels.
Under normal conditions, the entities of the proper level utilize only'
the amount of the elementneededto maintain the normal constants.The
rest of the elementusuallyis eliminated.An excessof the elementthus does
not induce a permanentexcessat the proper or higher levelsunder normal
conditions.Such an excessnormally is only temporary.A persistentexcess
of the elementat the proper level and at thc higher level indicatesan abnormal general availability. A persistentexcessat the higher level only
correspondsto a qualitativedeficiencyat the proper level. The organism
maintainsan excessof the elementat the higher level in an effort to compensatefor the qualitativedeficiencyat the proper level. Thus, an excess
of an element at the higher level indicatesa qualitative deficiencyat the
proper level, only if the value of this level is found low.
When there are low valuesat higher levels,we also need to know the
amount present at the proper level in order to interpret the abnormality.
Low valuesof the elementat higherlevelswith a low amount of the proper
level indicates a quantitative general deficiency,while a high value at
the proper level permits us to recognizea qualitativeexcessiveutilization
of the elementat the proper level.For example,in cancer,copper is low in
tumor cells and liver, but abnormallyhigh in blood. When a high amount
of copper appears in blood, the diagnosisof a qualitative deficiency in
utilization in cells or an abnormal generalamount can be made by investigating the amount in cells. A low amount of copper as found in tumors
and liver cells in subjectswith cancer make the qualitative nature of this
deficiencyevident.The trouble lies not in too Iittle copper availablein the
body but in the lack of the capacityof the cancerouscells to manufacture
the compoundthroughwhich copperbecomesactive,in this case,catalase.
A normal amount of an elementat its proper level reflectsnormal utilization.The pathologicalamountcan resultfrom a quantitativeor qualitative
abnormality,Any anomaly in an elementnot only means an inadequate
amount of it at its own proper level but in other levelsas well. A quantitative dcficiencyresultsin an insufficientamount at its proper level. A qualitativc deficiency-that is, incapacity of the entities to manufacture the
proper compound-also leads to a reduced amount of the element at its
own level.
Thus, in both cases,quantitativeand qualitativedeficiency,the amount

PHARM^CODYNAMTC ACTTvTTY

351

of the element present at its own level is low. However, in quantitative

defrciencythere is also a low amount of the element in the hierarchically
superiorlevel, while an increasedamount at this superiorlevel occurswhen
there is a qualitativedeficiencyat the proper level.
The same oppositevariationsbetweenthe amountsat the two levels is
seenin the caseof an excessiveutilizationof the elementat its proper level.
The amount of the elementat the superiorlevel is also high, if a quantitative excessis presentbut this amount at the superiorlevel is reduced as a
meansof controlling the excessiveutilization at the proper level if a qualitative anomaly occurs.This makesit possibleto recognizethe quantitative
or qualitative nature of the abnormalityin utilization of an element at its
proper level by determiningits amount both at this level and at the next
superior level. Too much of the elementpresentat the proper level indicates either cxcessivczmount present or excessiveutilization, while too
little at the proper level can be due either to a qualitativeor to a quantitative deficiency.A low amount at the higherlevel indicateseither a quantitative generaldeficiencyor a qua.litativeexcessiveutilization at the proper
level. An excessamount at the higher tevel indicateseither a quantitative
generalexcessor a qualitativedeficiencyat the proper level.
This relationship,which is also critical for the understandingof the
pharmacologyof the elements,can be summarizedin the fouowins table:
Amount of Element
at the ProperLevel

Amount of Element
at the Higher Level

high
Iow
high

high
high
low

low

low

Interpretation
: Occurrence
at the Proper Level
quantitativeexcess
qualitativeinsufficiency
qualitativeexcessive
utilization
quantitativedeficiency

From a practical point of view, we must have information on the

amount of the elementboth at the proper level and at a higher level. we
found that for the elementsproper to the cellular level, such information
can be obtained by comparing the amounts in plasma and red cells (or
total blood). It is not the ratio betweentheseynlus5-25 often supposedwhich is important, but rather the valuesthemselves.For changesat the
systemiclevel, the comparisoncan be made betweenblood and urine. the
latter corresponding to the level above the systemic.
The importanceof this conceptcan be seenin the following examples.
Potassiumis a cellular level element. In cancer, in offbalance type A,
potassium is present in abnormally higb quantity in proliferating cells. It

352

xEsEARcH tN pHystop^THoLocy

is also found in high amountsin blood red cells. In thesecases,potassium

is found in low valuesin the hierarchicallyhigher level in the blood plasma
or serum. The abnormality does not reside in a simple hypokaliemia,but
in excessiveutilization of potassiumat the cellular level, a low amount of
potassiumin red cells also would indicate a potassium deficiency. A high
amount of potassium in serum and red cells can be interpreted, as mentioned above, as correspondingto a quantitativeexcess.The reduction in
the quantity of potassiumin red cells, togetherwith an excessin the serum,
indicatesa qualitativedeficiencyat the proper level.
we have the true picture of the situation if we considerthat .,qualitative" excessor deficiencyis determinedby the ability to form the proper
compounds.While it is the elementas such which has to be administered
in order to correct a quantitative deficiency,other factors must be changed
to overcome qualitatively deficient utilization, excessiveutilization, and
quantitativeexcess.(FiS. 127)

trr

Excess

E
+

v,
E
:'
L

o)

(t)

TotalBloodK+rtq
F r c . 1 2 7 . T h e r e l a t j o n s h i pb e t w e e nt h e a m o u n t o f p o t a s s i u mi n s e r u m a n d i n t o t a l
b l o o d p e r m i t s t o i n d i c a t et h e e x i s t i n gc o n d i t i o n a s b e i n g i n n o r m a l l i m i t s , i n q u a n r i tative deficiencyor excess,or in an offbalancetype A or D.

PHARMACODYNAMIC ACTIVITY

,,/ 353

Although these relationshipsrepresentthe most important aspect of

the pharmacodynamyof the elements,still others must be considered.An
excessor deficiencyof an elementat a superiorlevel.even if it servesas a
biological defensemeans to combat an abnormality at a lower level, representsa problem by itself for the supcrior level.The fact that the element
doesnot belongto this levelgivesa noxiouscharacterto its influence.The
influenceexerted upon sensitiveorgansoften inducesimportant abnormal
manifestations.
Hyperkalemia,evenoriginatedby an abnormallylow utilization of potassiumat the cellularlevel,can lead to serioustroublesin the
function of the nervoussystemor of the heart.
Another importantaspctof the reactivityof an elementis its influence
upon levelsbelow its proper level. An elementacting at a lower level usually has a biologicallyoppositeeffectto what it has at the proper level and
thereforeis a noxious influence.At the lower level, the A or D type of
activity of the elementis reversed.
Sodium, for instance,which is an agent of the A type of the metazoic
compartment,producesan offbalanceof type D at the cellular level, which
is hierarchicallyinferior to its own. Similarly, Mg, which is a D agent at
its own metazoiclevel,has an A inducingactivityat the cellularlevel.
Analysisof the pharmacologyof elementsin terms of their A or D inducing activity, the level at which they belong and the compoundsthrough
which they act, is still only in its early stagesalthough it representsa program of great promise. ln the presentationwhich follows, we will try to
interpret the data concerningthe elementsin terms of A and D inducing
activity. We start with Hm elementshaving a D inducing activity. They
parallel in their action the lipoids, with a negative polar group. Tear-E
XVII lists Hm elementsor the D seriesand relatesthem through their
periodsto the organizationalcompartments.
T,rsI-e XVII
Hr'r Elevexrs
Compartments
Organic
Metazoic
Cellular
Nuclear
Submorphol.
Primary Biol.
Submolecular

Metals
Be
Mg
Ca Sc V
S r Y Nb
Ta
Ba
Ce N d
Ra Th U

Non-Metals

c
si
Mn
Tc
Re
Sm
Pu

Co C u
Rh Ag
Ir
Au
C d Dy
Cm Cf

Ge
Sn
Pb
Er

o
s
Se
Te

Yb

We have alreadydiscussedthe pharmacologicalactivity of sulphur and

seleniumthrough the compoundsin which they enter and will not discuss

354

REsEARcH rN pHysropATHoLocy

them again here. Before analyzing the other elements,it is of interqst to

emphasizeagain a principal characterof their activity. As most of the elements act through specific compounds, the factors which determine the
entry of elements into specific biological combinations appear to be of
capital importance. Availability of the element alone is only one factor in
its pharmacodynamy.with this in mind, we have investigatedsome of the
elementsof this Hm group.
we know little about any influenceof berryllium as a metal upon the
organismas an entity. Its toxic effectsare due to abnormal amountsactive
at lower levels.In the same period of the chart of elements,we have two
nonmetals,c and o, for which the organismrepresentsthe proper level.
The general pharmacologicalnature of oxygen is indicated by the role of
oxidation in metabolic changes.Oxidation representsthe first step toward
catabolic,homotropic changes.The respiratoryphasein the metabolismof
carbohydrates,the oxidative fissionof fatty acids, and the oxidative desaminationof amino acids representexamplesof the fundamentalhomotropic interventionof oxygen.
Acting at the systemic level, immediately inferior to its own level,
oxygen has a different action. According to the rule mentioned above,
oxygen,a D agentat its proper level, will have an A activity for the blood,
which we will study below togetherwith the other A agents.The homotropic relationshipof oxygcn to fatty acidswas discussedabove with the
study of the biological role of thesesubstances.
The relationshipbetweenCo2 and fatty acids also is interesting.Large
amounts of free fatty acids in the blood were seen to allow better fixation
of COz to hemoglobin,just as large amountsof sterolsdo for oxygen. We
have investigatedthis correlationbetweenCo2 and fatty acids by keeping
a fatty acid, such as linoleic acid, in an atmosphereof Co2 connectedto a
manometer.A manifest negativepressureresults.The venous blood, rich
in CO:, which also shows a predominanceof fatty acids, loses CO2 and
fatty acids during passagethrough the lungs.
Magnesium
Among offbalanceD inducing elementsof the II A seriesof the periodic
chart, we first studied magnesium,which belongs to the metazoic compartment and thus, is related to the sea as original environment.Much of
the activity of this element can be interpreted as D inducing activity. Mg
in many respectsis antagonisticto Na, the cation of the samecompartment,
a memberof a serieswith an oppositeA inducingcharacter.High valuesof

P H A R M A C O D Y N ^ M I CA C T I V I T Y

,/

355

Mg found in blood were related to adrenalinsufficiency,(127) while low

It'lglevelsusually occur when blood cholesterolis high. (128)
Excessof Mg was seento induce adrenaldeficiency.we explainedthis
actionthrough the antagonism betweenMg and Na. We could thus counteractthe salutary effect of NaCl in adrenalectomizedrats through administrationof magnesiumsulfate parenterally.A similar effect was obtained
evenwith oral use of magnesiumthiosulf.ate.(Note 9)
The relationshipof magnesiumto the defensemechanismis of special
interest.
Mg seemsto intervenein the lytic effectof sera upon ascitescancercells (extensivelystudied in our laboratory by R. Willheim, p. Fluss
andM. Auber) (129) which, as we have seen,representsa characteristic
featureof D inducing activity. Similarly, magnesiumpreventsthrombosis,
actingas an antithrombocyticagent.Not only does it prevent the appearanceof fibrin, partly preventing the destructionof thrombocytes(130);
but it also favors the lysis of already existing thrombi. ( l3l ) It appears,therefore, to be a valuable agent in the treatment of thrombosis.
( 132) Its concomitantaction againstcholesterolhas led to its use in the
preventionand treatmentof coronary thrombosis.
Magnesiumappearsto be part of anotherdefensemechanism,the nonspecificone, representedby the properdin system.Properdin is active only
in the presenceof magnesium;neitherca nor Na can replaceit. (133)
Higheramountsof magnesiumincreaseproperdin activity.
Magnesiumsometimesis seento parallel the action of cu, another D
inducingelement. In animals fed milk too long, leading to a type A offbalance,the amount of magnesiumfalls along with the amount of cu. The
quantityof magnesiumin the blood is low in humans with convulsion.
(134) Mg appearsto be espcciallyeffectivein the prevention and treatmentof "grass tetany" in animals, which often follows feeding on grass
withhigh potassiumcontent.( 135)
Magnesiumhas a similar antagonismtoward K and this can explain
its activity in cancer. Lower than normal valuesof magnesiumare found
in cancer,as opposedto high amountsof potassiumwhich is an A inducing element, and the low Mg seems to favor cancer growth. Moisture
increases
the amount of K and lowers the amount of Mg in plants, a fact
relatedto cancer frequency in various geographicregions. (136) The
difference
between the preventiveand curative actionsof magnesiumis of
specialinterest. Administered after a carcinogenhas becn applied, magnesiumreduces the percentageof cancers induced. (137, 138) It has
minimal influence, however, once the tumor has appearedor upon trans-

356

R E s E A R C Hl N

pHystopATHoLocy

planted or spontaneoustumors. We will discuss this occurrence below

togetherwith the effect of other elements.
It is necessaryto bear in mind, when we have to choose nonspecific
combinationsin which to administer it, that the D inducing activity of
magnesiumis particularly manifestedat the metazoic level. Magnesium
sulfate appears to be suitable for parenteral use, while the thiosulfate
appearsto be suitablefor oral administration.In theseforms, Mg has been
found to induce marked local alkalosisin the secondday wound crust pH
and to produce salutary effectsupon pain of the acid pattern. A marked
influenceupon thiamine-inducedconvulsionsin rats and mice has led us to
usemagnesiumthiosulfateas a tool not only in tetany but in the treatment
of convulsions.It appearsto be effectivein preventingepileptic seizures
and valuableeven in casesof statusepilepticus.We have utilized the same
preparationsuccessfullyin cancer when pain and preterminal conditions
correspondingto an A offbalancewere present.Less important effectsare
seenfor magnesiumsulfate and magnesiumthiosulfateat lower levels.
Calcium
The biological activity of calcium, another member of the II A series
and a D inducingelementbelongingto the cellular level, also is of interest.
In its absorptionby grass from the soil, Ca parallels Mg, another D
inducingelement,but opposesK, an A inducingelement.Grasstetany is
thus induced by high K and low Ca and Mg values.Ca, which is another
D inducing elementlike copper is antagonisticto zinc, an A inducing element. It has been observedthat cancr is less frequent in the so-called
calcerousclay regionswhere the soil is formed by limestone.(139) Together with other minerals, an optimum of calcium in soil may help to
prevent cancer. While SiO,,favors cancer,Ca appearsto prevent it. Calcium also is an antagonistto zinc which, in high doses,seemsto favor the
developmentof cancer. (140) The relationshipbetweenCa and K has
permitted us to be more precise about the role of Ca in cancer pathogenesis.As opposedto K, which increasesby as much as 60Vo in tumors,
the contentof calciumdecreases
by 44Vo. ( l4l -147)
Confronted with K and Ca changes,it appearedinterestingto see to
which element we could directly attribute the increasein malignancy. In
the regenerationof liver cells, where rapid growth without malignancy
takes place, potassium is increasedwhile the amount of calcium is unaltered.Similarly in other rapidly growing but normal cells, calcium is not
diminishedwhile K is increased.Potassiumthus appearsto be related to
the processof cellular growth and multiplicationwhich representsalso an

P H A R M ^ C o D Y N A M t C A C r l v r T Y

3 5 7

addedfactor in transformingnoninvasiveinto invasivccancer.However,

potassiumis not directly related to the cancerouscharacter of the cells.
on the other hand, reduced amountsof calcium appear to be peculiar to
the cancerousprocess
. (147 ) The reducedcalcium in cancer is not due to
a lack of the elementin the organismsince calcium is not only available
but even apparently present in excessat the systemiclevel. As we have
shown,a high urinary calcium index, indicating exaggeratedexcretion, is
presentin the type A offbalance.The anomaly residesin the low capacity
of the cancerouscells to fix and properly utilize calcium. As calcium acts
at the surfaceof the cell and its deficiencyreducescellular adhesiveness,
(148, 149) lack of cellularcalciumcan be seento increasethe invasivenessof cancercells and the tendencyto metastases.
Deficiencyof calcium
in cells appearsrelated to the characterof youth while excessseemsto
resultin rapid agng.
The anomaly induced by the qualitativedeficiencyin calcium thus appearsto be at the cellular surface.Relatedto it also are manifestationsat
the tissular level.
Administrationof any calciumsalt inducesa manifestincreasein local
alkalosis(Fis. i,28)of the secondday wound crust pH. It appearedinterestingthat in bone lesions,especiallyin bone cancermetastases,
the offbalancetype A is characterizedby an osteolyticprocess,the D type by an
osteoplasticone. The local acidosispresentin lesionswith an A type of
offbalanceexplainsthe mobilizationof calciumin theseosteolyticprocesses.
ca is depositedin important amountsin metastases
with a type D offbalance,a fact which can be related to the local alkalosisresultingfrom the
abnormal metabolism.This alkalosisrepresentsa condition favoring the
precipitationof calcium. Indirectly, the deposit of calcium in bone metastasesappearsto corrcspondto the D pattern of tissular abnormality.
Calciumhas a D inducingactivityevenin this case.
with calcium excreted in excessthrough the urine, the problem of
calcium pharmacologyin the A type of cancer is related to the form in
which it acts at the cellular level, which appean qualitativelyimpaired. As
the quantitativedecreasemust be considercdto be a consequenceof
qualitativeinsufficiencyand not a generalquantitativedeficiency,the problem is not to provide calcium but to find a way to insure better utilization
at the cellular level. It is for this reasonthat administrationof most calcium sals does not influencethe evolutionof experimentalor clinical cancer, but has a preventive effect upon the induction of tumors through
carcinogens.Administered after the injection of the carcinogen,calcium

358

xESEARcH rN pHysropATHoLocy

appearsto reduce the percentageof positive results.Administered after the

tumors have appeared,the influenceis minimal or nil.
As we have mentioned above, an excessivecalcium excretion is, in
itself,sufficientto indicatethe cxistenceof a deficiencyin calcium utilization at the cellular level without a calcium deficiencyin the organism.The
therapeuticindication is for agentsable to influencethe fixation of calcium
at the cellular level. Fatty acids which changethe cellular metabolismso
t o
>
t
o
i6
|
Fo
l r
6 C
I
r L
6 0
|
t ! l u o
-- l
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t
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G
($

F r c . 1 2 8 . T h e i n f l u e n c ee x e r t e d b y t w o e l e m e n t s ,p o t a s s i u ma n d c a l c i u m , u p o n t h e
second day wound crust pH shows a frank tendency toward acidification for potassium and alkalinization for calcium. For diflerent salts, the differences result from
t h e u n e q u a l i n f l u e n c ee x e r t e d b y t h e a n i o n w h i c h w o r k s a d d i t i v e t y t o t h a t o f t h e
cation.

as to induce local a.lkalosis,have appearedto be the most active agents.

Testosterone,and calciferolhave appearedhelpful but not nearly as active
as fatty acids.
while high urinary excretionof Ca indicatesa type A oftbalance,low
or no urinary excretioncan result either from an excessivecellular utilization of calcium or from a type D oftbalanceat the cellular level. other
analysescan be used to indicate the probable occurrence.With calcium
excretion low and other analysesindicating type D offbalance, all the
chancesare that the low excretionis part of the D offbalance.If, on the
contrary, only the ca excretionis low and the other values correspondto

358

r.ESEARcHrN pHysropATHoLocy

appearsto reducethe percentageof positiveresults.Administeredafter the

tumors have appeared,the influenceis minimal or nil.
As we have mentioned above, an excessivecalcium excretion is, in
itself,sufficientto indicatethe cxistenceof a deficiencyin calcium utilization at the cellular level without a calcium deficiencyin the organism.The
therapeuticindication is for agentsable to influencethe fixation of calcium
at the cellular level. Fatty acids which changethe cellular metabolismso
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i
|
o C
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F r c , 1 2 8 . T h e i n f l u e n c ec x c r t e d b y t w o e l e m e n t s ,p o t a s s i u ma n d c a l c i u m , u p o n t h e
second day wound crust pH shows a frank tendency toward acidification for potassium and alkalinization for calcium. For diflerent salts, the differences result from
t h e u n e q u a l i n f l u e n c ee x e r t e d b y t h e a n i o n w h i c h w o r k s a d d i t i v e l y t o t h a t o f t h e
cation.

as to induce local alkalosis,have appearedto be the most active agents.

Testosterone,and calciferolhave appearedhelpful but not nearly as active
as fatty acids,
While high urinary excretionof Ca indicatesa typ A offbalance,low
or no urinary excretioncan result either from an excessivecellular utilization of calcium or from a typ D offbalanceat the cellular level. Other
analysescan be used to indicate the probable occurrence.With calcium
excretion low and other analysesindicating type D offbalance, all the
chancesare that the low excretionis part of the D offbalance.lf, on the
contrary, only the Ca excretionis low and the other values correspondto

PH^RM^CODYNAMIC ACTIVITY

359

A type of offbalance,the probability is a quantitativelack of calcium in

the organism. This can be corrected by the administration of calcium
parenterallyor orally in any absorbableform. Low excretion as well as
somesymptoms can be overcome in a short time through the adminl563tion of sufficient calcium. In the opposite type of case, with metabolic
calciumretention,administrailonof calcium will induce an increasein the
intensityof symptoms.
Excessof calcium in the urine thus correspondsto lower values at the
cellularlevel and need for fixation of calcium. This indicatesagain that
the problem is not the amount of calcium presentbut the deficiencyin its
utilization.
Copper
copper, from the lB seriesand another anti-A elementof the cellular
level,appearsindispensablefor the synthesisof heme of catalase.A deficiencyof copper results in reducedactivity of this enzyme.Similarly, the
synthesisof hemoglobinis possibleonly in the presenceof copper. cytochrome oxidase contains Fe and Cu. Cu deficiencyreduces liver cytoc h r o m eo x i d a s e .( 1 5 0 , l 5 l , 1 5 2 ) c u i s p r e s e n it n b l o o d s e r u mb o u n d t o
a proteinto form ceruloplasminwhich also acts as an oxidase.(153) Cu,
while favoring the synthesisof these substances,intervenesultimately in
the processeswhich lead to the active catabolicintcrventionof oxygen. In
this way, cu acts as a D inducing agent. cu intervenesactively in the
metabolismof sulfur. (154, 155) The transformarionof sulfhydryl to
disulphideis slow and incomplctewhen there is a copper deficiency.The
samedeficiencyreducesthe formation of phospholipidsas seenin rat liver.
(156) Indirectly,Cu favorsanti-A activities.
The influenceof cu upon ca metabolismalso is indirect.we have seen
that a local depositof calcium in bonescorrespondsto a local D pattern.
Deficiencyin copper, a D inducing agent,permits the appearanceof local
A conditions,which in the case of bone, will result in a lack of calcium,
theoppositeof what is seenin local D offbalance.This correlationexplains
why, in spite of sufficientP and Ca, the lack of coppr inducesan osteomalaciawith bone fracture and symptomsof rickets, as seen in animals
with an indirect Cu deficiencycausedby an excessof molybdenum, ( 157,
158, 159) an A inducing agent.The administrationof copper helps to
repairthesefractures.(157) Parallelreductionsin Cu and Mg occur in
milk-fedcalves. (160) A richnessof zinc, like molybdenum,can provokea deficiencyin Cu and Ca. With copper deficiencyand low catalase,
resistance
to infectionsis lowered,In brucellosis,
a deficicncyof Cu and Ca

360

nEsEARcH rN pHystopATHoLocy

coincideswith reduced concentrationsof Mn and Co in the blood and

p i t u i t a r yg l a n d s .( 1 6 1 , 1 6 2 , 1 6 3 ) C u , C a , M n a n d C o a r e a l l m e t a l so f t h e
D inducinggroup.
In cancer,a qualitativedeficiencyof copper is found. A frank reduction of the catalasecontentis seenin cancerouscellsas well as in the liver
of canceroussubjects,( I 64, 165) on the other hand, cu contentis considerablyincreasedin the blood of thesesubjects,with valueseven three
times greater than normal, often encountered.(166. 167, 168) These
valuesreturn to normal if the canceroustumor disappears.(169)
In cancer,copper deficiencyat the level of the cells is manifest,shown
not only by reducedcatalasebut also by upsetsin the cytochromeoxidase
system,the heme system,the sH metabolites,
and the phospholipids.The
deficiency,however, is only local and consequent.lyqualitative since an
excessof Cu is found at the immediatelysuperiorlevel, the blood.
A local abnormalityresidingin inadequatecapacityto utilize copper
thus appearsrelatedto cancer.In normal animals,coppr in excesscan be
utilizedand is able to prevcntthe appearance
of tumors.This explainswhy
Cu, which protectsrats againstcarcinogenetic
azodyes,(170, l7l, 172)
does not influencethe tumorsonce they have been induced,i.e., once the
qualitative insufficiencyis present.The recognitionof this difterence between the form in which copperis utilizedby the normal animal and the
deficiencyin the cancerousentitics,hasbeenthe basisfor a seriesof studies
concerningthis key problemin the pathogenesis
of cancer.The therapeutic
useof copper-and of other elemcntswhich we will discussbelow-is not
a quantitativebut a qualitativeproblem.
Manganeseand Cobalt
Two other elementsof the cellularcompartmenthave appearedinteresting.Manganeseby interveningas a catalystin processes
resultingin an
activationof oxygen,indirectlymanifestsD inducingcharacter.Its presence
in smaller amounts in tumors or cancerousorgans than in controls, has
beenconsidered.Just as with copper,no effectsare seenin treating tumors
with manganesecompoundsalthougha certain degreeof preventiveaction
is obtainedin tumors inducedby carcinogens.Similarly,with cobalt we
have obtained a certain degreeof preventionagainsttumor induction by
carcinogensbut no effect upon the evolution of tumors once they have
appeared.No effectshave beenseenin transplantedtumors.

r
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n:!ii:
iil\r*#iiini

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P H A R M A C O D YN ^ M I C

A C T I V I T Y

361

Heavier Elements
we have studiedthe activity of elementscorrespondingto the lower
lgvsl5-5ugh as strontium and tin for the nuclear;barium, gold and lead
for the submorphologic;and cesium for rhe primary biological compartments. The lower the level of the element,the greater appearsto be the
preventiveeftect againstinduction o[ tumors by carcinogens.But the minimal, or completeabsenceof effect upon alreadyexistingtumor cells remains unchanged.
As mentionedabove,we conncctedthis paradoxicalactivityto the fact
that, while normalcellsare able to manufacturecompoundsthroughwhich
the appearanceof a cancerousentity can be prcvented,these compounds
are no longerformed if an cntity is alreadycancerous.
These have been the considerations
forming the basis for an entire
seriesof studiesof the role of variouselementsin the pathogenesis
of
cancer.
Elements in the Pathogene,sis
ol Canc'er
Investigationshave been made of the amount and form of the element presentin the normal animal as contparedto the cancerousanimal.
The quantitativeand especiallyqualitativedifferenceshave been seen to
indicatethe site of the abnormalitylargelyresponsible
for the lack of influenceexertedby the element.The resultsexplainwhy the administration
of the elementalone is unable to influencethe cvolution of an a-lready
existingcancerousprocess.More interesting,
they show what compoundof
the elementcould have an influence.This research,which is in progress,
opens the door for possibletherapeuticapplications.It is through such
compounds,presentin the normal and lackingin the cancer-stricken
animal, that attemptsare beingmadeto influencethe evolutionof cancer.An
important step has been the finding that suitablecompoundscan be obtained by the treatmentof fresh organsin vitro with some of the elements.
Their study may make possiblesyntheticpreparationof suitable compounds.The few resultsalreadyobtainedin experiments
with animalsconfirm that D-inducing activity representsa factor which the promising elementsshare.

C H A P T EI R5
PHARMACODYNAMIC
ACTTVTTY(PART TW O)
ANTI.FATTY ACID GROUP

ID
ll- nnellrL

To INVEsTTcATIoN
oF AcENTScapable of correcting offbal-

ancesof type A, attentionwas directedto agentsthat miglrt influencethe

oppositeoffbalance,type D. Since fatty acids are involved in the pathogenesisof type D offbalance,agentswith anti-fatty acid propertieshad to
be sought as correctives.Some of these are natural constituentsused by
the body to control normal and abnormalinterventionof fatty acids. They
were consequentlyisolated and studied. Synthetic agents also were obtained and studied,their choice largely inspiredby the control mechanism
used by the body.
Anti-Fatty Acids Constituents
We have seenthat a free fatty acid losesmost of its biological activity
rvhenits polar group is bound to anotherradical. This led us to inveswhich naturally are bound to fatty acids. It could be
tigate substances
shown further that each major group of fatty acids is bound in the organism to specificconstituents.Thc saturatedfatty acids are principdly bound
acidsas lecithins,
to glycerol,the low unsaturatedacidsto glycerophosphoric
and the high unsaturatedmembersto sterols.The conjugatedfatty acids,
found in abnormal conditions,appcaredto be opposedby neoglucogenic
corticoids.The constituents'were
conceivedof as being naturally occurring
first effort was to study how they interacid
substances
and
our
anti-fatty
vene to balance the activity of fatty acids, especiallywhen the latter act
as pathogenicfactors.

p H A R M A C o D y N A M T cA c r t v t r y

(p,rnr rwo)

363

one, a relatively
wereinvestigatcd:
In this study,two typesof influences
direct effect induced through a neutralizationof the energeticcenters resulting in a more or less advanceddegreeof inactivationof the fatty acid;
the second,an indirect effect achievedthrough changcsin the metabolic
processes
in which fatty acidsintervene,In a differentkind of intervention,
the anti-fatty acid to which a fatty acid is bound governs its ultimate
biological fate. For example,the bond to glycerol favors caloric metabolism. The bond to glycerophosphoricion converts a fatty acid, saturated
as well as unsaturated,into an organizationalconstituent.The bond to
sterolsfavors a functional role, even for monoethenoids.
We started the study of the naturally occurring anti-fatty acids with
those agents known to be bound to fatty acids in the organism. The
simplestsuch agentis glycerol.
Glycerol
Glycerolis the most ubiquitousfatty acid-bindingsubstancein nature.
We attempt to explain this fact on the basisof glycerol'sstructureand the
specialbiological role it confers.We have seen that fatty acid molecules
take reciprocal parallel positions when they form monomolecularlayers.
In their bond to glycerol,thesefatty acid moleculesconservethis recipro(Fig. 129)Thiscould cxplainwhy, in the body, the bond of
cal relationship.
being
fatty acid and glycerolalwaysis a triglyceride,mono and digJycerides
(?)

CD

(-)

c)

(-)
(J
O
O
c(\.1) o
- t -: E
a

(?)

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cf)
CJ

cf)
(J

(7)

(-)

Io

C\J

-(J

F t c . 1 2 9 . T h e f a t t y a c i d r a d i c a l st a k e i n t h e t r i g l y c c r i d em o l e c u l e sa s i m i l a r p a r a l l e l
position as when they form a monomolecular la1'erat the surface of water.

364

nESEARcHrN PHYsropATHoLocY

only intermediatesteps.The samefatty acid has different biological activities if administeredas free acid or as triglyceride.The combinationseems
to serveas an energy-furnishingmetabolite.Bound to glycerol,fatty acids
with long or short chains, saturated,monounsaturated,polyunsaturated,
and even conjugated,seem to representenergeticreserveswhich are utilized as caloric metabolites,especiallyin those specieswhich are able to
storethem.
With this relationshipin mind. we administeredglycerolwith two objectives: I ) to obtain, as an immediateeffect,the inactivationof the free
fatty acids presentin abnormalconditionsthrough the neutralizationof
their polar groups, and 2) to eliminatethesefatty acids by turning them
into caloric metabolites.
Studying the activity of glycerol at different levels, we could see no
influence upon phages. However, an indirect effect was observed upon
viruses.Glycerol is widely used as a specialmedium for the preservation
of virusesin tissucs.Its preservative
value can be correlated,at least in
part, with its influenceupon fatty acids.We have seenthat fatty acids have
a noxious effect upon viruses,leading to their disappearancein various
organs. The treatment with purc glycerol reduces autolysis of organs
through a dehydrationeffect.Curtailingthe lytic activity of the enrymes
active in autolysis,glycerol reduccsthe amount of fatty acids liberated
through such autolysis,and thus preventsthe destructionof the virus.
Glycerolmay also preservevirusesby acting antagonistically
to any fatty
acids which still manageto appear.
Glyccrol has a bacteriostaticeffectupon only a few speciesof microbes
and only when appliedin high concentrations.
A minimal influenceupon cells was seenfor glycerolin Tetrahymena
pyriformis and ascitestumor cells. To study its action at higher levels,
glycerol was administeredorally or parenterallyto animals or humans.
Solutionsof ZOVoglycerol were well tolerated when injected subcutaneously or intramuscularly.It should be noted that when glycerol was administeredto complcx organisms,it was largely absorbedand circulated
without alteration,a fact which would explain the effectiveness
of relatively small amounts.At the tissuelevel.glycerolinduceda changeof the
local pH of a lesiontoward the acid side,as seenin the secondday wound
crust. Figure 130 illustratesthis. The changeexplainsglycerol'saction in
increasingintensityof acid pattern pain and decreasingintensityof the
alkaline.This influenceupon pain was obtainedconstantlywith very small
amounts,permittingthe use of glycerolevenas a testfor diagnosisof pain
pattern.Intramuscularinjectionof t/z cc. of a 2Q% solutionor oral ad-

p H A R M A c o D y N A MAt c r r v r r y ( p , r n r r w o )

365

minisuation of. Vz cc. of a 5oo/osolution in water has been used for this
purpose. However, later, when other agcntswere found to produce even
more overt responses,we stoppedusing gJycerolas a routine test.
Glycerol has almost no beneficialinfluenceupon the healingof wounds
or radiationlesions.Healingwas evenretardedin someexperiments.
Various changesin the evolution of tumors occur when host or transplant are
trcated with g.lycerol.In some, these changesare minimal; in others an
obvious reduction in grofih occurs. In a high proportion (12/20), a
marked involution has been noted for Walker tumors in rats. Ascites Sa
180 in mice, after repcatedintraperitoneal
injectionsof a solution of.ZVo

GI ycerol

F r c . l l 0 . C l y c e r o l i n d u c e sa l o w e r i n go f t h e s e c o n dd a y
wound crust pH.

aoaa
aa
.O

glycerol, disappearedin TOVoof the cass.A lesser effect was seen in

Ehrlich and Krebsascitestumors and still lessin the solid tumorsobtained
with these ascitescells. (Fig. I3 I ) One of the most interestingeffectsof
glycerolwas that seenupon the tumorsin humanswherea manifestinvolution was obtainedin casesin which an offbalanceof type D was present.
This important effect will be discussedbelow with the therapeuticuse of
glycerol.Glycerol administrationhad an interestingeffectupon the amount
of cholesterolin the blood in a few subjects.When ten drops of glycerol
weregivenorally threetimesa day for a month or more,cholesterolvalues

366 /

RESEARCH IN

PHYSIOPATTIOLOOY

OO

Control

Start t reatnent

Ireatedrith
Glycero
I
rith
Treated
Glycerol
Ireatedrith
Glycerol

. l
. l
. 1

I -,.

F t c . l 3 l . C h a n g e si n d u c e di n W a l k e r t u r n o r si n r a t s by the treatnlcntsof the aninrals

with glycerol (daily subculaneousinjections with /: cc of 57a glycerol solution in
saline isotonic solution).

In somepatients,with no changein diet and no medicationother

decreased.
glycerol,
valuesoriginally above300 mgr./100 cc. serum fell to below
than
170 mgr. If thcsepaticntsalso were hypertensive,long-term administration
of glycerol produceda reductionof blood prcssure.
An impressivehcmorrhagiparcilect was noted, frequently ulccrated
lesionsstarting to bleed shortly after administrationof even a few drops of
glycerol. The relationship of hemorrhagingto glycerol was clear when in
the same subjects, repeated administration of this agent invariably was
followed by bleeding. The bleeding usually was arterial; only occasionally
was an oozing hemorrhagesecn.
Many years ago we became interested in studying, in a group of
severelyburned subjccts,the rolc of fatty acids in the pathogenesisof burn
complications.Glycerol was administercdto thess patients with good effects
upon pain. Before the use of antibiotics, one of the principal manifestations in burn patients with widely infectcd wounds was repeated chills.
These chills also were influencedby glycerol. (Note // A direct action upon
the parasympatheticsystemcould be attributed to glycerol and could explain the effect upon chills. This view has been confumed by studying the
eflects upon cardiac rhythm produced by intravenousadministration of
glycerol in rabbits. (Note 2)
Convulsions could be induced by glycerol in animals and also were
seen to occur in humans. (Note 3/ They could be induced, with much

pHARMAcoDyNAMtc

(p,rnr rwo)

^crtvrry

,/

367

smaller dosesin animalswhen, along with glycerol,an otherwiseharmless

dose of deoxycorticosterone
acetatewas administered.Injection of 0.1
mg. of this hormone in mice weighing25 to 30 grams, followed by an
injection of.Vc cc. of a 57o solutionof glycerol,inducedconvulsionswhich
r+'ereusually lethal. In terminal cancer patients,too, concomitantadministration of the cortical hormone and glycerol for a few days has produced
convulsiveseizures,which proved to be lethal in one subjectin whom no

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F r c . 1 3 2 . T h e a d m i n i s t r a t i o no f I c c g l y c e r o l d a i l y i n d u c e sa c h a n g eo f t h e u r i n a r y
pH toward thc type A, before changesin the other analysestake place.

368

REsEARcHrN pHysropATHoLocy

previous abnormal cerebral manifestationshad been observed, and who

had receivedthesetwo medicationsfor only a few days.
Of the different systemic manifestationsinfluenced by glycerol, the
eftect upon acid-basebalance is most striking. Even before any other effect
upon systemicanalysesbecomesapparent,an immediatechangeof urinary
pH from acid to alkaline values is induced by glycerol administeredin
sufficientquantity.(Fig. 132)
Glucose
The pharmacologicalactivity of glycerolhas raisedthe questionof the
relationshipbetweenthis substanceand glucosesince some of the manifestationsinduced by glycerol could be obtained with glucose.The same
eftect is seenon local pH in the secondday wound crust. Glucoseadministeredfor a few days prior to, and immediatelyfollowing, wound induction
shifts local pH toward more acid values.A similar effect is obtained on
the pH of tumors.(173) Glucosedecreases
intensityof pain of the alkaline
pattern and increasespain of acid pattcrn. We were able to induce convulsionsin rats by injecting20 to 25 c*. of an isotonicsolution of glucose
subcutaneouslytwice daily. After 4 to 6 days, convulsionsappearedand
often led to death. [n some terminal cancerpatientswith brain metastases,
who had had previousconvulsions,intravenousadministrationof glucoseas
a therapeuticprocedureproduced convulsions.We have even seen lethal
convulsionsin a patient after a few days of intravenousadministrationof
glucosein salinesolution in conjunctionwith intramusculardosesof I mg.
twice daily of deoxycorticosteronc.
Although rarer than for glycerol, hemorrhagesoccurred after each
glucoseadministrationin some patientswith previouslybleedingulcerated
lesions.Bleeding stopped when glucoseadministrationwas discontinued.
Renewedadministrationwas followed each time by renewedbleeding.We
want to emphasizethis relationshipof glucoseto hemorrhagebecauseof
its clinical importance.
It seemspossiblethat glyceric aldehydeand glyceric acid, which appear during glucosemetabolism,play a role in the manifestationsmentioned
above.
GlycerophosphoricA cid
The ability to relatethe pharmacologicalactivity of glycerolto its bond
with fatty acids,has led us to consideranothersubstanceable to bind fatty
acids,glycerophosphoricacid. The bond of this acid with quaternarybases
such as choline or ethanolamine,and the fatty acids results in phospho-

p H A R M A c o D y N A M T c^ c r t v t r y

( plnr

rwo)

369

lipids which take part rn the formation of boundaries and separating

membranes.Although variousfatty acids,both saturatedand especiallyunsaturated,enter into these phospholipids,resulting in a variety of compounds, their generalbiological behavior appearsto be the same. Certain
fatty acids, the di- and tri-ethenic,however, are preferentiallybound as
phospholipidswhen they passfrom the intestineinto the circulation.
In investigatingits influenceas an anti-fattyacid agent,we administered
glycerophosphoric
acid, 50 cc. of a n/10 solutiondiluted in 1000 cc. of
isotonic saline,glucoseor other solutions,intravenouslyor subcutaneously.
It had salutary effectsupon pain of an alkaline pattern and upon corresponding lesions. A manifest influence was exerted upon the systemic
acid-basebalance,especiallyin caseswith high urinary pH and with all
other analysesshowing an offbalanceof type D. No influencewas observed
upon evolution of tumors in animalsor humans.
The increasedbasal metabolismand a marked increasein work capacity, which are observed in subjectstaking sodium glycerophosphatefor
long periods,has made us suspecta possibleeffect upon thyroid secretion.
The appearanceof thyrotoxicosisin a subjectwho had inadvertentlytaken
seemedto confirm this view.
a large doseof sodiumg.lycerophosphate
Sterols
A third group of natural constituents,which act as anti-fatty acid
agentsis composedof sterols,which are absorbedand circulatedbound to
polyunsaturatedfatty acids. Except in brain and red cells where only free
sterolsare encountered,the sterolsare found both as estersand free substancesin aU cells,tissuesand organs.
Cholesterol,phytosteroland a few sterolswere utilized in pure form in
our research.In addition,sterolswere obtainedand used as mixtures,as
in the insaponifiablefractions of tissues,organs,organismsand biological
products.In some studies,thesefractionswere further separatedinto conWe usedcholesterolin
stituents,largelythroughcolumn chromatography.
(Note 4)[Natery or gum cellulosesuspensions
were
differentpreparations.
used for in vitro and in vivo studies.
Cholesterolwas seento inducea changein the shapeof some bacilli,
such as B. subtillis, B. megatheriumand B. anthracis,turning them into
irregularround formations.At the sametime, their Gram positivestaining
became abnormally intense. The agar cultures had a creamy asPect.
The influenceupon Gram positivity explains the fact that Gram positive
individualscould be obtainedin culturesof variousGram negativemicrobes
such as Esch. coli or Eb. typhy after repeatedtreatmentwith colloidal

370

nESEARcH rN PHYSToPATHoLocY

cholesterolpreparationsaddedto broth. The Gram positiveforms, however,

could not be isolated.
Cholesterolwas seento influencered cellsin form, shape,volume, sedimentation, velocity, and oxygen-combiningcapacity. A vermillion color,
which persistedfor a long time, was obtained through in vitro treatment
of blood with cholesterolor through intravenousinjectionsin animals.Although such injections were lethal, they induced the abnormal vermillion
color. Cholesterolproduceda manifestchangetoward lessalkalinevaluesin
the secondday wound crust pH. A favorableeffectupon rabbit skin wounds
was obtained,with abnormallyintensiveproliferationof the epithelium,
this healing effect, howevcr, was less manifest for irradiated wounds.
In subjectswith ulceratedlesions,prolongedadministrationof cholesterol
was frequently observedto induce hemorrhages,especiallyof an arterial
character.However, in patientswith coronary occlusionor endarterialobliterations,the administrationof cholesterolwas followed by an increaseof
symptomsapparently related to exaggerationof the degree of occlusion.
This effect upon blood vesselsalso could be seen in animal tumors. Administrationof cholesterolinduced zonesof necrosisin the tumors which
could be related to proliferation of the cndarterialcells leading to thrombosis and ischemicinfarct. The portions of tumor correspondingto these
ischemic infarcts showed characteristicnecrosiswith unaltered structure
but without the normal staining.
In animals injected with cholesteroland then submitted to trauma in
the Noble-Collipdrum, shockwas prevented.Injectionof cholestcrolprior
to the experiment reduced mortality to zero while in untreated controls
mortality was high. A similar but less constanteffect was obtained when
cholesterolwas administeredimmediatelyafter trauma. It is noteworthy
that in animals injected with cholesterolbefore being placed in the drum.
the blood not only did not becomeabnormallyblack but the usualbleeding
from the nose, mouth and paws (if not taped during the trauma) was
abnormally bright red.
The effectupon cxprimentaltumors was investigatedthrough the dipgenerations.
ping techniquerepeatedin successive
Changesin the evolution
of certain tumors. such as mammary adenocarcinomain mice, were induced. The effect of cholesterolwas similar to that of insaponifiablefracin more detail later.
tion but was lessmanifestand will be discussed
Cholesterol'seffect upon the ccntral nervous system was interesting.
Often, immcdiatelyafter administration,both in animals and humans,
uansitory somnolencewas observed.However. repeatedadministration
inducedconvulsions.(Note 5) Exophthalmiawas seenin mice after injec-

p H A R M A c o D y N A M I cA C r t v r T y ( n r n r

rwo)

371

tion of cholesteroland was most manifest24 hours thereafter.It contrasted

with normals and especiallywith animalsinjectedwith fatty acids or similar lipoids, who showcdenophthalmia.
An ether-oil cholesterolsolution induced paraplegiawith early foot
ulcerationsin rats and rabbits. This occurred particularly in females and
was related to a predominanceof stcrolsin this sex. Castrationor administration of sex hormonesdid not changethis specialsusceptibilityof the
nervous systemof femalesto cholestcrol.However, the administrationof
fatty acidsor of acid lipid preparationsof organsor tissuesdid suppressit.
(Note 21, Chapter VI)Changes in systcmicanalyseswere generallynot
obvious and, when prescnt,were slow in appearing.They correspondedto
changestoward offbalanceA.
UnsaponifiableFractions (lnsaponifiable or Non-saponifiable)
When insaponifiablefractions wcre preparedfrom various tissuesand
organs, big differencescould be seen in the quantity and the number of
sterol compoundsnaturally present.However, a certain specificityrelated
to the origin of these insaponifiablefractions apparedmost interesting.
The insaponifiablefractions of various materials were prepared by the
usual methods.Most of the fractionsare solublein oil in higher proportion
than cholesterol,with some of them even misciblewith oil. More concentrated solutionsin sesameoil could be preparedthan for cholestcrol.In
most of the experiments,5 or lOVo solutionswere used. Colloidal suspensionsalso were preparedin the samemanner as for cholesterol.
In spite of the extremevariationsin sourcesof the insaponifiablefractions, almost all have some propertiesin common. Some are similar to
cholesterol in their eflects particularly at the lower levels. The marked
differences
appearat higher levels.Thcy inducehemorrhages
in the adrenals betweenthe fascicularand reticularzones.
On the healing process,especiallyof radiation wounds, insaponifiable
fractions of placenta,embryos and buttcr-matcrials related to growthshow impressivelygreater activity than cholesterolor preparationsof insaponifiablefraction of other origin. They induce healing processesevcn
in standardized
radiationlesionswherc cholestcrolhas a weak effect.
In their influenceupon tumors, the preparationsof insaponifiablefractions of different organs differ markedly. No changesat all were obtained
with some preparationssuch as from pig intestine,for example,while
interesting results were obtained with others. The differencesappeared
especiallyevidentin experimentsin which a direct influenceupon the tumor

372

nEsEARcH rN PHYSropATHoLocy

was exerted. Transplants of Ehrlich mammary adenocarcinomain mice

were dipped in insaponifiablepreparationsand grafted. In general,no immediatevisible effectswere seenwith this techniquefor the first transplant
generation.By repeatingthe sameprocedurefor following transplant generations,changeswere obtained which varied with the preparationsused.
The insaponifiablefraction of human placenta, for instance, produced
a marked increasein malignancy,together with morphological changes,
the tumor changingfrom an adenocarcinoma
to an encephaloid.Further
treatmentof the transplantsled to still greatermalignancywith a sarcomatoid transformation.Thereafter,negativcresults were obtained with new
treatmentof the transplants.(Note 6/ With this procedure,plancentapreparations showeda manifestinfluenceeven at the third transplantgeneration,
and they were negativepassages
for the fifth to sixth transplantgenerations.
With pig intestinepreparations,even after ten successivepassages,malignancy was unchanged.
The specificityaccordingto origin was also seenin other experiments,
such as in the influenceexerted by thesepreparationsupon the development of specificlesionsproducedby smallpoxvirus in low-reactingspecies
such as mice or rats. Preparationsfrom receptiveanimals, and especially
from organs sensitiveto the virus, were more capable of inducing local
receptivitythan were preparationsfrom refractory animals. For instance,
positiveeffectswere obtained with vacciniavirus in mice and rats previously injectedsubcutaneously
with the insaponifiablefraction of rabbit skin
or brain, while no such effectswere seenwhen the insaponifiablefractions
of pig or hen intestineswere used.
Differenceswere observedbetweenthe insaponifiablefractions of different organsfor conditionsprincipallymanifestedat the organ level. Conditions affecting mainly one organ were treated with the insaponifiable
fraction correspondingto that organ. These preparationsoften appeared
much more activethan thosefrom other organs.We investigatedthe effects
of a heart insaponifiablefraction on patientswith myocardial insufficiency.
especiallywhen responsesto other therapeuticagentscould no longer be
obtained. In like manner, we used the insaponifiablefraction of liver for
manifestliver insufficiency.The rate of liver regenerationin rats after subtotal resectionwas found most acceleratedby liver insaponifiablefractions.
The good effectsobtained in the treatmentof intractablediarrhea with insaponifiablefractions from pig and hen intestineswill be discussedbelow.
We investigatedpreparationsobtained from lymph nodes and spleen for
the treatmentof shock, particularlyin its acute form. Similarly we also
used adrenal insaponifiablefraction to influence induced adrenal insufi-

p H A R M A c o D y N ^ M l c A c T - r v r T y( p , r n r r w o )

373

ciency,and brain insaponifiable

fraction in an attemptto influenceinsomnia. The resultsof thesestudieswill be discussed
in the sectiondealingwith
therapy. The changesobtainedwith the respectivepreparationsindicate
that they have a specificitywhich rcprescntsan importantfactor in normal
and abnormalphysiology.
In a secondgroup of researches,
constituents
of the rough preparations
of insaponifiablefractions from various sourceswere separatedby different methods.The ketonicand nonketonicconstitucntswere obtainedand,
when testedin animals,showedseveraldificrencesin biologicalproperties.
Further researchof specificitywas madeusingseparations
through the
chromatographic
column method.Most of thcsepreparations
are still under
laboratoryinvestigation.Experimentsare being conductedwith difterent
organ preparations,
somefractionsobtaincdbcing identifiedas commonto
all organs,while othersarc specificto onc organ or to a goup of organs.
Theseexperimentsalreadyhaverevealeda markedpluralityof constituents
for the insaponifiablcfractionsof organswhich has to be related to the
pluralityof constituents
found in the acid lipid fractionsof the sameorgans
and which was discussed
above.The specificityseenfor organswould thus
greatly concerntheir lipidic constitucnrswhich form the acid and the insaponifiablefractions.It is especiallyin rerms of specificitythat the acid
lipidic and insaponifiable
fractionsof variousorgansare being investigated
in researchnow in progress.(Note 7)
Corticoids
The study of the defenseof the organismagainstfatty acids focusses
attentiononce againon the adrenalswhoseconstituents
appearto be part
of the naturaldefensemechanism.To date,around 30 differentcrystallizable compoundshave been isolatedfrom less than a third of the total
cortical extract. The amorphouspart, biologicallymore active than the
crystallizedpart, would containother importantcompounds.Even if some
of them are intermediarycompoundsor artefacts,adrenalinterventionstill
is characterizedby plurality of its active agcnts.Furthermore,severalopposite tendenciesare recognizedbctween groups of adrenal compounds.
While all the corticoidsshow a certain antagonisticaction toward fatty
acids,mineralocorticoids
are, from severalpoints of view, antagonisticto
neoglucogenics.
With theseconsiderations
in mind, and recognizingthe adrenalsas one
of the principalmeansfor relativelyrapid defenseagainstnoxiousagents,
we have investigated
the relationshipof the adrenalsto lipids.
We have alreadVnoted the strikins richnessof the adrenalsin arachi-

374

x E S E A R C I {r N p H y s r o p A r H o L o c y

donicacid.About 25Vo of the total fatty acid contentof the glandsis made
up of this acid which is found only in smallquantitiesin other organs.We
alsofound pentaencs
and hcxaenes
prcscntin greateramountsthan in other
organs.These fatty acids nrust have a biological purpose and two hypothesescan be advanced.Accordingto one mcntionedpreviously,(Note I I,
Chapter VI), the corticoidswould be synthesized
from arachidonicacid
throughcyclization.Accordingto thc sccondhypothesis,arachidonicacid,
as well as other high fatty acidsprcsentin the adrenals,would be used as
activefunctionalfatty acids.Secreted
by the adrenals,they would passinto
the circulation and intcrvene as nccded by the organism, especiallyfor
immediatedefcnsepurposcs.
We have seenthat an interventionof polyunsaturatedfatty acids occurs
in the first defcnsercsponscof an organismto a noxious agent.These acids
for the exaggeratcd
are responsible
through which the
oxidationprocesses
organismattacksthe noxiousagentsthemselves
or the heterogenized
constituentsresultingfrom their action. Wc considerthat some of the fatty
acidsinterveningin this defensemcchanismare liberatedlocally,especially
if they appearin responscto a conditionlimited to a lower entity. In this
case,they would come from changesinduccd in the constituentsof the
entity itself. Thc local interventionof lipolytic enzymeswould lead to a
liberationof free fatty acids.In a defcnseresponsefor the organismas the
highestentity, thc activcly intervcningfatty acids appear in the general
circulationin the first phaseof the diphasicphenomenon.Some of these
fatty acids would be of adrcnal origin, liberatedat these moments.In the
secondphaseof thc defensemechanism,
a further liberationof steroidsby
the adrenalswould occur, aimed at counteractingthe effectsof fatty acids.
The diphasicsystemicprocesswhich rcsultscan be consideredto represent
which occur alternatelyand which, through
an exaggcrationof the proccsses
normal oscillations,insurethe dynamic systemicba.lance.
The adrenalsconccivablycontrol abnormalfatty acid activityby their
quantitativelyexaggcrated
interventionand by releaseof qualitativelyabnormal products that would pass into the circulation and result in offbalances.The activity of the adrcnirlsin counteractingthe influenceexercisedby fatty acids has been made the subjectof a specialinvestigation
in our laboratoriesby E. F. Taskier.
By comparingthe doscsof an agentrcquiredto kill normal and adrenalectomizedanimals, it has been possiblenot only to identify this intervention but to judge thc degreeof this specificdefensemechanism.The
"Adrenal DefenseIndex" for an agent-the ratio betweenthe minimal
animals represents
lethal dose in normal animalsand in adrenalectomized

p H A R M ^ c o D y N A MA
r cc r r v r r y ( r l n r

rwo)

375

a numerical estimatcof this rcsponse.It could bc shown that for certain

fatty acids, such as conjugatcdtricncs,uhich are related to trauma, or
alpha hydroxy fatty acidswhich are rclatedto microbialinvasion,a highly
effectiveinterventionof thc adrcnalsoccurs,through releaseof neoglucogeniccorticoids.The administrationof neoglucogenic
corticoidsmanifestly
increases
the resistance
of the organismto the noxiouseffectsof fatty acids.
This influenceis reducedfor thc mineralcrcorticoids
and is nil for sodium
chloride, otherwisean important factor related to adrenal intervention,
(Note 17, Chapter V I )
We will discusslatcr an inrportantdillcrcncc,evcn an antagonism,between thesetwo groupsof corticoidswhcn their influcnceis exertedconcomitantly with that of other agcnts.

Synthetic Anti-FaW Acids

Analysisof the natural anti-fattyacid agcntshas revealedthe importance
of their positivepolar groups.And this has guidedus in attemptsto obtain
synthetic agentswith anti-fatty acid cflccts.
An important step was a study of alcoholswith lipoidic properties,the
lipoalcohols, starting with the primary mono-alcoholhomologousseries.
This study alsohas pcrmittcdus to rccognizcthc importanceof the lipoidic
propertiesfor their biologicalactivit)'.We startedwith butanol which is
the first memberof thc homologoussericsof aJiphaticalcoholswith lipoid
characteristics.
Butanol
Butanol has a spccialplaceamongthc alcoholsthat have been utilized
as anti-fatty acid agents,not only by virtue of its physico-chemicaland
biological proprtiesbut also bccauseof intcrestingtherapeuticresults
obtainedin animalsand humans.
Extensivestudieswith butanolhavehelpedconsiderably
in definingthe
physico-chemicaland biologicaldiffcrenccsbctwccnlipoids and nonlipoids.
Accordingto the conceptadvanccdpreviously,lipoids and nonlipoidscan
be distinguishedby solubility charactcristicswhich are dcterminedby the
energeticrelationshipbctweentheir polar and nonpolargroups.The nonpolar group is predominantin a lipoid; thc polar group is predominantin
a hydroid.Lipoids have grcatersolubilityin neutralsolvcntsthan in water,
and this providesa simplecriterionfor their recognition.
Methyl, ethyl and propyl alcoholsare all equally ntore solublein water
than in neutral solventsand thereforeare rccognizedas nonlipoids.Butanol,
lp!{d}i}r!}-qrt'i:i.".:'/?:-,:ry?:r

tY"1,:'

i,*:',i

jgf;r-J{.9!1:;'.';.r

i}.\-'ifl..il'::r.:;;

?5sJJf,;.ftrtalfg'.?aG,E-*r

if

'-S

l'*
, i ;:16',"1'";

l?6

RDSIJA RCI{

I N

PI"IYSIOPA']'}IOLOGY

t{gc-9-ry
H. Butyl
Alcohol

S e c .B u t y l TertiaryButyl
Alcohol
AIcohoI

Q rch
\IH
I sobutyl
Alcohol

F l c . 1 3 3 . S c h e m r r t i cr c p r e s c n t a t i o no f t h e n r o l e c u l l r s u r f a c e so f t h e . l i s o m c r s t r f
b u t a n o l . T h e c o n s t a n t b o f t h c v a n t l e r W a a l s f o r c e s r c l a t c d t o t h e i r s u r f a c e sa r e
u n e q u a l .A n l i n i m u n r v a l t r c i s s e c n f o r a l n r o s ts p h e r i c n r o l c c u l eo f l c r t i a r y b u t a n o l ,
a f a c t w h i c h e x p l a i n st h e n o n p r c d o m i n a n c co f t h e p o l a r g r o u p i n t h i s m o l e c u l e .r e s p e c t i v e l yi t s n o n - l i p o i d i cc h a r a c t c r .

howcvcr.differs from the lowcr membcrsof the aliphatic alcohol scriesby

being a lipoid, nlorc solublcin ncutral solvcntsthan in water. This. horvcver. is truc only ior thrcc of thc four isonrcrsof butanol.n-Butanol,secb u t a n o la n t i i s o - b u t : t n oal r c a l l n r o r c s o l u b l ci n n c u t r a l s o l v e n t st h a n i n
watcr, r.vhcrclstcrt-butanolis equally solublein both. According to our
c r i t e r i o nt h c r c f o r eu. ' h i l ct h c f i r s tt h r c c : r r cl i p o i d s ,t c r t - b u t a n oi ls n o t .
Thcsc considerations
havc enablcdus to corrclatclipoidal propertics

o
g

(tt
=

co
I
c

o
g

(It

ct

I
(J
OJ

v,

m
I

(f,
.Fl

c)
c

rO
3

m
I

o,

F t c ; . l 3 l . T h c t l i f l c r c n c c sb r t w c c n t h c r o u n d r h a p c r . ' l ft e n i a r v b u t a n o l a n d the longcr

fttrm of thc othcr iserrrcr. i s e v i d e n t* ' i t h r r r o d c l so f n r o l c c u l e s .

p H A R . M A c o D y N A M t cA c r r v t r y

(p,lnr .rwo)

3jj

with one more preciseintermolecularfactor, the prcdominanceof one of

the van der Waalscohesionforces.Comparativeanalysisof the structural
f o r m u l ao f t h e f o u r i s o m e r so f b u t a n o l( F i S . 1 3 3 ) ,r e v e a l st h e i m p o r t a n c e
of forcesrelatedto the surfaceof the moleculesin determiningdiffcrences
i n t h e i r s o l u b i l i t y I. n c o n t r a s t o t h e t h r e c l i p o i d i s o m e r st,h e m o l e c u l eo f
tertiarybutanolis rounderand hencehas a smallersurface.The difference
betweentert-butanoland the other three isomersis apparentlydue to the
cohesionforcesrelatedto the surfaceareaof the moleculc.of the van der
Waalsforces,thosedescribedas relatedto the surfaceof the molecules.or

n - B u t y l s e c - t s u t tyel n t - B u t ylls o b u t y r

F r c . 1 3 5 . W h i l e t h e 3 i s o m e r so f b u t a n o lw h i c h a r e l i p o i d si n f l u e n c et h e s e c o n dd a y
w o u n dc r u s t p H . l o w e r i n gi t s v a l u e s .t c r t i a r y b u t l n o l w h i c h i s n o t a l i p o i d , d o e s n o t
i n f l u e n c ei t .

378

RESE^RcH tN pHysropATHoLocy

as the constant b of the cohesionforces, thus appearedto be most importantin determining

(FiS.I 3a)
lipoidicproperties.
study of the four isomersof butanol has confirmed the importanceof
lipoidic propertiesfor biological activity. Like the lower members of the
homologous series of aliphatic alcohols which arc not lipoids, tertiary
butanol does not influencepH of the secondday crust of a wound, while
the three other isomers,all with lipoidic characters,lower the pH as the
h i g h e rm e m b e r so f t h i ss e r i e sd o .( F i g . 1 3 5)

29o

Contro I s
Butanol

Agein reeks
F t c . 1 3 6 . E . f f e cot f 0 . 5 9 ' bs o l u t i o n o f .n - b u ! a n o la d m i n i s t e r e di n s t e a do f d r i n k i n g w a t e r
u p o n t h c i n c r e a s ei n w e i g h t o f y o u n g r a t s . T h e v a l u c s r e p r c s e n tt h e a v e r a g ef o r 2 0
f e m a l e s ( . . . . ) . N o d i f f e r e n c e sa r e s e e n f r o m n o n t r e a t e dc o n t r o l s ( - ) .

The fact that a saturatedwater solutionat 20oC still contains7.9Vo

n-butanol is of grcat practical importance.Becauseof its degreeof solubility in water, n-butanolcould be utilized in aqucoussolutionsin suffistudiesand could be used
ciently high concentrationfor pharnracological
as a therapeuticagcntin this forrn withoutnecd for an oily solventvehicle.
The acute toxicity dose for butanol correspondsto the narcotic dose
for the respiratorycenterswhich is relatedto interferencewith the aerobiotic life of thesecells.
The minimal lethal dose of n-butanoladministeredsubcutaneouslywas
f o u n dt o b e 4 . 6 - 6 . 4g m . , / K g n rf .o r m i c e ,3 . 7 - 5 . 9g m . / K g m .f o r r a t sa n d 3 . 3 5 , 6 g m . / K g m . f o r r a b b i t s ,g u i n c ap i g s a n d h a m s t e r sT. h e s ev a l u e sc l o s e l y
approximatethe findingsof other workers.The minimal lethal dose of
is very closeto that for subcutaneous
n-butanolinjectedintraperitoneally

p H A R M A C o D y N A M T cA c r t v r r y

(p,rnr rwo)

3'79

and intramuscularadministration,indicatingthat absorptionfrom the tissuesis almostas rapid as from scrouscavities.

W e h a v e a d m i n i s t e r ebdu t a n o li n l a r g ed o s e st o h u m a n subjects,and
theseclinical studieshavc confirmedthe laboratoryfindingsthat the toxic
effectis especiallymanifestthroughthe narcoticeffectand is a t t a i n e do n l y
with the use of very large doses.(ltlote 8)

Controls
Butanol

/6

20

24

29

J2

Ageln t|eeks

F r c . 1 3 7 .T h e s a m c d a i l y c h a n g es c e ni n m a l e r a t s

Long term usc of n-butanolhas virtually no influenceupon normal

physiologyin animals.Administercdcontinuouslyin the drinking water
of young animals,it had no effcct on growth (f iS. 136) It also did not
affect reproductioncapabilitiesof mature animalsor inliuencetheir offsprings.
n-Butanolshows a definiteinfluenceon white blood cells in rats. The
leucocytecount is increasedin adult rats receivingdaily injectionsof a
saturatedsolutionof n-butanol.(Note 9)
The influenceexcrtedby smalldosesof n-butanol,as for other lipoids,
appearsto be almostentirelyconfinedto abnormaltissuesand cells.This
is evident in the influenceupon the pH of experimentallyinduced wounds
in animals.Administcrcdbeforc wound inductions.n-butanolshowedno
influenceupon normal tissue,no differenceswere observedbetweenpH of
their normal tissuesin trcatcdand untreatedanimals.During the first day

380

./

RESEARCH IN

pHystopATltoLocy

following wound inductions,pH of the lesion in trcated animalswas no

differentfrom pH in untreatedcontrol animals.However,by the second
day. pH of the wound crust wasloweredby butanol,as seenin Figure 135.
n-Butanolaccelerated
the healingrate of wounds,althoughthe difterences between treated animals and controls was not striking. n-Butanol
enhancedhealingof radiation burns to some extent but the effect was not
constantin different groups of animals.In severalanimals treated with
n-butanol, radiation wounds hcalcd within two or three weeks. while in
controls healingtook more than four weeks.
Butanol, when administered
to patientswith pain of alkalinepattern,
has repeatedlyprovided relief within a very short time-in some cases
within three to five minutcs.In pain of an acid pattern,exacerbationoccurs, also within a few minutes.Its quick effecthas led to use of butanol
as a diagnosticmeansfor determiningthe pain pattern.
The anti-fattyacid actionof n-butanolhasled to the investigation
of its
effectupon shock since,as previouslynoted, shock appcarsto be related
to interventionof abnormalfatty acids.Administrationof butanol subcutaneously,eventogetherwith largeamountsof saline,is only slightlybeneficial for shockin mice with caloricburns.The additionof sodiumlactate
has markedly prolonged survival time in these animals. (Note I0) (FiS.
/-18l Still better effectsupon traumaticconditionsare obtainedby associating butanol with glycerophosphoric
acid in salineor in glucosesalinesolution. Especiallyeffectiveand well toleratcdis a solutioncontaining0.3-0.5
gm.Vo butanol with n/300 to n/200 glycerophosphoric
acid and with
5Vo glucosein saline,used for intravenousinfusions,as well as for subcutaneousclysis.
T h e a d m i n i s t r a t i oonf b u t a n o li n s u f f i c i e nat m o u n t st o m a n y p a t i e n t s
h a v i n gm a s s i v eh e m o r r h a g ehsa s c l e a r l yd e m o n s t r a t etdh a t t h i s s u b s t a n c e
has a hemostaticeffectwhich will be discussed
below.
After butanol studies,the effectsof other aliphatic alcoholswere investigatedand revealedthe importanceof the nonpolar group in their
biologicalactivity.
Higher Alcohols
T h e s t u d y o f a l i p h a t i ca l c o h o l sh a s s h o w n t h a t o n l y f e w m e m b e r so f
this homologousseries,bcginningwith butanol and ending with octanol,
havean effectupon the s.d.c.pH. For octanol,only half of the test animals
showedchangesin secondday wound crust pH. (Fig. 139) We thought it
worthwhile to study the biologicaleffectsexertedby thesemembersof the
series,including those which demonstratedno effect on the secondday

p H A R M A C o D y N A M t cA c r r v r T y ( p , r n r r w o )

3gl

wound crust pH. Comparativestudiesindicatedefinitedifferences

between
the two groupswith odd or even numbero[ carbonsin thcir ability to act
upon an existingoffbalanceand reducethe abnormalmetabolism.Along
with differences,
many common propertieswere recognizedthrough effects

loo
-No Loctole
./

90

, Untreofed

80
-No

70

=
o

Cl

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NoCl

50

ae

30

n- Bulonol
No loclole

' f r

t/,'

20

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ro

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/I,,,,
6

T i m e ( h r s ).
F t c . 1 3 8 . I n f l u e n c ee x e r t e d b y d i f f e r e n t a g e n t su p o n t h e m o r t a l i t y o f m i c e s c a l d e d
f o r 3 s e c o n d si n w a t e r a t 9 0 ' C . W h i l e s o d i u m l a c t a t e s e e m se v e D t o i n c r e a s et h e
m o r t a l i t y a n d N a C l a n d b u t a n o l i n s a l i n eh a v e l i t t l e i n f l u c n c e .a m a r k e d p r o l o n g a t i o n
o f t h e s u r v i v a l t i m e i s i n d u c e db y t h e m i x t u r e o f n - b u t a n o l a n d N a l a c t a t e .
ffi

"i. J.;' r'-r:; ..S ::-:If't{1l?tl.?ETffd

ir.i,-.."

::"v:''i:*a:.1?riaL:

*i.

;r:'rii:
*,
?'
.i J, .,' -; j ;,:):i I-r\1,:;-*
U0irir{., ;'--fj'",&'}.n':'.i;.q:

hqf*,SJt..'',{r

i:.:_*{fjlt

gj :r,f

382

p . E s E ^ R c l {r N p H y s r o p A T H o L o c y

induced at various levels. In general,the eftects are more profound for

memberswith longer chains.This is true for butanol comparedto hexanol
in the even carbon seriesand for pentanol compared to heptanol in the
odd carbon serics.For octanol, most of the effectsare diminished.In the
group with odd carbon numbers,nonanolhas very little or no influence.
on viruses,a protectiveeffect againstexternal influencessuch as heat
or fatty acids is evident. It is more striking for the even carbon group. In
microbes,little except an antibacterialeflect is produced by membersin
the even carbon group. The odd carbon group induces Gram positivity,
inegularitiesin form with a tendencytoward roundness,and vacuolization.

92

n-HeptyI

n-Octyl

n-l,lonyl

778
776

PH

oo
o
o

771
772

o-o
o
o

oo
7

ooo
1

o
o
oooo
oo
o
:6

F t c . 1 3 9 . T h e i n f l u e n c ee x e r t e db y h e p t a n o l ,o c t a n o la n d n o n a n o l u p o n t h e s . d . c .p H .
W h i l e h e p t a n o l i n d u c e s c o n s t a n t l ya l o w e r i n g o f t h e a l k a l i n i t y o f t h e s e c o n d d a 1 ,
w o u n d c r u s t , t h i s c f f e c t i s l c s sc o n s t a n tf o r o c t a n o la n d n i l f o r n o n a n o l .

p H A R M A C O D y N A M T CA C r r v r T y ( p ^ R T T w o )

393

No important changes occur at the cellular level. We have already

mentioned the difterent effectsupon wound crust pH for these alcohols.
On pain, nonanol has no effcct and hexanol and octanol relatively little.
But pentanol and especiallyheptanolshow a very marked influence,both
immediate and prolonged. It is intcrestingto note another striking effect
at the tissular level, observedonly for heptanol.It correspondsto an abnormal accumulationof fluid in certain abnormal tissues.This was first
seen in surgical scars even severalmonths old. Edema of the entire scar
occurred followed by blistering at thc surface,or even by formation of
fluid-filled cavities in the scar itself. The same phenomenonappearedin
other lesionssuch as tumors, especiallywhen they were infccted, although
there bad been no clinical indicationof infection before the administration
of heptanol.The effect sometimeswas very intense,transformingan entire
lesion, scar or tumor into a cavity with septic fluid exudate, but few
Ieucocytes.
The influence of high doses of heptanol upon inl'lamatoryprocesses
could be judged experimentallyin the gas pouch induced subcutaneously
in rats or mice by injecting nitrogen and subsequentlyinjected with a low
pathogenic microbe. In controls, no unfavorableeffects were noted. In
animals injected with heptanol subcutaneously,
fluid exudate accumulated
(Note
in the pouch in a fcw days.
I I) Subcutaneous
administrationof
heptanolalso inducedan exudatein the peritonealcavity in mice and rats
injected with the same microbial suspension.This did not occur in the
controls. It must be emphasizedthat thcse cffects wcre scen only with
relatively high dosesof heptanol.
At the organic level, while nonanol again showedno activity, the two
higher alcohols,heptanol and octanol, had an influcnceupon the central
nervous system.In humans, evcn in larger doscs,such as 200 mgr. six
times a day (2 cc. of a 10% solutionin oil everyfour hours) repcatedfor
ten days or more, the two higher alcoholsproduccd no abnormal central
neryous systemmanifestations.In some subjcctswho had previouslyhad
convulsiveattacks,administrationeven in small doses,such as 25-50 milligramsoncea day, did induceconvulsiveseizurcs.
If thescsubstances
were
given alongwith desoxycorticosterone,
the latter even in dosesof I mgr. a
day, severeand even fatal convulsionswere produced.Nonanol had no
such effect.Somnolencefollowed by coma was observedwith concomitant
administrationof cortisoneand heptanolor octanol, but nonanol did not
produce this effect either,
of interest was the influenceexerted by heptanol upon the different
analyses.Fig. 1a0 showshow thesevalueschangetoward the offbalanceA

384

,/

sESEARcHtN pHysropATHoLocy

under the influenceof heptanol.lt is to be noted that of all the analyses,the

urinary pH and the blood serumpotassiumare the first to be changed.They
are followed by specificgravity, while the urinary surfacetensionseemsto
be influencedlast.
M. Bier, in our laboratories,has shown that alcohols,when added in
vitro to freshly obtainedblood, reducedblood clot retractibility.It is interestingto note here the relationshipBier has shown betweenthis effect in

/0

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tt

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F l c . 1 4 0 . l n a p a t i e n tw i t h b o n e m e t a s t a t i cl e s . i o n tsh e a d m i n i s t r a t i o no f h e p t a n o l a n d
b u t a n o l s h o w s a p r o g r e s s i v ed c c r e a s ei n t h e v a l u e so f s e r u m p o t a s s i u ma n d i n c r e a s e
i n t h e u r i n a r y p H , t o w a r d o f f b a l a n c et y p e A .

p H A R M A c o D y N A M TAc c r r v r r y ( p e n r r w o )

385

retractibility and other propertiesof the alcohol seriesmembers.Thus, he

could demonstratethat there is a critical value for the concentrationof
each alcohol, when mixed with fresh blood: blood clot retraction is prevented only when this value is exceeded.The critical value varies with the
f e n g t ho f t h e c h a i n ,d e c r e a s i nfgo r t h e h i g h e rm e m b e r s(.F i e . I a 3 ) B i e r a l s o

o
o

to

lr3

/
/

ti
/

r----t/ " ,

Or
i
5

20

.02

.o4

06

.o8

.to

.t2

,t4

.t6

M o l o r C o n c e n l r o l t o n- E u l o n o l
F r c . l 4 l . C l o t r e t r a c t i o n ,m e a s u r e da s p e r c e n tw e i g h t o f c l o t / t o t a l weight of blood,
plotted against molar concentration of butanol in blood. Different symbols used for
blood samplcs of cach animal.

has shown that, since the toxicity of thesealcoholsseemsto be related to

the same factor, a correlationcan be establishedbetweencritical concentration valuesand lethal toxic doses.
This relationship,as shown in Fig, 142, appliesto the membersof
this seriesof saturatedalcohols,but not to alcoholsof another seriesalso
studied.For the latter, the toxic dose is higher than the critical dilution at
which the clot retraction is hfluenced, and this can be explained by the
interventionof the double bond in the molecules.
Systemiceftects were seen for these alcoholsif administeredin sufficient doses.Some specialeffectsalso were seen. Heptanol decreasedthe
sulfhydrylindex in urine analyses,especiallyif it had been high previously.
Octanol'saction was mainly to increasesurfacetensionif it had been low.
Nonanoldid not show any such activityat all.

386

RESEARCH IN

PHYSIOPATHOLOGY

o - One yeor old

t - Three nonlh old
o - One nonlh old

R
I

60

o
o
I

02

04

06

Molor

08

Concenlrolton

to

t?

t4

.t6

- Bulonol

Frc. 142. Clot retraction. measuredas percent weight of clot/total weiBht of blood,
p l o t t e d a g a i n s ts i m i l a r c o n c e n t r a t i o no f b u t a n o l i n b l o o d . A v e r a g e so f d i f f e r e n t a g e
g r o u p a n i m a l sw e r e s t u d i e d .

Jc""*J.$J
\

l"

as

\e 80

Eoo

't!

I
'20

04

08

t2

t6

20

.24

}--m

32

.48

Molor Conc Alcohol

F r c . 1 4 3 . C l o t r e t r a c t i o n ,m e a s u r e da s p e r c e n tw e i g h t o f c l o t / t o t a l w e i g h t o f b l o o d ,
p l o t t e d a g a i n s ts i m i l a r c o n c e n t r a t i o no f v a r i o u s a l c o h o l si n b l o o d .

p H A R M A c o D y N A M tA
ccrlvtry (p,rnrrwo)

387

Polyols
In another study, we consideredthe polyalcohols,bearing in mind the
important role playcd by glycerol in the biological acrivity of lipids. In
animals,ethylene-glycol
and diethylene-glycol
proved to be too toxic for
parenteraladministration.However, near toxic dosesproduced interesting
resultsespeciallyin tumors. Even in relativelysmall subcutaneousWalker
rat tumors, 2.5 cm. in diameter,for instance,necrosiswas constantlyinduced and followed by skin ulceration. The characteristicinfluence of
these alcohols was to induce a necrotic processnot limited to the tumor
alone but affectingsurroundingtissues,
1.2 Diols
By following the influenceexertedby more than one hydroxyl in the
molecule,we tried to relate propertiesof glycerol to those of aliphatic
lipoids.we preparedlipoids havinga polar formarionof 2 or 3 hydroxyls
bound to the first carbonsof an aliphaticchain. The lipoidic charactcr
was inducedby the lengthof the chain. To preparethcsesubstances,
we
started with correspondingalph-hydroxyfatty acids in which the carboxyl
was reduced to a primary alcohol by treatmentwith lithium aluminum
hydride.As a prototype,we studied1.2 octanediol.The lipoidic character
was recognizedby its high solubilityin neutralsolventsand a limited solubility in water.
There were no marked diflerencesbetweenthe effcctsof octanol and
1.2 octanediolin systemicanalyses.Both raisedsurfacetensionvaluesin
particular.However, the new componenthad an effect upon the central
nervoussystemdifferentfrom most other higher alcohols.As mentioned
above, thesealiphaticmono-alcoholsdo not induce convulsionswithout
the concomitantinterventionof anotherfactor. The secondfactor can be
a local condition in the nervoussystemitself, as in subjcctswith cerebral
tumors,or otherswho have had previousconvulsions.It can also be another substance;desoxycorticosterol,
coraminc,glycerolor glucose,when
administered
with octanol,for example,inducedconyulsionsin some subjects.However,1.2 octanediol,in rcpeatcddosesof around200 mgr. daily,
inducedconvulsionsby itself.This could be explainedby the facr that 1.2
octanediolcontainsin its moleculca group which energeticallyresemblcs
glycerol.Beyond this effect,thcre were no manifestdifferencesbetween
this substanceand correspondingmono-alcoholsin influenceupon pain,
tumor growth or systemicmanifestations.

388

x E S E ^ R C Ht N p H y s r o p A T H o L o c y

Lipoalcolwls x,ith EnergeticCentersin the Nonpolar Group

We also studiedother alcoholswith energeticcentersin their nonpolar
group. From various ethenic fatty acids, we prepared the corresponding
alcoholsby reducing the carboxylsto primary alcoholsthrough treatment
with lithium aluminum hydride. Thus we obtained,in addition to oleic
alcohol, linoleic and ricinoleic a.lcoholsfor the nonconjugatedfatty acids
and eleostearicalcohol for the conjugatedmembers,as well as the correspondingalcoholsof an entire seriesof mixtures of fatty acids from safflower oil, cotton seedoil, cod liver oil, acid lipids of organs,etc., and of
conjugatedfatty acidsderived from them. Having two seriesof substances
with the same common nonpolar gloup, but with difterent polar groups,
cooH and oH, we could relate the effect of these alcohols to their respectivefatty acids and thus ascertainonce more the fundamental role
played by the polar and nonpolar groups in determining the biological
effectsof lipoids.
Study of the effectsinducedupon skin viral infection has largely helped
to define the diffcrences.The polar group determinesthe direction of the
intervention-ins1s45edreceptivityor refractivity-but the extent of intervention is determinedby the nonpolar group. For example, the effect is
very much reducedfor oleic and even for linoleic alcoholsand it was similarly reduced, in the opposite direction, for the respectiveacids. Effects
were more apparentfor polyunsaturated
alcohols,ricinoleic and eleostearic
alcohols,polyconjugated
membersand corresponding
acids.
The same antagonismbetweenthe correspondingacids and alcohols
was very clear for systemicanalyses,pain, healingof wounds,and eftects
upon tumors.The extentof the effectsin eitherdirectionis generallydetermined by the natureof the nonpolargroup and its energeticcenters.
This comparativestudy of acids and alcoholic lipoids has permitted
us to arrive at an important conclusionconcerningthe general behavior
of lipoids. Thus, while the nonpolar group is extremelyimportant for the
extent of the changesinduced,it appearsto be secondaryin importanceto
the polar group which determinesthe direction of the changes.And this
explainsthe role attributed to the polar group in the biological activity of
theseagentsand their separationinto two fundamentalgroupswith antago n i s t i cb i o l o g i c apl r o p c r t i e sw, h i c h i s r h c b a s i so f o u r a p p r o a c ht o l i p o i d s .
We studiedthe effectsof alcoholsand mixturesof polyunsaturated
nonconjugated and polyconjugatedfatty alcohols on a larger scale. These
preparationswere obtainedfrom saffiowerand cod liver oil. The immediate
effectupon alkalinepain was nearlycompleterelief.with prolongedtreat-

pHARM^coDyN^Mr( ^crrvrry

(p,r*r rwo)

lgg

ment, the rclief pcrsistedin most cascs.No influcnccupon tumors was

n o t e d i n n r o s tc x p e r i m e n t isn a n i m a l s .I n h u m a n s t. u m o r a r r e s ti n a f e w
than with isolated
caseswas obtaincd.The cffect was more accentu.rted
memberssuchas eleostearic.
linoleicor oleicalcohols.The convulsanteffect
was much lower than for any other lipoalcoholof this group; even large
dosesdid not produceconvulsionsexceptin patientswho previouslyhad
had convulsions.
Effectsupon systemicanalyseswere the sameas for most
of the higheralcohols,manifcstespeciallyupon urinarysurfacetensionand
sulfhydryl index. The polyconjugated
alcoholmixture, as an agent acting
at the systemiclevel, producedeuphoriabut had very little eftect upon
growth and cvolutionof tumors.
With the intentionof loweringthe level at which alcoholswould act.
we studieda specialgroup characterized
by havinga doublebond between
Cz and C3. We were particularlyinterestedin two membersof this series,
allyl and crotyl alcohols.
Other Alcohols
Crotyl alcoholis a lipoid sinceit is misciblewith neutralsolventsand
only slightlysolublein water.Allyl alcohol,solubleboth in waterand neutral
solvents,appearsto be an intermediarysubstance.
Sincc the lVa solution
of crotyl alcohol in saline was painful when given by intramuscularinjection, an oily 2Vo solution was used. No marked differencesfrom the
effectsof previouslydiscussed
lipoalcoholswere seen.
We studiedpolyalcoholshaving,in additionto a primary alcohol,one
o r m o r e O H a t t a c h e dt o t h e m o l e c u l cs, u c h a s 9 . 1 O - d i h y d r o x y - s t e aarli-c
cohol. However, this substancedid not show any propertiesother than
those noted for oleic alcohol.The alcohol obtainedfrom ricinoleicacid,
in which the carboxylwas reducedto a primary alcohol,showeda limited
systemiceffect.Even in larger doscs,the changeswere slow and not intensive,althoughpassagesfrom one pattern to the other could be seen
more often than with other higher alcohols.A stateof euphoriaappeared
in some subjects.The immediatceffectupon pain was lessthan that obwas satisfactory.
There
tained with other alcoholsbut, in many instances.
was no favorableeffectupon growth or persistence
of tumorsin animalsor
humans.In severalcases,on the contrary,rapid growth of the tumor ocof well being.In general,
curred despitelack of pain and evcn sensations
ricinoleicalcoholseemsto act at the interstitiallevel and above.but not
below.

390

/'

RESEARCH IN

PHYSIoPATHoLOOY

Lipamines
Theoretically,it was to be expectedthat lipids with an amine as polar
group would have a marked anti-fatty acid action. We studied severalof
these substancesfrom the standpoint of their influence upon physiopathologicalchangesconsideredrelatedto lipoidic predominance.The first lipoid
of the aliphatic amine seriesis hexylamine.A nonpolar group of at least
6 carbons is required for predominanceover the potent amino radical.
From commercial sources,we obtained aminescorrespondingto the usual
saturatedfatty acids with cven numbersof carbons,ranging from 6 to l8;
a few unsaturatedwith 18 carbons;and heptylaminewith an odd number
of carbons.All thesecompounds,when injected in mice and rats produced
severelocal reactions,often followed by skin ulcerations,even when administeredin oily solutions.For this reason,we tried salts of theseamines
usuallyobtainedwith aceticor hydrochloricacid. Saltsof the lower members of the series,no longer had lipoidic character.However, we could use
hexylaminein an oily solution for intramuscularinjection. It appearedto
be rclatively well tolerated locally evcn in humans. No apparent changes
were seen, however, in systemicanalyses,and the immediate and long
range effects upon pain were minimal. No changes were obtained in experimental tumors except by local treatment,as in ascitestumor, or by
injectingthe product at the level of the transplantitself, in which case the
growth of the tumor was slowedor even halted. A similar effect was seen
when the transplantwas dipped in the oily solution of the product and the
procedurewas repcatedin successive
generations.
On a larger scale,we utilized, both in animals and humans. the salt
obtained from hexylaminewith nicotinic acid. It showed favorable influenceon pain of alkaline pattern, and exacerbatcdpain of acid pattern, but
had no other effects.Heptylamine has been used by others as a hypertensiveagent.In our studies,its hypertensiveactivity appearedto be weak
and transitory.
The study of lipamineswas the startingpoint for an entire seriesof
into the biologicalrole of the amino group, especiallyas it interresearches
venesin a complexmolecule.We have seenthat, like all other polar group6,
the amino group will act as a fixinggoup in a molecule.Its characteristics
appearto be relatedto its capacityto bind the moleculeto other molecules
in a relativelystrongerand more specificway than other polar groups.
The rcalization of more complex chemical polymers as biological entitiesevidentlyis relatedto this capacityof the amino group, This appean
clear when the amino goup is bound to various acidsin the alpha position

p H A R M A c o D y N A MA
T cc r r v r r y ( p r n r r w o )

391

to form the alpha amino acids that enter into the formation of complex
proteins.In amino sugars,the amino goup showsthe same property, producing the polymer formationscharacteristicof connectivetissues.Furthermore, it is the amino group, acting as a secondpolar group, which givesto
alkaline amino acids their fundamental role in the biological realm, as
mentioned above. The alkaline amino acids,like other amino acids,form
polymers through their amino acid group. However, as these polymers,
histonesor protaminesremain reactivethrough the terminal alkaline nitrogen-containinggroups and it is through these groups that they realize new
bonds, such as to nucleic acids,
In a molecule with two polar groups far apart the amino group will fix
the molecule, while the other active radical wi.ll provide reactivity. The
selectivefixation upon certain constituentsin various placesin the organization accomplishedby the amino group localizesthe intcrventionof the
other active groups of the molecule.
we have consideredmany of the biological substancescontaining an
amino group and a secondactive group. Typical examplesare the local
anestheticsin which the amino group acts to fix the molecule, while the
other energeticformationsintervenemore activelyto induce the anaesthetic
effect. Similarly, in epinephrine and ephedrine,the amino group serves
to fix the molecule,while the hydroxyls later intervenemore specifically.
In somemoleculessuch as the alkaline amino acids,the secondactive
group can be another amine or another nitrogen containing group. It was
for this reasonthat we first becameinterestedin investigatingnatural and
syntheticcompoundswith an amino acid to servefor fixation, and another
energetic center to intervene more actively. We studied amino-butanols,
but no particular activity could be found. These agents, however, were
not lipoids. We were consequentlyinterestedin substanceshaving a lipoid
character as well as two active polar groups of which one is an amine. A
study of theseagentsis now in progress.
Procaine
In this group of agentswe studiedprocaine.We becameespecially
interestedin procaine after we had seensome casesof ulcerationsdue to
varicoseveins healed in a few days with only two intra-arterialinjections
of procaine accordingto Leriche'smethod. With the idea in mind that an
we studiedprocainebase.
actionthroughits lipoidic propertiesintervenes,
procaine
in
base sesameoil, or suspensionin Tween or gum
Solutionsof
we
cellulosewere prepared.Parallelwith theseprocainebasepreparations,
srudiedalso the hydrochlorideas well as severalsaltsof procaine,such as

392

nESEARcH rN pHysIopATHoLocy

lactate,glycerophosphate,
where a further effect was sought through the
acidsat which procainewas bound. We studiedall theseagentsbut only
underthe aspectof their activityin relationshipto the dualisticoffbalances.
No specialeffectswere seenupon virusesand nricrobesfrom this point
view.
of
Injectedsubcutaneously
to rabbits,it was seento increasein the
skin correspondingto the place of injection,the manifestations
of a subsequentsmallpoxinoculation.Added to the medium for culturesof tetrahymenapyriformisin higherdoscs,it led to the appcarance
of almostround
forms. we studiedthe survival time through the capillary tubes method
mentionedabove.For progressive
amountsof procainehydrochlorideadded
to a culture of tetrahymena,no immediateeffect of thc life-spanof the
culturecould be seen.In anothergroup of experiments,the procainewas
added to the culture before its inoculationand thc life-spanof the tetrahymenagrown on this medium was studied.Under thesecircumstances
a
prolongationof the life of the cultureitselfcould bc seen,but only in the
cultureswhich were showingalso a slow growth.
Procainein general,and much more evidentin the basepreparations,
was seento reducethe pH valuesof the secondday wound crust. In micc
and rats, the lethal doseswere seento bc manifestcdthrough the appearanceof convulsions.
Sublethaldoseswcre seento lead to convulsionsafter
a few successive
daysof administration.
In humans,an intramuscularinjection with procainebasewas seento influencepain but this effectwas appare n t l yl i m i t e do n l y t o t h ec a s e sw i t h a n a l k a l i n ep a t t e r np r e s e n tT. h i s a n d t h e
fact that the effectswere no more manifestwith procainebase than with
other lipoidic preparationswith positivepolar group, even when administeredin relativelyhigh dosessuchas of 2 cc. of the 57o solutionof procaine
in oil, indicatesthat its actionother than at the placeof the injection,has
to be relatedlargely to the interventionof procaineas lipoid. A special
mention,however,has to be madefor the anginapain which seemsto be
more evidentlyinfluencedby the injectionwith procainebase than other
pains.
The immediateinfluenceexertedupon systemicanalyses,
if any. was in
generalreducedeven aftcr the administrationof | 0 cc. of. ZVo procainc
h y d r o c h l o r i d eI t. w a sm a n i f e s t epda r t i c u l a r l uy p o nt h e u r i n a r yp H . i n d u c i n g
a changetoward more alkalinevalues.This effectwas transitoryand especially seenfor the first injections.With rcpeateddoses,this effectdecreased
evento the point of disappearing.
T h e s t u d y o f t h e p h a r m a c o l o goyf p r o c a i n eh a s e m p h a s i z e tdh e c h a r acter of its activity as a lipoid with a positivepolar group rathc'rthan a
s .h i s v i e w i s c o n f i r m e db v t h e
d i r e c to n e t h r o u s hi t s c h e m i c acl o n s t i t u c n t T

p H A R M A c o D y N A M T cA c r r v t r y

(e,rnr rwo)

393

fact that the sameeffectsare not obtainedwith any one of thc two constituentsof procaine, para-aminobcnzoic
acid and diethylaminoethanot,
administeredseparatedor evcn togethcr.The only differencein this case
betrveenprocaincand its constituc'nts
residcsin the propertiesof procaine
as such, which we relatedlargely to its lipoidic properties.
Furthermore,the studyof the diflerenteffectsinducedby procainecan
be seen to result from a nonspecificeffect in which these agentsintervene
indirectlyupon the metabolicprocesses
whichoccurat the cellsfor instance,
rather than to a direct influenceexertedby procaineconstituentsupon
specificmetabolicprocesses.In this group would enter the effectsseen
upon different enzymatic processesindirectly influenced by procaine
throughthe nonspecific
changesinduced.
wc have investigatedundcr this specialaspcct the use of procaine
hydrochloridcin solutionswith a pH between3 and 4, as indicatedby A.
Aslan,againstthe manifestations
of old age.The low pH seemsto intervene
by preventing,as long as possiblc,the enzymatichydrolysisof procaineand
thus permittingits absorptionand action as a nondissociated
lipoid. we
found that the protectionexertedby the low pH would result from the
chemotacticnegativeinfluenceexerciscdby hydrosolubleacids upon the
whichthuswill reducethe interventionof the leucocyticenzymes
Ieucocytes,
in the processof hydrolysis.
Furthermore,the fact that the bond of procaine,as such, takes place
throughthe aminoradicalof the P A B A explainswhy a high aciditywould
preventthis local bond of procaincto occur at the placc of the injection
and thus would favor its action at other sitesin the organism.This action
is confirntedby the fact that the analgesicpropertiesof procainedecrease
through the low pH, while at thc same time, its other metaboliceffects
increase.
The fact that good effectswere observedwith procainein old age and
a r t h r i t i s .s c h i z o p h r c n iaan d m r n y o t h c r c o n d i t i o n si,n d i c a t c st h a t a n o n specificaction would take place. The fact that the effectsobtained are
similar to thoseobservedin the sameconditionswith other lipoidic agents
with a positivepolar group.pernritsus to seeas principalfactorin the pharmacologicalaction of procainc, its nonspecificinterventionas positive
lipoid. Besidesthis nonspecificlipoidic effect,some othcrs relatedto its
chemicalconstituentsand rvhich would rcpresentspecificadded factors
Thc relationto thc folic acid would be an example.
haveto be considered.
led us to study variouspreparationsin which
All theseconsiderations
was
procaine
bound to differentagents,with the aim to enhanceits nonspecificintervention.Through procaine lactate,malate and citrate, the
anl

itfi-FlIrjj*:clryr

;.is\r,

394 /

n E s E A R c Hr N p H y s r o p A T H o L o o y

chemotacticnegativeeffect was highly increasedand procaine activity enhanced.Bound to maleic or citraconicacids,the anti-fatty acid activity of
procainewas further directedtoward influencingmore specifically,the abnormalchangesoccurringleadingto the appearance
of abnormalfatty acids.
Tle Elements
with the recognition that elementsact biologically in two opposite directions,we becameinterestedin those with anti-D character,capable of
intervening in offbalance D. we studied the direct action of these agents
upon fatty acids, as well as their indirect action upon processesand substancesrelated to the metabolic changescharacterizing the offbalance D.
we separated these inducing elements-according to their series and
the compartments where they predominantly exercisetheir influence. The
following table showsthis systematization.
T,rsla XIX
InducirrgGroup or Anti-D Group
Metals

Non Metals

Series

Seric.s

Compartments

10_ I V B I V I B , V I I I

Organism

Li l

Metazoic

Cellular

N, I
K l TiiC.

Fe

Ni

Zn

Nuclear

Rb

Zr

Mo

Ru

Pd

cd

Submorphologic Cs

Hf

Os

PtlHe

Primary

VIII'

IIIA

_l_-1-

AI

vA furro
F" I
P l c r

Ca

As l B r

Tl

Bi

Fr

To theseelementswe can add othersfrom the lanthanumand actinium

serieswhich have anti-D characteristics.
They would act in the submorphological and primary compartments.
Compartment
Anti-D Elernenls
Submorphologic La
Pr
Pm
Primary
Ac
Pa Np

Eu
Am

Tb
Bk

Ho

Tm

Lu

Many of thesc elements are known to have influences antagonistic to

those of members of the D inducing group. we will consider them in more
detail below.

.irffifi$i

p H A R M A c o D y N A MA
T cc r r v r r y ( r r n r

rwo)

395

concerning their influenceon fatty acids,some memberssuch as those

of the VII A seriesare known to affectfatty acidswith an activenonpolar
group by inactivatingtheir double bonds, while other elementshave antifatty acid action by binding their polar groups. Some exert this influence
indirectly through the metabolismin which they take part,
We have studiedthesedifferentaspectsof the influenceof the elements
which will be presentedhereonly in condensed
form.
Monovalent Cations
One interestingaspectof monovalentcationsis their correlationwith
organizationalhierarchiccompartments.Study of this correlationwas first
suggestedby the selectivedistributionof monovalentcations accordingto
levels of organization.From the previous discussion,it seemsclear that
the distribution into compartmentscan be related to positions of these
elementsin differentperiods.Sodium is the principal cation at levels above
the cell, forming the metazoiccompartment.Potassium,which is the next
higher element in the series,is the predominantcation in cytoplasm.According to our hypothesis,ammonium,with propertiesresemblingthose of
rubidium, can be considcredto be the cation of the nuclear compartment.
We have only limited evidenceof direct anti-fatty acid activity for
membersof this I A series.However, they produce characteristicchanges
in conditionsin which an offbalanccexists.They induce the type "A," as
an antagonisticto the type "D" offbalance,The changesare especially
evident in terms of the function of the compartmentto which the cation
belongs.
Sodium
Sodium, the cation of the metazoiccompartment,correspondsto the
environmentof the seain which the entitiesof this compartmentdeveloped.
We have indicatedpreviously the relationshipbetweenthe time when the
metazoiccompartmentwas formed, the degreeof salinity of the sea, and
the concentrationof the cation establishedas a constant in the compartment. Any excessof sodium is eliminatedthrough the kidneys in order to
conservethe metazoicconstant.If an excessivesupply is retainedfor long,
it favors the appearanceof manifestations,
which correspondlargely to an
offbalanceof type A in the metazoiccompartment.
We could show that the appearanceof aorta atheromasin animals
receivingan excessof dietary cholesterolis promoted by concomitantadministrationof sodium. This occurs not only in rabbits but also in rats in
which a cholesterol-richdiet alone doesnot induce such lesions.Excessof

396

nESEARcHrN pHysropArHoLocy

sodium also favors the appearanceof thiamine-inducedconvulsionsin animals.In rats and mice kept on high salt intake,the convulsantdoseof thiam i n ef e l l f r o m a r o u n d1 5 0 m g r . / l 0 0 g r a m st o b e l o w1 0 0 m g r . / 1 0 0g r a m so f
body weight. Administration of sodium-in the form of sodium chloride
and especiallyas sodium lactate-favorably influencesthe state of shock
which, as we have seen,correspondsto a changetaking place principally
at the metazoic level. In superacuteand acute shock, we consideredthe
excessof sodiumpresentat still lower levels,suchas cellsand tissues.to be
one of the pathogenicfactors.The anomalylies not alone in the excessbut
in the fact that the excessis at a levelto which the cation doesnot belong.
The study of sodium mctabolismin thc light of organizationalsystcmatizationof elementshasrevealedthe importanceof two factors;the combinationwhich is "proper" for an elementin its normal compartmentand
distributionof the elementamongcompartments.
In abnormalconditions.
the unusualcombinationsoccur. At the metazoiccompartment,the bond
of sodiumto chloridecan be consideredto be normal. we have seenthat.
when this combinationdoesnot take place,the result is an anomalycharacterizedby accumulationof excessamountsof sodiumin the immediately
higher compartment.
In stateof shock,for example,the pathogenicanomalyin fatty acids
leadsthem to bind the chlorideion. This removesthe normal combining
factor for sodium which then passeson to the immediatelysuperiorcompartment,the gastro-intestinal
tract, This explainsnot only the excessivc'
passage
of sodiumto thc duodenumbut also its retentionthere. The fact
that sodium is present in reduced amounts in blood and in excessive
a m o u n t si n t h e d u o d e n u mi n t h e s t a t eo f s h o c k .i l l u s t r a t etsh e r u l e d i s c u s s e d
abovewhich we believegovernsthc distributionbetwcenlevelsof elements
in abnormalities.
Potassium
This same rule would explain the distributionof potassium,another
elementof the same series.Potassiumis the principal anti-D cation for
t h e c e l l u l a rc o m p a r t m e n ta, s i t i s o n c o f t h e p r i n c i p a lc o n s t i t u e n tosf t h e
earth'scrust, the environmentin which the nuclei developed.Excessof
potassiumin the cell resultsin a cellular A offbalancewith consequent
active proliferation.Potassiumin excessappearsthus to be the cellular
growth-inducing
factorand its role in cancerhasto be considered
especially
the
at
cellular level. Through the inducedgrorvth,the excessof cellular
potassiumwould thus representthe factor immediatelyresponsiblefor the
invasivephase.

,,|:j:
*dF.i:

p H A R M A c o D y N A M T c^ ( : T t v l r y ( p e n r

rwo)

397

F o l l o w i n gt h e r u l c o f d i s t r i b u t i o no f e l e m e n t sb c t w e c nt h e l e v e l sa n d
c o m p a r t m e n t sa. n e x c e s so f c e l l u l a rp o t a s s i u mw i l l r c s u l t i n l o w b l o o d
potassium.This permits us to associateexccssof cellular potassiumand
hypokalemiain cancerwith activecellularprolifcration.The oppositeoccurs in the stateof shock and in offbalanceD. The amount of potassium
decreases
in the cells. Consequently,
the elementaccumulatesand is retainedin excessivc
amounlsin the compartmcntimmediatclyabovethe cells.
the metazoic.Teleologicallyspcaking,whilc the excessof sodium in the
intestinesand the excessof potassiumin blood could be eliminatedeasily
by means availableto thc organism,they are kept in high amounts in
thesesuperiorcompartmentsas rcscrves,disposablewhcn and if the abnormality disappearsand they can be used properly again.We have seen
horv this samemechanismexplainsthe exccssivcamountsof coppcr in the
blood in cancerpatients.
This redistributionbetweencompartments
explainsanotherfact about
potassium.An excessof potassiumis found in cancercells: the greaterthe
degreeof malignancy,thc g:reaterthe excessof the element.Along with
the cellular excess,the anrountof potassiumin the blood drops to low
values,even below 4 mEq. Thc low blood valuecannot be relatedto lack
of potassium,sincequantitiesof the elementsare eliminatedin the urine.
It has to be consideredas a kind of defensethroughthc higherlevel to an
excessive
amount of the elementat its propcr level.This relationshiphas
led to the comparativestudy of thc potassiumcontent of red cells and
serum as a meansof obtainingindicationsconcerningthe interventionof
this metal at its propercellularlevel.The changesin the potassiumcontent
of red blood cclls are consideredto pararllelthoseoccurring in the cells in
g e n e r a l I. t i s n o t t h e r a t i o b e t w e e nt h c s ev a l u e sw h i c h i s o f i n t c r e s t b
, ut
each value by itself.Low amountsin red cellsand in serumcorrespondto
a quantitativedeficiency;high amountsin both, to an excessof the mctal;
low amountsin cellsand high in serumindicatea metabolicanomalycorrespondingto a depletionof potassiumin the cells as seenin offbalanceD;
while a high amount in cells and with low valuesin thc serum indicatea
cellular offbalanceof the typo A.
Administrationof sodiumand potassiumas therapeuticaids has to be
guided by thesefindings.Isotonicsalineappearsto be adequateas a replacementproduct but it is uselcssto administerit only becausehyponatremia exists, except if this hyponatremiaresults from a quantitative
perspiration.
as in excessive
insufficiency,
The problem to be consideredis
the
normal
bond
for
to
restore
the
cation
at its proper level. Often,
how
w h a t i s n e e d e di s c h l o r i d ei o n s f o r s o d i u m ,n o t m o r e s o d i u m .S i m i l a r l y ,

398

REsEARcH tN pHyslopATHoLocy

with hyperkalemia, if it exists in casesof type D oftbalance, it is not the

hyperkalemiawhich has to be directly attacked;eflorts must be made to
havo potassiumagain normally fixed in its own cellular compartment.The
useof glucose,insulin and ACTH seemsto accomplishthis for potassium.
while administrationof potassiumin casesof cancer with hypokalemia
possiblyproducessome immediatesubjectiveimprovement,it is constantly
followed by an exacerbationof tumor gowth as long as it does not correspond to a quantitativedeficiency.In caseswhere this potassiumquantitative deficiencycan be eliminatedas the causeof hypokalemia,beneficial
resultsare obtained through administrationof agentssuch as magnesium
sulfateor calcium salts.
Searchingfor a cation that might competewith potassiumand sodium.
we fust chose ammonium. Theoretically,it seemedto be a likely choice
sinceit penetratesinto the cell and nucleuswith ease.In thc pharmacology
of ammonium,the missinglink is the factor or factors-the substancesand
conditions-which dctcrmine the role of this cation in the nuclear compartment. Ammonium proved valuelessbecauseit was taken up by the
liver and transformedinto urea. Therefore,we resortedto the use of anothcrmonovalentcationof thc samehomotropicI A scriesbut with a higher
atomic weight, rubidium. This cation is very similar to ammoniunt ion.
Rubidium and ammonium,in nitrates,sulfatesand cspeciallyin double
sulfatesof aluminum or magnesium,are isomorphic. We tried rubidium
salts in animals and found them to have a very low degree of toxicity.
In order to study its influence upon sodium and water retention in
we administeredl-2 cc. of a 5%
lesionsin which fatty acidspredominate,
rubidium chloride solutionin water, two or three times daily. In several
c a s e st,h i sc a u s e dd i u r c s i sa n d s i g n i f i c a nr te d u c t i o ni n c d e m a .W i t h t h e i d e a
of havingrubidiumact at the nuclearlevels,wc consideredthe usc of rubidium compoundswith anions that seemto interveneat these levels. Rubidium nucleinatewas preparedand l-2 cc. of a l0% solution in water
administeredto subjectstwo to three times daily. Although thesestudies
are not yet sufficientlyadvancedto permit any conclusionsto be drawn, it
seemsthat the rubidium saltsmay be usefulin casesof intractableedema
relatedto a local condition.
These studiesof the distributionof cationsat different levelsof organization appear to be extremelyimportant if we want to reach cellular and
cspeciallynuclearlevelswith cations,particularlyas radio-activeisotopes.
We tried to go still further and utilize heaviercations.Experimentsu"ith
cesiumsaltsseemto presentmore difficulties,at leastfor the present,be-

p H A R M A c o D y N A M t cA c r r v t r y

(r,rnr rwo)

399

causeof the insolubility resultingfrom the high atomic weight of this element.
In the VI B series, we know little about chromium. Molybdenum
appears to be an active agent. The influencexerted by molybdenum is
neutralizedby the action of methionine,with its active thiol group. Excess
of molybdenumfound in somepasturesinducesa deficiencyin copper and
calcium in anima.lsfollowed by osteomalacyand bone fractures,just as it
induces low fertility. (174) The antagonismbetween these elements is
shown by the inhibitory effect of ammonium molybdateupon oxidaseactivity of ceruloplasmin,the form in which copper is bound to protein in
blood serum. (175)
The anti-D effect of molybdenumis especiallymarked in microbes,in
which it induces morphologicaland tinctorial changes.Bac. anthracis
treatedwith ammoniummolybdateshowsa cocciformchangeand abnormally intensiveGram positivestaining.
Iron
Iron, a member of the VIII seriesand an anti-D agent belongingalso
to the cellular level, is of specialinterest.Its form of activity as cytochrome
oxidaseor hemoglobinhas facilitatedthe understandingof its interveotion.
We could thus relate the high or low amountof the red cells in hemoglobin
which correspondsto the amount of iron ions to thc respcctivooffbalance.
Hypochromia correspondsto a type D offbalance and hyperchromia to a
typ A. These patterns, which were first recognizedthrough clinical investigationsare in accord with the anti-D characterof iron at the cellular
level where it belongs.The existenceof offbalancesof A and D types in
cancer explainsthe high and low values of Fe in the immediatelyhigher
level, the blood serum. Fever, as we have noted, correspondsto an offbalanceof type A at the metazoiccompartment.The fact that serum iron
is low during fever would indicate increasediron activity at the cellular
level, which accordswith its A-inducingrole. We administerediron compounds to cancer patientswith hypochromicanemiato correct the anemia
and also becauseof iron's anti-D effectwhich might act antagonisticallyon
sulfhydryl groups.In severalcases,with very high sulfhydryl index values,
ferrous sulfate has been administeredin dosesas large as severalgrams
daily. In addition to producing an increasein hemoglobinand color index,
ferrous sulfate in large doseshas been observedto have another effect in
some cases,reducingpain of an alkaline pattern.The salutary effect upon
pain was noted more often with reducediron than with ferrous sulfate.No
other influenceupon systemicchangeswas observed.

'irffi

400

nESEARcHrN pHysropATHoLocy

In tumors, iron produced an increase in rate of growth as expected.

This peculiar effect of iron administrationcould be clearly seen in a case
of lymphaticleukemia.
B.v., 4 years old, came under our care with the diagnosisof subacute
lymphaticleukemia.He had a count of 145,000leucocytes,of.whrch 96vo
were lymphoblasts.Butanol administeredin small dosesreducedthe number of leucocytesconsiderably,even after a few days of treatment. After
two weeks, the count was below 5,000 leucocytes,with the proportion of
lymphoblastsdecreasedto 6vo. A count of 3,200 leucocytesmade us discontinuethe administrationof butanol. In three weeksthe count rose progressivelyto previous values. Butanol was again administeredand again
the blood count showedthe samemarked decreasein leucocytes.
All through this "remission," intensivehypochromic anemia persisted
and led us to administerferrous sulfatein addition to butanol. Within
a few days after iron was added,the white cell count increasedto 110,000
feucocytes/cmm.,and the proportion of lymphoblastsrose to 90vo. when
the iron was stoppedand butanol continuedalone,the leucocytecount fell
again,to 8,000/cmm.Iron wasagainadministered
and the total white count
rosea secondtime in three days,from 8,000 to 80,000, with 96Vo lymphoblasts.when the iron was discontinued,the count again fell back within
a week to between6,000 and 7,000, this time with less than 307o lymphoblasts.This continued for another two weeks,when iron therapy was
institutedfor the third time, the count went up again to 38,000 within a
singleday. When butanol was administeredalone,it went back to 5,600 in
anotherfive days. The boy died a few weekslater in acute shock during a
blood transfusion.
While generalizations
cannot be made from thesefindings.it could be
definitelyestablishedin this particularcasethat the administrationof iron
was followed by a marked increasein the number of leucocytesand in the
proportion of lymphoblasts.We saw this repeated,although not so spectacularly,in another caseof lymphatic leukemia.
In severalcancerpatientswho receivediron in large dosesover a proIonged period of time, tumor growth seemedto be stimulated.In animal
experiments,tumor transplantsgrew only slightly faster in animalsfed iron
than in controls.Mice and rats usedin thesestudiesreceivedapproximately
0.05 grn. daily of reduced iron per 100 grams of body weight, mixed in
powderedPurina chow, or were given a correspondingamount of ferrous
sulfatein drinking water. Iron was added to the diet of the animals two or
three weeksbefore tumor transplant,and was fed after transplant.Tumor
growth was slightly enhancedand survival time shortened.

p H A R M A c o D y N A M T^cc r r v t T y ( p , r n r r w o )

401

Similar studieswere carried out usingnickel and zinc alone or mixtures

of them with iron. The metalsrcducedby hydrogcnwere obtainedand a
powder preparationwas incorporatedin powderedPurina chow in amounts
calculatedto provide approximately0.05 gr. metal/100 gr.of body weight
daily. Significantchangesoccurredin the evolutionof Walker tumor transplants in rats receiving0.05 gr. daily of zinc or nickel/I00 gr. of body
weight. In most experiments,two different results were noted. Tumor
growth was retarded in a significant proportion in one group of animals,
with the tumor disappearingin some cases.In another goup with the same
Walker tumors, tumor growth was stimulated.It is interestingto note that
retardationoccurred only in tumors with a necrotic, ulcerativecharacter.
while stimulation was noted in those with white masses.The correlation
of necroticand ulcerativetumors to an offbalanceof type D and of massive
non-ulceratedwhite ones to offbalanceof type A explains this paradoxical result.This would confirm the anti-D or A-inducing characterof these
metals.
Zinc
Zinc intervenes with a certain specificity in one group of metabolic
changes,those related to carbohydrates.Both pancreas"A" cells, which
manufactureglucagon,and B cells which produce insulin, contain large
amountsof zinc. Zinc regulatesglucosemetabolismthrough its influenceon
insulin. Znc and insulin give insolublecombinations,Zinc chloride retarding the action of insulin. (176) A diet rich in glucosedepletesthe Langerhaus isletsof zinc, (177) while protein and lipids, or even fat, increasethe
zinc contentof the pancreas.( 178) With alloxan,the isletslosetheir physiologicalcapacityto storezinc. (179) Indirectlyzinc apparsto haveanti-D
activity.
A similar anti-D activity for zinc can be scen in the prostate,which
is particularly rich in this element.Conceivably,zinc's role would be to
favor the persistenceof sperrnatozoids.
The capacity to utilize zinc is lost
in the abnormalprostate.In adenomatoushypertrophyof the prostate,zinc
valuesdecrease.(180) They decreasestill more with cancerof the prostate. (l8l) Although cancerhas beeninducedby excessive
administration
of zinc, the element'srole in pancreasand prostateseemsto be indirect,
through the metabolic changesit influences.calcium is antagonisticto
zinc, which is to be expectedconsideringthe opposite fundamental biological groups to which they belong.
It is interestingto note the influenceexertedby the oral administration
of zinc powder upon radiation effectsin mice of C3H strain receivinglethal

4OZ /

xESEARcH rN PHYSToPATHoLocY

dosesof 1500 r. In differentexperimentsin which 85 to L)OVoof the controls died in lessthan twelvedays,the mortality rate for animalsgiven zinc
rangedfrom 25 to 50%. In one g'oup, a mortality rate of ISVo was observed.There was a much weakereflect when nickel was incorporatedin
food. No effect at all could be obtainedwith iron.
Mercury
Mercury, a nrember of the IIB series was studied. Theoretically, it
shouldact upon the sulfhydrylgroups,and thus limit the processes
in which
thesegroups take part.
A seriesof compoundsin which mercury is presentin the anion, and
which are routinely utilized as therapeuticagentsbecauseof their diuretic
action and their effectson clectrolytemetabolism,was investigated.Because
of their well-known diuretic effect,we initially used them in patients with
generalizedanasarca.We extendedtheir use to caseswith localizededema
which could be related to tumors with local alkalosis.It was noted that
the mercurial diuretic, in addition to influencingwater and sodium excretion, changedother systemicanalysestoward type A, although only temporarily. However, when these substanceswere administeredover long
periodsof time, other effectswere observedwhich could not be attributed
to diuretic action. In animals with slowly growing tumors, mercury was
found fixed with a degrec of selectivitywithin thesc lesions.After treatment for a certain time, peritumoralfats becamerich in mercury, as recogniz*d macroscopicallythrough abnormal ash color and confirmed by
histochemicalanaiysis.It was interestingto note that mercury appeared
largelyin the necroticpart of tumors.A beneficialinfluenceupon evolution
of tumors was seen,if a type D offbalancewas present.
Bismuth
Bismuth, from the VA serieswas studied.We utilized available antilueticcompounds.Again localizationin peritumoralfats was seen,with the
fats this time becoming abnormally reddish. Neither in animals nor in
patientscould other important effectsbe recognized.
Arsenic
Arsenic, from the same VA series,belongsto the cellular compartment. Its manifest A inducing or anti-D eftect was noted in all compartments if the amount administeredwas high enough. It is interestingthat
Bac. anthracis,under the influenceof arseniousacid, changesto cocci

.'ilru-ff

.,
. i,$i(
i df,,

p H A R M A c o D y N A M TA
ccrrvrry (r,rnr rwo)

4O3

highly irregular in dimension,with intensiveGram positivestaining,and a

creamy characterof cultures. (182)
The carcinogeniceffectof arsenichas beenwidely investigatedby many
authors.In our experiments,arsenicin various preparationsdid not show
a carcinogeniceffect, It did, however,enhancethe effect of various carcinogens,and thus appearsto be an activeco-carcinogen.
Its action at thc
cellular level would explain this effect.
Aluminum, from the IIIA series,belongsto the metazoiccompartment. In minimal doses,it producesa D systemicoffbalance.Boron, as
boric acid, showsfew effectsother than in gastrointestinaldisorders.This
effect may be due to the fact that it belongsto the organismcompartment. Insoluble compoundsof boron appearedto be useful in the treatment of diarrhea.
The effect of membersof the VIIA seriesappearsto be largely correlated with a dircct action upon the double bonds of the nonpolar group of
the fatty acids.
Chlorine
We have noted previouslythe capacityof chloride ions to bind fatty
acids.The bond representsthe first step in abnormalmetabolismof sodium
chloride. In a secondstep, sodium forms alkaline compoundsby binding
with the carbonate anion and, in sufficientamounts, induces local alkalosis. Accumulatedin cells along with water, the sodium compound leads
to the appearanceof vacuoles.In interstitialfluid, the sameprocessinduces
edemaand pain of an alkalinepattern.At the systemiclevel, it resultsin a
state of shock.
In the last analysis,the influenceof the alkaline sodium compounds
can be consideredto result from the lack of anions other than carbonate
availableto bind sodium. For this reason,we were interestedin studying
the effectsof substances
able to furnishthe chlorideanion to the organism.
Through metabolismof ammoniumchloride and calcium chloride,chloride
ions are liberatedin the body. They have little effectsat lower levels.An
immediateinfluenceon local pH is seen at thc tissuelevel. Favorable influence on alkaline pain is part of this action. The effect of ammonium
chloride in shock is relatedto the fact that it furnishesthe neededchloride
ions.
The action of chloride upon alkaline sodium compounds,however, is
handicappedby anothcr aspectof its intervcntion.Studicsof the pathogenesisof shock has shown a noxious effect producedby compoundsresultingfrom the bond of chloride ions to fatty acids.The gastriculcerations

404

IESEARcH

rN

pHystopATHoLocy

seenin the stateof shock with the severeliver damageproducedby several

chlorine compounds----could
be largely rclated to the bond of chloride ions
to fatty acids. This led to the idea of trying such combinationsin vivo to
destroyabnormalentitiessuch as tumoral cells. By using lipoids rich in
chlorides,we hoped to achievethis without the noxious effect of free sodium ions.
We administeredtrichlorethylene
and chlorbutanol(the last also used
as an antisepticin many pharmaceutical
preparations).No effectwas seen
at any level in experimentson animalsand humans.In a secondstep, ws
added chlorine to various lipoids, especiallythose with negativepolar
g r o u p s .W e s t a r t c dw i t h 9 , l 0 - d i c h l o r o s t e a r iacn d 9 , 1 0 , l Z , l 3 - t e t r a chlorostearic
acid. The resultswith thesepreparationsin animalsand in a
few humans were not encouraging.Investigationof products obtained
throughfixationof chloridesat the doublebond of conjugatedfatty acids,
has shown that, in large amounts,they are able to induce gastriculcerations in rats and rabbits.This brought us back to the use of chloridesas
anti-D agents---ofwhich sodium chloride appearsto be the most effective.
Fluorineand Bromine
In anotherseriesof experiments,
we tried to replacethe chloride ion
with another halogen.we studiedthe influenceof fluorine,bromine and
iodinecompounds,this time upon the proccsses
that induceabnormalpatterns relatedto predominance
of abnormalfatty acids.The administration
of sodium fluoride and of other compoundscontainingfluorine did not
havc any appreciableeffecteither in animalswith tumors or other pathological conditions,or in humanswith alkaline pattern of pain. The fact
that fluorinebclongsto the organismlevelled us to investigate
it in terminal cases.Neither pain nor tumor evolutionwas changed.Bromine, too
except for a sedativecffect, did not influencesystemicchanges,pain or
tumor growth.
Iodine
The influenceof iodine was rather extensivelystudiedbecauseof its
relationshipto tumors and becauseit belongsto the nuclearcompartment.
Beforcthe daysof pathologicaldiagnosisof cancerand serologicalmethods
of detectingsyphilis,iodine saltswcre used to differentiatebetweengummatousand tonguecancers.This was the so-called"pierre de touche"treatment, since iodine w:rs assumedto favorablyinfluenceluetic lesionsbut
m a r k e d l ye n h a n c en e o p l u s t igc r o w t h s .
ln view of the dualisticconccptof cancer.wc were interestedin ascer,:n::
i.:l

p H A R M A C o D y N A MAI cc r r v r r y ( e , r n r r w o )

405

taining whether the eflects of iodine were related to one of the two offbalancespresent,and if this was so, to try to take therapeuticadvantage
of it. Iodine was administeredprincipallyin the form of potassiumiodide
solutionsand Lugol's solution.In most caseswith alkaline pain, the intensitydiminishedand pain evendisappeared
soon after iodine administration. In severalcases,dosesas low as 3-10 drops of Lugol's solutionwere
sufficientto induce such an effect for hours. However, larger dosesor repeated doses produced cffects that wcre distinctly undesirable.Edema
within the neoplasticlesionwas increasedby the administrationof iodine,
sometimesto such an extentas to requirediscontinuingits use before any
other changesin the tumor could be noted.
Researchalso was done with lipid and lipoid moleculesincorporating
iodine. The purposewas to determincwhethcrthesemoleculeswould act
more selectivelyupon abnormal cclls and induce local toxic effects.9,
l 0 - d i - i o d o s t e a r iacc i d a n d 9 , 1 0 , 1 2 . l 3 - t e t r a - i o d o s t e a rai c i d w e r e p r e paredand testedin animalsand humans.No differcnccs
wcre notedbetween
thesesubstances
and inorganiciodinc preparationsin influenceupon syst e m i c a n a l y s e sp, a i n o r t u m o r g r o w t h , i n b o t h a n i m a l s a n d h u m a n s .
Oxygen
Oxygen, an agent with a ncgativeanti-A charactcr,at the organism
level,acts like an anti-fattyacid agentat the lower tissularlcvel. For that
reason,we will discussit here.
At the beginningof our work, we werc intercstedin determiningthe
relationshipbetweencellularmembranepermeabilityand the pathogenesis
of the two offbalances.
If a changein the permeabilityof cell membrane
was the primary mechanisminvolvcd,changingthe oxygentensionin or
around entitieswhere offbalancesoccur misht result in thc correctionof
the abnormal manifestationsprcsent.
Clinical studieswere made in which oxygen was administeredto patients with acid pain pattern.The pain was not relievedas expected.Indeed, its intensitywas even increased.These experimentsindicate that
impairedcell permeability.if it exists,is not thc major factor in the pathogenic mechanisminvolvedin the acid pain pattern.Actually, thesestudies
indicatedthat another pathogenicmechanismwas involved since oxygen
administrationincreasedthis pain. According:ly,
it had an oppositeeffect
the pain being
on alkalinepain. The intensityof alkalinepain decreased,
often entirely relievedby administrationof oxygen.We found that pain
producedby traumatic injuries.which was subsequently
identifiedas inpattern,
the
alkaline
could
be
satisfactorily
variablyof
relievedby oxygen,

406

xESEARcHrN pHysropATHoLocy

Such relief occurred in patients who had suffered all sorts of traumatic
injuries, from superficialwounds to severecomminutedfractures.
A curious phenomenonwas seento occur which limited the practical
usefulnessof oxygen. After fust relieving pain, continuation of oxygen
administrationled to appearanceof a new pain. The patient was able to
distinguishbetweenoriginal and new pain by is localization,by the different quality of the sensation,and also by the fact that insteadof being releved by oxygen, the new pain tended to increasewith the continued
administrationof oxygcn.It disappeared
soon after oxygenadministration
was discontinued.If oxygen were administeredagain, the new pain returned within a short time. The new pain might becomesevereand even
unbearablervith continuationof oxygenadministration.
on the other hand,
when administrationof oxygen was stopped,the original pain again appearedwithin l0-20 minutes.
The appearanceof a "new pain" and the resemblancebetween thcse
changesand those observedwith the use of lipids, suggesteda change in
the pain pattern itself, although the changesinduced by oxygen evolved
over minutesinsteadof days.This was confirmedby following the responsc
to acidifyingand alkalizingagents,the only adequatemeansto investigate
the patternin thesecases.By this test the new pain was found to be of an
acid pattern.
Bccauseof the possibilityof inducinga period of calm betweenthe
old alkalinc and new acid pain, oxygenadministrationstill seemedto bc
useful.In casesof traumaticpain, alwayswith an alkalinepattern,successful resuls were obtained.A necessary
condition appearedto be the physical
and mentalability of the patientto guidethe administrationof oxygen.He
had to recognizeif too little or too much was being administeredon the
basisof the di{Iercntsensations
felt, and consequently,
to adjust the administrationto the optimum amount. Extreme pain causedby extensive
traumatic injuries was controlledvery successfullywhen the patient could
be taught to utilize oxygenproperly.
In addition to pain relief, an effectupon cvolutionof the lesion iself
was manifcst.In a few dayswith this form of oxygentreatment,guidedby
the subjectivesensations,
thc healingprocessitself was observedto be sufto
make
ficiently advanced
the pain disappearentirely. Healing of the
woundsseemedto be greatlyenhancedby oxygen therapyguided by the
patient.In severalcasesof open comminutedfracturesin which amputation was considcrcdinevitableon admission,unexpectedimprovementwas
noted. Atonic rvoundswere transformed,becomingrich in granulations.
and hcalingwas rapid.

p H A R M A c o D y N A MA
T cc r r v r r y ( p , r n r r w o )

40j

However, the nature of the treatmentwas such that it could only be

successfulwhen properly applied. If dosageswere too lorv, thcre was no
sedativeeffect; and if doses were too large, new pain was induced. The
physicaland mental statusof the patient thus appearedto be the determining factor for success.The use of an oxygen tent, in which the amount of
oxygencan be carefully adjusted,helpedin a few casesto maintain proper
dosage.But evenin this situation,the patienthimselfmust furnish information not so much about intensityof pain, as about changesin the charactcr
of pain.
The relationshipbetweenthe amountof oxygenadministeredand thc
clinical results,especiallyin pain, has suggestedthat this factor may be
signfficantin other conditions in which oxygen therapy is used. The fact
that, despiteits generalusefulnessin the acute stageof myocardial infarction, oxygendoesnot alleviatethe pain in somecasesand may even increase
it, suggeststhat the amount administeredmight not be adequate.In such
cases,a decreasein pain intensityfollowingtemporarydiscontinuationof
the use of oxygenwould indicatethat the amount utilized was too high. If
of the oxygenadministration
the suppression
is followcdimmediatelyby an
increasein pain intensity,the amount prcviouslyadministercdhas to be
consideredtoo low.
The possibility that too much oxygen can inducc a proliferation of
vesselsand connectivetissue,as seenin the fibroblasticretinopathyof premature babies kept in an atmospheretoo rich in oxygen, fits in with the
data mentionedabove.We have noted that too much oxygeninducedan
anoxybioticprocesswith anaboliccharacter.This explainsthe abnormal
type of offbalancewith proliferativetendenciesseen in fibroblasticretinopathy. We will return later to a discussionof this important factor in
oxygen therapy.
On the basisof findingsin traumatic pain, we studiedoxygen in cases
of painful cancer.Attempts were made to employ it as a diagnosticaid to
help determinethe acid or alkalinecharacterof pain presenton the assumptionthat oxygenwould intensifythe first and would relievethe second
pain. In a group of subjects,we comparedthe diagnosisof the existing
pattern through concomitantvariationsin pain intensityand urine pH, the
responseto acidifying and alkalizing substances,
and responseto oxygen.
In most of thesecases,accuracyof the informationfurnishedby the last
methodwas confirmed.However,this methodhas showngreat limitations.
Whereasmost of the patientswere able to recognizean immediatechange,
they were less precise about a secondchange when it occurred. At the
presentstageof this research,it appearsthat judgment concerningthe de-

408

R E s E ^ R C Hr N p n y s t o p A T H o L o c y

velopmentof changcscould be inrprovedby rcducingthc concenrationof

o x y g e na d m i n i s t e r c dt h
. u s i n c r e a s i n tgh c l e n g t ho f t i m e d u r i n g w h i c h t h e
changeswould appear.In gcneral,thc resultswhen appliedas routine were
insufficiently
clcar to bc usedas a practicalmeansfor the diagnosisof the
pattern.
Oxygenalso has been tricd as a therapeuticagentfor controllingalkaline pain pattern in cancercases.Unfortunately,even in patientswho are
able to analyzethe variationsin pain character,the time betweenthe decreasein intensityof the originalpain and the appearance
and increasein
the intensityof the ncw pain is so variablethat it is almost impossibleto
adjust the dosagcof oxygensatisfactorily
to obtain a long enoughperiod
of calm.
We soughta theoreticalexplanationfor oxygcn'seffcctsupon pain. As
mentioncdabove,thc rcductionof pain of an alkalinepatternand the appearanccof a pain of an acid pattern.are in accordwith our view of the
pathogencsis
of thescpains throughthc intcrvcntionof the two groups of
l i p i d s ,f a t t y a c i d sa n d a n t i - f a t t ya c i d s .A t e n t a t i v e x p l a n a t i o cn a n b e f o u n d
in the activerole of oxygcnupon fatty acids.In the presenceof increased
oxygentcnsion,it is possiblcthat oxygcn is fixed in greateramountson
abnormalfatty acids,thus rcducingtheir intcrventionin chloride metabolism. With lesschloridesfixed by fatty acids,alkalinecompoundswould
b e r e d u c e dl.' h i s w o u l d e x p l a i nt h e i n f l u e n c co f o x y g e nu p o n p a i n w i t h a n
alkalinepattern.Thc appearancc
of an acid pain patternproducedby increasedanoxybiosissccmsto be cxplaincdby thc fact that inactivationof
fatty acids,if it gocs bcyondccrtain limits, changesthe balancctoward a
predominance
of sterols.Besidesthis mechanism.
anotheralsocan be considercd.Any action upon fatty acidsthenrsclves.
would rcducetheir availability as activeagcnts.It seemspossiblcthat under highertension,oxygen
is bound to thcscfatty acidsin a wav diflcrentthan the bond which leads
to the appearance
of activatedoxygen.Through it, the role of the unsaturated fatty acid in activatingoxygen for the cells would be to decrease
intracellularactivatcd oxygen and thus change cell metabolismto the
anoxybiotictypc. Sincethe bonds are labilc. return of oxygen tensionto
normal allowsthe fatty acidsto recovcrtheir functionof activatingoxygen.
Teleologically,
this processcan bc interprctedas a mechanismto prevent
passageof excessoxygcn into thc cells when the externaloxygen tension
increases.
Studiesof oxygcn in normal and abnormalphysiologyhave led us to
consideralso the possibleutilizationof variousoxygcnatedcompoundsas
therapeuticagcnts.A distinctionmust be nradc betweenhydroperoxides,
which occur normallyin orqitnisnts.
and pcroxidesand epoxides.Peroxides

pHARMAcoDyNAMTc Acrtvlry

(p,rnr rwo)

409

result from the binding of molecularoxygen, while the epoxidesresult

from binding of atomic oxygen,both undcr abnormalconditions.peroxides
were administeredas adjunct agentsin caseshighly refractory to therapy
with lipoids having negativepolar groups. In preliminary experiments,it
could be seenthat a certainconditionwas necessaryto influencethe desired
processes.The peroxide used has to be a lipoid if it is to have influence
upon the lipidic system.some lipoidic peroxideswere prepared,and their
therapeutic value is still under investigation.The influence of lipoidic
epoxidesupon the processof carcinogenesis
also is under study.
Peroxidases
It has been previously noted that hydroperoxidesresulting from the
oxidation of mono- and polyunsaturatedfatty acids are found normally in
the organism,but pcroxides appear only under abnormal conditions and
particularlywhen abnormalfatty acidsintervene.The manifestationsof the
type D offbalancein its oxygenphasethus can be attributedto the presence
of peroxides.Biologically,the interventionof peroxideswould be counteracted by peroxidasesand catalases.It was interestingto study a clinical
curiosity which could be connectedwith a probableinterventionof peroxidases.
In severalpatients with frequent headaches,whose analysesshowed a
typical acid pain pattern, pain was repeatedlyintensifiedor even induced
by eating pears. No other fruit had such effect; some had the opposite
eftect.This led us to considerthat the pain intensifyingaction was not due
to an acid-basechange.The large amount of peroxidasesin pears led us
to isolate this enzyme in order to study its direct influence upon pain.
Peroxidasecould be obtained from pearsin relativelysmall amounts,and
showed reduced activity upon peroxideseven in vitro. We were able to
prep:r.remuch larger quantities of a highly active peroxidase from horse
radish.
After being purified and tested for antiperoxideactivity, preparations
were administeredorally to patients with acid or alkaline pain pattern.
While the former was definitelyintensified,there was no relief of the latter.
It seemedthat the effectsobtained through the administrationof isolated
pexoridaseswere the same as those obtainedwhen pears were eaten.
Antioxidants
The relationshipbetweenfixation of oxygen and chlorides has led to
the study of antioxidantscapableof acting in situationsof abnormal oxidation. It was hoped that thesesubstancas
also would be able to influence
tbe

410

xESEARcH rN pHysropATHoLocy

We have utilized several groups of known antioxidants, starting with

manganesecompounds,such as inorganicsalts,and later binding these to
lipids. It is too early yet to draw any definiteconclusionsfrom animal experiments,and we have not used the compoundsclinically. However, our
studiesup to now do not show any influencethat could be interpretedas
sufficientto warrant hope that these compoundscan control the fixation
of chlorideson fatty acids.
In the sameseriesof researches,
other antioxidants-some of them used
for the preservationof edible fats and others for the control of oxidation
in other substancessuch as rubber-were tried. We investigatedthe influencc of tocopherols,the natural antioxidantsfor vegetableoils. Alpha
tocopherolin dosesof 100 mg. was administercdseveraltimes a day to
patients having symptoms and signs correspondingto an intervention of
abnormal fatty acids. A decreasein the intensity of pain of an alkaline
pattern was observed.
Along the same lines, we investigatedthe influenceexerted by maleic
acid,usedto preventthe rancidityof ediblefats.In proportionof l/10,000
this acid conservesthesefats for months.Curiously enough,maleic and
citraconic acid havc shown an influenccupon the abnormal manifestation
of the type D.
For thesereasonswe utilized thesetwo acids-maleic and citraconicas anti-D agents.In one study, the acids were injectedintravenouslyin
proportion of from 0.1 to I mgr./100 cc. of saline.In others, the sodium
salts of the acid were used,while in still others the butyl esterswere prep a r e d a n d a d m i n i s t e r eidn t r a m u s c u l a r il n
y o i l y s o l u t i o n sF. o r t h e p r c s c n t
it is difficult to judge the effectsobtained.
All the abovc mentionedattemptswere made on the basisof a direct
action upon fatty acidsand other lipoidic constituents
with negativepolar
groups which intervcnein inducing offbalances.For the present,it seems
that no singlc agentcan resolvethe problemsthat result from the plural
intervention of various abnormal fatty acids at the different levels. The
use oI various asents acting selectivelyat the different levels involved
seems to be the only available path by which therapeutic intervention
againstthe multiplemanifestations
at differentlevelscan be accomplished.
going
Beforc
further. wc thoughtit usefulto have a synopticview of
t h i s s p c c i a lp a r t o f t h e p h a r m a c o l o g i c a cl t i v i t ya s o b t a i n e dt h r o u g h t h e
studyof the influenceupon pain and the systemiclevel analysesas seenin
humans.To this rve addedthe effectseenupon tumors in humans.Tables
XX and XXI which give this inforntationin a very condensedform, were
limitcd to the most imporrantagentstestedfor each group studied.The
tation.

p H A R M A C o D y N A M t cA c r r v r r y

(nrrnr rwo)

4lI

TrsLr XX
CLrlrc,rLRssulrs Wtrtr AcrNrs Tur
T H e O n n s , { t - A N c ET y p t

Group

Agent

Fatty Acicls Saturated

Polyunsaturated

.,A"

Systemic
Level

Pain

None
Slight

None
Fair

Mixtures from organs

" from cod liver
oil
Irradiated
Conjugated
a -OH
Polyhydroxy
Chloro-derivativcs

None
Slight
Fair

Slight
Fair

Oleic

SIight

None

Slight

Slight

Good

Good

Acr Upox

lnfluenccsExerted
Upon Tunror
None
Some,not consistent,
not persistent

Good I
Fair
Fair, not consistent,
not persrstent

.:

rr

{t

ta

None I None
I

None
Some, not consistent
not perslstent
None

I S l i g h t ,n o t c o n s l s t e n t ,
not persistent

Fair

Good l S l i g h t F a i r
Good I Cood I F a r r
None
None N o n e

Methyl thioglycolate
Tetrahydronaphthalene oersulfides
Srk"lr^
| Alkyldiselenide
Cctmpoundsl
I Perselenide

-l

Peracids

Perborate
Perchlorate

Slight

Slight

Fair

Fair

Slight
Fair

Fair
Fair

Fair

Slight

Fair

Cood

i=;-

1-'F

---.l
Hormones

i Testosterone

Slight

None

Mustards

I Sulfur mustard

Fh"

Fair

Fair
to good

Slight

Hydrines

Epichlorohydrin

Ciood, consistent,
persistent
Fair, not consistent,
not perslstent
Good, consistcnt
and pcrsistcnt

Good, consistent,

.f".ti:,"n,..
S o m e ,n o t c o n s i s t e n t ,
not persistent

Seldom, not consiste n t ,n o t p e r s i s t e n t

Fair, "
Fair, consisrcnt.
persistent

4t2 /

RESEARCH IN

PHYSIOPATHOLOGY

Trsl-e XXI
Cr-rNrcrl Results Wtrs AceNts THr.r Acr Upox
THe OnrsrLANcETYpe "D"

Group
Sterols

Agent
Cholesterol
Insapon.fraction
of organs
of eggs
of milk

Alcohols

Octanol

Slight

Polyconjugatcd
Crotonic
Ricinoleic
Salicylic
Hormones

Estrogens

Amines

Aminobutanol
Hexylamine
Heptylamine
Glucosamine

Halogen

Iodine
i Oxvcen
'|

Fair

Seldom,not consistent, not persistent

aa

aa

ar

ra

Good
Fair
Slight

None

None

Slight

Good

None

None

Slight

Slight
Fair

F^t;:

,,

Slight
Fair
Slight

.:h
a,

Good
Good

Fair

iI N"*

:.

.:

None
None
Slight, not consistent,
not persistent

,: .:

Slight,
r'
Fair,

aa

aa

ra

aa

Slight

Fair,
None

Good

None

Slight
None

None

Fair
Good

Good, consistent,persistent
None

ra

Slight

aa

Good, consistent,persistent
Fair, not consistent,
not persistent
Slight
None

t?I'

Niketamidc

Iron
Mercury
Bismuth

lnfluences Exerted
Upon Tumor

aa

Good
Fair

Inositol
U nsaturated
Oleic
Linoleic
Polyunsaturated

Metals

Fair

Aliphatic saturated
Butanol
Pentanol
Heptanol

Octanediol
Nonanol
Polyalcohol
Glycerol

Nicotinic
acid deriv.

Systemic
Level

None

"

aa

C H A P T ELR1
1 ' H E R A P E U T I CA P P R O A ( ] H

BIOLOGICALLY GUIDED THERAPY

ln
lLHe

sruDrESpREvroust-yDEscRrBEo
have identifiedmany factors involved in the pathogenesisof abnormal conditions and have shown that
there are similar factors which also govern the pharmacologicalactivity of
variousagents.Out of this peculiarrelationshipbetweenabnormalconditions and agentshas evolveda therapeuticmethodwhich is fundamentally
different from methodsin common use.
In conventionaltherapeuticefforts againstdisease-in chemotherapy
of cancer for example-methods as standardizedas possibleare sought.
The goal is to find agentscapableof influencingthe cancercells,if not in
all tumors,then at least in tumors of the sameorigin. Within the limitations imposedby the generaltoxicityof thc agent,the conductof treatment
is standardizedas far as possible.The criterion of value of a treatment is
its ability to overcomeindividual variations.
In our approach,in clear contradistinction,the effort is to influencethe
complexcondition as it is presentat differentlevelsof the organization.It
correspondsto treatment individualizedas far as possible.Therapeutic
effortsare guided in all their aspects--{hoiceof agents,dosesto be usedwith their multiple quantitativeand qualitative
by existingmanifestations
variations.The fact that many of thesevariationsoccur during treatment,
as a responseof the organisminducedby the medicationitself, increases
the individual characterof treatment.No predictioncan be made about
the whole course of treatmentto be used for an individual patient becausethe individualclualitativeand quantitativechangeswhich occur during the treatmentcannotbe foreseen.

414 /

n E S E ^ R C Hl N

pHysropArHoLocy

In this approach to therapy, the patternsof manifestationsconstitute

the criteria which determinethe choice of the group of agents,the specific
membersof the group, and the dosagesto be used. Throughout, clinical
and analytical manifestationsare the major factors that guide treatment.
The clear systematizationof the relationshipbetween analytical findings
and therapeuticindicationsis vital.
Thc fundamentaloffbalances,their patternsand deviations,thus guide
the therapeuticapproach. The nature of the fundamental offbalance,in
general,indicateswhich of the two basicgoups of agentsis to be used; the
patternsshow which substances
from thesegroups are desirable;while the
quantitative changesin these patterns, especiallyunder the influence of
the agents,determinethe dosesto be administered.
Any changein biological
manifestationsis followed by changein therapy. We call this biologically
guided therapy.
Through the years,althoughthis biologicallyguided treatment has remainedfundamentallyunchangedas an approachitself, applicationshave
evolved.Efforts have been made to find increasinglyprecisecriteria and
more efficientagents.
Each of the basic physiopathologicalconceptshas greatly helped to
developstandardsfor the guidanceof treatment.For instance,the concept
of organizedconditions has led to the concomitantuse of several agents
from the samcgroup insteadof just one.The ideafor this was derivedfrom
the fact that the concomitantmanifestations
at differentlevelsshow relative
independence;they must be influencedseparately.The concept of physiopathologicaldualism has indicatedthe need to use, ac@rding to the offbalance present, agents from one or the other of the two groups with
antagonisticproperties.The role of thc lipids in dualism has underscored
the importanceof thesesubstancesas active agentsable to induce fundamental changes.The dualism of other constituentshas been the basis for
their use. The mechanismof natural dcfensehas shown the advantageof
using two systemssimulating,in part, the kind of dualistic responsewhich
occurs naturally. As more has been learned about these basic concepts,
the therapeuticapproach,while it has remainedfundamentallythe same,
has evolved and becomeincreasinglyeffective.
The value of biologicallyguided chemotherapyappearsclear when we
comparethe effectsobtainedby using agenlswith and without the guidance
furnishedby this method. Severalpatientstreated at another center of researchwith a well-known agent, such as a nitrogen mustard gas derivative, have been studied.Analytical data showingthe patternspresentin
these patients were obtained but were not used to influence the conduct

THERAPEUTIC

APPROACH

4I5

of treatment.Favorable resultswere seen in a few of thesesa5s5-whgn.

by happy accident,the treatmentapplied was suitablcboth to the original
problem and to the changesoccurring during treatment, as revealedby
analyses.In the failures, there was no such fortunate coincidence.The
fundamentaloffbalancepresentcalled for an agent other than what was
used.Or changesoccurringduring treatmentindicatedthat useof the agent
should be discontinued,althoughthis was not done.
We have employedthe sameagentsin somepatientsbut with treatment
guided by individual manifestations.In some cases,the analysesindicated
the need not only for more prolongedtreatment,but even for the use of
dosageshigher than thoseusuallyaccepted.In others,on the contrary, they
indicatedthe needto reducedosageor evento stop the treatment,although
the doseswere, accordingto the usual posology,too small. It is interesting
to note that in some cases,the trcatment has been stopped and restartcd
severaltimes on the basisof analyses.The clinical results-immediate and
long-range-were definitelybetter than in patientsgiven unguidedtherapy.
In analyzingthe developmentof clinical applicationsof this form of
biologicallyguided chemotherapy,progresscan be seento have resultedas
knowledgehas increasedin three areas:criteria,agentsused,and method
of application.
Criteria
We have discussedthe different symptomsand analyticalchangeswith
dualistic character encounteredin pathologicalconditions. At various
stages,all were consideredas potentialcriteria for guided therapy. The
value attributed to them as indicationsfor treatment has changedas research has progressed.Some of thc analytical rcsts have been found to
reflectchangeslimited to a speciallevel,while othersof more generalvalue
have been found to furnish indicationsof broader offbalancesthemselves.
The importance of some criteria was establishedonly after many years.
Others, once consideredof major usefulness,have appearedto be less reliable as time went on. Body temperature,for example,in spite of the frank
dualistic aspectoften seen,has bcen consideredfor a long time to be influencedby too many factors to furnish any valuable information. With
the developmentof our research,however, temperaturehas become an
important criterionfor administrationof agentsbut limited toward manifestationsat the level of the organism.
Urinary specificgravity and pH; excrction of chlorides,sodium, calcium and sulfhydryl; total blood and serum potassium;and the count of
in circulatingblood-all havebeen usedindividuallywith the
eosinophiles

416 /

xEsEARcH tN pHyslopArHoLocy

hope that they would indicatethe typc of fundamentaloffbalancepresent,

and consequently,
which group of agentsshould be used. Recently,however,we havecome to usethesemeasurements
rather as criteriaof changes
at different levels.Urinary surfacetension has thus become important as
an indicationof the oftbalanceat the levelsabovethe cells while potassium
in blood indicatesthe offbalanceat the level of the cells. Years of investigation have shown conclusivelythat proper criteria representthe principal
condition for successin treatmentby guidedchemotherapy.
The Agents
The relative independence
of the variouslevelsinvolved in a condition
like cancer,which originatcsbelow the level of the nuclei and progressively
involveshigher levelsas it evolvesuntil it terminatesas a systemiccondition, necessitates
having severalagentschosenspecificallyfor their ability
to work at thesedifferent levels.
Over the years,we have utilized many agents.The level at which they
are most active, the nature of activity from the point of view of dualistic
intervention,and integrationof activity in the frame of the defensemechanism, representthe factorswhich determinethe specificuse of theseagents.
It must be emphasizedthat, in the study of pharmacodynamicactivity, the
value of information furnished by tests in laboratory animals is limited.
Becausechangesin patternsin animalscannot be followed, we have been
obligedto follow such changesin humans.
Method ol Application
The critical role played by each level of organizationin a complex
condition-and the independence
of theselevels-underscoresthe urgency
of applying therapeuticagentscapable of acting at specific levels where
they are needed.Such applicationshave beenmade possiblethrough special
affinitiesof various agentsfor different levcls.Their 5t1us[u1g-and especially their content of specificelementswhich belong to specific levelsaccount for their tendencyto act at specificlevels. However, it has often
beenjudged necessaryto insureinterventionof an agcnt at a given level by
applyingit directly at the level itself.The injectionof an agentinto a tumor
or into a regionof the body, through surgicalchemotherapy,representsone
type of procedureinsuring level activity. Another procedurehas been to
inject an agent prepared in vitro so that it is bound to specificbody constituents,such as cellsor nuclei. In the latter procedure,the specificdefense
mechanismis seen to take place at the level of the body constituentand
thus desiredlevel activitvis obtained.

i;riiNF]

T H E R A P E U T T CA P P R O A C H /

417

In the developmentof this research,we faced another problem which

is basicto all therapeuticattemptsbut especiallyimportantin cancer:evaluation of the resultsobtained.
Evaluation
Incontestably,favorableresultshavebeenobtainedin cancerwith many
agents.However, becauseof the variety of manifestations,
evaluationof
the changesobtained has been thc subject of much controversy. Each
clinical worker in cancerhas his own ideasabout the importanceof various
changesobtained and often utilizes as criterion changesless acceptedby
other workers.A systematization
of evaluatingstandardsappearednecessary and it seemspossibleto do so under the conceptof the diseaseas an
organizedcomplexcondition.
In the minds of most workers in the field of cancer today, any therapeutic procedureaimed at the control of cancermust prove itself in tumors
in animals.we have already mentionedthe reasonwhy control of human
and animal tumors representtwo different problems,Failure to recognize
that the resultsobtained in animal cancer do not necessarityapply at all
in human cancer has led to failure of the tremendousproject of screening
virtually all known substances
for their effectsupon grafted animal tumors.
The unguidedtreatmentof cancerin animals,as carriedon today,i.e.,
without any considerationof existing offbalances,seems to explain the
limited importance placed upon many changesobtained in these experiments. But even if evaluation is confined solely to the influence exerted
upon the cancerouscells in unguidedtreatment,the sameagentcan appear
active or inactive,dependingmainly on coincidentalcircumstances.Its influencemay changefrom favorableto unfavorableduring, and due to, the
treatment.Becauseof this, it is quite probablethat many useful agentsare
not being recognizedin the simple screeningmethod used today. Most of
the agentswhich we have found incontestablyvaluable in guided therapy
appearedto be entirely ineffectivewhen used in unguided treatmentson
small animals.As long as not enoughvaluablecriteria are availableto
permit guided therapy in experimentsin animals, the results would have
only a relativevalue: to furnish useful informationabout limited problems
such as toxicity, specialpharmacodynamicactivity, etc.
Under the conceptof canceras a complexcondition,all manifestations,
not just those directly related to the anatomicalpresenceof cancerous
tumors, must be regardedas important.
The tendencyof almost all cancerworkers is to limit evaluationexclusivelyto changesobservedin tumors. Without underestimatingin any

418

xEsEARcH rN pHysropATHoLocy

way the importance of these changes,we must emphasizethe importance

of other manifestations.For a subjectin the systemicterminal phase, any
changeobtained in the tumor will have little immediateimportance, whereas
a beneficial change in systemic metabolic anomalies will be of great immediatevalue. Similarly,for a subjectin the invasiveor painful phase,any
influence exerted upon the organic or systemic functions will be of less
immediate importance than a beneficial influence upon pain or upon the
cancerouslesion. With cancer recognizedas a complex condition, the decisionof many scientists,especiallyclinicians,to limit evaluationto changes
only in the tumors would appearto be unilateraland unrealistic.
We tried to systematizeevalution of results obtained in cancer treatmcnt by consideringall the manifestationspresent in the patient and by
assigningto each its relative value. In addition to any decreaseor disappearance of tumor masses,various analytical changes, improvement in
general well-being, gain in weigbt and control of pain represent accomplishments,the importanceof which dependson the severityof thesemanifestationsin the individual case.
With all these factors in mind, we have evaluatedthe various results
obtained through biologically guided treatment over a period o[ many
years,relating them to agentsand criteria which were changed,of course.
as researchprogressed.
The study of differentcomplexconditionshas emphasizedthe fact that
many of the manifestationspresent are common to difterent diseases.
Through the variety of pathologicalmanifestationspresent in these conditions, we had the opportunity to study many problems specificfor these
conditionsor of generalinterest.The therapeuticmethod consequentlyhas
been appliedin many noncancerous
conditions.Before presentingan analysis of the progressof this researchin the field of cancer,it may be of interest to review resultsof the sametherapeuticapproachin other conditions,
each with its specificproblems.

THERAPEUTIC APPLICATIONS IN CONDITIONS

OTHER THAN CANCER
From the beginningof this study, manilestations,clinical and analytical,
presentin a variety of abnormal conditions,have appearedto conform to
the basicphysiopathological
conceptspresentedabove.In thesevaried conditions, however,morc often than in advancedcancer,some analyseshave
shown no abnormal patternswhile others have shown them so constantly
as to indicate that certain anomalies,often limited to specfficlevels of or-

$:in
,ii ;

:J...i,

THERAPEUTIC

^PPROACH

,/

419

ganization,are of geat importance.In spite of individual variations,some

of the conditions studied have shown patterns and interrelationshipsof
patternsthat provide characteristicanalyticpictureswhich will be analyzed
in further publications.Furthermore,some of thesepatterns appear to be
so strongly related to a given condition that our attention was directed to
the idea of a fundamentalrelationshipbetweenthe processesrelated to the
patterns and the pathogenesisof the condition itself. The following examples illustrate this.
An impressiverelationshipwas found betweenthe presenceof peroxides
in urine and schizophrenia.We mentionedpreviously the reaction which
rve devisedfor detectionof thesesubstancesin urine. This reaction would
indicatethe existencein the body of abnormalprocessesin oxygenmetabolism, leadingto the appearanceof peroxidesin the urine. we saw that these
processesultimately could be related to an abnormal interventionof fatty
acids,correspondingto the oxygenphaseof offbalanceD.
Daily urinary analytical patterns were studied in a group of. 27 advancedschizophrenicsover a period of three years. Over 27,000 urinary
sampleswereexaminedand more than 135,000testsperformed.Oxidizing
were found in 87% of thcsesamplcs.This appearsto be highly
substances
significantwhen compared with only 2Vo positivevalues in subjectsconsideredclinically normal, and 4Vo in cancerouscasessubmittedto various
treatments.
Not a singlenegativeanalysiswas scenin somcof theseschizophrenics during the three years of daily testing. This suggestedthat the
by the appearanccof peroxides,might
metabolicabnormality,characterized
play an important pathogenicrole in this disease.(221)
Another exampleof a pattern revealingcharacteristicpathogenicprocesseswas seenin geriatric cases.Old people often have a manifest abnormality in urinary S. T. Figure 261, page648 showsthc valuesencountered
in old peoplecomparedto thosein subjectsof mixed ages.The high surface
tensionin the aged group is related to predominanceof sterolsat the systemic level. Such predominanceis also found at the organic level of the
A clinical test,whealresorptiontime, indicatedabnormal
skin, for i.nstance.
valuesfor almost all the aged subjectsstudied,as shown in Figure 68.
These data, integratedinto the generalconceptof complex conditions,led
us to the pathogenicconceptof old age presentedabove.
The urinary chloride retention index in subjectsin a state of shock
consistentlyshows such exceptionallyhigh values that our attention was
directed to the study of the role of abnormal metabolismof chlorides in
the pathogenesisof the condition itself. This view has been confirmed by
further research,as mentioned previously.

420

nESEARcHtN pHyslopATHoLocy

The study of the differentconditionsin terms of fundamentalpathogenicconceptshas had other consequences.

It has established
clearerrelationshipsbetweentheseconditionsand cancerthan previouslyrecognized.
In fact, many of the data first obtainedin the studiesof theseother conditions have been applied specificallyto cancer.
One example is shock. Occurring in the terminal stage and usually
leadingto death, shock often has been consideredto be one of the "complications" of advancedcancer. Under the concept of organizedcomplex
condition,however,it could be seenthat, in cancer,shock is related to oftbalance type D of the terminal systemicphase. The study of shock has
greatlycontributedto the knowledgeof this offbalancein cancer.
The therapeuticapproach,in which the choice of agentsand the doses
used are determinedby the different patterns present,has been useful in
many noncancerousconditionsin which the anomaly apparentlyis limited
to one level of organizationor even to a specialgroup of entities. As examples here, we will discussconditionsin which manifestationsare producedby an acid-basetissularabnormality.

ABNORMAL LOCAL ACID-BASE

MANIFESTATIONS
Pain, itching, vertigo and dyspneaare symptomsof many conditions
often far apart etiologically.we have seenthat, accordingto our research,
thesesymptomsare related to an acid-baseabnormalityat the tissue level.
The ideai treatmentwould be to remove the causeof the symptom, to act
upon the etiologicalfactor. This ideal treatmentwould be entirely difterent
if the symptom stemsfrom a systemictoxic condition, a cancerouslesion,
a local inflammatoryprocessor a local a-llergy,for example.But the great
varietyof causesthat can inducesuch symptomsas vertigo,pain, itching
and dyspnea,often makes this ideal therapy almost impossible,particularly in everydaymedicalpractice.
Another approachto controlling thesesymptomswould be one which
is not especiallyconcernedwith the etiologicalfactors involved but with
influencingthe specificpathogenicchangesunderlyingeach of thesesymptoms. But as such specificchangeshave not yet been defined,symptomatic
treatmenthas remainedas the only practicalrecoursein theseinstances.
Recognitionin the physiopathological
changespresentin pain, itching,
vertigo and dyspneaof an acid-basepattern, has permitted another therapeutic approachon a different, more precisebasis.These symptoms,with
have been treated accordingto the two possibilitheir dual pathogenesis,
ties: acid or alkalinepattern.Identification
of the specificpattern,as noted

THERAPEUTTc APPRoACH

421

previously,is simplc.Furthermore,pain resultsfrom an abnormalitypresent

at one level of organization,the tissues.With two groups of antagonistic
agents,both capableof acting at the tissularlevel, the therapeuticproblem
is reduced to the choice between these agents according to the pattern
present.The therapeuticproblem-previously so complex becauseof the
need to consider so many possibleetiologicalfactors so often unrecognizable-is simplifiedby this approachto a choice betweentwo groups of
agents,determinedby one easily idcntifiablefactor.
The means used to recognizethe acid-basepattern vary with each
symptom.We presenthere a resumeof the researchmade in this direction.
Pain
We have seenthat pain can bc sensorialor symptomatic,and that the
latter has a dual pathogenesis
with an acid or a.lkalinepattern.Reducedto
the problemof an acid or alkalineoffbalanceoccurringat the tissularlevel,
the treatmentof pain is grcatlysimplified.
The first problem was to determinethe pattern present.The relationship between variations in the curve of pain intensity and concomitant
changesin the urinary pH, previouslydiscussed,
has been used for such
determinations.The simplified method of comparing two urine samples,
one correspondingto a period of pain and the other to one of calm, has
proven very useful.
Among other analyses,only changesin serum potassiumcontent have
appearedhelpful in the recognitionof the pain pattern. This can be explained by the fact that most of the other routine analysesdo not inform
us specificallyabout changesat the tissuelevel where pain occurs.A high
serum potassiumlevel, increasingduring pain exacerbation,indicatesan
alkaline pattern; a low level, decreasingduring pain exacerbation,indicates an acid pattern.
Another, sometimessimpler, techniquefor recognizingthe pattern is
the responseof pain to administrationof acidifying or alkalizing agents,
as seen above. Also, some substances,while being used as therapeutic
agents,indicatethe pattern through the responsesthey induce.Butanol and
5sdirrmthiosulfatein adequatedosages( I cc. of.a 6.5% solutionof butanol
or I cc. of.a 4Vo solution of sodium thiosulfateinjccted intramuscularly)
are examples.The intensity of an acid pattern of pain is increasedby
butanol, and decreasedby thiosulfate.The inverseoccurs in pain of an
alkaline pattern.
With the pattern of the pain recognizedthrough one of these means,
agentsare chosen from the two groups,anti-A for acid pattern and anti-D
u however.some
ts have been found to be
for alkaline. In each

422

xESEARcH rN pHysropATHoLocy

more effectivethan othersbecauseof their gleateractivity at the tissuelevel.

The foUowing agents have been found to be most effective against acid
pain, in the order presented:lipoaldehydes
and especiallypropionic aldehyde, sodium thiosulfatc,sulfurizedtetrahydronaphthalene,
the acid lipid
fractionof variousorgans,polyunsaturated
fatty acids,epichlorohydrin,and
seleniumin the form of perselenide.
For an alkalinepattern,agentswith a
positivepolar group, such as butanol,nikethamide,insaponifiablefractions,
glyceroland heptanolare effectivein that order.
Once the agent to be used is chosen,the dose which can vary greatly
from caseto case,is easilydeterminedfrom the clinical response.In practice, the patient is given a small dose.If the intensity of the symptom decreasesbut is not completelycontrolled, the same dose is usually given
three hours later. If the symptom intensity remainsthe same, the dose is
continuouslyincreasedmore or lessrapidly, in accordancewith the severity
of the symptoms,the medicationbeing repeatedthis time at intervalsvarying from a half hour to everythreehours,
On the other hand, if the symptom disappearsafter the first administration of the agcnt, medication is not repeated until the symptom
reappears.If the symptom is controlledfor lessthan six hours by the dose
used, the same dose is given when the pain reappears.If more than six
hours of relief follow a dose,the next doseis usually reducedin proportion
to the pcriod of time the effectlasted.If 24-hour relief occurred,the dose
is reducedby half; if relief was of two days' duration,one-quarterof the
last dose is given. However,if, with three succcssive
dosesof medication,
the symptom increascsin intensityeach time, always during the first half
hour after the medicationis administered,it is discontinuedand the entire
group of agentsto be used is reconsidered.A new test for the pattern is
performed.If the patternis the sameas before,other agentsfrom the same
group are tried. However, if the pattern now is opposite to the original
has to be changed.
pattern,the group of substances
This techniquehas produced,with few exceptions,excellentpain relief.
This approach to pain has been the object of severalcontrolled studies.
S . A . B a r r a g a nM a r t i n c z ( 1 8 3 ) a n d E . S t o o p e n ,( 1 8 4 ) i n t h e j o u r n a l
"Pasteur," confirmed our conclusions.From Stoope
n's publication we
q u o t e t w o c h a r a c t c r i s t iocb s c r v a t i o nisn N o t e / .
An intensivcstudy of head and neck pain was made by B. Welt and
publishedin the American Medical Assn. Journal of Laryngology(185).
with the followingconclusions:

THER^PEUTIC

APPROACH

423

" Summary and Conclusions

A seriesof 120 patientshavingthe symptomof pain in the regionof the
head and neck has been studied.
The symptomof pain has been analyzedaccordingto Revici's concept
concerningthe alkalineor acid patternof the painful symptom.
Eleven caseswere eliminatedbecauseno patternwas identified;109
casesshowedan acid or alkalinepattern.The resultsshoweda satisfactory
resultin 84% of the vascularheadaches,
100% in migraineheadaches,
and
75Vo in the neuralgicgroup.Theseresultsindicatea correlationof Revici's
concePtsand the resultsachieved.
The simplicityof this methodis indicated,
Additional data about Revici's views are given. The relationshipto
homeostasis
and the biologyof thc cell is indicated.
The problem confrontingthe clinician in treatingcasesof this type,
on accountof the many factorsinvolved,is simplifiedaccordingto Revici's
concept of dualism.
Two additionalactiveproductsare prescnted,onc acid and the other
alcoholic.The data here presentedconcernedthe incidenceof the control
of the acutesymptomswhile under observation.Further experiencewith
this methodwill be necessary.
As the methodstands,it is a practicalmethod
of therapyfor the control of pain. Recurrences
were seenand controlled."
Our methodof controlling pain was the subjectof a panel discussionat
the American Academy of Ophthalmologyand Otolaryngology,New York,
N. Y., September
2l and 22, 1954,and of an articlepublishedby B. Welt
M.
Welt
in
and
Modern Problemsof Ophthalmology.(186) (Note 2)

Trauma
A particularly interestingapplicationof the method describedabove is in
the treatment of pain related to traumatic lesions,The study of wounds
from the standpointof the offbalancepresenthas revcaleda definite pattern in their evolution. In all wounds, there is an initial period during
which the offbalanceis alwaystypc D, correspondingto a predominanceof
fatty acids. This can be recognizedindirectly by measuringthe local pH
which shows alkalosis,the chloride content which shows a manifest increase,and by cytologicaland histologicalstudieswhich indicatea rapid
cellular agtng process, with necrosis and sloughing. In uncomplicated
wounds,in humans, the painful period usually lasts a few days. The pain
in this initial period is of an alkaline pattern.
The constancyof the alkalinepatternin pain in traumaticlesionselimi-

:let

424

RESEARcH tN

pHysropATHoLo6y

natesneed for any tests, thus simplifying therapeuticchoice by reducing

agentsto be usedto one group--those with positivecharacter.
As alreadynoted,someof the agentsiue more effectivethan othersin
influencingalkalinepain. Butanol and heptanolare particularlyuseful in
traumaticpain, followed in order of activity by polyunsaturated
alcohols,
nikethamideand glycerol.
One importantapplicationof thesefindingsis in routine treatmentof
postoperative
pain.For surgicalwounds,whichrepresent
typicaltraumatic
lesions,butanol appearsto be especiallybenehcial.Its use has been the
subjectof extensive
studiesby B,welt (187) in surgeryin otorhinotaryn-

W'TH
BUTYL
ALCAHOL

WITH
PHYSrcLOGICAL
SAL'TVE

4O cosas

25 cosas

O' little or no pain

7 , poitr
Flc. 144. Rcsulls obtaincd with bulanol in postoperativepain-compared with rub"
jects recciving saline as placebo-(From B. welt-AMA
Archives of Ololaryngology.

t 2 . 5 4 Ar.9 5 0 . )

T H E R A P E U T I C^ P P R O A C H

425

PA'N 12 HOURS AFTER

THE LASI q-BUTYL ALCOHOL,NJECT,ON

WITH
BUTYL ALCOHOL
362 coses

fvo

TREATMENT
585 coses

9t%

+
O = minimol or no po,n

* " moderoteor seyerepoin

F r c . 1 4 5 . R e s u l t s I 2 h o u r s a f t e r t h e b u t a n o l i n j e c t i o n s .( F r o m B . W e l t , A M A A r cbivesof Otolaryngology, 50, 590, 1950.)

426

xESEARcH rN pHysropATHoLocy

gology,(Note 2) (Fig. laa)by M. Welt in ophthalmology(188), by A.

Ravichin urology ( 189) (Note3), andby S. Sheerin plasticsurgery.(190)
with more than 10,000 casestreated,butanol seemsto be completely
safe as well as highly effective.Not a single case of undesirablereaction
due to the medicationhas been noted. The dose necessaryto obtain pain
relief varies with the amount of traumatizedtissue,With an adequatedose
of butanol,the patienthas a minimum of pain, is conscious,and without
impairment of intestinaland bladder function. Butanol also has a marked
preventiveaction againsthemorrhage,which will be discussedbelow.
successwith this medicationdependsupon adequatedosage.The addition of butanol to the salinesolutionscustomarilyused in postoperative
care has done much to simplify the problem of administeringit in adequate
doseswithout resortingto supplementaryinjections.Doses as high as 100
cc. or even200 cc. of 6.5% solutionof butanolhavebeen administeredin
the fust days following surgery,with good resultsand no inconvenience
to the patient.
Administeredby injection or orally in only very small doses,heptanol
is helpful. A mixture of heptanol, butanol and polyunsaturatedalcohols
constitutesa valuable preparationfor controlling postoperativepain.
Butanol, heptanol and glyssTophosphoricacid added to glucose and saline
hasprovedvery helpfulto controlnot only pain but alsothe other disagreeable post-operativemanifestations.(Note 4)
In other forms of trauma, such as accidentalwounds or fractures,the
sameagentshavebeencquallysuccessful
if in sufficientdoses.
With their use,the healingperiod seemedto be shortened,as shown in
experimentsin rats and rabbits. In animalswith standardizedwounds, the
healingof connectivetissuein particularis accelerated.
In woundstreated
with preparationsof cholesterol,sterolsor insaponifiablefractions of organs, the healed epithelium shows abnormally high proliferation.

Burns
The pain of burns has been successfullytreated with higher alcohols such
as butanol, heptanolor polyunsaturatedalcohols.However, here the prob.
lem is not so simple since pain is only one, and not the most important,
of the manifestations.Systemicfunctions are often so greatly impaired as
to representthe immediatecauseof death.The higher alcoholsare effective
in controlling pain and local troubles,while other agentsare neededwhen
the condition also involves the systemic level. Glycerol and especially
sterolswhich are presentin the insaponifiablefractions of organs such as

.,i,i:.
4:li,r,i

T H E R A P E U T T CA P P R O A C H /

427

placenta,are very helpful in thesecases.Anuria of severaldays' duration

has been influencedin severalcases,with abundant diuresis occurring only
a few hours after the administrationof 2-4 cc. of. a l0% solution of the
insaponifiablefraction of placenta, an effect not obtained with butanol
alone,even in large doses.The effectwas still more impressivewhen large
dosesof butanol, such as l0 cc. of the 6.5% solution every hour, were
administeredfor a few hours before injection of the insaponifiablefraction.
The addition of sodium lactateto butanol in sufficientamount has been
seen to favorably influence systemicproblems related to burns. Experimentally, we have shown the value of this preparation in extending the
survival time of mice.
Under ether anesthesia,adult mice were scaldedin water at 90oC for
three secondsup to the xyphoid appendix.The survival period for untreated animals was short. Of the various preparationsused, the butanol-sodium lactate mixture seemedto be the most active in prolonging
survival.(SeeNote I3,Chapter I0 and f iS. /,38)
Ultraviolet ray burns, such as occur in sunburn,have been treatedwith
butanol in severalhundred cases.We startedwith 5 cc. of the 6.5Vo solution orally, and repeatedthe dose every 10-15 minutes until the painful
sensationdisappeared.When the burning sensationreappeared,treatment was resumed.It could be seenthat butanol not only controlled pain
rapidly and completelybut also that the skin lesionshealed in a shorter
time than in untreatedsubjects.
X-ray and radium burns in animalsand humanshave respondedto the
srme substancesas mentioned abovc. Standardizedulcerated radiation
Iesions were inflicted in rats by inserting,betweenthe lips of a skin incision, a needlewith radium in platinum or monel metal. A needlewith
25 mg. radium in monel metal was kept in place for two hours, one with
l0 mg. in platinum for 96 hours. Ulcerationsresultedand took four weeks
or more to heal. In animals treated with sterol preparations,such as insaponifiablefractions of placenta,wounds healed in much shorter time.
Similar effectscould be obtained with butanol, although less consistently.
Good resultswere obtainedwith the sametreatmentin radiation burns
in humans. In relatively new lesions,pain was relievedafter a day or two
and rapid healingusuallyfollowed.Even in old burns which had not healed
for many years, pain was controlled in less than a week and the lesions
started to heal. In some of these,scarsdevelopedin lessthan a month.

428

nesEARcH rN PHYsroPATHoLocy

Vertigo
Another interestingapplicationof the same therapeuticmethod has been
in otological conditions.B, Welt has made a stimulatingcontribution
to the study of ear conditionsin terms of the pathogenicmechanisminvolved and an even more valuablecontributionto therapy.
Vertigowas one of the conditionsstudied.In a few cases,by comparing
concomitantvariationsin the intensityof the symptomand changesin the
urinary pH, we had been able to show that acid and alkaline pathogenic
patternsexist,similar to thoseseenin pain.
Going on from there, B. Welt, in an extensivestudy, has shown thc
presenceof this dualism and has successfullyapplied the therapeutic
methoddiscussedabove,to a significantnumber of casesof vertigo.The
major advantageof this method over other treatmentsfor this condition
lies in its extremesimplicity.
For vertigo,which has so many difTerentcauses,an etiologicalapproach
althoughideal,has seemedimpossibleto all workersin the field especially
as a routine procedurein medical practice.An initial simplificationof the
approachcould be made by relating vertigo to acid and alkaline patterns.
thus reducingtreatmentchoiceto two groupsof agents.Welt, in a further
step, simplified even this procedure,making the method highly applicable
in routine medicalpractice.He administercdto patientsone or the other
of two agentschosenfrom each group, such as butanol or sodium thiosulfate,being guided in his choice by clinical aspectssuch as changesin
the symptomwith the time of day or the intakeof food. A favorablechange
in the symptomwas consideredto be a confirmationof this tentativediagnosisof the pattern.The treatmentwas then continuedwith the samegroup
of agents.An increasein vertigo led to the use of the opposite group of
agents.The clinical resultsobtainedby Welt with this simple method arc
impressive.(190) He arrivesat the following conclusions:
" Summaryand Conclusions
A seriesof 44 patientshaving the symptom of dizzinesshas been
studied.
The symptom of dizzinesshas been analyzed according to Revici's
conceptconcerningthe alkaline or acid patternof the symptoms.
In 80% of the analyzedcases,the responseshave shown the existence
of an alkalineor an acid pattern;lTVo showedan inconsistent
relationshipl
8% showedno result.

T H E R A P E U T T CA P P R o A c H

429

A thcrapeutic approach has been devised which correspondsto the

presentedresults.
The control of thc symptom of dizzinessby this thcrapeuticapproach
has been obtainedin 80Vo of the cascs.
As comparedwith other methodsof treatment,which are prolonged
and involved, Revici's approach is simple in application and effcctive in
its results.
This therapeuticapproachbasedon thc dualisticpattern of the physiopathologyof the pathologicaltissuesis especiallyapplicableto the symptom of vertigo, which becauseof the multiplicity of the ctiologic and
pathogenicfactorsmake a direct mcdicalattack almostimpossible.
The evidencepresentedin this communicationrelatesto the control of
the acute attacksof vertigo. Recurrenceswcre observedin this seriesand
successfullycontrolled.
In the casessubmittedin this communicationand in subsequent
cases
subjectedto this form of therapy,further observationby this method will
indicatewhetheror not the lesionis permanentlycontrolled."
Welt completcdhis vertigo study by using many othcr membersof the
same groups of agentsas thcy were studiedby us for othcr conditions.
The following conclusionsappear in Welt's secondpublicationon vertigo.

(lel)
" Sum mary and C oncl usi ons
A seriesof 106 caseswith the symptomof vertigohas beenpresentedin
this and a previouspaperand analyzedaccordingto Revici'sconceptof a
metabolic imbalancewhich exists in pathologicalfoci and manifestsitself
as a local alkalosisor acidosis.
Sevencaseswere elinrinatedfor lack of data. This left a total of 99
case studies.This figure represents
the sum of 42 casestermed the 1953
seriesand the presentseriesof 57 cases.The two serieswere combined
and analyzedaccordingto sex, alkalineor acid characterof the symptom,
and clinicaldiagnosis.
The analysisshowed an averagegood result of. 80% for both series.
This percentageindicates a consistentcorrelation of the use of Revici's
method and substanceswith the results achieved.Additional substances
from various sourceswere utilized in the present study. It is here again
emphasizedthat I have submittedthis article for the practical reasonthat
the method describedis simple and has bcen successfulin its application.
In addition, it gives a new systematizationto clinical data in the field of
otolaryngology.Finally, this communicationpoints to a vast group of sub-

430

RESEARcHtN pHyslopATHoLocy

stancesfrom widely differing sourcesand of different structureswhich have

similar biochemicalactivities.The substances
presentedhere are but a few,
but they open a pathway for the developmentof other substanceshaving
similar biochemicalactivities.It is to be hoped that thesetoo, may have a
practical application in the field of otolaryngology.
views on cell permeability and their relationship to fatty acids and
sterolsare added.
The role of foci of infectionand psychologicalaspectsin their relationship to the symptom of vertigo is amplified.
The simplicity of this method of therapy is emphasized,and its integration with a new method of systematizationof clinical knowledge is
indicated."
Hearing Impairment
Deafnessremains the most important problem in otology despite good
resultsrecently obtainedwith surgicalproceduressuch as fenestrationand
stapes mobilization. Since deafnessresults from long evolving processes,
a logical approach to the problem would be not to wait for the advanced
stagewhere surgerymust be usedbut to try to control the processesbefore
they reachthis ultimate statewhich usuallycorrespondsto sclerosis.Studies
of the morphologicalchangesrelatedto progressivedeafnesshave been too
limited to present a basis for a biologicalor biochemicaltherapeuticapproach that would have a chance of success.The nature of the lesions,
their minusculedimensions,and their chronic evolution, offer no opportunity for direct information. Consequently,we tried to adapt to this situation the knowledge acquired through researchin the field of abnormal
processesin general.
Just as for pain, it was observedthat some patients with hearing impairment are aware of differencesin their acousticacuity accordingto the
time of day, i.e. some hear better in the morning, some in the evening.
There seemedalso to be a correlationwith the intake of food. Based on
these observations,casesof various degreesof surdity were investigated
for relationshipto acid-basechanges.Some subjectsshowed audiometric
variationsunder the influenceof acidifyingor alkalizingagents,while others
did not respond.when two antagonisticagentswere used in the same subject and hearingacuity was analyzed,the changesoccurredin two opposite
directions.we could interpret this only by analogyto changesin pain and
other acid-basesymptoms.Accordingly, we postulatedthat in some cases
no activity is going on in the lesionssince no changesare induced in thc

T H E R A P E U T T CA P P R o A C H

431

audiogram by administration of acidifying or alkalizing agents; in others,

there is activity which is acid or alkalinein pattern.
Thus, the existenceof a responsewas consideredto indicate a still
active lesion.Time is not the determiningfactor. Even in relatively recent
lesions,biologicalactivity may have ceasedwhile in other cases,even with
very advancedhearingimpairment,lesionscan still be evolving,with processesnot yet advancedto the scleroticscar phase.This aspect has been
extensivelystudied becauseof the therapeuticimplications,since only an
evolving statecould possiblybe influencedby this method of therapy.
The influenceof agentsused in other conditionswith acid-basepathogenesiswas studied in casesof impaired hearing.The first attempt made
years ago, with fatty acids and sterol preparations,was not successful.
Much better resultshave been obtainedwith more recentlydevelopedsynthetic agents.In the last five years,Welt has devotedmuch time and effort
to this research.He has made thousandsof completeaudiogramsand has
used a seriesof active agcntswith placebosfor control. (l9z) welt has
studied approximately460 casesto date.(Note 5)rn the simplified form,
an agent from one or the other group was used for a short time and improvementin the audiogriimwas taken to indicatethat a suspectedpattern,
acid or alkaline,was actually present.
Clinical resultshave largely confumcd theoreticalviews. Regardlessof
degreeof impairment, improvementrangedfrom nil or almost nil in cases
with no more signsof activity,to goodand excellentin thosewith still active
lesions. In young patients in whom the proportion of active lesions is
high, resultswere particularly impressive.Hearing impairmentsof 60-70
decibelsor more have been overcome,sometimesin only a few weeks of
treatment.Theseresultshavebeenobservedto persistin many cases,some
thus far for more than three years.However, a tendencyfor impairments
to recur, usually following rhino-pharyngealinfections, has been noted.
Generally, renewedtreatmenthas been able to restorehearing to previous
valuesin a shorttime. (Figs. 146 to i,50)
It appearssuperfluousto emphasizehere the importanceof this contribution to the medical and social problem of deafness.It representsthe
first approach to treat successfullythe pathologicalprocessesrelated to
impaired hearingbeforc they lead to advancedlesionswhich can no longer
be influencedexcept by surgery.Every caseof incipient or even advanced
hearing loss shouldbe investigatedby the simpletechniquedevisedby Welt
to see if correction by this method is still possible. Surgical intervention
should be reserved principally for patients with lesions which correspond
to sequelsand who no longerrespondto biochemicaltreatment.But even

432

RESE^RCH IN

PHYSIOPATHOLOGY

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THER^PEUTIC ^PPROACH

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434

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THERAPT,UTTA
c PPRoACH

435

sugical casescan benefit from biochemicaltreatmentwhich could prevent

further sclerosiswhich often follows surgcry. Application of this method
in time to prevent deafnesswould be even more important than restoring
hearingin far advancedcases.

Itching
Acid and alkalinepatternsof itching havc beenrecognized,and treatment
basedupon dual pathogenesis
has been applied.The agentto be used is
determinedby the pattern found through analyses.Dosage,as for pain, is
established
accordingto the subjectiveresponse.
Following thesemethods,
many casesof intractableitching have becn completelycontrolled. of thc
group of agentswith positivecharactcr,butanol and heptanolhave produced the best results;in the oppositcgroup, cpichlorohydrinand sodium
thiosulfatehave been most effectivc.

Gallbladder and Renal Colic

Attempts were made to influencercnal and gallbladdercolic by using the
same method.Butanol has shown little cffect upon renal colic at least in
the doseswe tried. But the resultsin gallbladdercolic were impressive.A
6.5% solutionof butanolin salineinjcctedintramuscularly,
or even l5-30
cc. (one or two tablespoonsful)of 6.5% solutionof butanol in water administeredorally when the patient was not vomiting, controlledgallbladder
colic rapidly. only twice in more than 20 casesin which butanol was used
was it necessaryto give largcr doscs,such as 50 cc. of the same solution,
to obtain the desiredeffect.

A*hritis
The analysisof the pattern of pain has shown that often in rheumatoid
arthritisthe patternis alkaline,while in osteoarthritis,
it is acid. Comparison of variationsin pain intensitywith changesin urinary surfacetension
helps to determine the pattern. However, with urinary surface tension
values usedas a guide both for agentand dosage,favorableclinical results
have been obtained in casesresistantto other therapy. Arthritis, painful
for many months, was controlled in many instanceswith severaldosesof
heptanol or of sodium thiosulfatewith epichlorohydrinand more recntly
with propionic aldehydeor sulfurizedtetralin. The simplicity of treatment,
total lack of undesirableside effects,and the long period of improvement

436

nESEARcHrN pHysropATHoLocy

after even brief trcatmcnt in -rrn',ccascs.indicates that this method is

worthy of further invcstigation.Scn: illustrationsof the results are containedin the followingobservaticrs:
Case $l
Mr. M. R., 63 years old rvirh r long history of arthritic pains. Four
years ago he was obliged to stri' in brd for three months for severepains
in the right sacroiliacregion. T*,r )'carsago he was again bedriddenfor a
month and a half becauseof ihc same type of pain. A year ago, pain
startedagain in the samesacroilirc rcgion, to be followed by a very severe
sciaticawhich kept him in bed for two months.RepeatedX-ray examinationsshowedadvancedarthriticlesionsin the lumbar spineand aroundthe
sacroiliacarticulation.About tcn monthsago, he sufteredanothersimilar
attack, with severepain, in thc snme rcgion. Urine analysismade at this
time showedan offbalanceof rhc t1,peD with low surfacetension,low pH
and high specificgravity.The blood serumpotassiumwas high (5.3 mEq.).
The patient was treated with agcntsof the group D-l
mg. of heptanol
and 30 mg. butanol threetimesa day. The patientremainedfree of symptoms for four months, when similar scverepains were again present.This
time analysisshowed an offbalanccof type A, with high urinary surface
tension,high pH, Iow specificgravity and low blood serum potassium(4.0
mEq.). For this rcason,diflcrent medicationof the group A-5 mg. epichlorohydrinand 50 mg. sodiumthiosulfate-wereadministered.The responsewas as rapid as previouslyand pain disappearedwithin a few hours.
Sincethen the paticnt has becn taking 5 mg. epichlorohydrinand 50 mg.
sodium thiosulfatetwice a day wheneverthe slightestpain appears.The
clinical resultsare excellent,thc patient being now able to move about
entirelyfree of pain, as he has not been able to do for many years.
Case $2
Mrs. F. R., 60 yearsold, was bedriddenfor one and a half yearsabout
six years ago, with lumbar and sacroiliacarthritic pains. X-ray treatments,
short wave, ionizationwith histamineand ultrasonictreatmentsappeared
entirely ineffcctive.The administrationof urolytic agentshad helped at this
time. The pains recurredrepeatedlyin the last three years keeping the patient in bed for from one week to three monthsat a time. For the past eight
months,the patienthas beenundertreatmentusingmedicationof the group
A or D. accordingto the urine analysis.With this treatmentthe pain has
beencompletelyand rapidlycontrolled,neverlastingmore than a few hours.

THER^PEUTIC APPRO^CH

437

Case $3
M r . N . K . , 5 5 y e a r so l d , s u f f e r e df o r m a n y y e a r sw i t h g e n e r a l i z eadn d
very painful rheumatoidarthritis, Prior to coming under our care, hc
had beenbedriddenfor six months,entirelyimmobilizedwith severepain
in arms,back and legs.Thc patientwas entirelyincapacitated,
unableeven
to feed himself.Treatmentswith differentcortisoneprcparations,ACTH,
g o l d ,e t c , ,p r o v i d e dp r a c t i c a l l n
y o r e l i e fo f t h e s e v e r cp a i n s .H e c a m eu n d e r
o u r c a r e ,e n t i r e l yi m m o b i l i z e da n d i n s c v c r ep a i n . A c c o r d i n gt o t h e a n a l ysis which showcda low urinaryspccificgravity.and pH, high surfacetcnsion and a low blood scrum pot:rssium,
the paticntreccivedmedicationsof
t h e g r o u pA , s o d i u mt h i o s u l f a t ee,p i c h l o r o h y d r iann d h e p t y l d i s e l e n i dUen. der this medicationthc patientmadc a most dranraticrecovery.within a
f e w d a y s ,t h e p a t i e n tw a s o u t o f b c d w i t h o u tp a i n . w a l k i n gn o r m a l l ya n d
w i t h f u l l f u n c t i o n acl a p a c i t yo f h i s a r m sa n d l e g s .W h i l c s t i l l u n d e rt r e a t m e n t
t h e p a t i e n tw e n t b a c k t o e n t i r e l yn o r m a la c t i v i t y .H e g a i n e d3 5 l b s . i n t h e
Iastmonths.

ConnectiveTissueConditions
We have discussedpreviouslythe hypothesisthat corticoidsactively intervenein the synthesisof an cntire group of substances
in the organism.
The energcticformationbetwcenCrr and C3, o[ thesecorticoidswould act
as a mold or templateto attractand keep variousradicalsin specialreciprocal position,thus favoring,as a secondstep,the synthesisof new substances.Thesesubstances
would vary accordingto the cnergeticformation
of the corticoid.In one group,whichcould be callcdthe "gluco" group,wc
identifiedglucose,glucuronicacid, glucosaminc,gaJactosamine,
glucosaminic acid,galactosaminic
acid,and someotheruronic acids.Even ascorbic
acid could be relatedto such a synthesis
mechanism.
we investigatedsome of thesesubstances,
particularlyin relation to
t h e t h e r a p e u t ipcr o p er t i e so f c o r t i c o i d sI.n 1 9 5 1 ,w e c h o s eg l u c o s a m i naes
a substance
closelycorresponding
to the templateof cortisone.Glucosamine
treatmentwas usedin patientswith pemphigus.rheumatoidarthritis.psoriasis,ocularconditionand allergy.The followingcasesillustratethe results
obtained.
Mr. H. S., 46 years old, had generalizedpsoriasis.In the last
twelveyea$, the lesionshad progressed
continuouslyto cover more
thanT5Vo of his skin. Local treatmenthad had little effect.Cortisone
injections cleared the skin in two weeks but not completely. With

438

n E S E A R C Hr N p H y s t o p A T H o L o c y

cessationof treatment,the condition returned.The patient remained

without treatmentfor a few monthsbefore beginningour treatment:
5 cc. injectionsof glucosaminel0o/o solution administereddaily.
Changeswerc seenin 3 days,with the lesionsbecomingpaler and less
scabby,With two injectionsdaily, the skin was almostentirely normal
in about I0 days, only a few lesionsremaining.Continuationof the
treatmentfor anotherweek did not make thesedisappear.Upon cessation of treatment,the lesionsregressedto their previous dimensions.
In a secondcourseof treatment,with two daily injectionsof 5 cc. of
a l0% solutionof glucosaminein saline,the lesionsclearedbut reappearedwhen treatmentwas discontinued.
We also treateda seriesof casesof rheumatoidarthritis and found
that subjects,who had previouslyrespondedwell to cortisone,also respondedto glucosamine.
Mrs. B. B. had rheumatoidarthritis of the left knee and both
hands. Pain and swellingwere severe.The patient had responded
fairly well to cortisone,but due to retentionof water, this treatment
was discontinued.In lcss than l0 days, the arthritis returned.The
patientwas given glucosamine
injections.At first, 5 cc. of the lOVo
solutionin salinewas injectedonce a day. Later, 3 injectionsdaily
were used.The symptoms,which had decreasedwith the lower doses,
were fully controlledby the 3 daily injections.Pain and swellingfully
disappeared.Moon face was not obscrved.The treatment was continuedwith 2 injectionsdaily and, after one week,one injectiondaily.
With this dose,pain and swellingreturned.Resumptionof 3 injections
a day controlledthem again.The trcatmentwas discontinuedbecause
of the apprehension
of thc patientovcr receiving3 injectionsdaily of
5 cc. each.A mixed treatmentwith a lower doseof cortisoneand one
injectionof glucosamine
daily was continuedwith satisfactoryresults
for a few weeks,after which the patient stoppedher visits.
We treated casesof rhcumatoid arthritis with oral glucosaminewith
fair to good results.Four gramsof glucosamine
a day appearedto be the
minimum requirement.Good resultscould be seenin somecasesafter doses
of 5 to l0 gramsdaily, Theseresultswerc confirmcdby B. Welt who, following his research,usedthe samepreparationof glucosamineon a series
of l5 patientsbedriddenwith arthritisat the GreenpointHospitalin Brooklyn, New York. Subjectiveand objectiveimprovementoccurredin patients
with rheumatoidarthritiswhile no chanqeswere seenfor thosewith osteoarthritis.
For more than one year, Welt has treatcda case of pemphiguswith
injectionsof glucosamineand has beenable to preventthe appearanceof
newlesions.

THERAPEUTTC APPROACH

439

In a few casesof iridocyclitis,treatmentwith glucosamine(2 injections

of 5 cc. of the l0% solutiondaily) healcdthe lesionsin two or threedays.
producedresultsalmostsimilarto thoscof glucosamine.
Galactosamine
Still better resultswere seenwith glucosaminicacid, usedin the samedoses
as glucosamine.Some strikingly good results in severalcasesof arthritis
were obtained with gluconic acid administeredorally in doses of a few
grams a day (four tablespoonsof.a 25% solution).

Hemorrhage
In studying hemorrhage,it seemednecessaryfirst to define preciselythe
conditionsunder which this importantepisodeoccurs,as we did for pain,
itching,etc., sincethe coursco[ hcmorrhage,and particularlyits response
to treatment,seemsto dependgreatly upon certain peculiaritiesrelated to
its pathogenesis.Hemorrhageresultsfrom a break in the continuity of a
blood vessel,which can be inducedby an externalinfluenceon a previously
normal vessel,or can occur as a result of proccssestaking place in abnormal vessels.we havc called the first type of hemorrhage"accidental" or
"traumatic" and reservedthe ternr "pathological"for the second,which
apparsto be a direct result of pathologicalchangesin a vessel.The latter
term is used in the samc senscas it is usedfor fractureswhere "pathological" indicatesonly preexistinglesionsin the bones.
In a traumatichemorrhage,the therapeuticproblem is limited to stopping the flow of blood. Mechanicalmeansto close the bleedingvascular
wound or agentsable to increasethe capacityof the blood to form clots
can be used.They representthe only approachin accidentalhemorrhage.
In pathologicalhemorrhage,other problems arise. The knowledgeof
the local pathologicalchanges,which lead to the appearanceof bleeding,
is importantboth for prcventingand controllinghemorrhage.The pathogenic factor involved will be consideredalong with the problem of hemostasis.
H emorrhagiparousA gents
Local and generalfactorsare involvedin pathologicalhemorrhage.Ulceratedlesionsprovide a favorablecondition.So doesloca.linfection.Anticoagulantsmay act indirectly upon blood vessels.
propcrtiesof somesubstances
Hemorrhagiparous
bccameevidentduring researchon their therapeuticuse in cancerpatients.For example:
P. T., a patient with carcinomaof ttre floor of the mouth widely
ulceratedin the submaxillarregion,bled sporadicallyfrom the lesion.

440

nESEARCH tN

PHYSToPATHoLocY

When a dose of 20 drops of Coramine(brand of nikethamide)was

given for his general condition, a relatively severehemorrhage aF
peared immediatelyafterward.There was another hemorrhagewhen
a similar dose was administered12 hourslater. Suspectinga correlation between hemorrhage and medication, medication was discontinued and there was no hemorrhageduring the following two weeks.
When only l0 drops of Coramine solution was given again, a new
hemorrhageappeared.After two more weeks without Coramine and
without hemorrhage,a new dose was given to check the correlation
betweenmedicationand bleeding,and was followed again by hemorrhage.
Reviewing cases lost through hemorrhage,we could nna s6Eiiffi
which lethal bleedingwas precedcdby administrationof Coramine in the
usualdose.This hemorrhagiparous
effectwas noted so consistentlyas to
us
discontinue
use
of
make
Coraminein caseswith ulceratedlesions.
Alerted by this expcriencc,we noted hemorrhagiparousactivity in
other agents.Glycerol,we found, could induceseverebleedingeven when
administeredin dosesas low as 5-10 drops, and two lethal hemorrhages
werc tracedto such doscs.Thiamine chloridein therapeuticdoses-such
as 100 mg. injections-producedhcmorrhages.
A similarhemorrhagiparous
effectwas noted for isamincblue. Glucose,administeredintravenouslyin
largc dosesto paticntswith ulceratedand infectedcancerouslesionswho
had previouslyhemorrhaged,inducednew blceding.We must emphasize
that all the hemorrhagiparousagentsproduced the bleeding effect, especially in subjectswho had previouslyhad hemorrhages
from their lesions.
This would indicate the importanceof local changes.In such patients,
other agentsalso inducedblecding.In somecasesof gastro-intestinal
cancer, bleedingfolloweda normal doseof aspirin.But even in lesionswhich
had never bled, thc first group of hemorrhagiparousagents-{oramine,
glyceroland thiamins-geulcl inducebleeding.
In trying to correlate the pathogenesis
of these hemorrhagiparouseffects with existingoffbalances,an interestingrelationshipwas noted. A
tendencyto bleed was found to be promoted by sterols.Administration of
cholesterolin dosesas high as 5 cc. of a ZaZ solutionin oil, two or three
timesa day, for severaldays,was followcdby hemorrhagein patientswith
ulceratedlesions.This occurredonly in a few cases.But a high sensitivity
to other hemorrhagiparousagentsoftcn developed.The administrationof
minimal dosesof glycerolproducedbleedingin subjectstreatedwith cholesteroland did so even more frequentlywhen large amountsof insaponifiable fraction of organswere administered.

'f r' t,
:.:.5: i

T H E R A P E T J T t cA P P R o A c H

441

Hemorrhageswhich follow administrationof thesc hemorrhagiparous

agentsare usually severeand of arterialcharacter.Examinationof such
bleedinglesionshas revealedtransverse
severance
of smallor evenmedium
arteries.only rarely was there oozing bleedingfrom capillariesor small
veins.In severalcascs,administrationof hemorrhagiparous
substances
induced petechiaeor purpura, but this occurredonly when thrombopenia
was also present.The petechiaewere scenat siteswhcre local circulatory
impairmentalready was present.We bclievethat this hemorrhagiparous
effect must be emphasizedfor its clinical importance.Productssuch as
C o r a m i n e( n i k e t h a m i d ea) n d t h i a n t i n ca r e w i d c l yu s c da s t h c r a p e u t iacg e n t s
and glyccrolis a common vehiclefor pharmaceutical
preparations.
Therefore, we must bear in mind their possiblerole in hemorrhages.
In subjects
with ulceratedlcsions,their administrationhas to bc banned or special
precautionsmust be used.The samc precautionshave to be taken for the
use of glucosein casesin which a previoushemorrhagchas not beencontrolled bv mechanicalmeans.

Antihemorrhagic A gents
Butanol
I n 1 9 4 3 , d u r i n g r c s e a r c hs t u d i e sc o n c c r n i n gt h e p h a r m a c o l o g i c a l
activity of glycerol on abnormal foci, its hemorrhagiparouseftect
appearedas a serious handicap.Various hemostypticsubstanceswere
testedwithout sufficienteffcct. At this time, a new product, to which a
hemostaticeffect was attributcd.appearcdon the market. It was a very
w e a k s o l u t i o n( a r o u n d l / 1 0 , 0 0 0 ) o f o c t a n o li n s a l i n e .w e c o u l d f i n d n o
therapeuticeflect for it. Howevcr, we were studyingbutanol and other
higher aliphaticalcoholswith lipoidic propcrties,and we decidedto test
butanol for its antihemorrhagicactivity, hoping that it might counteract
the undesirablehemorrhagiparous
effectof glycerol.It did and we have
sinceaddedbutanolto glycerolfor this purposc.
We observedthc remarkablehemostaticeffectof butanolyearslater in
a patient with severehemorrhage,to whom dosesof l0 cc. of a 6.5Vo
solutionwere given intravenously.
Henrorrhagestoppedin a few minutes.
Sincethen, wc have succcssfully
appliedbutanolclinicallyin hemorrhages
of variousorigins.
A s a n a n t i h e m o r r h a ga
i cg e n t .b u t a n o li s a d m i n i s t e r eedi t h e rp a r e n t e r ally as a 6.5vo solutionin saline,or orally as 6.59,i solutionin water.The
r o u t eo f a d m i n i s t r a t i o n - i n t r a v c n o uisn,t r a m u s c u l asr .u b c u t a n e o uosr o r a l

'liijr.t,i!'

442

RESEARcHrN pHysrop^THoLocy

-is

chosenaccordingto the severityof the hemorrhage.Dosesof 5-20 cc.

given
are
and repeated,if necessary,at intervalsof a few minutes. Since
butanol is not at all toxic in theselarge doses,we usually glve them with
good resultsin severeemergencies.
The following casesillustratethe styptic
effect.
R. E., a 64-year-oldman with an extensiveulceratedepidermoid
carcinomaof the floor of the mouth and largebilateralcerviial metastases,had receivedintensiveradiotherapy,occasionally,there was a
small amount of bleedingfrom the oral lesion,but a suddenhemorrhageof about 500 cc. of blood during a harf hour period occurred
l:te at night while the patientwas at home. pressure,appliedto the
floor of the mouth, was of value but bleedingrecurred-immediately
upon release.oxidized gauze,adrcnalinsoakedgauze,vitamin K and
vitamin C in large quantitieswere of no value.n-Butanolsolutionin
salinewas finaliy obtainedand 5 cc. injectedintravenously.Bleeding
ceasedduring the injection.A secondequallyseverehemorrhag"o.curred one week later and again could not be adequatelycontrolled
by pressureor oxidizedgauze. 10 cc. of n-butanol solution administeredintramuscularly
stoppedthe bleedingwithin 2-3 minutes.Three
hours later, the floor of the mouth was carefullyexaminedpreliminary to right externalcarotidarteryligation,and thc lesionwas found
to be free of bleeding.Despitethe ligation,bleedinglater recurredbut
was controlled each time by n-butanol administeredorally.
S. S., a 30-year-oldman, had an adenocarcinoma
involving the
right maxillary sinus with cervicalmetastases.
During the period of
observation,this patientcxperienced
a profusehemorrhagefrom nose
and mouth. Blood flowed at the rate of approximately5-6 cc. per
minute, and pressuregave little or no relief. 5 cc. of n-butanol in
salinesolutionwas injectcdintravcnously
and within two minutes,the
profusehemorrhageceasedand did not recur at that time. on several
other occasions,bleedingwas controlledfollowing the administration
o f o r a l d o s e so f 5 - 1 0 c c . o f 6 . 5 % n - b u t a n osl o l u t i o ni n w a t e r .
A. M., a 36-year-oldman, had multiple pulmonary metastases
from a primary malignantmelanomaof the left foot, On severaloccasions,hemoptysisoccurred and during three of these episodes
bleedingwas profuse.5 cc. of n-butanolin saline administeredintramuscularlystoppedtwo of theseepisodesrapidly,but in the third, an
injection of l0 cc. intravenouslywas neededten minutes after an
initial intranrusculardose. Although the intramuscularinjection was
ineftective,thc bleedingwas halted within two minutes after intrav e n o u sa d m i n i s t r a t i o n .

I n a r e p o r ti n l 9 5 l i n " A n g i o l o g y , (" 1 9 3 ) we presentedthe following

statisticsconcerningthe control of hemorrhagein cancercases:

THERAPEUTIC ^PPRoACH

Number of casesobserved
Number of caseswith profuse hemorrhage
Percent of caseswith profuse hemorrhage
Number of deaths attributed to hemorrhage
Percentof deathsin caseswith profuse hemorrhaget

443

Group I
Group II
Untreated
n
-Butanol
- -256-344
l8
25
j
7
lz
l
67
4

r Death attributeddirectly to hemorrhage.

Since then, theseresultshave been consistentlyconfirmed.

Additional progresswas achievcdwith organic acids added to butanol
in adequate amounts. Completely non-toxic in the dosagesused, they
were obseryedto enhancethe hemostaticeffectof butanol. This hemostatic
effect has been confirmed many times, particularly in Europe where, following our research,butanol has been widely used as a hemostypticagent.
A hemostaticeffect was also evident when butanol was used with the
principal aim of controlling pain in postoperativecases.The hemostatic
eftect was especiallyimportant in cascswhere pathologicalbleedingusually
representeda major complication,either becauseof the impossibility of
obtaininghemostasisduring operationor becausethe surgicalwound could
not be kept aseptic,as in tonsillectomy,prostatectomyand plastic surgery
of the nose.In a study on the use of butanol for postoperativecare in tonsillectomies,B. Welt has been able to show a preventiveeffect, and more
important, a hemostaticone if hemorrhageoccurs. (187) (Fig. 9l) ln
prostatic surgery,the amount of bleedingwas so reduced,that of a group
of 40 cases,only one neededtransfusionwhile in a similar number of
c o n t r o l s , 8b a d t o h a v et r a n s f u s i o n s( 1. 8 9 )
Still more impressiveresults have been noted in pathologicalhemorrhagesfollowing plastic surgery,especiallyof the nose. In a significantly
largenumber of cases,severehemorrhages
tend to occur around the 7th day
We
have discussedabovethe pathogenesis
after operation.
of thesehemorthe
relationship
to
the allergicdefensemechanism.Such hemorrhagesand
rhages have been difficult to control. The use of antibiotics has only
partially reducedtheir frequencyand gravity. S. Scher has obtained very
good results with the administrationof butanol in such cases, using
an injectionof 5 cc. of a 6.5% solutionof butanolonce beforesurgicalintervention and four times daily for two days afterward, followed by oral
of l5 cc. or one tablespoonfour timesa day for eigfrtdays.
administration
No hemorrhagesoccurred in more than two thousandcasestreated. In a

444

xESEARcH IN pHysropATHoLocy

few patientswho neglectedto takc the mcdication,and in whom a hemorrhageappeared,bleedingwas rapidly conrrolledby butanol. (Note 6)
ExperimerrtulResearchon the Henro.stl'ptic'
Eflet't; ln a seriesof studies
we soughtto find the mechanismby which butanol controlshemorrhage.
In an investigationcarried out in our laboratory,M. Bier and p. Teitelbaum (194) showedthat individualmembersof the homologousseriesof
aliphaticalcoholsdecreasethc degreeof rctractionof clots when added in
v i t r o t o b l o o d .( F i g s .l 4 l t o 1 4 3 ) B y v a r y i n gt h e a n r o u n t sa d d e d ,r h i se f f e c r
was observedto occur only at valuesabovea critical concentrationof the
alcoholin blood. Bier could show that the critical value.which diffcrs for
nrembersof the homologousseries,is proportionalto the lethal toxic dose
in mice.
This non-retractionof clots in vitro also can be rccognizedin cascs
treatedwith butanol through the gelatinousaspectof the clots at the moment whenthc hemorrhage
s t o p s .H o w e v e r ,i t i s i n t e r c s t i n gt o n o t e t h a t
t h e a m o u n to f b u t a n o li n j c c t e d c, o n s i d e r e idn t e r m so f t h c a m o u n to f t h e
circulatingblood. resultsin concentrations
considcrablybelow the critical
valuesneedcdto producethis effcct in vitro. However,concentrationof
butanolat the sitc of the wound may explainthis. lt is also of interestthat
the samc gelatinouscharactcrtypifiesthc clots which remain attachedto
b l e e d i n gl e s i o n si n a n i n r a l st r e a t e dw i t h b u t a n o l .I n m i c c , w h e n a p o r t i o n
of tail was cut and butanol was used,the abnormallylong clot remained
adherentto the wound,differingfrom that secnin controls.
ln spiteof changesin the clot, it scenrsthat the effcct upon the blood
itsclf, and its coagulationin a wound, representsonly one of the means
throughwhich butanolcontrolshemorrhagc.
The spcedwith which butanol
acts,often within secondsaftcr intravcnousinjcction,is nruch greaterthan
blood coagulationtime. Consequently,
changesin clot formation alone do
not appearsufficientto cxplain the mcchanismthrough which the rapid
henrostasis
occurs.
M. Bier and H. Lerner in our laboratory studied the influence
exertedby butanol upon hcmorrhageinducedby the highly activeprotcolytic enzyme,ficine.(195) Thcy were able to inducestandardized
hemorrhagesby injectingficinesolutionsunder the skin of the abdomenof white
n t i c c .( F i g . l 5 l a ) ( l n o t h e ra n i m a l sa n d w i t h o t h c rs i t e so f i n j c c t i o ni n m i c e .
the individualvariationswere too greatto make the resultingbleedingusef ul as nraterialfor tcsting the cffect of agcntsupon hemorrhage.
) with
adequatcdosesof ficinc, sevcrehcmorrhages
followcd by skin ulcerations
were induccd.The bleeding.and eventhe ulcerations,
were almostentirely
preventcdwhen butanolwas administcred.
Figurc I 5 I b showsthe resultsof

"r'!:l{-i1r!::,
. i . . .

, .

T H E R . { I } I ]U T I C

A PTROAC}I

445

F r c . l-5la. Shorvsgradcd extcnt of hemorrhagic i n l i l t r a t i o n o b t a i n a b l ew i t h p r o g r c s s i v e a m o u n t s o f f i c i n i n j c c t c d s u b c u l a n c o u s l yi n t h c a b d o n . r i n ; rrlc g i o n i n m i c c .

F : c . l 5 l b . T " h ci n f l u e n c ee x er t e d b l ' b t r t a n o lu p o n l h c h e m o r r h a g ci n d u c e dh y f i c i n i n iection.

T o p R o w . ( i o n t p a r a t i v eg r o u p t r c a t e d w i t h h u l a n o l . l - a s l a n i r n a l i n c a c h r o w
shows tbe exlernal appearanceo f t h c s i t c o f t h e i n j r c t i o n r v h i l e i n o t h e r s . t h e
skin flap was separaled.
Bollom Row: Ficin iniectedanimals.

446

xESEARcH rN pHysropATHoLocy

this experiment, with the manifest difterences,following ficine injections,

betweenanimals receivingbutanol intraperitoneallyand controls.
The antifibrinolytic activity of butanol (196) could explain its intervention in protracted bleeding, and especiallyin caseswhere the effect of
butanol appears after hours or days. However, inhibition of fibrinolytic
activity cannot be conceivedto intervenein an action taking place in less
than a few minutes.
In trying to explain butanol'santihemorrhagiceffects,we also considered its pharmacological
activities.We havenoted the actionof butanol upon
the acid-basebalanceof abnormaltissues,as shownby changesin the pH of
the secondday wound crust. In abnormaltissues,butanol reduceslocal pH
but it does not do this to any great extent in normal tissues.This led us
to investigatethe difterencein the influcnceof butanol upon pathological
hemorrhages.
The immediatcbleedinginducedby standardcutting of the
tail in mice was not constantand, in general,was not markedly influenced
by administrationof butanol. But there was a marked effect upon hemorrhage induced by the displaccmentof the clots through mechanicalmaneuversone to two hours after cutting of the tail. In controls,the bleeding
lastedalmostthe samelengthof time as bleedingfrom fresh lesions,while
in animalstreatedwith butanol, it oftcn stoppedin a very short time.
These data, althoughinteresting,did not seem to ofter a completely
satisfactoryexplanationof the mechanismby which butanol stops bleeding, especiallyin casesin which it acts within minutesor less. It appeared
improbablethat the formation of a clot alone would stop the hemorrhage
under theseconditions.An interestingobservationled us to another hypothesis.
In severalpatientswith wide ulccrations,we were able to examinethe
hemorrhagingvesselafter bleedinghad been stoppedby butanol. Contrary
to all expectations,we found that the artery, which usually was severed
transversely,was not buried in a clot but remained almost isolated and
somehowseparatedfrom it. It seemedthat blood vesselsthemselvesmight
have a role in hemostasis.The interventionof a spasm of the smooth
musclesof the vessclswas considered.This was indirectly confirmed when
a patient with severeand prolongedbleedingfrom the bed of a prostate
after ablation,was treatedwith butanol.An intravenousinjectionof 40 cc.
of butanol was followed by such violent contractionof the bladder as to
expel, with great force, the cathetertogetherwith clots and urine present
in the bladder. At the same moment, bleeding,which had persistedfor
more than a week, stopped suddenly. We connected this sudden spasmodic contraction of the smooth musclesin the abnormal bladder with

rnlfin,

r:r1

iiii

THERAPEUTTC APPROAcH

447

the injectionof butanol and consideredthat butanol might produce a spasm

of the muscularwalls of abnormalblood vg55sl5-3[normal bccauseof the
hemorrhagiparous
condition itself.This would explain the rapidity of the
hemostaticeflect and the selectiveaction upon small and medium arteries
with important muscular walls. In fact, we saw that, while very severe
arterial hemorrhageswere completelystoppedin lessthan one minute after
an intravenousinjection of butanol, the same effect was not obtained in
oozing bleeding.
We tried to investigatefurther this spasticeffect upon blood vesselsas
the mechanism in butanol hemostasis.Experiments with isolated aorta
preparationsof rabbits and rats showedthat a spasticeffect cannot be induced by butanol, even if the vesselspreviouslyhave been harmed by
manipulation.This can be explainedby the fact that the aorta does not
have muscular fibers. The same lack of spastic effect is seen in normal
arteriesof the hind legsof rats and frogs.
The administrationof butanolto an animalafter a smaUbranchof the
mesentericartery was first crushedand, after some timc, cut, produced a
spasticeffect which transformedthe jet-like hemorrhageinto an oozing
one, greatly reducingthe blood loss. The hypothesisof vascularspastic
contractionas the mechanismin butanol hemostasis
has receivedfurther
indirect confirmationthroughthe studyof the cffectof butanol upon hemorrhage induced by single traumatic lesionsin various organs. Differences
were observedaccordingto organsor tissuesaffected.While bleedingIiver
wounds are influencedonly to a small degreeby intravenousadministration of butanol, hemorrhagefrom a kidney wound stopped rapidly. The
fundamentaldifferencebetweenthe liver portal circulationwith minimum
muscularityof the vessels,
and that of the kidney,wherehighly developed
artery muscular layers are seen, can explain the unequal responseto
butanol.
As we can see in thesecases,the contractionof a pathologicalartery
can irsure rapid hemostasisand accordswith the fact that the artery is
not buried in a clot. This explainswhy, especiallyin clinical application,
butanol is more active upon arterialhemorrhageand much lessactive upon
capillarybleeding.This mechanismalso would explain the samegood effect
upon hemorrhagefrom veins with important walls. Failure of butanol seen
in tfuee casesof hemorrhagcfrom varicesof the inferior esophagealveins
can be explainedby the almost completelack of muscularlayers in these
varicoseveins,which would bar a vasculo-muscular
contraction.
Parallel to these studies,other possiblehemostaticmechanismshave
been investigated.We tried to interpret the unusual fact that butanol

448 /

R E S E A R c Hr N p H y s r o p A T H o L o c y

stoppedhemorrhagewhile agentssuch as nikethamide,thiamine, isaurine

blue, sterols and glycerol induced bleeding-yet all have positive polar
groups in their molecules,reprcsentedby an amine or amide radical for
most of them and by hydroxyls for glyccrol and sterols.As we have mentioned before, all agentswith such positive radicalsinduce a shift toward
less alkaline values for the second-day-wound-crust
pH, Therefore, this
could not be consideredto be the factor that determinesthe antagonistic
effectson bleeding.The aliphaticor cyclic characterof the nonpolar goup
doesnot seemto be a factor sinceglyceroland butanol both have aliphatic
chains.
However, correlationwith anotherbiologicaleftectcould be noted. All
the substanceswith hemorrhagiparouseffect, also produce a convulsive
effect if administeredin sufficientlyhigh doses.The convulsive effect of
coramine and thiamine has been known for a long time and we used these
substancesany time we wanted to induce experimentalconvulsions.We
also have seen that convulsionscan be consistentlyinduced in animals
through the use of relativelylarge dosesof glycerol or sterols.Convulsions
in severalhumancaseshavc followedthc useof theseagents.Dosesas high
as 20 cc. of a 57o solutionof the insaponifiable
fraction of placentaor of
the 2Vo solution of cholesterolin oil, administeredfor therapeutic purposeshave induced convulsionsin patients,with a previous history of
convulsivescizures.A convulsiveeffect in animals has been noted for all
the agents mentioned above which also have hemorrhagiparousactivity.
we were able to induce convulsionsin rats, even by injecting 20 cc. of a
solutionof loo/o glucosesubcutancously
once a day for a few days. In
humans,we also saw convulsionsappearingafter glucosewas administered
intravenouslyin an amount of 100 gm. in a saline preparationto patients
treatedconcomitantlywith desoxycorticosterol,
althoughthe last substance
has no convulsivanteffectby itself.
On the other hand, the antihemorrhagic
butanol producesa hypnotic
effectif administeredin high doses.The doseof butanol neededto prevent
hemorrhageinducedby ficine in mice, for example,was enough to provoke hypnotic activity comparableto that of barbiturates,chloroform and
even ether. very interestingis the fact that these agentsalso are able to
prevent ficine-inducedhemorrhageif administeredin doses sufficiently
high to produce hypnotic or narcotic effects,The peculiar correlation between convulsiveand hemorrhagiparous
effect on the one hand, and
hypnoticand hemostaticeffecton the other, providessome further understandingof butanol hemostasis,
but does not furnish the explanationfor
the mechanismthrough which this hemostasistakesplace.The most plausi-

T H E R A P E U T T CA P P R O A C H

449

ble conclusion is that butanol intervenesthrough several mechanisms,

some inducing immediatehemostasisthrough arterial contraction,while
others have a later effect through changesin the blood clotting process,
with lack of retraction of the clot and an influenceupon fibrinolysis.
Contraction would cxplain why, with only few exceptions,it is the
pathologicalarterialhemorrhagewhich respondsmost favorablyto butanol.
An immediateinfluenceupon capillary or venoushemorrhageis lessmanifestand, in a very few cases,absent.The sameis true for traumatichemorrhageswhere,althoughthere are somegood immediateresults,in general
they are lessrapid than for pathologicalarterialhemorrhage.Becauseof its
immediateeffectupon arterialbleeding,butanolbecamethe agentof choice
for those pathologicalhemorrhageswhich, through thcir arterial origin,
could be fatal in a short time.
One of the big advantages
of hemostasis
inducedby butanol over that
obtained through other agentsresidesin the fact that there is no manifest
increasein blood coagulability.Only in concentrations
which nevercan be
attainedin vivo has butanol beenseento changethe coagulabilityof blood.
The inherentdangerof thrombosisrcsultingfrom high btood coagulability
Iimits the amountof the other agcntsto be administcred.
This dangerdoes
not exist for butanol and no suchlimitationsare placedupon its use.The
fact that normal prothrombintime is uninflucncedby butanol while high
prothrombintime is reducedtoward normal rcprescntsanotheradvantage.
In further studics,we tried to enhanccthe hemostaticactionof butanol
without increasing
blood coagulability.
The additionof calciumsaltswas of
value
no
but an enhancement
was secnrvith potassiumsalts,
In a seriesof expcriments,
it wasnotcd that. when a solutionof butanol
is kept for a long time in a stopperedbottlc. its activity increases.For
long-standingpreparations,50% smallerdoseswere sufficientto protect
mice againstthe action of ficine. The narcotic and toxic effectsof these
preparationsalso increasedin the same proportion. This led us to add
butyric aldehyde,the product of immediate oxidation of butanol, which
increasedbutanol's coagulatingeffectsonly very slightly but enhancedits
hemostaticeftect considerably.The addition of hydrogen peroxide did
much the same.
Blood Mixed with Butanol, usedagainstHemorrhages
S. Akad, working in our laboratory,showedthat the coagulationtime
of blood is alsoincreasedif butanol,nrixedprcviouslywith blood, is added.
In the clinical applicationof this observation.u,e used the patient'sown
mixed with butanol.In a syringecontaining,for
blood extemporaneously

450

REsEARcH rN pHysropATHoLocy

instence,l0 cc. of the butanol solution, 2-5 cc, of the patient's blood is
withdrawn. After mixing them, and without removing the needlefrom the
vein, the contentsof the syringeare injectedintravenously.A similar mixture can be injected also intramuscularlyor subcutaneously.The results
have been very good. In some casesin which butanol alone was not able
to stop a hemorrhage,the blood-butanolmixture did. with this special
technique,we have becn able, in recent times, to bring most hemorrhages
under control within a few minutes.
The fact that agentswith positivepolar groups,such as sterols,glycerol,
coramine,thiamine and others, have hemorrhagiparousactivity led us to
try to influencehemorrhagewith agentsconsideredbiologicallyantagonistic
through their negativepolar group.
Many years ago, experimentingwith cilorine solutionsin saline, we
observeda manifcst effcct upon coagulationtime. Intravenousinjection of
such solutionsbrought coagulationtime to values as low as one minute.
The addition of thesesolutionsto butanol greatly increaseits eftectin vitro
upon coagulationtime but had lesseftectupon hemostasisin vivo. Similar
but somewhatless manifest effectswere obtained by adding hydrochloric
acid to butanol solution. On the other hand, organic acids such as oxalic,
malonic,citric, lactic maleic or citraconicshoweda favorable effect.Without changing coagulability of the blood, these acids were seen to increasethe hemostaticeffect.
Fatty Acids
The same antagonismto sterolsand glycerol led us to use fatty acids
from cod liver oil. (197) While resultsin severelarge arterial hemorrhages
were not impressive,the effect upon oozing capillary, venous and small
arterialhemorrhagcs
was vcry good in a largeproportionof cases.For example:
N. V., 57 years old, with multiple pulmonarymetastaticlesions
from a hypernephroma,
had frequenthemoptysis.
At times,the bleeding becamemore accentuated,the patient expectoratingclots as well
as uncoagulatedfresh blood. Intramuscular administration of two
dosesof 10 cc. of a 6.57o solutionof butanol at half hour intervals
had little influence upon the bleeding.Intramuscular administration
of I cc. of a solutionof l0% of the mixture of fatty acids obtained
from cod liver oil stoppedthe bleedingin less than 20 minutes, with
the effect persistingfor more than two months. A new episode of
oozing bleedingwas again immediatelycontrolledby injection of the
fatty acid preparation.

; i,ii:;

Sillj r;;rlri:ir8

..i.i

THERAPEUTTC APPROACH

45L

other caseswith hemoptoic sputum, prolonged bleeding from gastric

or duodenal ulcers or from rectal or uterine turnors, all correspondingto
oozing rather than to acute massivehemorrhage,have respondedto administrationof this fatty acid preparation.Almost uniformly, theseoozing
hemorrhages,
which had not respondedto butanol,were rapidly controlled.
we now use butanol mixed with small amountsof hydrogenperoxideand
organicacidsmentionedabove,to controlseverearterialhemorrhages
while
for the oozingtype, fatty acidsfrom cod livcr oil are used.

Allergic Conditions
Allergic conditions, which as seen above, are related to the defense
mechanism,also have been integratedinto the general therapeutic approach. This has largely permittedus to apply to such conditionsthe same
therapeuticmeasuresused in general,The study of the manifestationshas,
however,shown that exceptfor the anaphylacticshock, the clinical allergy
correspondsto the prolongedphasewith the predominanceeither of sterols
or lipoacids.The urinary surfacetension has appearedas a valuablecriterion to indicate the occurring offbalance.[n caseswith a high urinary
surfacetension,sodiumthiosulfate,aldehydesor epichlorohydrinhave given
particularly good results,while the caseswith low surfacetension showed
favorable responseto heptanol and butanol. Food allcrgies,asthma,urticaria which had persistedfor years and were insufficientlyinfluencedeven
by corticoids-have respondedwith the complete disappearanceof their
manifestationswith an adequatedosc of one or the other of the medications mentionedabove.In most of the caseskept under continuoustreatment with minimal doses,favorableresultspcrsistedeven after the subjects
were again under the influence of antigens,The following observations
illustratethese results.
Mrs. M. S., 45 years old, with skin and eye allergic manifestations,
higbly sensitivefor the past six years to fish, eggs, alcoholic beverages,
some vegetablesand fruits, and especiallyto dogs, showed no favorable
response to corticoids. With high urinary surface tension and high
the patient was treatedwith 50 mg. sodium thiosulfateand
eosinophiles,
2 mg.epichlorohydrin,four times a day. The symptomsdecreasedprogressivelyto disappearin a week. After a month's treatment,she was able
to take-without any inconvenience-foodsand alcohol to which she had
been previouslysensitive.After another two months, the acquisitionof
three puppiesproducedno disagreeable
effects.The patient has continued

:"f-

452 /

REsEARcH rN pHystopATHoLoGy

on the sametrcatment,but reducedto twicc a day, for the past year without recurrenceof the manifestations.
Mr. A. L., 58 yearsold, had frequentattacksof asthmafor over four
and a half ycars which left him unableto work for the past year. Because
of low urinary surfacetension,the patient was put on butanol-S cc. of a
6.5Vo solutionIn wn1s1-to be taken every six hours. Although the first
dosesshowedmarked objectiveand subjectivechanges,the treatmentwas
continued.The patient was free of attacksfor four months at which time
he stoppedmedication.Two weeks after stopping medication,he had an
attack,the first in four and a half months and this was followed by another
the following day. By resumingthe medication,he has been free of symptoms for more than a year.
The possibilityof preventingallergicmanifestations
with butanol,which
appearedso clear in the traumaticlesions,opensan entirelynew view for
many conditions where such a pathogenesis
intervenes.
The conceptof a nervoustissueallergyintcrveningin the pathogenesis
of multiple sclerosis,Ied us to try similar antiallergictreatments.The low
surfacetensionled us to utilizeas agcnts,lipoidswith a positivecharacter.
Interesting-although not constant-results were obtained with insaponifiablefractionof cow brain. Good resultswereobtainedwith butanol-100
mgr. four times a day-together with an antihistamine
preparation.

ARTERIOSCLEROSIS
One of the most important medicalproblcmsfrom all points of viewpathogenic,therapeuticand even social-since it still representsthe chief
direct causeof death in most civilizcdsocieties,
is that of arteriosclerosis.
The fact that lipids seemto intervenein its pathogenesis,
has led us to consider this condition from the point of view of biologicaloffbalances.With
the developmentof our rcsearch,wc tried to apply to this conditiona systematicanalysisin accordancewith the basic conceptspresentedabove.
This attempthas permittedus to arrive at some new views which will be
discussed
briefly here.
The analysisof the specificmanifestationsof arteriosclerosis
from the
point of view of organiTatiorr,
that is, as relatedto the differentlevels,led
us to recognizethat it represents
a conditionprincipallylimited to the level
more
organism,and
especiallyto its secondarypart, the circulatorysystem.
Many of the fundamentalcharactersof the condition could be explainedby
the relationshipof this lcvel to other levclsof the organization,as we will
seebelow.

T H E R A P E U T T CA P P R O A c H

453

From the point of view of the dualisticconcept,it was easy to see that
arteriosclerosis
corresponds
to an offbalancetype A. All the analysesconcerning the systemiclevel show patternswhich indicatc this offbalance.
High urinary surfacetension,low urinaryspecificgravity,high urinary pH,
low urinary sulfhydryl index, found in the routine analysespoint to such
an offbalance.The long persistenceof the skin wheal seen in the subjects
analyzed,confirms this diagnosis.The fact that some manifestations
of
arteriosclerosis
can be induced experimentallyin animals through the administrationof cholesterol,has placedinto limelight the pathogenicrole of
cholesterol.While its administrationin high amountsto rabbitsor cockerels
inducesatheromas,it doesnot inducethe complexconditionitself.Usually
the animalsreturn to normal, even with a rapid healingof their atheromas
after the suppression
of the administration
of cholesterol,Although hypercholesteremia
representsthus only one pathogenicfactor, it appearedinterestingto investigateits interventionin the condition.
The relationshipof cholesteremia
to arteriosclerosis
is certainlynot a
simpleone.The total amountof cholesterol
in blood alone,in its free form,
and the macromolecules
of certain dimensionsrcsultingfrom their bond
to the other constituents
of thc blood, althoughrelatedto arteriosclerosis,
do not seemto representby themselves
the pathogenicfactor of this condition. This latter seemsto be relatedto a more specificinterventionof this
substanceat the level of the blood and circulatorysystcm.Researchin
this field has led us to recognizethis specialintervention.
It is a known fact that the appearance
of crystalsof cholesterolin the
cellsof the intima of the arteriesand in the cellsof the vasavasorumrepresentsan essentialfactor in the pathogenesis
of atheromas.Study of the
capacity of blood serum to dissolveor, on the contrary, to precipitate
cholesterolhas permittedto link it with the appearanceof specificarterial
lesionsin this condition.
In this study, we used the procedureoriginally devisedby Policard
the relationshipof cholesteroland arthritis.Blood is
when he investigated
veinous
punctureand the serumis separatedaseptically.
through
obtained
The amountof total cholesterolis determinedin the blood serum.A sterile
crystal of cholesterolis addedto the serum.The treatedserum is incubated
throughfiltrationfrom
for 6-12 hoursat 37"C. The serumis thenseparated
the crystal added,or from those formed during the incubation.The difference between the amount of cholesterolin the serum before and after
incubationwith the crystal.showsthat while someseraincreasetheir content in cholesterolthroughthis treatment,othersdecreaseit.
We could show that the serum of those rabbits which were fed with

454 /

n E s E A R c Hr N p H y s r o p A T H o L o c y

two grams of cholesteroldaily and which have a tendencyto make atheromas,precipitates

cholesterol.
Oppositely,the resorptionof atheromasseen
in rabbitsafter the suppression
of the feedingwith cholesterol,was seento
occur togetherwith the blood scrum capacityto dissolvecholesterol.The
relationshipbetweenthis capacityto precipiratecholesreroland the appearanceof atheromaswas confirmedby the fact that it could not be seen
in rats fed with cholesterol,where atheromasappear very seldom. Further
it was seenprescntin subjectsprone to make vascularocclusions.The administrationof high amountsof cholesterolto animalswith tunrorshas led
to the appearanceof vascularocclusionsfollowed by ischemicinfarcts in the
tumors.The occlusioncould be relatedto the proliferationof the arterial
endotheliumand the capacityto precipitatecholesterol.

tt

2tc

g
o
t
o /to
I

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L

a@

c
(J

o
o

o
0
a

F l c . 1 5 2 . T h e i n c u b a t i o no f c i t r a t e db l o o d a t 3 7 " C c h a n g e so n l y l i t t l e t h e a m o u n t o f
c h o l e s t e r o li n C a s e ( a ) , w h i l e i t d e t e r m i n e sa r a p i d d e c r e a s ei n C a s e ( b ) .

In a further study of the cholesterolin blood, we investigatedthe

capacityof the differentsamplesof blood to causetheir cholesterolto disappearafter incubationat 37'C. We could show that while some samples
of citratcd blood kcpt at 37"C under stcrile conditions,decreasetheir
cholesterolcontentrapidly,othersdo not changeit even after days of incubation.Figure 152 showstwo such cxamples.We could also show that
in rabbits fed with cholesteroland having atheromas,their blood lacked
the capacityto make cholesteroldisappearafter incubation,diffcring from
what was observedin thc majorityof the controls.
We then studiedin anothergroup of researches,
the relationshipbecontent.We could showthat by washtweenred cellsand their cholesterol
ing red cells with saline,they lose their cholesterol.By using saline in
amountscorrespondingto the plasma,this effectcould be measured.How-

THERAPEUTTC APPROACH

455

ever,while in somebloods a manifestlossin cholesteroloccurswith the fust

or secondwashing,in others the loss occurs very slowly. Sometimesten
such washingsare necessarybefore the cholesterolis low enough to start
hemolysis.Figure 153 illustratestwo such examples.we could show that
the red cells of rabbits fed with cholesteroldemonstrateda higher capacity
to retain cholesterolthroughout saline washings,than did red cells of
normal animals.
The analysisof arterioscleroticcasesfrom the point of view of this
relationshipbetweenblood and cholesterolhas shown that the tendency
to precipitatecholesterol,coupled with a low capacityto make it disappear
through incubation,is encounteredin thosecasesprone to acute episodes
of vascularocclusion.

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# of rashinos

f of rashinqs

F t c . 1 5 3 . T h e w a s h i n g o f h u m a n r c d c c l l s w i t h s a l i n e i n d u c e sa v e r y s l o w d e c r e a s e
i n t h e i r c o n t e n t i n c h o l e s t e r o li n C a s e ( c ) . T h e h e m o l y s i ss t a r t s i n t h i s c a s e a t t h e
lOth washing. In Case (d), such an effcct appears after the second washing.

The big differencesseen in the behavior of blood towards cholesterol

has raised the problem of its relationshipfrom an organizationalpoint of
view. The applicationof the conceptof a proper level rn the organization
to cholesterolhas permitted us to interpret the occurring changes.
Cholesterolrepresentsa cellularconstantand the classicalstudiesof
Shafferhaveshownthe importanceof its amountin the cells.The quantityof
water retainedby the cellsis largelydeterminedby the ratio betweencholesterol and fatty acidsof the cells.Under this aspect,we have hypothetically
consideredhypercholesteremia
as corresponding
to a kind of defenseresponseof the blood level toward the changesoccurringat the level below it,
respectivelytoward an oppositelipidic offbalanceoccurringat cells, tissues
under this specific
or even organs. We investigatedhypercholesteremia

456 /

RESEARcHtN pHysropArHoLocy

aspect.We have seenabovethat in the lipidic system,sterolscorrespond

to an anti-fattyacid agcnt. This Ied to the suppositionthat hypercholesteremiacould representa responseat the blood level to changestaking
placeat a lower lcvelin thc rclationshipbctweenfatty acidsand cholesterol.
A first fundamentalfinding in this directionwas the fact that the amount
of cholesterolin the blood represents-asshown for potassium,copper.
hydrogenion conccntration,
etc.-valucs which, while maintainedconstant
by a regulatorysystcm,vary vcry widcly under differentcircumstances.
These wide variationswould correspondrather to a secondaryresponse.
The amountof blood cholesterolwould changeas a secondaryconstant.A
balancedsystemof manufacturingand cxcretinginsuresthe maintenance
of this secondaryblood cholesterolamount.In the caseof cholesterol,the
manufacturingfactor secmsto be represented
by the R E S cetls,while the
excretingfactor by the liver,primarilyby the Kuppfer cellsand secondarity
by the hepaticcclls, thc cholesterolbeing ultimatelyexcretedin the bile.
we do not know the factor which directly changcsthe manufacturingor
excretionof cholesterolin blood and which consequentlymaintains its
balance.By analogywe considerthe changesin blood cholesterotvalue
to result from thosc crccurringat the lowcr levelsas seenfor other substancessuch as potassium,or copper.for which blood does not represent
the properlevel.Accordingto thisview, with cholesterolhavingthe cellular
level as its propcr lcvel and the blood as superiorlevel,the blood changes
would reflectsecondaryresponses
to thoseoccurringat lower levels.We
saw such changesat the cellularlevel in old age.
We could find thus that organsof old animals,perfusedwith salineto
wash them from their blood, are richer in fatty acids and poorer in sterols
than those of young individuals.The cellular lipidic abnormality,corresponding to aged persons,would correspondto a quantitativepredominanceof fatty acids.Thcir analysisshowedthat thesefatty acidscorrespond
qualitativelyto thosenormally encounteredin the organismsand especially
to the polyunsaturated
members.It shouldbe noted at this time that this
predominancediffersfrom that found in abnormalconditionswhich correspondsto offbalanceD. In this latter case.the fatty acids are abnormal,
With the conceptthat cholesterol
belongsto thecellularlevel,we studied
as we did for potassium(SeeChapter5 Note 2), the concomitantchanges
in the amountof free cholesterolin plasmaand in red cells.This was carried out to obtain informationconcerningthe relationshipbetweencholesterol and abnormal conditions.While a high or low amount in both
plasmaticand cellularcholesterolwould indicatean excessor lack of this
the discordancebetweenthesetwo data would correspondto
substance,

THERAPEUTTc APPROACH

457

a n o f f b a l a n c eA. h i g h c c l l r l a r . w i l h a r e l a t i v e l yn o r m a l o r l o w p l a s m a t i c
cholesterol,correspondsthu:; to an abnormalcholcstcroloffbalanceprcd o m i n a n ta t t h e c c l l u l a r I c v s l , 1 1 ' h i 1ag r c l a t i v e l yl o w c e l l u l a r a n d h i g h
plasmaticcholesterol,
to thc opposireoffbalance.
By now considering
cholesterol in its anti-fattyacid rolc, thcscchangcspermit further interpretation.
As we haveseenabovc,cholestcrolcorresponds
ratherto the anti-fatty
acid constituentscontrollingthc normal polyunsaturated
fatty acids and
thus differsfrom the corticoids*hich representagentsopposingthe abnormal fatty acids.The ultimatccauseof hypercholesteremia
would logically
b e s o u g h ti n a q u a n t i t a t i v c layb n o r m a l l h
y i g ha m o u n to f q u a l i t a t i v e lnyo r m a l
fatty acidspresentat thc low'crlcvcls.
A l t h o u g hh y p c r c h o l c s t c r c n r ci ap r c s c n rbsy i t s e l fo n l y a r e s p o n s et o a
fatry acid offbalancctaking placcat lowcr levels,the high amount of sterol
presentin blood can cause,by itsclf,a scricsof disagreeable
manifestations.
Hypercholesteremia
can thus inducenoxiousmanifestations
although,teleologicallyspeaking,it is directedto correctan offbalanceat a lower level.
The effectof this chronicrichncssin blood cholesterolis manifestedin the
circulatorysystemand athcromatosis
appcarsas an immediateresult.The
precipitatingtendencyfor any ncw incrcascin bloodcholesterolexplainsthe
appearanceof crystalsin the cclls of the intima and secondarilythe appearanceof atheromas.
Therapeutic Attempts
This pathogenicconccptof artcriosclerosis
presented
above.has guided
. s c a r l y a s 1 9 4 2 ,w e u s e da c i d l i p i d i c f r a c t i o n s
o u r t h e r a p e u t iac p p r o a c hA
of organs in a tentativethcrapy to control hyperrensionand indirectly
with intcrestingimmcdiatercsults.We learned however,
arteriosclerosis,
that not only were thc effectstcmporarybut also that after sometime, the
fatty acidsinduceda progressive
administrationof unsaturated
increasein
t h e a m o u n to f c h o l c s t c r opl r c s e n ti n b l o o d .
as a secondaryblood responseto
The conceptof hypcrcholcsteremia
a cellular fatty acid predominancccxplaincdthis occurrenceand led us
also to the further developmentof this approach.With cholesterolas an
anti-fattyacid agentwhich appearsin high amountsin blood, as a response
to an excessof fatty acids at the cellularlevel,we tried to influencethis
secondaryresponseby acting on thc existingprintary cellular offbalance.
could be obtainedby supplyingsubstances
other
A decreaseof this response
than cholesterol,acting as anti-fattyacid agcnts.If the cellular fatty acid
offbalancecan be controlled,the organismas an entity would no longer
be obliged to respondto the occurringoffbalanceand manufactureblood

,.. . S'3T- l.*

458

REsEARcHrN pHysropATHoLocy

cholesterolin excess.without this need,hypercholesteremia

will no longer
appear.Under thescconditions,the administrationof anti-fattyacids has
appearedas the logical way to preventand even to combat an existing
hypercholesteremia
and its consequences.
Choice ol the Anti-Fatty Acid Agent
Thc next problem was the choiceof an adequateanti-fattyacid agent.
It appearedadvisableto use more than one such agent.one would interveneat the lower cellular level where the primary factor-predominance
of fatty acids----exists;
another would act at the level of the tissue and
blooditselfwherethcir presence
would act as an anti-fattyacid and prevent
more directly the further appearanceof cholesterol.
Theseconsiderations
led us to utilizeas adive agents,heptanol,glycerol
and polyunsaturated
alcoholsfor the tissuelevel.Butanolwas added,being
more active at the organ and systemiclevels.We found that mixturesof
thesealcoholswere advisablealso in view of the plurality of fatty acids
interveningin this abnormalcondition,Mixtures of polyunsaturated
fatty
alcoholswere obtainedthroughtreatmentwith lithium aluminum hydride,
of the fatty acidspresentin cod liver oil, fish oil, saffioweroil, sesameoil.
or evenin the lipoacidfractionof organs.
Clinical Results
Thc best clinical resultswere obtainedwith a preparationhaving in
its constitution,glycerol, polyunsaturatedfatty alcohols, heptanol and
butanol.In a seriesof subjectswith persistenthigh amountsof cholesterol
in blood, the administrationof this preparationbroughtthe blood cholesterol to low values,The followingobservations
are characteristic.
Mr. M. R., 60 years old, had high cholesterolin the blood for ten
years,with valuesabove 350 mgr. o/o,in spite of severediet poor in fats
and cholesterol.Administrationof unsaturatedfatty acids brought the
blood cholcsterol,for a short time, to valuesbetwecn260 mgr. and 300,
returningto valuesabove350 after cessationof medication.wtih the mixture-glycerol, polyunsaturatcd
fatty alcohols,butanol and heptanol-the
went down to 150 mgr. vo in lessthan two weekswithout
blood cholesterol
any sidc cffectsor restricteddiet, and with a manifestlygood generalcondition. It remainedat this level for the 5 monthsof observationwith only
minimal and irregularlytakcn medication.
L. N,, 70 years old, with blood cholesterolvarying in recent years
between400 mgr. and 280 mgr. in spiteof low fat and low cholesteroldiet
and differenttreatments,The mixture of glycerol,fatty alcohols,heptanol

THER^PEUTIC

APPROACH

459

and butanol,brought it down in three weeks to 160 mgr. It remained

around this value for more than the six months of observationwith no
specialdiet and with only a very small amountof medication.
In a groupof 20 subjectswith cholesterol
above350 mgr. vo, a d.escent
of thc cholesterol
to valuesbelow250 mgr. Vo wasobtainedin all the cases,
with a treatmentrangingfrom l0 daysto 3 weeks.No inconveniences
were
observed.
Coronary Occlusion
The coronary occlusion with the consequentmyocardial infarction
represents
the most importantcomplicationsof arteriosclerosis.
Death can
occur instantaneously.
In thesecasesit can be consideredto result from
a lesionof such localizationor dimensionas to be entirely incompatible
with the functionof the heart.When deathoccurs,not instantaneously
but
at any time after the occlusionhas taken place,other factors have to be
consideredas interveningand leadingto the fatal issue.Shock appearsto
be the most importantone. Superacute
shock,with deathin a few minutes,
an acuteshock with deathrn l-2 hours,or stateof shock with death within
hours or days,representthe other importantinterveningpathogenicfactors
added to the infarction itself. As seenabove, thesetypes of shock correspond to offbalancesof the type D, this time with a predominanceof abnormal lipoacids,A therapywith anti-fattyacid agentsrepresentsas shown
above, the interventionwhich could preventor reduceshock. while the
administrationof sterolsappearscontra-indicated,
sinceit would increase
vascular
the
occlusion,that of the non-sterolicanti-fatty acids is especially
advisable.
The effect of these anti-fatty acid agentsupon pain and the other
symptomswhich are of alkalinepattcrn,as well as upon the evolutionof
the condition,has fulfilled our expectations.
we attributedan important
glycerol
role to
in thesecases.Its action similar to an anti-coagulant
but
limited to the level of the existinglesions,was able to preventfurther local
thrombosiswithout, however.the dangerof a generalreductionof blood
coagulability.Administeredin adequateamounts,guided by the pattern
analyses,the mixture of the anti-fattyacid agentsmentionedabove has
been followed by manifestclinical improvements.
L. K., 58 yearsold, with a history of severalmyocardialinfarctions,
was seen in a very severestate of shock, The electrocardiogramshowed
that besidesthe old lesion in the posteriorwall, a new infarction of the
anteriorwall was present.Butanolwas administered
in dosesof 5 cc. of the
6.5% solutiontogetherwith 0.1 cc. glycerolevery hour until the complete

460

R t , S E A R C H l N

P H Y S I O P A T H O L O c Y

cessationof symptomsand was continued4 times a day afterward.The

favorablecffectscontinuedin an unexpccrcd
form, the patientbeingwithout
p a i n i n l e s st h a n 3 h o u r sa n d w i t h o u ts y m p r o m st h c t h i r d d a y . T h c c h o l e s terol was found to be 135 mgr. Vo the fifth day.
This view of arteriosclerosis
opensa new way for further researchconcerningmany pathogcnicproblems,and a logicaltherapeuticintervention
i n t h i sc o n d i t i o n .

C H A P T ETR 5
THERAPEUTICAPPROA(IH
TO CANCER
/\
o F T H Eu L T T M A TnEr u s o f o u r r e s e a r c h a s b c c n t o t r y t o u t i l i z e
L/*E
in cancerthe knowledgeobtainedfrom investigations
of the gencralprobIems of pathologyand therapy.Encouragedby the resultsof biologically
guided therapyin many other conditions,we have appliedit to the treatment of malignancy.
As we have mentionedbefore, differencesbetween animal cancers,
both experimentaland spontaneous,
and human cancersrepresentone reason why an agent,howevergood its resultsin animals,may not apply to
human malignancy.Another factor, conductof treatment,is no less important.The main characteristic
of our therapeuticapproachresidesin the
fact that treatmcntis continuouslyguidedby data rcprescntingthe actual
conditionof the subject.At leastfor the moment,it appearsimpossibleto
recognize,through suitabletests,the patternspresentin animalsso as to
apply them to guidcd therapy.Thcrefore,we have becn obligedto do our
therapeuticresearchin humans,reservinganimal studiesfor limited problems.This situationhad led us to emphasize,
always,the experimentalnature
o[ our thcrapeuticeflorts in humans.Although we startedwith desperate
terminal cases,frank immediatesubjectiveand objectivebenefits,even
though temporary,were obtainedfrequentlyenoughto encourageus to go
on. Together with the above mentionedconsiderations,
they seemedto
justify the continuationof therapeuticresearchin human patients.We will
try to reviewas objectivelyas possiblethe resultsobtainedwith therapeutic
methodsand agentsevolvedover the years.
In 1927, a 33-yearold woman with typical preterminalcancerof the
stomach came under our care. In the highly emaciatedpatient, a hard,
461

462

nEsEARcH tN pHysropATHoLoGy

irregular mass frlling the entire epigasterwas palpable. Radiological cxamination indicated a prepyloric gastric tumor. Laparotomy showed an
inoperabletumor of the stomach,with the omentumand lymphaticgJands
greatly involved and evidenceof multiple peritonealand liver metastases.
In view of the patient'sgeneralconditionand the fact that the pyloruswas
only partially obstructed,no surgicalprocedurewas performed other than
biopsy of one of the metastasesin the omentum. The biopsy showed an
adenocarcinoma
Grade III of gastricorigin. Treatmentwas not prescribcd.
I saw the paticnt two yearslater in apparentlygood health.clinical and
radiologicalexaminationat that time showedno tumor. The patient attested
to receivingno treatment.At thc time of the operation she had been trvo
months'pregnant.we had attributedher amenorrheaat that time to the
advancedcachccticcondition.She had given birth at terrn to a normal grl.
Hers was one of thosecasesusuallycatalogued
as "spontaneous
remission."
Since then, I have analyzedmany of the published observationsof
casesof so-callcd"spontaneousrcmission"of cancer alwaysto find a turnilg point that coincided with the interventionof some event usually considered to have no possiblesignificancefor malignancy. The fact that such
eventshavenot inducedsimilarchangesin other cancerpatientshas made
them seemunimportant to many investigators.
While not regardingthem as the only causeof favorable changes,we
havc not eliminatedthe possibititythat sucheventsmay have a contributory
role. We must recognizethat, if such events in themselvesappear to be
powerlessto changethe courseof cancer,they may intervenein conjunction
with, and potentiate,another factor also powerlessin itself to induce a
change.
It was with this concept in mind that we reviewed the case of the
woman with stomach cancer. We consideredthe possibleeffects of two
factorswhich apparentlyintervenedconcomitantly:pregnancyand surgery.
We then begana seriesof experiments.
Ehrlich mammarycarcinomawas graftedin two groupsof female mice,
onc pregnantand the other not. In eachgroup,half of the mice were kept
as controlswhile the other half was submittedto a sham surgicalprocedure
consistingof a laparotomy in which multiple ligatures were performed.
Growth of the tumors and survival times were noted. Compared with nonoperated,nonprcgnantmice servingas controls,both the pregnantmice
and the surgicallytreatedmice showeda slowingdown h the evolution of
cancer lesions.It was in the group of mice, both pregnant and surgically
treated,that a temporary arrest in tumor evolution was seen.In some ani-

:;tiitl)r,l:.i

liti,i

'
i,,I'{,tr,l

THERAPEUTIC

APPROACH TO

cANcER

463

'n
malseventemporaryregression
was noted; 1/20 the tumor regressed
entirely.
PlacentaExtracts
Theseexperimentsled us to try an extractobtainedfrom placentaautolysateswhich, in our opinion at the time, would reproduce,up to a point,
some of the conditions present in these experiments.Human placentas
were autolyzedby being maintainedfrom severalhours to a few days at
37" C, and an alcoholicextractwas obtained.The alcoholwas eliminated
tfuough distillation in vacuum. The residueproved non-toxic in animals
and was injectedintramuscularlyin somc terminal cancercases.
Imprcssiveresultswere observedin the first cases.Pain was markedly
diminishedand, in someinstances,
disappeared
entirely.objective changes
in the tumors could be noted. while only temporary resultswere seenfor
most of thesecases,for some the resultsappearedto last a long time. Their
number would excludepure coincidence.
Mr. H. 8., 56 yearsold, came under our care with a cancerinvolving
more than half of the right parr of the tongue.Multiple large submaxillar
and cervical gland metastaseswere present,two of them being approximately 8 cm. in diameter.The mouth lesion was very painful and bled
occasionally;there was moderate pain in the ganglionar metastases.A
biopsy performed at a much earlier stagehad shown a squamouscarcinoma. Consideredinoperable,the subjecthad not receivedany treatment
except for pain palliation.
We administereddaily intramuscularinjectionsof 5 cc. of the placenta
extract. Except for a limited local reaction at the site of injection, no disagreeableeffectswere seen.On the contrary, after each injection, the pain
in the tongue was reduced for a few hours. It disappearedentirely after
one week of treatment.During the secondweek of treatment,the tumor of
the tongue, as well as the metastases,
beganto decreasein size. The local
reaction at the site of the injectionsincreased,however,to such an extent
that we were obliged to stop treatmentafter 5 weeks.In spite of this, the
lesionscontinuedto decreaseso that the tongue tumor was no longer
palpableafter two months.At that time, the gland metastases
were reduced
to approximatelyone and a half centimetersin diameter.The patient's
general condition was much improved and he gained weight. In another
month, exceptfor a scar on the tongue,no other pathologycould be found.
We followed this casewithout treatmentfor anotheryear and a half during
which time there was no recurrence.After that, the patient left town and
$'e were unableto reestablish
contactwith him.

464

REsEARcH rN pHysropATHoLocy

Mrs. B. A., 44 yearsold, cl..:i u;r 'cr our care with a massivetumor
filling the entire vagina.The crnrl;:;rn had becn diagnosedg months previouslyas carcinomaof the ccr'..x "1;.:h had invadedthe parametriaand
was propagatingtoward thc .,ig.;- r. .', Liopsy made at that time indicated
squamouscarcinomaGradc III. .\: ll c patientwas consideredinoperable
and refusedany other treatment,only scdationwas prescribed.when we
examinedher, the tumor was pr-iilrdirg from the vaginaas a hard mass.
Rectal examinationrevealedin.,'asionof the entire recto-vaginalwall. The
patient received a daily injection cl the placenta extract preparation for
45 days, after which she intcrrupleJ the treatment.The pain had been
entirely controlled in less than a w'cck, but no other changeshad been
observed.
She returnedthree months latcr, having receivedno treatmentin the
interval. Examination revealed complcte disappearanceof the vaginal
tumor, with the cervix entirely rcplaccd by soft scar tissue. we followed
this casefor two years,during u,hich no further treatment was given and
the patient showedno recurrencc.
Mr. A. N., 40 years old had an extensivecancerof the cheek, with
massiveulcerationresultingin a largc communicationbetweenoral cavity
and exterior. Occasionallysmall hemorrhagicepisodeswere experienced.
The patient had had severalcoursesof radium therapy. when he came
under our care, he had multiple lesions,and severalbiopsiesperformed at
that time revealedactive carcinoma in atl lesionstested.The patient receivedintramuscularinjectionsof 5 cc. of the placentaextract preparation
daily for l7 days. A massivehemorrhageoccurredat this point, treatment
was stopped, and he went home without further medication. when the
patient returnedthree months later, scar tissuecoveredall areaswhere the
tumor had been seen previously.clinically no trace of tumor could be
found. In a few months,the patient'scondition was good enough to allow
his surgeonto attempt a skin graft to cover the big opening in the cheek.
This was not successful.The graft from the skin of the neck unfortunately
underwentnecrosis.
In severalother cases,similar subjectiveand objective changeswere
observedwith use of the same alcoholicextract of human placentaautolysates.During this time, we attemptedto substitutecow placenta,utilizing
both fetal and maternalparts, which are easily separablein the cow. In a
relatively small number of casesin which these products were used, we
could see no differencesin the influenceof placentaextract accordingto
origin. Poorer resultswere obtained with extractsusing fresh placentain-

rHER^pEUrrc ^ppRoAcrrTO cANCER

465

s t e a do f t h e a u t o l ; ' s a t cH. o u ' l ' c r , c c r t a i ni n t c r r ' s t i n cgl i n i c a l r e s u l t si n d i c a t c d t h a t f r e s h p l a c e n t ar t i l l h r s a c a p r c i t l ' t o i n l ' l u c n c ew h a t c a n b e

c o n s i d e r e dt o b c t h c n o r r : r u cl o r r r s cr . f c a n c c r .T h c f o l l o w i n gc a s ei s a n
example.
M r s . C . , 5 4 y e a r so l c , h , r c i r t u m o ro f t h c r c c t u mr v h i c hw a s c o n s i d e r e d
i n o p e r a b l eo.n l y a c o l o s t o m ) ' \ \ ' apsc r f o r n r c dP. a i n w a s s l i g h ta n d n o o t h e r
treatmentwas institutcd.Scvcralmonths after the colostom!, the patient
c a m e u n d e ro u r c a r c . A t t h u t t i m c , t h c t u m o r f i l l c d t h c e n t i r c r e c t a la m pulla. Trcatmentwith thc frcsh co*' placcntairlcoholicextractwas started,
using intramuscularinjectionsof 5 cc. daily. After lessthan a month o[
t r e a t m e n tt,h c t u m o r d i m i n i s h c di n s i z c ,I c a v i n ga s m a l l p a s s a g e
for the
e x a m i n i n gf i n g e r .A f t e r a n o r h e rm o n r ha n d a h a l f , t h e t u m o r h a d e n t i r e l y
disappearcdand the rcctal ampullawas wide open. Proctoscopic
examination showedonly normal mucosa.This casc was followcd for two and a
h a l f y e a r sw i t h n o e v i d e n c co f r c c u r r c - n cTc h. e r e a f t e rb, e c a u s e
of the war.
wc lost touch with her.
I n s p i t e o f s u c h r e s u l t si n a f c w c a s c sh o w e v e r ,e x t r a c t so f f r e s h
placentawere judged to bc much less effcctivein ggneralthan extracts
of placentaautolysates.
A t t h e b e g i n n i n go f o u r r c s e a r c hs, t i l l u n a w a r co f t h e d u a l i s mi n t e r veningin cancerpathogenesis,
we observeda seriesof casesin which the
placentapreparationsin gcneralproduccdundcsirableresults.such as increasein pain intensity.Furthermorc.whcn uscd in higher dosesover a
longer period of time, it inducednew pains which clearly increasedwith
e a c h s u b s e q u c ni nt j e c t i o n F
. o r a w h i l c ,t h i s f a c t m a d e u s l i m i t t h e u s e o f
the product to only those paticntsshowingfavorableresponsesin pain,
until we could find an explanationfor theseparadoxicalresults.
With the progressof our researchand recognitionof dualism in the
pathogenesis
of pain, we limited the useof the placentaextractsto patients
with an acid patternof pain. This improvedthe subjectiveand objective
results,and reducedthe casesin which undesirableeffectsoccurred.In
o v e r 1 0 0 t e r m i n a lp a t i e n t st r e a t c dw i t h t h i s p r e p a r a t i o n
b e t w e e n1 9 3 5 a n d
1938 in differenthospitalsin Paris,objectiveimprovcmentwas observed
in 20%. In a few, tumors disappearcd.
Acid patternpain was relieved.In
many of thesecases,however,after a periodin which the tumor decreased
in size,or even clinicallydisappeared,
it startedto grow again and could
not be influencedby further treatment.Furthermore,when the dose was
other pathologicalmanifestations
appeared.The followingcases
increased,
are examples.
Mn. B. 8., 42 yearsold, came under our care with severepain result-

..":i;"

:..:.rl;i

:j

l"'&:{lr.,l i rs.":r

466

nESEARcH rN pHysropATHoLooy

ing from a widely ulceratedcancerof the cervix involving the parametria

and the vagina.5 cc. of the cow placentaextract was administereddaily
and the patient remainedwithout pain for almost three weeks,after which
time the pain returned.An increasein dosage-to two injectionsof 5 cc.
daiiy and then to two injectionsof l0 cc. daily-resulted not only in an
increasein pain but also causedthe appearanceof an abundant watery
vaginal discharge.In a few days this reachedseveralliters a day. Despite
the fact that we stoppedtreatment,the exudatecontinued to increase.At
one point, it amountedto 8 liters in 24 hours. The very concentratedurine
was reducedto lessthan 200 cc. in 24 hours. The patient died in ten days
in spite of all attemptsto stop the excessivesecretion.
Mrs. G. L., 48 years old, had a radical mastectomyfor a left breast
adenocarcinoma.A rapidly growing local recurrencewas seen 6 months
later.The patientcame under our care with an ulceratedtumor occupying
the entire left half of the chest.Administration of 5 cc. of cow ptacenta
extractfor two weeksnot only increasedthe burning sensationpresentbut
causedthe appearanceof an abnormallyabundant watery exudate.As is
often true in such cases,an infectionwith B. pyocyaneuswas seen.A clear
fluid was observedsurgingin drops from the ulceratedlesion. By weighing
the dressing,the amount excretedwas measuredand found to exceedl0
kilos a day. Despiteuse of salineinfusions,calcium preparations,vitamin
c in high doses,atropine, and other measures,the patient expired in less
than a week. The appearanceof such complications,the frcquent changes
toward alkaline patternsof pain, and the increaseof intensity of alkaline
pattern pain, made us reduce and ultimately stop use of these placenta
cxtractsin spite of some good resuJtsobtained.
Cod Liver Oil Fatty Acids ond Sterols
At the same time, progressin our researchhad led us to recognize,
in addition to dualism in the pathogenesisof many manifestations,the
specialrole played by lipids. In 1938, we began to use two groups of antagonisticlipids, fatty acids and sterols.We startedwith a mixture of fatty
acids prepared from cod liver oil for one group and with cholesterolfor
the other. Later we utilized only the polyunsaturatedmembers from the
group of cod liver oil fatty acids.
Fatty acids were administeredintramuscularlyin oily solutions or in
gelatinouscapsulesby mouth. As with administrationof placentaextract.
the immediateeffect was favorableon pain of an acid pattern, and adverse
on alkalinepain. In both cases,the effectoccurredin a few minutes.Therapeutic attemptswith fatty acids were consequentlylimited to patientswith

THER^pE,UTlCAppROACH To

CANCER

467

an acidpatternpain and with this restriction,pain was efficientlycontrolled.

We usedthe efiectupon pain as a criterion,and we discontinued
treatment
in any casein which fatty acidsinducedor increasedpain.
Subsequently,
alongwith the effectupon pain, we usedurinary pH an<J
specificgravity as criteriafor treatmentwith fatty acids.A persistenthigh
urinary pH and a low specificgravity were indicationsfor the use of these
substances.
In additionto the control of severepain, intcrestingobjective
changesoccurred.Unfortunately,most of them were only tcmporary.The
followingtwo examplestaken from a goup of l5 similarcasesare illustrative.
L. B., 66 yearsold, had cancerof the right lung for which he had receivedonly symptomatictreatment.For more than a month the patienthad
complainedof pain in the right chest,with increasingbreathingdilficulty,
ChestX-ray examinationrevealeda tumor of the right lung extendingfrom
the mediastinuminto the mediumlobe.A diagnosisof bronchogenic
cancer
was ntade.Subsequcnt
X-ray cxaminations
showedrapid growth with sevcral tumorsin the upper lobe and in the left lung. Thc gencralcondition
was rapidly and progtessively
deteriorating,
the dyspneaand pain increasing. Two monthsafter first symptoms,the patientwas bedridden.
When the patient came under our care a few wceks later, he was
dyspneic,slightly cyanotic,had persistentcough, was extremelyfatigued
and in almostcontinuouspain. By this time, we had startedto use urinary
specificgravity and pH as critcria for the recognitionof the offbalancc
present.Becauseof low specificgravity and high urinary pH, the patient
was given oral treatmentwith cod liver oil fatty acids.Gelatinouscapsules
containing 0.25 gm. of the fatty acid mixture were used in a starting dose
o f 0 . 5 g m . a d a y , a n d w e r ei n c r e a s epdr o g r e s s i v e tl o
y 1.5 gm. a day.The
patient madean impressivegain in a few daysof trcatnrent.The pain disappearedentirely,as did the dyspnea.The cough also almostdisappeared
in a few days,and in two weeksthe patientwas ableto get out of bed. The
improvementcontinued,and in lessthan two months,the patientwas even
able to go horse-backriding. Radiologically,
the tumors also showcdprogressiveregression.We continuedthe treatment with a relatively high
dosage-2 grams of cod liver oil fatty acids daily-for a total of two
months, with evidenceof continuedimprovement.Then, suddenly,symptoms of pulmonarycongestionbecameapparentand thc generalcondition
rapidly becameworse.Urine analysesnow showeda high specificgravity
and a low pH. In spite of discontinuingthe medication,rhe patient was
back in bed with increasingdyspnea.He died two weckslatcr with symptoms of pulmonaryedema.

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Mrs. D. A., 68 yearsold, had a cancerof the left breastfor which she
had undergonea radical mastectomyfour years previously.pathological
examinationof the lesionhad shown an adenocarcinoma
Grade IV, with
g a n g J i o n ai n
r v o l v e m c n tW
. h e n t h e p a t i e n tc a m c u n d e r o u r c a r e s h e w a s
bedriddenwith a diagnosisof multiple bone metastases.
Radiologicalexaminationshowed multiple osteolyticlesionsin the pelvis, femur, lower
spine.ribs and skull. We institutedtreatmentwith cod liver oily fatty acids
i n g e l a t i n ec a p s u l e sT. h e d o s e w a s p r o g r e s s i v e il n
y c r e a s e db. y 0 . 2 5 g m .
rncrements,until it reached3 grams a day. ortho-phosphoricacid was
addedorally in dosesof vq cc. of a Sovo solutiongiven in water in order
to controlthe pain which appearedafter administration
of the capsulesand
wasof an alkalinepattern.Improvementbeganin a few daysand continued
so satisfactorilythat in lessthan six weeksthe patient was up and about.
Five monthslater, with bone lesionshealed,the patientwent home. I saw
h e r i n I 9 4 1, a l m o s tt w o a n d a h a l f y e a r sl a t e r ,d u r i n gw h i c h t i m e n o t r e a t m e n th a d b e e ng i v e n .w h e n e x a m i n c da t t h a t t i m e ,s h e a p p e a r e di n e x c e l lent conditionS
. u b s e q u e n t l by e
, cause
o f t h e w a r , I l o s t c o n t a c tw i t h h e r .
The increaseof pain, and especiallythe frcquent appearanceof pain
o [ a n a l k a l i n ep a t t e r na f t e r e x t e n d e dt r c a t m e n t c, o n s i d e r a b l lyi m i t e d t h c
use of thesecod livcr oil fatty acid preparations.
Furthermore,an incons i s t e n ciyn o b j e c t i v c h a n g ew
s a ss e e ne v e nw h c na d m i n i s t r a t i ow
n a sg u i d e d
b y t h e a c i d o r a l k a l i n ec h a r a c t eor f t h e p a i n , I n m o s t p a t i e n t s f, a v o r a b l e
objectivechangcswere only temporary.
During this research,wc observeda very favorablcrcsponscin some
casesof hemorrhage,
especiallyof the long-termoozingtype, treatedwith
thesepreparations.Bleedingusuallystoppedafter one injection of I cc.
of a l0olc solutionof unsaturatcdmembersof cod livcr oil fatty acids.We
still use this preparationfor this purpose,as mentionedpreviously.
Among thc group of lipids opposedto fatty acids,we first usedcholest e r o l w i t h t h e i n t e n t i o no f t r y i n g t o i n f l u e n c ep a i n h a v i n ga n a l k a l i n ep a t tern. The effect was much less impressivethan that obtained with fatty
a c i d si n p a i n o f a c i d p a t t e r n .I n s o m ec a s e so. b j e c t i v ec h a n g e sa l s o w e r e
o b s e r v e da l t h o u g ht h e y w c r e l c s s f r e q u e n ta n d l e s sp r o f o u n dt h a n t h o s e
secn with the fatty acid preparations.Cholesterolalone never produced
t o t a lc l i n i c a ld i s a p p e a r a n coef t u m o r s .
Acid Lipidic' Fractions and Unsuponifiable
The developmcntof the conceptof dualisticpathogenicintervention
o f t w o g r o u p so f l i p i d s l e d u s t o a t r e a t m e net m p l o y i n gt h e s et w o t y p e s
o f l i p i d c o n s t i t u e n tosf t h c b o d y c h o s e na c c o r d i n gt o t h e c h a r a c t e ro f t h e

T H E R A P N U T I C

A P P R O A C H T O

C A N C E R

469

manifestations.
We obtainedfrom differentorganicsourcesthe insaponifia b l ef r a c t i o na s w e l l a s t h e a c i d l i p i d f r a c t i o n H
. u m a n p l a c e n t aw a s w i d e l y
employed.The two preparations,acid lipid and insaponifiablefraction,
w e r e u s e d u n t i l 1 9 4 3 o n a b o u t 2 0 0 p a t i e n t s( i n F r a n c e ,a n d M e x i c o ) .
Somegood subjectiveand oblectiveresultswere observed.The subjective
changeswere most impressive.Frequently,an injectionof only I cc. of
the human placentaacid Iipid prepararion(5c/c in oil) controlledpain in
a few minutes,with relicf lastingfor hours or sometimeseven days. with
the samcpreparation,alkalinepain increased
after only a few minutesand
sometimesbecameunbearable.In casestreatcdwith placentaacid preparation, usingacid pain patternas a criterion,we observedsome significant
objectivechanges.
Mrs. B. B., 54 yearsold, with a papillaryadenocarcinoma
of the ovary
and multiple peritonealmetastases
found during exploratorylaparotomy,
.l
had rapidlyreproducingascites. hc patientrequircdrepeatedparacenteses
a t s h o r ti n t e r v a l sI.n t h c m o n t hj u s t p r i o r t o c o m i n gu n d e ro u r c a r e ,i t h a d
beennecessary
to tap her once a week or evenevery five days,Treatment
w i t h a c i d l i p i d s o f p l a c e n t aw a s i n s t i t u t e dw, i t h d a i l y i n j e c t i o n sf,i r s t o f I
c c . a n d t h e n o f 2 c c . o f t h e 5 v c o i l s o l u t i o n .I n l c s st h a n t w o w e e k s ,t h e
pain was controlledand much lcsslluid accumulated.
The patienthad two
more paracenteses
at two and three week intervals,after which fluid no
longerwas a problem.The multipletumor masses,which wcre very easily
felt throughthe skin after cach paracentesis,
were seento decrcaserapidly
and disappcared
in about two and a half monthsof treatment.After four
m o n t h s ,t r e a t m e n tw a s d i s c o n t i n u e a
d n d n o r e c u r r e n c ew a s s e e nd u r i n g
years
we
the three
followedthis patient.
W e s a w M r s . L . S . N . . 7 3 y e a r so l d , i n 1 9 4 2 i n a s u b c o m a t o usst a t e ,
w i t h d e e pj a u n d i c ea n d w i t h a h i s t o r yo f p r i m a r yt u m o r o f t h e s t o m a c ha n d
m u l t i p l e b i g m e t a s t a t i cl c s i o n so f t h e l i v e r . H e r c o n d i t i o n ,w h i c h h a d
s t a r t c da f e w m o n t h se a r l i e r w
, a sg c t t i n gr a p i d l yw o r s e .w h e n w e e x a m i n e d
h e r . t h e I i v e r w a s o c c u p y i n gt h e a b d o m e nu n t i l t h e p u b i s ,a n d p r a c t i c a l l y
c a c ho f t h e i n d i v i d u a m
l e t a s t a t itcu m o r sp r e s c n a
t t t h c s u r f a c co f t h e l i v e r
w a s e a s i l yp a l p a b l et h r o u g ht h c t h i n a b d o m i n awl a l l .
G u i d e db y t h e u r i n c a n l l y s e s - w i t h a h i g h p H a n d a l o w o x i r e d u c t i o n
i n d e x - w e s t a r t e dw i t h a t r c a t m e n w
t i t h l 0 % s o l u t i o ni n o i l o f a h u m a n
p l a c e n t al i p o a c i dp r e p a r a t i o nT. h c d o s e sw c r e i n c r e a s e d
a c c o r d i n gt o t h e
a n a l y s e su n t i l t h e y r e a c h e d3 i n j e c t i o n sd a i l y o f 2 c c . e a c h .T h e p a t i e n t
i m p r o v e d a, n d i n l e s st h a n u w e e ks h ew a s c o n s c i o u as g a i n .H e r c o n d i t i o n
c o n t i n u e dt o i m p r o v ef o r m o r e t h a n a m o n t h w h e n a r a p i d c h a n g ef o r t h e
\,\'orse
took place.As the analysesat this time showedthe oppositeoffbal-

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R . E s E A R crHN p H y s r o p A T H o L o c y

ancepresent,the treatmentwas changed.In additionto infusionswith glucose and saline,glycerol in a dose of g drops daily with 20 drops of
coreminewas given.Again the response
was impressivcly
good.The general
conditionimprovedrapidlyand in lessthan a month, the patientwas out of
bed and startedto take care o[ her home. By this timc, the jaundicehad
almostcompletelydisappeared,
althoughthe liver remainedalmost of the
same dimension.The patient continuedto take glycerol for more than 6
months,lcadingan absolutelynormal life. It took almostone year for the
liver to come back to normal dimensions.
The patient remainedin perfect
healthfor the next 9 years.Severalcoronaryocclusions
led to her death at
the age of 83, from a myocardiarinfarction.She showedno objectiveor
subjectivesigns of recurrenceof either her stomachor liver condition.
Unfortunately,uncontrollablechangestoward rapid tumor gowth ultimately occurred in most casestreatcd with placentaacid lipids despite
favorableobjectivechangesat the beginningof treatment.
Clinlcaluseof the insaponifiable
fractionpreparationssupportedobservationsmade in animals,indicatingthe importanceof other factors for
obtainingfavorablechangesin tumors.With the insaponifiablefractions of
placenta,marked clinical effcctscould be obtaincdonly when the conclition
of the patient permittedthe treatmentto be continuedfor a long time.
Temporaryregressionor evenclinical disappearance
was effectedin several
cases,only to have the tumors start growing again,this time bcyond control
by the mcdication.Even at this point, it was obviousthat favorableinfluence with thesc two antagonisticgroups of lipids was dependenton using
the lipid which corresponded
to the patternpresent.It becameincreasingly
evidentthat changesin the patternoccurredduring treatment.Administration of acid lipidic and insaponifiable
fractionsinduced unfavorableresponses
in patientswith tumorsin which a predominance
of the samelipids
was indicatedby analyses.Pain increasedand unfavorablechangesoccurredin the evolutionof the discase.For cxamplc:
Mrs. A. D., a 42-year-oldwoman,operatedon for an adenocarcinoma
of the breast l8 monthsprior to comingunder our care, presenteda few
skin lesionsnear the operationscar.Radiologicalstudy revealeda few small
osteolyticlesions in two ribs, and one in the skull. The analytical data
showedlow urinary pH, high specificgravity and high chloride index, indicatingan offbalancewhich we attributedto predominanceof fatty acids.
Treatmentwith 2 injectionsdaily of i cc. of a 5o/ooily solution of the insaponifiable
fraction of placentawas started.This was later increasedto
2 cc. twice a day. The patientleft the city for a summervacationand took
the medicationwith her. Disregardingour instructionsthat treatment must

THERApEUTTC AppROACH TO CANCER

4 71

be guidedby further analysis,she continuedit without interruption for four

weeks,despitea rapid changein her condition and a constant incrcasein
the pain after each injection.when we saw her again a month later, all the
analyseshad changedmarkedly, indicatingthe appearanceof an opposite
offbalance.clhically, the condition also had changed.The skin was extensivelyinvolved in the vicinity of the operativescar and multiple, rapidly
growing metastases
were seenall over. Radiologicalstudy showedextensive
new lesionsin many bones.The condition had progressedin one month in
a manner never seenbefore in any patient and we had to relate it to continued use of medicationafter a changein the offbalance.'I-hischangewas
similar to that seenin animalswith massiveadministrationof the insaponifiable lipidic preparation. Switching to placenta acid lipid preparations
quickly changedthe evolutionof the conditionin this patient. pain was
controliedand for three monthsthe conditionseemedarrested.In spite of
treatment,however, it started to evolve rapidly again after that and the
patientdied five monthsfollowingher rcturn from vacation.
Similar deleteriouseffectswere seen with acid lipid preparationsobtained from organs when their administrationapparentlywas at variance
with the pattern present.
w. s., 56 yearsold, had a carcinomaof the cheekmucousmembrane
which led to wide perforation.Biopsy of the edgesof the ulcerationshowed
squamouscanceroustissue.Basedupon the urine analysis,which indicatcd
low specificgravity and high pH, treatmentconsistedof two daily injections
of I cc. of the acid lipid fractionof placentain a 57o solutionin oil. As
the patientwas treatedon an ambulatorybasis,he was advisedto sce us
in a few days. He continued the treatment without any control for two
weeks.when next seen,a marked gelatinousedema of the tissuessurrounding the ulceration was found and the patient complaincclof severe
pain. Biopsy at this time revealed,in additionto intersririaledema,a high
vacuolizationof the cancerouscells which had not been seenin the biopsy
done the day prior to beginningtreatment.The pain bccamcunbcarablea
few minutesafter eachinjection.Theselocal changeswere accompaniedby
a marked deteriorationof the generalcondition,the patient complaining
of a sensationof weakness.The unfavorablechangeswhich occurredin
only two weekswere very impressive.
Favorableresultswere obtainedevenin terminalcaseswith thesefraction preparations.In some cases,arrestor disappearance
of tumors was
noted. In most cases,however,tiese eftectswere only temporary. After
being arrestedfor months or even yearsby theselipid preparations,some
tumors began to grow and to becomepainful and could not be as reaclily

All
1 t L

R E S E A R C H

I N

P H Y S I O P A T H O T - O C Y

c o n t r o l l e da g a i nb y t h e s a m ep r e p a r a t i o nI .n a s m a l l n u m b e ro f p a t i e n t s ,
about 3Vo of the group of 200 treatedwith thesepreparations.
the favorableresultscould be maintainedover a numbcrof years
Even with the relativelystrict guidanceof therapy by the analyses
availableat the time, resultswere not alwaysfavorable.We attributedthis
both to the agentsused for therapyand the criteria employedfor recognition of offbalanccs.
For a long time, researchwas devotedto developing
meansto permit betterrecognitionof offbalances
and to ascertainthe value
of the variousanalysesusedas criteriafor the conductof treatment.Each
new urine or blood tcst was investigatcdas a criterion for the group of
lipids to be administcred.This lcd to bettcr resulrsin controlling pain,
i m p r o v i n gt h e g e n e r a cl o n d i t i o na n d e v e n i n o b j e c t i v cc h a n g e si n t u m o r s .
H o w e v e r ,t h e t e m p o r a r yc h a r a c t c ro f t h e c f T e c t o
s b t a i n e dw i t h l i p i d s
derivedfrom nornralorgansappearcdmore and nrorc evidentin long-term
appraisalof results.We changcd from human placenta to other sources
for both acid and insaponifiablc
fractions.We prcparedand used lipids
from differentorS,ansof cow, pig, tish. and chicken.We also used molI u s k s ,c h i c k e ne m b r y o s ,m o l d s a n d c v e n m i c r o b e s a. s w e l l a s m i l k a n d
eggsas sourcefor thcselipids.In one group of invcstigations,
we even tried
to use lipids of the organ from which the tumor derived.Most of these
preparationssatisfactorily
controlledpain, and in somecases,good results
wereseenin the growth of tumors.
T h e f o l l o w i n go b s e r v a t i o cno n c c r n sa c a s et r e a t e dw i t h t h e l i p o a c i d s
of humanblood.
A . M . , a 5 6 - y e a r - o l dn r a n .w a s r e f e r r e dt o u s b y h i s p h y s i c i a nw i t h a
diagnosisof cancerof the rcctum.Difficultyin defecation,mucosanguinolent discharges,and pain in thc rectal region had been increasingin the
t h r e em o n t h sp r i o r t o t h e d i a g n o s i sE. x a m i n a t i o nh a d r e v e a l e da t u m o r o f
a cauliflowertype, startingat about 4 cm. from the anal orifice and almost
entirelyfilling the rcctal anrpulla.A biopsyhad shown it to be an adenocarcinoma,Crade III. The patient had refusedsurgicalinterventionbecause,yearsbefore,a minor operationon his right hand had led to local
infectionfollowedby amputationof the hand.
At the time he was referredto us. his main complaintswere pain in
the rectum,radiatingto the left lc'g,and tenesmuswith frequentmucosanguinolentdischarges.
We employeddaily injectionsof I cc. of a 5 % olly
solutionof the acid lipidic fructionobtainedfrom human blood. The treatm e n t w a s c o n t i n u e df o r s i x w e c k s ,t h e i n j e c t i o nb e i n gg i v e n d a i l y d u r i n g
the first two weeksand twice a week thercafter.After one week. the pain
and tenesmusdisappcarcdand thcre was a decrease
in the mucosanguinol-

T H E R A p F . U ' , r t CAt p p R o A ( ' l t

ro

cANCE.R

473

ent discharge.
There was also an obviousdecrease
in the sizeof the tumor.
In lessthan a month the tumor regressed
to one-fourthits originalsize;in
six weeks,digital and proctoscopic
examinationshowedno clinical tumor.
A whitish scar could be observedin the posteriorwall of the rectum.
Thereafter,the condition of the paticnt was followed indirectly through
reportsfrom his physician.Therc was no tumor recurrencein spite of the
fact that he receivedno further treatment.He died six vearslater from an
acuteparatyphoidinfection.
Our clinicalexperienceprovidedamplecvidenccthat the preparations
rich in polyunsaturated
fatty acids would influencepain as well as the
growth and evolutionof human cancers.Using the sameamountsof polyethenicfatty acidspreparedfrom varioussources,no differencesin efiects
could be noted. The effcct upon tumors in all instanceswas relatively
limited.
our next effort was to try fatty acidsunlike thosefound in the organism. They includednorbixine-the monomcthylester of the bicarboxylic
acid, bixine-which we uscd in a group of patientsbetween 1938 and
1940.with urinary specificgravityand pH as crireria,daily dosesof from
I mgr. to 100 mgr. were administered
to 30 preterminaland terminalpatients with pattcrnscorrespondingto prcdominanceof sterols.Even with
small doses,the changestoward a predominanccof fatty acids were impressive.Superficial,massivetumors were often seento melt away within
a f e w d a y s ,u s u a l l yl e a v i n gu l c e r a t i o ni n t h e i r p l a c e . - f h i sr a p i d c h a n g eo f
a m a s s i v ct u m o r i n t o a n u l c e r a t e o
d n e ,h o w e v e ru, s u a l l yw a s f o l l o w e db y a
manifestdeteriorationof thc generalcondition. With this preparation,
once the offbalancewas changcdfrom thc original to the oppositetype,
attemptsto control the new offbalance*'ere usuallyunsuccessful.
This led
us to discontinue
i t s u s e a t t h i s t i m e i n s p i t co f t h e r a p i d a n d i n t e n s i v e
c h a n g eist i n d u c e di n t u n r o r s .
Croups ol Agents
Parallelto theseresearchcs
on fatty acids,the usc of agentswith positive polar groups also underwentchanges.As previouslymentioned,it
appearedincreasing.ly
clear that no singleagentcould be effectivein itself
if an offbalancecorrespondine
to predominance
of fatty acidswas present.
use
Quite early in this researchit could be observcdthat the simultaneous
of two agentsfrom the samegroup appearcdto be a bctter procedurethan
u s i n ge i t h e ra g e n ta l o n e .I n a l i m i t e dn u m b e ro f p a t i e n t st r e a t e dw i t h s i n g ) e
agents,without impressivcclinical changes.bctter effectswere obtained
when combinationswere tried. Although it was difficult to ascertainin

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R E S E A R c Hr N p H y s r o p A T H o L o c y

individual cases that favorable effects were due exclusively to change in

medication,the following casesare interesting.
A. Ch., a 53-year-oldwoman, had a radical mastectomyfor an adenocarcinoma of the right breast with axillar and supra clavicular ganglionar
involvement.Almost two yearslater--during the three months immediately
prior to the time we first 53w hs1-5he showedrapidly developingmultiple
metastases
for which only symptomatictreatmentwas applied. Along with
multiple bone metastases,
and a recentpathologicalfracture of the inferior
third of the right femur, there were liver metastasesand a right pleural
effusionfor which she had been tappedthree times. When she came under
our care, she complainedespeciallyof pain in the lower back causedby
lumbar and sacrum metastases.
Her condition was consideredterminal so
that the surgeondid not think it advisableeven to apply traction for the
fractureof the femur. It was under theseconditionsthat we startedto trcat
her with two injections a day of 1 cc. of a 5% solution in oil of the
insaponifiablefraction of human placenta.Except for the unexpectedsurvival of the patient, no apparentchangewas seenafter two weeksof treatment. The pain, pleural effusionand generalcondition remainedthe same.
The treatmentwas changedto 5 drops of glycerol three times a day,
and the dosagewas progressively
increasedto l5 drops of glycerolt.i.d.
After ten days without changc,Coraminein doscsof l5 drops was given
when necessaryto control the typical alkalinepain. After another weck
therc was still no change.The pain remainedalmost the same, except for
a decreasein intensityimmediatclyfollowingadministrationof Coramine.
When all three substances
were given concomitantly-the insaponifiable
fraction by injection and glycerol and coramine orally-the situation
changedimpressively.Within a few hours, pain disappearedcompletely.
and within one week other manifestations
had totally changed.The pleural
effusion,for which the patient had been tapped regularly each week or
every 6 days, disappeared.
The fracturc which, until then, had appeared
entirely inactive, showed a consolidationso rapid that in less than trvo
weeksa solidcalluswas present.In threeweeksthe patientwas out of bed
on crutches.X-ray picturestaken two monthsafter the changein medication showedmost of the osteolyticlesionsreplacedby new bone tissueand
the fracturereplacedby an abnormallysolid callus,There was no fluid in
the pleura.All treatment.exceptthc glycerol,was discontinuedafter anothcr threemonthswhen,at the start of the war, the patientleft Paris,we
heardthat shc continuedin good healthwihout further treatmentfor three
more years,at the end of which time she developeda recurrence,with
and died shortlythereafter.
Iiver metastases,

THER^PEUTIC APPRO^CH TO

C^NCER

475

M. R., a 58-year old woman, the wife of a professorof gynecology,

had an ulcerationof the cervix two years before coming under our carc.
Biopsy had revealedsquamouscarcinoma,Grade IIL After local treatment
rvith radium,she underwenttotal hysterectomy.
Six monthsbeforewe saw
hcr, she developedmultiple abdominalmetastases
for which only symptomatictreatmentwas prescribed.
When shecameunder our care,she had
a distcndedabdomenin which massesof variousdimensionswere easily
palpable.Besidesseverallarge tumors,two of them about 15-20 cm. in
diameter,there were many smalleroneswhich gave the distendedabdomen
a very irregular appearance.The abdominalpain, her generallypoor condition, as well as sevcreedemaof the legs,kept the patient bedridden.She
showeda higl urinary specificgravity and low pH, and treatmentconsisting of a daily injectionof 2 cc. of.a 25% solutionof cholesterolin oil
was prescribed.
When no subjectiveor objectivechangeswere secnin three
weeks, the treatment was changedto 10 drops of glycerol orally, three
timesa day, for anotherthreeweeks.Thcre was still no obviouschange.
After anotherthree week period, this time without treatment,during
which her generalconditiondeteriorated,
mixed treatmentwith cholesterol
and glycerolwas started.The patientnow made a suddenrcmarkablerecovery.The edcmaof the legsdisappearedrapidly and in lessthan a month
the tumorswere no longerpalpablc.The abdomen,however,continuedto
be distended,but insteadof the previousirregularitywith multiple wclldelineatedtumors, a singlehuge mass was recognized.It filled practically
the entireabdomen.we made the diagnosisof a large ovariancyst which
had probably been presentbefore but had been obscuredby the multiplc
tumors.The patientwas operatedon three monthslatcr and the cyst was
rcmoved.Not only were no tumorsfound in the abdomen,but there were
which the surgeonhad fearcd.White patcheswere seenat the
no adhcsions
sitespreviouslyoccupiedby the tumors.There were no recurrences
during
severalyearsof follow-up,after which we lost track of the paticnt.
Similarresultswere obtainedduring the followingyears,indicatingthe
value of mixed therapy.We made it our standardc[nical treatmentexcept
whcn the pharmacodynamic
effectof a specificagentwas beinginvcstigatcd.
As an exampleof the objectivechangesobtainedthroughmixed treatment for type D offbalance,we give an abbreviatedform of the following
observation.
Miss S., 39 years of age, came under our care with bone metastases
from an adenocarcinomaof the breast, for which she had a radical
mastectomylt/2 yearspreviously.At admissionin very severepain, besides
sheshowedespeciallya markeddestruction
other multiplebone metastases,

476

RESEARCfI IN

P}IYSIOPATHOLOCY

s
#

F r t i . 1 . 5 4 .A r t e r i o p o s t c r i o rr i c * o f t h c c h c s t o f p a t i c n t S a t t i m e o f a d n r i s - s i u rnh o $ ing an osteoll'tic processin thc' { r t h a n d 9 t h l c f t r i h s .

F l c . l - q 5 . , , \ r t e r i o p ( ) \ t e r i t r r v i s r r . o f t h c c h c r t o f p : r ti c n t 5 m o n t h s l n t e r . w i t h t h e
y
i r p p f a r a n c c o f t h e n e r v b o n c s r e p l a r ; i n g t h c p i l r t o f t h c r i h r p r c v i e i u s ldcstroyetl.

T H E R A p L - L i T t cA p p R O A C H T o

c A N C E R

4 7 7

of the 6th and 9th left ribs. (F'ig. 154)The analysesshowedan offbalance
t y p e D a n d c o n s e q u e n t lsyh e w a s t r e a t e dw i t h b u t a n o la n d g l y c e r o l .T h e
patientshowedrapid changcsnot only in her subjectivefeeling,but also
o b j e c t i v e l yF.i g . 1 5 5 s h o w st h e r i b o s t e o l y t ilce s i o n sh e a l e da n d t h e m i s s i n g
bonereplacedby new bone.
In general,however,in spiteof somefavorablecases,the resultswere
not satisfactory,Even in patientsin whom tumors regressedand disappeared,recurrences
wereseenanywherefrom a few monthsto severalyears
after discontinuation
of treatment.By prolongingtreatmentover a period
o f y e a r s ,t h e p e r i o d o f i m p r o v e n l e nwt a s l e n g t h e n e idn s o m e c a s e s S
. till,
all too often the responseto treatmentwas only temporary.
Mercaptans
Other substances
with lipoidiccharacterbut with a polar group different from thc carboxyl were tested.Mercaptans,as lipoids with a thiolic
negativepolar group, were used.In 1942. we startedto study the therapeuticeftectof ethyl mercaptaninjectedintramuscularly,
in a l0o/osolution
in oil. (326)
Use of ethyl mercaptanhad to be limited to subjectswith an offbalancc
corresponding
to a predominance
of stcrols.Although only a few patients
were treatedbecauseof the offcnsiveodor, thc resultswere satisfactoryas
indicatedby the followingcasehistories.
F . C . - T h i s p a t i e n t ,a t t h e a g eo f 6 6 , h a d a s m a l lc a r c i n o m ao f t h e t i p
o f t h e t o n g u er e s e c t e a
d t C o l u m b i aP r e s b y t e r i aHno s p i t a lo n M a y 6 , 1 9 4 1 .
At the same time, a secondcarcinomaof the left lateral bordcr of the
t o n g u e w a s f o u n d i n f i l t r a t i n gi n t o t h e d e e p e rp o r t i o n s .T h i s l e s i o nw a s
t r e a t e dw i t h r a d i u mn c e d l e w
s i t h a t o t a ld o s eo f 2 5 0 0 m g . h o u r s .I n S e p t e n r b e r 1 9 4 1 ,a p r o p h y l a c t i cl e f t r a d i c a ln c c k d i s s e c t i o w
. hc
n a s p c r f o r m e dT
patient was well for 8 months thereafter,and then began to experience
sorenessin the regionof the scar on the lateralside of thc tongue.
I n A u g u s t 1 9 4 2 , l 6 m o n t h sa f t e r t h e t u m o r h a d b e e n i r r a d i a t e dt,h e
p a t i e n tc a m e u n d c r o u r c a r e . A t t h c j u n c t u r eo f t h c a n t e r i o ra n d m i d d l c
t h i r d o f t h e l e f t l a t e r a lb o r d e r o f t h e t o n g u e ,t h e r c w a s a n i n d u r a t i o no f
2 / l c m . i n d i m e n s i o nw i t h a n u l c c r a t i o no f 1 / 0 . , 5c m . c o v e r c db y n e c r o t i c
tissue.The induratedbasewas especially
developedanteriorlyto the ulcerated lesion.There was no evidenceof recurrcnceat the scar of the tip of
the tonguenor at the left sideof the neck.The latterlesionwas very tender.
A W a s s e r m a nann a l y s i sw a s n e g a t i v eB. i o p s yo f t h e e d g eo f t h e u l c e r a t e d
s q u a m o u cs e l l e p i t h e l i o m a .
tumorrevealed
T h e p a t i e n tw a s t r e a t e dw i t h p l a c e n t aal c i d l i p i d f r a c t i o nf o r t h e f i r s t

478 /

REsEARcH lN

pHysropATHoLocy

week without any subjective or objective change. The treatment was

changedand the patient received I cc. of a l\vo solution of ethyl mercaptan in oil intramuscularly,three times a day. The dose was increased
after three days to 2 cc., three times a day. By the end of one week under
this treatment,pain had disappeared,althoughthere was still some burning sensation.The appearanceof the lesionwas unchanged.The treatment
was continued.By the end of the secondweek, the edema was reduced,
and after another week of the same treatment,the induration was gone.
After one month of treatment with ethyl mercaptan,epithelizationof the
lesion was noted. The treatmentwas discontinued.The lesion appeared
completelyhealed,without induration,burningsensationor pain, lessthan
a week later. The patienthas remainedwithout treatmentsince,and there
has been no recurrencein the last 18 years.Three yearsago, the patient
had a basalcarcinomaof the skin of the nose,which respondedwell to
local treatmcntwith radium in Mexico.
Mrs. L. F.-In Junc 1942, the patientthen 50 yearsold, observedan
induration on the right border of her tongue.Biopsy revealeda squamous
cell carcinoma.After the biopsy, the lesion progressedso rapidly that it
was judged inoperablc.Without having receivedany other treatment,the
patientcame under our care in August 1942.At that time, the tumor had
involved the right half of the tongue and multiple submaxillaryand neck
nodes,the biggestbeing 2 cm. in diameter.The lesionwas extremelypainful. urine analysesshowedlow specificgravity and high pH and the parient
was treated with 1jVo ethyl mercaptanin vegetableoil. we started with
three t,/zcc. injectionsdaily, The dose was progressivelyincreaseduntil it
reached6 cc. daily. The pain disappearedentirely in less than a week.
Treatmentwas continuedfor only 372 weeksat which time there was
a marked improvementin the tonguelesion and lymph nodes. Except for
an inflammatoryreactionat the site of injection,no side effectswere seen.
A month after the start of treatment,the lesionshad completelydisappeared.No treatmenthas beengiven since.The patientis still well today,
free of recurrence,18 yearsafter treatmentwith ethyl mercaptan.
H. A.-In May 1942, at the age of 52, the patient developedhematuria, pyuria and tenesmus.In June 1942 an infiltrating tumor of the right
side of the trigone was revealedby cystoscopyand cystogram.The patient
came under our care at thc end of August 1942. Having high urinary
specificgravity,he was treatedfirst with insaponifiablefraction of placenta.
receiving 1 cc. of a 5% preparationin oil three times a day. Hematuria
and the other symptomsincreasedunder the treatmentwhich was then
changedto ethyl mercaptan,ljvo solutionin oil, startingwith 15 cc. three

THERApEUTTC AppROACH To

CANCER

479

times a day. T'he dose was progressivelyincrcascdto 3 cc., threc timcs a

day by the tenth day. Hematuriadecreascdin the first 4tl hours and ceased
completeiyon the fourlh day. Ot-hersymptomsdisappcared.T he treatrnent
was continuedwith this dose for another2)5 weeks.Disappcaranceof the
tuntor was seen in follow-up cystograms.Without further trcatnrent,the
patientremainedwell until the beginningof 1955, l2Fz yearslater, when
a recurrenceof the tumor of the bladderwas notcd. Bcing in Mexico, hc
did not receivetreatment by this method and dicd from generalizedcarc i n o m a t o s iasf t e r 9 m o n t h s .( F i g s . 1 5 6u n d 1 5 7 )

F I c . l - J 6 . C y s t o g r a mo f p a t i e n tH , A , b e f o r et r e a t r n e n l .

Similarfavorablercsultswere obtainedfor other patientswith patterns

correspondingto sterol prcdonrinancc.Wc trcatcd only thrce paticntswith
'I'hc
usc of cthyl mercaptan
patternsindicatingfatty acid predominance.
in thcsecasescauseclan cxacerbationof symptomsand rapid deterioration
of the generalcondition.
The odor of ethyl mercoptanwas so oflcnsivcthat its use pxrscdinsurmouotableproblems. Patientswere forced to bc social reclusesand it
was practicallyimpossiblcto get nursesto adnrinistcrthe injcctionsbecausc
clf persistenceof the odor on skin and clothes.Wc had nrany complaints
that the odor of thc nredicationpollutcd the atmosphcrcof a large area for

4lt0

R E S I J A R C } I I N

P } I } ' S I O P A T I I O L O C Y

a long tinrc. obliged to discontinueits use.we soughrother preparations

containingthiol groupsor bivalentsulfur that we hopedwould havesimilar
biok:gicaleffccts.
we investigated
a lirrgenunrberof such substances
in animals.only a
few wcrc cxtensivclystudicdin patients.of the homologousscriesof aliphatic mercaptans,
rve utilizedpropyl. butyl and amyl nrercaptans.
T'hese
otlcredno advantag,e
over ethyl rnerclptan. beinglessactivebut hardtv less
oflcnsivein odor.

t . l L , . 1 5 7 . ( y r t o g r a n r o f p a t i c n t H . A . . a f t c r t r c a t n r c n tu ' i t l r e t h y ' l n l c r c l p t u n .

Hex-ylrncrcaptanrvasllte lirst of thc scriesthat had a ntorc bearablt-r

odor but it was dclinitclylessactivc.Althoughobjectivcchangesin cancer
were observedrvith its use, thc tumors tjid not disapprear
as thcy did in
sotnecascstreatcd tvith ethyl rnr'rcaptan.and manv favclrableresponses
w'crconly tentpr)rtrv.
Of the highcrntercaptuns,
dodecyland hcxadccylwere nrostextensivell
studied.Whilc thcir odor rvas far lcss objcctitxable,thcy produced lcss
favorablercsultsthan the lowcr homologueslv{uchlarger doseswcre required to inllucnccpain. and cvcn tht:n th{rslTectswcre rcducedand tcm-

r H E R A p E U r r cA p p R o A c HT o c A N C E R

4gl

porary. The striking resultsobtainedwith ethyl mercaptancould not be

duplicatedwith any of the higher homologues.For this reason,we reluctantJyabandonedthe useof mercaptans.
SulluriTedOil
During the years that followed,we tried to achieveresultssimilar to
those obtainedwith ethyl mercaptanby using other agentswith bivalent
sulfur. The one most commonly used was thc so-called"sulfurizedoil"
containingfatty acid hydropersulfides,
whose pharmacological
characteristics have been discussedpreviously.Sulfurizedoil's effectson pain and
systemicmanifestations
were lessimpressivethan thosc of the mercaptans
but better than those of unsaturatedfatty acids and their derivatives.In
severalcases,tumor disappearance
was actuallyobserved.Generallythe
clinical resultswcre neither as consistentnor as persistcntas with the
mercaptans.with the use of this product alone, howcver. long-termfavorableresponses
werethe exception.
For example:
Mr. I. G.-White male, was operatedon at MaimonidesHospital in
January 1950 at the age of -56.A pyloric masswas found and a subtotal
gastrectomy
was performed.Diagnosisof adenocarcinoma
of the stomach
was made.Microscopicexaminationshoweda largeareaof replacement
of
gastricmucosaby atypicalglands,with a grcat massof abnormalcells invading the submucosa.
Thesecellsextendedinto the first part of the duodenum. There were post-operative
complicationswith abscessformations
in the wound, which were incisedand drained.with new complaintsof
pain in the upper abdomen,and rapid loss of weight,the patientwas admitted to MonticelloHospitalin July 1950,and to Kings county Hospital
twelvedayslater. He was then transferredto a nursinghome with a diagnosisof terminalcancerwith recurrenttumor in the upper abdomenand
metastases
to the chest.He remainedat the nursinghomefor sevenweeks.
In September,
he came under our care.
A t t h i s t i m e ,h e h a d l o s t 3 9 l b s . ,a n d c o m p l a i n e o
d f e x t r e m ew e a k n e s s ,
pain in the lower chestand upper abdominalregion,and cough with hemoptoic sputum.The pain was only slightlyrelievedby narcoticsand the
generalconditionof the patient was consideredvery poor. A large mass
was found occupyingthe entireupper abdomen;X-ray examinationof the
chestshowedmassesin the right lung. No fluid was obtainedfrom several
chest punctures.
Basedon the urinary analyses,
the patientwas treatedwith a preparaThe response
was excellent.Not only was his pain
tion of hydro-persulfide.
rapidly relievedbut the mass in the abdomenprogressively
decreasedin

482

RESEARcH rN pHysropATHoLocy

size. Evidence of lung involvementslowly disappearedin X-ray studies.

After November 1950, he continued the treatment at home for over 13
months after which he resumedhis old job as a millinery cutter. He had
gained 55 lbs. since beginningtreatment.For the past 7v5 years,he has
worked without interruption and there has been no clinica.levidenceof
malignancy.The extremecondition under which the patient came into our
care can explain the length of time neededfor general recovery and his
inability, despitedisappearance
of the tumoral masseswithin a ferv months,
job
to resumehis
for more than a year.
Thiosullates
we also utilized sodium thiosulfatein many patientswhere symptoms
suchas pain, vertigo,itching,etc,,could be relatedto a local acid pattern.
In these specific cases, the results were generally satisfactory.Sodium
thiosulfatewas administeredeither orally in drops of a 10% solution in
water, or parenterallyin a 4o/osolution in water. Intramuscularand subcutaneousinjectionswere well toleratedeven when doseswere as high as
10 cc. Dosesas small as lOmgs,were observedto influencesymptoms in
certain patients.However, in somecasesit was necessaryto give as much
as 5 gr. of the 5u!5tnnsg-125 cc. of the parenteralsolution-n24 hours
to obtain any effect.In thesecasesthere were no apparentside effectseven
when this dosagewas continuedfor many days. Effects upon tumors with
the useof thiosulfatealone were seenin severalcasesbut the treatmentdid
not producecompletedisappearance
and resultsusually were only tempo-

rary.
The use of sulfurized oil in conjunction with sodium thiosulfate has
beentestedin a sufficientnumberof cases(more than 75) to enableus to
recognizethat the combinationproduceschangesin pain and systemic
analysis,as well as reductionin the sizeof tumors,especiallywhen given in
adequateamountsover a long enougbperiod of time. In severalcases,
tumors disappearedfor many yearsfollowing this treatment.The following
are illustrativecases:
G. M.-In March 1944, this patient had an ulcerationof the cervix.
Biopsyshowedsquamouscell epithelioma,
Grade III. A total hysterectomy
was performed.She was treatedwith 3,600 mg. hours of radium in and
aroundthe ccrvix in April 1944.Five monthslater, there was evidenceof
local recurrenceand the patient was given 1800 r. of deep X-ray therapy.
In July 1945,examinationrevealedno evidenceof disease.
On October28,
1945,the patient was examinedat the Scott and White clinic in Temple,
Texas, and a diagnosisof extensivemetastaticcarcinoma of the para-

rHERApEUrtc AppRoAcH TO CANCER i

493

metria was made and further deep X-ray therapywas advised.The patient
refused this.
She cameunder our care in November1945.She was extremelyweak
and showedevidenceof considerableweight loss, weighingonly 86 lbs.
Hematuriaand dysuriawere the principalcomplaints.Multiple largetumor
masseswere palpablein the pelvisand extendedinto the abdomenabove
the umbilicus. The largest mass palpable,about the size of a big grapefruit, was in the right lower quadrant.The tumor was found to have invaded the bladder, too.
The treatmentwith which we started,t.i.d. intramuscularinjectionsof
I cc. of placentafatty acids, Ljvo in oil, was changedafter one week, in
view of the preterminalconditionof the patient.A hydropersulfide
preparation containingI o/csulfur,and sodiumthiosulfatelOTo wasadministered
orally. we startedwith a doseof lq cc.threetimesdaily of hydropersulfide
and Yz cc. thiosulfateand increasedit progressively,with ameliorationof
the generalconditionand disappearance
of the hcmaturia.After a month,
the dosagereached3 cc. of the first and 9 cc. of the latterpreparationdaily.
Under this treatment,the patientcontinuouslygainedstrenglhand weight.
The hematuriadid not reappear.
In May 1946,she was admittedto the Universityof Chicagoclinic. A
large fixed, firm, irregular tumor masswas still presentin the lower abdomen risingfrom the pelvisto the umbilicus.Althoughher generalcondition
had improved,the patient againhad urinary frequencyand urgency,and
cystoscopyrevealeda severecystitisand severalsmall stonesin the bladder.
An intravenouspyelogramshoweda right hydro-nephrosrs.
The treatmentwith hydropersulfide
and sodium thiosulfatewas continued for 4 months in Chicagoduring which time the abdominalniass
becamesmaller,softer, less fixed and was no longer tender. By August
1946, her weight was 136 lbs., a gain of 50 lbs. sincestart of treatment.
The treatmentwas continuedduring 1947, althoughabdominalexamination did not revealany palpablemasses.On rectalexamination,however,
the pelvis appearedto be frozen but no definitemass was felt. Cystoscopy
showedseverecystitis,bladder calculi and a distortedbladder.Although
shepassedseveralstonesand gravel,her urinarysymptomspersisted.
In December1948,an attemptwas madein Texasto removea bladder
after lithotrity.The bladderwas perforatedduring
calculustransuretherally
this procedureand a recto-vasicalfistularesulted.The patient'slocal physician in Texas believedthat the patient was terminal, but we insistedupon
This was done, and the surgeonreportedthat there was no evidenceof any
a colostomy to divert the fecal stream as an immediate first procedurc.

{84

R E S h A R (

l N

P H Y s l ( ) P d T H O t . ( } C t ,

'fhe
pelvic clr abdominal nrasses.
patient made a slow rccovery.According
to rcportsto this date. l6 ycarsaftcr start of treatntent,no recurrencehas
beennoted. Recentlythc colostomywits closed.the patientbeing in gtxxl
condition.
M r s . M . L . - l n N o v e m b e r1 9 4 1 ,a t t h c a g e o f 4 0 . t h e p a r i e n rh a d i r
lcft oophorectontyfor a multi-loculatedovarian tumor with ascitesand
peritonealinrplants.Thc pathologicalfinding was papillary cyst adenocarcinoma of the left ovary. Without any othcr treatnrent,she rentainedfree

Flc. l.5tt, Photonticrograph of l;-nrph node fronr ('ase [:.H. showing metastatic adenoc a r c i n o m aC r l l t ( 4 o O r ) .

of symptomsuntil thc beginningof 1945.whcn she startedto complainof

abdclntinaldiscomfortin the lorver left quadranl. A tnass was found at
thc site of operation.Grorvthrvasnoted in furthcr examinations.The putient had no treatmentuntil DecenrberI945, when she came under our
c a r e . O n e x a m i n a t i o na, t u m o r o f l l , / 6 c n t . r v i t h l i m i t e d m o b i l i t y w a s
found. No abdominallluid was prcsentat this time. She was startedon
anrylmercaptan
in dosesincreasingto 6 cc. daily of a !07o solutionin oil.
"rhe treatmentwas discontinued
aftcr a week becauscof the odor. No
manifcstchangeseould bc secn.A preparationof hydropersulfide
containing I {i sulfur. in a dose of I cc. threc timesa duy orally. was used.The
nrassin thc left puramcterdisappearedentirely in about 2 months. She

T H E R A p L U ' r l c A p p R O A C r rT u c A N C C . n /

495

continuedwith the $amenledicationfor unother l4 nronths,There has been

no recurrenceto datc.
Mrs. E. H.-In April 1947.at thc age of 46, thi.sparienthad a right
radical mastectomy. The pathological diagnosis was adenocarcinoma,
Grade III, rvithmetastases
to axillarylymph nodcs.(r'ig. t58) A courseof
post-operativeirradiation was administered.lvlenscshad been internrpted
l{ years before by a total hysterectornyand bilatcral salpingo-oophorec[(]nlv.

:$
r1ffi

i$

's
s,x

:*

TF

"J

2 2/ 4 8
l ' t t ; . l - 5 9 . A n t e r o p o s l e r i o rv i c w o f c h e s t i n ( ' i l s e F . . H . a t c o n c l u s i o no f l c s l o s t e r g n e
l . n d d c e p X - r a y t h c r a p ! 't o \ p i n c . s h o w i n gd c r t r u c t i o no f n r e d i a l ' . r o f l e f t c l a v i c l e
r n d m e t a r t a t i cr i h l e . ' i o n r .

The patientwas free of symptonlsuntil Julv 1948, when she beganto

complain of back pain and difticulty in walking. X-ray examinarionrevealedosteclyticmetastaticlesionsin the nredialthird of the left clavicle,
pelvis,thoracicand lumbar vcrtebrac,rvith collapxeof the ninth thoracic
and third iumbar vertebrae.She was hospitalized
and dcep X-ray therapy
was appliedto the thoracicand lunrbarvertcbralregionsand the right hip,
l80O r. beingdeiiveredto sx.h of thc thrcc helds.A total of 600 mgm, of
rerJin four wceks(50 mgm.three
trrtosteroncpropionatswasalsoatJministe

486

R E S I ] A R C H

I N

P H Y S I O P A ' IH O L O C Y

timesa week). Clinically.therervasinrprovenentand the patienttvasable

to rvalkwith thc aid of a back braceund canc upon discharge.
X-ray examination on August 22. 1948. at the conclusionof this period of therapy.
revealedcontinuedspreadof osteolyticlcsions.involvingthe bodiesof the
lower cervical,lorverthoracicand lunrbarvertcbraelnd pelvis.Numerous
lesionswereobservedin the leit ribs and in thc upperthirdsof both femurs.
T h e r c w a s f u r t h c rd e s t r u c t i oonf t h c "l e f r c l u v i c l c(. F i e . l 5 g )

tQ/ 24/48
F t c . 1 6 0 . A n t e r o p o s t c r r ovr i c u ' o f c h c s l i n C ' a s c L . H . t * ' o n r o n t h s a f t c r r t o p p i n g
t e s t o s t e r o n et n d X - r a 1 ' t h e r a p y . s h o w i n g f u r t h e r i n v o l v e m c n t o f m e d i a l r . . r o f l c f t
clavicle.

Aftcr discharge,the patientdid not receivclny further X-ray therapy'

or testosterone.
Thc clinical improvenrentlastedfor crnlya short time. the
back pain and difficultyin walking rrcurring within a few weeks,Radiographic exarninationat the end of October revealedfurther increaseul
t h e p r e v i o u s l y , d e s c r i lbccsdi o n sr.v i t hn e n ' a r c a o
s f i n v o l v e m e n(tF. i g . l 6 i l i
The patient receivedseve.ralinjectionsof an unknown medicationat
homc but her conditionc()ntinucdto grow worsc. Pain was more severe.
The patientwas confinedto
and rcquiredincrcasingirnlountsof narceitics.
bed and there was evidenceof increasingnurve invr:lvement.culminating,

T H I R A P I T U T I C

A P I T R O A C i I T O

( ' A N C L R

4ti7

by the cnd of Novembcr. in paralysisof both lower tinrbs with loss of

sensationbelow the levcl of the ninth thoracic vcrtebrae.There was no
bladderor recta.ldysfunction.
The patientcame under our care in this conditionon December21.
1948. Pain was so sevcrethat a body cast appcaredindicatcd.but the orthopedicconsultantconsideredthe paticnt'sconditiontrxr far acJvanced
to
warrantthis.Thc urinary pH was alkaline.

F t u . l 6 l . L a t e r i r lv i c w o f t h o r a c i cs p i n ei n ( l a s c F - . H .a t t i r r r eo f : r d m i s s i o ns.h o w i n g
e r l e n s i v e m e t a s t a t i ci n v o l v e m e n lo f v c r l e b r a ew i t h c o l l a n r c o f l i f t h a n d n i n t h v e r t e brae.

A h y d r o p e r s u l f i dpcr c p a r a t i o nu ' i t h 0 . 5 f i s u l f u r * a s a d m i n i s t e r e idn

dosesrangingfrom severaldropsby rnouthto I cc. twicc a day intramuscularly. Sodiumthiosuifatein a 109'i aqueoussolutionrvasgiven at thr same
timc by mouth. and aftcr a fcw weeks',t4l/r solutionadministercdintramuscularlywas substitutedand was givcn in rncreasingdoses up to as
m u c h a s l l c c . c v c r y f e w h o u r s .F o r a s h o r t p c r i o d .c o l l o i d a ls u l f u r w a s
a d m i n i s t e r cidn d o s e so f 1 0 0 n r g n r .o r a l l vc v c r yt h r c e h o u r s .
During thc first few il'eeks.rvith small oral dosr-:s
of sulfur in oil and
was
thiosulfatc.
thcre
n()
cvidencc
<lf
inrprroverncnt.2
cc, dclsesof
sodium
sodiumthiosulfateprovideddefinitcpain rclief for u short timc after each
injcction and the gradual increaseof the individual dosc to I I cc, ovsr ii

413tt

R E S E A R C } TI N

PHYSIOPATHOI.-OGY

twelve week period gave complete pain relief. During this time, the patient's generalcondition improved rapidly, Motion and sensationreturned
to the toes,feet and finally to the wholc leg. By April, the patient was able
to sit clutof bed, By the end of May, shc was ambulatorywithout requiring
a braceor cane.Hcr only complaintwas a mild facial acnethat developed
durins the courseof treatment.

F t t ; . l 6 l . , , \ n t er o p o ' t e t i o r v i 6 ' 1 1r,t f l e f t s h t r u l d c r r e g i o n i n f 1 s e F . H . f . u r n r ' n t h r

a f t c r s l o p p i n ! i t e s t ( ) s t e l (n c u n c l X - r a ; - t h e r i r p l ' u t l i r l c o f u r - l r r r i s s i r r r rh, o w i n g i n l o l v c .
ntcnt t:f entire left clin'iclc lnd continucd rplead in lcft scrcnth and c.ighth ribr po.
teriorlv.

Radiographicexlntinationswcrc nradc at thc beginningof treatment

and each month thcreafter.At thc bcginning,four nronthsafter thc last
wcre clbX-ray trcatmentand dosesof testosterone
, osteolyticmctastases
scrvedin all thc vertebraeof the ccrvicalspineand the txrdiesof the lorver
friictureserf thc third.
thoracic and lunrbar vertebrne,w'ith conrprcssir)n
f i f t h a n d n i n t h t h o r a c i cv e r t e b r a cT. ' h ec n t r r cp c l r i s r i l s i n v o l v c di F i g . ^ l 6 J t
and therc was dcstructionthroughoutthe cntirc lengthof thc left clavicle.
in the lcft acromial process,botl hunieri and s*'cral ribs on the left side.
r a r k i n g sw e r er n o d c r a tl ye i n c r c a s c idn b o t h l u n g f i e l d s .
t f i g . 1 6 2 )V a s c u l a m

I'llLRAPlll".itt(

At't,R()iclt

I () l ANCI:R

.lse

F l c ; . I 6 J . , r . n t e r o p o s t e r i ol 'ri c r v o f p e t v i sa n i l u p p c r i , o f h o t h f c t n u r s i n ( ' a s e l . ' . H

a t l r n t c r i f a d n r i s r i o n . h o r . l i n gr r ' i d e s p r c l r(d) \ t c ( ) l t t i cp r()cc\\.

F l o . 1 6 . { . [ - t t e r a l r i c u o f t h o r . c r c : p i n c i n ( ' . r r e l ' l l . f o u r r r . n . h r a f t c r h c g r n n i n gt r f
in ull reltchr;re.
trealment, shetwing considelalrlc bone 1c*a..tr,r.t

490

R F . S E A R C HI N

PHYSIOPATHOLOGY

Comparison with previous films showed definite evidence of continued

spreadof the metastltic process.
Films in Februaryshowedno changc,but in March, slight regressive
changcswerc cbserved.espc'ciallyin the left clavicle, left humerus and
pelvis.During this tinte the urine becameacid and rcmainedso alnrost
continuously.ln April regressivt-'
changeswere observedin the ribs. pelvis,

F ' t c . 1 6 , 5 .A n t e r o p o s l c r i o rv i e r + o f l ef t s h o u l d er r eg i o n i n C l s c E . I l . f o u r n r t i r r t h r
a f t e r b e g i n n i n go f t h er a p y . r l r o * i n g c o n s i d c r a h l cr e p a i r o f l e f t c l a v i c u l a r a n r l r r h
lesions.

fcnrurs and left shouldergirdle. Elscwhcrcnr,rfurther involvenrent\r'as

notcd.
ln July 1949.regrc.ssive
changeswere found to be continuingin all thc
involved
b o n e s(.F - i s s1.6 4 , 1 6 5 ,l 6 6 t
At this timc. all mcdicationrvasdiscontinuedand the patient returnerj
home. [n August. the patient againcomplainedof pain in the back. righr
thigh and left shoulder,and had increasingtroublc walking becauseof
di{liculty in moving the right leg. The urinary pH again was alkaline. Sodium thiosulfateand sulfur in oil again were administeredwith rclief of

t H L RA P l: Lj't l(:

A P P R0AClt

TO

CANC[,R

491

pain and considsrableimprovcmentin the ability to walk, A few months

l&terthe patienthad a strokefrom which shc died i n a few days.

F r c . 1 6 6 . A n t c r t t p o r l e r i otr' i * r ' o f p c l v i sa n d u p p e r 1 r o f h o t h f e n r u r si n ( ' a s c F - . 1 { .

f o u r n r o n t h sa f t c r h e g i n n i n go f t r e a t n r e n t s. h r t w r n gc ( ) n \ i ( l e r ' r h l ch o n e r e p a i r .

HvdrcrnaphtlralencPersulfides
good resultsobtaincdwith ntercaptans
Thc exceptionallv
in trcatingcanccr
in humanscould only partiallybe rcproducedby thc differentagentswith
bivalentsulfur in the polar group we tested.In addirionro the hydropersulfidesof the falty acids,we tricd to prepareand study other groups of
persulfidus.Thc great ability of the products of hydrogcnizationof
( tetralin) and dccahydronaphthalene.such as tctrahydronaphthalcnc
(decalin)
to
fix
peroxides,
oxygenas
naphthalene
led us to try to fix sulfur
on their mr:leculcs.
Thescsubstances
wcre trcatedwith sulfur
as pcrsulfides
i n c o n d i t i o n s i m i l a rt o t h o s cw h i c h l c d t o f i x a t i o no f s u l f u r t o f a t t y a c i d s .
The agentsobtained through this fixation on tetralin wcre particularly
studied.
We havementionedthe pharmacological
studyof theseproducts.Basr-'d
on their biological action. they appearedto bc intcrnrediarybctweenmercaptansand fatty acids pcrsulfides,with the big aclvantageof having a
bearableodor. In humans.we utilizedthis producein casesof rvhich.most
are still under obscrvation.In gencral,the rcsultsobtained were good.

492 /

R E S E A R c Ht N p H y s r o p A T H o L o c y

Subjectivechangesand objectiveones upon the tumors were obtained. We

found that only minimal dosesare to be used,correspondingto micrograms
of sulfur and milligrams of tetralin. With thesedoses,no side effectswere
observed.When given slightlyhigherdoses,some patientsfelt a sensation
of weaknessand also,in a few cases,of dizziness.
This agenthas showna markedinfluenceupon the existingpatternswith
a specialfacility to induce, in most cases,an offbalanceof type D at the
cellularlevel.with higherdoses,the systemicoffbalancewhich is also influenced,can be changedback to offbalancerype A only with difficulty.This
fact indicatesthe need for reduceddoses,actingalmostexclusivelyat the
cellularlevel.The dosagemust be guidedby a continuouscontrol of the
analyses,
and especiallyby thoseof serumpotassiumfor the cellularlevel
and of urinary surfacetcnsionfor the systemic.with valuesabove5 mEq
potassium
i n s e r u m( a n d l o w i n b l o o dr e d c e l l s )a n d u r i n a r ys u r f a c et e n s i o n
of lessthan 68 dynes/cm.,corresponding
to a systemicoffbalanceof thc
t y p e D , t h e d e c r e a soer e v e ns u p p r e s s i oonf t h e m c d i c a t i o ni s i n d i c a t e d .
we utilizedthe agentin canccrcaseswherethe rapid disappearance
of
t h e t u m o r a p p e a r e Ja s t h e c a p i t a la i m . M o r e t i n r e i s n e e d e dt o j u d g e t h e
v a l u eo f t h o s er e s u l t sa l r e a d yo b t a i n e dA
. m o n g t h e c a s c st r e a t e d t, h e f o l l o w i n go b s e r v a t i o rne p r e s e n tasn i n t e r e s t i negx a m p l e .
B. P., 32 year old male, was operatedfor a wart at the right ear Zth
yearsbeforecomingundcrour care.Pathological
examinationhad revealed
malignantmelanoma.A month later, a dissectionof cervicalglands was
made.The analyses
of the obtainedglandsshowedno cancerouscells.Two
monthsbeforecomingunder our care,a checkupreveateda new tumor in
the right side of the larynx. with thc growth of thc tumor, edema of the
right face was also increasing.
Examinationshoweda tumor 4 cm, in diameteron the right sideof the larynx.The tumor was adherentto the skin.
Laryngologicalexaminationshowedthe tumor protrudingin the pyriform
sinus.Becauseextensivesurgerywould have beennecessary
for the tumor
removal,the patientrefusedsuchsurgery,He was admittedand first treated
w i t h o n l y l i m i t e dc h a n g ei n t h e t u m o r . T r e a t m e n w
t a s c h a n g e dt o t e t r a l i n
persulfides
in a doseof l0 drops of a l0olo solutionin oil, administered3
times a day. Under this treatment,the tumor involutedvery rapidly. and
practicallydisappearedin 3 weeks.Subjectivefeeling,such as dizziness,
made us discontinuethe administration
of the preparation.The treatment
was changed to epichlorohydrinand sulfur in the form of fatty acid
hydropersulfides,
administeredin small doses.The patient has continued
this treatmentat home for the past2t/z yearswith no apparentrecurrences.

THl:RAt,EliTtc

Af l'l{()A('l{ 1 (1 (.AN(,tR

491

Butanol, Glycerri
During the periotj when mercaptansand other sulfur-containingagents
were being studied.attentionalso was ccntcredon butanol in the group of
anti-fatty acid agcnts.while butanol'seffecr upon pain and other subjective manifest:rtionsappearedevident from thc beginning of its use, the
inffucnceuFJn tumors seemed.snrall.'fogelher with glycerol, however, it
producedsevcrallong-lastingobjectivcchanges.Characteristically,
in nost
of thesecasesrecurrenccsapgraredonly after ntany yearsof normal activc
life during which therc was no clinical ntanifestation
of cancer.In some
cases,howevcr.therc werc no recurrcnccs.

l - l a . 1 6 7 . f v l y e l o g r a mo f p a t i c n t \ 1 . H . r h o w i n g t h e l l o w o f l i p o i d o l a r r e s r e da t t h e
ievel of {-T.

Mrs. M. H.-This 45-ycar-oldpatient cxperienced,in June 1,943,

scnsoryand motor disturbancrsu'hich progrcsscdso rapidly that in September 1943 she presentcda conrplcteparaplcgiabelow 4-t. she came
undcrour care in February l9rl4. a paraplcgiclor 5r,,i nronths.The myelograrn taken at that time showed ceimpletcobstruction.with the flow of
l i p o i d o la r r c s t e dw i t h i n t h c s p i n a lc a n a la t t h e l c v e lo f t h c 4 - T . ( t ' i g . l b 7 1

.{9.1

R[,SEARCH IN

PHYSIOPATHOLOCY

'l'he

urinc analysesshowed an o{Ibalanceof the type A, and she was

trcated with the acid lipid fraction of hunran placcnta,and with hydropersulfidcs."fhe symptomscontinuedto progressand the pain was more
scvere.Following a clrangein the urine analysestoward the offbalanscD,
the treatmentwaschangedto butanoland glycerol.The pain was controlled
in a ferv days.and a slow regressionof the paraplcgiaoccurred.After four
months of this treatnlcnt,thcrc was completeremissionand the patient
was again anibulatory.Thc ntyelogranrrcpeatcdat thc cnd of Novembcr
t 9 4 4 . s h o r v e dc o m p l c t ed i s a p p e a r a n coef t h c o b s t r u c t i o ni .f i g . / 6 d ) l - h r

I - ' r t ; .l 6 t { . l Ul c l r t g r a r n o f t h c p l t i c n t t r | . H . , r h o i ri n g t h e c o n r p l c l c d i s a p p c a r a n c ct r {
thr obstruction.

piltientrenrairtcdentirclywell, without trcatnrentand frce clf symptomsfor

clcvcnyears.Wc rvercinforniedthat, subsequently.
a recurrenceappearcd.
'fhe
follorvcdlglin by parlplcgia and nrultiplc lunq ntcttstases,
patient
clicdaftcr ftlur rtronthsof paraplt:gia.
l v l r L H . - l n 1 9 3 8 ,r i t t h e a g c o f 3 0 , t h e p a t i e n tu n d e n v e n st u r g e r ) '
for a tuntor of tltc right partttid diagnosedirs chontlronrvxosarcont.i;
in
was
ivas
I9"lt),a recurrent tunror
rcmovedanil
followedthis tinrEbv facisl

Ill[:RAPLUII('AP]'ROACtt

T() CANCER

4q.5

par&lysis.
In 1943.anotherrecurrcncew&strcatedsurgically.ln Junc 1945,
the same procedure was repcated.Immediately after thc last operation,
there was still another recurrenceand the tumor this time started to grow
rapidly. Severepain was only stighrlyrelievedby narcotics.Radiothcrapy
was refused,in spiteof the massivetumor and pain.
Thc patientcanrc under our care in Decernber1945. u,ith scveraltumors occupyingthe right parotid region and cxtcndingbckrw thc nrirndibula. He was using variousnarcoticsr.r,rth
lirtlc cllccr. Wirh the urinary
chlorideretcntionindcx, pH and specificgravityls critcria.trcatnlentwith

l - t t ; . 1 6 9 . P l t t i c n t l . l l . w i t h a r c c u r r c n tc h o n u r o n t v r r l s i i r r ( ) t o
r lfl t h c r i g h t p i r r t l t i r , l
g l a n d b e f o r c ; t n d a f t c r t r c a i n t c n l . - l ' h es c a r i r f r o n r r c p c a t c dp r e v i o r r s u r g i c i r il n t c r ,
v c n ti o n s .

s o d i u mt h i o s u l f a t a
c nd hydropcrsulfidw
e sa s s t a r t c d .t . n d c r t h i s t r c a t n r c n t ,
which lasteda rveck,the lesionappcaredto be unfavorablvinfluenccdand
pain increascd.With the urine chlorideindex usedas u critcrion.the treatnrent was changed.I cc. of butanol 6.,5r;lwas administercdurally threc
timesa day alongwith 0.3 cc. of glyccrol.After 4 da;-s,the doscof butanol
was incrcasedto 2 cc. three timcs a day. Pain was rclisvedin a fcrv days.
Rapid disappearancc
of the tunror masscsfoliowed.The same trcatnrcnt
rvascontinuedfor one year. Sincethen. the paticnt has been u,cll and is
cnjoying good health without any rccurrcnccas of this date. {F'ig. l69 )
E . M . - l n 1 9 3 . 5a, t t h e l g e o f 4 2 , t h i s p a r i e n rh a d a r i g h r r a d i c a ln r u s tectomyfor adenocarcinorna
of the brcast.ln I 940. recurrentnoduiesappearedin the linc of thc scar. One of thesewas bit-rpsied
and showeda
recurrent adenercarcinoma.
Cradc IIl. The pirticnt wAS then treated
with deep X-ray therapy,3800 r, beingdeliveredthroughfive {ieldsto the

496 /

R E , s E A R c Hr N p H y s t o p A T H o L o c y

right chest and axillary regions.A daily dose of 200 r. was given for nincteen days betweenDecenrber1940 and January 19.11,using tbe following
f a c t o r s :2 0 0 k v , . 2 5 m a . , 5 0 c m . , l , i m m . C u l n d I m m . A l f i l t e r . I n
May 1941, the wound arca and recurrentnodules(/.'is,.170) were excised
and a skin graft was used to repair the defect.
In July 1943, skeletalmetastases.
predominantlyosteolytic in nature,
were reported in the fourth, fifth and twelfth thoracic vertabraeand first,
second,fourth and fifth lumbar vertebrae,the first sacral segmentand the
l c f t ; r l l ro f t h e \ i r c r u n t l. l t e r c r r l s r t l . r 'i n r o l l c r n c ntlr f 1 [ g o u t e rp o r t i t t n< , f
t h c l t f t i i i t i t t tr r r l r -t il t e i r r n e rP t r r t i o nt , i t h c r i g h t i l i u n r n e l r t h e s a c r o i i i a c
'l
i o i n t . h c l ; r t r u n lr c c c i r e r rl r : u c o n cul o u r s ct r l d c e p\ , r a l t h c r a p yo v e r t h c
\ p i t ) ci t n d I ) ( ) s t c r i (I ) rc l rr r . t i r r 'l c t ; r ld , r " ch c i n ul g o ( ) r . * i t h t h e s a m cf a c -l
I t r r : . I j o l l c r r i r r tt h r s .h t ' r 'r ] r t ' n r t 'cr , ; l r r c r i . h c | ; 1 i n i n h c r h u c k . w h i c h h a d
c t t n l i n c t ' ll t f t l ' , r l r c t i . x l r r c o t t r r t l c n r l ' r l lr r : l i r - ' r e tlli n r . lr l r c b e c a n l c a n r b u l i l t e r r t ' .
I l r J l r t t r t ; r r r l ( l ' 1I l t o l t r .r . l . p r i r r r l r r r , . r1r ' r u l r c r l . t t l r i t h c p a t i e n t r v a s
; t g l t i t l e , r l t i t n r ' r "kl \ h r ' L Jl t l ( ) s l r r l t h r t t n t c \

n r r ' \ l L t d i L ' \ s h r l 1 | ' e dn f C t i t s { u t i c

t n v o l t ' c t t t e t l tl t t l r r i r c l t c . r l l rc, r r ' r . \ ' l h ( ) r i r a t !\ e l l ( ' l ) l ' ; t c .c \ n c c i x l l v m a r k e t l i n

t i t c i i r t t r t i r -l i t t h . e i r : h l l t . , r t , lt * c l f ' t l r , \ l i t h r l r u r r h . r r c r t r ' h r ; r cr i c r c i n v t r lv c r { .

a=

.t'r'

4lts{t-$.-

#r

=. ..- ELtil

-'

-{ f --i 1rfi- E

-of,:ryi
'
' fuf
,--f',
-t
_-.z _- !

d$$i${

-*.
rn*-hl1*
"
fu$ e**t:l: " "'
*t

*i*;.

*'&

{F
i*.j
wr*"f-;

'':i'
._;ffiSp$
*

tuoI

*.-x,*

l - t c . 1 7 0 . P h o t r . r n r i c r r r g r a p hr r f r e c u r r c n t r L i n n t r t l u l c i n ( ' l r e
c a r c i n o n t a .C r l l l ( 4 0 0 x ) .

[:.\1 .. rhowirrg lrJfntr

T H [ . R A p F . L r T t ( ' A p p R ( ] ^ ( ' H1 ( ] c A N C E R

497

especially the frrst, fourth and lifth bodics and transverseprocesses.De;xlsitswere presentthroughoutthe sacrum,in txlth iliac, the right ischiurn,
left acetabulumand upper fcmurs. Another seriesof deep x-ray treatments rvas uscd on the same areas of thc spinc and pxrsteriorpelvis as
before,the total dose this time being2000 r., usingthc samefactclrs.There

F r c . l 7 l . A n t e r o p o s l e r r o rv i e w s
of thoracic spine in Case E.M.
a l t i m c r . r f a d m i s s i o n ,s h o r v i n g
m o t a s t a t i ci n v o l v e n r e not f v e r t c brae.

F t r ; . I 1 2 . 1 . . ; rrtuel v i c w 6 f t h e
t h o r a c i cs p i n e i n ( a r e E . l v { .a t
tirnc of irtlnrisrion. showing
n r c t a s t i - t t iicn r , o l v c n r e n to f v c r tcbrac.

was againrelief of pain. althoughthe paticntu'askcpt in bed and a body

bracc was applied to rcclucepain associated*'ith ntotion and to avoid
fracture.
We lirst saw thc patienton March 9, 1944.She had nrodcratelyscvere
back pain, was confined to bed with a back brace. and contplained of
weakness.Blood pressurcwas 90 systolic,6tl diastolic.Operativescarsof

4q8

R I " : S t : A R ( : HI N

PllYSl{)PATllOt,ocY

the riglrt chcst region were well healedand therc was no evidenceof local
recurrenccof the tumor. I'here was no superficiallymphadenopa$y.The
liver area was tender to pressureon dcep inspirationbut the liver was not
palpable.There was tcnderncssover most of the vertebraewith pressure.
Patellar rellcxeswere hyperactivebilatcrllly. X-rays showed widespnead
skeletalmctastases
involvingthe dorsaland lumbar vertcbrae,the sacrum
a n d t h c p e l v i s .T h c l c s i o n sw c r e p r c d o m i n a n t l yo s t c o p l a s t i c( F
. igs.l7l,
1 7 2 .I 7 - t)

F r c ; . 1 7 " 1 ,. ' \ n l c r o p o r t c r t t l r i c * r r f p c l r ' i si n ( . ' i r s cI r . f r l. s h r " ri,n' rg g ' i d e s p r e a dn t el a s l a l i r

i n v o l v c n r e n li n l i f t h l u n r h a r v c r t e h r l c , \ i l ! t u n r . h o t h i l r a c u n r J f e n r u r s . l n o m o n t h r
irfler onsct of trcatntctt[.

"l he paticnt u'ashospitalized

and trcatctlexpcrirncntally
fron N,larch!i
to Deccmber5. 194.1,a total of ninc months.The substances
employed
weren-butyl alcohol.cholesteroland glycerin,administered
singlyand conc u r r c n t l yl t d i t l e r c n tp c r i r l d s .n - l l u t y l a l c o l t o lw a s a d n r i n i s t c r eads s l t u rilted witter or salincsolution{"l.9ct ) in dosesof from 3 drops every two
hoursto J0 cc. three tinresa day by mouth.or from I to 5 cc, three tines
a d a y . ? , 5 c i c h o l c s t e r oiln n c u t r a l o i l s o l u t i t l nw a s a d m i n i s t e r e ti ln t r a muscularlyin dosesof I to il cc. thrcc tinrcsa day. (ilycerin was cnrplo.v-eti
or:rlly in 0^5 to I cc. drtscsthrce tintesa day. l)osagcsr-rfall nredication
wcre incrcascdprogressivcly,
tirc airn being to bring about rnd naint:rin
of
the
unnc.
aikalizatron
At the tirrreof adnrission.thc daily urinc pH *'as almost constantly

THER^PEUTIC ^PPRO^CH TO

C^NCER

499

acid. After five months of treatment,the urine became alkaline and remained so until the patient was dischargedfrom the hospital.(FiS. 174)
Clinically, pain was completely relieved after about one month. The
patient's appetite improved and she began to gain weight and to feel
stronger.After five months, she was mobilized with a brace and showed
good progressin the ability to walk.
No changeswere observedin monthly X-ray studiesduring the first
four monthsof treatment.In the X-rays taken in July, the fifth month, at
the time of urinary pH change,a few lesionsbegan to show areasof decreaseddensity.These changesprogressedfairly rapidly thereafter.
zura
aa
co
7a
r2
6C
6'
al
30
56
5l
aC
aa
.,O

Ftc.2
F t c . 1 7 4 . p H o f d a i l y m o r n i n g u r i n e s p e c i m e n si n C a s e E . M . d e t e r m i n e dc o l o r i m e t r i c a l l y ,s h o w i n gc h a n g e sf r o m a c i d t o a l k a l i n e5 m o n t h s a f t e r t h e b e g i n n i n go f t h e
treatment.

Following her dischargefrom the hospital,the patient continuedto

take 0.25 cc. of glycerolthree times a day for another six months. All
medication was discontinuedafter that and the patient has received no
further treatmentsince.She returnedto her old clerical position and after
severalyears, when the X-ray changesappearedto warrant it, the brace
was removed.
X-ray picturestaken fifty-sevenmonths after beginningof the experimentaltreatment,(Figs.175, 176, 177) no longershowedosseouspathology
and indicatedvirtually completerestitutionof normal appearingbone. The
c l i n i c a cl o n d i t i o nw a se x c e l l e nat n d r e m a i n e ds o i n 1 9 5 6 ,w h e n w e s a w h e r .
we were indirectlyinformed that later, in Texas,she suffereda right pleural
effusionand died shortly thereafter.
M. H.-In February 1948, at the age of 18 months,this patient was
admitted to the Good Samaritan Hospital in Dayton, Ohio, becauseof

-500

'

Rr.sEARCt{ tN

pHysl()t,A'rHot.()cy

abdominalpain. An cxplclratoryoperationrt-'vealcd
nn obstructioncaused
by a tumor of the bowel that had spreadto the lymph nodesthroughoutthe
abdomen.A by-passingoperationwils performedto rclieve the obstruction,
A picceof the massand someof thc involvedlymph nodes were removed.
The pathologicaldiagnosiswa.sfibro-sarcoma.

F r c . l 7 - 5 .A n t e r o p o s t e r i o rv i e w s
o f t h o r a c r cs p i n e i n C a s e E . N l.
l i f t l ' - s c l c nn r o n t h si r f l c r o n s e to f
thcrapl', showing almclstcomp l c t c r er t i t u t i o n o f n o r m a l h o n e
5 tr u c tu r c .

Fri;. 176. I-ateral view of thoracic spine in (-ase E.M., fifi1'\cvcn rtronlhs lftcr onsct of
therapl. shou'ing almosl cttnr.
p l e l e r e s l i t r r t i o no f n o r m a l b o n e
\ l r U C t rUC .

On lvtarch3, l94ll, the patientcan)eunder our care.At that time, alnlost a nonth aftcr surgicalintervcntion,a nrassthe sizeclf a tangerinewas
found in the right side of the abdomen.Basedupon her analyses,the patient was trcated with ',a cc, glyccrol and 2 cc. of butanol solution thre.e
times a day. Shc continuedthe samc trcatmcnt without interruptionfor

r H F R A P I T J T I C A P P R ( I A C I I ' I ' ( }( ' A N C T i R

501

two years, gained weig$t and grew, and no recurrenceswere noted. lt is

now | 2 years since this little girl started treatment.She is well, attending
school,and carrying on all the usual activitiesof a child of her age.
Parallclingtheseclinicalinvestigations,
a study was madeof the elimi'fhis
nation of surface-active
substances.
led to lhe use of surfacetension
as a criterionfor the rccognitionof ollballnce pattcrns.A seriesof clser
rvastreatcdaccordingto this criterion.With thc progrsssof rcsearch,nc\e,
agentsalso were utilized.

Fto. 177. Antcroposteriorview of pelvis in case E.M. fifty-scvenmonths after onrel of treatment,
showingalmostcompleterestitution
of bone structrlre.

F'uttt',4 cids
C'otrjugatetl
ln 1947, rve startedtherapcutictrials of conjugatcdfatry acids,lirst
using eleostearicacid, the conjugatcdtrienc obtainedfrom tung oil, ad'fhc
ministeredorally or parr'nterally.
elTcctsupon pain, systemicchanges
and particularlytumor evolution,werenot up to cxpectationalthoughsubjectivcchangeswcre immediatelymorc nranifestthan for unconjugatcd
fattv
acids. [n a short timc, howevcr.it was found neccssaryto continuously
increasethe dosagein order to niaintainthe effects.'I'he intcrventionof a
defensemechanismagainstthese prcparationsoftcn rvas evident. Canccr
patients who had responcledto administrationo[ ulcostearicacid with
relicf of pain and even with an arrest of tuntor grou,th rvcre founcl to
requireincrcasingamountsof this substance.Aftcr a rvhilc.they no longcr

501

RtiSl:AR('l{

lN

Pl{YSl()PAl

ltOl.o(i}

responded.Even very large anrountsof this conjugatcd fatty acid, wcll

tolerated in thesecases,no longer had an eflect upon the tumor and its
manifestations.This fading eflect limited the clinical usefulnessof eler-rstearicacid. In this rcspcct.it appcaredto resemblemany other constituents or even heterogeneous
agentswhich have therapeuticeffcctsthat fade
rapidly.
We have noted previouslythat intcrventionof thc adrenalsis dirccted
especiallyagainstthe conjugatcdtricncs.substanccsrclated to traumatic
noxiousinlluences.Thcrcfore.rvc tried to utilizc eitherconjugatedmemtrcrs.
s o m ew i t h a h i g h c r n u n r b e ro f d o u b l c b o n d s .i n t l r c i r l y r gt h ' ' 1 t h e t r n t l ]
would not bc able to cllicicntlvfight their intervr.rui.n \\'e uhtrrrnctlrr''nj u g a t e df a t t y a c i d s w i t h f o u r d o u b l c b o n d s b y t r , , , t i r r 1rtr r \ l u r c s t . i l ' l r t n '
a c i d sr i c h i n a r a c h i c l o nlicci d . s u c hl s s u l n r o nt , , l l i i r ' r rl r ; r c t r l n ; r t i ntqi r e
m i x t u r c st h r o u g ht h c i r s o l u b i l i t yi n s o l v c n t sc. s 1 , r . , , , ' ,;rr c L l ( ) n c; r. t i o u
temperaturcs.
We also obtaincdthe samctype ol ('(]nrJ)()rrnel
r.lirrctlr'frcrrr
p a r i n a r u ml a u r i n u n tn u c c sa s p a r i n a r i ca c i d .a t r t r . r e t r r r . i t r g "lrct ci dd. L ' r " r n jugatcd pentaenicand hexacnicacids wcrc isolutcrllrorrr thc nri.rturcrrf
conjugatcdfatty acids obtainedfrom salmon,siirrlir'].lnri rlrd liver cil:.
W h c n t h e s cp r c p a r : r t i o nwse r e t r i c d i n p a t i e n t sn i r i r . , r n c c r .n o : r p p a r e n l
i m p r o v c m e not v c r t h e r e s u l t so b t a i n e dr v i t h n r i \ l u i ! . r ' l n r l n - c u n j u g ; t t c r i
firttv acid wls seL'n.
H a v i n g i n n r i n d t h c p l u r a l i t yo f l e v e l sa t * , l r r l r r l r c r r t o u l r . a
l ct. unri
c s p e c i a l l yc o n s i d c r i n gt h e i n i l u c n c ec x e r l t ' db y i r r , " ' l l reih r ' l l r n t h r c n cu 1 : r , ' ' n
c a r c i n o g c n iacc t i v i t y m
. i . r t u r c 'osf f a t t y a c i d sf r o m . , ' t l i i r c r o i l . s ; r r c l i nte. r i i i
a n d f r o m n c t r m aol r q a n sa n d t i s s u c sw e r cc o n j u g , r t r '.(rln r Jr r r c d .W h i i e t h r "
e f f c c t so n p a i n a n d s v s t e n r icch a n g c sw c r c n l o r c i r r t t : r r e. r l r r il r . n g c rl l s t i n g .
t h e c f f e c t .us p o n t u n t o r s\ \ c r e n o t s t r i k i n g l yd i l l c r r . ' nl1r rr n r t h t i s eo b t l i n c r l
w i t h t h c n o n - c t ; n j u l l t c di s o m e r s O
. f a p p r o x i r n i rtir. i i 1 ) c ; r s r r i n r i h i c h
thescconjugatcdfatty acid prcparations
wereused..1..'. 'h,r*ci-lsuhjr.ctir'*
c h a n g e sI .n 2 5 9 t , o b j c c t i v ec h a n g e o
s c c u r r e di,n c l L r J r nel l r n i e ; rdl i s a p p e l r a n c eo f m a l i g n a n t u m o r si n a f c w c a s c s M
. o s t ( | 1 t"l r c ' e r e s u l t sh. n u e r , r r .
w e r e t e m p o r l r y . T ' h e t u n r o r sl l r t e r g r c w a g a i n . , n t i r r r r l r , r i l S { l ' c o u k tl r *
contrtllled
b y a d m i n i s t r a t i oonf t h e s el i p i d s .I n s o n r . ' . . . r \ c t\ h
. c goodresults
p er s i s t c da n d t h c f o l l o w i n gi l l u s t r a t etsh r e eo f t h c s cc a s c s .
B. T.. 46 yelirsolcl, hld a lcft nrastectonty
in 19.18.ft:r an adenoc;irc i n o m a .1 t , : 1 ' ' e a rasf t c r t h e o p c r a t i o np. r o g r c s s i l ' c li rn,c r c a s i n g c n c r a l i z c d
pain appcarcdw
, i t h t h c t c n e r a l c o n d i t i o ng o i n g r l p i d l v d o r v n h i l l .P a i n "
more than the generalconditron,t'rbligcdhcr to hccometotally bedridden.
S u c c e s s i v eX - r a y c x a n r i n r t i o n ss h o w c d r a p i d l y p r o g r c s s i n go s t e o l v ( i c
rr'q[ffi&-

T H T R A P N [ J T I CA F P R O A C H T O C A N C E R

503

lesions.X-ray treatmentfor thrcc rcgionswas startcdwith the intentionto

control the pain which was most severein skull, ribs. spine. pelvis and
femurs.Bccauseof thc gencralcondition,this was discontinuedafter a few
treatments.Whcn the paticntcameunderour carcrshe rvasentirclyimniopain, X-rays revcalcd(1.i1.s.
bilizedand in scvcrL.
l7S, 179) nrultiplcosteolytic metastases
in skull, fcnrur.pclvic boncs,spinc.ln viciv of thc analysis

F r c . 1 7 8 . l - a t e r a l v i e w o f t h e s k u t l o f p a t i e n t ( t l . . f , ) u t t h e t i n r c o f a d n r i s s i o nr, h o w i n g m u l t i p l e o r t e o l y t i cm e t a s t & $ e s .

504

R r . s l - t A R C l t{ N

pHvstopAlnot.oc't'

as a lypical A oflbalancc,trertmcnt with hydropersulfidcand conjugated

f a t t ya c i d sw a s i n s t i t u t c d .
In her casc tve saw it pcculiarfornr of resp(lnscencounteredalso in
scvcralothcr subjccls,und rvhichw'econsidcrcdin gcncralas corrcsponding
to as r iuvorabrlc
responsc.During thc trcatment,whilc pain in generalwas
relieved,one lesion was seento becomeprogressivelymore painful. lt re-I"he
nrained.scvcrely
painful for 2-3 davsaftcr which the pain disappeared.
samcchangewils scento occur successively
in one lesionafter another with
tlrc sanrctemporory increascof pain until it becanrcvery scvere.followecl
h i r l i . l i l r l r e i r n l nt cr lel c rl - . 1 t l , r r ' \ L r t o n l t ' t l i i lt i r c p r u i rf r. r i lt o r c t u r ni n t h e
r ; t t l l l l r : t i o t l . l r t l l t l \ l . l , r l l f t h . : I r - : r i i r 1\ \1i l \ \ t ' e l t l 6 i n r , t r l U l t l t f t C r s U C l ta C h a n g * "
\ \ ' j t l r t l t i r [ 1 1 1 r' .)1l t t f . t t l l f t t l i h e p ; 1 1 i r ' 1pt l . r d c . t \ f r \ r l r p i { r e C O v e r ya n d
' , \ , t \( r l i l r r J f r , ' . 1i t l l r ' ' . l l l r l t i \ \ ( \ n r r , r r l h ' . I l r t r : r t l i r ' l . ' r g i u cl rl lt i r t t g c $ .
illthou$h
rittrlllllg iril,il!\\t\f
I ' r { r . 1 1 1 ' r ri l t r . i t r l t l l r t i r
ld.r,rnr, trrtrk nl(}fC tinft ttl
l r t . , r t l l l r l r ' i . i l I t : , l r ] ' l ] , " , \ r i l i t ' l i l . r l i r r l r r l t l r c I ' r r r 1 cl p t t i S 1 A S C iSn p f O g -l'lfC
t ( ' \ \ . \ \ l r i i t ' I i : :. I \ { ) , 1 1 1 ,i .S1 l 1 i t , ' r . J \ 1 l t \ r l l t , r ', l l r 1 1 1 r r: l \ c l t f \ .
patient

I-rc. 179. .,\ntcroposterior

of thc pelvisand upper parl\ of the fenrurs of the
"'rew
p a t i e n ({ 8 . 1 . ) r h o w i n gm u l t i p l co s t e o l y t i cm c t & s t r s c s .

T H E R A p E L T T T cA F p R O A C H T O C A N C E R

505

resumednormal life for 3 yearsafter rvhichrccurrcncesappearedon skin.

lung and liver. Theserespondcdlessfavorablvto thc sametrcatment.The
patientleft our care and died a fervnonths latcr.
M r s . S . T . , 4 7 y e a r so l d . c a m cu n d e ro u r c a r c i n a s u b c o m a t c l us st a t c ,
2t/z yearsafter a lcft brcastnrastectomyfor an adcnocarcinoma.For thrce
monthsbefore.the patientcomplaincdof generalized
pains and especially
of severeheadaches.
and for a month had synrptomsof diabetesinsipidus.

Frc. ltt0. I-atcral view of the skull of paticnt (8.T.) aftcr ll

M o s t o f t h e l e s i o n sh a v e d i s a p p c a r e d .

r n o n t h so f l r c a t r n e n l .

506

RTsEARCH rN pHvsrclpArHot.0fir,

An X-ray examinationof thc skull, madeprior to hcr admission{f ig. 1fi.]}

showedcxtcnsiveskull metastases
with an advanceddeslructionof the
clinoid bones.
Becauscof the diabctesinsipidus,thc urinc analyscscould nol furnish
thc neededindicationfor thc trcatmcntand wc recurrcd.therefore.to the
numbcr of blood lcucocytesand to the body tempcrature.as tests able ta

l : : t t ; . l f { I . , . \ r t t c r o p c r rrlieo r v i c r v o f t h e p el l i r a n t l f e n r r r r st r f p a l i e n t { l l . - f . ) t f l e r f o u r
n r o n l h r o f t r e u l n r c n t .r h e i r v i n qt h e l e l i c r n sd e c r c l r i n g ,

T H L R A P L L ' T I C

A [ tP R O A C H

T O

C A N C E R

507

indicate the existing offbalance

. With 14.500 lcucocytesand a c$nstant
temperatureof above 98.6'F, wc considercdthe oflbaliinccto bc of thc
type A and administeredconjugatedfatty acids obtainedfrom cod liver
oil. and sodium thiosulfatetocctherwith posteriorpituitary hormone for
hcr diabctcs insipidus.Probablyduc largclyalso to hcr clectrolyticbalancc
the patient regainedconsciousness
und madc I rapid recovery.In lessthan

F r c ; . l 8 ? . . A n t e r o p o s t e r r ovr i e w o f t h e p c l v i s a n d f c n r u r so f p ; r t i c n t{ 8 . ' l ' . ) 2 1 . . e a r r i

irrctl.
l a t er . s h o w i n g m ( ) s to f t h e l c \ i ( ) n :d i r i r p p e

two wceks shc was out of bed and resumccla nornral life. She continusd
with the same treatmcnton an ambulatorybasis.An X-ray cxanination
four monthslater showeda manifesthealingof thc previous lesions.ff i.rt.
I 84)
After anothcr four nreinthsholcvcr. without ;lnv recurrcnceof her
malignancythe diabctesinsipiduscriuld not bc adcquutcll'conlrollcd.Shc
r e f u s e dh o s p i t a l i z a t i oann d l e i t o u r c a r c .S h e d i c c lu s i r o r tt i n r c l a t c r u ' i t l r
symptomsof clcctrolyticoffbalancc.
M r . L . N . , 6 . 1y e a r so l d , h a d r l u n g h i s t o r vo f v c s i c a lt r o u b l e s w
. ith

508 I

rrEsEARcH
tN prrysropATlror.ocv

biopsiesshorvingcanccrouslcsions.In spitc of repeatedfulgurations,tht:

vesicaltunrorsgrew rapidly with constanthcnraturiaand tenesmus.Threc
monthsbefore coming undcr our care, thc patient sufleretJseverepains in
the left groin which X-ray examinationshowcd to be due to a bcnc
m e t a s t a t ilcc s i o n .F i g . 1 8 5 d c p i c t sr h c l c s i o n su p o n a d m i s s i o nT. h e a n a l yses showedan offbalancctypc A, and a rrcatnlcnt with conjugatedfatry
;rcidobtainedfrom cod liver oil and sodiunrthiosulfatewas institutcd.The
pain disappeared
in a few days.as did the hcnraturiaand dysuria.The paticnt continued to improve. An X-ray examination.four months after
t r c . t t l l l l n l . ' lt t r r l ; t r t u t l . s l t , r i ic r l t l r c ; t i r p c l l r i t n c c o l ; r c : r l l u s l t t h c p l a c C r . l f
l h c b f t n c I l t d l l t s t i i s c \ .( l : t i : 1 6 , \l | [ 1 g p l r l i c n t c r r n l i n u c d t l r c t r * a t m e n t f $ r : r
l c q t n o r c r t t o t t l l . tt:r f l e r r i l l i e h t n t ) { rr r e l o r i l r ' . r e Lo l } t i n t
I / | t t'l't tlt'tlt'rltil

. ' 1g r ' t t l r

I J u u . t L t r rel l l t t ( ) l { l t n r \ n } ' r\ l ( ' l e r r r r . r r i r i n st }t r c l " r t t r : l c i d sr v i t h * h i c h i t

c ( l l l t c \ t t t c o r l l . r c l L r n t l e tI r { } r . n r ' r: rl n t l . r [ r n o r r ] r l rul , r n r - l i t t o n sn,l o r g h g t e r t r .

,'$
t

#;:
'*,

F t c , 1 8 3 . L a t e r a l v i e w o f t h e skull of patient (S..I'.) shor.r'ingmultiple ostcoly'ric

procetses.

T H E R A p E U T T CA p p R O A C HT o c A N C E R

509

geneousagent$were sought,.We first resorted to the alpha hydroxy fatty

acids. lsolaled membersand mixtures of the acids. after being prepared
and testedfor toxicity in animals,were usedin patients.we have discussed
previouslythc striking and speciliceffectobtainedupon l;-mphosarcomain
cxH mice with alpha hydroxycaprylicacid. Although ivc werc not ablc to
accountfor this effect.we did attemptto dcterminewhetherthe acid would
have a similar favorable influence upon human lymphomas, especially

F r c . 1 8 4 . l - a t e r a l v i e w o f t h e s k r i l l o f p a t i c n t { S . T . ) s h o w i n g m o s l o f t b c l e r i o n sd i s appeared.

5t0

R L S E A R C H I N

P H Y S I O P A T H O L O G Y

Horlgkins'diseasc.Scvcralpatientswith Hodgkins'diseasewere trcated by

oral administrationof preparations
of alpha hydroxy fatty acids from caproic to stcaric acids,or with mixturesof them. Only in a few subjects
were very limitcd effects.such as a small decrcascin thc lesions.obscrved.
changes were not considcredto excc'edthosc known to occur spontaneously in such cascs.Thesc effectswerc inferior to those obtained with thc

l 1 1 , .I h i

' \ t l i r ' t , , p \ ! e ' r ' r r 1 r\ rt r ' r , ., t l r i r r t

i . t . l r i . t r 1 ; r I , ; ' l r c r i ! r r t i l t t L n , i r l g n r r c ; t f u i n n f t i ltr' l

t h e b l a d d e r .s h o w i n g t h e d e s t r u c t i r r no f r h e l c f t i s c h i o n h o n e .

fatty acid prcparationspreviousl)-testcd.'l'herc\rcrc no efTcctsupon evol u t i o n o f o t h e r l y m p h o m a so r i n o t h e r t y p c s o f c i r n c e r .A l p h a h y d r o x y

preparationsproduced limitcd subjcctivcchangcsin lcss than l0qi nf
c a n c e rp a t i c n t sa, n d n o m a r k e do b j e c t i v ec h a n g c sa t a l l .
c)ther hctcrogeneous
fatty aciclpreparationswcrc obtainedand" aftcr
study of their pharnracological
activitics.werc appiied in humans. Polyhydroxy frtty acids, pcroxidesof fatty acids. and fatty acids in which
chlorincwas fixed at thc doublebonds.wcrc rcstedin only a limited nunrber of paticnts,but cnough cascsto show that effectswerc no dilTerent
from thoscobtaint:dwith conjugatedfarty acids,for instancc.

T H F . R A P I L I T I C

A P P R ( ) A C I I T ( )

C A N C E R

5il

Parallel to thcse eflorts to lind new agentsfor patients with the one
typc of offbalance,othcr agentsfor usc againstthe opposite type of ofTbalancewerc investigated.
Sterolsand nonsaponifiable
fractionswere treatcd in variousways to
obtain heterogeneous
substanccs
not founelin living organisms,Wc used
heat at 300'C or ultravioletlighr, accordingro thc prcrcedures
employcd
by Roffcl.in ordcr to inducc changcsin sterolswhich would tcrnd.accord-

Flc. 186. Anteroptlttericlr view of the pelris of patient of Fig. lll.\. d rneinthr luter.
showing the healing of thc lesion.

ing to him ( l9ti) to makc them carcinogenic.

We considcredtheseheat or
ultraviolct trcated stcrols and nonsaponifiable
fractionsto rcprcsentab,
"l'hey
normal lipoids.
rvcreadministcrcdin oily solutionsto animalsand to
rvith
advanccdmalignancics.
a few patients
lnjectedintrlnruscularlydaily
for a few wceks,theyinducedno undcsirable
rcsults.T'hcirctlectsupon pain
and systemicchangeswere not signilicantlydifferent front those of corrcspondinguntreatedpreparations."l'hc studicsof othcr changesin canccr
patientstreatedwith thcse prcparatir>ns
do not allow iinl' conclusionsto
be drawn at this tirne.

512

x E S E A R C Hr N p H y s t o p A T H o L o c y

We prepared and studied various sulfurizedfatty acids, of which the

conjugatedwere used on a broad scale in therapeutictrials. Clinically.
thesesulfurizedfatty acids producedmarked resultsin severalcases,but
did not constitutea significantadvanceover sulfurizedoil in which sulfur
was bound to triglyceridesrather than to free fatty acids.
In the searchfor more activethiolipoids,we preparedseveralproducts,
one of which, methylthioglycolate,
was given a broad clinical trial. Although
it producedsome interestingobjectiveresults,they were neither sufficiently
intensenor consistentlyreproduciblenor persistentenoughto make methylthioglycolatea distinct advanceover the other sulfur preparationsused.
The compoundalso had the disadvantage
of disagreeableodor although it
is not as obnoxiousas the mercaptans.As a result, we abandonedthe use
of this substanceafter a year of clinical experiments.
of l3l patientstreatedwith methylthioglycolate,
39----or30zo showed
subjective
changesand l9----orl5vo, alsoshowedobjectivepositivechanges.
Another synthetic thiolipoid, hexylthionic acid, was utilized in a few
clinical cases.Only a small number of patientsshowed objective clinical
resultsand thesewere neither consistentnor persistentenough to warrant
using this substancefor further research.While the resultsobtained with
agentsother than mercaptanshaving a thiol polar group were interesting,
thesecompoundshad too little influenceupon tumors, especiallyin cases
where the pattern indicatedpersistentpredominanceof sterols.
Selenium Preparutiors
we have previously noted the considerationswhich led us to study
lipoidic compoundscontainingbivalentselenium.The compound used in
clinical researchwas hexyldiselenide,
a lipoid with an -Se-Se-as a polar
group. Oily solutionsin variousconcentrationswere given by subcuteneous
or intramuscularinjection. Doses as low as 4 microgramsor as high as
400 milligramswere employedseveraltimes a day. For oral administration,
capsulescontainingthe product in solutionin hydrogenatedoil in amounts
from 4 microgramsto 100 milligramswere employed.A short time before
the experimentaltherapeuticuse of selenium compounds,the means of
recognizingthe existhg oftbalancewere implementedwith the sulfhydryl
index, a measureof urinary eliminationof the sulfhydryl group.
Despite negativeresultsin animal tumors, hexyldiserenide
was used
clinically in the hope that, with treatmentguided by the data furnished by
urinalyses,satisfactoryresults could be obtained. The sulfhydryl index
served,at the beginningof this study as the principal indication for the
administrationof the seleniumcompound. Later we used urinary surface

T H E R A p E U T T cA p p R o ^ c H T o c A N c E R

513

tensionas the criterion, and lately we have usedthe changesin serum and
total blood pntassium.In general,we administeredthe medicationonly if
the sulfhydryl index was below 1.5, the surface tension above 68. A
marked influenceupon the tumor itself was seen in a relatively high proportion of cases.The resultsalso are of theoreticalinterestsince the compound had lessinfluenceupon symptomsat the tissuelevel,such as pain,
or at the systemiclevel,and more on thoseat the cellularlevel.The effect
on pain was slow to appear,often requiringdays.But once relicf of pain
was achieved,it persistedfor a long time, in contrastto the brief effect
p r o d u c e db y o t h e r a g e n t ss u c h a s t h i o s u l f a t eo r h y d r o p e r s u l f i d casc t i n g
d i r e c t l y a t t h e t i s s u el e v e l . P r i m a r i l yb e c a u s eh e x y l d i s e l e n i dacl t e r s t h e
patternpresentat the cellularlevel,it must bc emphasized
that determining
proper dosageat least in the beginning,appearedmorc difficult than for
any other substance
with which we have had experience.
A t t h e b e g i n n i n gw
, e u s e dd o s e si n t h c r a n g eo f l 0 - t t 0 m g . b u t i n a
numberof casesa pcrsistentchangeto the oppositcpatternoccurredafter
o n e o r t w o d o s e sT
. h i s l e d t o t h e u t i l i z a t i o no f s m a l l e ra n d s m a l l e rd o s e s
in order to avoid too rapid changcto the oppositepattern.Also with small
d o s e sw
, e h o p e dt o l i m i t t h e t h c r a p e u t ircc s p o n s teo t h e c e l l u l a rl e v e lo n l y .
w e d e c r e a s ctdh e d a i l y d o s c t o 5 m g . , t h e n t o I m g . a n d e v e n t u a l l ye v e n
t o m i c r o g r a m sW
. i t h t h e s es m a l l a m o u n t s .t h e i m m e d i a t ec l i n i c a le f l e c t s
s e e m e dt o b e a l m o s te n t i r e l yl i m i t e dt o t h e c c l l u l a rl e v c l .T h e c h a n g e si n
pain and the systenric
patternwereminimal.The choiccof doseto be given
w a sd e t e r m i n c idn t h e s e c o n dp a r t o f t h i s r e s e a r c p
h r i m a r i l yb y u r i n a r ys u r face tcnsion.It was observedthat microgramdosesmay influencelesions
without changingthe valuesof the other analysescorrespondingto the
systemiclevel.
The clinicalresultsobtainedwith the useof hexyldiselenide
in humans
warrantsdetailedconsideration.
Cood resultswere obtainedwith greater
consistencyafter the problem of dosagewas resolvedand the relationship
to urinary analyseswas established.
Important objectivechangescould be
achievedby using urinary analysesas a guidefor dosage.as illustratedby
the following cases.
A . 8 . , 6 8 y e a r so l d , m a l e .I n O c t o b e r1 9 4 9 .t h e p a t i e n th a d a n a m p u tation of the left leg at the hip-joint performedat the Memorial Hospital
becauseof a tumor of the femur. The pathologicaldiagnosiswas sarcoma.
H e w a s w e l l u n t i l J u l y 1 9 5 5 w h e n h e b e g a nt o c o u g ha n d h a v e r e c u r r e n t
e p i s o d eo
s f h e m o p t y s i sX. - r a y e x a n t i n a t i o rne v e a l e dr i g h t p l c u r a le f f u s i o n
a n d i n f i l t r a t i o no f t h e r i g h t l o w c r l o b e . H e w a s r e u d n r i t t e d
to Memorial
Hospital in october 1955 and thoracentesis
revealedbloody fluid. Bron-

514

./

n r , s E A R C Ht N p n y s r o p A r H o l , o c y

chr:scopy
showcdpartialnarrowingof the right lower lobe bronchus.Examinatiein
of pleuralfluid and bronchialwashingby the Papanicolau
methodshowedceltsthat wcre suspicious
for mirlignantdisease.but not
canclusivc.
It couldnertbe dctermincd
rvhether
a primarylung canceror
metastatic
silrconta
wasprescnt.An exploratory
thoracotonly
wasadviscd
but refuscdby the patient.who signcdhimsclfout.

2-5-s5

f r t t ; . l l { 7 . r \ n l e r o p o s t c r i r t rr ' r c u r r f c h c r t r n ( ' ; r s c { . 4 . 8 . ) a t t i n r c o f a t l n r i r s i o n s h o * . i n s
l h c p r e r c n u : co f l r m a s r i n t h e k r w e r p a r t o f t h e r i g h t h c n r i r h o r u x .

Two g'eekslatcr. hc canrcundcr our care and has beenan anrbulatory

paticntsincc thcn" His chestpain had bcen considcrablyrelicvedrr'ith the
thorlccntcsisand thcrc wits no blccding.He conrplaincdof distress:rncl
t i g h t n e s isn t h e c h c s ti l s r v e l la s p c r s p i r a t i oant n i g h t .A p h y s i c a e
l xaminatton rcvclrleddullncssin thc'lorvcrrirht hcnrithorirxbeh-rwthc scvcnthrib.
[Jlooclple\surc *'ls 2(]fi 165. X-rly cxarninationrr-ve
aled a large round
occupying
mass
thc riqht lorverlobc. 1/jiq.187/
Lrinalysesshowedu low surfucetcnsion,low specificgravity, high pH
a n d l o r v s u l f h y d r v lc x c r c t i o n .H c x y l d i s e l e n i d e ' , v nc sh o s c n a s t h e o n l !
chcnrothcrapeutic
agcnt."l-rc:ltment
was startedwith a tenth of a rnilligranr

THf-RAPI:-t;TI(

4t'PRt)4(lll

t'() ('AN(:hR

5r5

twice a day. and was increascdgrlduirlly to half a milligram twicc a duy.

C h e s tp a i n a n d h e m o p t v s idse c r r ; r s c idn t h c f o l l o r v i n {u c c k s . ' l ' h eg e n e r a l
conditionimproved.Hc fclt bcttcr and had no cough.X-rays il month and
a half later showcdthat the right lowcr lobc massrvasslightly snrallerin
size.
I n J a n u a r y1 9 5 6 ,a s t r i k i n gr e d u c t i o ni n s i z ca n c ld c n s i t yo f t h e l o w e r
right lobc masswas seen.lt wASnow abr.lut60{,i as big as a month beforc.

,.*.."
*Wrt

.,,.rsqffiF*

".*n
,r'

a - 2 6- 5 6

iYR*.

,$
rs:
';*

F t c . 1 8 8 . A n t e r o p o \ t e r i o rv i e w o f t h c c h u r t o f t h e p a t i e n t . { . 8 . a f t e r t r . n m o n t h s
o f t r e a t m e n tw i t h h c x v h l i s c l e n i d c .

At the beginningof Februarv.thc nrassin the right lrls'cr lobe was alnrost
entirelygonc. The intcrlobularthickcningof thc left ktucr lung still pers i s t e dl.n A p r i l . f u r t h e rc l e a r i n go l t h c r i g h tl c l w c rl o b c w a ss s c n .l f i S . 1 8 8 /
for a fel morc nronths.
The patientreceivcdtreatnlcntwith hcxyldisclenidc
H e c o n t i n u e di n e x c e l l e nct o n d i t i o n* i t h o u t f u r t h c rt r e a t m c n ft o r t h e n c " t
three years.At thc cnd of that tinrc. thcrc tus il recurrcnceof pain and
thc massin thc right lowcr lobe rcappearcdlnd grcw rlpidly. Trsatment
was rcsumcd.Thc pain disuppearcdagain within a
with hcxyldiselenide
shorttime. and thc tumor againregrcsscd.

516

x E S E , A R c Ht N p H y s l o p A T H o L o c y

I. A.-This 58-year-oldpatient had diabetesbeginningat the age of

30. tt was controlledby insulin. At 35, he had rheumaticfever and remainedin bed for 6 months.Angina developedat 53. ln January 1956, he
was awakened by a pain in the right sideof the abdomen,The pain continued to be severefor months and the patient lost weight. Barium enema
revealednothing. He entered Jewish Hospital in Brooklyn with jaundice
in April 1956. Upon operation, the gall bladder was found enlarged to
twice its normal size and the common duct was dilated. With a finger insertedthrough the foramen of Winslow, a hard, stony mass involving the
head, body and most of the tail of the pancreas,was felt. The tumor involved almost the entire pancreas.The lymph node of the common duct
was enlarged.The liver showedno evidenceof metastases.
The generalcondition of the patient, in spite of the extensiveinvolvementof almost the
entire pancreas,did not justify a total pancreatectomy.
Therefore,a palliative surgicalprocedurewas done. The gall bladder was anastamosedto
the jejunum.
The post-operativecourse was good. No other treatment was prescribed.When the patientcame under our care at the end of May 1956.
the jaundicehad disappeared
but the pain was the same as it had been
before operation. A mass occupyingthe upper abdomen was felt. Treatment was startedwith a dose of 30 microgramsof hexyldiselenide
a day.
This dose was increasedlater to 300 microgramsa day. The abdominal
pain in the upper abdomendisappeared.
discomfortlessened;
The patient
remainedon this treatmentuntil 1957 when epichlorohydrinwas added to
the hexyldiselenide.
With the mixed treatment,the patient continued to
improve and the tumor, which had been palpablein the upper aMomen,
disappeared
after a few months.Exccpt for diarrhea,which occurs from
time to time, the patienthas madea very good recovery.He has continued
and epichlorohydrinand, at present,four years later, is
on hexyldiselenide
in goodgeneralcondition.No tumor can be felt and he is at his regularjob.
C. M., 52 yearsold, male. ln 1946,the left breastof this patient was
removedbecauseof a malignanttumor. ln 1947, the right breast was removed but the tumor proved to be benign.In 1954, the patient,showing
blood in the urine, was admittedto the PresbyterianMedical Center where
In February 1955, the patientundera right kidney tumor was diagnosed.
went surgery. A tumor of the right kidney with extensivelymph node
involvementabove and below the renal vesselswas found. A simple nephrectomywas performed,the lymph nodesbeing consideredinoperable.Tbe
pathologicalreport revealed"clear cell carcinomaof the kidney with ex-

T H E R A p E U T t CA p p R O ^ C H T o

C A N C E R

5 1 7

tcnsioninto the renal vesselsand surroundingtissue."There was no evidenceof pulmonaryor other metastases.
The patient did not undergo any treatment at this time and felt well
until September1955, when he experiencedincreasedfatigue,flatulence,
slight pain in the right kidney region,and some abdominalpressure.No
abdominal mass or lymph node involvementwas found. He was treated
with hexyldiselenide,
starting with 200 microgramsa day. This dose was
increasedprogressivelyuntil it reachedZVz mrlligramsa day.
Under this treatment, most of the patient's pain disappearedand no
palpablemasscould be found. The patientcontinuedtreatment,with the
samedose,until October 1957,when epichlorohydrinwas added.At first,
with the new rnedication,the patient was more tired and there was a noticeablcincreasein perspiration.However,after a short time, hc continued
to improve.At present,he is still taking l5 milligramsof epichlorohydrin,
and 100 microgramsof hexyldiselenide
daily. He feelswell, continuesnorwork,
palpable
mal
and no tumor is
now four and a half years since the
beginningof the treatment.
W. H.-ln March 1954,at the ageof I l, this boy had the first of three
brain operationsat the JerseyCity MedicalCenter.He had complainedof
persistentheadaches
for about a year.A brain tumor was removedand on
pathologicalexamination,was first thoughtto be benign,but later proved
t o b e m a l i g n a n t( s p o n g i o - b l a s t o m aB) .y A u g u s to f 1 9 5 4 , h i s s y m p t o m s
recurredand a swellingappearedin the area of the scar. At the second
operation,it was possibleto removeonly a part of the rccurrenttumor. In
November 1954, local swellingand headaches
returned.X-ray treatments
failed to give relief and a third operationwas performed,but only a piece
of tumor was removed.lt showedthe sanrepathology.
H e c a m eu n d c ro u r c a r eM a y 3 , 1 9 5 5 ,a n d w a st r e a t e db, e c a u s e
of low
with
hexyldiselenide,
sulfhydrylurinary indcx,
in dosagesrangingfrom 300
-l'his
microgramsto I milligrampcr day.
treatmentwas continuedfor one
year. He has done rcmarkablywell since.He has had no headaches,is
more alert and lessdrowsythan at any time sincehe first becameill, There
has been no evidenceof recurrence,Since treatnrcnt,there has been a
definitereductionin involuntarymovementsof his head, extremitiesand
body, which appearto havebeen relatedto the tumor growth. There have
to date,five yearssincethe beginningof
beenno abnormalmanifestations
the treatment.The patient goes to school and engagesin all the activities
of a normal boy of his age,
Mrs. A. L.-This patientfirst noteda lump in her neck in the summer
of 1953 when she was 26 years old. In February 1954, a secondlump

518

R E s E A R c Hr N p H y s r o p ^ T H o L o c y

appeared.Surgerywas performedat the Ottawa Civic Hospital.The pathologicalexaminationshoweda cancerof the thyroid (papillary adenocarcinoma). It involvedboth sidesof the glandand many of the lymph nodes.
The entire thyroid gJandwas removedand bilaterallymph node dissection
was done in two stages,Sincethe surgeonfelt that he had not rcmoved
all the affectedareas,the patient receiveddeep neck X-ray therapy following the operation.However,by July 1954, a new massdevelopedon the
right side of the neck and this was removed in November.Thc pathologicalexaminationproving it to be the same type of tumor.
T h e p a t i e n tw a s f i r s t s e e nb y u s o n J a n u a r y1 0 , 1 9 5 5 .A l t h o u g hi t w a s
only two months since the last operation,there were severalrecurrent
tumor massesin the lateralright side of the back of the neck, as well as
infiltrationsinto the area of the last operativewound. The patient was
treatedwith hexyldiselenide,
in dosesrangingfrom 300 microgramsto I
milligramdaily. Under this treatment,the massesprogressively
decreased
and, after 3 months,disappeared.
Shecontinuedthe treatmcntfor another
six months.Now, after 5 t/z years,she is feelingwell. has had no recurrcnces,and is carryingon her usual activitiesas a housewifeand mother.
Hexyldiselenide,
althoughit producesimpressiveresults,very often is
not of itself able to provide enduringbenefit.Many other patients have
had recurrences,
some in spiteof impressivefirst resultsand continuation
o f t r e a t m e na
t s s h o w ni n t h e f o l l o w i n go b s e r v a t i o nJ.. D . , l 0 y e a r s o l d ,
cameundcr our care in a preterminalstate,aftcr an exploratorylaparotomy
revealingan extensivecarcinomaof the liver. (Fig. 189) The responseto
hexyldeselenide
treatmentwas impressive
with the patientmaking a perfect
rccovery.His livcr which had filled the abdomen,returnedto normal dim e n s i o n sT. h e p a t i c n tc o n t i n u e da n o r m a ll i f c f o r 2 y e a n sw h c n , i n s p i t e
of the continuationof the treatment,gcneralizedrecurrencesappcared.
Thesccould no longerbe controlled.
Tetralin Perselenide
The good cffectsobtained with persulfideson one hand, and with
seleniumon the other, have led us to investigate
the correspondingcomp o u n d - t e t r a l i n ep e r s e l c n i d cw.i t h i t s l o w t o x i c i t y ,t h c c o m p o u n dw a s a d m i n i s t e r c dt o h u m a n si n w h i c h t h e d e s t r u c t i o o
n f t h e t u m o r a p p e a r e dt h c
immediateaim. The cffectupon pain was good althoughnot immediate,thc
s a m ea s f o r t h e s y s t e m i cl e v e l .I t w a s a t t h e c e l l u l a rl e v e l sw h e r e t h e s e
were the most manifcst.Researchwith this agent is still in progressand
for the momentthe influenceexertedupon the tumorsseemsto be very fa-

.IH[:RAPEU'TIC
APPR0^CII TO

CANCER

519

vorable.Similargood resultswerc obtainedwith naphthalene

perselenide
andothersimilarpreparations
of aromatichydrocarbons.
Parallelto studieswith syntheticncgarivelipoids, syntheticposirive
lipoids wcre investigated.Thc problem was quitc diflercnt becauseof the
fundamentaldiflercnccsin biologicalroles o[ the two antagonisticgroups.
Arnongnegativelipoidswe singledout iln etlectagainststerolpredominance
and a destructivcactivity through thc inducrionof rapid ccllular aging.

F r c . 1 8 9 . P h o t o m i c r o g r a p ho f a l i v er h i o p s yo f p a t i c n tJ . D .

lipoids.wc soughtagentsablc to correct the noxious

Anrong the positivr,effcctsinducedhy futty acirjs.As notcd prcviously,this lcd to a searchin
one group of expcrintents
for substances
with highcrspecificityfor binding
particularfatty acids and in other expcrimcntsfor substances
with broad
spctrumactingagainstacid lipids in general.We havediscussed
previously
charactcristics
the pharmacodynamic
of many of thesesyntheticpositivelipoids.Clinical resultshave improvedwith thc dcvelopmentnot only of new
substanccsbut also of new meansof recognizingthe pattern presentand
of following the changesthat take place. Wc have mcnt^ionedthe results
-rd&ltr-r

520

xESEARcHrN pHysropATHoLocy

obtainedwith butanol,the fint of this group of syntheticlipids with positive

characterto be usedin humans.
Butanol was unableby itselfto influencethe growth of tumors,although
it was effectivein controlling pain of an alkaline pattern. This beneficial
eftect upon pain was still more manifestfor other aliphatic alcohols, and
especiallyfor heptanol. In high doses,however, heptanol probably acs
beyond the tissue level. Besidesinfluencingpain, it inducessevereedema
and changesin the evolutionof the tumor. It has little influenceat the
systemiclevel upon hemorrhage,even with high doses.For tumors with
offbalanceof type D, a mixed treatmentappearedindicated.The use of
heptanol-butanol,
howevcr,was not fully satisfactoryalthoughit has been
employedin somecaseswith goodclinicalresults.
During thc progressof this research,it becameincreasinglyimportant
to have an accurateknowledgeof the existingoffbalance,and of the adequate use of the availabletherapeuticagents.The following observations
showhow thc rcsultsobtainedby the treatmcntare a functionof the correct
applicationof this conceptof guidedchemotherapy.
M. S.-This 52 year old woman startedto lose wcight in September
1953 and her abdomenbecamevery distendedseveralmonths later. An
exploratorylaparotomywas done in July 1954 at Brooklyn Hospital and
revealeda large mass in the lower abdomenwith metastases.
A lymph
node biopsy was performedand showedlymphosarcoma.
(Fig. 190) Subsequently,she had 36 X-ray treatments,
followedby anothercourseof l2
X-rays, the last in December1954. She felt relativelywell until the first
week in February when abdominalpainsrecurred.A mass in the middle
abdomen,the size of a large grapefruit,could be felt.
Under treatment with sterols,the tumor first increasedslightly in size
so that by the middle of April, it extended,filling up the enrireleft side of
the abdomen.At the end of the month, the pain becamestrongerwhile
the abdominalmassremainedunchanged.
The patientwas treated,according to her analyses,with I mgm. of hexyldiselenide
daily. The tumor became much reducedin size.Pain recurredat intervalsand, althoughonly
a small masswas still palpableafter lt/z yearsof treatment,the patient's
generalconditionstartedto deteriorateat that point. Analysesthen revealed
a changein the pattern present.The treatmentwas changedto a mixture
of higher alcohols-octanol, heptanoland polyconjugatedalcohols.The
tumor rapidly regressed
and, at the beginningof 1957, the mass had entirely disappeared.She continued in good general condition until August
1957 when she had a coronaryocclusion.At present,two years later, the

l.llllRAPF.trTl(

At,t'R(]A( l{

1'o

( ANctiR

5lr

palicnt is in goeldgenerll conditionanti uithout anv sign of recurrcnceof

the tumor.
ln this case. the first treatmentwith stenrls.using urinary pH as a
critcrion,broughtsomesubjectiveinrprovement
but this could also bc consideredto bc the result of the radiation.The improvementwas transitory
and thc appeilranceof more symptoms.probablyrcllectcdthe rvaningcfTect
of radiation.Thc changeto hcxyldiselcnide
broughtimprovenrcntbut could

F t c ; . 1 9 0 . P h u t o n r i c r o l i r a p ho f i r l r n t p h n o d e o f p u t i c n t \ 1 . S . s h o ! , , ' i n ga l - v r n p h o .
\a rconta.

not cntirely control thc condition. It was rvith highcr alcoholsthat irn-fhc
portant objectivcas well as subjectivcchangeswcrc achievcd.
tunlor
decreased
rapidly. Intcrruptionof treatment.duc to fie cardiaccondition"
did not seemto inlluencethe favorablcproercssof the condition.
M. B.-Toward the'end of 1950.at thc agc of 4-5,thc patientbcgan
to complain of right lower quadrant and i.ntcrnrittentlcft upper quadrant
p a i n .S h cd i s c o v e r e a
d n a b d o n r i n anl r a s sb y h e r s c l f I. n F c b r u a r yl 9 - 5 1 .a n
exploratorylaporatomywas pcrformedat Lsnox Hill Hospital rvith the
following findings: "Situated in the left side of the pelvis, a huge cystic
structurcthe sizeof a football was found. 1'hc supcrioru'all oi this nrirss
was attachedto thc nrcsentery
of thc kxrp of the snrallbori'cl.The anterior

522

nEsEARcH tN pHystopArHoLocy

cyst was attachedto the posteriorleaf of the broad ligamentand contained

the left ovary. on the ovary and within the wall of the cyst were several
papillomatousstructures.On the right side of the pelvis was another cystic
masscontaininga greenishturbid fluid. This cyst containedthe right tube
and ovary and on its walls were severalpapillomatousprojections. The
parietalperitoneum,the liver and omentumwere studdedwith tumor implants and severalsectionsof bowel were matted togetherby the same
tumoral tissue.Biopsy specimenswere obtainedfrom severalareas."
The grosspathologicaldiagnosiswas papillarycarcinomaof the ovary
with metastases
to peritoneum,liver, intestinesand duodenum.The report
on the frozen sectionwas malignant.Microscopicexaminationconfirmed
the diagnosisgiven at the time of the operationand revealeda fibroid fatty
tissueextensivelyinfiltratedby a malignantneoplasnro[ epithclialorigin.
The paticntcame under our care I I daysafter her operation.The abdominalmasswas palpable,risingfrom the pelvisto a level slightlyabove
the umbilicus.Vaginalexaminationrevealedlargecysticmassesfilling both
fornicisand cul-de-sac.
The pelvicstructureswere partiallyfixed. The urinary specimcnshoweda pH betweenJ.4 and 7.8 and the surfacetension
w a s 6 8 t o 7 l d y n c s / c c n t i m c t eTr .r e a t m e n w
t a s i n s t i t u t e dw i t h a m i x t u r e
of conjugatedfatty acids,derivedfrom cod liver oils, and a preparationof
hydropersulfides
as well as sodium thiosulfate.The first preparationwas
administercdintramuscularly
in a -5o/aoi| solutionin dosesof from 2 to 4
( c o r r e s p o n d i nt o
g 0.5% sulfur) and lovr
c c . d a i l y .T h e s u l f u rp r e p a r a t i o n
solutionof thiosulfatewcre administered
in dosesrangingup to 6 cc. daily.
The treatmcntwas continuedwithout changefor I ycar and 8 months.
The abdominalmasswas found somewhatsmallera month after starting
treatment.A month later. a surgeon'sreport showedthat the tumor was
about vt the size at the timc of operation.After sevcralmonths,no mlss
could be felt on abdominaland vaginalexamination.After twenty months.
the treatmentwas discontinuedfor one year. At the conclusionof this
period, the patient began to relapseand a cystic mass in the culde-sac,
palpableunder finger examination,was found. Treatmentwith the same
medicationsas bcfore was institutedbut did not changethe dimensionof
the recurringtumor. Despitetreatment,the tumor continuedto grow and
in April 1954, the abdomenwas distended.X-ray also revealeda large
liver and elevationof the right diaphragm,The patient was readmittedto
Lenox Hill Hospita.lwherea smallabdominalincisionrevealedtwo rnasses.
one above the livcr and anotherone in the lower abdomen.Fluid reaccumulatedrapidly and the patient'sgeneralcondition becamepoor. on
severaloccasionsit was necessaryto tap the subdiaphragmatic
cyst and

THIN,AP}]UTIC AI'PROA{]II^rO CANCF,R

.i

-523

lo*'er abdomenseparatelyas emergencyproceduresbecauseof acutc distre$s.

Becauseanalysesnow showeda low pH (around 5.2) and low surfacc
tension(around6l dynes/centimcters).
and alsobccauseshehad obviously
failed to respond to conjugatedfatty acids. trcatment with butanol and
nonsaponifiable
lipids of intestinewas insrituted.By July the situationappeared to havc been brclughtundcr control again. Fluid was no longcr

F r c . 1 9 l . P h o t o m i c r o g r a p ho f a l y m p h n o d e b i o p s i e df r o m p a t i e n t M . B . s h o w i n g
meta$taticadenocarcinoma.

accumulatingand lcg edcma,which had becn pronounced,was gone. At

the end of 1954, no massescould be felt. Thc subjectivecomplaintsdisappearedslowly.In February 1955,thc nonsaponifiable
medicationswere
replaccd by heptanoland octaniil. The patient continucd to improve with
lhis treatnrent.In March, the abdomenwas a a little swollen.By the end
of April, a masscould bc found in thc abdumcnjust abovethc umbilicus.
At the end of May the masshad incrcasedand the patient was hospitalized for aboutthreewceks.A largcbloodycyst wasdrainccl.A smalllymph
node was biopsied and microscopicexiiminationshowed the same meta(Fig. 191)The patientwas put on epichlorohydrin
staticadenocarcinoma.
and the mass did not grow back. she has remained on epichlorohydrin

524

nEsEARcH rN pHystopATHoLoGy

and for four years has had no subjectivediscomfort.No mass can be felt
in the aMomen and the patient is well now.
Radiation and Chcmotherapy
Parallel to the guided chemotherapy,the idea of mixed radio- and
chemotherapyhas led us over the yearsto severaltentatives.In one of them,
in collaborationwith Dr. L. Goldman on a larger scale,chemotherapywas
addedto radiotherapy.(Note I)ln spite of the interestingimmediateresults, l/ chemotherapywas not further pursued, the ultimate fate of the
patientsthus treatedwas no better than that of the patientswho had received radiationalone. Analysisof thesevarious attemptshas permitted
us, however,to establishthe conditionsunder which such a mixed treatment would appear especiallyindicated.consequently,it is the type of
offbalancepresentwhich is seento determinethe relationshipbetweenthe
two therapies,and respectivelythe procedureto be followed. The samc
relationhas permittedto definethe conditionsunder which chemotherapy
would be indicatedand addedto radiotherapy.It should not be forgotten
that throughits action upon the fatty acids,radiationrepresentsindirectly
an agentof the samegroup to be addedas the negativelipoids.
The caseswith a type A offbalancewhich seemto respondinsufficiently
to medicationwith agents,representan indicationfor the mixed treatment.
Acting in conjunctionwith radiation,chemotherapy
has appearedespecially
active. Chemotherapystartedbefore,is continuedactively during radiotherapyand followingradiation,for a long time, even for years.The radiation acting as an adjuvantagentis usuallyapplied in small doses.fronr
only severalhundredr to a few thousandr, in generalmuch belorvthosc
dosagesknown to induceby themselves
alone therapeuticeffect upon tumors. In caseswith type D, as the radiotherapyrepresentsan agent increasingthe actualoffbalance,
an addedchemotherapy
with positivelipoids
would have the role of correctingnot only the existingmanifestations
but
of preventingsome of those which would be inducedby radiotherapy.It
w o u l d i n f a c t h a v e t h e s p e c i f i ca i m t o p e r m i t t h e c o n t i n u a t i o no f r a d i o t h e r a p yw h i l e l i m i t i n gi t s n o x i o u si n f l u e n c e .
The mixed therapyhas to be guidedby the routinetest, with frequent
analyses,in view of the rapid important changeswhich have been seen to
occur. In thesecases,it appearsusefulto follow especiallythe changesof
the ctrlorideindex, thoseof peroxidesin the urine which togetherwith the
surfacetension,are seento be not only particularlyinfluencedby radiation
but able to indicatethe importantchangein offbalance.

THERApEUTIC AppRoACH

'ro

CANCER

525

Exceptionallygood resultswcre seenin thc'casesin oflbalancetype A

treatedby this mixed method whcn chenrothcrapyhas been added to
radiotherapy.These casesare cspcciallyinterestingbecauseof the pers i s t e n tr e s u l to b t a i n e d .T h c a b n o r n r a l l ys n r a l la m o u n t o f r a d i a t i o no f t e n
used, appliedonly on a localizedspot, has servedin these casesas an
adjuvantto help to resolveparticularlocal problems,while the chemotherapy,continuedfor a long time thereafter,to preventthc appearance
of recurrences.
In the caseof offbalanceD, as the radiotherapywill increasethe actual
offbalanceand the role of chemotherapyis to rcducethe intensityof this
offbalanceand thus permit thc continuationof radiationfor its direct target
effectthe resultsare lessinteresting.
difleringlessthan in the first casefrom
thoseobtainedby radiationalone.I he continuationof chemotherapy
howe v e ra f t e rt h e r a d i a t i o nh a sb c e nc o n r p l e t c dh,a sa p p c a r c de s p c c i a l lhy e l p f u l .
Level Chemotherapy
I n r e c e n yt e a r s t, h e c o u r s eo f o u r r e s e a r c h a sb c e nd i r e c t e dp a r a l l e tl o ward the preciseapplicationof availableagentsas well as finding new
substances.
For it has bccomeincreasingly
evidentto us that it is the basic
independence
of the differentlevelsof organizationin the body which representsthe critical factor in the pathogenesis
of cancerand its manifestations and which is also critical to the most effectiveuse of therapeutic
agents.This concepthasled us to focusour rescelrch
effortsin certainareas.
Attemptsare madeto find criteriacapableof indicatingchangesat specific
levels of organization.Similarly,a growing awarencssthat each element
belongsto a specificlevel of biologicalorganizationand exerts its biological activity primarily at this level has led us to try to use specific
elementsin correctingdysfunctionsat variouslevels.Becauseof the independence
of the levels,we have attemptedto triggerthe defensemechanism at appropriateindividuallevclsrather than indiscriminately.
This is
especiallyimportantwhen the failureof the defenseto operateat someone
level represents
a major factor in allowingpathologicalstatesto advance.
While each of thesedevelopments
has beenimportant,it has been the
use of all three which has brought marked progressin the clinical application of the methodin the last years.
With the conceptof indepcndence
of levels,the use of more than one
agentis not simply a matterof synergistic
activity,but rathcr of employing
proper level in complexconof
agents,
capable
acting
at
its
several
each
ditions where more than one level is involved.To make this possible,it

526

RESEARCHtN

pHysropATHoLocy

has beennecessaryto relate analyticaldata to changesoccurringat specific

levels.For example,changesin potassiumin serum and red cells have
beenrelatedto cellularlevelabnormalities;
urinarysurfacetensionchanges,
to metazoic;specificgravityto systemic;urinarypH and blood eosinophiles,
to tissular.This has guided the choice of agents:seleniumpreparations,
epichlorohydrinand heptanolfor the cellular level; sulfurizedhydronaphthaienes,lipoacids, unsaponifiablefractions and glycerol for the tissue
level;magnesiumand sodiumthiosulfates,
propionicaldehydeand butanol
lor the organ and organismlevels.Through this approach,therapy has
evolvcd toward what we can now considerto be biologic'allyguided levelchemotherapy.
The following observations
show the role of theselevel indicationsin
the conductof the treatmcnt.
F . R . , i n M a r c h o f 1 9 5 7 ,h a d a l e f t m a s t e c t o mfyo r a n a d e n o c a r c i n o m a
of a mammarygland.In July of 1958,she beganto experiencepain in the
lower back and legs.In the thrce monthsprior to conringunder our care,
severalskin nodulesand someprogressive
difficultyin breathingwas evident and therewas a lossof 25 lbs. in weight.on admission,multiple skin
and subcutaneous
lesions,someof themmeasuring3-4 cm in diameterwerc
found, as well as 3 or 4 nodulesin the right breast.The ambulantpatient
startedtreatmentwith seleniumand epichlorohydrin.Although treatment
wasfollowedwith irregularity,improvementwas seenin the local lesions.In
December,1959, howcver, the difficulty in breathinghad markedly inc r e a s e da n d o x y g e na d m i n i s t r a t i ow
n a s n e e d e d I. n l e s s t h a n a w e e k h e r
conditionworsened.The treatmentwith more seleniumand epichlorohydrin
appearedunableto control thc situation.She was admittedto the hospital
in extremcdyspnea.The suppression
of oxygenfor even a few minuteswas
followedby convulsions.Radiologicalexaminationshoweda very limited
exudateespeciallyin the right hemithorax,but muitiplematastases
in both
lungs.Alt thc analysesshowedan offbalanceof rypc A. considering the
conditionas manifestedalso at the organicand systcmiclevel,to the treatment with epichlorohydrin,acting at the cellular level, we added bixine
and propionicaldehyde.The first agcntwas considered
to act at the tissular
the
latter
the
level and
at
systemiclevel. With this treatmentthe patient
madea vcry good and constantrecovery.In lessthan two weeksit became
unnecessary
to use oxygen and her generalimprovementprogressedso
t h a t 3 m o n t h sa f t e r t r e a t m e nw
t a s s t a r t c d t, h e p a t i e n tw a s a b l e t o r e s u m e
part of her housework.With mixcd treatntent-epichlorohydrinacting at

T H E R A p E U T t cA p p R o A c H r o

cANcER

527

the cellular level, and bixine and propionic aldehydeat the organic and
systemiclevel-the conditionseemscontrolled.I'he pulmonarymetastases
seenin previousX-rays have alsodisappeared
and all thc skin nodulesdecreasedrapidly, most of them having complctelydisappeared.she is at
presentin good stateof health,doing all her own housework.
Mrs. c. H., 60 yearsold, camc under our care two years ago with a
historyof lymphosarcoma
with two positivebiopsiesof thc inguinalglands,
complaining
e s p e c i a l lo
y f p a i n i n t h e a b d o m c nO
. n e x a m i n a t i o ni ,n g u i n a l ,
axillar and cervicalglandswere present,someof thenr 4 crn in diameter.
With treatmentof sodium thiosulfateand fatty acid hydropersulfides
in
relativelysmalldoses,the pain wassufficiently
controlled.After two months
of treatment,the patientexperienced
extremepain and a tumor of the head
of the right humerus.In view of the local lesionswith analyseswhich all
showedan intensiveoffbalanceof type A, the treatmentwas changcdto
epichlorohydrinand heptyldiselcnide.
Administeringtheseagentsin relativelyhigh doses,it was possiblcto not only controlthc pain in a few hours,
but to have the iesiondisappearin lessthan two weeks,as did also all the
abnormalgJands.
However,in spiteof theseobjectivevery good results,the
generalconditionbecameunsatisfactory,
cspeciallywith marked weakncss.
C h a n g e si n t h e d o s e so f m e d i c a t i o o
n r t h e t e m p o r a r yd i s c o n t i n u a t i oonf t h e
m e d i c a t i o nf.a i l e dt o c o r r e c ti t . I t w a s o n l y w h e n p r o p i o n i ca l d e h y d ec o n s i d e r e dt o a c t u p o nt h e o r g a n i s ml c v c lw a sa d m i n i s t c r e tdh. a t a r a p i dc h a n g c
t o w a r da f e e l i n go f g e n e r aw
l ell-being
w a s o b t a i n e dA. t p r e s e n tw
, ith small
dosesof propionicaldehydethe patientis entirclyfree from any subjective
or objectiveabnormalmanifcstations
and has bccn so for almost a year,
M r s . E . R . , 4 2 y e a r so l d , h a d a r a d i c a lm a s t c c t o m fyo r a n a d e n o c a r cinomaof the breast.After I t/z years,shc experienccd
persistent
back pain
w h i c hw a sf i r s td i a g n o s e ad s a r t h r i t i sX
. - r a y s t u d i c sa n d a m y e l o g r a mm a d e
at MontefioreHospital showedtwo metastaticlesionsat the I st and 2nd
lumbar.Surgicalinterventionwas thoughtto be not indicated.X-ray therapy
neitherrelievedthe pain nor made it possiblcfor her to lcave her bed. At
a d m i s s i o un n d e ro u r c a r e ,t h e p a t i e n w
t a si n s e v e r ep a i n a n d u n a b l et o e v e n
turn in bed, althoughshewas still ablc to moveher legsand thcir sensitivity
w a s c o n s e r v e dA. t r e a t m e nw
t i t h h e p t y l d i s c l c n i dpcr,o p i o n i ca l d e h y d ea n d
bixine made thc pain disappearand up to date the patient is leading a
normal life, after havingbeenbedriddenfor 8 months.
M r s . M . M c B . , 6 2 y e a r so l d , c a m c u n d e ro u r c a r ef o r a b a s o c e l l u l a r
carcinomaof the left side of the face,near the inferior eyelid.Advised to

.,..
,.:,:,.4t
:ltt ,
'.:j:.,!:;:ti

528

RESEARCH tN

PHt'StOPATItOLoCY

undergosurgcry,which would cnucleatcthe left cyc. shc rcfusedthe operation. A hiopsy u,as performcd (l'ig. tq2) showing the presenceof a
basocellularcarcinoma.She u,as treatr-'dlvith hcxyldisclcnidcand with
sodium thiosulfate,and the lesion clislppearedwithin a few weeks. No
rccurrcnccrr,asobscrvcddurint thc nlrsl 6 r'cars.

l"l
. t

r,s
t;r'*
tl"

rl
ti

rd
'r\
A

F l t ; . l 9 l , P h o t o m i c r o g r a p ho f t h c h i o p : y n r l d c o n p a t i c n t l t l . N l c B . s h o w i n g u
b l s t l c e l I u l a rc u r c i n o l n t r .

M r . C ' 2 . . 5 6 Y c a r so l d c ; . t n t u
c n d c ro u r c a r c f r r r a l c s i o no f t h e u p p e r
l i p . B i o p s y p r o v c d i t t o b c x s q u a n l ( ) ucsc l l c a r c i n o m a . ( f i g . / 9 J l T h e
patientrefusedsurgeryor nrdiation*hich r,''cadvised.Followingthe biops.v
t h c t u n r o rs t l r t c d t o s r o $ r ; r p i d l r .u \ s c c ni n F i g . 1 9 4 . A c c o r d i n gt o t h e
anall'scs.
the paticnt*'us trertctl rrith flttv acid hydropersulfidc.
The lcsion
(
i
n
d i s a p p c a r c d l c s st h u n I u c c k s t : t g . 1 9 ,/5N o r c c u r r c n c cw a s o b s c r r v c c l
a l t h o u g ht h e p a t i c n tw a s n o l o n s c r u n d e r l r c a t m e n t T
. h c p a t i e n td i e d i i
m o n t h sl l t c r f r r l n t i t c ( ) r ( ) n i l r ov c c l L r s i o n
"l'hcability to induccchangcsat sinqle
specificlevelshas alteredcompletclyi,l'hatrvus()nceit grossduulistrcapprolch. Llntil rccently,we had
c o n s i c l e r ci d
t i n l c l r i s a b l el o i n d u c et h c r l p c L r t icch a n g c su t a s i n g l el c v c l i f
r-rflbalances
\\,crcprcscntat nrorethan onc levcl. With the conceptof level
independcncc.it has become part of the merhod to attempt to influcncc

T T I E R A P L U T I C A P P R ( ) A C I I" T OC A t \ i C I R

l9l.

P a t i e n t ( i . 2 . h e f < t r cl r c a r r n c n t

F t c ; . 1 9 4 . T h c t u m o r g r c w r a p i d l l f o l l r r u i n gh i o p s y

F r ; . 1 9 5 . P a t i e n tt i . Z , a f t er r r c ; r r n r c n r

-5l()

530

* E S E A R C Hr N p H y s r o p A T H o L o c y

the level in which the predominantabnormality is present rather than to

induce gross changesat all levels.In caseswith generaloffbalanceA but
predominantat the cellular level, therapy to induce offbalanceD limited
to the cellular level alone, with the metazoicremaining at offbalanceA,
has producedinterestingobjectiveclinical results.Similarly,good results
have been obtainedin caseswith predominantoffbalanceD at the cellular
level and in the metazoiccompartment,with treatmentdirected at changing the systemicand tissularoffbalanceD into type A without influencing
the oftbalanceD at the cellularlevel.It is interestingto note that in both
typesof cascsthe changeslead ultimatelyto the same overall pattern of
offbalance-D for the cellularlcvel and A for the mctazoiccompartment.
Level chemotherapywith selenium,heptanol, propionic adlehyde and
otheragents,hasgivenhighlygratifyingresults.It led us to the presentform
o f t r e a t m e n tw
, i t h p a r t i c u l a r l yg o o d r e s u l t s I. n a v a r i e t yo f t u m o r s ,s o m e
impressivebecauseof their huge dimensionsor becauseof their high degree of malignancy,the immediate objective responsehas been striking.
Massivelung metastases
from breastcancer,generalized
abdominalmetastasesfrom colon carcinoma,bone metastases
from breastor prostate,and
melanoma
metastatic
are among the caseswhich have respondedand we
are now waitingfor time to indicatewhetherthe resultshave lastingvalue.
While we were convincedin the past of the inherentpotentialityof the
method in gencral, becauseof resultswith many different agents and
criteria,we have beengreatlyencouraged
by the most recentapplications.
Wc belicvethe method now providesa meansof controllinga significant
proportioneven of preterminaland terminalcasesof malignancyconsidcred otherwiseentirelybeyondany hope.The particularlyfavorableresults
o b t a i n c di n t h e c a s c sw h i c h c a m e u n d e ro u r c a r e b c f o r e t h e d i s e a s eh a d
progressed
to advancedstagcsindicatethat we are fully entitledto prefer
this therapeuticmethod evcn for those caseswhere the presentJyused
p r o c e d u r em
s i g h t s t i l l h a v c a c h a n c et o h e l p .

C H A P T EI R6
PRESENT F-0RM
OF TREATT{ENT
rTr\t
l L n , , " u * A p E U r r c M E A S U R EaSg a r n sct a n c c rw h i c h h a v e e v o l v e dt h u s
far out of the pathogenicand pharmacodynamic
conceptsdescribedin these
pageshave provcd, as we have shown above,to bc useful.All along in
their development,thcy have fallen short of producinguniformly,the full
measureof benefit which, we have continuouslyhoped, one day will be
attainedin most and possiblyin all casesof malignancy.
Yet the resultsachicvedwith biologicallyguidedtherapy,imperfectas
t h e y h a v e b e e n , h a v e c o n t i n u o u s l yi n d i c a t e dt h c g r e a t p o t e n t i a lo f t h e
method.
W h a t h a s b e e na c c o m p l i s h eddu r i n gt h c y c a r sh a s b c e n h a r d l yc o n s i d ered as a linal measureof the effectiveness
of this basicnew approachto
treatmentbut onJy of the availablecriteria and agents.With thc criteria
and agentsthe method itself has been evolving.In its presentform, the
method representsa valuabletool for helping many cancer patientswho
are entirelybeyondhelp with presentday methods.In spite of its accomplishmentsfurther advancesalong the same lines-both in techniquesfor
recognizing
fundamentaloffbalances
at diflerentlevelsof organizationand
cven in new compoundsmore effectivein correctingthem-are to be
expected.
Equal in importancewith the current resultswith biologicallyguided
therapyis the fact that new applicationsarc evolving;that this approach
helpsus to betterunderstanding
and treatmentof malignancyand of other
pathological
well;
conditionsas
and that, by its very nature,the methodhas
the capacityto furnish the guidanceneededfor improvingit.
We are presentinghere the form of treatmentwhich we currentlyuse.
531
: 1 , :
.

| . .

532

n E S E A R C Hr N P H Y S r o p A T H o L o c y

In this form, when correctly applied, biologically guided therapy can, in

many cases,bring under control even far-advancedmalignancies.
Thc importanceof the correct applicationof agentsbased upon specificcriteria
makesit necessaryto emphasizehow agentsare chosenand used.
Criteria Used: Recognition that no test is able to indicate by itself the
existenceof more than an offbalanceat a specificlevel, and that cancer is
a complexconditioninvolvingmany levels,has led to the use of a group
of tests able to offer the necessaryinformation on offbalancesat different
levels.We use the following routinely:
U r i n a r y - s p e c i f i cg r a v i t y
pH
surfacetension
c a l c i u me x c r e t i o n
index
B l o o d - p o t a s s i u mi n s e r u m
potassium
i n t o t a lb l o o d
Painpattern
Body temperature
I n c x c c p t r o n ai nl s t a n c cw
s h e nt h c r o u t i n ct c s t sd o n o t s u f f i c ew
, c use.
Urinary----chloride
retentionindex
B l o o d - c h l o r i d e si n s e r u m
l e u c o c y t ecso u n t
c o s i n o p h i l ecso u n t
T h e o l l b a l a n c ei n d i c a t i o nps r o v i d e db y t h e s et e s t sa r e s h o w ni n T r s L t
XXII.
T,rnr.c XXII
Test
Specificgravity
pH
S u r f a c et e n s i o n
Serumpotassiunr
Total blood potassium
Body temperature
[-eucocytcs
Eosinophiles
C h l o r i d ei n d e x
C a l c i u mi n d e x
C h l o r i d e si n s e r u m
P a i np a t t e r n

OffbalanceD
high
low
low
high
low
low
low
low
high
Iow
high
alkaline

Average
Values
I .016
6.2
68
4.5 mEq
38 mEq
37'C
7000'cmm
1 0 0/ c m m
, , I

2.5

s::'s

Offbalance
low
high
high
low

hish
high
high
high
low
high
low
acid

PRESENT FoRM

oF

TRE^TMENT

533

whereas the other analyscsgive direct information concerningthe offb a l a n c e st ,h e v a l u c so f p o t a s s i u mi n s e r u mh a v et o b e c o r r e l a t e dw i t h t h e

valuesin red cells or in total blood potassiumfor a recognitionof the
offbalancepresent.A serum potassrumabove 4.5 mEq ard a low total
blood potassiumbelow 3tt mEq indicatc an offbalanceD, while a low
s e r u mp o t a s s i u mb e l o w 4 . 2 m E q w i t h a h i g h t o t a l b l o o d p o t a s s i u ma b o v e
4 0 m E q . a n o f f b a l a n cA
c . L o w v u l u e sf o r b o t h a n a l y s eisn d i c a t ea q u a n t i t a t i v e d e f i c i e n c yo f t h i s e l e m e n ti n g e n e r a l w
, h i l e h i g h v a l u e sf o r b o t h a n
e x c e s so f t h i s e l e m e n itn t h c o r g a n i s m .
As relatedto the diffcrcnt levclsof the organization,spccificgravity
changesreflectprocesses
taking placcespcciallyat the systemiclevel.The
pH reveals,indirectly,changesat the tissuelevel.Surfacetensionindicates
offbalancesat the metazoic compartmcnt;both serum and total blood
potassiumindicate cellular level offbalances.However, while potassium
m e a s u r e m e ng
t si v e a n i n d i c a t i o no f c y t o p l a s mc h a n g e sc, a l c i u mi n d i c a t e s
c c l l u l a rI e v e lc h a n g e sc, s p e c i a l l iyn t h c m c m b r a n c sE. o s i n o p h i l c o u n t si n d i c a t ea c i d - b a sceh a n g e sa t t h e t i s s u el c v e l .a n d t c m p c r a t u r cs, y s t e m i co f l b a la n c e s .
A s w e h a v e s e e n ,a n y a b n o r m a vl a l u ei s m o s t l i k e l y t o a c c u r a t e l yr e flect a fundamcntaloffbalanceat its corrcspondinglevcl when the same
value is obtainedin severalrepetitionsof the test. Tests on two or threc
successive
days are most helpful in asccrtaining
the existenceof an oftbala n c e ,e s p e c i a l liyf i t i s l i m i t e dt o o n l y a f e w l c v e l s .
The offbalancedetermineswhich acentsare to be used.
As anti-A asentsw
. e now use:
propionicaldehyde
cpichlorohydrin
hexylor heptyldiselenide
t e t r a l i np c r s e l e n i d c
t e t r a l i np e r s u l f i d e
thiosulfate
s o d i u mo r m a g n c s i u m
m i x t u r eo f l i p o a c i d s
, e
A s a n t i - D a g e n t sw
heptanol
polyunsaturated
alcohols
butanol
glycerol
fractionof liver
insaponifiable
g l y c e r o p h o s p h oa
r icci d

534

n E S E A R C Hr N P H y s r o p A T H o L o c y

The preparationsand usualdosesare:

Propionicadlehyde l0% sol.-for oral administration,usedfrom 501000 mgr. daily; for parenteraladministration:2o/o sol. in saline.
E p i c h l o r o h y d r i n 0: . 5 o / os o l u t i o ni n i s o t o n i cs a l i n e ,u s e d f r o m l / l O
mgr. to hundredsof milligramsdaily for parenteraland oral administration.
Hexyl or heptyl diselenide:in solutionsfrom 20 microgramsto 50
mgr./cc. for intramuscularinjection.Orally, we use either capsulescontainingfrom l0 microgramsto l0 mgr. or, still better,a solutionof O.4Vo
in oil, whichcorresponds
to 100 microgramsper drop. The dosesvary from
l0 microgramsto 50 mgr. or more daily.
Tetralinperselenidc:
This preparationhas 40 mgr. of seleniumfor 100
g r . o f t e t r a l i na n d i s u s e do r a l l ya s d r o p s ,f r o m a s o l u t i o no f l O % o r l V o i n
o i l . F o r i n j e c t i o n sa, s o l u t i o nc o n t a i n i n gl 0 % i n 1 0 0 c c . o f s e s a m eo i l , i s
u s e d i n d o s e sr a n g i n gf r o m 0 . 1 c c . t o 1 0 c c . d a i l y . I n s t e a do f t c t r a l i n e ,
n a p h t h a l e noer o t h e r a r o m a t i ch y d r o c a r b o nasr e u s e d .O n e c c . o f a l O V c
s o l u t i o no f t h c s ep r e p a r a t i o ni sn o i l , c o n t a i n s0 , 1 n r g .o f s e l c n i u mt;h e d o s e s
u s e dr a n g ef r o m 0 . 1 c c . t o 1 0 c c . d a i l y .
Sodium or magnesiumthiosulfateis used in a loo/o solution for oral
administrationor in a 47o solutionfor intramuscularinjection in doses
rangingfrom 25 mgr. to 5 gramsor more daily.
Tetra.linepersulfide:This preparationcontains59'c sulfur in oil and is
usedorally from lr',.,,
cc, to more than l0 cc. daily. In intramuscularinjectionsit is usedfrom ,1,{
o cc. to 2 cc. daily.
The acid lipids mixture is a solutionin oil of lo/o bixin, 2Vo cow liver
lipoacidsand 7Vo cod liver oil fatty acids.It is administeredin dosesof
f.romr/z cc. to 6 cc. daily by intramuscular
injcction.
Heptanol in a solution of 0.57o or 5Va in oil is used for parenteral
a d m i n i s t r a t i o na; s o l u t i o no f 5 % i n o i l i s e m p l o y e df o r o r a l a d m i n i s t r a tion. The dosesrangefrom 1 mg. to hundredsof mgs. a day.
Polyunsaturated
alcoholsare saffioweroil fatty acidswith the carboryl
primary
to
changed a
alcohol.They are usedin a lOVo solutionin oil with
V+ to 2 cc. injectedintramuscularly
up to four times a day.
Butanol in a 6.5Vo water solutionis used for oral administration.A
6.5% salinesolutionis employedfor parenteraladministration.
Usual doses
are from Y+ cc. to more than 100 cc. daily.
Glycerolis usedas a 50Vosolutionfor oral administrationor as a 2OVo
solutionfor parenteraladministrationin dosesrangingfrom lessthan 0.2
gram to a few gramsdaily.
Insaponifiablefraction of pork liver in a 5Vo solution in oil is used
for intramuscularinjectionsin dosesfrom Vz cc. to 8 cc. daily.

PRESENT FORM

OF TREATMENT

,,i .535

A preparationof a mixture of polyunsaturated

alcohol,heptanoland
butanolalso is usedfor oral or parenteraladministration.
If, during treatment,any other medicationsapparnecessaryfor various concomitantconditions,it is preferableto choosethem with consideration given to the influencethey will exert upon the patternspresent.
TlgLs XXIII shows the effectsof some commonly used therapeutic
agentsupon the fundamentaloffbalances,
which would indicatetheir use
in caseswith one or the other offbalance.
Trsle XXIII
E r n e c r s U p o N O r n s e L , { t , / c rA
-s rNo D or Vrr,tous Acexrs
U s t o T ' x r ' n l p L ,nuc A L L y
Having an anti A tflect
Dicoumarol
Digitalis
Antipyrine
Aminopyrine
Acetophenetidine
Atropine
Quinine

Huving on anti D efJect

Glycerol
Glucose
Coramine
Acctyl salicl,'licacid

Caffeine

Procaine
('odeine
Morphine
Dcnrcrol
A m i n o p h yl l i n e

Liver extracts
vit. A, D, 8,,

I ron
Vit. 8,, B,, K. E

Testosterone

I)rogesteronc
Sti Ibesterol
Dcsox.v-cort
icoslerone
G I ucosanri ne

Epinephrine
Penicillin
streptomycin
Aureomycin

Cortisonc

Sulfas

Insulin
B a r bi tu r a t e s
lv{ercuhydrine
Benzedrine
B e na dr r I

536

pHysropATHoLocy

xESEARcH tN

In tuing the various agents,we have to keep irt mind the eftect upon
the existingoffbalances.
Conductol Treatmenl;Although there may be somevariationsdepending upon the circumstancesof individual cases,treatment is conducrcd
generallyas follows:
Urine and blood analysesare performed.Values for two or three consecutivedays are usuallydeterminedbeforestartingtreatment.If all values
indicate the same pattern, the diagnosisof the offbalanceis clear. If the
analysesindicate the presenceof different patterns, an interpretation is
made on the basisof analysesshowingthe offbalanceat specificlevels.For
patients in whom manifestationsinvolve the metazoic compartment, urinary surfacetensionis the important criterion. Urinary specificgravity and
urinary pH, are important when systemicor tissular manifestationsare
present.Potassiumis the criterion when changesat the cellular level are
most important.The body temperatureand the other complementaryanalysesare usedwhen discordantpatternsare encountered.
They help to recognizethe type of offbalanceat the different correspondinglevels.
Once the level oftbalancesare determined.suitabletreatmentis instituted.The agentis chosenfrom the proper group accordingto the level
TrsLE XXIV
Acexrs CHosrNAcconorNcro rHE Tr,srs
AceNrs
Test

Level

OftbalanceA

OffbalanceD

Cellular

Potassiumin I S e l e n i u mp r e p a r a t i o n s Heptanol
blood
lEpichlorohydrin
Urinary
calcium

Tissue

U r i n a r yp H
Lipoacids
Blood
tTetralinpersulfides
eosinophiles
Hydropersu
lfides
Mg Thiosulfate
Surface
Tension
P a i np a t t e r n

Polyunsaturated alcohols
Unsaponifiable
fraction
liver
Glycerol
Butanol

Organ and
U r i n e s p e c i f i cM g o r N a T h i o s u l f a t e
gravrty
Organisnr
P r o p i o n i ca l d c h y d e
Glycerophosphoric
or

I
I

Surface.
l ensron

I Body

l_

I
t"lo"'"i"':]

lactic acid

PRESENT FORM

OF TRE^TMENT

537

to be influenced. Since, in invasive cancer, the offbalance occurs at tbe

cellular level, the agentsused are seleniumpreparationsand epichlorohydrin, for the offbalanceA and heptanolfor the offbalanceD.
For the tissuelevel, indicatedby pain as well as by urinary pH or
eosinophiles,lipoacid preparations,Mg thiosulfateand Tetralinpersulfides
are used for offbalance A, and polyunsaturatedalcohols, unsaponifrable
fractionsand glycerol and butanol for offbalanceD.
For the organ and systemic levels, with the offbalance recognized
through urinary specific gravity, surface tension and body temperature,
magnesiumor sodium thiosulfateand propionic aldehydeare used for offbalanceA. and glycerophosphoric
acid for offbalanceD. (Table XXIV)
Conduct ol Treatment
If the patternsof the differenttestsand clinicalmanifestations
are concordant,concerningthe offbalancepresent,the agentschosenare from the
respectivegroup. Specialattentionis given however to the level which,
clinicallyor analytically,showsthe most abnormality,so that for a patient
with a limited tumor, but without pain and in good generalcondition,the
factor guidingthe therapywill be the analysesrelatedto the cellular level
( a s r e v e a l e db y t h e p o t a s s i u mi n b l o o d ) . l f t h e g e n e r acl o n d i t i o ni s p o o r ,
indicatingrather a predominantsystemiccondition,the trcatmentwill be
directedespeciallyby the abnormalcyat thc organismlevel revealedby the
correspondinganalyticaltests-such as urinary specificgravity, surface
tensionand body temperaturc.
This interpretationof the most neededrnterventionbecomesstill more
importantwhen the data obtained-clinical and analyticalare discordant.
The treatmentwill follow the indicationfurnishedby thc levclwhich appars
the most important.In a casewith a limited tumor, and no other clinical
manifestations,
the patternof the cellularlevcl, will determile the nature
of the treatment.even if the othcr analysesshow diflerentpatterns.
For a patientwith a tumor and severepain-it is the patternof the pain
which will indicatethe agent to be used---+venif this is discordantwith
that of other analyses.
The sameis true for a systcmicseverecondition,the
respectiveanalysesdeterminethe agentto be used,cven if theseare discordantwith the analysesconcerningthe other levels.In general,the decision of what level will representthe guiding factor in the treatment,
represents
seldoma problem,the conditionof the patientdirectingthe attention towardthe principalanomaly.
In general,treatmentis startedwith small doses.If the offbalances,
and especiallythe clinical manifestationspersist,increaseddosesare indi-

538

*ESEARcHrN pHysropATHoLocy

cated.Larger amountsare usedand are administeredmore often. Once t-he

desiredanalytical and clinical effect is obtained, dosageis maintained. If
analysescorrespondingto the offbalancepass far enough on the opposite
side,the amount of the medicationis first reduced.If this changepersistsor
increases,
treatmentis discontinued
for severaldays. If the new offbalance
still persists,and especiallyif new clinical symptomsdevelop, use of the
oppositegroup of agentsis to be considered.A slight passageof an offbalanceinto the oppositeis usuallysalutaryand treatmentis continuedas long
asclinicalimprovementpersists.
It appearsto be of great importancethat treatment be continued for
severalmonths after all subjectiveand objectivemanifestations
have disappeared.In a number of patients,we have been investigatingthe value of
continuingmedicationfor yearsin very smalldosesas a prophylacticmeasure to prevent recurrences.The resultshave been highly encouraging.No
have been noted with controlled continued use of any of
inconveniences
the substances.
Resultsobtained
With this form of trcatmentmost of the resultsobtainedare striking.
perselenide,epichlorohydrin
With naphthaleneor tetrahydronaphthalene
and bixine as principalagentsfor the type A and heptanol-glycerol
for the
typeD, the subjective
wereseento be well conand objectivemanifcstations
trolled.Pain,if prescnt,disappcarcd
in a few days,the tumorsprogressively
diminishinguntil they disappear,
even in casesconsideredas far advanced.
The followingfew recentobservations
of suchcases,give an idea of these
r e s u l t s u, p t o d a t c .
Mrs. M. C., 59 yearsold, had 3 years ago a mastectomyfor adenoc a r c i n o m aS. h e c a m e u n d e r o u r c a r e w i t h m e t a s t a s ei ns t h e r i g h t 8 t h r i b
ascitesand
and in the I lth and l2th dorsal vertebrae,liver metastases,
With
pains
dyspnea,
severe
and
pleuraleffusion.
almostcontinuousvomitwas pering, her generalcondition was judged very poor. Paracentesis
formed, but thc fluid accumulatedrapidly. needinga secondparacentesis
l0 days later. In offbalanceA, the patient was treatedwith perselenide,
bixine propanal and epichlorohydrin.Shortly after the beginningof the
t r e a t m e n t .h e p a i n d c c r e a s cidn i n t e n s i t ya n d l a t e rd i s a p p e a r ecdo m p l e t e l y ,
the ascites
The pleural effusiondecreasedand after the two paracentesis,
no longerreproduced.The gcneralconditionchangedrapidly for the best,
Liver reducedin sizeafter threemonths
togetherwith the objectivechanges.
t a s w i t h i n t h e n o r m a l l i m i t s .R a d i o l o g i c ael x a m i n a t i o nt,w o
o f t r e a t m e nw
completelyhealed.The general
monthslater showedthe bone metastases

P R E S E N T F O R M O F T R I ] A T M E N T

539

conditioncontinuedto improve.Thc patientresumcdher normal life and

n o w d o e sn o t s h o w a n y c l i n i c a la b n o r m a l i t y .
M r . J . s . , 7 l y e a r so l d . T w o y c a r sp r i o r t o a d m i s s i o nt,h c p a t i e n th a d
persistent
hematuria,for which a prostatcctomy
was performcd.Hematuria
reappeared.
Cystoscopicexaminationrcvcaledtunrorsof the bladder,for
which he was operatedlvz yearsago. Sincethen,he has had almostcons t a n th e m a t u r i aw
, i t h f r e q u e n tm i c t u r i t i o n sd u r i n g t h e n i g h t a n d a l m o s t
everyhour duringthe day. Pain in thc lower abdomenbecameprogressively
s t r o n g e rT
. h e p a t i e n tw a s a d m i t t c da n d t r c a t e d ,i n a c c o r d a n c ew i t h t h c
analyses,
with perselenide,
bixine and epichlorohydrin.
In lessthan a week,
t h e h e m a t u r i ad i s a p p e a r e da,s d i d t h c d y s u r i a ,t h e p a t i e n tb c i n g a b l e t o
passclear urine every 6 to tJ hours.With the interruptionof the treatment
f o r a l m o s ta m o n t h ,t h c s y m p t o m sr c a p p e a r c d* ,' i t h h e m a t u r i aa n d d y s u r i a .
with the treatmentresumed,the conditionrespondedwell again,the hematuria and dysuriabeingcontrolledand pain disappeared.
Mrs. R. A., for five years,had symptomsof gastriculcers,more acc e n t u a t e idn t h e s u m m e r w
, h i c h i m p r o v e dw i t h t r e a t m e n t sF.o r s i x m o n t h s
p r i o r t o c o m i n gu n d e ro u r c a r e ,t h e p a t i e n th a d p r o g r e s s i v em
l ya r k e dd i f f i c u l t yi n s w a l l o w i n ga n y t h i n go t h c r t h a n f l u i d s .L , v e na f r c rt a k i n gt l u i d s ,s h e
very severeretrosternal
experienced
pain, almostalwaysfollowedby vomiti n g . X - r a y e x a m i n a t i o ns h o w c ds l i g h td i l a t a t i o no f t h e c s o p h a g u w
s ith a
c l e a r l yv i s i b l eg r o w t hi n t h es t o m a c hn e a rt h ec a r d i a .A c c o r d i n gt o t h e a n a l y s e st,h e p a t i e n w
t a st r e a t e dw i t h p e r s e l e n i di nc i n j c c t i o na, n d b i x i n a n d e p i chlorohydrineorally. Two weeksafter treatmcntwas startcd,the patient
was able to swallownot only fluids but also finely ground food. The intp r o v e m e nct o n t i n u e dt,h e p a t i e n tb e i n ga b l e a f t e r5 w e e k so f t r e a t m e ntto
swallowfood of almost normal consistency.
M r . J . R . , 5 6 y e a r so l d , c a m e u n d e ro u r c a r e w i t h s c v e r ep a i n si n t h c
. t e x a m i n a t i o na, s u b m a x i l l a r
r i g h ts i d eo f t h e n e c ka n d h e m o p t o i sc p u t u m A
g l a n do f 6 c m . d i a m e t c rw a s s e e n .L a r y n g o s c o p iecx a m i n a t i o ns h o w e da
tumor in the right pyrifelp f655n-Biopsy revealeda squamouscell carc i n o m aT
. h e v e r y s e v e r ea n d c o n s t a npt a i n a n d t h e c o n s t a nbt l e e d i n gc a u s e d
t h e p a t i e n tt o b e h o s p i t a l i z e dA. c c o r d i n gt o t h c a n a l y s c sp. e r s c l e n i dbey
injection,epichlorohydrinand bixine werc administcred.ln 24 hours the
p a i n w a s f u l l y c o n t r o l l e dT. h e b l e e d i n gs t o p p e da f t c r4 d a y s .a n d t h e g l a n d
'fhc
startedto decrease-l0 days after the bcginningof the Lrcatrnent.
l a r y n g o s c o p iccx a m i n a t i o ns h o w e dt h e t u m o r t r a n f o r m c di n t o a g r a y i n g
decreasing.
mass,which was progressively
l i t h d y s p n e ac, o u g h
M r s . A . D . , 6 0 y e a r so l d , a d m i t t e dt o t h c h o s p i t aw
and pain in the right hypochondrium,epigastriumand generalizedweak-

540

REsEARcH tN pHysropATHoLocy

ness.For 30 years,the patienthad complainedof pain in the right hypochonder,related to the prescnceof gall btadder stones.In February of
1960, a laparatomywas performedand a tumor of the gall bladder with
metastases
to liver was found. Only a biopsywas performedwhich showed
a carcinoma.At admissionunder our carc, the patient was in very poor
generalcondition with marked dyspnea,dcep jaundice and severepains
in the upper abdomen,emaciatcd.A right pleural effusionwas found and
a thoracentesis
performed.An irregular masswas found in the right hypochondriumarrivinguntil the umbilicus.Shehad clay coloredstools,typical
for obstructivcjaundice.In spiteof thoracentesis
the dyspneacontinuedto
be severeand the patientwas kept under oxygen.The patient was placed
under chemotherapeutictreatment with epichlorohydrin,bixine and perselenidein accordance
with her urinalyses.
The patient'sconditionimproved
progressively.
The stool returncdto normal color; fluid in the right chest
did not reproduceand the mass on the right hypochondriumdecreased
p r o g r e s s i v etloy h a v et h e l i v c r i n n o r n r a d
l irncnsionA
s .t p r e s c n tt,h e p a t i e n t
with all the subjectiveand objectivcsympromsimprovcd considerably,is
ambulatory.
M r . S . S . , 6 4 y e a r so l d , w a so p e r a t e di n l 9 _ 5 3f o r a h y p e r n e p h r o moaf
the right kidney.Two yearslater, massivemetastases
were seenin the left
femur and pelvic bones.Pathological
fractureof the neck of the left femur
was treated surgically.Further X-ray cxaminationsrevealed extensive
metastases
of the femura,and pelvicbones,with multiplelung metastascs.
When the paticnt came undcr our carc, he was sufferingagonizingpain
in the left hip. An X-ray cxaminationshowedan almostcomplete
especially
disappearance
of thc upper part r)f thc lcft fenrur,with multiple metastases
in the right femur and pelvicbones,and multiplenretastases
in both lungs.
Accordingto the analyses,
the patientwas treatedwith perselenide,
bixine,
and epichlorohydrin.The condition improvcd rapidly, the patient being
ableto sit up and evento walk a little. Hc rvasdischarged
from the hospital
to follow the treatmcntat home which was done very irregularly.He was
readmittea
d n r o n t hl a t c r w i t h v e r y s e v e r ep a i n a n d t h e t r e a t m e nrt e s u m e d .
g r a d u a l l ya n d t h e g e n c r acl o n d i t i o ni m p r o v e dm a r k e d l y .
The painsubsided
RccentX-rays revealeda manifestrecalcification
of the upper part of the
s ,a d n o l o n g e rb e e nv i s i l e f t f e m u rw h i c h ,i n p r e v i o u sX - r a y e x a m i n a t i o n h
ble. At the sametimc, many of the metastatic
lesionsof the lung disappeared
while in others,a markeddecreasein their sizewas seen.Thesesubjective
and objectiveimprovementsare continuingconstantly,up to date.
F . G . , 6 1 y e a r o l d f e m a l c .i n J u l y 1 9 6 0 h a d p a r t i a lc e c o s t o m yf o r
The mcsentericlymph nodes were found involved. A
adenocarcinoma.

P R E S E N T F ' o R M o l ' T R E A T M E N T

5 4 1

month later, becauseof vaginalblecdingand an erosionof the cervix, a

biopsywas performed,showingthe samemalignancy.with constantbleeding and pain in the abdomen,the paticntcameunderour care.On examination the tumor was seento occupy all the upper part of the vagina,with
infiltrationof the recto-vaginal
wall. Accordingto the analyses,
a treatment
rvith perselenidein injection,epichlorohydrinand bixine was instituted.
The bleedingstoppedcompletelyaftcr onc w,eck,as did the pain. While
t h e p a t i e n ti s s t i l l u n d e r t r e a t m e n tt,h e l e s i o nh a s b e e n s e e n t o r e g r e s s
constantJy,
up to date.
W . M . , 4 8 y e a r o l d m a l c . I n 1 9 - 5 6h e h a d a l e f t n e p h r e c t o m yf o r
h y p e r n c p h r o m aH.e w a s w e l l u n t i l e a r l y i n 1 9 6 0 w h e n a m a s sw a s f o u n d
i n t h e l e f t s i d eo f t h e a b d o m e nw h i c h w a s p r o g r e s s i v e g
l yr o w i n g .A t t h e
s 3 m et i m e ,h e h a d h e m o p t o i cs p u t u m .T h c x - r a y c x a m i n a t i o no f t h e c h e s t
s h o w e dm u l t i p l e l u n g m e t a s t a s e F
s .i v e w e e k sb e f o r ec o m i n g u n d e r o u r
c a r e ,a v e r y m a r k c de d e m ao f t h e l e f t l c g w i t h v c r y s e v c r ep a i n i n t h e b a c k
a n d l e g a p p e a r e dH
. e c a m e u n d e r o u r c a r e e s p e c i a l l f. oy r t h e u n b e a r a b l e
p . r i n .A c c o r d i n gt o t h c a n a l y s e sa. t r c a t m c n tw i t h h c p t a n o l ,b u t a n o la n d
g l y c e r o lw a s i n s t i t u t e dT
. h e p a i n d i s a p p e a r c idn 2 - 3 d a y s a n d h a s n o t
r e t u r n e di n t h e t w o m o n t h sw h i c h h a v ee l a p s e ds i n c et h e n .T h e e d e m ao f
t h e l e g a l s od i s a p p e a r e dT.h e t u m o r w h i c h ,a t t h e t i m e o f a d m i n i s t r a t i o n ,
was occupyingthe entirc spacebctwecnthc ribs and thc ileac crust, was
seento becomefirst much softer.and progressively
to reduceits dimension.
A c t u a l l yt w o m o n t h sa f t e rt h e t r e a t m e nw
t a s s t a r t e dt.h e p a t i e n ti s l e a d i n g
a n o r m a l I i f c w i t h t h c t u m o r d c c r c a s i n pg r o g r e s s i v e l y .
We want to emphasizethat benefits,often impressiveeven in terminal
cases,have been obtainedonly by following thc above rules. Treatment
guided closelyby changesobservedin the patternsindicatedby analyses
appearsto be the conditionsinequa non f.orthe attainmentof good results.
, venin far
T h e r e s u l t so b t a i n e da n d e s p c c i a l l tyh e i r h i g h p r o p o r t i o n e
advancedcases,permitsa fair judgementof the placc of the presentform
of applicationof this method in the fight againstcanccr. Basedon these
rcsults,we are fully entitled to considerit, not only a highly beneticial
treatmentwhich can be offercd now for this disease,but even a major
step nearer to the solution of the problcm of the therapy of cancer.

NOTES

C h a p t e rI , N o t e l . S u b n u c l e a O
r rganization
T h e a n a l y s i os f t h e a v a i l a b l e
d a t ac o n c e r n i nsgu b n u c l c apra r t i c l c ss h o w s
that while an unsuspccted
number of diffcrentparticlesari progressively
discovered,no satisfactoryrelationshipbetweenthem-from the point of
view of the organization---can
be established.
The rccognitionof a pattern
c o n c e r n i n gt h e i r o r g a n i z a t i o n ar le l a t i o n s h i p
w o u l d f i l l a n i m p o r t a n tg a p
in the knowledgeof this cntire field.Thc fact that the samepatterngoverns
t h e o r g a n i z a t i o ni n g e n e r a l ,f r o m a t o n r i cn u c l c i u p , h a s i n d u c c d u s t o
a t t e m p tt h r o u g ha n c x t r a p o l a t i o nt,o s c a r c hi t f o r t h e s u b n u c l e a re a l m .
A s s e e na b o v e .t h e s t u d yo f t h c o r g a n i z a t i ohna s p c r m i t t c du s t o d c f i n c
t h e f o l l o w i n gc o n c e p t sa s c h a r a c t e r i s tfi o
cr the organizationa
p la t t e r n :
I ) A l l t h c e n t i t i e si n n a r u r ec a n b e i d e n t i f i e db y t h e i r p l a c e i n a
h i e r a r c h i co r g a n i z a t i o ni,n w h i c h t h c c n t i t i c sa r c c o n n e c t c dt h r o u g h a
s u p e r i o r - i n f e r i or re l a t i o n s h i pA. n e n t i t y e n t e r si n t h e f o r m a t i o no f o t h e r
e n t i t i e sw h i c h a r e c o n s i d e r e d" s u p e r i o r "t o i t . a n d i s f o r n r e db y e n t i t i e s
w h i c h a r e h i e r a r c h i c a l l"yi n f e r i o r " t o i t .
2 ) E a c h e n t i t y i s f o r m e d b y a p r i n c i p a la n d a s e c o n d a r yp a r t , t h c
p r i n c i p a lp a r t b e i n g r e p r e s e n t ebdy e n t i t i e sh r e r a r c h i c a l liyn f e r i o r t o i r ,
r v h i l et h e s c c o n d a r yp a r t , b y c l c n r c n t st a k e nf r o n r t h c i m m e d i a t cc n v i r o n m e n t i n w h i c h t h e e n t i t i e sf o r m i n gt h e p r i n c i p a p
l a r t h a v ee x i s t e d .
3 ) E n t i t i c sw i t h s i m i l a rp r i n c i p a lp a r t sb c l o n gt o a s a m cl c v c l . I n t h c
h i e r a r c h ipc r o g r c s s i o nt h, e r ea r ee n t i t i e so f t h es a m el e v e lw h i c ha r eg r o u p e d
t o g e t h etro c o n s t i t u t e
t h e p r i n c i p a lp a r t o f a n e n t i t yo f a l e v e li m m e d i a t e l y
superior.
4) From the energeticpoint of vicw, thc principalpart in the organization of eachentity appearsmore positivethan the secondarypart to which
it is bound.
5 ) The hierarchicprogressionof the organizationfrom one entity to
t h a t i m m e d i a t e l ys u p e r i o rt o i t , i s m a d e t h r o u g ht w o p r c r c c s s ewsi t h t w o
d i f f e r e n ti n t e r v e n i n gf o r c e s .F o r c c so f c o l u m b i a nn a t u r e b r i n g a n d k e e p
the electrostatically
opposite principal and secondaryparts together,A
n e w e n t i t ya p p e a r sh o w e v e ro n l y w h e nq u a n t u mf o r c e si n t e r v e n eo, r g a n i z i n g t h e r e l a t i o n s h i op f t h e t w o c o n s t i t u e n tasn d e s p e c i a l l tyh e i r r e c i p r o c a l
m o v e m e n tT
. h e i m m e d i a t ea i m o f t h i s o r g a n i z a t i o ni s t o p r e v e n ta r e c i p r o c a lt o t a l a n n i h i l a t i o no f t h e t w o p a r t s ,p o s i t i v cp r i n c i p a la n d n e g a t i v e
secondarywhich would occur if the electrostaticforces alone would be
present.
6) In the hierarchicprogrcssivcdevclopmcntof the organizationfor
543

544

n . E s E ^ R C Hr N p H y s r o p A T H o L o c y

eachpassage
from one levelto the immediatelysuperior,thesetwo kinds of
forces---electrostatic
and quantum-were seento intervenealternatively.The
fulfrllmentof one force is seento inducethe appearanceof the other. From
an energeticpoint of view, an entity will appearinactivewhen its electrostaticforcesare fulfilled but with quantumforcespresent,or energetically
active with quantum forces fulfilled and the electrostaticforces present.
The exampleof atomsand ions is a typicalillustrationof this relaiionship
for the atom level.
As a work hypothesis,we tried to apply the above schematically
presentedconcept of hierarchicorganization,to the subnuclearrealm. lt
is only by using analogiesthat such an attemptcan be made. The scarce
data availableseem to confirm however,this view. According to it, the
electronand positronwould representthe lowestentitiesof the subnuclear
realm with the smallestmassand oppositecharge.If thesetwo corpuscles.
when attracted, encounter one another, they will annihilate each other
with liberationof two photons.This annihilationis preventedhowever,
althoughthe two corpusclesremain bound togetherthrough their electrostatic forces by the interventionof quantum forces organizingtheir reciprocal movements.The rcsult is a new entity, of a level immediately
superiorto the positron,in which the electronis kept electrostatically
bound
to the positronbut kept into an orbital movement.Hypothetically,it can
evenbe conceivedthat throughdifferences
in the resultingmovement,more
than one solutionwould exist.
Residual Clurge
Due to the interventionof this movement.the resultingelectrostatic
neutralizationbetweenpositronand electronis incomplete.A "residual"
positivechargewould characterize
the new entity.This chargealone would
not be sufficientto keep anotherelectronby neutralizingits charge.However severalsuch entitiesgrouped togethercan have the sum of their
"residua.lpositivecharge" such as to be compensated
by a new electron.
The two electrostatic
forces,that of the groupof entitiesand that of the new
electronwill keep thesetwo parts together,while the quantum forces will
again organizethe movementof this new added electron,preventingthis
t i m e a g a i nt h e a n n i h i l a t i o o
n f t h e s et w o p a r t s .A n e w l e v e l ,t h i s t i m e o f t h e
third order, is thus realized.It is casyto conceivethat severalsolutionscan
existfor each case,sincethe sum of the residualpositivechargesdoes not
Severalsolutionsappear
correspondexactlyto that of thc negativeelectrons.
thus possible.Besidesthis in which a small group of entitieswould be
by one electron,a higher number of entitieswould be kept
compensated
togetherwith the sum of their residualchargesapproachingthat of two
or more electrons.For each level,severalsuch solutionsare conceivable.
passage
toward higherlevels,the number of the soluWith the progressive
tions increases.
positiveand negativeparts of the
The fact that the two electrostatic
entitiesdo not compensateperfectly,leadsto the possibilitythat the compensationtakes place either with an excessor lack of negativecharge.

The differenceis induced by an acrditionarerectron.:"::r,r

:Jt:
positiveor negativechargewould result.with one or the other chargeprev.ailing,two energeticallyactive, positiveor negativeforms, ditrerlng by
the massof an electronexist,for eachentity.The analysisconfirmsthii existence.
Accordingto the hierarchicorganization,the differcntparticlesof the
subnuclearrealm can be separatedin variouslevels.promezons,mezons,
protons would representsuch levels-cach onc with respectiveneutral,
positiveand negativeentities.The same as for the higher lsygl5-3tern5
and molccules-the differententitiesformingeachof the subnuclearlevels
w i l l d i f f e rt h r o u g ht h e n u m b e ro f t h c e n t i t i e sc n t e r i n gi n t h e p r i n c i p a lp a r t s .
A systematization
of the subnuclearrealm on this basiscan be conf,rmed
b y t h e f a c t t h a t t h e d i f f e r e ne
t n t i t i c so f a l e v c lr c p r c s e nst u m so r m u l t i p l e s
o f t h ee n t i t i e so f t h e i ri n f e r i o rl e v e li n t h e s a m ew a y t h a t t h e d i f f e r e n nt u c l e i
r c p r e s e nm
t u l t i p l e so f e n t i t i c so f t h c p r o t o n i cl e v e lp r o t o n sa n d n e u t r o n so r
t h e d i f f e r e n tm o l e c u l em u l t i p l eo f c n t i t i c so f t h e a t o m i cl e v e l .
I n t h e p r o g r e s s i o on f t h e h i c r a r c h i co r g a n i z a t i o ni t, i s s e e n t h a t t h c
passagefrom one level to the other resultsin an exponentialincreasein
t h e n u m b e r so f t h e k i n d s o f e n t i t i c s( f r o m a r o u n d 1 0 0 d i f f e r e n ta t o m si t
p a s s e tso a r o u n d1 0 0 , 0 0 0d i f f e r e n kt i n d so f m o l e c u l e st o
, m i l l i o n so f k i n d s
o f g e n e sa n d t r i l l i o n so f i n d i v i d u a l sT. h i s f a c t s u p p o s e tsh a t t h e n u m b e r
o f t h e e x i s t i n gp a r t i c l e sd e c r e a sw
e i t h e a c hi n f e r i o rl e v e l ,i n t h e s u b n u c l e a r
rcalm,to arriveto two-positron and clectroi-nt the bottomof this realm.
C h a p t e r2 , N o t c l . C - N - C - N
As far back as 1905, Kosscl has had indicatedthe existencefor the
i m p o r t a n ta l k a l i n e a m i n o a c i d sa. r g i n i n ea n d h i s t i d i n e ,o f t h e C - N - C - N
g r o u p , f o u n d a l s o i n t h e n i t r o g e nc o n t a i n i n gb a s e so f n u c l e i ca c i d . T h e
hypothesiswhich we advancethat this C-N-C-N group would representthe
s t a r t i n gp o i n t o f t h e b i o l o g i c arl c a l m i t s c l f ,c a n s u r c l yb e s u b j e c t o d i s c u s sion.
Progressively
more evidenccis bcing obtainedthat importantorganic
compoundscan appearfrom the constituents
itself,under
of the atmosphere
or of ionizingirradiation.While Henthe influenceof electricaldischarges
riet (268) was the first to show that formic acid is presentin rainwater,
it was Loew (269) who obtainedglycine from the constituentsof the
By utilizing,under the same
atmospheresubmittedto electricaldischarges.
t t the
c o n d i t i o nm
, i x t u r c ss i m i l a rt o t h o s ec o n s i d e r etdo h a v eb e e np r e s e n a
time when life is supposedto have started,Miller obtained,through elecmany amino acidsand other substances
especiallyglycine
trical discharges.
(
2
7
0
,
w
e
r
e
2 7 1 ) . M i l l e r ' sr e s u l t s
e x t e n s i v e l cy o n f i r m e d
and formic acid
(272 to 279).
The inadiation of the mixturesof gasesconsideredpresentin the atmospheremillionsof yearsago has led to the synthcsisof many other subs t a n c e s u c h a s f o r m i c , a c e t i c ,p r o p i o n i c ,s u c c i n i ca n d e v e n t r i c a r b o x i l i c
acids (280, 287). From these,we considerof especialimportancethe

nEsEARcH rN pHystopATHoLocy

546 /

memberswith a secondnitrogen group far in the molecule, as diaminosuccinicacid, iminodiaceticor iminoacetic-propionic

acid.
The synthesisof the strongjy positivec-N-c-N group which we consider as the startingpoint of the biologicalrealm, seemsthus to have taken
place rather under the influenceof radiation. This fact appearsespecially
important sinceit would relatemorc directly the beginningof tne Uiotoglcat
realm to the interventionof the radioactiveelements,which according to
our systematizationof the elements,form the period which corresponds
to the fowest levelsof the hierarchicorganization.(see chapters 2 od 5)
ALKAII AI{IGAC|DS
Arglnlnc

Frc.,,,r-ot-&**
Hlsttdtnc

@'-@,*

l'ilIR0GEmt6
8ASS
Prlncs

Adeninc

furlrtdincs
Thplnc

ol
I

ot
61zcrr,
,\/
H
Cytostrr

tfu-ot-o,z-o,Sro.,

ffi
H

F l c . 2 0 1 . T h e N H , a n d c - N - c - N g r o u p sa p p e a ra s e n t i t i c st a k i n g p a r t i n t h e f o r m a t i o n o f a l k a l i n e a m i n o - a c i d sa s w e l l a s o f n i t r o g e n o u sb a s e s .T h e b o n d t o a c h a i n
h a v i n g a n a m i n o a c i d g r o u p i n t h e f i r s t c a s e ,r e s u l t s i n a n e w e n t i t y - a n a l k a l i n e
amino acid-which polymerizesthrough the amino-acid group, Through the alkalinc
group it conserves its positive electrical character. In the nitrogenous bases, the
C-N-C-N group is part of the cycle. Bound to phosphoric acid, the results are acid
e n t i t i e sw i t h n e g a t i v ec h a r g e s .

The furthcr evolutionof the C-N-C-N formation seemsto have taken

place in two directions----one
in which one or two such groups have formed
a cycle and given rise to the nitrogenousbases,purinesand pyrimidines,
and the other in which this energeticgroup has bound an aliphaticamino
acidchain,this lastprobablyoriginatedundcrthe influenceof electricaldischarges.The two principal alkaline amino acids, arginine and histidine,
h a v et h u sa p p e a r e d(.F i g . 2 0 l l T h e d o u b l ec a p a c i r yo f t h e a l k a l i n ea m i n o .
acids,to bond other amino-acidsthroughtheir amino-acidgroupsand thus
to form polymers,and to bond acid substances
throughtheir alkalinepolar
groupsand make new hierarchicentities,has given thesesubstances
their
peculiarorganizationalrole, C-N-C-N, alkaline amino-acidsand histones

NOTES

541

( o r i n f i s h ,t h e p r o t a m i n s )w o u l d t h u s r e p r e s e ntth e f l r s t h i e r a r c h i cs t e p s
i n t h e p r o g r c s s i oonf t h c b i o l o g i c asl c r i c s .
A f e w w o r d ss h o u l db e s a i da b o u tl y s i n c .t h e a l k a l i n ea m i n o - a c i dw i t h
a n a m i n og r o u p a s a l k a l i n et e r m i n a g
l r o u p .A l t h o u g ht o g c t h c rw i t h t h c
o t h e r a l k a l i n ea m i n o - a c i d si t, e n t e r si n t h e f o r m a t i o no f h i s t o n e si,t s e e m s
t o h a v e a n o t h e ri m p o r t a n tb i o l o g i c arl o l c , t h a t o f a n a g c n ti n r c r v c n i n gi n
t h e m e t a b o l i s mo f l i p i d s .
Chapter2, Note 2. Distribution o[ Potassiumand Sodiunr
A s m e n t i o n e da b o v c , p o t a s s i u mi s t h c c a t i o n o f t h c c y t o p l a s m t, h e
secondarypart of cells, while sodium is the cation of the secondarypart
of the metazoiccompartment,that is of the lluids of this compartment.
A c c o r d i n gt o t h e v i e w p r c s e n t e da b o v e .t h e p e c u l i a rd i s t r i b u t i o no f t h e s c
t w o c a t i o n si n t h e b i o l o g i c a rl e a l m r c s u l t sf r o m t h e i r s i m i l a rd i s t r i b u t i o n
i n t h e e n v i r o n m e n tfsr o m w h i c h t h c s cr e s p e c t i vsec c o n d a r yp a r t s ,c e l l u l a r
a n d m e t a z o i c a, r c c o n s i d e r c dt o h a v c b e e n d e r i v e d .A s w e r e l a t e dt h e
c y t o p l a s mt o m u d , r c s p e c t i v c ltyo t h e l i t h o s p h c r ca. n d t h c f l u i d s o f t h c
m e t a z o i c o m p a r t m e nt to t h e s ea , w e I o o k e df o r a c o n f i r n r a t i oonf t h i s v i e w
i n t h ec o m p a r i s o b
n e t w c c nt h c a m o u n to f t h c s ec a t i o n si n t h c t w o b i o l o g i c a l
c o m p a r t m e n t as n d i n t h e t w o e n v i r o n m e n tw
s h i c h w e c o n s i d e rt o c o r respondto them.
A l t h o u g hp o t a s s i u ma n d s o d i u n ta r c i n a l m o s te q u a l a m o u n t si n t h c
g e n e r acl o n s t i t u t i o o
n f t h e c a r t h ' sc r u s t .p o t a s s i u ni s) f o u n d a l n r o s e
t ntirely
i n t h e s o l i d p a r t sw h i l e s o d i u mf o r n r st h e p r i n c i p a cl o n s t i t u e not f t h e s a l t s
o f t h e f l u i d p a r t o f t h e c a r t h .T h c d i s t r i b u t i o n
of potassium
foundbetween
c c l l s a n d e x t r a c c l l u l afrl u i d ss e e m sv e r y n e a r t o t h a t w h i c h e x i s t sb e t w e e n
l i t h o s p h e raen d h y d r o s p h c r cP. o t a s s i u m
i s f o u n di n a p r o p o r t i o no f 2 . 4 6 0 / o
o f t h e l i t h o s p h e raen d o n l y i n 0 . 0 4 f t o f t h c h y d r o s p h e r(e2 O l ) . T h e r a t i o
o f t h c s er e s p c c t i vceo n c c n t r a t i o ncso r r c s p o n dtso a r i ; , v a l u e .T h i s s e e m s
n e a re n o u g ftro t h e r a t i of o u n di n b i o l o g y W
. h i l et h e e x t r a c e l l u l apro t a s s i u m
r e p r e s e n tosn l y 5 m E q p e r l i t c r , w i t h a t o t a l o f 7 0 m E q ( 2 . 7 g m . ) f o r a
n o r m a l b o d y , t h e i n t r a c e l l u l apr a r t c o r r e s p o n dtso I l 5 m E q p e r l i t e r o f
c c l f s ,w i t h a t o t a lo f 4 . 0 0 0 m E q ( 1 6 0 g m . ) f o r t h e b o d y ( 2 0 2 ) . T h e r a t i o
o f t / r , , f o r t o t a l e x t r a c c l l u l aar n d i n t r a c e l l u l airs a c c e p t e dt o d a y a l t h o u g h
g e n e r a l l cy o n s i d e r etdo o h i g h w h e nc o m p a r c dw i t h t h e p r e v i o u sd a t a g i v e n
y e a r sa g o b y S h o h l ( 2 6 5 ) . T h i s v a l u eo f r . , , a p p c a r si m p r e s s i v e n
l ye a r
t h e r a t i o o f l , rr; f o u n d i n t h e c o m p a r i s o no f t h e p o t a s s i u mc o n t e n to f t h e
lithospherw
e ith that of the hydrosphere.
d h c n c o m p a r i n gt h e p r o p o r t i o n
A s i m i l a rr e s e m b l a n ci es e n c o u n t e r ew
b e t w e e ns o d i u ma n d p o t a s s i u mi n t w o l l u i d s : t h c i n t c r s t i t i afl l u i d o f t h c
. sp p c a rc l o s ee n o u g h .
b o d y a n d o f t h e s e a .T h c t w o r a t i o so f t h e s ee l e m e n t a
g
m
.
h
a
s
1
0
.
4
6
4
o
f
s
o d i u mp c r t h o u s a n d
A
t
l
a
n
t
i
c
Ocean
F o r i n s t a n c et,h e
, h i l e t h e P a c i f i cO c e a n h a s
a n d 0 . 7 2 5 g r . p e r t h o u s a n do f p o t a s s i u mw
1 0 . 2 3 3g r . p e r t h o u s a n do f s o d i u ma n d 0 . 6 3 4 g r . p c r t h o u s a n do f s o d i u n r
( ? 6 6 ) . T h e r a t i o sb e t w e e ns o d i u ma n d p o t a s s i u m
a r e r c s p e c t i v e lly4 a n d
w
h
e
n
t
h
e
a
v e - r a gies c o n s i d c r c da s
i
s
l
6
1 6 , I n t h e b l o o d s e r u m ,t h e r a t i o

548

nESEARcHtN pHysropATHoLocy

3 2 0 m g . o / ao f N a , a n d 2 0 m g . ( / ro f K . T a b l e X X V s h o w st h e s ec o m p a r a tive values.
T n s L EX X V
(a) Comparison
b e t w e e nt h e E x t r a c e l l u l aarn d I n t r a c e l l u l aPr o t a s s i u m
and the amount presentin Hydrosphereand Lithosphere.
Rstio
Potassiumextracellular-total
body 2.7 gm. )
P o t a s s i u mi n t r a c e l l u l a r - r o rbaol d y 1 6 0 g n r . )
Potassiumin the hydrosphere 0.04ch )
Potassiumin the lithosphere
2.46% )

1,59
Ii6l

( b ) P o t a s s i u ma n d S o d i u m i n t h e s e a a n d i n t h e body fluids.
Na
A t l a n t i cO c e a n 1 0 . 4 6 . 1 g r . 0 0
P a c i f i cO c e a n
1 0 . 2 3 3g r . 0 0 0
Blood Serum
320 nrer. 0 0

K
0 . 7 2 5g r . 0 0 0
0 . 6 3 4g r . 0 0 0
2 0 m g r . 0 , ,0

Ratio
14.4
16.0
16.0

T h e s ev a l u e ss e e mt o b i n d t h e d i s t r i b u t i o no f K a n d N a , s e e n b e t w e e n
the cellularand metazoiccompartments,
to that which existsbetweenthc'
environmentsfrom which we considertheserespectivesecondaryparts to
have been derivcd.
Chaptcr 2, Note 3. Social Hierarchic Organization
The organizationof nature with the characteristichierarchic structure
so evidentin the biologicalrealm, has suggested
a similar structurein social
organization.
All the factors which were seento characterizehierarchicorganization
a p p e a rc l e a r l yi n a n a n a l y s i so f s o c i a lo r g a n i z a t i o n( F
. is. 202)The entity,
immediatelyabove that of the individual,is the family. Here, parents and
children-as a grouping of entities of the same level form the principal
part. The secondarypart is made up of elementsof the immediateenvironment, which are kept organizedaround this principal part, and as such are
integratedin the new entity, the family. Housing,goods,even psychological
factors, ideas and habits, characterizethese added factors. A boundary
formation is often much more visible than expected.Living quarters and
comrnon possessionsare well delineated,and charactertznthe farnily. As
expected,most of them are not consideredto belong to an individual but
the family as entity. "This is family property" is a common expression.
Almost always,numerousfamilies are grouped in nearby dwellings.
althoughthis fact alone does not lead to the immediatesuperior entity. the
community.When the group of familiesorganizestogetherand limits certain possessions
taken from the environment,as common to the group.
the entity "community"appears.The principalpart is madeup of the group
of families,the secondaryby the material and even moral goodswhich are
attachedto the goup of families in common. The community has prop-

NorEs

549

erties which belong only to the community-streets, for example-as it

has, by definition, a boundary. The limits of thesesocial entities are well
definedand thesethree factors-principal part, secondaryadded part from
the environmentand boundary-characterize these entities as they character'unthe entities in the entire biological realm. The same pattern applies
f o r t h e c o u n t yw h e r eg r o u p so f c o m m u n i t i efso r m t h e p r i n c i p a lp a r t , a n d

lndividualsG
' oods--

Families'Goods
- ---rt.
C o m m ui e
n si t' G o o d s ' C o u ni et s ' G o o d_,/_
s
S t a t e s 'G o o d ts'

i l a t i o n ' sG o o d s
l n t e r n a t i o n aGl o o d s -

In d i v i d u a l s
F a mI i e s
C o n m u inei st
C o u ni et s

- States

N aito n s
H u m aSno c i e t Y

F r c . 2 0 2 . T h e s c , c ' i ohl i e r u r c h i co r g u n i z u ! i o nf o l l o w s t h e s a m e p a t t c r n a s t h e o r g a n i z a t i o n o f m a t t e r o r o f t h e b i o l o g i c a l r e a l m . E a c h e n t i t y r e s u l t sf r o m t h e b o n d o f a
group of lower entities with a secondarypart taken from the environment and limited
by a proper boundary.

proper parts taken from the environmentand common only to this new
entity form the secondarypart. This entity also is defined through its
boundary. It is easy to see how through the same tuerarchic pattern we
passfrom countiesto states,nations,hemispheresand world which reprehigher hierarchicentities.It is interestingto see how, in
sent successively
which we have
each one of thesesocialentities,the samemanifestations
found to characterizethe biologicalentitiesalso exist. The relationship
betweenentities and especiallymany of their functions shows that the

550

RESEARCH IN

PHYSIOPATHOLOCY

social entities are not artificial mental concepts,but are the result of the
interventionof the same forcesin which heterotropicorganizationopposes
the lawlesshomotropy. It has appearedinterestingto seehow much of the
knowledgeof the physiologicaland especiallythe pathologicalmanifestations of the lower entities,we can apply to understandmanifestationsoccurring at the social hierarchiclevels.
Under this aspect,sociologyfinds a new basisnot only for the analysis
of many of its problems,but can have an insight to how nature,through its
own organization,has tried and often succeededin resolving problems.
With the concept of unity in all organization,from subatomic to social
entities,we can understandhow the evolution of the environment,representedby material and intellectualgoods, can produce changesin social
entities.The conceptof highersocialentities,organizedso as to conserve
the characteristicsof the lowest social entities,gives a new aspct to the
relationshipbetween individual, family and society. A scienceof social
physiologycan be createdby systematizinghierarchicsocial entities much
as we did for entitiesin the biologicalrealm. The same approachcan be
applied to social pathology and social therapy as well. Such an approach,
will be the subjectof other presentations.
Chapter 3, Note 1. PrecancerousLesions
Precancerous
lesionswere identifiedespeciallyin casesin which cancerous lesionswere inducedand wherea manifestpolycentricityof lesionswas
present.(203) Polycentriclesionspermit us to study the entiresuccessive
changesfrom normal to invasivecancer.
Induction of cancer in the stomach of rats through carcinogensand
detergents(204) has furnishedexcellentmaterialfor such study; it has
also permitted us to characterizethe specificchanges.Among cells which
appeargrosslynormal, there are somein which certain morphologicalcharacteristicsof the nucleus,notably size and form, appear abnormal. The
existenceof an anomaly is much more evident when the cell divides. lt
may be limited to just a few chromosomeswhich are abnormal in their
dimension and form. This chromosomialabnormality appea.rsstill more
evidentwhen comparedto cellsin mitosisin controlswith normal mucous
membranes.( 309 )
Chapter 3, Note 2. Non-invasiveCancer
We have emphasizedthe characterof the cytoplasm of the cells in
non-invasivecancer,The nuclei show a number of changeswhich, together
a cancerousentity: an irregular shape
rather than separatelycharacterize
of the nucleus with a manifest increasein size; a sharp nuclear border
formed by a dark pigmentednuclear membrane having fine chromatine
particles; a hyperchromatism with clumps of chromatin separated in
bizarre, irregular fashion; and an uneven, irregular distribution of these
chromatin clumps, concentratednear the nuclear membrane. Also often

NOTES

,,, 55I

encounteredis the presenceof one or more irregularenlargednucleoli,with

a distinctnucleolarborder and especiallywith a manifestacidophilicstaining.
In the non-invasivecancer, all these nuclear anomaliescontrast with
the relativelynormal cytoplasm,which has not only an acidophilicreaction
--+olored in orange with Papanicolau'strichromic staining-but also a
welldefined cell membrane with fairly clear cell border. The size of the
cytoplasm----compared
to other cells-is normal, although the nuclearcytoplasmicratio is increaseddue to the big nucleus.Due to the character
of the cytoplasm,thesecells were calledthe "third type difterentiatedcells"
by Graham. (205 ) Wc emphasized
the "normal" aspectof the cytoplasm
of thesenon-invasivecells, in contrastto the invasivecells where the abnormality includesboth the nucleusand the cytoplasm.This explainswhy
most of the invasivecellshavelittlc cytoplasm,an indistinctcellularborder
a n d a b a s o p h i lc y t o p l a s m i cs t a i n i n g .( 2 0 6 ) B u t b e s i d e st h e s ec e l l s w i t h
totally abnormalcytoplasm,there are someinvasivecellswith an apparently
difterentiatedcytoplasm. Although their staining is orthochromatic,their
cytoplasmshowsmarked abnormalityin form, The tadpolecells found in
the exfoliativecytologyin epidermoidcarcinoma,(207) or the fiber cells
(208) with abnormallylong fibrillar cytoplasmrevealedin other forms of
invasivecancer indicate a participationof the cytoplasmin the abnormality. The cells found in so-called"Bukhead's disease"with minimal abnormal cytoplasmthus appearsto be at the boundarybetweennon-invasive
and invasivecancerouscells,
Chapter 3, Note 3. Abnormal Arnino Acids
We have seenabovehow the conceptof hierarchicorganizationbrought
us to considerthe alkaline amino acidsand the histoneswhich they form as
one of the first membersof the biological realm. Anomalies can be conceived to result from a processof rcsonancewhich occurs constantlyon a
statisticalbasis,As work hypothesis,we considersuch resonanceentities
as correspondingto these abnormalforms, which in hierarchicdevelopment would lead to cancerousentities
The naturallyexistinglevorotatoryalkalineamino acids representthe
which, throughtheir number and role in further organization,
constituents
representnormal entities. Opposed to them, the dextrorotatory alkaline
amino acids would representabnormal entities.Their existenceand their
without, however,bringing
role has made the object of many discussions
sufficientlight to this problem. The constantexistencein the body of
specificenzymesagainstthese dextrorotatoryamino acids in spite of the
fact that they are not recognizableanalytically,indicatesa certain defense
as a resonancephenomeagainstthem, The conceptof their appearance
amino acids,alDextrorotatory
easily
this
occurrence.
non would explain
though abnormalfor the organism.exist in practicallyall individualsas a
resonanceform, but they are not able to develo5--or developin extremely
reducedform-because of the enzymeswhich attack them. They are, how-

-552 .'

RESEARCH tN

pHystopATHoLocy

ever, able to developthe lowest levelsof cancerousentitiesas they are

recognized
to exist practicallyin all normal individualsparticularlyafter a
certainage.There are theseconsiderations
which lead us to believethat it
is the dextrorotatoryresonanceform of alkalineamino acidswhich representthe abnormalentitiesat the lower levels.
Chapter 4, Note l. Physiologicaland PathologicalPain
PhysiologicalPain
Physiologicalpain may be definedas a specificsensorialsensationinduced in normal tissueswhen externalstimuli arc applied rvith sufficient
intensityto endangertissueintegrity.Becausepain may be induced by a
w i d ev a r i e t yo [ s t i m u l i ,i t h a sn o t a l w a y sb e e na c c c p t e d
a s c o n s t i t u t i nag s e n s o r i a l s e n s a t i o nC. o n s i d e r a b l e v i d e n c ce x i s t s ,h o w e v e r .t o i n d i c a t et h a t
physiological
pain is a specificsensation,
similar to the other sensorialsens a t i o n sO
. n e i n d i c a t i o nt h a t p a i n i s a p r o p c r s e n s o r i asl e n s a t i o lni e s i n t h e
fact that its hasits specialnervesystem.
B l i x ( 5 ) a n d G o l d s c h e i d e(r6 ) f o u n d t h a t c e r t a i na r e a so f t h e s k i n
were sensitiveto painful stimuli from a pinprick while others were not.
S t r u g h o l d( 7 ) d e m o n s t r a t e tdh a t , i n v a r i o u sa r e a so f s k i n , p a i n p o i n t s
were concentratedin varying degrce.Microscopicstudy of areasof skin
showinghigh aggregations
of spotsof spccificforms of sensibilityhas indicatedthat spccialsensorynerveand organ structuresare apparentlyassoc i a t e d w i t h d i f f e r e n tt y p e s o f s e n s a t i o nT. h u s , K r a u s e ' sc o r p u s c l c sa r e
consideredas reccptorsfor cold; Rullini's cndingsand Golgi-Mazzonicorp u s c l e sf o r h e a t ;a n d M e i s s n c r ' cs o r p u s c l c sM, e r k e l ' sd i s c sa n d t h e b a s k e t
c n d i n g sa r o u n dh a i r r o o t sf o r t o u c h .( t l ) W o o l a r d( 9 ) d e s c r i b e u
dnmyelinated,finely bcaded,branchedfree endingsas the specificnerveend organs
believedto bc rcsponsible
for thc reccptionof pain impulses.Certain areas
such as thc corneaand the mucousmembraneof the nose,which are gcne r a l l yc o n s i d e r e d
s c n s i t i v et o p a i n a l o n c ,h a v e b e e n s h o w n t o h a v e t h e s c
f r e ee n d i n g sa s t h c c h r r a c t c r i s t incc r v ec n d i n g sa t t h e s es i t e s .W e d d e l lh a s
f o u n do n l y t h i s t y p eo f e n d s t r u c t u r ei n a r e a so f s k i n s e n s i t i vseo l e l yt o p a i n
d u r i n gn c r v cr c g c n e r a t i o n .
1'hatpain constitutes
a specificform of sensation
is further indicatedby
the evidcncethat its impulsesare carricd along dcfinitenerve pathwaysto
s p e c i a cl e n t e r si n t h e t h a l a m u sB
. y t e m p o r a r ya s p h y x i ab, y c o c a i n i z a t i o n ,
o r b y c o o l i n gd
, i f f e r e n t i ai nl t e r f e r e n cwc i t h c o n d u c t i o no f t h e s p e c i a sl e n s a tions of pain, touch, hcat and cold along a nerve can be produced.Thc
e x i s t e n co
e f i n d i v i d u a l sw i t h o u tt h e s c n s a t i o o
nf pain,but with sensations
of touch,cold and heat, has confirmedthis view of pain as a proper sens o r i a ls e n s a t i o n .
I t i s c h a r a c t c r i s to
i cf t h e s e n s a t i oonf p a i n t h a t i t m a y b e e l i c i t e db y a
w i d e v a r i e t yo f s t i m u l i .B c l o w t h e p a i n t h r e s h o l d t, h e i n c i t a t i o ni n d u c e s
s p e c i f i cs e n s o r i a sl c n s a t i o nas c c o r d i n gt o t h e s t i m u l u su s e d . A b o v e t h i s
t h r e s h o l dt ,h e s e n s a t i ofnc l t i s p a i n .W h e nd i f f e r e nnt o x i o u ss t i m u l ip r o d u c e
p a i n , t h e s u b j c c tc a n n o td i s t i n g u i s thh e n a t u r co f t h e i n c i t a t i o n .I n e f f e c t .

NorEs

553

when it is below the threshold,the incitation informs about the nature of

the stimulus;above the threshold,the individualis consciousof anothcr
fact: that the stimulusis of suchintensityas to endangerthe integrityof the
tissues.Pain thus appearsto be the sensorialsensationof a specificcharacter of 5timuli-sufficicnt intensity to representa danger for the tissues,
pain from the other sensorialsensations,
and it is this which differentiates
and puts it into a specialcategory.Pain is indepcndentof the natureof the
s t i m u l i .B y c o n s t i t u t i n a
g w a r n i n gt o t h e b o d y , t h a t i t s t i s s u c sa r e i n j e o p ardy, physiologicalpain inducesa generalrcsponseinvolvingbrisk, rapid
movements,a rise in pulserate, and a senseof invigoration.(10)
The fact that sensorialpain resultsfrom thc intensityof the external
incitationshas promptedinvestigators
to study this kind of pain largelyin
terms of the thresholdof incitation.It must be emphasizedthat for each
stimulusthere exist two thresholds,one for intensityvalues rcquired to
producespecificsensations,
and the secondfor intensityneededto produce
pain. There is a considerable
diflerence,for example,betweenthe heat intensitynecessary
to producea sensation
of warmthand the amountthat will
producea sensationof pain.
Parhological Pain
Pathologicalpain differs profoundly from physiologicalpain. It is a
psychicresponseto impulsesoriginatingin tissueswhich are abnormaleither becauseof damageproducedby externalstimulior becauseof inflamWhen pain is presentas
or other proccsses.
matory,circulatory,neoplastic,
it can no longerbe consideredas
of tissuedamageor disease,
a consequence
a signof injury,
a warningof dangerbut constitutes
responseto pathologicalpain is totaUydifferentfrom the
The genera.l
pain. Instcadof the organismbeing preparedfor
responseto physiological
fight or flight, its efforts are directedtoward placing the painful injured
a r e a ,o r t h ee n t i r eb o d y a t r e s t ;a n d ,t o p r o t c c t h e p a i n f u la r e af r o m f u r t h e r
injury, the pulse rate generallyslows,the blood pressurefalls and often
t h e r ei s s w e a t i n ga n d n a u s e a (. l 0 )
for pathologicalpain has
The local nature of the changesresponsible
possible
problem
of
the
several
mechanisms
of actionwhich may
raisedthe
intervenein inducingthis pain.
l. Locally originatedstimuli producedby damagedtissuesthemselves
may act directlyupon the pain end organsto inducepain impulses.Lewis
with tissuedamageis a resultof the
that the pain associated
has suggested
( l0) This posactionof locallyelaboratedabnormalchemicalsubstances.
sibilitywas first consideredby von Frey ( I I ) althoughactuallythe second
pain describedby him was due to differentratesof pain impulsetransmiss i o n .[ - e w i s( 1 0 ) a n d h i s a s s o c i a t chsa v es t u d i e dp a i n i n e r y t h r a l g i aw h i c h
r e p r e s e n tas t y p i c a lf o r m o f p a t h o l o g i c apla i n . ( 1 2 . 1 3 ) T h e y h a v e s h o w n
t h a t w h e n s k i n h a s b e e n i n j u r e d a n d t h u s r e n d e r e dh y p e r a l g e s ibcu t n o t
actuallypainful,sintplearrestof circuiationto this injured area may induce
pain. A similar phenomenonis evidentwhen a muscleis exercisedvigorouslywhile its circulationis arrested.If the constrictingblood pressurecuff

554

nESEARcH IN

pHysropATHoLocy

is released,the pain that is experiencedduring the period of ischemia disappears,but if the cuff is reinflated,pain may recur without further exercise.In both instances,
no new stimulusis requiredto arousepain. It has
been found that in erythralgia,ncither vasodilatationnor changein skin
temperatureis the factor responsiblefor lowering the pain threshold. According to Lewis, when the circulation to the affectedarea is severelyreduced,accumulatedstablechemicalsubstances
elaboratedby the damaged
tissuesmay act directlyas the pain stimulus.No definiteevidencehas been
offered by these researchers,however, as to the chemical nature of the
elaborated
substances
involved.
2. Local changesin damagedtissuesmay bring about a lowering of
the nerve threshold for pain. Lewis has demonstratedthe spread of the
lowered threshold to nerves far beyond the site of the lesion itself. He
studiedthe cutaneoushyperalgesia
followingtissue-damaglng
excitationof
a tiny area of skin by a tapcredforcepsor faradic current. By producing
damagein a previouslyanaesthetized
area,he found that the local changes
brought about by the damagedid not producehyperalgesiain the surrounding skin until the effectsof the local anaestheticwore off. The localized
nerve changesthen createda wide zone of hyperalgesiafor prolonged
periods.Tower (14) has presented
evidenceto show that the receptorend
structuresfor pain have an arborizingrather than a plexiform arrangement,
thus making unnecessary
the postulationof an autonomicallyunidentified
"nocifensor"nervesystem,as proposedby Lewis, ( l5 ) to accountfor the
type of spreadof the hyperalgesia.
The cxtent and especiallythe distribution of this areaof hyperalgesia
has clearlyindicatedthat it is the result of
a loweredthresholdin the arborizingbranchesof the cutaneousnerve, a
few branchesof which were originally intenselystimulated.When a few
fibers of a cutaneousnerve were directly stimulated,the same effect was
observed.The findings suggestedthat a local tissue change lowers the
thresholdfor pain for the nerveendingsof the damagedarea,and that this
effectmay spreadthrough other branchesof the cutaneousnerve involved
as well as through larger nerve trunks so that the resultantarea of hyperalgesiabecomesvery extensive.
3. Local changesmay alter end organsordinarilyconcernedwith other
forms of sensationin such a way that the impulsesoriginatedby them
evoke the sensationof pain. Certain areassuch as the appendixand the
mucosaof the stomachapparentlycannot,under normal circumstances,
bc
incitedto respondpainfullyto any form of stimulation.(16) However, in
the presenceof inflammation,the same stimuli may give rise to pain in
theseareas.The rclationshipbetweenthe end organsor nervesordinarily
concernedwith the receptionof other forms of sensationand those of pain
has been consideredby severalauthors.Weddell (17) has demonstrated
that the various complex end organ structuresare suppliedwith accessory
fibers,unmyelinatedand beaded.analogousto thoseconsideredto be pain
roceptors.Head ( I 8 ) has shown in experimentson the glans penis that
theremay be a fusionof varioussensations
into a singleconceptand that
one sensationmay inhibit another.Accordingto Feng, (19) the balancein

NOTES

555

excitabilitybetweentouch and pain receptorsmay be upet peripherallyby

Iiberationof a chemicalsubstance
as the resultof injury.
Considerable
evidenceexiststo indicatethat pain is the most primitive
form of sensation.It is possiblethat in the presenceof pathologicaldisturbances, through dedifferentiationcomplex systemsfor the reception and
transmission
of other modalitiesof sensation
come to act as pain receptors.
Whateverthe exact mechanismsmay be, the findingsof differentinvestigatorshave led them to the conclusionthat abnormalchemicalsubstancesare releasedfrom pathologicallyaffectedtissuesand that these
chemical substancesmay play an important role in the production of
pathological
pain.
Chaptcr 4, Notc 2. Blood Titrimetric Alkalinity and Urinary pH
The important role of the kidney in regulatingthe acid-basebalanceof
t h e b l o o d h a s b e e n d e s c r i b e d( 2 0 9 ) a n d a g e n e r a lr e l a t i o n s h i p
between
d a i l y a c i d e x c r e t i o na n d p l a s m ab i c a r b o n a t e
h a s b e e n r e c o g n i z e d(.2 1 0 )
However,a consistentrelationshipbetweenblood acid-basevariationsand
urine changeshas not beenclearlyestablished.
The determinationsof blood pH and COr combiningpower are the
most commonly employedmethodsfor following acid-basechanges.However, they indicateonly certaindistinctivefactorsinterveningin acid-base
balance.The pH is a measurementof the dissociatedelementsin the blood
and is maintainedwithin narrow limits by the buffer mechanisms,while
the CO: combining power is a measurementof only one of the multiple
factorsin the buffer system,thc bicarbonategroup. (2ll) The inconsistent
relationshipbetweenvariations in urinary pH and blood values indicate
that changesin urinary pH dcpcnd upon factors othcr than the blood's
dissociatedsubstancesand bicarbonate-carbonicacid buffer mechanism.
The phosphates,proteinsand hemoglobinare amongthe membersof other
important bufrer systemsthat have a role in the control of the acid-base
balance
o f t h e b l o o d .( F i g s . 2 0 3 , 2 0 4 )
The titrimetric alkalinity of the blood representsa measurementof the
totality (reserve supply) of the substances,both dissociatedand nondissociated,that are involved in the maintenanceof the acid-basebalance
of the blood. (212) We consideredit intercstingto examinethe relationship between blood titrimetric alkalinity and urinary pH, Concomitant
variationswere compared.
Human subjectsand dogs without apparent kidney dysfunction were
used.Blood was obtainedby venipuncturewith an accuratelycalibrateddry
syringe. After the needle was introduced into the vein and before withdrawing blood, the tourniquet was releasedfor severalminutes to avoid
changesdue to stasis.
Exactly 5 cc. of blood was introduceddirectly into a flask containing
3 0 c c . o f a 0 . 0 0 1 N s o d i u mh y d r o x i d es o l u t i o nT. h e f l a s kw a s i r n m e d i a t e l y
closedwith a rubber stopperand the mixture agitatedsufficientlyto assure
homogeneity.If determinationswere not carried out at once, flasks were

556

n E S E A R c Hr N p H Y s l o p A T H o L o c y

storedat 5oC and then brought to room temperature(20"C) before analysis.Fifty cc. of distilledwater was used as a wash when ffask contentswere
transferredto a beaker for titration. The total alkalinity was determined
b y e l e c t r o m e t r itci t r a t i o nt o p H 7 . 0 a g a i n sat . 0 l N h y d r o c h l o r i ca c i d s o l u ,s6
75r
e

75o
?t.
,t6

z' rr.
,t2
,90
/o

c -

cc. KI nzlC0

6t
56
6tt

3:

!3
.-';

6L
60
5l'
_56
Stoo
f,ors

@
::

I fr soto
1 'Y. so2r
60 00
7tl

JrS

too

tf

us

a -

V)o

ner 7
\

67t

2!o

zts 6?5
-----lll-After
6r
50
I

6q

: 6 0
qa
1

s6
5r
O

JQ

t{Irul ts

&

tra

.l!o

Ja l

F t c . 2 0 1 . T h e c o m p a r i s o n b e t w e e nt h e c o n c o m i t a n tc h a n g e ss e c n i n v a r i o u s b l o o d
and urine analysesconccrning lhe ocid-hasebalance of the body. It shows that rhe
variations of the titrimetric alkalinity of the total bloo'd are thc only values which
c o n s t a n t l yp a r a l l e l t h o s e o f t h e u r i n a r y p H . ( P a t i e n t w i t h b r e a s t c a n c e r )

tion usinga BeckmanpH meter,Model G, and a mechanicalstirrer.Blanks

of 30 cc. of the sodiumhydroxidesolutionwere storedand treatedin the
samemanner.
U r i n e s p e c i m e n sf r o n r t h e h u m a n s u b j e c t sw e r e o b t a i n e d t h r o u g h
completeemptyingof bladder contentsby voluntary micturition. An indwellingcatheterwas used for dogs and some humans.Specimenswere
placedin containenclosedwith rubber stoppersand storedat 5"C. They

NOTES

557

were brought to room temperaturebeforetests.The pH valueswere determined electrometrically.

Blood specimenswere collectedeach hour for at least five consecutive
hours. Urine sampleswere obtainedevery thirty minutes,as specimensac,JO

715

9
o
o
@

?3s
,o cc. fl n/ltlO
78
!

7l

t s
! o <

22
7o
68
66
6e

E .

t9.
(8

< G

'/O

JQr
@

-----'---'------

r l
-t?-

6\'

tJo

60,)

/cp

510

K . L . z

7tl

e
L
a
c

7o
64

rAf{r

6 ^

st
Sa
60'o

- J.

Frc. 204. In some casesthe changes in several blood analysesparallel the cbanges
of urinary pH. (Subject with metastaticmelanoma)

cumulated in the bladder during the half hour precedingbleeding and again
during the half hour following bleeding.The pH values of the urine specimens were plotted separatelyon graphs as A and B curves. Comparison
was then made betweenthe two urine curves and the curve representingthe
valuesof the blood titrimetric alkalinity, using time as abscissa.In another
group of experimentsusing dogs, the bladdercontentswere drained at intervals of from five to ten minutes and blood specimenswere obtained
every thirty minutes.

558

pHysropATHoLocy

. , ' R E , s E A R c Ht N

Severalpreliminary tests were carried out to determine the degreeof

a c c u r a c yo f t h e m e t h o d su s e d .B y e m p l o y i n ga 0 . 0 0 1 N N a O H s o l u t i o n ,l
probableerror of no more than 0.1 cc. was found.
Comparisonsbetweentitrimetricalkalinityof hourly blood specimens
and half-hourly urine specimenswere made in thirty human and seven

cc. HCIn/100

oa

3 . .
a
a

I
O

a a

>

'erc

a l
O

(U
_)a

'io

ltl

(u

t
a

(F r n
l n
E
o
o
@

. ?.

a
a
i

a
a

o
a
oa

a
a

ara
o

a l '

a
a

a
aa

U r r n a r yp H
F r c . 2 0 5 . ' I h e c o m p a r i s o nb e t w e e nt h e c o n c o m r t a n tv a l u e s o f u r i n a r y p H a n d o f
t h e t i t r i m e t r i ca l k a l i n i t yo f t h e b l o o d s h o w st h a t t h i s r e l a t i o n s h i pc o n c e r n sm o r e t h e
o c c u r r i n g c h a n g e sa n d l e s s t h e a b s o l u t ev a l u e s o f t h e f i n d i n g s .U r i n e s a m p l e s ' * , i t h
, r e s e e nt o c o r r e s p o n dt o b l o o d s a m p l e sw i t h d i f t h e s a m e p H . i n d i f f e r e n ts u b j e c t s a
ferenttitrimetricalkalinitY.

NOTES

,i

559

caninesubjects.In all cases,the curvcsof blood titrimetricalkalinityvalues

showeda consistentparallelismto the pH curvesof urinc specimensthat
accumulatedin the bladderduring the thirty minutesprecedingthe bleeding
a n d w e r ec o l l e c t e a
d t t h c t i n r eo f v e n i p u n c t u r (eF. i g . 2 0 6 ) T h e c u r v e sr e p r e sentingthe pH valuesof urine specimens
accumulated
in the bladderdurItg
I

cc. C\

4e
947

c,
d

3+a
(lJ

c . -4t

?
J

++
4!

HCIn/100
;c{

3ee
:60
L

?,0
L

? l o

7tl

7,1

7.t
7.2

e
>L

7,1
7.<

D.

Minuteso

f 6rr !'lo

24o

F r c . 2 0 6 . C o m p a r i s o no f t h e h o u r l y b l o o d t i t r a t a b l ea l k a l i n i t y ( i n t e r m s o f c c . o f
. O I N H C I ) a n d h a l f h o u r l y p H c u r v e s i n a h u m a n s u b j e c t .U r i n e c u r v e s A a n d B
r e p r e s e n tt h e b e f o r e a n d a f t e r b l e e d i n g s p e c i m e n s . ' I h e p a r a l l e l i s m b e t w e e n t h e
t i t r a t a b l ea l k a l i n i t y o f b l o o d a n d t h e u r i n e c u r v e A i s c l e a r l y s h o w n , C u r v e B s h o w s
no correlation.

560

nESEARcHrN pHysropArHoLocy

ing the thirty minutes after each bleeding (B curves) did not show the
s a m ec o n s i s t e nct o r r e l a t i o nt o t h c b l o o d t i t r i m e t r i ca l k a l i n i t y .( F i e . 2 0 7 )
In severaltests,collectionsof half-hourly urine specimenswere made
f,fteen minutes before and flfteen minutes after each bleeding.The same
parallelismwas found betweenthc pH of urine accumulatedin the bladder
during the period from fifteenminutesbeforeto fifteenminutesafter bleeding, and the titrimetric alkalinity of blood specimensdrawn in the middle

Elood

cc.

(o, N HCI) u--

to
9
I
7
6.O
5.8
56
5.4

\---

Hour

-'

F r c . 2 0 7 . C o m p a r i s o n o f t h e c u r v e s r e p r e s e n t i n gh o u r l y b l o o d t i t r a t a b l e a l k a l i n i r l ,
( i n t c r m s o f c c . o f . O l N H C I ) a n d u r i n e p H i n a h u m a n s u b j e c t .U r i n e p H c u r v e
" B e f o r e " s h o w s v a l u e s o f s p e c i m e n sa c c u m u l a t e di n b l a d d e r d u r i n g t h e h a l f h o u r
i m m e d i a t e l yp r e c e d i n ga n d c o l l e c t e da t t h e t i m e o f b l e e d i n g ,w h i l e u r i n e p H c u n ' e
" A f t e r " i s f o r s p e c i m e n sc o l l e c t e do n e h a l f h o u r a f t e r b l e e d i n g .T h e r e i s a definite
p a r a l l e l i s mo n l y b e t w e e nt h e c u r v e so f t h e t i t r a t a b l ea l k a l i n i t y o f b l o o d a n d the urine
pH curve A.

of the urine collectionperiod.The pH curvesof urine specimensaccumulated during the period from fifteen to forty-five minutes after each bleeding showedno consistentcorrelation.
When urine and blood specimenswere obtainedat shorter intervals.
the sametendencyof urine pH changesto precedethe changesin blood
t i t r i m e t r i ca l k a l i n i t yw a s o b s e r v e dI.n F i g u r e2 0 8 a r a p i d r i s e i n u r i n e p H
is seen to begin within twenty minutesof the time of administrationof
sodium bicarbonate.The blood titrimetric alkalinity does not sho,*,an
elevationfor at least forty-five minutes,

NorEs

561

Thesestudieshave shownthat urinary pH variationscorrespondclosely

to changesin the valuesof an important factor reflectingacid-basebalance
changesof the blood, the titrimetric alkalinity.As a result, it has been
possibleto employ variations in urinary pH as indicationsof qualitative
changesin acid-basebalanceof the blood for other studies.

cc
Elood
(.ot N Hct)t7 O

t6.o
r{n

59n..NoHCO,

t,

t40
130

t2.o
9H
Urinc

8.5
8.O
7.5
7.O
6.5
6.O
40

70rc^.90
Minules

ilo

t9

t@

F r c . 2 0 8 . T h e e f f e c to f t h e a d n r i n i s t r a t i oonf 5 g r a m s o f s o d i u n rb i c a r b o n a t eu p o n
the blood titratahle ulkalinitv and urine pH curt'es in dog. The elevation in urine
p H i s s e e n t o p r e c e d eb y t w e n t y - f i v em i n u t e s t h a t o f t h e t i t r a t a b l c a l k a l i n i t y o f t h e
blood.

Chapter4, Note 3. Acid Pattenr of Pain and Lactic Acid

We have investigatedthc relationshipbetweenpain of acid pattern
and the appearanceof lactic acid resultingfrom abnormal metabolismof
For this purpose we used the techniqueof Friedemann,
carbohydrates.
l a t i e n t si t a p p e a r e dp o s s i b l et o e s t a b l i s h
C o t o n i oa n d S h a f f e r .I n s e v e r a p
b y m e a s u r i n gt h c l a c t i c a c i d c o n t e n to f e f l c r c n tb l o o d
this relationship
f r o m t u m o r sd u r i n g i n t e n s i v ep a i n o f a c i d p a t t e r n .I n a y o u n g m a n w i t h
a h u g e s a r c o m ao f t h e k n c e ' ,f o r w h o n t t h c a c i d p a t t c r n o f t h c p a i n
throughits relationshipto the changesin thc urinary
had beenestablished
pH, suchanalysescould be carriedout in the blood obtainedfrom the big
efferentveins,At the momentsof very severepain, the
easilyaccessible
amount of lactic acid had markedly increased.Values as high as 128
mg./100 cc. blood werc found during thesc painful periods,contrasting
with valuesaround30 mg,/100 cc. blood in the period of calm.

562

RESEARCH IN

PHYSIOPATHOLOGY

Chapter4, Note 4. Itching

Although we were especiallyconccrnedwith pathologicalitching, *'e
a l s ob e c a m ei n t c r e s t c idn p h y s i o l o g i c a
i tlc h i n g e
, s p e c i a l liyn i t s r e l a t i o n s h i p
to sensorialsensations
in general.Physiological
itching can be regardedas
a d i s t i n c ts e n s a t i o nn, o t j u s t a g r a d a t i o no f a n o t h e r s e n s a t i o nI.t c h i n g
seemsto have its own end organs.The propensityof ccrtain regions of
the body-such as the nasalmucousmembranes,
the skin near the nostrils.
and the perioraland perianalskin-to itch in responseto externalstimuli
can be correlatedwith the presenccof suchend organs.The proteopathic
c h a r a c t eor f i t c h i n gw o u l d a l s o p r o n r p tu s t o c o n s i d e rl e s sm y e l i n i z e do r
e v e n n o n m y e l i n i z endc r v e sa s i t s c o n d u c t o r sB. y a n a l o g yw i t h p a i n . t h e
e x i s t e n co
e f p r o p e rc c n t r a lc e n t e r sc o u l d b e c o n c c i v e d .
The most importantcharacteristic
of itching is that it can be induced
b y s t i m u l iw h i c h ,a t o t h e ri n t c n s i t i e sr e, s u l ti n a d i f f e r e n st e n s a t i o sn u c h a s
t o u c h ,f o r i n s t a n c cA. l t h o u g hl e s sm a n i f c s to, t h e r s t i m u l i ,s u c ha s h e a t a n d
c o l d , a l s oc a n i n d u c ci t c h i n g .w c h a v es c e nt h a t s t i m u l iw h i c h u s u a l l yi n d u c e o t h e r s e n s a t i o ncsa n p r o d u c ep h y s i o l o g i c apla i n i f t h e y h a v e a n i n t e n s i t ya b o v ea t h r e s h o l dl e v c l . I t i s t h e i n t e n s i t yo f t h e s t i n t u l u sw h i c h
determincw
s h e t h e ri t c a u s c sp a i n o r a s e n s a t i o n
o f t o u c h ,h e a t o r c o l d .
Sincepain appcarsif the stimulusis abovethc thresholdlevel,it servesas
a w a r n i n go f a d a m a g i n gi n c i t a t i o n .
I n s t u d y i n gi t c h i n gu n d e r a s i m i l a ra s p e c ti,t c a n b e s e e nt h a t i t , t o o .
i s i n d u c c d b y n o n s p c c i f i cs t i m u l i .B u t , f o r i t c h i n g ,t h c i n t e n s i t yo f t h e
s t i m u l ii s l o w . E v e r y b o d yk n o w st h a t a n e s s e n t i aclo n d i t i o nf o r t h e i n d u c t i o n o f i t c h i n gi s t h a t t h c i n c i t a t i o nb c s l i g h t T
. h i s i s c a s i l ys c c nf o r t h e s k i n .
a n d e s p e c i a l l tyh e n a s a lm u c o u sm e m b r a n ew
, h e r ea s t r o n g e rs t i m u l a t i o n
w i l l n o t i n d u c ei t c h i n gb u t a t o u c h s e n s a t i o nJ.u s t a s t h e i n t e n s i t yo f a
s t i m u l u sd c t e r m i n e sr v h e t h c rp a i n o r t o u c h i s p r o d u c e d ,s o t h e i n t e n s i t y
a l s od c t c r m i n c sw h e t h c ri t c h i n go r t o u c hi s f c l t . W h i l e t h e s e n s o r i asl c n s a t i o n o f t o u c hi s i n d u c e db y s t i m u l iw i t h i n t e n s i t i cbse l o wt h o s er e q u i r e df o r
p a i n , i t c h i n gi s i n d u c c di f i n t e n s i t i cas r e b c l o w t h o s er e q u i r e df o r t o u c h .
T h e r e l a t i o n s h iopf i n t e n s i t yo f s t i n r u l u st o i t c h i n g ,s e n s o r i asl e n s a t i o n
o f t o u c h .a n d p a i n i s s h o w ni n F i g u r e2 0 8 b i s . T h i s c o r r e l a t i o ne x p l a i n s
w h y i t c h i n gi s p r c s c n ts o m e t i m cfso r a b r i e f p c r i o d w h c n s k i n o r m u c o u s
n t e m b r a n es e n s o r i asl e n s a t i o o
n r c v e n p a i n i s i n d u c e d .l m m e d i a t e l ya f t e r
a n i n j u r y , f o r c x a m p l c ,i t c h i n gm a y b c f e l t f o r a s h o r t t i m e o n l y t o d i s a p p e a rj u s t p r i o r t o t h c d e v e l o p n r e notf p a i n . - l - h c l o w i n t e n s i t yo f t h e
s t i m u l u sr e q u i r c de x p l a i n sa s t r i k i n gc h a r a c t c r i s toi cf i t c h i n g :i t s d i s a p p c a r a n c c w h e n a s t r o n g e rs t i n t u l u si s a p p l i e d .T h u s s c r a t c h i n gw
, hich adds
m o r e i n t c n s i v cs t i m u l a t i o nm
, a k e si t c h i n gd i s a p p c a rT. h e n r o r ev i o l e n tt h c
s c r a t c h i n ge,v e nt o t h e p o i n t o f i n d u c i n gp a i n . t h e m o r e c f f e c t i v ei t c a n b c
i n e l i m i n a t i ni gt c h i n g .
T h e g e n e r a lr e a c t i o nt o w a r d i t c h i n ga l s o a p p e a r sr e l a t e dt o t h e c h a r a c t e ro f i t s i n d u c t i o nT. h c i n d i v i d u arl e s p o n dtso p a i n b y f l e e i n go r f i g h t i n g
i n o r d c r t o e s c a p ct h e i n t e n s i v en o x i o u si n c i t a t i o nA
. s t h c i n c i t a t i o nt h a t
p r o d u c c si t c h i n gi s m i n i n r a lb y d e f i n i t i o n - t h c p r c s c n c eo n t h e s k i n o f a

NorEs

563

minor irritant such as a fly or mosquito,for instance-scratching is sufficient to eliminateit. with a fly on his skin, the individualneed not flee or
fight, but only scratch.With the conceptthat itching can result from exactly
the sametype of stimuli as pain and touch,we integrateit in the group of
sensorialsensations.
We can then establisha separatesensorialsubgroup
for itching and pain. whi.le other sensorialsensationsinform us of the
nature of the gxsilsli6n-heat, cold, sound,taste,etg.-itching and pain
inform us only aboutthe intensityof the stimulus,not its nature.
Pathological
itching,like pathological
pain, is relatedto the existenceof
abnormalities,
In addition to the differences
in stimulusintensityrequired
to induceitching and pain, their differentnervousformationshelp explain
their clinical separation.No patient we have studied has ever indicated
any confusionas to whetherhis discomfortwas due to severeitching or
pain. The two sensationsare seldomconcomitant;usually they sucieed
one another.The fact that proteopathicpain and itching both seemto be
conductedthrough unmyelinizednervesindicateswhy they can appear
under similarconditions,as in nerveregcneration.
This semsto have led
to confusionbetweenitching and pain. However, itching and pain observedduring nerve regeneration
can be clearly differentiated
by the patient. The fact that the itching sensationis producedby stimuli of low
intensityalso explainswhy itching is so oftcn presenton skin or mucous
membraneswithout appreciablepathology.Minimal changesappearsufficientto inducethe sensation.

Pain

Touch
It c h i n g
I n t e n s i t yo f e x c i t aito n
Ftc. 208 bis. Similar to pain, itching representsa special kind of sensation,with the
a i m t o i n f o r m a b o u t t h e i n t e n s i t yo f t h e e x c i t a t i o n ,I f t h i s i s v e r y s l i g h t , i t i n d u c e s
i t c h i n g .I f a b o v e t h e t h r e s h o l d ,a t w h i c h t h e i n c i t a t i o na c q u i r e sa n o x i o u s c h a r a c t e r ,
it inducespain.

564

RESEARCH IN

PHYSIOPATHOLOGY

CL

v)
(It

z.

uays
F t c . 2 0 9 . T h e n a s a l p l l m e a s u r e dw i t h a g l a s s e l e c t r o d e i n t r o d u c e d d e e p i n t h e
n o s e s h o w s t h e s a m e d u a l i s m a s t h e o t h e r a n a l y s e sW
. i t h 6 . 5 a s t h e a v e r a g ev a l u e ,
t h c c u r v e so f t h e n a s a lp H h a s m o r e r a p i d a n d b r o a d e r v a r i a t i o n st h a n o t h e r a n a l y s c s .
C u r v e o f d a i l y a n a l y s e ss h o w s v a l u e sa b o v e t h e a v e r a g el i n e i n a c a s e o f g e n e r a l i z e d
melanoma.

7tr
70
o(tt

u,
(u
z.

f 6 7 t

? ro tr (2 rt tr lu r fr /C /9

Days

F r c . 2 1 0 . T h e n a s a lp H s h o w sp e r s i s t e nl to w v a l u e si n a c a s eo f c a n c e ro f t h e l i v e r .

NOTES

565

C h a p t e r4 , N o t e 5 , N a s a l p H
Nasal pH was measuredusing a portableBeckman pH meter and a
glasselectrodesmall enoughto penetratedeepinto the nose.In a research
m a d e w i t h N . B u c h a n a ni t w a s f o u n d t h a t v a l i d d a t a c o u l d b e o b t a i n e d
only if the electrodetouchedthe turbinate,otherwisemarked differencesin
valueswere noted.

7
7
o
ct
(n

(!

z, 6
q

t
Zro
T

(t
lL

6
f

5to

,s

IQ

lt

/3

/1

/6

Uays
b e t w e e nd a i l y c h a n g e so f t h e n a s a la n d u r i n a r y p H s h o r v s
Frc. 2l l. The relationship
oppositevariations.

566

../ RESEARCH tN

pHyslopATHoLocy

In a simplified method, cotton applicatorswere soaked in Guillaumin

indicator solution with methyl red and bromothymol blue and left to dry.
They were easilyintroducedto sufficientdepth in the nose and left in place
for at leasttwo minutes.Color of the wct spotswas checkedwith a colorimetric scale.Data obtainedwith glasselectrodeand colormetric applicator
were found to coincide closely.
Two offbalancescould be seen,one with the pH elevatedsometimes
even above8, Figures209 and 210 show curvesof the offba.lance
in two
patients.It is interestingto note that the changesin the nasal pH values
parallel these seen at the level of lesionsand are opposite to those concomitantlyoccurringin the urinary pH (Fis.2ll) which parallelthose of
the titrimetricalkalinitvof the blood.
Chapter 4, Notc 6. Wheal Resorption
Interestinginformation could be obtainedby analyzingthe absorption
of fluid injected intradermicallyin various subjects,and correlating the
resultswith the existenceof metabolicoffbalances.we used the technique proposedby Mcclure and Aldrich, in which they measuredthe
time needed for the disappearanceof a wheal, resulting from an intracutaneousinjection of a saline solution. A relatively extensivestudy of
absorptionwas made in more than 500 subjects-both normal and abnormal, We presenthere a few of the conclusionsfrom this study.
The averagetime necessaryfor resolutionof the wheal obtainedby the
injectionof .2 cc. of lVo NaCl solution,in normal subjects,was 23 minutes;the range was from 15 to 30 minutes.when deviationsfrom these
valueswere observed,they were consistent,in the sensethat testsrepeated
at short intervalsin the samegeneralarea in the same subjectgave values
in the same abnormal range.Abnormal valuesoccurred in two directions.
Resorptiontime was shortenedin somecasesand valuesas low as I to 2
minuteswere noted. In the opppsitedirection,valuesas high as 90 minutes were observed.Thesedeviationsfrom normal time could be related to
local and general conditions. The presenceof local or regional edema
shortensthe resorption time so much, that in some caseswith massive
edema,no wheal could even be realized.Shorteningof time was found to
be true for an edema,regardless
6f s3u5s-inflammation,impaired local
circulationas in phlcbitis,impairedgcncralcirculationas in cardiacsor in
renal failure. Lengtheningof resorptiontime in casesof phlebitis provides
valuable information on the evolution of the condition. The return of
resorptiontime to normal valuesseemsto indicate sufficientimprovement
to permit mobilizationof the patient.
In subjectsin whom no local factor could be consideredto be responsible for changesin the resorptiontime of the wheal, we could see that
abnormal variations had a direct relationshipwith the general offbalance
present.In some subjectswith a manifcstoffbalanceof type D, wheal
resorptiontime was shortened.Values as low as 4 to 5 minutes were obtained. Analysis of a number of casesindicated that this shortening of

NorEs

567

resorption time meant a bad prognosis.A few patients with only values of
2 to 4 minutes died within a few days although other symptomsgave no
indicationof a fatal outcomewithin a short tlmc.
Extendedresorptiontime hasbcenfound in subjcctswith an offbalance
of type A. Values as high as 60 to 90 minutes were found in subjects in whom all other analysesindicated this oftbalance. It is also
interestingto note the existenceof slow resorptiontime for aged subjects.
In a group of 80 patientsrangingfrom 70 to 90 years of age,^n average
resorptiontime of 90 minuteswas found. (FiS. 68)
Chapter4, Note 7. Eosinophiles
The role of the blood as the secondarypart of the entity organismhas
explainedmany of the peculiaritiesof its cells.Aside frorn the phagocytary
functions that can be consideredas a particular form of capturation,the
leucocyteshave to be recognizedas acting as holocrine monocellularformations whose specificconstituentsare liberated by cellular lysis. We have
seen that in the case of the neutrophilicgranulocytes,the hydrolytic enrymes so liberated,stronglyresemblethe externalsecretionof the Dancreas.
Under this aspect,we have investigated
the blood eosinophiles
with a role
similarto the Panethcellsof thc duodenum.
The physiologyof theseleucocytes
has to be soughtin the acidophilic
character of their granules. Morphological analysis of the eosinophile
granule shows that it is formed by a content and a membrane,the last
clearly seenin preparationsin which the granuleshave lost their content.
Like many other membranes,that of the eosinophilegranulecan be easily
identifiedas being made partially at leastby lipids being stainedwith dyes
dissolvingin lipids, such as black Sudan or Scharlach.However, it is the
content of the granule with its ability to combinewith acid dyes that indicatesits specificcharacters.Under certain circumstances,
when the blood
is maintainedin vitro betweenthe slide and cover object for a certain time,
the membraneand granulecontentare seento separate.Before this occurrence,a lysis of the eosinophileleucocyteitself takes place. This is manifested through the breakdown of the cellular membranewith the lysis of
the nucleus.It is in a secondstepthat the eosinophilegranulelosesits content. Following it, besidesthe empty glanulesand lysed eosinophiles,characteristicCharcot-Leydencrystals appear. The correlation betweenthese
crystalsand eosinophileshas been recognizedand is generallyacceptedas
occurringin vivo and in vitro.
A y e r ( 2 1 5 ) h a s s h o r t e n c dt h e p r o c e s so f l y s i s o f t h e e o s i n o p h i l eisn
vitro by treatingthe blood preparationswith a detergent,aerosol.By repeating Ayer's experiments,tle relationshipbetween the appearanceof
the Charcot-Leydencrystalsand the more complex processof lysis of the
eosinophileshas becomeapparent.It could be seenthat the crystalswould
appearat the site where the nuclei of the eosinophilesdisappearedthrough
lysis, and where careful observationof the granulesreveals the loss of
their eosinophilic content. The presenceof empty granule membranes

568

R E S E ^ R c Hr N P H Y S T o P ^ T H o L o c y

stainedby the fatty dyes,in additionto the lysednuclei,would indicatethe

conditionsunder which Charcot-Leydencrystalsappear.The eosinophilic
contentof the granuleand productsinducedby the nuclearlysis represent
the two factorsthat togetherresultin thesecrystals.
Concerningthe relationshipbetweenCharcot-Leydencrystalsand eosinophiles,
it is interestingto note the diffcrencethat existsbetwecneosinophilic granulesin variousanimals.Besidesthe morphologicalaspectwhich
can be very different,apparentlyno Charcot-Leydencrystalsare obtained
from speciesother than humansand certain simians.This indicatesthat
when the biologicalrole of the eosinophiles
is considered,
we have to seek
anothercommonfactor in additionto the morphological
and chemicalones.
It would seemthat it is in their basicreactivity,i.e. in their capacityto bind
substances
of acid character,that the commoncharacterof all eosinophile
't'his
g r a n u l e sh a st o b e s o u g h t .
i s a l s ot r u c f o r t h e d u o d e n acl e l l s .
Followingthis vicw, we initially tendedto accordmore importanceto
an antacidpropertythan to any other, seeminglyagrecingwith other data
obtainedfrom this study.Among the substanccs
found to be the principal
constituentsof these eosinophilegranules,the alkaline amino acids, of
which arginineis the the principalone, assumea very importantrole. According to the hypothesiswe advance,thcse alkaline amino acids would
representthe activefactor of thcsegranulesand would be liberatedby the
when they disintegratc.
eosinophiles
The eosinophileswould intervenein
physiologyfor the specificpurpose of furnishingcertain alkaline compoundsin whoscconstitutionthe alkalineamino acidsenter.The solubility
of the granulecontent,when liberated,and the Charcot-Leydencrystals
indicate,accordingto this view, that the main characterof the eosinophile
granuleis its capacityto furnish alkalinecompounds.Under specialcirthey are able to act againstsubstances
cumstances,
with acid properties
that resultfrom the lysisof the nuclei,and togetherto form the CharcotLeydencrystals.
The relationshipof the disintegration
of the eosinophileto the surface
t e n s i o nl o w c r i n gi l g c n t si s a l s o i n t c r c s t i n g
f o r t h c f u r t h c r l i b e r a t i o no f t h e
granules.
contentof these
Just as for other granulocytes
and lymphocytes,
fate of thesecellsand would constitutetheir most
lysisis the characteristic
important character.As seenabove,it can be related to the role of blood
in the organization,i,e. as the secondarypart of the organismlevel.
As for the other lcucocytcs,an importantfactor in the holocrinicrole
of the eosinophileshas bcen seenin the necessityof a maturation of these
granulesfor their activeintcrvention.Whcn lysiswas induced,it was seen
to affect only the cells that had reacheda certain degreeof maturity, not
only for the cellsthemselves
but also for the granules.Young cells,recogn i z e db y m o r e i n t e n s cb a s o p h i l yo f t h e c y t o p l a s mb, y l a c k o f . o r r e d u c e d
lobulationof the nucleus,and especiallyby a neutrophilicor even basophilic characterof the granules,do not break down. As in the circulating
blood, immature elementsare seen,the delay observedin inducing eosinopeniaby various agcntscan be intcrpretedas correspondingto the
time neededfor the circulatingeosinophiles
to reachmaturity,as an essen-

NorEs

559

tial condition for their lysis. This situation was apparcnt in a particular
case, that of eosinopeniainduced through administrationof the adrenal
corticoids.Although this appearsto have a direct effect upon the eosinophiles,a definitetime, often even24 hours, is seenessentialin order to
achievethe disappearanceof the eosinophilesfrom the circulating blood.
This delay has been related to the presenceof eosinophiles,allegedly"resistant"to the corticoids.In fact, in studyingthe eosinophileswhich persist
after the administrationof thesehormones,we could see that they represent only immature elements,probably prematurely liberated in larger
amountsfrom the bone marrow. The cells from which some will persist
even for 24 hours after administrationof corticoneoglucogenic
hormones,
do not show lysis in vitro nor the appearanceof Charcot-Leydencrystals,
and they presentthe tinctorial charactersof immaturity for the cytoplasm
and especiallyfor the granules.
Granule maturation,which correspondsto the acidophiliccharacter,
seemsto be the essentialconditionfor the lytic interventionof thesecells.
In the physiologicalrole of eosinophiles,
an importantaspectof maturation was seenin the relationshipbetweenthe richnessof theseelementsin
the circulatingblood and the processesin which a manifest local eosinophilia is induced, as through injection of parasitelarvae or vegetal oils.
A direct relationshipbetweenlocal and sanguineeosinophiliawas apparent, the value of the former being the function of the latter. The ability of
bone marrow to rapidly compensatethe transitory eosinopeniafollowing
the passageof these cells into the tissues,has further directly connected
the local richnessin eosinophilesto the bone marrow's capacity to send
new cells into the circulation.In all thesechanges,the prevailingfactor
has apparentlybeen the degreeof maturity of the eosinophilegranulae,
which seemsto require a certain time to reach thc desired degreewhich
is the principalconditionalso for their physiological
intervention.
Correlationbetweenthe biologicalinterventionof the eosinophiles
and
the acidophiliccharacterof the granulesand their richnessin alkaline
a m i n oa c i d sh a s b e e nc o n f i r m e di n a s t u d yo f t h c b a s o p h i l eosf t h e b l o o d
cells with granuleshaving an oppositecharacter.These granuleshave an
acid content, as seen by their tinctorial affinity for alkaline dyes. They
were also observedto contain heparine.a polysulfonatedmucoid of frank
acid character.The biologicalantagonismbetweenheparineand alkaline
proteinsis well known. Protamines,the correspondentof histonesfor fish,
are usedto correctthe exccsses
of hcparinin thc body. cspeciallyas therapeutic measures.
Therefore,the antagonismbetweeneosinophilicand basophilic granulocytesgoesbeyond their tinctorial characters.
Through the alkaline reactivityof eosinophilesas related to the fundamental separationof interveningconstituentsaccording to their positive
or negativecharacter,the antacid eosinophilescould be consideredto be
in the former group, while the basophiles,
rich in lytic heparin,are in the
latter.
We shall more fully discussbelow the nature of the interventionof the
eosinophilesafter studyingthe role of a specialgroup of constituents.For

570

nESEARcH rN pHysropATHoLocy

the present,it seemsthat under abnormal circumstances,exaggerationin

the amount of eosinophileswould indicate an existing predominanceof
conditionsthat correspondto agentsof positivecharacter,i.e., with heterotropic tendency.The more preciseantacidcharacterof these cells further
indicatesthe place that has to be reservedfor the eosinophilesin the group
of heterotropicagents.Under this aspect,the eosinophilewould be seen as
an agent of anti-acid characterin the blood and tissues,conceivedto act
as a holocrinic cellular gland, i.e., through the lysis of the corresponding
cell. Therefore, the richnessof the blood and tissuesin eosinophileswould
indicatea predominance
of heterotropictendency,while paucityin eosinophilesor their absencewould indicatea homotropictrend.

E
E

C,

o
C'
u,J

0ays
F t o . 2 1 2 . C u r v e o f b l o o d e o s i n o p h i l e si n a c a s e o f b r e a s t a d e n o c a r c i n o m aw i t h
m u l t i p l e m e t a s t a s e ss,h o w i n g v a l u e sp e r s i s t e n t l yb e l o w t h e a v e r a g el i n e o f 1 0 0 .

In order to understandthis aspectof the eoshophiles,we tried to follow the changesin their amount in the blood in relation to normal and
abnormal physiology.Study of the changesin the number of eosinophiles
in the circulating blood under physiologicalconditions has indicated the
existenceof the same 24-hour oscillationsas seen for many other constituentsof the blood. A relationshipis apparentbetweenthe periodscorrespondingto higher or lower quantiticsof circulating eosinophilesand the
degreeof activity of the individual.This appearsto be oppositein humans
who show diurnal activity, and mice and rats that show nocturnal activity.
By experimentallychangingthe hours of light and dark for mice and rats,
and through it the time of rest and of activity, the rhythm of change was
reversed,
Following the concept of the interventionof eosinophilesin biological
balance,we further investigatedthis aspectof the problem in relation to
the dualism in abnormalconditions.Just as for other tests,we obtained an

NorEs

571

averagevalue in a large seriesof normal human subjects.Utilizing the

Dungar technique for a direct count of eosinophiles,the value of 100
cells/l cmm. was found to be the averagevalue. An impressivedirect
correlationcould be found betweenthe amount of circulatingeosinophiles
and the two patternsof abnormaiity.In one group, that correspondingto
type A, the number of eosinophiles
appearednot only high but with their
values fixed above the averagevalue. High values were observed to
persistfor long periodsof time. Figure 213 shows such a case.For the
oppositepattern, correspondingto the fundamentaltype D, these values
appearedto be below 100 and very often 0, persistingfor a long period of
time. (Fr.g.212)ln thesedual patterns,the degreeof abnormalitycould be
relatedto the deviationin the number o[ theseelementsfrom the averaqe
valueof 100 elements/l cmm.

E
(.)

an
o
UJ

Days
F t c . 2 1 3 . C u r v e o f t h e b l o o d e o s i n o p h i l e si n a c a s eo f g e n e r a l i z e dm e l a n o m a s h o w ing values persistentlyabove the averageline of 100.

The relativefacility with which the number of eosinophilesin the blood

of an individual can be determined,has made it an important researchtool
for information regardingthe balancebetweenthe two fundamentalbiological tendencies.
Chapter 4, Note 8. Total Blood Potassium
For a large-scaleinvestigation-requiring as many as one hundred determinationsa day-the technique of separatingred cells from plasma
appearedto be impractical.In view of the relativelyminute amountsof
potassiumin plasma as compared to cells, we could utilize total blood
insteadof the cells.It was also found that by diluting the blood 1/10, the
values obtained were in the same range as for serum potassium,a fact
which permittedthe use of the flame photometerwithout any changein the

572

R E S E A R C H

I N

P H Y S T O P A T H O L O G Y

E.

a"!"(:i{llii:','l:":':'."tt'"r'-'r'2trr',",'-i'i'/i','i'i't'i.J.;l'l!l
, . t n { t

,.5
,.4
at

5r,

Ett
2tr

a ,

- . , .

t t 4 , ' 4 ' t t . . t . r

? .

t a n t . ' , 2 , ,

N^
A
.\ rA' \ nA
'r \ VA\ ^ n
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\
/
\
V
\
^rA
r
V V V ^-J \ i V
\r /
VV

< tt
t

.;
?.'

""'^.-'
)r
C r

6 t
L A

- ' \a A r

lo
J '
t o
I t

tr

jl!

F I c . 2 1 4 . R e l a t i o n s h i pb e t w e e n s e r u m K + a n d t o t a l b l o o d K + prmrts to recogn i z e t h e n a t u r e o f t h e c h a n g e sc o n c e r n i n gt h e i n t e r v e n t i o no f t h i s e l e m e n t . I n a c a s e
of periarteritis nodosa, the bigh values of serum potassium and low values of tbc
total blood potassium indicate an offbalancetype D.

,\!L1t:t4u1Jldt

n /\

i\

V \iV
\

F t c . 2 1 5 . L o w v a l u e so f s e r u m p o t a s s i u ma n d h i g h v a l u e s o f t o t a l b l o o d p o t a s s i u m
i n d i c a t ea n o f f b a l a n c et y p e A i n a c a s eo f c a n c e r o f t h e g a l l b l a d d e r .

NorEs

573

set-up of the apparatus.The blood was diluted with a lvo aceticacid solut i o n i n t h e p i p e t t cu s e df o r c o u n t i n gw h i t c c e l l s .T h e p i p e r t ew a s s h a k e na s
for the count of cells, and the necessaryamount taken from the diluted
content.The potassiumamount was determinedand the result multiplied
by 10. while the averagevalue for the total blood was found to be around
3 8 m E q . ,v a l u e sa s l o w a s 2 0 o r a s h i g h a s 6 0 w e r es c e n (. F i S s . 2 l 4 , 2 l S ,
and 216)

tr4

G
J

60
.ft

7,

on

l l l l | | l | l

(u

a
g

\z

1e
4o
l5
3o
2t

o
o

fo

1o
4z
(g
JE
o

lt

:P9

c
+
Y

F t c . 2 1 6 . L o w v a l u e so f p o t a s s i u mi n s e r u m a n d i n t o t a l b l o o d i n d i c a t ea q u a n t i t a t i v e
d e f i c i e n c y i, n a s u b j e c tw i t h a l i v e r a d e n o c a r c i n o m aT. h e a d m i n i s t r a t i o no f 4 0 m E q
KCI daily, for 9 days brought the two curves to normal.

Chapter 4, Note 9. Sulfhydryl Determination

The catalytic effect of sulfhydryl groups on the oxidation of sodium
a z i d eb y i o d i n ew a sf i r s td e s c r i b ebdy F . R a s c h i g( 2 1 4 ) , a n d F . F e i g l ( 2 1 7 )
utilizedit to developthe most sensitivequalitativetest for the presenceof
s u l f h y d r y lc o n t a i n i n gc o m p o u n d sT. h e r e a c t i o n i, n i t i a t e db y m e r c a p t a n s ,
s u l f i d e st ,h i o s u l f a t easn d t h i o c y a n a t etsa, k e sp l a c ea s f o l l o w s
2 NaN', + I,r =, 2 Nal * 3 N,,
While this equation implies that the sulfhydryl compounds do not take
part in the reaction,this is not entirelycorrect,becausesimultaneously,the
sulfhydrylgroupsare oxidizedby the free iodine. Accordingly,the reaction
is carriedout in a Warburgapparatus,where I ml. of 0.2 M sodium azide

574

R F . S E A R C } II N

P I I Y S I ( ) T ' AT T I O L O C Y

Poly+rurtrrrtd

Bhlne

alcohols

Aftrr
Aftrr

Beforc

Codliver oll
fatty aclds

Octanol
t

.i

.1

l
^
LJ\4Jr'*'Lq"/

Aftr

Before

i.^^.^----^**

After

Before

Eeforc

lable
Ursapontf
frrctlon of

Epichlorohydrtn
II

,llvqr.r^.,^.-.*.rrJ

rr.-rb,t+!*L\L
:

Beforc

'

!.

h^tn^n",ra"y
Aftcr

I.

::.

l
i
F*l.\fA+,trfl
Eefoie

r4^/tr4.A/

After

Aftr
Sodtm Thl0sulfate

t-:

{+^,'--- t ,

Beforc

I : r c . 2 1 6 l J . l i l c c t r o c a r t l i o g n r n r si n. { i r r t l e i r d i n r u b b i t r i n j c c t c d i n t r a p t r i t o n e a l l y w i t h
s u b l e t h ; rdl o s c sr . r fd i l l c r c n t i i g c n l \ . l n t h e g r o u p u ' i t h l i p o i d s * ' i t h p o s i t i v ep o l a r g r o u p s
b e s i d c so t h c r c h a n g c s .i t l l a t t c n i n go f l h c r l a v c T i s i n r l u c e d ,w h i c h c o n t r a s t s w i t h
n m o r e e l e v a t c dT f o r t h e g r o r r p o f l i p o i t h i v i t h n c g a t i v cc h a r a c t e r .

ioclidcsolutionsarc rnixcd.while the

and I ml. of 0.1 IVtiodinc-potlssiunr
sulfhydrylcontainingsolutionis kcpt in thc sidearm,and then addedto the
rcagentsafter tentpcraturecquilibriumis reuchcd,Upon completemixing,
therc is rapid nitrrxcn cvolution,which, lrowevcr.ceaseswithin l3 minutes.Thc anlount of nitrosrn cvolvcdis found to bc linearly proportional
of the sulfhl'drylgri)ups.lrnd on the avcrage,1 ml. of urine,
to the soRtL-nt
ml.
of
blood
or I nrl. of a 3.x l0 1 lv{ sulfhydrylcontainingsolution
0.05
are antplysufiicicntfor irn irs\av.J'hc nrethodis thus rvellapplicableto the
of suifhrdrl'l levcls,providcdthe nrcthodis standardized
detcrnrination
with
thc appropriatccontpoundto trc testcd.ls thc catalyticctlect of all merc i l p t i l n si s n o t t l r c s a r n c .{ : i N )

NOTES

5'ts

Chapter4, Note 10. Calcium in Urine

I cc. of the urine was diluted in a test tube with 8 cc. of distilled water
and the optical density of the mixture was determined.To the mixture,
I cc. of a lVo solution of potassiumoxalate and 3 Va of.oxalic acid was
added.After standingfor 5 minutes,the tube was shakenand the optical
density again was read. The difference,multiplied by 10, was divided by
the two figures of the specificgavity of the sample. The value obtained
was called the calcium index.
Chapter4, Note I l. Urinary SurfaceTension (ST)
The role of changesin the surfacetension of various body fluids in
normal and abnormal physiologyhas becomeof increasinginterest.Some
authorshave gone so far as to considerthe surfacetcnsion forces present
at the interfacesseparating
entitics,to be the most importantfactorsin the
boundaryformationswhich serveto individualizetheseentities.
Consideringmultiple aspectsof the problem, it appearedinterestingto
attempt,as a first step, to obtain information about the surfacetensionof
different body fluids. It was as part of this program rhat urinary surface
tensionwas investigatedwith the intention of utilizing the data to gain insigbt into changesrelatedto the dualisticoffbalances.Before we could proceed,it was necessaryto resolvescveralproblems,including the technical
difrculties in measuringsurfacetensionthat result from the specialconstitution of the urine.
Technical Problems
Successivemeasurementsof surface tension, when made on fluids
formed by a single substance,consistentlyfurnish the same value. But for
fluids composedof two or more constituents,values vary from one moment to the next. This is explainedby the fact that moleculesof constituent
substanceshave a tendency to migrate in the fluid, some accumulating at
the surface,others concentratingin the bulk. (Gibbs dictum) The surface
tension of difterent complex fluids has been found to vary according to
the nature and amount of tensio-activesubstancespresent.And, study of
the variations has furnished information about the nature of these substances.
In a fluid such as urhe, containingmany differentsubstances,
the problem of variations in surface tension is a major one. ST measurements,
made without consideringthese variations, would be subject to serious
errors. Examination of different samplesof urine has shown great difterencesbetweenvaluesobtainedat differenttimes.Using Lecomtedu Noiiy's
tensiometer(215) it could be seenthat, for the sameurine sample,values
vary accordingto the length of time the sample is left to stand. Values
progressivelydecreaseas standingtime increases.Similar changesare seen
when the pendantdrop method is used. (216)
Becauseof the fact that a certain time is neededfor changesto take

576 /

n E S E A R c Hr N P H Y S r o p A T H o L o o y

place, the relationship bctwccn change and time was investigated.The

study of variousurine sanrpleshas emphasized
the inequalitywhich exists
betweenthem not only in the intensityof changesbut also in the time
necessary
for the changesto take place.This fact has rendereduselessthe
measurement
of the surfacetcnsionof differentsamplesif all are made at
somegiven moment.Except for measurcments
made at frequentintervals.
u s e o f d u N o i i y ' s t e n s i o m c i c hr a s a p p e a r e dt o b e i n a d e q u a t ef o r u r i n e .
Traube's sta.lagmometer
also is unable to furnish values that take these
changesinto account.
Theoretically,it would appear possibleto obtain measurementsthat
would correspondto the surfacetension for each drop at a desired moment by changingthe rate of flow of the urine through the apparatus.But
the differencesbetweenurines, related to changesin distribution of components,have made this inadequate.
With the pendant drop method, progressivechangeswhich occur in
the shapeof the drop would appearto indicatethe changesin surfacetension. (216) Technically,it would appearnecessaryto obtain data as frequently as possiblein order to follow changeswhich occur at various times.
By usingserialpictures,the changes,the momentof their occurrence,and
their intensitycan be studiedaccurately.Unfortunately,the complexityof
the method,with the needfor frequentpicturesand involvedcalculations.
prohibitsits usefor routinemeasurements
and.consequently,
for any broad
c l i n i c a la n d e x p e r i m e n t ar e
l search.
It was under these circumstances
that we returned to the capillary
methodwhich we considered
capableof furnishingthe desireddata. Classically, the heightof the ascendingcolumn in a calibratedcapillary is used
to calculatethe surfacetension.Height alone,however,is unsatisfactory,
sinceit doesnot rcvealthe changesthat takc place.It was by studyingthe
descentof the column in a capillarythat we were able to obtain the data
which we were seeking.We could show that the column does not descend
with uniform velocity. It stops or slows down perceptivelyseveral times
before it comesto rest at a fixed value. We could recognizethat, for most
urine samples,there is a first stop usuallyof severalsecondsduration. In
some urines,this first stop is replacedby a marked slowdownin velocity
of descent.
The stopor slowdownis followedby renewedbut slowerdescent
and a secondstop somewhatlongcr than the first. After anotherdescent,
often lastingmore than 20 minutes.a new stop occurs.
The time of descent,the duration of the stops, and especiallythe
heightsof the column at which the stopsoccur, while reproduciblefor the
sameurine, vary widely with differentsamples.They would thus indicate
differentrepartitionsand the times whcn they occur. This techniqueof
using the capillary consequently
appearsto be adequatefor the study of
the surfacetension of complex solutionsand particularlyfor the study
of urine.
Each of the heightsat which the descending
column stopswould indicate the surfacetensionfor a particularstagein the repartition of the constituents.In studyingthis problem further, it appearedadvisableto try to

NorEs

5'17

have the capillary so calibrated as to permit a direct reading of surface

tensionvaluesat thesestops.The study of the relationshipbetweenthe
surfacetensionof a fluid and the heightof the column has indicatedthe
nature of interveningfactors,their values,and under what conditionsa
direct readingis possible.
The fluid column remainsstationaryin a capillarytube when the surface forceswhich bind the column of fluid to the wall of the capillary are
equal to the weight of the fluid column.
With o representingthe surfacetension; r, thc radius of the capillary
tube; h, the heightof the column; A, the specificgravity of the fluid; and g,
the acceleration
of gravity,we havc 2 zrr ,r
zrrr hg A. It can be seenthat
the specificgravity is the only factor relatedto the sample,other than the
surfacetension,which intervenesin determiningthe height of the ffuid
column.
Accordingto this formula,the relationshipbetweenthe surfacetension
o 'lt
and the specificgravity of thc specimcnis. ,,
The sameheiglrtof
the column is obtainedif the relationshipbetween,,lrfu.. tensiono arrdo'
of two difterentliquids with the specificgraviticsA and ,r' fulfills the conclltton: n =

a'L'
A

lf mcasurements
with a capillarytube havinga bore radiusof 0.5 mm.
are madein New York City, whcre the acceleration
of gravity is 981 upon
water which at 18"C has a surfacetension73 dynes/cm.,the heightof the
fluid column is found to be 6.0 cm. and the rclationshipbetweeno and A,
expressedin the cgs. systemis o : 73 A.
A capillary tube thus can be calibratedto permit the direct readingof
the surfacetension in dynes/cm. for any liquid having the same specific
gravity, For fluids of different specificgravity, rhe samecapillary tube can
be usedif a correctionof 0.073 dynes/cm.,is made for each 0.001 increment of the specificgravity.
Urinary specific gravity values encounteredclinically range between
1 . 0 0 1a n d 1 . 0 3 5 ,w i t h a n a v e r a g ev a l u e a r o u n d 1 . 0 1 5 .T u b e s c a l i b r a t e d
to measureurine specimenswith specificgravity valuesat either extreme
can yield errorsin the surfacetensionof as much as 2 dynes/cm.In order
to minimize the degreeof error for routine laboratory use, the capillary
tube has beencalibratedto correspondto a fluid with a specificgravityof
1.015.The maximum error of the surfacetensionvaluesfor the extremes
of specificgravityclinicallyobscrvedwill be reducedto approximately+ I
dyne/cm.in this way. Furthermore,the fact that sodiumchlorideconcentration is one of the importantfactorsinducingdiffcrentvaluesfor urinary
specificgravity reducesthe influenceexercisedby specificgravityupon the
hcightof the column. Sodiumchloriderepresents
a negativesurfaceactive
It will raise the surfacetensionvaluesas its concentrationinsubstance.
e f i t s t c n d c n c yt o n r i u r a t cf r o m t h c s u r f a c ct o w a r dt h e b u l k
c r e a s cbs c c a u s o
of the fluid.This will partiallydecreasc
the influenceexertedby the specific
gravityof the urine, Sincethe surfacetensionvaluesof human urine speci-

578

R E s [ . A R c Ht N p H y s r o p A T H o L o c y

mensmeasuredby this methodhavebeenfound to vary between73 and 50

dynes/cm.,the error for extremevaluesof specificgravity is less than 5%
and is not clinically significant.If more precise values are desired, the
necessary
correctioncan be madeby addingor subtracting0.073 dynes/cm.
for every .001 differencein the sampleaboveor below the specificgravity
f o r w h i c ht h e t u b e i s c a l i b r a t e d( i . e . 1 . 0 1 5 ) .
The temperatureof the urine to be testedis anotherfactor which intervenes.Although the fluid rapidly attainsthe sametempratureas the capillary walls, it is advisableto perform measurements
when the temperature
of the fluid is around l8oC, sincethe surfacetensionof a liquid decreases
as its temperatureincreases.For clinical use, correctionsfor differencesin
temperatureare not considerednecessary.
Design and Calibration ol the Urotensiometer
In order to obtain direct readingsof the urinary surfacetensionvalues
with a maximum error of + I dyne/cm.,the previouslydiscussedfacton
were taken into consideration
in designingand calibratingthe urotensiomwhich
we
(FiS.
eter
conceived.
217) The glasscapillary tube has a bore

14cm.

Ftc.2l7. Urolensiometer-calibratedto indicate by drrect readings in dyne/cm

t h e s u r f a c et e o s i o no f f l u i d sh a v i n ga s p e c i f i cg r a v i t yo f 1 . 0 1 5 .

diameterof0.5 mm. and is approximatelyl4 cm. in length.It is calibrated

to indicatethe surfacetensionof a fluid with a specificgravity of 1.015
directly in terms of dynes/cm, in the following manner: a continuous
columnof distilledwater at 18"C is drawn up to about three-fourthsof the
height of the tube and allowed to descendwith the tube maintainedvertically. The point at which the top of the column stops is marked. It representsa surfacetensionof 73 dynes/cm.for water having a specificgravity
of 1.000. In order to make the necessarycorrection for a fluid having a
specificgravity of 1.015, thc distancebetweenthis point and the tip of the
tube is dividedinto 74 (insteadof 73) equal parts.The tube is calibrated
down to 50 dynes/cm.sincelower valueshavc not been encountered.In
the tubes manufacturedby Clay Adams, New York, the markings are permanent.The split line featureof the scalepermits easyvisualizationof the
meniscus.The encirclinglines help in maintainingthe tube in the vertical
position.
The Measurementol UrinarS,Surlace Tension with the Urotensiometer
To determine surface tension by means of the Urotensiometer,tbe
taperedend of the tube is introducedinto the bulk of the urine specimen.

NorEs

579

The fluid is drawn slightly above the highestmark by mouth suction and
evacuatedseveraltimes by positive pressure.The tube is again filled to the
same point, care being taken this time that no air bubbles interrupt the
continuity of the fluid column. The tube is removedfrom betweenthe lips,
and the tip of the capillary is then gradually raised toward the surfaceof
the fluid. When the top of the column descendsto the top line (T) of the
scale, the tip of the tube is removed from the fluid and maintained in a
vertical positionat eye level. The descentof the top of the column can best
be observedby viewing the meniscusbetweenthe ends of the split line
calibration markings. The top of the column descendswithin one or two
secondsto an initial point (P1) where it comesto a temporary halt or its
rate of descentsuddenlyslows perceptibly.The column again slowly descends,coming to rest after severalminutesat a secondpoint (P2). After
some time, the descentmay again be resumedat a much slower rate until
a third and final stopping point (P3) is reachedafter more than fi.fteen
minutes.For routine measurements,
the first reading (Pr) is consideredas
the surface tension value of the urine. This correspondsrougNy to the
surface tension value of the specimenbefore any important secondary redistribution of moleculeshas taken place.
The capillary tube should be thoroughly cleaned with distilled water
after use. It is well to check the tube before each seriesof measurements,
using distilled water at room temperature.If the check readingsare above
74 or below 73 on the scale,the tube must be carefully flushed through
with distilledwater by meansof a suctionpump. Occasionally,water alone
may not be srrfficientand it will be necessaryto clean the tube with sulfuricochromiccleaningsolution,followed by thoroughflushingwith water,
in order to obtain correct check readings.when the tube is not in use, it is
best left standingin a gJassbeakercontainingdistilledwater.
Surlace Tension in Clinical and Experimental Research
The Urotensiometerfor the first time makespossibledeterminationsof
the surfacetensionof urine and other physiologicalsolutionsas a routine
laboratory procedure. The highest surface tension value for urine encounteredclinicallyis 73 dynes/cm.,and this is correlatedwith a minimal
quantity of surface-activesubstances.
The lower the surfacetensionof the
urine in dynes/cm.,the greaterthe amount of tensio-activeagentspresent
in the specimen.A surfacetcnsion of 52 dynes/cm. is the lowest clinical
value that we have found by this method in more than 100,000 measurementsmade during the last 12 years.
The first problemconcerningthe meaningof the differentvaluesof urinary surface tension arose when it was observedthat usually the urines
with low specificgravity have high surfacetension,while those with high
specificgravity have low surfacc tcnsion. The direct correlation between
the valuesof surfacetension and specificgravity of the samplesthus had
to be investigatedwith the suppositionthat the amount of water in the
urine will have a great influence,by itself, on surface tension. While a
correlationbetween surface tension and water content is often observed.

580

x E S E ^ R C Ht N p H Y s l o p ^ T H o L o c y

it is not a causeand effect one. Urinary sampleswith a specificgravity as

l o w a s 1 . 0 0 3 w e r c s e c n w i t h a s u r f a c ct e n s i o no f 5 8 d y n e s / c m .w h i l e
samplew
s ith a specifig
c r a v i t ya s h i g ha s 1 . 0 3 0h a d a s u r f a c et e n s i o no f 7 0 .
Although very seldomencountered.thesevalueshave invalidatedthe supposition that it is the amount of water in the urine which determinesthe
value of the surfacetension,so that from the analyticalpoint of view determination cannot be substituted.
The Nature ol tlu Intervening Substances
The existenceof severalvaluesfor the surfacetensionof urine has suggestedthe interventionof differentsubstances
in the dcterminationof surface
tension.we used the study of the changesinducedin the three valuesof P
obtainedfor a sample.Differentrepartitioncapacitieswere consideredas
correspondingto differentgroupsof substances.
Severalmethodswere used
in order to identify thesesubstances.
In one method, different constituents
of the urine were separatedby usingsolventsor absorbents,
or by allowing
the constituents
to assemble
at the surfacc.
The fact that the solvents,if they remain in the fluid even in very
minute amounts,influencethc surfacetcnsion,has largely handicapped
their use.However,when lipid solventswere usedand could be thoroughly
eliminated,the treatedurine showeda changein surfacetension,especially
in P1 values.With the use of activatedanimal charcoalabsorption,all the
P valueswere changedtoward highervalues.
M . B i e r i n o u r l a b o r a t o r i e sh a s s t u d i e dt h e n a t u r e o f t h e s u r f a c e activeconstituents,separatingthem from urine by using the fact that they
assembleat the surface.Urine was made to foam by passingan inert gas
throughit. The foam-and, with it. a high proportionof surface-active
substanceswas separated.
By repeatingthe procedure,the separationcould be
pushedfar enoughso that it could be seenthat the ST values,especially
thoseof P3, were influenced.Analysesof the fractionsobtainedindicated
that lipids would intervenein determiningthe surfacetension revealedby
the Pr value,while proteinswould intervenefor the Ps, i.e., after a repartition requiring a specifictime. we have tried to confirm these preliminary
databy addingthe agentsto urine and followingthe changesinduced.
The additionof minimal amountsof soapsto urine has been found to
inducea changein all P valuesand especiallyin P1.The additionof billiar
saltschangedPr values,while the addition of proteins,such as albumin,
influcnccdthe valuesof P3.lt would appearfrom this preliminaryresearch
that while P1 changesare relatcdto an increasein fatty acid derivatives,
P3 changesare relatedmore to the interventionof billiar acids.while prot c i n sa n d a m i n oa c i d se x e r tg r e a t e ri n f l u e n c e
o n t h e v a l u e so f P 3 .
This explainswhy surfacetension,corresponding
to P,. is still high in
urinesrich in albumin, and sometimesalso in those with billiar acids. It
would also explainthe observationin the Hay's Test with sulfur flower in
urine,that the sulfur startsto fall quickly if the urinc is left standingfor a
while, but for the sameurine this fall occursonly after a certain time if
the sulfuris addedto urine inrmediatclyafter stirring.With surfacetension

N o r Es

581

affectedby fatty acidseven in minimal amounts,ST changesin relationship

to conditionswhere thesesubstancesinterveneare particularly interesting.
It is chiefly with these data in mind that we tried to investigatesurface
tensionin relationto normal and abnormalphysiology.
Surlace Tension and Normal ond Abnormal Physiology
The ability to measure surface tension rapidly and accurately enough,
even for very small amounts of fluid, has made it a preferred method for
many investigations.In addition to clinical applications, where the information furnishedhas beenespeciallyvaluable,we have utilized this method
in experimentsin animals.
Time ol the Day and Urinary Surface Tension
Urinary surface tension measurementswere made in several normal
subjectsat hourly intervals.In order to eliminatethe influenceexertedby
exerciseand food, the subjectswere kept in bed for a few hours preceding

:p

olChlqhl r-

I.-

F t c . 2 l E . 2 4 - h o u r s h o u r l y u r i n a r y s u r f a c e t e n s i o n v a l u e o f a 3 O - y e a ro l d n o r m a l
male, kept resting and with a constant hourly food intake, showing a maximum in
the afternoon and a minimum around 5 a.m.

measurements.
During this period, and the entire period of the experiment,
the subjectswere perrnitted to leave their beds each hour to void. Throughout the experiment,they were given the samekind and amountof food each
hour. This eliminatedas variablethe influenceof food and activity.Figures
218,219 and 220 and Table XXVI show samplesof the curvesof surface
tensionin suchcases.A 24-hourdiphasiccurve can be noted.
Surfacetensionin mice undcr similar conditions,however,showsdifterences.A group of 20 mice kept in cageswere used.By slight squeezingof

s82

R E S E A R C H

I N

P H Y S I O P A T H O L O C Y

26 yr mon on stondordrzed
hwrly fad oN lluid rntotc

E
(,
o
c
D
c
o
6
c

e
c,
f

v)

t2

t2
fVoon

lrloon

F t c . 2 1 9 . C u r v e o f t h e u r i n a r y s u r f a c el e n s i o ni n a 2 6 - y e a ro l d m a l e o n s t a n d a r d i z e d
hourly food and fluid intake showinga maximum toward the early morning hours
and a minimum toward the evening.

the lower abdomen,a few drops of urine were obtainedin a little cup and
usedfor surfacetensionmeasuremcnt.
Changesseenin Figure 221 show
that with the passage
of time,thereis a dampeningeffecton the curve.This
has made us doubt that the interventionof strcssin thesecasescan be responsiblefor the changes.In order to eliminatestressas a factor, a second
group of experimentswas done in which urinary sampleswere obtained

rcon

e---

o
D!.

/
o
'----

'

t
----J

a
rta^toht

'

+I.------J

Ftc. 220. Curve of the urinary surface tension in a 27-year old female on stAndard
hourly feeding, with a maximum in the afternoon and a minimum in the morning.

NOrES

,/

583

65

64

:63
E
()

v)

i62

s
;

6l

(l)

()

60
f,

u)

59

58

t2
Noon

g t o 1 2
2 4
I|idnjgh!

t o 1 2
Noon

F t o , 2 2 l . A v e r a g e v a l u e o f s u r f a c el e n s i o n i n t h e u r i n e o f 2 0 m i c e . o b t a i n e d e v e r v
h o u r , s h o w i n g v a r i a t i o n sw i t h a d a m p e n i n gc h a r a c t e r .

eachhour from a differentanimal.Thus,eachanimalprovideda sampleof

urine only once and was not under stress.Under these conditions, the
dampeningeffect was not present.A curve showingonly two phasesn 24
hours was obtaincd.The ST curvcs for humansand mice are opposite.
During the period when high valuesoccur in humans,low valuesoccur in
mice, and vice-versa.Becausesuch oppositevariationsbetweenhumans
and mice were found for many other analyticaldata and were considered
relatedto the nocturnalactivityof mice, a third group of experimentswas
performed in which the mice were kept in darknessduring the day and
under Iight during the night in an attempt to changethe rhythm of their
activity. After three weeks, there were no marked changesin analytical

584

nESEARcHrN pHysropATHoLocy

dataobtainedin thesemice.It is possible,

however,that more time is requiredto inducechanges
in surfacetensionby alteringthe rhythmof mouse
activity.(Fig. 222)
T,'rsLe XXVI
Sunrrce TeNsloN rx NoRprrrl HerlrHy 32 ye.rn Or_oM,rx rNo
27 Yern Oln Feurlr ox SrrNolno HounLy Ft'eolxc

!g!r
7 a.m.
8 a.m.
9 a.m.
l0 a.m.
I I a.m.
12 Noon
I p.m.
2 p.m.
3 P.m.
4 p.m.
5 p.m.
6 p.m.
7 p.m.
8 p.m.
9 p.m.
l0 p.m.
I I p.m.
l2 Midnight
I a.m.
2 a.m.
3 a.m.
4 a.m.
5 a.m.
6 a.m.

Male
S.T. in Dynes/cm.
65
64
65
67
69
'70
7l
7l
1t l^

72
73
73
72
70
69
68
67
67
66
66
65
65
63
64

Female
S.T.in Dynes,cm.
66
61
67
69
70
70
72
73
73
72
73
72
7l
7l
'lo
IU

67
67
67
65
65
66
66
65

Surlace Tension in Normal Humans and Animals

From the first analysesof urinary surfacetensionin groups of individuals it could be seenthat certainchangesconlmon for all were taking place.
There were days when all subjectshad higherrelativevaluesand other days
when lower valuesprevailed.Sincctherewas no common dieteticor habit
factor for all the subjectsstudied,we searchcdfor environmentalchanges
that might be the immediate cause of these variations. In collaboration
with P. Teitelbaum we made the following experiment,using 80 rats
divided into four groups.Onc group consistedof fcmalesof the Wistar
strain and a secondconsistedof nralcsof thc sarnestrain.The remainin.e
two groupswere composedof 20 femalesand 20 malesof a black hooded
strain.The animalswere maintainedin groupsof five in separatecageson
Purina and water ad lib. They were kept in a nonconditionedroom. The
experimentwas conductedfor one month.from May to June.

!
'
'

t. I
,
I

:,r
:.:1': '

i'l
:

NorEs

585

o
o

F.

c
.s
q,

U
o

(,

tz
,\roon

t2
Mtdnght

to

t2
IVoo.t

F t c . 2 2 2 . A v e r a g e h o u r l y v a l u e si n t h c u r i n a r y s u r f a c et e n s i o no f g r o u p s o f 5 m i c e ,
t h c g r o u p b e i n g c h a n g e de a c h h o u r .

Urine was collectedin a small vesselby kecpingthe animal firm and

pinchingthe lower abdominalskin. Surfacetensionwas measureda few
minuteslater. In each group, animalswhich did not give urine under this
procedurefor severalconsecutive
days were replacedby others.
Samplesof urine wereobtained6 daysa week,between9 and 10:30 in
the mornrng.From the data obtained,an averagevalue was calculatedfor
each group, and the respectivevalues were plotted in curves having the
days as abscissae.
The valuesfor the femalegroup were higher than for
males.No differenceswere seen betweenthe two strains.And all four
curvesshowedthe samevariationsat the sametimes.Thus it appearedclear
that the variationswere relatedto someexternalfactor acting upon all the
animals.We compared the ST curves with others traced for different environmentalvaluespresentat the tinte of observation.Such y3lus5-!s1smetric pressure,electrostaticvalue and temperature-wcre obtained fronr
the WeatherBureauand the curvesfor the arca at the hour of the experim e n tt r a c e dO
. f t h e ma l l , o n l y t h c c u r v cf o r t c n r p e r a t u rceh u n s ew a s s i g n i f i c a n t .T h e S T w a s s c c nt o r i s c c a c h t i m c t h l t t h c t c m p c r a t u r cf c l l a n d f a l l
w h e nt h e t e m p e r a t u rreo s e .( F i g . 2 2 - )l
This correlationwas further studiedby usinginduccdratherthan natural temperaturechangesin the followingexperimentwhich was made in
collaborationwith E. F. Taskicr.
Adult female CFr strain mice were divided into three groups of 20
mice each. They had ad lib accessto food and water. One group was
placedin an incubatorin which the temperaturewas maintainedat 37oC.
A secondgroup was kept in a refrigeratorat 8"C. The third group served
f o r c o n t r o la n d w a s m a i n t a i n c da t o r d i n a r yI a b o r a t o r yt c m p c r a t u r ew h i c h
ranged between2O-25"C.

586

RESEARCH IN

PHYSIOPATHOLOCY

Becauseof the diurnal patternof surfacetensionvariations,urine specimens were collectedat the same hour every day. During a period of. 22
days, daily urine specimenswere obtained between9 and I I A.M. This
was easily accomplishedby firmly gripping a mouse in one hand by the
scruft of the neck and tail. with the finger of the other hand, the lower

q)

6
a

6
O

{!
aI

(o 6 t
E
l!

a
g

t:

6a

JZ

o
L
:f

o
L
o

E
o

h'

tt

(u
tJ
q
0
o
L

: 99

o
E
o
L

:Y9rn

4)

So
>\

6o
E
E
l

1o
2o

22

2'
UAVS

2t

at

h t c . 2 2 3 . C o m p a r i s o n b e t w e e nw e a t h e r d a t a a n d t h e a v e r a g e v a l u e o f t h e s u r f a c e
t e n s i o ni n 4 0 m a l e r a t s a n d 4 0 f e m a l e r a t s . I t s h o w s a r e l a t i v e p a r a l l e l i s m w i t h t b e
c u r v e o f t h e b a r o m e t r i cp r e s s u r ea n d a m o r e c o n s i s t e n rt e l a t i o n s b i pw i t h t h e i n v c r s e
curve of the temperature.

NorEs

587

aMomen was gently massaged,causingthe animal to void 2-5 drops of

urine into a small glasscup.
The surface tension of each specimenwas determinedwithin a few
minutesafter it was obtained.
The valuesobtainedfrom day to day were charted for each individual
mo'|rse;the averagefor each group maintainedunder different temperature
conditionsalso was determined.

F t c . 2 2 4 . A v e r a g ev a l u e o f s u r f a c el e n s i o no f t h e u r i n e i n c o n t r o l m i c e o v e r a p e r i o d
of 3 weeks.

The averagesurfacetension readingsin the control groups are shown

in Figure 224. It can bc sccn that thesevalueslluctuatedin an irregrlar
fashionbetween58 and 63 dynes/cm.The surfacetensionvaluesof the
group maintainedat 37'c show a steadysustainedrise from 6l to 65
dynes/cm.(Fig.225) The mice kcpt at 8oc showedan initial slightfall in
surfacetension,with a gradualreturnto the originallevels.(225)
After severaldays in the incubator,the micebeganto loseweight,their
fur becamesparse,and snout areaswere constantlywet. The urinary output
was scantyas comparedwith the two other groups.Death beganto occur
in the mice kept a high temperatureon the 12th day. The animalsin the
refrigeratordevelopedthick luxuriant coats and huddled closely together
at most times.None died from exposureto this temperature.
The fact that the animalsmaintainedat 37oC showeda steadyrise in
urinary surfacetensionwas especiallysignificant.As expected,the urine of
theseanimalswas scantyand more concentratedthan for the other groups.
With this diminution in volume, it would be expectedthat the concentration
of surface-active
substances
would rise and the surfacetensionwould be
lowered.The fact that the exact oppositeoccurredindicatesthat the observedchangehas to be consideredas an effect of the high temperature.
This appearsespeciallyinterestingfor the relationshipbetween temperatureand the two ST patterns.While higher temperatureinducesone

588

RESEARCH IN

PHYSIOPATHOLOOY

lemole fi,'Ee kepl 6

tncubolor ot 37oC
^

41

962
;
o

--

:50

/
Y--

-r'

2O temole mtce hegt n

relrgerotot ot FC

O
Doys

Frc. 225. Tbc average values of the urinary surface tension of 20 female mice kcpt
in the incubator at 37"c. and of 20 female mice kept in refrigerator at 8"C. The
values are progressivelyincreasing for the animals kept in the incubator until the
a n i m a l d i e d . F o r t h e a n i m a l s k e p t i n r e f r i g e r a t o r ,a f t e r a n i n i t i a l d e s c e n t .t h e v a t u e s
a s c e n dt o w a r d n o r m a l .

pattern,cold inducesthe other. It must be noted that the organismcannot

defend itself against the pattern induced by higher temperatureand the
animal dies after a certain time, but for the pattern inducedby cold, defenseis possible.The body seemsto be able to overcomethe change.The
surfacetension returns to normal and the animal becomesadapted to the
Not one of the animalskept in the refrigeratordied, while all
temperature.
i n t h e i n c u b a t o rw e r e d e a d a f t e ra m o n t h .A d r e n a l e c t o miyn d u c e sa n i m m e d i a t ci n c r e a s e
i n t h e s u r f a c ct e n s i o no f t h e u r i n e .( F i 5 . 2 2 6 )

l,:-

()
qJ

.cc

t(-r,

at

att
c

/E

-x-/

67
/-l+

Pne-ac'rx.
Cays

Post- adr x.
days

Ftc. 226. Tbe surfacetensionof tbe urine increasesto high values after adrenalcctomv.

5E9

NOTES

r
I

lent/or) chong{$ rf, urrn;

:jurfote
rf\e(inp^
ol 16 rt ol4 O.eg@l
tonon

:
'
F

b).

:
Oeltrarg4

c CtL).
0

;
c

lday 7 i

t
l
I
e 6iJ

5 0 tt
Ut

\ I

I
Y

. -,
l

lwv

Oec

1l

2r

'rvn

aC?r

6
Fab

t4

t?

t
il,or

t t

2 t

5 u t 5 2 Q t f ? f

Aq

tloy

Frc. 127. Curvc of the surface lension changes in urine specinrens of a 36-yr.
old prcgnant woman shows a manifest change toward low values, starting with thc
4th month, and becoming especiallylow in the last three months.

I
l4J
(t)
J
q

<t

o
d.9

+7O 69-66 65+7O 69-66 65(Oynx/cn)

URINARY SURFACETENSTON
Frg. ??8. The average value of the surface lensi()n in pregnant wonlen shows a
manifcst changc toward low values.

590

xEsEARcH rN pHyslopATHoLocy

Colloids in Urine ud Surlace Tension

An interestingrelationshipbetween urinary surface tension and the
presenceof "colloids" in the urine was noted by Butt and his associates.
(240) Using the pendantdrop merhodfor the ST, direct examinationin
dark field for the presenceof colloids in the urine, electrophoresisfor determinationof electricalcharges,and the study of evaporatedurine smears,
they showed that urines with a high content of colloids, have a low ST;
those with a low colloid content, a high ST. They have further correlated
a low amount of colloids with a tendencyof urine to precipitateand form
stones.(242) They examinedthe colloid particlesin the urine of different
groupsof individualsand found them high in Negroes,and especiallyhigh
in pregnant women, which is in accord with the low surface tension of the
urine which we found in these cases(Frgs. 227 and 228) and the low
tendencyof both groups to form urinary stones.
We were interestedin the relationshipbetweenvariationsin the urinary
content of colloids and systemicpatternscorrespondingto high and low
sr. R. Ravich in our laboratory has confirmed the correlation between
presenceof colloids and ST by using our urotensiometer.(219)
Chapter 4, Note 12. Urinary Oxidoreduction Potential
For the study of the oxidoreductionpotential of the urine, we used
a BeckmanpH meter with platinum electrodes.We measuredthe potential
at the pH of the sampleand also at pH 7. For this purpose,the platinum
and the respectivecalomel electrodesused for these measurementswere
introduced into the beaker of the Fisher titrimeter together with the electrodes of the potentiometer. After stirring, the pH of the sample and its
oxidoreductionvalueswere measured.The pH then was brought to 7 with
HCI or NaOH solution, and the value of the oxidoreductionpotential
was again measured.Four valueswere thus obtained: the original pH, the
titrimetricacidityor alkalinity,and the oxidoreductionvaluesat the original pH and at pH 7. Figures229 and 230 show a sampleof such curves.
Chapter 4, Note 13. Oxidoreduction Potential of the Urine
We have tried to determinethc oxidoreductionpotential of urine
samplesby usingthe changeof a color indicatorin its leuco base.We chose
toluidineblue which, with a rH2 of 14, is at the middle of the scaleof the
rH2 values. In order to eliminate two of the important factors which intervene in the oxidoreductionpotential-differencesin pH and temperaturc
-we used a fixed temperatureand very low pH. The degreeof oxidorcduction potential was determinedby the time necessaryto obtain the discolorationfor a standardamount of the color indicator. The reactive used
was a solution of toluidine blue in a normal solution of hydrochloric acid.
The amount was chosenso as to give a discolorationat 100 secondsfor the
normal individual. 1.5 cc. of a saturatedsolution of toluidine blue in al-

NOTES

59r

rr rt r.,lrr r5 ;7 rl r1

F r c . 2 2 9 . T h e c u r v e o f t h e o x i d o r e d u c t i o np o t e n t i a l o f t h e u r i n e m e a s u r e de l e c t r i cally. The curve of the measurementsmade directly on the urine (El) show big variations which are smaller if the pH of the sample is brought to 7 (E2). In a case of
cancer of thc breast,the curve remains constantly below the 0 valuc.

Frc. 230. Curve of the oxidoreduction potential values of the curve brougbt to pH 7
of a case of cancer of the breast,shows valuesaround or above the 0 value.

592

REsEARcH rN pHysropArHoLocy

cohol was addedto 100 cc, of n/10 hydrochloricacid. 1 cc. of this reagent
was addedto 4 cc. of urine in a test tube kept for a while in boiling water.
The time necessaryfor the discolorationwas marked. values as low as 3-4
secondsor as high as above420 secondswereseen.A high oxidoreduction
potentialinducing a rapid discolorationwas found to correspondto a pattern of the oftbalance typ A while a low discoloration was seen to corre-

C'
(t
-9f
q,

eI

x
o
to
aO
b

F t c . 2 3 l . C u r v e o f u r i n a r y o x i d o r c d u c t i o nv a l u e s i n a c a s e o f c a r c i n o m a o f t h e
breast with multiple bone metastases.The values are establishedas the time ncccssary to obtain the reduction at 100"C and with a pH around 2, of a solution of
toluidinc blue so chosen as to have 100 secondsas the average value for groups of
normal individuals. In this case thc values remain fixed low bclow 100 scconds,correspondingto a patterD,of the offbatancetype A.

spondto the patternof type D. Figures231 and 232 show two suchcurves.
We used this test for many years as main analysesto determinethe exist(220)
ing oftbalances.
Chapter4, Note 14. Peroxidesin the Urine
The hypothesis
of the existence
of a phase"oxygen"of offbalanceD led
us to study the appcarancein urinc of productsrcsultingfrom abnormal
oxidation.We were especiallyinterested
in the existenceof substances
having peroxide properties.We found that addition of sulfuric acid to urine
of certain subjectsinduced appearanceof indigotin and indigo'rubin. In
order to hvestigatethe reaction,we have utilized the solubility of indigotin
and indigo-rubin in neutral solvents.Through their extraction it appears
possiblefirst to preventtheir transformationin colorlessisatin and, second,

NorEs

593

to evaluatethe rclativeamountswhen they appearduring the reaction.To

4 cc. of urine,one centimeterof toluenewas added.After shakingthe mixture, I cc, o[ pure sulfuricacid was addedand the mixturewas immediately
s h a k e na g a i n .w h e n t h e m i x t u r ew a s a l l o w e dt o s t a n d ,t h e t o l u e n es e p a rated and its color, blue or violet,indicatedthe presenceand also the relative amountsof indigotin and indigo-rubin.
Another methodusedto detectperoxideswas the acidificationof urine
followed by addition of potassiumiodide.For ioclometricevalution,starch

1a

2oo

o
60
C'
f
E

o,

?Y
I

o
-E
>\
x

'@

9o
b
D
h
1o

Days
Fto. 232. The urinary oxidoreduction values in a case of cancer of the colon with
abdominal metastases.The values remain the wholc time above 100 scconds,corrcsponding to the pattern present in the offbalancetype D.

solution was added. The amount of iodine liberatedcould be determined

titrimetrically.
The form in which peroxidesare presentin the urine is not clearly established.Although the distillationof the urine givesperoxidesin the first
d i s t i l l a t et,h e s ca r e n o t i n t h e f o r m o f h y d r o g e np e r o x i d e ,s i n c ec a t a l a s e
doesnot inducetheir disappearance.
The valuesobtainedwith both methods, sulfuricacid and iodometric,are relativelyparallel.The sulfuric acid
method,however,produceda higherpercentage
of positiveresults.
The presence
of slightamountsof peroxidein the urine has beenfound
in about 3Vo of.normal subjects.[n contrast,we found peroxidein the urine
of.87Vo of a group of.27 schizophrenics
studiedthrough daily analyses
over a periodof threeyears.In someof the subjects,throughoutthe entire
three-yearperiod with more than 1,000analyses,
not a singlenegativere(
F
i
g
.
(
2
2
1
,
w
a
s
2-13)
action
seen.
222)
We have also found positive reactionsduring streptococcicinfection,

594

xEsEARcH rN pHystopATHoLocy

erysipelasor tonsillitis.In radiation sicknessthe reaction is positive especially when tissuelesionsare manifestsuch as mucositisor epidermitis.In
general,treatmentwith seleniumhas given a relatively high proportion of
positiveresults,especiallyat the beginningof treatment.While positive reaction appearedto be consistentwith a favorableevolution of tumors, an
extremelyintensivereaction appearedrelatedto a bad prognosis.The following observationis characteristic.

o
L

F I c . 2 3 3 . T h e r e a c t i o n f o r p e r o x i d e sr e m a i n s c o n s i s t e n t l yp o s i t i v e i n t h e u r i n e o f a
schizopbrenicin daily analysesduring a period of 3 years. (part of the curve.)

Mrs. N. C., 28 years old, with Hodgkin's disease,had received three

treatmentsof teleradiotherapy,with the generalcondition completely unchanged. The patient presentedan extremely intensive urine peroxide reaction.Dilution of the urine to l/50 still showeda marked blue color after
treatmentwith sulfuric acid and toluene.We informed the attendingphysician about the finding and advisedthe discontinuance,at least for the moment, of the treatment.The perfectgeneralcondition of the patient induced
the radiologistto disregardour advice.A new reatment of teleradiotherapy
was administered. Three hours later the patient, who only that morning
had been shopping,went into a shock and died during the night.
we want to emphasize,however,the correlation between the lack of
peroxidesin the urine and a poor prognosisduring radiotherapy.The cases
in which a positivereactiondisappeared,
were alwaysfollowedby a change
for the worse in the general condition. The persistenceof this lack of
peroxideswas seenin thc caseswith a rapid lethal termination.
Chapter 4, Note 15. Index of Excretion and of Retention
one of the most important aspectsof the relationshipof an entity to
its environmentis givenby its intakeand output.The conceptof hierarchic
organizationwith emphasison the individualityof the entities,has given a
specialmeaningto the study of theseprocesses.
For each entity its proper
environmentis representedby the secondarypart of the entity immediately
superiorto it. The nuclearsap represents
thus the environmentfrom which
the chromosomestake the material necessaryfor their metabolism and
wherethey rejectthesesubstances
which are no longer needed.Similarly,
the cytoplasmrepresentsthe environmentfor the nucleus,the interstitial
fluid for cells,the lymph for tissues,the blood for organsand the actual
environmentfor the organism.This systematization,
basedon the organizational individualityof the entities,has guidedthe study of the relationship

NorEs

595

betweenentitiesand their environmentsunder normal and abnormal conditions. It is under this aspectthat we have investigatedthe renal excretion,
which accordingto the hierarchicorganization,correspondsto the relationship betweenthe organismas an entity and its environment.
some of the substancesexcretedcome from the metabolismof lower
entities.Relatedto blood, they would representoften noxiousundesirable
substances,
if the higher mechanismof the blood would not intervene.Although that which we see as urine is the resultof the relationshipbetween
the organism as an entity and its environment,the origin of the different
substancesforming it, as related to the different other levels, has to be
considered.While for certain elementsthis origin is evident,for many substancesonly suppositionsare availabletoday.
when a systematicanalysisof theseconstituents
was attempted,other
difficultiesarose.The isolatedurine sampleseasily available,have only
a very relativevalue for many of theseinvestigations.
The titrimetricdata
expressedas concentrationof various substancesare all functions of the
amountof watereliminatedin the sample.As this often varieswidely,the
informationsobtainedare only relative.Balanceanalysesconcerningentire
intakesand outputs representsuch technicaldifficultiesas to make them
unavailablefor routine investigation,in which hundredsof subjectsare daily
studied. We tried to bypassthis difficulty by eliminating the factor water
excretion,from the considereddata. The fact that the concentrationof a
substanceand the specificgravity of the urine, are both direct function of
the amount of water present,has permittedto eliminatethis factor. The
ratio betweenthem appearsthus independentof the amount of water
present.It relatesthe amount of a substanceto that of the bulk of the
substances
eliminatedthroughthe kidney.An index of excretionwas thus
obtainedby dividing the concentrationof the substanceby the specific
gravity.The oppositeratio would correspondto an index of retention.
From the physiologicalpoint of view, these ratios are not aftected
by the factors which govern the glomerular filtration, which are acting
present.They are little or not affectedalso
similarlyfor all the substances
by the back resorption,wherethe differences
betweenthe varioussubstances
are reduced.They will show consequently,
big variationsas resultingfrom
which takesplacein the distal portionsof the conthe activereabsorption,
volutedtubes.It is this characterwhich givesthe indexesof excretionor
retention,as we calculatethem, their value. We have utilized for years
theseindexesfor chlorides,sodium, potassium,phosphoricion, sulfhydryl,
calcium, to obtain valuable information which otherwisecould not be
furnishedby the simpleanalysisof the isolatedurinesamples.We will come
back to theseindicesduring further analyses.
Chapter 4, Note 16. Water and Nitrogen Metabolism
The analysesof different urine sampleshave shown that the amount
of water presentin urine seemsto influenceindirectlyits constitution.It
could thus be seenthat while very diluted urine, correspondingto a large

596

nEsEARcH tN pHysropArHoLocy

amount of water excreted, is usually alkaline, concentratedurine, correspondingto small amountsof water excreted,is generallyacid. Furthermore, it could be seen that thesechangesare related to more profound
metabolic differences,When related to the nitrogen metabolism,it could
be seen that diluted urines are rich in free ammonia while concentrated
urinesin uric acid.We havethusinvestigated
the relationshipbetweenthese
two factors,the amount of water excretedand the form under which the
nitrogenis eliminated.
The comparativephysiologyshowsus that the manner under which
nitrogenis excretedvaries for differentanimalsaccordingto the amount
of water availablein the surroundingenvironment.In fish, with water
almostunlimited,nitrogenis excretedin the form of ammonia.The high
toxicityin this form of nitrogenexcretionis counter-balanced
by the amount
of water in which the excretaare diluted.Fish are ammonioselic.In terrestrialmammals,wherethe amountof water availableis more limited, the
excretionof nitrogenis made in the form of urea which is much lesstoxic
than ammonia.The dangerof poisoningthe drinking water throughexcreta
is thus reduced.Mammalsare ureoselicanimals.For birds,for whom water
is scarce,the form of nitrogenexcretionis of uric acid. which through
its low solubility in water, has little chanceto contaminatethe drinking
water.Birds are uricoselic.Basedon this relationshipbetweenwater availability and the type of nitrogenexcretion,we looked for a similar relationship in humansbetweenthe excretableamount of water and the rype of
nitrogenmetabolized
undernormaland abnormalconditions.An immediate
confirmationwas obtainedin thoseabnormalconditionswherethe amount
of water excretedis abnormal.As alreadyseenabove,in subjectshaving
a high diuresis,which in generalwould correspondto a high amount of
water availableto be excretedfrom the body, the urine is usuallyalkaline,
the alkalinitydue to ammonia.On the oppositeside of the normal, there
are subjectswith a very reducedurinary cxcretion.In patientseliminaring
only two to three hundredcc. in 24 hours,the amount of uric acid in the
urine is manifestlyincreased.Upon standing,theseurines alwaysshow a
reddishdepositformed mostly by uric acid. Relatingthis to comparative
physiology,while the normal subjectswould appear ureoselic,those with
polyuria can be consideredas ammonioselicand those with oliguria,
uricoselic.
We tried to see if a changcin the form under which the nitrogen is
eliminatedcan be inducedby changingthe amount of water availableto
be excreted.Normal subjectswhoseurineswere testedfor certain periods
of time for their contentof ammonia,urea and uric acid,were given I to 2
litcrs of water to drink. The highly diluted urine which was subsequently
. h e t o t a l a m o u n to f a m m o n i ai n c r e a s e dw h i l e
e x c r e t e db, e c a m ea l k a l i n e T
urea and uric acid were slightlyreduced.The same subjectswere later
given a dry diet for 12 hours or more. The specificgravity of the urine in
m o s to f t h e s ec a s e sw a s a b o v e1 . 0 2 6 ,i n d i c a t i n ga k i d n e yw i t h n o r m a l c o n d anifestly
c e n t r a t i o cn a p a c i t yA. l t h o u g ht h e c o n t e n it n a m m o n i ad e c r e a s em
in theseurines,the increasein uric acid excretedwas minimal. We sub-

NorEs

59'7

mitted the samesubjectsto a diet with fluidsreducedto a minimum, for

3 to 4 days. Under theseconditions,the excretionof uric acid startedto
increase.
Greatintakeof waterwould thustransforma normal ureoselicindividualinto an ammonioselic
in only a few minutes.A similarpassage
into
uricoselicwas seento need days, and even then it showedonly minimal
changes.This can be explaincdby the fact that while ammonia in urine
apparslargelythroughchangestakingplacein the kidneycellsthemselves,
uric acid resultsfrom more profound mctabolicchanges,which concern
especiallythe nitrogenousbases,particularlythe purines.
Chapter 5, Note l. Second Day Wound Crust pH
The metabolicprocesses
characterizingabnormalfoci have beeninvestigated in various *iys. One method was devisedin order to study the acidbase changeswhich take place within abnormal foci createdby surgically
producedwoundsunder the influenceof variouschemical,physicaland biologicalfactors.This rcscarchwas made in collaborationwith C. HuescaMejia. (212)
Adult Carworth Farm female and male albino rats weighing 150 to
200 grams were employed.The animalswere separatedaccordingto sex
and all receiveda standarddiet of Purina Chow and water ad lib. Tests
were run on groups of twenty animals,with from two to four control animals in each group.
The animals were divided into groups that were subjectedto various
experimentalconditionsfor three days,after which time, under ether anesthesia,a wide area of the back of each animal was carefully depilatedby
hand.A I squarecm. wound was then producedin the depilatedareadown
to the dorsalaponeurosis.
The wound was kept free of blood with dry gauze
until bleedinghad entirelyceased,and was then left uncovered.All wounds
w e r em a d eb e t w e e n8 : 0 0 a . m . a n d l 0 : 0 0 a . m .
A glasselectrodemade accordingto the specifications
of Mclnnes and
Dole (223) was used.As reference,an electrodecontainingnormal saline
was employed,(224) The electrodeswere mounted on a stand in a fixed
position.A model H Beckman pH meter, whosesignal was amplifiedby a
6H6 and 6SN7 push-pull,was usedas the first amplifier.The readingswere
made on a 200 micro-ampermeteradjustedso as to have the full scale
representone pH unit. The experimentalerror using this apparatuswas
found to be lessthan + .01.
pH determinations
were carried out by bringingthe wound area into
contactwith the tips of the electrodes.
The animalswere held gently but
frrmly in one hand until they were completelyimmobile,at which time a
firm contactwas establishedbetweenthe center of the wound area and
the tip of the glasselectrode,and betweenthe referenceelectrodeand the
wound periphery.Readingswere carried out on the surfaceof the freshly
exposedaponeurosiswithin ten to fifteen minutes after the surgical proreadingswere madeeverytwenty-fourhours upon the
cedure,Subsequent
wound crust, which was washedand then moistenedwith a drop of an

598

.,i

pHysropATHoLocy

RESEARcH rN

0.1% sodium ch.loridesolution freshly adjustedto pH 5 as suggestedby

Blank for use in skin pH determinations.(225) The electrodeswere
standardizedwith buffers of difterent pH before and during each period
of testing.
The effect of variouschemicalagentsupon the wound pH was studied
by daily oral administrationfor three days precedingthe operation and
thereafteruntil the conclusionof the seriesof pH determinations.
In general,
the substanceswere administeredaccordingto weight in quantitiescorresponding to the therapeuticdoses acceptedfor humans or in amounts
equal to 10Vo of the daily toxic dose.Water solublesubstances
were administeredin drinking water. Oil solublesubstanceswere administeredon
piecesof bread. In some cases,substances
were introduceddirectly into
the stomachthrough a catheter.In a few cases,substances
were injected
subcutaneously,
In the 164 controlanimals,the pH of the wound surfacevariedbetrveen
7.30 and 7.33 when measuredten to fifteenminutesafter the completion
of the surgicalprocedure.The pH remainedat this level for at least three
hours.
Twenty-four hours after the wounds were produced,the pH of the
m o i s tc r u s tc o v e r i n gt h e a r e aw a s f o u n d t o b e b e t w e e n7 . 7 2 a n d 7 . 7 6 i n
a l l u n t r e a t e da n i m a l s(. F i S .2 3 4 )
/86
f84
782

7&
778
+ -

776
o

f74
.

- .
- a

- F a - a
- a

- - a
- a - a a a - - . -

- . - . - - - - - - -

7to
78
76
7e4
762

F r c . 7 3 4 . S e c o n dd a y w o u n d c r u s t p H v a l u e sr e m a i n i n a r a n g e f . r o m 7 . 7 2 a n d 7 . 7 6 i n
n o r m a l r a t s a s s e e n i n t h e a n i m a l s w h i c h s e r v e da s c o n t r o l s f o r t h e m u l t i p l e e x p e r i .
ments.

At forty-eighthours,the pH of the crustwas found to be betwcen7.42

and 7.60, this being the period of greatestvariability.At seventy-twoand
ninety-sixhours, all the readingswere between7.28 and 7.32. After the
fifth day, similarvalueswere found and further readingswere not taken.
No consistentcorrelation was found between the values on different
days for individualanimals.For example,in some animalswith a wound
pH of 7.30-the lowest value-immediately after the operation. pH
reached7.76, the highestvaluefor the normal range,in twenty-fourhours.
In males,valuesappearedto be slightlyhigherthan in females.

NorEs

599

I n 4 1 0 o f t h e 8 6 0 a n i m a l se x p o s e dt o v a r i o u sc h e m i c a l sp H v a l u eo f
the woundswas determinedseveralminutesafter the operationand found
to be between7.28and 7.35. only 22 showeda deviationfrom the control
range of 7.30 to 7.33. No correlationwas observedbetweenthe minimal
changesin theseanimalsand the type of treatmentadministered.
while the pH values found several minutes after wound induction
showedvery little or no variation from the control rangeand no correlation
with the variousexperimentalconditionsemployed,the findingsat twentyfour hours showedconsiderablesignificancefor the agentsused. At fortyeight hours and thereafter,no important differenceswere observedbetween
valuesfor controlsand thosefor groupstrcatedin differentways. In continuing thesestudiestherefore,determinationswere carried out only several
minutesafter the productionof the experimentalwounds and then again
24 hours later. Actually, only valuesfor the twenty-fourhour readingappearedsignificantand will be discussed.
we made 24 hour measurements
f o r a l l o f t h e 8 6 0 a n i m a l st r c a t e dw i t h d i f l e r e n ta g c n t s F
. or convenience,
we refer to the pH value of the crust at twenty-fourhours after creation
of the cxperimentalwound as the s.d.c.pH (secondday crust pH).
From four to twelve animalswere employedin the assaysof the activity
of each agent.By applyingtie sameexperimental
conditionsto animalsin
groups testedat different times, it was possibleto determinewhether the
changesobservedwere due to some externalfactor such as temperature,
humidity, etc., or were actuallydue to the imposedexperimentalconditions. The s.d.c. pH has proven to be of considerableinterest because
consistentlysimilar changeshave beenfound to be producedby the same
agentswhcn appliedto animalsin differentgroupstcstedweeksor months
apart.
Consideringall the animalstrcatedwith variousagents,three possibilities have been found to exist: I ) There may be no effect upon the s.d.c.
pH, in which casethe valueswill all fall within the control rangeof 7.72
a n d 7 . 7 6 f o u n d i n u n t r e a t e da n i m a l s ;2 ) s . d . c .p H m a y b e e l e v a t e dt o
v a l u e sb e t w e e n7 , 1 7 t o 7 . 8 5 ;o r 3 ) t h e s . d . c .p H m a y b e r e d u c e dt o v a l u e s
of 7.70 to 7.60.
We will presenthere only the conclusionsof thesestudiesas related
to the variousagentsinvestigated.
CHEMICAL

FACTORS

Cationsand Anions
It was interestingto investigatethe influenceexertcdby some cations
and anions by first using the same anion with various cations and then
usingdifferentanionswith the samecation.It was apparentin all the experimentsthat the immediatepH of the wound doesnot differ from that of
the untreatcdanimals,and that the s.d.c.pH data obtainedare concordant.
We studiedthe influenceexertedby anionsfirst by investigatingthc
effectsof admhistrationof acids.With even strong inorganicacids, no

600

R E S E , ^ R c Ht N p H y s r o p A T H o L o G v

changeswere seenin the normal tissues.Definite and opposite effects u'ere

thus obtainedwith acids correspondingto the elementsof the VIlth series
and with thoseof the VIth series.The first group of acids,especiallyHCl,
induced a frank acidifying eftect while the secondproduced an alkalizing
eftect,The phosphateand nitric ions showedstrong acidification,the iodide
and bromide ions were weaker than the chloride ion. The bicarbonate ion
clearly alkalized, as did sulfate and selenate.The thiosulfate ion had an
obvious alkalizing effect, with pH valuesas high as 7.85. Among the organic acids,citric acid producedone of the strongesteffects,even stronger
than that of the inorganic acids. The fact that citric acid is only slightJy
metabolizedcould explain its strength.The gluconateion induced a slight
acidification.The cacodylateion seemedto induceno changesalthough not
enoughanimals were utilizcd to judgc its effect thoroughly.
In studyingthe effectsof salts,the roles of both cation and anion were
considered.Using different anions for the same cation, it was possible to
judge the effect of the cation.
Sodium and lithium produced relativelyslight acidification.Potassium
manifestlyacidifiedas did ammonium,the latter, however,to a lessmarked
extent.Marked acidifyingeffectswere seenfor iron, mercury and bismuth.
A lessereffectwas obtainedwith molybdenumand aluminum.On the other
hand, a manifest alkalization was found for bivalent calcium, strontium.
@pper, barium and cobalt. Manganeseand silver cations seemedto influencethe secondday wound pH only slightly toward alkalization.It
se'emsthat there is an additiveeffect for the differentelementsin their
acidifying or alkalizing influence.This has permittcd us to judge the
eftectsof the different ions. Potassiuminduces greater acidification than
sodium,and still greateracidificationthan ammonium.Potassiumchloride.
in which the two ions have an additiveeffect,is thus more frankly acidifl'ing. The same is true for the acid phosphate.The alkalizingtendencyof
sulfateion opposesthe acidifyingeffectof potassiumand explainsthe slight
acidifyinginfluenceof potassiumsulfate.We must arrive at the relatively
strong alkalizingtendencyof the carbonateion to find an anion able to
counteractthe acidifyingeftectof potassium.The s.d.c.pH effectof potassium carbonateis in the normal range.The data obtained through this
study led to the researchon the interventionof the elementsin biology.
which is the subjectof Chapter5. Informationconcerningthe effect upon
the s.d.c. pH of theseelementsas well as the relationshipbetween this
effectand the structureof the elementsis discussed
in this chapter.
Calcium ion has an alkalizinginfluencestrong enough to counteract
the acidifyingtendencies
of suchanionsas chlorideand phosphate.Weaker
acidifyinganions,such as lactateand gluconate,are not sufficientlystrong
t o c o u n t e r a ctth c a l k a l i z i n gt c n d e n c yo f c a l c i u m a
, n d c a l c i u ms a l t so f t h e s e
acidshave a strongalkalizingeffect.
This analysisindicatesthat the effectof a salt upon the local pH of
abnormaltissuescan be judgedby considering
the additiveinfluenceexerted
by anion and cation. the effect increasingif both have tendenciesin the
samedirection.and decreasing
are opposed,We will not
if the tendencies

NorEs

601

emphasizehere the other antagonisticbiological eftects of cations and

anions,as they pass from acidifying (citric acid and potassium) to alkalizing (calciumand thiosulfate).Wewill discusstheseeffectsin connection
with the pharmacologicalstudiesof these agents.For the moment, we
want only to emphasizethe valueof this investigational
method.
Acid Lipoids
The next step in the useof the s.d.c.pH was the study of the eftectsof
a specialgroup of acidsin which we were interested,the fatty acids.Analysis of the results obtained clearly shows the importance of the nonpolar
group. A carboxyl, when bound to a long chain as in the fatty acids,does
not by itself seem able to induce a changein the secondday wound crust
pH, the valuesremainingin the normal range.The substanceappearsinactive when the nonpolar group does not have its own energeticcenter or
formation.The saturatedfatty acidswith 10-16 carbonsdo not influence
the s.d.c.pH. The presenceof doublebondsin the nonpolargroup changes
the influenceexerted. All the nonsaturatedfatty acids studied show an
alkalizingeffect,with relativelyslight differencesfor the higher desaturated
memberssuch as linolenicor arachidonicfatty acids,or for the fatty acid
mixture obtainedfrom cod liver oil. However,the conjugatedfatty acids
acid, or the mixture of acidsstartingfrom cod
studied,such as eleostearic
liver oil, showedthe highestvaluesin this group, even for so small an
a m o u n ta s 5 m g r . p e r a n i m a lp c r d a y . T h e s ed a t ai n d i c a t et h e r o l e o f e n e r getic formationsin the nonpolargroupsof fatty acids.
We must emphasizethe differencebetweenthe hydrosolubleorganic
acids and the group of fatty acids mentioned above. In the former, the
action seemsdue to the interventionof the carboxyl, probably explaining
the strongactivityof tricarboxyliccitric acid. With an importantnonpolar
group, this carboxyl seemsto be unable to carry the molecule,and therefore cannot act. The intervention of a double bond could serve in two
ways: I ) by bendingthe molecule,thus increasingits mobility as it also
reducesthe melting point; 2) as an energeticcenter where reactionstake
place.The fact that the 10 carbonlauric acid is inactivewould indicatethe
slight influenceto be expectedfrom the bendingof the moleculealone, as
in oleic acid. The influenceexertedby metabolic changesin which fatty
acidsintervenethrough the energeticcentersin the nonpolar goup explains
the fact that theyhave a local alkalizingeffectinsteadof the acidifyingeffect
o f m a n y o t h e ro r g a n i ca c i d s .( F i g . 2 - 1 5 )
Alcohols
The role of the relationshipbetweenpolar and nonpolar groups appearedvery clear in the study of the seriesof aliphatic alcohols.
We have considercdseparatelya group of agentsextensivelyused in
that have a radical with a
our research.These are organic substances
bivalent sulfur as the polar group. We have mentionedabove that an
inorganic substancewith similar constitution,sodium thiosulfate,has a
strongalkalizingeffectupon the s.d.c.pH. Such an effectupon the s.d.c.

602

nESEARcHrN pHysropATHoLocy

pH has beenobtainedwith all preparations

containinga polar thiol group
when administered
orally or parenterally.We must emphasizethe unusual
uniformity of results,rarely encounteredin other biologicalexperiments,
It appearsthat the alkalizingeflect of thesepreparationsis sufficientto
overridethe individual differencesin subjectsreceivingthem. The same
arJkalizing
cffecthas beenapparentfor sodiumthiosulfate,and seemsto be
a common characteristic
for substances
having a bivalent sulfur in their
polar group.
These alcohols exert no influenceupon the pH of normal tissues,as
seen in measurementstaken immediatelyafter the skin is cut. The first
membersof the series,the methyl to propyl alcohols,do not influencethe

Exlrelad
Lr9@t

786
784
7A2
780
778
775
774
7f?
7 n

- , ^ - _
,o.C .."+il ..*
.fr' urt'itf ,p".

r.

- - -i - l - -J
i

+
.

L.

Lbtotrs,ld
L,pc6

\
*
.d/
F

s{tuat.d
L,@d!

.*." n"*s,
"s

:l
1

,+
!

"C""a I

=-

--

l l
-l-l -1- ---.- l - i rr--.1
F

rtO

na""o
alo

--

c.

_I_.

'

.t --la
r

?''d

768
766
761
762
760

F r c . 2 3 5 . T h e a d m i n i s t r a t i o no f v a r i o u s l i p o a c i d s u p o n t h e s . d . c . p H s h o w s t h a t
while the saturated fatty acids do not influence it, the non-saturatedfatty acids as
w e l l a s t h e l i p o a c i d p r e p a r a t i o n so b t a i n e df r o m d i f t e r e n ts o u r c e si n d u c e a n e l e v a t i o n
of the local pH.

s.d.c.pH. From butyl alcoholto nonyl alcohol,the membersof the serics

show a consistentacidifyinginfluence.However.a very important observation was made when the four isomersof butyl alcohol were studied.
'l-hree
showedthe acidifyingeffectwhile one, the tertiary isorner.like rhc
lower alcohols,did not influencethe secondday wound crust pH. This
could be correlatedto a specialcharacteristicof these substances.
their
relative solubility in water and in neutral solvents.Just as do the lower
alcohols,tertiarybutyl alcoholmixeswith water and neutralsolvents,while
the other three butyl alcohols,like the higher membersof the series,arc
n r o r es o l u b l ci n n c u t r a ls o l v e n t st h a n i n w a t e r .( F i s . 2 3 6 ) T h i s c h a r a c t e r the polar-nonpolarsubstances,
istic, which was used in systematizing
appearsto determinethe activityof the aliphaticalcoholsupon the s.d.c.pH.
We must again emphasizethat this activity is not direct but influencescersince the active alcohol induceslocal acidificatain metabolicprocesses,
tion rathcr than the alkalizationto be expectedwith a direct effect.

NorEs

60-3

Another factor which seemsto influencethe activity of this alcohol

seriesis the lengthof the carbon chain.Whilc heptanolinducesthe characteristicacidification,octyl alcoholdoes so in only some animals.Nonyl
and decylalcoholsappcarinactive.
The possibilityof a biologicalcompetitionbetweenthese agentsand
fatty acids has led to researchwith other alcoholscapableof combining
with fatty acids, particularlyin vivo. We studiedglycerol,glycerophosphoric ion and sterols,which are frequentlyfound combinedwith fatty
acids.We addedglucoseto this group becauseof its metabolicrelationship
to glycerolderivatives,althoughit is apparentlynot relatedto fatty acids.
It is intcrestingto note that glycerolproducedonly minimal influence
upon the immediatepH. The highestvalueswere still in the normal range.
The acidifyingeffectof g.lycerolupon the s.d.c.pH, secmedro be subject
786
784

Alcohols

78?
7n
7/8
716
774
77?
774
/a

.tr'

et'

l
(t

+_
tlt
rur

{_
tt
I

cf
((I

;-

''fl

-----|-

76

a
I

764

nn

Ju

"*'*
-;

c q.'c

-j

--

--T-

Co^t6t

I
I

a
a
a

IF

7&

F t c . 2 3 6 . S e c o n dd a y w o u n d c r u s t p H v a l u e sf o r v a r i o u s a l c o h o l ss h o w s t h e r e l a t i o n s h i p b e t w e e na c t i v i t y a n d l i p o i d i c p r o p e r t y .A l l t h e a l c o h o l s b e l o u , b u t a n o l a n d
t e r t . b u t a n o l .w h i c h a r e n o t l i p o i d s ,a r e i n a c t i v eT. h i s a l s o h o l d s t r u e f o r a l c o h o l sw i t h
c h a i n sl o n g e rt h a n e i g h tc a r b o n s .

to individualvariationsbut also with valuesat the lower limit of normal,

such as 7.72. Glucoseshoweda slight acidifyingeffect,whilc cholestcrol
showeddefiniteacidifyingactivityupon the s.d.c.pH, as did all the preparations of the insaponifiablefraction of various organs or tissues.The
fraction preparationsis more
acidificationproducedby the insaponifiable
intensethan for cholesterolalone,indicatingthat the other constituentsof
thesepreparationsalso have an acidifyinginfluence.
Other Agents
We investigatedsome of the hormonesthat are often used clinically
and found that the biologicalantagonismbetweenmale and female sex
hormonesis also apparentin their influenceupon the s.d.c.pH, the male
hormone having an alkalizingeffect,the female hormone, an acidifying
one. We must note herethat the acidifyingeffectis also apparentfor pro-

604

R E s E A R c Hr N p H y s r o p A T H o L o c y

gesterol.This eftect is opposite to that of the male hormone and similar to

that of stilbestrol,contrary to what we expected.There is sirnilarantagonism betweendesoxycorticosterol
and suprarenin,the former being acidifying and the latter slightly alkalizing. The liver antianemic extract also
has an alkdizing effect.
Severalvitamils also were investigated.We were able to seethat while
vitamins Br, Br, E and K have an acidifyingeffect,vitamins A, D and Bu
have an alkalizing action. Ascorbic acid seemsto have no effect upon the
s.d.c.pH.
A few alkaloids and glucosides,important for their pharmacodynamis
activity, were studied. The two opium alkaloids and a similar synthetic
agent are moderately acidifying. Atropine, caffeine, and quinine have a
slight alkalizingeftect.The difterenteffectsof digrtalineand saponinewere
unexpected;both show slightbut opposedaction,the first acidifyingand the
secondalkalizing.
Becauseof their eftect upon the central nervoussystem,narcoticsand
hypnoticswere studied. While ether and chloroform are slight alkalizing,
the two barbiturateswhich we testedshowed an acidifying effect. This is
interesting,especiallywhen related to the opium alkaloids and demerol
which also induce acidification,althoughto a lesserextent than the barbiturates.
Various other agents were studied. Among pyretogenicsand antipyretics, there is an obvious antagonismin influence upon the s.d.c. pH.
While the pyretogenic,methyleneblue, inducesa frank acidification,the
three antipyreticswe examinedproducedalkalization.However, acetylsalicyclic acid does not follow this mle-it has an acidifying effect.
Among antimicrobial agents,a parallel action was apparent for the
three antibioticsof fungal origin and the two sulfa drugs studied.All have
an alkalizing effect, like the antipyretics.The acidifying effect noted for
benzedrinedoes not accord with a similar eftect observedfor substances
with hypnotic and sedativeactivity. This discordancebetween principal
pharmacologicalactivity and effect upon the s.d.c. pH indicates that the
latter must, on many occasions,be consideredto be due to a secondary'
influenceexertedat the interstitiallevel.
The antagonisticbiologicalactivity of anti-anemicliver extract and iron
preparation,the former favorably influencingthe hyperchromic anemias.
the latter favorably influencingthe hypochromicforms, is reflectedin their
oppositeeffect upon the s.d.c.pH. The anti-anemicliver extract induces
alkalization,while iron inducesacidification,The same antagonismis seen
with two agents having an opposite action on blood coagulation.Whilc
vitamin K induces acidification,dicumarol is alkalizing.Although rutin
actsupon other factorswhen it intervenesin bleeding.it has the sameeffect
as vitamin K on the s.d.c.pH.
Aminophyllin inducesacidificationin contrastto caffeine,which producesslightalkalization.On the other hand, procaine'seffectis like that of
This can be
all inducingacidification.
the opium alkaloidsand barbiturates,

xores

605

relatedto the effectof higher alcoholswhich alsohave narcoticactivityand,

as seenabove,also induce acidifcation.
TrsLn XXVII showsthe effectsupon the s.d.c.pH of all of the substancesexamined.
T,rsle XXVII
THr Errpcr

or V,urous Sunsr^NcEsUpoN rHe S.D.C. pH.

--=Elevate
Reduce
No Effect

Vitamins.

A, D, B.

B ' , B r ,K ' E

Hormones

Testosterone,
Epinephrine

Stilbestrol
Progesterone,
Desoxycorticosterone

Alkaloids . . .

Atropine,
Quinine

Codeine,
Morphine

Antibiotics . .

Penicillin,
Streptomycin,
Aureomycin

Sulfonamides. .

Sulfathiazole,
Sulfamerazine

Antipyretics and
analgesics

Acetophenetidin,
Aminopyrine
Antipyrine

Acetyl-salicylic
acid

Narcotics....

Chloroform.....

Phenobarbital,
Pentobarbital

Liverextract....
A n t i - a n e m i c s. .
X a n t h i n e s . . . . Caffeine
Miscellaneous. . Dicoumerol,
Benzene,
Toluol,
Saponin,
Pteryl-glutamic
acid Teropterin)

Iron (reduced)
Aminophylline
Procaine,
Benadryl,
Coramine,
Glucose,
Glycerine,
Sodium gluconate,
Rutin,
Methyleneblue,
Mercuhydrin,
Benzedrine,
Demerol

Basedupon the findings for such different groups of agents,the s.d.c.

pH method appearsto be an interestingtool for the study of pharrracoat
logical activity.We must emphasizethat not one of thesesubstances,
least in the dosagegiven, has had any influencein changing the pH of
normal tissuesfrom normal range, as indicatedby pH values obtained
immediatelyafter wound production.As our researchconcernsonly ab-

606

xESEARcH rN pHystopATHoLocy

7.76

7.74

772

7.68

7.6

7.64

7.62

7.60

7
Ooys

to

d a l k e r t u m o r o n t h c s . d . c .p H d c F r c . 2 3 8 . T h e c f f e c t o f a g r o w i n gt r a n s p l a n t e W
w
o
u
n
d
s
p
r
o
d
u
c
e
d
s
e
r
i
a
l
l
yo n t h e d a y o f t r a n s p l a n ta n d e v e r y
i
n
s
u
r
g
i
c
a
l
terminalions
t
h
e
s . d . c .p H w a s o b s e r v e di n a l l a n i A
m
a
r
k
e
d
r
e
d
u
c
t
i
o
n
i
n
t
h
e
r
e
a
f
t
e
r
.
three days
mals.

NorEs

607

normal tissues,the s.d.c. pH is of use. Howover, in integratingthe pH

valuesinto the generalpicture of the offbalances,it must not be forgotten
that they representchangesin the interstitialfluids, which correspondto
the secondarypart of the tissuelevel.
PHYSICAL AGENTS

Animals kept in an incubatorat 38o C. showeda definitetendencytoward

a loweringof the s.d.c.pH, The effectof cold upon animalsmaintainedin
a refrigeratorfor forty-eight hours before operationwas less apparent.

776

7.74

7.72

7.70

7.66

Doys
F r c . 2 3 9 . S e r i a l s . d . c . p H c h a n g e si n a n i m a l s w i t h t r a n s p l a n t e dt u m o r s t h a t r e g r e s s e dr a p i d l y o r f a i l e d t o g r o w , A s l i g h t f a l l i s s e e n t o o c c u r i n t h e f o u r t h a n d
seventh post-transplantday determinations,with a r e t u r n t o t h e c o n t r o l r a n g e i n a l l
the animals by the tenth day.
ffi

-?tLw#--t!lt?l!:aFl

I!+5Effry

608

xESEARCH rN

PHYSToPATHoLocY

BIOLOCICALFACTORS

TrarcplantedTumors.Preliminarystudiesregardingthe effectof a transwound have

plantedWalkertumor upon the s.d.c.pH of an experimental
the control s.d.c.pH was debeencarriedout. Prior to transplantation,
terminedfor eachanimal.Tumorswerethen transplanted
subcutaneously
left
to the
flank region by the trocar method.After the tumor had been
transplanted
to the animals,experimentaloperativewounds were produced at intervalsof three days and the serial s.d,c pH values on the

774
- ---

--

7.72

7.70

7.66

Ooys
F r o . 2 4 0 , T h e s . d . c . p H c h a n g c si n t w o r a t s w i t h g r o w i n g W a l k c r t u m o r s ( - ) .
from which the tumor w8s removed on tbe seventh port-transplant day. Followio3
tho rcmoval, tbc e.d.c. pH began to return towards the control r&n8e. A similar largc
inc.irion was madc in two animals without tumors and no effcct was observed upon
t h e s , d . cp. H i n t b e s ea n i m a l s( , . . . ) .

fourth,seventh,
tenthandthirteenth
post-transpr.",
O"r, ;;;:,J
;::
pH valucsdetermined
eachtime immediately
wound
proafterthe
was
duced,did not differ from the valuesfound in untreatedcontrols. As in all
other anim4ls,the s,d.c,pH only showedsignificantchanges.

775

7.74

7.73

7.7t

4
Doys

F r c . 2 4 1 . S e r i a l s . d . c .p H d e t e r m i n a t i o n isn r a t s w i t h o u t t r a n s p l a n t e dt u m o r s . T h e
scrial small surgical proceduresdid not alter the s.d.c.pH from the control range.

F i g u r e 2 3 8 i l l u s t r a t e sg r a p h i c a l l yt h e c h a n g e si n t h e s . d . c .p H i n a
group of twelve animals with successfultumor transplants.A steady and
maintainedlowering of the s.d.c. pH was observedin each animal, with a
tendencyfor it to rise slightly in thosc animalssurvivingthirteen days.
In some of our animalsthe tumor has shown a tendencyto regress
or fail to take. In six animalsin which the tumor failed to
spontaneously
the s,d.c,pH in the
take or underwentearly regressionand disappeared,

610

xESEARcH rN PHYSToPATHoLocY

fourth and seventh post-transplantday tests fell slightly, but thereafter returnedto control levels.(Fig. 239)
When transplantedtumors that were growing well, were surgically removed following the secondpost-transplantdeterminationon the 7th day,
the s.d.c.pH was seento increaseat the next test.(FiS.2a0)
Surgical wounds. The same incision as for the removal of the tumors
was producedin control animals,but the incision alone did not aftect the
s.d.c.pH. (Fis. 2a0)
In order to further control these experiments,repeateds.d.c. pH determinationswere carried out upon wounds produced serially as in the
tumor bearing animals.When no tumor was present the serial s.d.c. pH
valuesdid not vary from one examinationto the next. (Fis.241)
The serial small surgicalproceduresdid not alter the s.d.c.pH from the
control range.
Chapter 5, Note 2. Potassium
We have seen in Chapter 2, Note I how the place occupied in nature
sodium
and potassiumcan explaintheir peculiardistributionin the body.
by
Proper to the earth crust, one of the environmentsthrough which the
complexindividual has passedduring its phylogeneticevolution, potassium
part of the cellularcompartmentin
appearsas an elementof the secondary
heterotropicelement, potasAs
a
monovalent
the hierarchicorganization.
cation of this compartment.
sium representsthe principal organizational
Its influenceexertedin normal and abnormalphysiologycan be understood
throughits specificinterventionat the cellularlevel, the changesin potassium contentof the other compartmentsof the hierarchicorganizationbeing
sccondaryto thoseoccurringat the cellularlevel.
Potassium,absentat the nuclear compartment,is thus found in the
nuclearor chromosomalsap,only in minimal amounts.We have no direct
information about a passageof potassiumfrom cytoplasminto the nuclear
compartmentunder abnormalconditions.Judging by analogy,it appears
probablethat such a passagewould occur and result in the appearanceof
nuclearvacuoles.
Ample data are availableconcerningthe relationshipbetweenpotassium
of the cellular and of the metazoiccompartments.This information receives
a specialinterpretationwhen relatedto the above mentionedhierarchic
distributionof the elements.
The cells maintain a proPer amount of potassiumin the cytoplasm
which correspondsto a cellularprimary constant.This constantinsuresa
normal cellular metabolismand is controlledin part by the selectiveinterventionof the cellularmcmbranc.Under normal conditions,only a slow
passageof potassiumthrough this membranetakes place as compared to
such as of water, for example.On using radioactive
bther-constituents.
potassium,Moore has shown that it takes about frfteen hours to bring it
into balancewith the intracellularpotassium,while for heavy water, such
an equilibrium was reached in less than two hours. (254) Due to the

NorEs

6ll

interventionof the membrane,only minimal changesresult in the cellular

potassiumeven though rapid potassiumvariaLionstake place in the extracellularcompartment.Under normal circumstances,
the body is insured
through a regulatorymechanismagainsttoo strongor too rapid systemic
changes.Normally, no potassiumis storedin the body beyondthat which
is containedin the cell and metazoiccompartment,Followinga high intake,
potassiumis rapidlyexcreted.A very smallamountis lost throughperspiration, an additionalamount,of around ljva is lost in the stool (252), and
the remainderis lost throughthe kidney. (253) For insufficientintake,or
an abnormallossthroughexcessive
diuresis,prolongeddiarrheaor vomiting, the organismtries to reducethis loss of potassiumto a minimum. A
prolonged systemicpotassiumdefrciencybrings about an increasein the
rveightof the kidney with tubular hyperplasia,which can be interpretedas
a compensatoryhyperthrophy,in order to insure the reabsorptionof very
small amountsof potassiumfrom the glomerularfiltrate,and thus savethis
importantelementfor the organism.Consequently,
a quantitativeabnormal
intakeor lossof potassiumwill resultin an abnormalamount,eithertoo high
or too low in the metazoicfluids, only in caseof a concomitantdeficiency
of the regulatorysystem.Prolongedquantitativechangesin the amountof
potassiumavailableto the total body, influenceonly up to a certain point,
the potassiumcontent of the cells. Thus when the total body potassium
remainslow over a periodof time, the cellstoo lose potassium.(250) On
the other hand, the potassiumin the cells increasesafter an abundant,
p r o l o n g e da d m i n i s t r a t i oonf t h e c a t i o n .( 2 5 1 )
Besidesthesequantitativechangeswe have to considerabnormalconditions as affectingthe specificinterventionof this elcment, The proper
level to which potassiumbelongsand the characteristic
changeswhich take
placeat the cellularlevel,could be translatedin too high or too low values.
We tried to further interpret its interventionthrougb the heterotropiccharacter of this element.
The changesseen to occur in muscleshave permittedus to relate this
dual occurrenceobservedfor the potassiumcontentof the cells to the two
abnormalmetabolicconditionswhich take place in the cells.A musclein
anabolic metabolismand characterizedby a processof glycogenogenesis,
shows an increaseof its potassiumcontent. On the contrary, catabolic
processes
such as those occurringduring muscularexercise(255) or in
tissuesin agonicstates,are relatedto a loss of potassiumfrom the cells.
This metabolicloss of potassiumis differentfrom that seento result from
the death of the cells.
The destruction of cells in general, results in a liberation of their
potassiuminto the interstitialfluids.However,such a destructionexplains
only in part the progressiveloss of potassiumencounteredin abnormalconditions. The erythrocytesof stored blood lose their potassiumto plasma
The sameis true for patients
throughanotherprocessthan their destruction.
undergoingsurgery.They experiencea constantloss of cellular potassium
into the metazoicfluids,for which the processof destructionis only partially
The breakdownof cellsas it occursin starvationor dehydration
responsible.

612 /

n E S E A R c Hr N p H y s r o p A r H o L o c y

for instance,releasesa proportion of.2.4 gm. of potassiumfor every gram

of nitrogen which is liberatedunder theseconditions. (256) However, in
surgical patients,the potassiumloss was found to be two or three times
higherthan expected,consideringthe nitrogenloss. (257) This indicates
a passage
of potassiumout of the cellswithoutcellulardestruction.
The study of the red cell potassiumchangesin stored blood and in
the diphasicbiologicalphenomenonsuch as in hemo-shock,has increased
our knowledgeof the conditionswhich correspondto these changes.We
found thus that in the first phaseof hemo-shockthere are lower amounts
of potassiumin the red cells,followed by a secondphasewhere this cellular
potassiumcontent increases.These two changeswere thus connectedwith
the characteristicoffbalancesoccurring in hemo-shock:type D in the first
phase,and type A in the second.Further studiesof tissuesin offbalanceA
or D in pathologicalconditions have confirmed this correlation between
the two fundamentaloftbalancesand the two oppositevariationsin cellular
potassium.
The heterotropiccharacterof potassiumas seen through the test of
the secondday wound crust pH, (SeeNote I, Chapter 5,), has correlated
the changesin potassiumdistribution to the heterotropic or homotropic
characterof the occurring processes.
A low cellular content in potassium
would correspondthus to the homotropiccharacterof the processescharacterizingthe offbalanceD. Those correspondingto a high cellular conteDt
concord with the group of heterotropicprocesses.Increasedintracellular
potassiumresults in a heterotropicanabolic effect, while loss of cellular
potassiumcorrespondsto catabolicmetabolism.
The relationship between the hierarchic compartments explains the
changesunder abnormal conditions in the amouot of potassium present
at the different compartments.The study of the diphasic phenomenon of
hemo-shockhas permitted us to follow this relationshipbetween cellular
and plasmaticpotassium.In the first phaseof a hemo-shock,to the low
potassiumof the cells correspondsa hyperkalemia,while in the second
phase,to a higher cellularpotassiumcorrespondsa hypokalemia.
With potassium,a cellularelement,the changesseen in the metazoic
compartmentcan thus be consideredto be secondaryto those occurring
at the cellular level. An abnormal cellular condition, with an increase in
the cellularpotassiumcontcnt,would thushavean oppositeeftectupon the
amountof potassiumpresentin the metazoicfluids.The fact that plasma
potassiumvaluesare kept below normal, has to be interpretedas a means
the high valuespresentin the cells.Theselow values in
of compensating
plasmawould resultin a reductionof the cellularpotassiumand favor its
return to normal valueswhen possible.It is especiallythrough changes
in the urinary cxcretionthat the respcctivehypokalemiaare induced and
maintained.On the other hand, an abnormalchangein the cellular condition resultingin a low amountof potassiumin the cells is seento induce
a prolongedcompensatoryincreasein the potassiumcontentof the metazoic
fluids.Oliguriawith reducedlossof urinary potassium,permitsthe creation
and maintenanceof this secondaryhyperkalemia.

NOTBS

613

Potessium and Ofl bulance.r

We tried to utilize the informationconcerningthe relationshipbetween
the two types of offbalanccsand the changesin potassium distribution
betweencells and metazoicfluids, in order to recognizethe existenceand
nature of theseoffbalances.This led us to compare the amount of potassium presentin red cells and in serum as an indication of this distribution
betweencclls and metazoicfluids. From a practical point of view, we
utilized the total blood insteadof the separatedred cells. (Chapter 4, Note
5/ From the relationshipbetweenthe two values,we could interpretthe
nature of the occurringchangcsas corresponding
either to a quantitative
abnormality excessof deficiency---orto a qualitative abnormality due to
an offbalanceA or D. Fig. 127 showsthis correlation.With the values of
serumpotassiumaround 4.5 mEq and of total blood around 38 mEq, the
condition is considerednormal from the point of view of the potassium
intervention.Low valuesin serumand total blood correspondto a quantitative deficiency,while high valuesfor both, a quantitativeexcess.High
serum with low total blood potassiumcorrespondto the offbalancetype
D, while low serum potassiumand high values in total blood to the offbalance type A.
In chronic conditions, such oflbalancesare seen to persist over long
periodsof time. Fig.214 showsan exampleof a typical D offbalance,with
the serumpotassiumhigh and the totalblood potassiumlow. Fig. 215 shows
an offbalancetype A, with low serum potassiumand relatively high total
Fig.216 showsan exampleof lack of potassium,with low
blood potassium.
valuesin both serum and total blood.
Potassium and Lipids
The correlationbetweenpotassiumdistribution and the offbalancesA
and D, has linked this information to the lipids and lipoids. The administrationof agentsof one or the other of the two groups, positive or
negative,has produccdoppositechangesin thc distributionof potassium
total blood and serum. These experimcntswere made in rabbits in collaboration with Ismail Eroglu, Patricia Mclachlan and Lee Weston. Administeredin large amounts,all the positivelipoids and especiallyheptanol
were seenable to reducepotassiumin blood and increaseit in the red cells.
The negativelipoids have an opposite effect. Administered in reduced
amounts, big differencescould be sec between the agents of the same
g:oup, many having no influenceon the potassiumand only few showing
manifesteffects.Among the negativelipoids,thc most active agentshave
appearedto be heptyldiscnide,sulfur and seleniumtetra-hydronaphthalene
and epichlorohydrin.
Potassiumand Sodium
The changesin the amount of potassiumcould be connectedto its
in the frameof the hierarchic
to sodium,and furtherinterpreted
relationship
organization.Both are membersof the same seriesin the periodic chart,

614

. , ' R E , s E A R c Hr N p H y s r o p A T H o L o c y

but they correspondto differentcompartments

of the organization.Sodiunr
i s t h e c a t i o n o f t h e m e t a z o i cc o m p a r t m e n ta, n d p o t a s s i u mt h a t o f t h e
cellular.They would consequently
act differentlytowardscells.for instance.
U n d e ra b n o r m acl o n d i t i o n ss,o d i u mi s a b l et o e n t c rt h e c e l l s .F o r e x a m p l c .
following an injury, the cellular membranewhich is almost impermeable
to sodium,lets it passthrough.A penetrationof sodium into cells occurs.
as demonstrated
b y t h e u s e o f r a d i o a c t i v es o d i u m . ( 4 2 ) A s a c o n sequence,potassiumis releasedfrom the cells in order to maintain thcnecessary
osmoticpressureconstant,Many abnormalprocesses
occur foll o w i n g t h c p e n e t r a t i o no f t h e s o d i u mi n t h e c e l l s a n d o t h c r s o c c u r d u e
to the releaseof potassium.The sodiumwhich enteredin the cells is partly
isolatedtogetherwith water, to form cellular vacuolcs.The responseof
the metazoiccompartmentto thosechangesare interesting.Hyperkalemia.
with a simultaneous
hyponatremiaoccursin the first phaseof the diphasic
phenomenon.
The oppositehappensin the secondphase,when hypokalemia
coincideswith a hypernatruria,
This antagonismbetweensodiumand potassiumis seenfurther in the
pharmacologic
activityof theseelements.The administrationof potassium
salts inducesa greatereliminationof sodium and water, which explains
its diureticactionwith dehydrationof tissuesand progressive
alkalinization
of the urine. The effectupon the diflcrentorgansis also antagonistic.For
instance,in their effect upon the heart, as Merrill and co-workers have
shown, the characteristicclectrocardiographic
cffects of hyporkalcmia
a p p e a ro n l y w h e n t h e s o d i u ml e v e l o f b l o o d i s l o w e r e d .( 2 5 9) A h i g h
sodiumcontentpreventsthe cardiaceflectsof hyperkalemia.
I n o p p o s i t i o nt o t h c a t r o p h yo f t h e a d r e n a g
l l o m e r u l o s ian d u c e db y a n
p
o
t
a
s
s
i
u
mo v e r d o s ec a n c a u s e a n
e x c e s so f s o d i u m a d m i n i s t r a t i o na,
of thesezoncsin rat adrenals.
enlargement
The hyperkalemiadue to externalintakesuch as in potassiumpoisoning, or due to systemicresponseto low cellular potassium,specificallyinfluencescertain functions.The peripheralvascularcollapsewith lorvered
and mentalconfusion,
cold clammyskin,pallor,listlessness
blood pressure,
are symptomsrelatedto thc offbalanceD, and encounteredin high plasma
potassium.They appear in conditionssuch as shock and burns rvhich
correspondto a prolongedfirst phaseof the diphasicphenomenon.Parest h e s i sa n d f l a c c i dp a r a l y s i s( 2 6 0 ) ( 2 6 1 ) a r e a l s o i m p o r t a n t r e s u l t so f
hyperkalemia.The most importantchangcsappearin cardiac physiology
and can be interpretedto correspondto low cellular automatism.Hypetkalemiawill thus inducedromotropicpositivechanges,such as the increase
of the duration of the Q R S complex,or that of the P-R interval, with
a delay in the ventricularcontraction,or a block rvhich can lead to thc'
llp
y p e a r st h c
a r r e s to f t h c h c a r ti n d i a s t o l c (. 2 6 2 ) , ( 2 6 3) C h a r a c t e r i s t i c a a
T
h
e
c
h
a n g es e e n
e
v
e
n
a
n
g
u
l
a
r
.
w
h
i
c
h
b
e
c
a
m
e
w
a
v
e
i
n
c
r
e
a
s
e
d
T.
c h a n g e isn
in offbalanceD. are similarto thoseinducedby a prolongedadministration
activity of various agents
of potassium.ln a study of the pharmacological
with I. Eroglu.we usedthesechang.-s
upon the heart,madein collaboration
as an indicationof the type of offbalancethey induce'

NorEs

615

Fig. 2168 shows some examplesof this effecton rabbits,One group

comprisesagentsconsidered
as ableto inducean offbalanceD. They induce
changesin the T wave which high amplitudeis characteristic
for a hyperkalemia.An oppositecffectis seenin thc othergroup,formedby agentsable
to induce an offbalanceA. and where the T waves are remarkedlvdep r e s s c d(. S e ep a g e5 7 4 )
Po Iassi ttrtt and T her ap.v
A l l t h c s ec o n s i d e r a t i o nhsa v e l e d t o a m o r e p r e c i s eu s e i n t h e r a p yo f
t h e i n f o r m a t i o nf u r n i s h e db y t h e s t u d y o f p o t a s s i u md i s t r i b u t i o n T
. he
quantitativedeficiency-recognizedby low potassiumin total blood and
5s1urn-15controlledby oral and parenteraladministrationof potassium.
A q u a n t i t a t i v ee x c e s s - w i t h h i g h v a l u c s i n s c r u m a n d t o t a l b l o o d - i s
t r e a t e dw i t h t h e a d m i n i s t r a t i oonf i o n i ce x c h a n gceo m p o u n d sd, i e t ,l a x a t i v e s
a n d m o r c s o d i u mi n t a k c .F o r t h c o f f b a l a n c c st h, c t r e a t m e n ti s c s p e c i a l l y
directedto thoseagentswhich seemto influencemore stronglythe cellular
p o t a s s i u mm c t a b o l i s mS
. c l e n i u ml i p o i d s a n d s u l f u r i z e dt c t r a - h y d r o n a p h thalencare usedfor offbalanceA, irnd hcptanolfor offbalanceD.
T h c f a c t t h a t t h e c e l l u l a rm c m b r a n ep l a y s a n i m p o r t a n tr o l e i n t h e
metabolismthrough which the abnormalchangesin potassiumoccur, has
l e d u s t o u s e a g c n t sa c t i n gu p o n t h e s em e m b r a n e sA. d r e n a l i n eh a s b e e n
s e e nt o f a v o rt h e d i s c h a r goef p o t a s s i u m
f r o m t h e l i v e rc e l l s .s i m u l t a n e o u s l y
rvith glycogen.We thus usedadrenalinefor casesof offbalanceA in which
w a sp r e s e n tO
. n t h e o p p o s i t es i d e ,t h e
a n e s p e c i a l lhyi g hc e l l u l a rp o t a s s i u m
r i t h a n i n c r e a s e idn t a k eo f g l u c o s ew a s
a d m i n i s t r a t i oonf i n s u l i nt o g e t h e w
s e e nt o i n c r e a s et h e c e l l u l a rp o t a s s i u m(.2 - 5 8 ) A s i m i l a re f f e c tw a s s c e n
for cortisonc,and ACTH, for heptanol,cholesteroland also for the unfractionsof organs.Theseagentshave been usedin the treatsaponifiable
ment of offbalanceD at the cellularIevel.where abnormalhish valuesof
arepresent.
s e r u mp o t a s s i u m
Chapter 6, Note l. I)efinition of the Lipids
Accordingto Bloor: "Lipids may be definedas a group of naturally
consistingof the higher fatty acids, their naturally
occurring substances
found naturallyin chemicalassociaoccurringcompoundsand substances
in generalby insolubilityin
tion with them. The group is characterized
"
f
a
t
w a t e r a n d s o l u b i l i t yi n
s o l v e n t s , "e . g . , c t h e r , c h l o r o f o r m ,b e n z e n e ,
etc." (226\
Chapter 6, Note 2. Deffnition of Lipoids (221)
analysisof our definitiono f l i p o i d s ,J .
From a physicomathernatical
M a r i a n i a r r i v e st o t h e f o l l o w i n gc o n c l u s i o n s :
Any substancewill behavelike a lipoid with respectto a polar solvent
w h e ni n :
l . ,
l ; o l ,
at:*
?: z,,,ir.fit-2a
2"u,?".."'

616

REsEARcH rN pHysropATHoLocy

g will be positive.The electricalattractionsgiven by:

2ana.zz
will not be able to balancethe attractiveforces:
I

, ,
)air *

1 , )
?2ai,

due to van der Waals forces (and to the polar groups of A). But we can
simplify this definition.Let us considera solventthe moleculesof which are
dipolar and form polar groupssimilar to thosecontainedin the solute. We
can call those substancesIipoids for which the dipole interaction energy in
the cavity occupiedby the moleculeis lessthan the energyarisingfrom van
der Waals forces.
We get:

* - I,,,,',^""
(o"*,) -' (o"*' * o ) = ) " - e
where Dow o representsvan der Waals forces in a cavity occupied by the
molecule and f the dipolar forces at the surface of the cavity.
Chapter 6, Note 3. Separating Membrane Between Aqueous Media
The possibility of having a membrane with polar groups at both surfaces, as shown in mitochondria, representsone of the most important
meansused in biology to separatetwo aqueoussolutions. (228) One of
the characteristicsof such a membraneis that often it representsan isolating boundary rather than a membranewith functional activity. This is
seen in the fact that very often such a membranebreaks when the two
separatedaqueousmedia have to mix.
Chapter 6, Note 4. Fatry Acids Break Down
In a work hypothesisof the biological breakdown of a long chain
moleculefor caloric metabolism,we considertwo factors as being of capital
importance: 1 ) that the result be an even carbon-numberchain molecule.
since the Knoop oxidation leads to complete caloric utilization only for
suchmolecules,and 2) that the lengthof the chain of carbonsbound to the
carboxyl in the new moleculebe not higher than l1 carbons in order to
permit direct beta oxidation. To fulfill the first condition, the breakdown
processdoes not occur between the carbons of the double bonds themselves,since in natural fatty acids the double bonds separateportions of
the chains,usually with odd number of carbons.Such a brcakdown would
lead to moleculesresultingin further incompleteKnoop oxidation.
According to the hypothesisthe breakdownof the molecule occurs at
the carbon ncarest the double bond. Through the energetic influence
exerted by the double bond, the even numbered carbon nearby appears
strongly positive. As a first step, this carbon was seen to fix a molecule of

NorEs

6l'1

oxygenresultingin a hydroperoxideas shown by Farmer and co-workers

first for rubber (30) and later for fats in vitro. (31) It is with the passage,
in a second step, of this hydroperoxidegroup into a carboxyl that the
breakdownof the molcculeoccursat this level, as shown in Fisure 242.

, 7
.

(a)

(b)

\/v A/
\

OoH

(c)

F t o . 2 1 7 . ' f h e o x i d a t i v eb r e a k i n gd o w n o f a f a t t y ' a c i dm o l e c u i e( a ) o c c u r si n v i v o
t a k i n g p l a c et h r o u g h t h e a p p e a r a n c eo f a h y d r o p e r o x i d ea t t h e c a r b o n a d j a c e n tt o t h e
d o u b l e b o n d ( b ) . I t l e a d s u l t i m a t e l yt o a c a r b o x y l f o r m a t i o n ( c ) a t t h i s a d j a c e n t
c a r b o n a o d r e s u l t st h u s i n c h a i n sw i t h e v e n n u m b e r o f c a r b o n s .

If the even numberedcarbon near the double bond is toward the tcrminal methyl group, a monocarboxylicacid will result. A similar process
taking place at the other carbon adjacentto the double bond toward the
carboxyl will lead to a dicarboxylic molecule.The metabolic changesin
vitro-and also i:r vivo-have shown the appearanceof thesetwo groups
of evencarbonmono and dibasicfatty acids.By binding two moleculcsof
water the remaining 2-carbon chain linked by the double bond would re. u c hc h a n g e so c c u r r i n gi n t h e c a l o r i cm o n o s u l t i n a c e t i ca c i d m o l e c u l eS
ethenoidspermitted us to explain one of the baffiing peculiaritiesseen in
the constitutionof the monoethenicfatty acids.
In Note 5, we discussthe positionof the doublebond in the principal
naturallyoccurringmonoethenoids
as it followsa charactcristic
pattern.In
moleculeswith l6 or lesscarbonsthe doublebond is more often placedso
as to separatea group with 9 carbonstoward thc carboxyl end, while in
moleculesof 18 carbonsor more, the doublebond separatesalmost constantlya group of 9 carbonstowardthe methylend. Figure 243 showstwo
characteristicexamples.
t*t
*ooa
t*t
**",
e m p l a c e m c n t sa r c s e e n f o r t h e d o u b l e b o n d i n t w o m o n o Oro. ,or.T*ic
ethenoids.The double bond separatestwo groups with odd number of carbons. For
t h e m y r i s t o l e i ca c i d , i t s e p a r a t e sa g r o u p o f 9 c a r b o n st o l v a r d t h e c a r b o x y l i ce n d . a n d
a g r o u p w i t h a s h o r t 5 c a r b o n c h a i n t o w a r d t h e m e t h y l e n d . F o r h e x a c o s e n o i ca c i d
( 2 6 C ) a c h a i n w i t h l 7 c a r b o n s i s s e p a r a t e dt o w a r d t h e c a r b o x y l e n d a n d o n e o f 9
carbons loward the methvl end.

618

nesEARcH tN pHYsropATHoLocy

The breakdown of the molecule accordingto the processnrentioned

a b o v ee x p l a i n st h c p e c u l i a r i t yF. o r t h e c h a i nw i t h l 8 o r l e s sc a r b o n s ,b i o logicalfissionwould resultin a diacid with 8 carbonsand another monoacid with 8 or lesscarbons,both subjectto Knoop oxidation. In the long
chain fatty acid the double bond separatinga 9 carbon fraction toward the
methyl-endof the moleculewill result in an 8-carbonchain as monoacid
havingthe methyl group at the other end. The other part of the molecule,
with more than 8 carbonsand which corresponds
to the long fraction hav'^t
HOOC

,"*AA,/\,/*
i **/y'*'

F t c . 2 4 4 . T h r o u g h t h e e m p l a c e m e n to f t h e d o u b l e b o n d i n t h e m o l e c u l e s o f t h e
m o n o e t h e n o i d st,h e b r e a k i n g d o w n o f t h e m o l e c u l e so f f a t t y a c i d s a t t h e c a r b o n s
adjacent to the double bond leads to the sppearanceof a dicarboxylic acid for the
part of the chain having more than l0 carbons.

i n g t h e c a r b o x y la t i t s e n d , r e s u l t si n a d i a c i d . ( F i 1 . 2 4 4 ) T h i s a l s o w i l l
be subjectto Knoop oxidation,even with a long chain. In the diacid, this
process,which is relatedto the interventionof the carboxyl, can take place
at both endswherecarboxylsare present.
Chapter 6, Note 5. Position of the Double Bond in lUonoethenoids
The comparativeanalysisof the principal naturally occurring monoethenicfatty acids has shown a curiousconfigurationdue to the peculiar
r e l a t i v ep o s i t i o no f t h e d o u b l eb o n d i n t h e s em o l e c u l e sF. i g u r e2 4 5 s h o * s
t h e p o s i t i o no f t h i s d o u b l eb o n d i n t h c p r i n c i p a lm o n o e t h e n o i d s( .2 6 7 )
A n d r e h a ss h o w nt h a t t h e d o u b l eb o n d i s m o r e o f t e n p l a c e ds o a s t o s c p r rate a group of 9 carbonsendingwith the carboxyl.We could show that
this group is presenttoward the extremityending with the carboxyl especially if the chain has l8 carbonsor less.For the longerchain, this group
of 9 carbonsis presentbut usuallytoward the extremity,ending rvith the
methyl. The importanceof this configurationfor the Knoop beta-oxidation
of theseacids is discussedin the previousNote.

NorES

//

619

Similar separationin groups of carbonstotaling 9 carbons,are seen

i n p o l y u n s a t u r a t ef adt t y a c i d s .( F i g . 2 a 6 )
Chapter 6, Note 6. Saturation-DesaturationBalance in the Liver
The total numbcr of doublebondsdoesnot changein the simultaneous
processes
of saturationand desaturation
occurringin the liver. To a preparation of liver cells,saturatedand polyunsaturated
fatty acidswere added.
The iodine numbcr of the fatty acid mixture presentwas determined,as

ic llame
I'lame Systemat
Common

Formula

A 4 , 5 D e c e n o i c *oo"A4AA4'*'
O b t u sI i c
C a p r o l e i c A 9 , 1 0D e c e n o i c xooc&s\AAl"'
A 9 ,l 0 D o d e c e n o i cHoocAAAAI\/,:"
L a i n Ioe i c
A 5 , 6 T e t n a d e c e nxoo<
o i [fuArr1.r41r17c"r
c
o i cn,An.AAA/'n'
l.lyristoleic A 9 ,l 0 T e t r a d e c e n u*
nrlav\'\l'Wvcxs
xooc
P a l m i t o l e i cA 9,l0 Hexadecenoic
ic
P e t r o s ienl i c A 6 r ? O c t a d e c e n o,,*.AAAAAAAA/'''
,.oo.nz\4lA.1yl^yV.cut
0leic
A 9,l0 0ctadecenoic
ic
Vaccenic A l l , l 2 0 c t a d e c e n o-*AA.z,\AAqA/NAIc*r
Gadoleic A 9 , 1 0E i c o s e n o i c ,*"Wt"t

A l l , l 2 E c o s e n o i c eoo(W(*t
C e t r loei c A l i , 1 2 D o c o s e n o i c* o o . W t " t
Eonicic
A i 3 , 1 4D o c o s e n o i c*-.Wc*'
x oiocc W t " l
, 6T e t r a c o s e n o
S e l a c h o l e iAc 1 5 1
A 1 7 , 1 6H e x a c o s e n ou oi ocW t * t
Zr i c o s e n o i c
A 2 1 , 2 'T

Ct{g

F l c . 2 4 - 5 . P r i n c i p a l n a t u r a l l y e x i s t i n g m < l n o e t h e n o i dssh o w t h a t f o r t h e m e m b e r s
w i t h a s h o r t c a r b o n c h a i n t h e d o u b l e b o n d s e p a r a t e sa g r o u p o f 9 c a r b o n s t o w a r d
t h e e n d o f t h e m o l e c u l eh a v i n g t h e c a r b o x y l .F o r f a t t y a c i d s w i t h m o r e t h a n l E c a r bons, the group of 9 carbons separatedis toward the end with the mcthyl 8roup.

620

RESEARCH IN

PHYSIOPATHOLOGY

well as the quantity of mono-unsaturated

members.After incubation at
37oC, the amount of monoethenicmemberswas seento have increased
greatly.Analysisof thetotalfatty acidspresentin the preparation,
however,

u.a

a gtz
cta

A rt,n
Cro

+LaJ.

r*- A

Agrz.tt

G1

+a+
t

Czo

qt,t
cto

-f1r

'

tJgtJ-g

L 2,,.,t
czo

,!

L , z .o . ' t
1zr-

,!

ta

22

20

+]Jr

711rJ

Ttro- A
Igt.rt.rt
cao

f i z r o r cr c

Czz

A",z,o.,,
cla

{Js

C
JJs

&t(n'A
l 6 r
A<,0.r;. ,1tt
c zz'
4arL44L41LJJL

S".

t-g
t

J
r

'

to

ta

t'

tL

tt

'o

Ar ro.u,to.,g
czz

H < r a -A

i-:IJ1 l-tO./t /6 /9
czt

,rqJJglJL

2
Lt

6
'gJ

5!.'

to

,LtJ-rJL

/2

/a

s
r.t

JJs

.Ll_

L,+t.to n to
,A
,/ v \,/
c,L4+Lr-Lr!L_rJu!:-!+

)o

2z

)o

22

&-

/a

"

J!qt-

Fro. 246. In the polycthenoids, groups of 9 carbons are recoSnized, formed usuatly by the sum of two or three groups,multiple of 3 (3 or 6). The group of 9 carbons
i s u s u a l l yp l a c e dt o w a r d t h e m e t h y l e n d o f t h e m o l e c u l e .

showed the same iodine number, indicating that in the changeswhich had
taken place, the saturation and desaturationprocesseshad comPensated
each other through a transfer of double bonds from the polyunsaturatedto
the saturatedmembers.

NorEs

621

Chapter 6, Note 7. Essential Fatty Acids

A strong positive characterof the carbon of the carboryl resultsfrom
its bond to two oxygens.This inducesan alternatingpolarity, with the odd
carbonspositive in the chain. On the other hand, the influenceexertedby
a double bond in the molecule correspondsto an enhancementof the
propr charge of the adjacentcarbons.When a carbon is situated in an
intermediary position between two double bonds, the influence resulting
from the two double bonds is highly increased.Thesetwo factors-positive
characteras an odd carbon and intermediaryposition betweentwo double
bonds-make Crr of linoleic acid a particularlystrong positive carbon
which apparsto be espcciallyable to bond a negativeoxygen.We consider
this stronglypositivenrcthylcniccarbon to bc thc condition which determineswhethera fatty acid has the characterof an "essential"fatty acid.
ttt
Hooc

(+)

(+)

(-)

Linoleicacid

L i n o l e n i ca c i d
(+)

(+)

A r a c h i d o na
i cc i d

F t c . 2 4 7 . R e l a t i o n s h i pb e t w e e n t h e p o s i t i v e c h a r g e o f t h e m e t h y l e n i c c a r b o n a n d
'l-hc
similarity which exists between linoleic
the character of essential fatty acid.
a n d l i n o l e n i c f a t t y a c i d s a s e s s e n t i a lf a t t y a c i d s ,c a n b e e r p l a i n e d b y t h e f a c t t h a t
b o t h h a v e o n l y o n e p o s i t i v e m e t h y l e n i cc a r b o n . A r a c h i d o n i c a c i d , w i t h t w o s u c h
p o s i t i v e l yc h a n g e dc a r b o n s ,h a s t h i s c h a r a c t e ro f e s s e n t i a lf a t t y a c i d s m a r k e d l y i n creased.

This explainsfurther a peculiarrelationshipbetweenthe three important essentialfatty acids. No differencesare seen,from the point of view
of activity as essentialfatty acid, betweenlinoleic and linolenic acid, although the last has 2 intermediarycarbons,one at Crr and another at
C1a.This can be explainedby the fact that the secondintermediarycarbon,
the C1a,as an even carbon, has a negativeelectricalcharacter.From this
point of view of strongly positiventethyleniccarbon, there is no difference between linoleic and linolenic acids. The relationshipbetween the
characterof essentialfatty acid and intermediarypositivecarbon is further
confirmedby the fact that arachidonicacid, with two positiveand one negative intermediarycarbons,is also a more active essentialfatty acid than
linoleic and linolenic acids,each of which has only one positiveintermediary carbon.(FiS. 2a7)

622

xESEARcHtN pHyslopATHoLocy

Chapter 6, Notc 8D. Fixation of Halogens on Coniugated

Double Bonds
The fact that the fixation of halogensat the conjugateddouble bonds
occursin two stepswould explainthe relativedifficultyin a reversereaction.
As seenfor butanediene,the halogenions are first attachedto the external
carbonsof the conjugatedformation with the appearanceof a double bond
betweenthe centralcarbons.It is only in a secondstepthat two other halogens are bound to thesecentral carbonstoo, thus completing the bond of
h a l o g e n tso a l l t h c c a r b o n so f t h e c o n i u g a t e fdo r m a t i o n .( F i g . 2 4 8 )

ctA/"

,rxr[

F t c . 2 4 8 . F i x a t i o n o f h a l o g e n sa t t h e c o n j u g a t e dd o u b l e b o n d s t a k e s p l a c e in t*'o
steps, with a displacementof the double bond in an intermediary position in the
f i r s t p h a s e ,a f a c t w h i c h e x p l a i n st h e n o n r e v e r s i b i l i t yo f t h e p r o c e s s .

Chapter 6, Note 8A. Solvent Fractionation of the Lipidic Constituents

In studyingthe biologicalrole of the lipids, we recognizedthe importanceof the fornts under which the differentlipids are presentin the
body, forms which seemto determinegreatlytheir activity.A lipid changes
its reactivitywhen it passesfrom the free form to one bound to other
A first stepin this studywas the analyticalseparationof these
constituents.
thus four fundamentalforms for acid lipids as well
forms.We distinguished
fractions:
for
unsaponifiable
as
I. Free lipids, or thosebound in such a labile physicalform as to be
able to take part dircctly in differentreactionsthrough their polar groups.
w i t h o t h e r c o n s t i t u e n t st h, a t i s , i n a r e l a I I . L i p i d s b o u n di n c e n a p s e
form.
tively labile
I I I . L i p i d s i n c o m b i n a t i o ntsh r o u g ht h e i rp o l a r g r o u p sa s e s t e r so r f a t s .
usuallya reserveor an inactivecirculatingform which
This form represents
can becomeactivethroughhydrolysis,
as to be inseparable
IV. Lipids bound so firmly to other constituents
of thc materialin order to be
throughsolventsand to need saponification
liberated.
The first form would representthe functionalform. the seconda rapid
availablefunctionalreserve,and the third, a reserve,The last would represent a stablecombincdform as part of thc buildingof entitiesthcmselves.
in solubilityof thesevariousforms of
We have utilizedthe differences
lipids in order to separatethem from the materialto be studied,and thus to
s t u d yt h e i ri n t c r v e n t i o inn d i f f e r e nnt o r m a la n d a b n o r m acl o n d i t i o n sI.t m u s t
that this separationconcernsonly the form under rvhichthc'
be emphasized
lipids are presentin the organismand not their chemical constitution.
the
In spitc of only a relativedegreeof accuracyin some separations,

NorES

/'

623

differences
noted from one sampleto anotherare so manifestand so consistentthat this mcthod can be consideredas an interestingand reliable
sourceof information.We have,therefore,usedthis techniqueof separation
for thousandsof samplesthroughthe years.
Accordingto the techniquewe devised.the materialto be analyzedtissues,organs, entire organismsor only biological products-is finely
divided in a blender.It is then extractedseveraltimes with ether under
stirring,or in a Soxhletapparatus.Under theseconditions,ether removes
the lipidspresentin the form of free lipids and neutralfats.This represents
a mixtureof the fractionsI + Il. The rcsidueis againextracted,this time
w i t h a m i x t u r eo f l 0 % e t h y l a l c o h o li n e t h e rw h i c h b r c a k st h e c e n a p s e s
a n d s e p a r a t etsh e l i p i d sp r e v i o u s l b
y o u n di n c e n a p s eT. h e r e s u l tr e p r e s e n t s
fractionIII. Thc residucis saponifiedwith l}c/o KOH and extractedwith
ether.This represents
fraction lV US or the unsaponifiable
lipids of fraction IV. After acidification
w i t h t a r t a r i ca c i d . a n e w c x t r a c t i o ni s m a d e
which represents
fractionIV LA or the acid lipids of this fraction.
The ether of fraction I + tl is distilledand the resiductreatedwith
85-90Voalcohol concentrationwhich dissolvesselectivelythe free lipids
as fraction I, and leavesthe part formcd by neutral fats as fraction II.
FractionsI, II and III are then saponifiedseparately.The unsaponifiable fractionsarc extractedwith ether, giving respectivelythe fractions
n i t h t a r t a r i ca c i d , o t h e r e x I U S , I I U S a n d I I I U S . A f t e r a c i d i f i c a t i ow
which
representfractionst LA,
with
made
respectively
ethcr
are
tractions
I I L A a n d I I I L A . E a c h o f t h e s ef r a c t i o n si s w a s h e dw i t h d i s t i l l e dw a t e r ,
dried with anhydroussodiumsulfateand, after the etheris distilledoff, the
fractionsare weighed.
For each material,we obtainedthus four different fractionsfor the
part which we called the US fractions,and four for the
unsaponifiable
saponifiablecalled the LA fractions.They correspondrespcctivelyto:
I , f r e e l i p i d s ; I I , l i p i d s p r e s e n ta s e s t c r so r f a t s ; I I I . l i p i d s b o u n d i n
s h i c h a r e l i b e r a t c do n l y t h r o u g h
c e n a p s eI;V , l i p i d sf i x e di n c o m b i n a t i o nw
In the followingexamples,we chosedifferentmaterialsto
saponification.
illustratethe kind of informationobtainedthrough this method.Fig. 249
the resultsobtainedin normal rats, in rats under abnormalconrepresents
w
d i t i o n sa
. s e l l a s i n t u m o r s ,a t l e x p r e s s eads t h e 8 l i p i d i c f r a c t i o n s .
In order to facilitatethe comparisonbetweennormal and abnormal
c o n d i t i o n sw, e c h o s ef o r t h i s e x a m p l e r, a t s b e t w e e n1 8 0 a n d 2 0 0 g m s ' o f
w e i g h t ,a l l m a l e sw i t h o n e e x c e p t i o n( c a s c b ) . T h e d a t a o b t a i n c dw c r c
thus comparedwith that obtainedfor case (a) which correspondsto a
normal male rat killed by ethcr.Thc analysisfor thc lipoacidfractionsof
t h i sc a s es h o w sa f r a c t i o n( l V ) a t 3 . 5 m g . / p e rg r a mo f a n i m a l ,t h e f r a c t i o n
a t . 2 m g . i p e rg r a m ,l I c o r r c s p o n d i ntgo f a t s a r l ' 2 m g . / p e r
III in cenapse
g r a m a n d t h e f r a c t i o n - la s f r e e l i p i d sa t 8 . 1 m g . / p e rg r a m o f a n i m a l 'F o r
ih. unruponifiablefraction. the fixed part IV is at 0.8 mg./per gram-of
frec
a n i m a l ,t h e I I I i n c e n a p s ca t 2 m g . , t h e I l a s f a t s a t 0 ' 8 m g ' ' a n d t h c
part I at 7 mg./gramof animal.
The female rat (b) showsin general,lower valuesfor the acid frac-

624

RESEARCH IN

PH}'SIOPATHOLOOY

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-t{

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f ' r r l i ; . - , 1; n , ; l i t p t , l r

Ftc. 249. Solvent separalion between difrerent lipoacids and unsaponifiable fractions
in various aninrids and tumors. l'hc vlrlucs cxprcssedin mgr,,'grof animal show big
varialions concerning espccialli' the frcc lipidr. (Top fraction)

t i o n sa n c ih i g h c rf o r c u c hf r a c t i o no f t h r u n s i . r p r ) n i R apbl rl ct . I t i s i n t e r e s t i n g
to note that the fixed part (lV) in acid iipids is lorverthan in any male,
'l'hc
a fact conflrnrcd lry otht-.ranalyscs.
increascin unsaponifiablepart
concernsespeciallythe ccnapse"fats and frce lipicl fractions.These differencesare in &grcementwith the data conccrningthe rclationshipbet$recn
lipids in males and females,lrs discussedin Chapter 6.
It is interestingto nc,tethe changr'sin starved animals a$ $hown in casc

NorE s

625

(c) where a marked decreasein the unsaponifiablefractions is seen,but

with an increasehowever,in the acid part, especiallyin the free fraction
(which reachesvaluesof 13 mg./l gm.). Similir changesare seenin acute
shock (d) inducedby the intravenousinjecrionof a rich culture of Esch.
coli in broth. Although death occurredin less than 40 minutes,a marked
increasein free fatty acids is seen with a correspond.ing
decreasein the
unsaponifiablefractions.The deviationfrom normal in the same direction
appearsstill more manifest for the animal in a traumatic state of shock
(inducedby 700 falls in the collip-Noble drum), (e).The dying animal
showeda markedincreasein free farty acid (37ng./l gm.) wiih inotable
reductionin the unsaponifiable
fractions.cases*ith ov-.r 70 mg./l gm. of
free fatty acid were also found. A lethal dose of radiation (1500r) (f),
and caloricburns (g), were seento inducesimilarchanges.
AIso of interestwas the influenceexertedby the intraperitoneal
injection
of gJyceroland choline, with an impressingincreasein all the fractions
of the acid and unsaponifiablelipids, and of those in cenapseand fat
fractionsin (h). we choseexample (i) of anotheranimal submittedto
the sametreatmentto show the extentof thc concordance
of thc information furnished by this method.
It can be seenthat almostunder all conditions,the amount of fraction
IV as obtainedthrough saponification,
changesvery little. The amount
presentin cenapseand as neutralfats show more variations.Varying considerablyfrom one caseto the other, are the free lipids as fraction I which
thus could be connectedmorc directly with a pathogenicinterventionof
the lipids.
Another conclusioncan be drawn from the analysisof theseresults.
The high amount of free lipids often obtainedunder abnormal conditions
cannot be considcredto result only from a liberationof thcselipids from
the pre-existingreserveas neutral fats or from the more labile cenapse
form, sincethe total amount of the free lipids found is much greaterthan
the sum of theseforms. An appearance
of new lipids, through synthesis,
has become evident in theseconditions.
Example (j) is the analysisof an animal with a l24ay old Walker
tumor. Comparison with the normal controls shows a reduction of the
acid lipidic part, with an increasein all the four fractions.The study of the
tumor itself (k) showsstill greaterdifferences.The fixed fraction is smaller
than in the total body of the animals,while the cenapsepart is greatly
The free lipoacidicfractionis almostnil. For the unsaponifiable
increased.
fractions,that in cenapse,and especiallythat corresponding
to free lipids,
is greatlyincreased.In the necrotictumor all four lipoacid and unsaponifiable fractions arc increased.Thc latter however, to a greater degree.
Though the cenapseamount is high, an increaseis most evident in the
free fraction.Case (m) of an animal treatedwith sulfur mustard applied
The animal, dying the 14th
on the skin, appearsparticularlyinteresting.
fractionsleft (less than I mg./l gm.).
day, has almostno unsaponifiable
We used this method extensivelythroughoutthe years,in spite of its
one limitation-the imperfect separationof neutral fats from frce lipids.

626

x E S E A R C Hr N p H y s r o p A T H o L o c y

The greatconcordance
for all variationshowever,has practicallyoverruled
any objectionof a failure of the separationof thesefree lipidic fractions.
Chapter 6, Note 8B, Spectral Analysis
In collaborationwith Carlos Huesca-Mejiaand PriscillaTeitelbaum.
we studiedseveralthousandsamplesof lipid preparationsthrough spectral
analysis,in ultravioletand first portion of visible light, and more rarely
in infrared,usingthe Beckmanspectrophotometer.
We will limit ourselves
to note here only some of the principalconclusionsreached.
I ) Concerningthc chemicalisomerization
procedures,we could show
the importanceof the temperatureused when a mixture o[ fatty acids is
treated.Thc conjugationin vitro as it is usuallycarriedout, with ethyleneglycol or gJycerolas solvents,was seen to result in preparationswith too
low amountsof tetra-,penta-and hcxacnicconjugatedmembers.A relatively rapid disapparance
of the conjugatedformationswith 4, 5 and 6
doublebonds was seento bc induccdby the high temperaturcused. This
led us to utilizea new methodof conjugation,at lower temperatures.
Using
cthyl alcohol as solvent,prcparationswith high conjugatedformations
were obtained.
2) We utilizedthe spectralanalysisfor quantitativedeterminationnot
only for di-, tri- and tetraencsas usuallyemployed,but also for pentaenes
and hexaenes.For this purposewe determinedthe extinctioncoefficient
to thesepentaeneand hexaeneformations.This was made
corresponding
possibleby isolating the respectivepentaenicand hexaenicconjugated
membersthroughappropriatcsolvents.
3 ) We studied various nraterialsand especiallydifferent organs in
order to corrclatctheir richnessin difTerentfatty acids to thcir biological
activity,by usingthe spectralanalysisof the in-vitro conjugatedfatty acids.
as mentionedabovc.
4 ) S i m i l a r l yw
, c t r i e dt o c o r r c l a t et h e e x i s t e n c o
e f c h a r a c t e r i s t ipce a k s
fractionsof organsto their
in the spectralanalysiscurve of unsaponifiable
biologicalactivity.
5 ) We utilizedspectralanalysisfor the study of the effectsof various
agentssuchas chlorine,sulfur,sulfuricacid or oxygcnupon the conjugated
fatty acids.
6 ) We showedthat minimalchangesare inducedin thenonpolargroups
of conjugatedfatty acidsby changingtheir polar grouPfrom carboxyl into
hydride.
t ith lithium-aluminum
a p r i m a r ya l c o h o l ,b y t r e a t m e nw
7) We studiedthe influenceexertedby conjugatedfatty acids upon
This can be partiallyrevealcdby the quenchingaction induced
carcinogens.
of theselatter agents.
the
fluorescence
upon
8) In an extensivestudy we investigatedthe influenceexerted by
w a sc h a r a c t e r r a d i a t i o nu p o nf a t t ya c i d si n - v i t r oa n d i n - v i v o .T h i s i n f l u e n c e
of conjugatedtrienes,and is presentedin Chapter
ized by the appearance
i
n
" R a d i a t i o n . "a n d o t h e r N o t c s .

NorES

627

Chapter 6, Note 8C. Vapor Fractionation of F"tty Acids

In a group of expcrimentswe apptiedthe gaschromatographymethod
to the studyof fatty acids,The principalaim was to investigate
the valueo[
the information furnished by this method concerningthe presenceof
conjugatedfatty acids. This study was made in collaborationwith Ivan
B i e r a n d w i t h W i n s t o n D i n d i a l w h o p r e p a r e dt h e s a m p l e s .
Methyl esters of eleostearicacid, linoleic acid and its conjugated
isomers;of linseedoil fatty acids and the conjugatedpreparation;of cod
Iiver oil fatty acids and the conjugatedpreparations;and of samplesof
fatty acidsobtainedfrom animalsand tissuesunder normal and abnormal
conditionswere obtained.We analyzedthroughvapor fractionation,these
differentpreparations
as such,the preparations
obtaincdthrougihcondensation on cold fingersduring distillationin vacuumat differenttemperatures,
and the differentfractionsobtaincd through distillationin vacuum. For
all thesetestswe usedthe Perkin-Elmervapor fractionerwith a column of
succinylpolymersheatedup to 235"C. Under thescconditions,no differencescould be seenbetweenthe respective
conjugatedand nonconjugated
s a m p l e sF. i g s . 2 5 0 a n d 2 5 1 s h o w e x a m p l e so f s u c h a n a l y s e so f a c o d
liver oil fatty acid preparationand of a preparationobtainedafter treatment with KOH in butyl alcohol. Fig. 252 shows the spectralanalysis
of this last product.
Under the conditionof analysisused,the gas chromotographymethod
does not permit the identificationof the conjugatedisomerspresent.This
is the reasonwhy the analyticalmethodcould not indicatethe presence
o f s u c h m c m b e r si n m a t c r i a l so b t a i n c dd u r i n g a b n o r m a lc o n d i t i o n sT. h e
conjugatedisomerscan be identifiedby other methods-such as spectral
y x a l i ci n d c x - a f t e r o x i d a t i v ef i s s i o n .
a n a l y s i sa n d e s p c c i a l l o
W e a r e n o w t r y i n g t o o b t a i n c o l u m n st h a t w o u l d p c r m i t w o r k i n g a t
much highertemperatures
and would pcrmit us to identifytheseconjugated
fatty acids.In view of the minimal amountof materialneededfor analyses
and the precisionof thc resultsusually obtaincd,an adaptationof this
methodfor the identificationof conjugatedmemberswould be of especial
g r c a tv a l u c .
Chapter 6, Note 9. Twin Formation
The odd number of carbonsin a cyclic moleculerepresentsone of the
conditionswhich always would result in the appearanceof a twin formation, since the alternation between positive and negative signs gives the
sameelectricalchargeto two nearbycarbons.(Fig. 254) The correlation
of the positiveand negativechargesof nearbycarbonsto the fact that acetic
acid moleculeshave been utilized in the synthesisof the moleculerepresentsan additionalfactor for inducingtwin formationseven in cycleswith
an even number of carbons.

(x

$l
o
lt
N
G

9,

't

"r
ti

s'

Llnolenate

o
q1
o
ct
o
CL

l
c

O
t

o
o,

o
cl
0

Llnoleate
o

0leate
Stearate

Palal
toleate
Palmltate
Ityristate

O . m l H l S a m p l2e2 0 c C2. UP S I G3 H " P "

Ftc. 250. Gas chromatographic analysis of a sample of cod liver oil fatty acids
showing some of the different constituents.

3
t

L i n ol e n a t e
l

o
o
o

r
1
c\

Lrnoleate
Uleate
5teanate

Palnrioleale
i'almii.ate

l,lyrist at e

0 , 0 0 1l { l S a m p l2e2 0 0 C2.0 P S I G3 l { " P "

F r c . 2 5 1 . G a s c h r o m a t o g r a p h i ca n a l y s i so f a s a m p l eo f t h e f a t t y a c i d s o f c o d l i v e r
oil after chemical conjugation. No differencesare seen between the curve of Fig. 250
and this curve, indicating that in the conditions under which the analysis was made,
aar= aarr rt

-,:'
. i

630

R E S E A R C HI N

P H Y S I O P A T H O L O C Y

cod I ivenoi I f atty acids

Hethylesterscon.,;r.rqated
0 . 0 0 2 t n c t h y l a l c o h o l / e t h yall c o h o l

c
o

E
c
d
L
!l

;ave Length ("V)

F r c . 2 5 2 . S p e c t r a l a n a l y s i so f t h e s a m p l e u s e d f o r g a s c h r o m a t o g r a p h y .s h o w n i n
F r c . 2 5 1 , i n d i c a t i n gl h e p r e s e n c eo f d i - , t r i - , t e t r a - ,p e n t a - a n d h e x a e n i cc o n j u g a t e d
members.

Frc, 254. A twin formation appears in cyclopentaDe as a

r e s u l to f t h e a l t e r n a t es i g n o f a d j a c e n t c a r b o n s .

NOTES

631

Chapter6, Note 10. ArachidonicAcid-SterolRelationship

The relationshipbetween arachidonicacid and sterols,both of which
are presentin the adrenals,has been followed through the changeswhich
occur in the amount of these substancesin the adrenalsduring certain
pathologicalprocesses.
Rabbits were slowly injectedintravenouslywith a
suspension
of microbesuntil symptomsof deepacuteshockappeared.The
animalswere sacrificedimmediatelyby bleedingthem and the adrenals
and blood analyzed.The amount of polyunsaturated
fatty acids of the
adrenalshad markedly decreased,and, in some animals, had almost
disappeared
from theseorgans.At the sametime, the amount of the same
fatty acidsof the circulatrngblood had manifestlyincreased.The amount
of insaponifiable
fractionsand sterolsin thc adrenalswas unchangedas
comparedwith controls.It seemsthat fatty acids have passedfrom the
adrenals into the general circulation as a first responseto the noxious
intervention.
Chapter 6, Note I l. Steroids Deriving from Arachidonic Acid
The study of the relationshipbetweensteroidsand certain fatty acids
led us to the hypothesis,
accordingto which somesteroidsare derivedfrom
fatty acidsthemselves.
Although the synthesisof cholesterolfrom scalene
(230) is highly plausible,it would not representthe only origin for all the
steroids.The membcrswith a two carbon chain would derivc from other
substances.
We presentthis hypothesishere becauseit also representsan
exampleof anotherimportantrole playedby the double bonds which appear to intervenein the processof cyclizationin the organism.
Accordingto the hypothesiswhich we have advanced,polyunsaturated
fatty acidslead to allopregnanc,
thc parentsteroidwith a two carbonlateral
c h a i n . F i g u r e 2 5 5 s h o w st h e d i f f c r c n tp h a s e so f s u c h a t r a n s f o r m a t i o n
startingfrom arachidonicacid.The prescnceof a carboxyland four double
bondsin the arachidonicacid molcculeresultsin someof its carbonshaving a particularlystrongenergeticvalue.The stronglychargedcarbonsare:
l) Cr, throughits bond to : O and - OH; 2) Cz and C3, through an induction processsincethey are closeto the carboxyl;and 3) Cc and Co,
Ca and Co, Ctr and C13,and C16and C17,respectively
bound by double
bonds.
Due to alternateinduction.all the odd-numbered
carbonshave a positive signand the even-numbercd
havea negativeone. as indicatedin Figure
2 5 5 a .B e c a u s o
e f t h e h i g h f l e x i b i l i t yo f t h e a l i p h a t i c h a i n .a n d t h e p r e s e n c e
of carbonswith strong positiveand negativecharacterin the same molecule, attractionsbetweenthe stronglychargedcarbonswith oppositeelectrical signsin the samemoleculewould occur.This would lead, as a first
step,to a bendingof the chain so that the stronglychargedcarbonswith
o p p o s i t es i g n sw o u l d f a c c o n e a n o t h e r (. F i S . 2 5 5 b )
In a secondstep, as thesecarbonsare bound by double bonds, the
respectivezr electronsof the double bonds would serve to form a new

632

REsEARcH rN pHysropATHoLocy

bond betweenthe facing carbonsand thus to close cycles.This would ocrur

without any loss or gain of electrons.The three double bonds between C5
and C6, C11and C12,afld Crr and Crr would serveto closethe three cycles.
(Fig.255cl The double bond betweenCs and Ce would seryeto make Cb
highly reactive. It can be seen that Ce of arachidonic acid corresponds to
C,, of cyclopentanophenanthrene,
which explainswhy this carbon has a
high positive character, with an oxygen fixed on it. The carbon of the carboxyl could be used either to form the methyl goup at Cra or, more plausibly, could be lost in a processof decarboxylation,which by itself, in this
case,would induce the bond betweenCz and Crs of the arachidonic molecule. Through the intervention of the strong energetic center, the pentanic
cycle would be closed and would have two carbons with the same charge.
(FiS. 255d) The two methyls, correspondingto C6 and C1e of steroid
molecules, would result from a further process of methylation afrcr the
polycyclic molecule was formed. C4 and Ce of arachidonic acid would be
especiallylikely to have a methyl $oup fixed on them through their electronic displacementdue to the new bond.
-lo

t cHt

(a)

(d)
Ftc. 255. Hypothesis of the synthesis of the allopregnane radicd from arachidonic
acid. Fig. (a) shows the relative position of tbc double bonds in thc arachidonic acid
molecule. Fig. (b) shows how the molecule bends,due to the attraction betwecn tbc
energeticallyoppositely charged C: and C*, C., and C',, and G and C,'. ln Fig. (c)
the cycles are closed with the electronsfurnished by the double bonds. (d) Tbc closing of the cyclopentane occurs with the appearanceof a twin formation. The electrons
of tbe double bond between C, and C, become available at G (G of allopregnan)
to realizc Oe bond with an oxygen at this carbon.

NorBs

633

This hypothesisexplainsthe biological relationshipbetweenarachidonic

acid and corticoids. The adrenalsare especiallyrich in both. The synthesis,
which occtus with a minimum changein the richnessin electronsor atoms,
explainstwo of the most important characteristicsof these corticoids,the
high energeticvalue of Cs and of the cyclopentaxewith its twin formation.
Preliminary experimentsseem to show that, in a preparation of adrenal
tissue, arachidonic acid can be transformed to corticoids under the influence of the adrenocorticotrophichormoneof the hypophysis(AgtH).
Chapter 6, Note 12. Steric Coupling
Through reciprocal influence, the energeticcenters present in the nonpolar parts of two moleculeswhich combine, largely lose their activity.
Steric coupling is possible if the two opposite molecules,once bound
through the combination of their polar groups,also adheretogetherthrough
their nonpolar groups via the opposing energetic centers or formations
present in the two molecules.In this way, steric coupling completesthe
partial reciprocalneutralizationof the moleculesobtainedthrough the combination of polar groups. The bond betweenthe polar groups that keeps
the two moleculesin a reciprocalposition is thus an important condition
for steric coupling.
Chapter 6, Note 13. Luteoid Function
It appearsthat the luteoid patterncan be correlatedto a specificaspect
of the energeticpicturc of this 5uf5fangg-the presenceof two relatively
strongnucleophilicccntersin thc characteristic
oppositepositions,one at
C3 and the other at C2e,as in progesterone.
In fact, any changein the energeticpictureof this steroidwill decrease
luteoidproperties.The relationship
of the luteoid property to the nuclcophilyat C3 is easily seen.The lack of
t h e n u c l e o p h i l icce n t e ra t t h e c a r b o n3 , a s i n p r e g n a n e
one 20, (Fig.256a)
Ieadsto an inactive substance,the energeticpicture being entirely changed.
A singlenucleophiliccenter thus appearsto be insufhcientfor the luteoid
property.
The substancebecomes inactive if, instead of a nucleophilic center
througb : O at the carbon 3, an electrophiliccenter is present,as in the
pregnaneol 3 one 20, so that the energeticpicture no longer corresponds
t o t h e l u t e o i dp a t t e r n .( F i g . 2 5 6 b )
A n y c h a n g ei n t h e n u c l e o p h i l o
yf thc C
O c c n t e ra t C r w i l l d e c r ea s c
the luteoid propertiesof the substance.The presenceof a seconddouble
bond betweencarbons 6 and 7, as in the 4, 6 pregnandiene-dione
32O, (Fig.256c) in spite of the fact that it will increasethe value of the
nucleophilyof the carbonyl at the carbon 3, will changethe energeticpattern and thus also decreasethe luteoid character.On the other hand. the
reductionof the nucleophilyof the centerproduces,through a similar influenceupon the energeticpattern,a dccreasein the lutcoid property.In
t h e c a s eo f p r e g n a n e - t r i o n3c- 6 - 2 0 ,( F i g . 2 5 6 d ) t h c l a c k o f i r d o u b l cb o n d

634

nEsE^RcH tN pHystopATHoLocy

in the cycle and the reciprocalinductionrealizedby the parallel double

bonds of the two carbonyl,wil.l reducethe nucleophilyof the C. centcr.
and with it, the luteoidproperties.This alsois true for A 5 pregnenedione3 - 2 0 - o l6 , ( F i g . 2 5 6 e ) w h e r et h e p r c s c n c eo f t h e h y d r o x y la r t h e c , , . a s
well as a nonparalleldoublebond betweenC5 and C6, decreases
the ionic
characterof the c - o at the carbon3. In s 5 pregnene-dione
3-zo, (Fis.
2 5 6 1 .w) h e r eo n l y t h e d o u b l eb o n d b e t w e e nc a r b o n s5 a n d 6 i s c h a n g e d .
beingoppositeto that of the carbonylat the carbon 3, the ionic value of
c : o at c3 is decreased
insteadof increased.
The luteoidproperty seems
to have almostdisappeared.
The secondconditionfor the luteoidpropertyis the nucleophiliccenter
at czo.Any changein its characteror valuewill influencethe luteoid prope r t yo f t h e s u b s t a n c eA.n d r o s t a n d i o n(eF. i g .2 5 6 9 ) w i t ha n u c l e o p h i l icce n t e r
at carbon 3 similar to that of progesterone,
but with an : O directly at-

,\^) -.ar--,
lnacttve (a)

Decreasedactrvitv

ll tl G )

Inactrve

t i
\?v

(b)

D e c r e a s e da c t ' " , 1 y G )

./-A/., ")yy (r
?(,u,ty
Decreased

Vsry low actrv'ty (g)

-0x

Inactrve

activrty(jl
Decreased

lr.actrve

(,)

activity
Decreased
- 0

l n aaccti
t r vve
e 0
0))

lnactive

( m)

F r c . 1 5 6 . A n y c h a n g e i n t h e e n e r g e t i cc h a r a c t e r i s t i c so f t h e c e n t e r s w h i c h a p p e a r
r e l a t e d t o t h e s p e c i f i cl u t e o i d a c t , i v i t yl e a d s t o a d e c r e a s eo r d i s a p p e a r a n c eo f t b i s
activity.

NOTES

6]5

tached to carbon 17 and without any side chain, presentsan energetic

picturewith two nucleophiliccenters.But the nucleophiliccenter : O attacheddirectlyat carbon 17, givesa differentcharacterto this part of the
m o l e c u l eT. h e s u b s t a n ctch u s h a s o n l y s o m el u t e o i dp r o p e r t i c sA. 4 p r e g n e n e - o n e3 - o l - 2 0 , ( F i g . 2 5 6 h ) i n w h i c h t h e c a r b o n y l a t c a r b o n 2 0 i s
changedby a hydroxyl,is, on the otherhand,inactive,the substance
having
a picturedifferentfrom that of the dinucleophilicpattern.Also inactiveis
J 5 prcgnone--<Jionc-3-20-ol
17, (Fig. 256i) in which the presenceof the
hydroxyl at carbon l7 changesthe energeticpicture at this extremity of
the steroid.
T h e p r e s e n coef a d o u b l eb o n d b e t w e e nc a r b o n s1 l a n d 1 2 ( F i g . 2 5 6 i )
will decreaseluteoid activity through an induction and field influencebetweenthe nonparalleldouble bonds and a decreaseof the ionic character
of C26bound to oxygen.The influenceexertedby an hydroxyl in the neighborhoodof carbon 17 is interesting.Through an inductiveeffect,thc OH
attached
a t c a r b o nl 2 ( F i g . 2 5 6 k ) c h a n g etsh e i o n i cv a l u eo f t h e C : O a t tachedat carbon20 and thus decreases
luteoidactivity.An hydroxylbond
to carbon I I has a much more intensivecffect,changingthe ionic character of the bonds betweenit and the : o and thus inactivatingthe luteoid
p r o p e r t y .( F i g . 2 5 6 1 )
For most of the steroidswith : O ccnters.metabolismleads to a
changeof this center in an hydroxyl.The product of the changeof both
C : O into C : OH, which represents
the form in which this hormoneis
e x c r e t e di,s i n a c t i v e P
. regnane-dio
l
3 - 2 0 , ( f - ' i g . 2 5 6 n l )w i t h a s t r u c t u r ef a r
removed from the pattern required for luteoid activity, has no luteoid
properties.
Chapter 6, Note 14. Energetic Center in Steroids
Energetically,
as a consequence
of the presenceof a nucleophiliccenter
in this situation,there is a strongpositivechargefor the carbon 20 of the
chain itself. Consequcntly,
a strongnegativechargeappearsfor carbon 21.
The ionic value of the bond linking OH to C:r is thus incrcascd.With the
reactivityof this radical increased,the electrophiliccharacterof this OH
centerbecomesstill stronger.The respectiveactivityof two opposedenergeticcentersnear to eachother is increasedinductively,
Chapter 6, Note 15. RelationshipBetrveenCorticoids
Corticoidscan be viewedin termsof changestakingplacein the general
metabolismin the organism.In the evolutionof thesesteroids,a processof
oxidationwould interveneat the principalenergeticcentcrs.To the hydrocarbon,first an hydroxyl and then an oxygenwould be attached.An inverseprocessof reductionwould take place as a metabolicprocess,the
OH being the usual form through which steroids are eliminated, often
b o u n d t o g l u c u r o n i ca c i d . T h i s p r o c e s so f o x i d a t i o nw o u l d o c c u r a t C r r .
(e 4 pregnene-21-ol-3:20Attaching an OH to desoxycorticosterone

636

RESEARcH rN pHysropATHoLocy

dione), would lead to the appearanceof corticosterone(a 4-pregnenell:21-diol-3:20-dione). A further oxidationwould changethis OH into
an O, resulting in dehydrocorticosterone( A pregnene-21-ol-3
: I I :20trione). All theseare mineralocorticoid
compounds.
With a further change, this time at C17,where an hydroxyl would be
attached,all three compounds-desoxycorticosterone,
corticosteroneand
dehydrocorticosterone-displayneoglucogenicproperties. They represent
17 hydroxy derivativesas l7 hydroxy-desoxycorticosterone
(a 4-pregnenel7-9 (beta):21-diol-3:20-dione);17 hydroxycorticosterone
(A 4-pregn e n e - l l ( b e t a ) : 1 7 ( b e t a ) : 2 1 - t r i o l - 3 : 2 O - d i o noer )c, o m p o u n dF ; a n d l 7
hydroxy-1I dehydrocorticosterone
(a 4-pregnene-17:2ldiol-3:I I :20-rrio n e ) , o r c o m p o u n dE o r c o r t i s o n e .
The fact that the presenceof an hydroxyl at Crz greatly changesthe
propertiesof the entire group makesit likely that the energeticformation
of which Crz is a part intervenesin the specificactivity of thesesubstances.
The analysisof the constitutionof the corticoidshas further shown that
they have the charactersof lipoids-being polar-nonpolarsubstanceswith tie nonpolar group predominant,The membersstudied,mineralo- as
well as neoglucogenic
corticoids,have beenshown to induce a changetoward lower valuesin the secondday wound crust pH, indicatingthus a
tendencyto induce an offbalanceof the type A.
Chapter 6, Note 16. The Template Hypothesis
Figure 257 showsthc templateformationin cortisonc,which cxtcnds
from C11to C:r. Each one of thesesix carbonswill attract a carbon from
the radical in front of it. The energeticcharacterof each of the six carbons
of the templatewill determinethc electrophilicor nucleophilic character
of the carbon so attracted.This attraction is easily induced when acetic
radicals,with an clectrophilicand a nucleophiliccarbon,form thesegroups.
Furthermore,the value of the carbonsof the template also will determine
which polar radical will be bound to the respectivecarbon kept in front of
it. In general,the carbon kept in front of a carbon of the templatewill have
an oppositeelectricalsign. Thc polar group bound to the carbon kept in
placewill be oppositein sign to the polar group bound to the carbon of the
template.When the first polar group takesa position parallel to that of the
polar group of the template,both will have the sameelectricalsign.
lt can bc sccnthat C2t within the templatehas an OH, the group being
electrophilic.This will causethe carbon kept in front to preferablybind
an oxygen,realizinga nucleophiliccenter.Czoof the template,which correspondsto a carbonyl, representsa nucleophiliccenter. With an oxygen
bound through a double bond, it has strong reactivity.The positivity of
C3ealso is highly enhancedthrough its bond to the two strongly negative
carbons,C:o and C17,each being bound respectivelyto a hydroxyl. With
this high positivity,C3swill induce a strongreactivity in the carbon kept in
front of it. This will be strongenoughto bind a radical energeticallyopposite to oxygenand strongerthan the hydroxyl;that is, an amino group.

NorEs

637

The specialposition of the oH bound to crz as relatedto the template

will induce the carbon kept in front of it to bind another hydroxyl. The
sameappliesto carbon 13. This is the result of the relativelystrong molecular reactiviryof thesetwo carbons,due to the twin formation which they

H2

o=,/'\:->
HL

E,Q

"

O=
o\oH
F r c . 2 5 7 . H y p o t h e t i c a l v i e w o f t h e t e n r p l a t ef o r m a t i o n b e t w e e nc a r b o n l l a n d c a r bon 2l of the corticoid molecule. The groups kept in front of the different carbons
. t h e e x a m p l ep r e s e n t e da b o v e ,
o f t h e f o r m a t i o n s e r v c t o s y n t h e s i z en e w s u b s t a n c e sI n
t h e t e r n p l a t eo f t h e c o r t i s o n em o l e c u l ew o u l d l e a d t o t h e a p p e a r a n c eo f a g l u c o s a m i n e
molecule.

realize.The methyl bound to C'3 will determinethe steric position of the

hydroxylbound to the respectivecarbon kept in front of this carbon.
The effectof carbon 12 is different.It is highly influencedby the twin
f o r m a t i o na n d h a s a n o p p o s i t ce n c r g c t i c h a n r c t etro c a r b o n sl 3 a n d 1 7 ,
consequently,it will favor the binding of the carbon kept in front of it to
an oxygenwhich is the samewhich binds the carbonin front of Czr. Positionally, the carbon in front of Crz also will be nearestto the C kept in

638

pESEARCH rN

pHysropATHoLocy

front of C31,which alsowas inducedto bind an oxygenas seenabove.This

will make it possiblefor a similar oxygento be bound by the two carbons
kept in front of Crz ard Cer, thus closingan hexagonictycle. The radical
bound to carbon I I will causethe carbonin front of it to bind an opposite
radical.In the casesin which C11hasan oxygenbound to it, as in cortisone.
the carbon kept in front of it will bind a hydroxyl. The presenceof an
hydroxyl bound to Crr, ?S in hydrocortisone,
will causea carboxyl to be
bound to the respectivecarbon kept in front of it. It is thus seenthat the
synthesisinducedby the templateformation of cortisonewill result in a
moleculeof glucosamine,
while for hydrocortisone,
the synthesized
molecule will correspondto glucosaminic
acid.
Chaptcr 6, Note 17. Adrenal Defense Index
E. F. Taskier in collaborationwith the author, has studiedthe interv e n t i o no f a d r e n a l si n t h e d e f e n s e
a g a i n svt a r i o u sf a t t y a c i d s .( 2 3 1 )
It is known that the adrenalshave an important role in the defenseof
the organismagainstnoxiousagents.a normal animal being more resistant
to toxic effectsthan an adrenalectomized
onc. Systematicstudy of adrenal
interventionhas shownthat it occurswith a certaindegreeof specificity.
T h e m c t h o do f i n v e s t i g a t i ouns e dw a s t h e f o l l o w i n g :i n b r e dW i s t a r r a t s
of the samesex and approximatcbody weightof 150 gr. were adrenalectomized.The surgicalprocedurewas carefullystandardized,
lastinglessthan
two minutes,and producinga minimumof trauma.This was madepossible
by the Noyes Fixation Forcepswhich is utilized in ocular surgeryand is
well suitedto pick out the adrenalgland without damageto it or to neighboring tissue.As controls,animalsof the sameweightand sex were shamoperated.On the third postoperative
day, the agent to be testedwas injectedintraperitoneally,
At this time, the organismhad recoveredfrom the
immediatetraumaticcffectsof the operationand no manifestadrenaldeficiencysymptomswere as yet present,Only thosedeathsoccurringup to
48 hours postchallengewere consideredto be due to the direct toxic effects
of the substance,
Deathsoccurringmore than 5 days after adrenalectomy
could be attributed to effectsof the adrenal deficiencyitself, and for this
reason,were not includedin the experiment.
The minimal Lethal Dosefor eachagentwas determinedby using progressively
increasingdoses.This was done separatelyfor adrenalectomized
and for control animals.It was the differencein the toxicity in shamoperatedand in adrenalectomized
animalsthat was consideredrather than
the toxicity of the substanceitself.
T h e r a t i o o f t h c M i n i n r a l L c t h : r lD o s e f o r t h e t w o g r o u p so f r a t s .
was calculated. It furnished a
sham-operatedand adrenalectomized,
of the degreeof adrenal interventionand was
numericalrepresentation
called the "Adrenal DefenseIndex" or A.D.I. We utilized this index,
MLD Sham-ope_rated
as a measureof the relativeadrenalparticipation
MLD Adrenalect";r.d.
in the body defenseagainstdifferentsubstances.
A low index points to a

NOTES

,'

639

general nonspecificresponse;a higher index indicatesa more significant

intervention.
We obtainedthe adrenaldefenseindex for variousgroupsof fatty acids.
They includedhomologousseriesof saturated,unsaturated,
alpha-OH and
conjugatedfatty acids.Over 900 rats wereusedin this study.
For the saturatedfattv acids we found:
FATTY ACID
Caproic Acid
Caprylic Acid
Capric Acid

Lauric Acid
Myristic Acid
Palmitic Acid
Stearic Acid

A.D.I.

2.5
5
5
1.5

2
t2
6

We interpret this to mean that the defensecapacity of the organism

against these fatty acids was only slightly more effective in the shamoperatedthan in the adrenalectomized
animals.The adrenalsdid not seem
to be especiallyactive in the defenseagainstthesesubstances.
For the unsaturatedfattv acids we found:
FA]TY A9]D
Oleic
Linoleic
Linolenic

A.D.I.
6
9
5

These valucs indicate more active adrenal intervention against these

fatty acids.
For the saturatedalpha-OHfatty acidsthe resultswere:
FATTY ACID
alpha-OH Caproic Acid
alpha-OH Caprylic Acid
a l p h a - O HC a p r i c A c i d
alpha-OH Lauric Acid .
alpha-OH Myristic Acid
alpha-OH Palmitic Acid
alpha-OH StearicAcid

A.D.I.

4.5
4
3
20
9
3
50

We choosethis seriesof substances

becauseCamien and Dunn, and
have
the
importance
shown
others,
of thesefatty acidsfor bacterialgrowth,
as well as the presenceof alpha-OH lauric and alpha-OH myristic acids
as part of the lipido-polysaccharide
fractionof bacteria.
Through related research,we were particularlyinterestedin the A.D.I.
value of fatty acid moleculeswith conjugateddouble bonds. As conjugated
diene,we administeredconjugatedlinoleic acid. The A.D.I. of this substancewas similar in value to that of its nonconjugatedisomer.The index
was 5. For the conjugatedtriene, we used eleostearicacid obtained from
tung oil. The resultswere striking. We found an A.D.I. value of 120. The

640

R E S E A R c Hr N p H y s r o p A T H o L o c y

A.D.L of this acid thus showeda 24-fold increaseover the index of its
nonconjugatedisomer.
The data indicatea degreeof specificityfor the adrenaldefensemechanism. A.D.[. va]uesof 3 or lesscould be interpretedto correspondto a
nonspecific
adrenalinterventiontoward fatty acidsin general,while higher
valuesindicatea larger,probablyspecificadrcnal activity. In the case of
alpha-OHlauric and alpha-OHstearicacids,the high A.D.I. valuescould
be releatedto the fact that thesefatty acids have been found to be part of
the constitutionof bacteria.
The intensivedefenseevidenced
by the high A.D.I. valuefor eleostearic
acid is relatedto the appearanceof conjugatedtrienesduring the course of
certainpathologicalconditions,especially
trauma,The continuousincrease
,
especiallyof conjugatedtrienes,in thc adrenalectomizrd
animalssuggests
that thesefatty acids appearin the organismbut are destroyedunder normal conditions.They accumulate,howevcr,in adrenalectomized
anima-ls
and may contributeto deathwhen they reach a certaincritical concentration. It is possiblethat with additionaladministrationof theseconjugated
trienestheir criticalconccntrationwould be reachedand the animalswould
die.
In the light of thesedata, we investigatedthe role of different adrenal
factors in thesc responscs.Cortisone, desoxycorticosteroneacetatc
(DOCA), and sodium chloride were tested for their protective action
againstthe toxic effectsof oleic and eleostearic
acids.Groups of rats were
treatedimmediatelyafter adrenalectomy
with daily dosesof I mg. of cortisone,two-tenthsof a cc. of DOCA, or l7o NaCl drinking water ad libitum.
Control adrenalcctomized
rats wcre given no sustainingtherapy. Three
days after adrenalectomy,
a challengingintraperitonealdose of 1 cc. of
lUVo oleic acid pcr 150 gr. of body weight was administered.
Whereasthe mortality of the control rats was 90Va, it was 25% in
t h c c o r t i s o n et r e a t e da n i m a l sD
. O C A a d m i n i s t r a t i odne c r e a s etdh e m o r t a l ity only to 65Va and NaCl had little effcct,decreasing
it only to 85Vo.
The protectiveeffectagainstoleic acid in adrenalectomy
is seenin thc'
followingtable:
AGENT
C--ontrol
Cortisone
DOCA
NaCl

To MORTALITY
90
25
65
85

T h i s s u g g e s ttsh a t t h c n c o g l u c o g e nhi co r m o n ep l a y s a s i g n i f i c a npt r o tectiverole in the defenseof the organismagainstthe noxious effectsof
fatty acid.The mineralocorticoid,
on the other hand,seemsto play a lesser
role in this mechanism,a fact which is confirmedby the ineflectiveness
of
s o d i u ma d m i n i s t r a t i o n .
A similarpreliminaryexperimentcarriedout for eleostearicacid shorvs
the sameprotectiveeffectiveness
of cortisone,lessereffectiveness
of DOCA
and almost no effect at all of sodium ctrloride.

NOTES

641

Chapter 6, Note 19. Bonds of Glucuronic Acid

The study of the detoxifying-cxcretionof different agentsled to the
following conclusions.Primary aliphatic alcohols-except methyl alcohol
-are eliminatedcoupledby glucuronicacid; the samefor the secondary
aliphaticalcohols;tertiary aliphaticalcoholsand glycolsfrom propylene
glycol up. Of the aliphaticaldehydesonly a few are coupledand only after
transformationin vivo. While most of the aliphatickctonesare coupled,
phenol and more emphatica.llythe cresols and salicylic acid are in part
excretedas sulfo+oupled,and only in part as glucurono-coupled.
Resorcin o l . c a t e c h o lo, r c i n o l ,p h c n o l p h t h a l e i npch. l o r i d z i na r e m o s t l y e l i m i n a t e d
as glucurono-coupled,
while adrenalinealmost solely as sulfate.Most of
the aromatichydrocarbonsare also bound to glucuronicacid, but only
after having been changedin vivo.
For the aromaticacids,the bond to glucuronicacid is conditionedby
the presenceof secondpolar groups, usually one or more hydroxyls. The
aromaticnitrogencompoundsare first changedinto amino groupsbefore
they are coupledwith the glucuronicacid. Only relativelysmall amounts
of sulfonamidcs
were cxcretedbound to glucuronicacid.
Many of the heterocycliccompoundsare bound to glucuronic acid
with the conditionto have amino or hydroxylgroups;the samefor the sex
hormones,the estrogens
being the group cspeciallyexcretedas such.
The generalcharactcristicof the substances
excretedas coupled with
glucuronicacid is the prcsenccin their moleculeof one or more positive
polar groups hydroxyl or amino. As with fcw exceptionsall these substanceshave also lipoidic propcrties,the excretioncoupledto glucuronic
a c i d a p p e a r sa s a m c a n st o e l i m i n i r t cp o s i t i v cl i p o i d s .
Chapter 6, Notc 20. Cltrctrronic Acid-N{echanismof Coupling
The analysisof urine specimens
containingperoxides,also has revealed
significantamountsof glucuronicacid compounds.We utilized a slightly
changedTollens technicluefor the dosificationof glucuronicacid in the
urine,basedon the reactionof this acid with naphthoresorcine
in an acid
medium.To 5 cc. of urinc,0.5 cc. of a I 7o solutionin alcoholofnaphthoresorcine(1.3 dioxynaphthalene)
and 5 cc. of concentrated
hydrochloricacid
were added. The mixture was boiled for onc minute, allowed to stand
for another five minutes,thcn cooled. preferablyin an ice water bath.
Whcn cold, a nrixtureof.90c/oethcr and l0o/o alcoholwas added,agitated,
and the blue-violetcolor of the cther-alcoholmcasured,using a spectrophotometcr.The valucsobtainedin diflerentsubjectshaveshowna definite
increasein the amountof glucuronicacid in the urincscontainingperoxides,
indicatinga probablcrclationbetwccnthcm. Basedon this correlation,we
one of the roles of glucuronicacid in the organism,that of
investigated
detoxifyingagent.
Glucuronicacid could be consideredto result, at least in part, from
oxidation of glucose.Usually oxygen intervenesin glucosemetabolism

642

REsEARcH rN pHysropATHoLocy

t (232), correspondingto the fermentaonly after the desmolyseprocesses

tive phase has progressedto the appearanceof pyruvic-acid. Glucuronic
acid could be consideredto appearfrom a more direct fixation of oxygen
to the glucosemolecule.This fixation has to take place upon c6 in oraer
to lead to the appearanceof glucuronic acid. Theoretically, the oxygen
fixation might be expectedto occur at cr in view of the aidehydegroup
presentat that carbon. This takes place in vitro. It would lead to the ap
pearanceof gluconic acid. However, il a phosphoric or other radical is
bound to cr in vivo, fixation of the oxygen at this carbon is prevented.
Oxygenattachesitself then at C6 which is the next most reactivecarbon in
the molecule.This reactivity at Cs is seenwhen oxidation in vitro is continued beyond gluconic acid, leadingto saccharicacid, a bicarboxylic acid
with one carboxyl at C1 and anotherat Co.
Glucuronic acid intervenesin the physiologicaldefenseprocessesby
combiningwith certain noxious productsand helping to eliminate them in
non-toxicforms. The resultingcompoundbetweenglucuronicacid radicals
and various substances-and for sulfuric acid radicals as well-has been
called "conjugation," the substances
being sulfo- or glucurono conjugated
compounds.Becauseof the specialattentiongiven in this publicationto
the conjugationof the double bonds, we will use the term "coupled" for
this bond to sulfuric or glucuronicacid.
A certain parallelismexistsand has always been emphasizedbetween
the detoxifying and eliminating function exerted by the sulfuric and glucuronicradicals.Not only the two derivativesappearin the urine, but it is
often noted that glucuronicacid interveneswhen large amounts of certain
substances,such as menthol or phenol, are presentand the sulfuric acid
radicalsare not in sufficientquantity to insure detoxificationand elimination. By administeringmineral sulfatesto thc subject, the proportion of
sulfo{erivatives is seen to increase.This parallelism appears especially
interestingwhen we recognizethat sulfuric acid representsthe final stage
of the oxidation of sulfur introducedinto the organismin combinationsin
which it entersas a bivalentnegativeelement.Both sulfuric and glucuronic
acid result from oxidative processes,
one involving the thiol group and the
other, glucose.
While glucuronic acid often substitutesfor the sulfuric radical in the
excretedsubstances,
qualitativedifferencesexist.For example,phenol as
well as indoxyl is bound to the sulfuronicradical,while the higher alcohols
and especiallythe cyclic oxyacidsdo not combinewith this acid. For many
substances
the parallelismthat existsin the bond to sulfuric and glucuronic
radicalsextendsonly up to a certain point, after which the amount of the
sulfuricesterno longer increases.
This fact has raisedthe problem of the
actionof thesetwo radicalsand the differences
betrveenthcm. and can be
+ T h e t e r m " d e s m o l y s e . "w i d e l y u s e d b y s o m e a u t h o r s a b r o a d . c o r r e s p o n d st o
the changes occurring in metabolites under the action of hydrolases, whicb, when
completed, result in a splitting apart of the moleculesand the liberation of the energy
they possess.

NorEs

643

understood only by consideringthe substancesthat are bound by these

acids and eliminatedas sulfo- or glucurono-coupled
derivatives.
We haveseenin Note l9 that,with very few exceptions,
suchas benzoic
acid, almost all the substancesexcretedor bound to glucuronic acid have
an OH as polar group, However, many have more than one polar group.
The bonding with the sulfuric radical, as it occurs in the organism,
follows a relativelysimplepattern.(Fig.258a) In a first step,the bonding
of one acid function of the sulfuric radical to the substanceproduces an
acid sulfuric ester. This is further changedby combination with a metal,
usually potassium,which producesa highly hydrosolublesalt and representsan excrementalderivative.

,-0H- [ster

s0,f
'\

sl

Salt

S u l f u r i cA c i d
[)erivatives
(a)

CH
0 - Glycoside
tJH
CH
(JH
CH
\
tH [st ers
CH
CH
CH
c00l-l SaI t

G l u c u r o n iAc c i d
Derivatives
(b)
F t c . 2 5 8 a . T h e b o n d o f s u l f u r i c a c i d i n t h e o r g a n i s mc a n r e s u l t i n a n e s t e r a n d a s a l t .
F I c . 2 5 8 b . T h e b o n d o f g l u c u r o n i ca c i d c a n r e s u l t i n a g l u c o s i d e ,a n e s t e r o r a s a l t
combination.

The binding is more complicatedfor glucuronicacid. Glucuronic acid

does not realizethe bonds,sinceit is eliminatedas such if administered.
The bond occurs at the aldehydegroup of glucose,forming a glucoside,
which is passedfurther in glucuronicacid. As is seenin Figure 258b, glucosecan realizedifterentbonds.It can bind a radical either to the adlehyde
group at Ct to form a glucoside,or to its alcoholic hydroxyls to form an
ester.The glucuronicacid can bind a metal to its carboxylto form a salt.
The possibilityof realizinga glucosidiccouplingat Cr or an estericat C2has
beenrevealedby Quick. (233) The ability to realizeconcomitantlya multiple coupling,for which glucuronicacid seemsto be highly suitablethrough
its multiple and various functions,appearsmore interesting,The study of
eliminationof differentoxybenzoicacids shows that it is bound to gly-

644

R E S E A R c Hr N p H y s r o p A T H o L o o y

cocoll,(FiS 259a) while the para isomeris eliminatedcoupledby the gluc u r o n i cr a d i c a l .( F i g . 2 5 9 b )

The difterencebetweensulfuric and glucuronicacid thus appearsto be
related to double coupling especiallyif the secondfunction is acid. The
presenceof two oppositepolar groupswill thus preventthe bond to sulfuric
acid and favor the bond with the glucuronicradical. organic oxyacids are
thus not bound by sulfuric radical as long as their carboxylsare free. The
amides,such as salicylamides-and, the esterssuch as methyl salicylatesare coupled by the sulfuric radical, while the acids are not. It is through
these multiple coupling possibilities-and, in addition the possibility of
forming a salt especiallywith potassium-thatglucuronicacid corresponds
to a broader activity as a detoxifying and eliminating agent than sulfuric
acid.

C00H HrN...

hr--Hocc)"'

Ortho
(a)

K- H00C
001-l-H0C
I

H0c
I
H0c
I

fH\,il!
Para
(b)

F t o . 2 5 9 . T h e o r t h o - o x y b e n z o iacc i d ( s a l i c y l i ca c i d ) i s b o u n d i n t h e o r g a n i s m r o
g l y c o c o l l ,( a ) w h i l e t h e p a r a - i s t r m cirr c o u p l e db y t h e g l u c u r o n i cr a d i c a l . ( b ) .

The capacity to utilize glucuronic acid for excretion of noxious sub.

stancesvaries widely between species.Benzoic acid has been shown to
appear as a glucuronic derivative in animals by Csouka, Brakefield, and
Schmidt (234), but never in humans, a fact that is not confirmed by
Quick. (235) Rabbits eliminatethe tertiary alcoholsas glucuronic derivatives more completelythan dogs.While thyroidectomyin rabbits decreases
the synthesisof camphoroglucuronicacid (236), the administration of
thyroid extract increasesit.
Chapter 6, Note 21. ParaplegiaInduced by Cholesterol
Male and female rats, weighing approximately250 grams, were injected with 5 cc. of 70Vo cholesterolonly partially dissolvedin a mixture
of 3 parts oil and I part ether.The next day paraplegiawith some ulcera-

NorEs

645

tion of the hind legs was seen. Ulceration becameaccentuatedin the

following days. Curiously enough,this occurredonly in the females.In
repeatedexperiments
in rats,and alsoin rabbits,guineapigsand evenmice,
the samesex differentiationhas beenobserved.Most of the femaleanimals
died from retentionof urine as a principalcomplicationof the paraplegia,
To investigatethe nature of this sex diffcrence,groups of males and
femaleswere castratedand spayed.then testedwith the samecholesterol
injection at various times from a few days to four months afterward.Castrationhad no influencein male rats;nor did the spayingof fenralesreduce
the incidenceof paraplegia.The administrationof 5 mgm. o[ testosterone
daily for ten daysto femalerats, spayedor unspayed,also did not prevent
the appearance
of paraplegiaafter the injectionof cholesterolin ether-oil.
The administrationof Vz mgm.of stilbestroldaily for l0 days to males,
castratedor not, did not break down their resistanceto the cholesterol
injection.However, the administrationof thc insaponifiablefraction of
placentaor of the total body of rats, in dosesof 2 cc. of a 5olosolution in
oil daily for ten days, madc the malesrespondwith paraplegiato the inject i o n o f c h o l e s t e r oiln e t h e r - o i ls o l u t i o n .T h e a d m i n i s t r a t i o n
to females.
spayedor unspayed,of a l07o solutionin oil of the acid-lipidfraction of
the sameorigin in dosesof 2 cc. daily for tcn dayspreventedthis effect.
Chapter 6, Note 22. Adipose Cells and Sulfur Nlustard
In collaborationwith the late Prof. R. Leroux, Professorof Pathology of the Facultyof Medicinein Paris,we studiedthe influenceexerted
by sulfur mustardappliedon the skin. The followingtechniquewas used:
was depositedon the cxternalside of the
One drop of the pure substance
ear pavilionof adult white rats. left in placefor l0 minutes,and wiped off.
A part of their ear pavilion was cxcisedin a V form. The fragmentso
obtainedwas immediatelyprocessedthrough the specialtechniqueused
f o r t h e c x t e m p o r a n c o uesx a m i n a t i o o
n f o p e r a t o r yb i o p s i e sA. l o n g i t u d i n a l
sectionof the pavilionwls madein the frozenfreshmaterial,and this surface treatedfirst with formalin and then staincdwith scarletred or black
Sudanand hematein.The sectionwas kept in water coveredwith a glass
c o v e r a n d i m m c d i a t c l ye x a m i n c du n d e r i n c i d e n tl i g h t , u s i n g t h e Z c i s s
"Ultropak" dispositive.A water immersionobjectivewas used for higher
amplification.From thc same material,fixed in formaldehydeat loo/o,
frozr,nand paraffinimbeddedsectionswere also obtained.
In nontreatedcontrols, cxcept for the small fatty droplets in the
the only fatty cellsfound wcre at the baseof the
cartilagecellsthemselves,
pavilion.In the mustardtreatedanimals,two or threelayersof adiposecells
were seento appcar beneaththe treatedskin, near the cartilage,around
2 0 m i n u t e sa f t e r t h e a p p l i c a t i o no f t h e s u l f u r m u s t a r d .
. r i o u s l ye n o u g h .t h i s p h e n o m e n o n
was
I n o u r g r o u p o f e x p e r i n r e n t cs u
seento occur only in the treatedfemale rats and not in the males.This
differencein the rcsponsebetweenmalesand fcmalcswas not influenced
either by castrationor by treatmentof the animalswith sex hormones.It

646

'

R E S E A R c Hr N P H Y s t o P A T H o L o c y

wasinduccdin maleshowever,by the administration

of unsaponifiablc
fractionsobtainedfrom the total body of rats,or from human or cow placenta.
a n d a d m i n i s t e r eidn d o s e so f I c c . o f a l 0 % s o l u t i o ni n o i l f o r a t l c a s t
o n e w e e k p r i o r t o t h e a p p l i c a t i o no f s u l f u r m u s t a r d .I n f e n r a l e st,h e a p
pearanceo[ adiposecells after the sulfur mustard applicationswas seen
to be prevented,if the animalswere treateddaily for 10 days prior to
this applicationwith 2 cc. of a l}Vo solutionin oil of lipidic acid fraction
of human or cow placentaor of cod liver oil fatty acids.
Chaptcr 6, Note 23. Fatty Acids and Old Tetrah)'mena
Culturesof differentagesof tetrahymena
pyriformisin proteosepeptone
medium,were analyzedfor thc fatty acids present.The richnessin fatty
acidswas seento increasewith the ageof the culture,the old culturesbeing
the richest.
Chapter 6, Note 2a. Lipids and Old Age
In the changesrelatedto old age, we have recognizeda relationship
betweenthe amount of insaponifiable
fractionsand fatty acids presentat
different levcls of the organization.A certain degrce of opposite changes
in two successive
hierarchiclevelscan explain certain peculiaritiesfound
in the variationsof sterolsand fatty acids in old age.
In old rats we could establishthat changesoccur in two oppositcdirections betweenthc cells and the metazoiccompartment.Serum cholesterol
amountsincrease,as an increaseof fatty acids and particularly polyunsaturatedmembersoccursin the cells.The contrastbetweenthe two compartmentsbecomcsprogressivelymore evident with old age. We tried to
apply this data to the study of the relationshipbetweenthe samecompartmentsin humans.Insteadof other cells,we analyzedred blood cells. In
rats, rabbits and humans,the cholesterolin serum increasesprogressively
with agebut does not do so in the red cells.The amount of fatty acids in
red cells is relativelyincreasedwith age at the same time as cholesterol
content is decrcased.The comparisonbetwecnthe cholesterolof serum
and cholesterolof the red cells as well as fatty acid content of red cells
seemsto furnish informationrelatedto thc progression
of abnormalitiesof
preliminary
research,
however,
needsmore confirmation.
old age.This
C h a p t c r6 , N o t e 2 5 . S u r f a c eT e n s i o no f U r i n e i n O l d A g e ( 2 3 7 )
During the courseof a study of the biologicalchangesin old age, the
was found to be considerably
substances
renal excretionof surface-active
below the levels seen in younger age groups. Thcse findings were considered important since ( I ) they were observedin individualswith no evidenceof significantrcnal impairment,(2) the samedecreasewas obsened
in all subjectsexamined,and (3) informationon this subjectis lacking in
the absenceo[ a suitableanalvticmethod.

The capitlary devicc has made it possiblc to determi;;::.-"r:;

within a few seconds.as part of routine urinalysis,without need for complicated apparatusor involved formulas.
A group af.?3 inmatcs of an old agc institution wcrc studied.Ages of
these 14 men and 9 women rangedfrom 70 to 8T years. All appearedin
good general condition and showed no evidenceof significantrenal pa,
thologr. Morning urine specimenswere obtaincd from each patient for
severalweeks. These werc examined within several hours after voiding.

Femolc
Male

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UR,fVARY SURFAC TENSION (Oynat/ cm )

F t o . 2 6 0 . D i s l r i h u t i o n o f u r i n a r l ' s u r f a c e l e n s i o n v a l u c si n t h e g r < - r uo
pf o l d p e o p l c
studied.

Surface tension values for the entire group avcragcd70.9 dynes,/cm.

T h e a v e r a g ev a l u e f o r t h c w o m e n u ' a s 7 1 , 5 a n d . f o r t h e m a l e s , 7 0 . 4
dynes/cm. When surfacetensionvaiues*'ere dctermincdin serial samplcs
from individual subjccts.variationsof only 2 to 6 dynes,,'cnt.
were fclund.
None of the samplesfrom the women showed a surfacetension below
69 dynestcm,All but 3 of the specimens
from the men had surfacetension
v a l u e so f 6 8 d y n c s ' c n l o. r h i g h e r (. F i g . 2 6 0 l S p c c i l i cg r u v i t vv a r i c df r o n r
1 . 0 0 6t o 1 . 0 3 0 ,w i t h a n a v e r a g eo f l . 0 l ? f o r t h e g r o u p .
In control groups,thc averagesurfacetcnsionis 67 to 6lt dynes/cm.
Diurnal and day-to-dayvariationsof "5to I I dynesT'cm..
rvith valuesranging betwecn 72 to 62 dynes,/cm.,arc found characteristic:.rlly
in healthy
subjects.
It is evident that thc pattern of excretion of surface-activematerial
in old age is quite differcnt. This is indicatedby ( t ) thc high averagicsurface tension value clf 7l dynes/cm. for the group. which is at least 3
dynes/cm.highcr than for the controls; (2) the limited day-to-day range
of variations,which in no casewas greaterthan 6 dynes,'cm.;and (3) the
striking disparity in the distributionof valuesin healthy young adults and
in old age. (fig. ?6// Eighty-eightpercent of thc urinc specinrensfrom

648

x E S E A R C Hr N P H Y S r o p A r H o L o c y

old individualshad surfacetensionvaluesof 70 dynes/cm. or higher, and

only I percent were below 66 dynes/cm. In contrast, 23 percent of the
specimensfrom the control group had a surfacetension of 70 dynes/cm.
or higher, and 23 percentwere below 66 dynes/cm.These findingsindicate
that the amount of surface-activesubstancesexcretedin the urine of old
people of 70 to 87 years is greatly reduced as compared with that in
healthy young adults. The fact that specificgravity values for the old age

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TOord 69-66 65ond

over
under
UR,NARYSURFACE
TENSION( Dynes
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696

65oN
under

Fro. 261. Distribution of surfacetension values in healthy adults and in old age.
s h o w st h e h i g h p r o p o r l i o n o f v a l u e so f 7 0 d y n c / c m a n d a b o v e f o r t h e l a s t g r o u p .

group were not significantlydifferentfrom the controlsindicatesthat this

changecannotbe explainedon the basisof a failureof the kidneysto concentrateby reabsorbingwater.
The average surface tension value for the females was I dyne/cm.
higherthan for males.Thc samesexdifferencehasbeennoted in studiesof
healthyyoung adults and in groupsof patientswith a variety of diseases.
It is of interestto find the differencepersistingin theseold people.
In the light of current conceptsof the possiblerole of surface-active
"colloidal" substancesin the urine, it is interestingto speculateon the
possiblesignificance
of thc observcddecreased
excretionin terms of renal
Various authorshave contendedthat the crystalloidsof
physiopathology.
the urine are maintainedin solution by the colloids, which prevent ag-

NorEs

649

glomerationand precipitationof salts.(238, 239) Somesuch mechanism

is undoubtedlypresentin the complex physiologicalsolution finally excretedby the kidneys,sincemost of the urinarysaltsare presentin proportions far exceedingtheir ordinary limits of solubilityin aqucoussolution.
Low levelsof colloidal activity and high surfacetensionvalueshave recentlybeencorrelatedwith an increasedtendencyto stoneformationin the
urinary tract. (240) lt is possiblethat this reductionin surface-active
substancesin the urine of old people,combincdwith the obstructionat the
vesicalneckdue to prostatichypertrophy,nlay lead to retentionof precipitated matcrial in the bladder, accountingfor the increasedincidenceof
vesicalcalculi during the latcr decades.In women, incidenceof bladder
stoneis relativelylow, even thoughthe quantityof excretedsurface-active
s u b s t a n c ei s s m a l l ,b c c a u s et h e r ei s n o a n a t o m i c aol b s t r u c t i o nt o u r i n a r y
flow.
Administrationof hyaluronidaseraises the level of colloid activity
(241) and lowersthe surfacetensionof thc urine to a slight degree.This
enrymehasbcenutilizedin thc treatmcntof chronicrecurrentstoneformers
(242) The reducedexcretionof tensio-active
with apparenteffectiveness.
substances
in the urine may be relatcdto the decreased
enrymaticactivity
within the tissues.
Chapter6, Note 26. EnvironmentalInfuences
I n m o s to f t h e e x p e r i m e n ti sn a n i m a l st,h e a n a l y t i c adl a t af o l l o w e do v e r
a c e r t a i nl e n g t ho f t i m e s h o wv a r i a t i o n sw h i c hc a n n o tb e e x p l a i n e db y t h e
e x p e r i m e n ti t s e l f . A d i r e c t r c l a t i o n s h i po f s u c h v a r i a t i o n st o c h a n g e s
occurringin the environmentcould be seen in the following experiment.
S i x g r o u p so f 2 0 f c m a l ew h i t c W i s t a rr a t s e a c h ,w e r c i n j e c t e do n t h e
s a m ed a y s ,e a c h g r o u p w i t h a d i f f c r c n ta g c n t .O n e g r o u p o f a n i m a l sr e ceivedneutraloil as control while the othersreceiveddifferentfatty acid
preparations.The urinary surface tension was measureddaily at approximatelythe same time of day for all the animals.The averagevalue
of thcsedaily datawasobtaincdfor cachgroupand uscdto tracethe respective curves.
By comparingthcsccurvcs,two characterswere recognized.One was
from one curve to another,and consequently,
seento concerndifferences
would be consideredas rcsultingfrom dilTcrenccs
in the dircct cffectof the
medicationsused.The other group of changesconcerningvariationsfrom
one day to the other, was sccnto cxist in all thc cLlrves,the curveshaving
t h u s p a r a l l e lv a r i a t i o n s(. F i S . 2 6 2 ) T h e s e v a r i a t i o n sc, o m m o n t o a l l t h e
c u r v e s ,w e r e c o n s i d e r c di n d u c e db y a g c n c r a li n f l u e n c eT. h e a n a l y s i so f
thesecurvesshowsthat the first kind of changes,
relatedto the agentsused,
concernsdifferencesin the levels of the curves themselves,when comp a r e dw i t h t h a to f t h c c o n t r o l T
. r e a t m c nw
t i t h s t e a r i ca c i dd o e sn o t i n f l u e n c e
t h e l e v e l , a n d t r e a t n r e nrtv i t h o l e i c a c i d h a s o n l y a s l i g h t i n f l u e n c e A
.
manifestchangeis sccn for thc othcr curvcs.Thc. urinary surfacevalues
correspondto lowestvaluesfor linoleicacid (d) and for cod liver oil fatty

650

RESEARCH IN

PHYSIOPATHOLOCY

acid (e). Some less marked difterencesfrom the control curve is seen for
the fatty acid preparationobtainedfrom cow spleen(f ).
Independentof these level differences,all the curves of the controls
as well as of the treatedgroupsshow parallel daily changes.An exception
I

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treated with different fatty acid preparations(l cc of lo/o in oil daily). Thc parallel
changesin the curves, except for oleic acid when the variations are opposit+indicate a common external influence.The differencesin the relationship of the curvs lo
the average value line, corresponds to the direct influence exerted by tbc atsnts.

NorEs

651

is seenfor the curve for oleic acid, which showsoppositevariations,we

do not havc an explanationfor this discrepancy.
The parallelchangewould
indicatethe interventionof a common factor, independentof the experiment itself.By rclatingthcsechangcsto thosctaking placc in thc environment,the commonvariationsin the curvescould be recognized
to followi n a n o p p o 5 i l g5 g n 5 s - t h o s co f t h c c n v i r o n m e n t at el m p c r a t u r c .
It shouldalsobe notedthat this environmental
influenceis progressively
more accentuatedfor thc curves where the level of the surfacetension
is lowered as a result of the agent administered.This correlationwould
suggcstthc possibilitythat the action exertedby the cnvironmentwould
take place largely through changesin the interventionof the fatty acids
of the organismsthemselves.
Chapter 6, Note 27. Environmental Influences Upon Urinary
SurfaceTension
The morning urinary surfacetensionwas measuredin four groupsof
4o rats each:one group of 20 males,one of 20 femalesof the white wistar
strain,one group of 20 malesand one of 20 femalesof a black "hooded"
strain. From the data obtainedon each group, the averagevalue was
calculatedand its respectivecurve traced.The four curveswere seen to
b e p a r a l l e ls, u g g e s t i nt gh a t t h e v a r i a t i o n n
s o t e dr e s u l tf r o m t h e i n t e r v e n t i o n
of a commonexternalfactor.We soughtthis factorin the changesoccurring
i n t h e e n v i r o n m e n tF. o r t h i s r e a s o n w
, e c o m p a r c dt h c v a r i a t i o n sp r . t e n l
in the surfacetensioncurvcs with the meteorological
data, furnishedby
the U. s. weather Bureau,corresponding
to the time of this experiment.
Such a relationshipwas seen to only partially
parallel the barometric
-th!
changes,but appeired more closelyrcliitcd to
tempcraturechanges.
The observedrelationshiphowever,is inverse,that is, for higher environmentaltempcraturcs
thc surfacctensionvaluesare low whereasfor lower
environmentaltemperatures,
the surfacetensionvalues are high, as seen
i n F i g , 2 2 3 . ( P a g e5 8 6 )
This corrclationappearsstill more intercstingwhen it is comparedto
that inducedby keepinganimalsat a constanttemperature,
as in an incubator or in a rcfrigerator.The cffect of such an induccd temperatureis
oppositeto that causedby the environment.The high temperatureof the
i n c u b a t o ri n d u c e sp r o g r c s s i v c lhy i g h c r s u r f a c ct e n s i o nv a l u e sw h i l e t h e
l o w t e m p e r a t u roef t h e r e f r i g e r a t olro w e r st h e s u r f a c et e n s i o n ,a t l e a s ta t
f i r s t .F i g . 2 2 4 s h o w st h c a v c r a g cv a l u e o f t h e u r i n a r y s u r f a c et e n s i o ni n
c o n t r o l l e da n i m a l sw
, h i l e F i g . 2 2 5 t h a t o f t h e a n i m a l sk e p t i n a n i n c u b a t o r
a t a t e m p e r a t u roef 3 7 ' C a n d i n a r e f r i g e r a t oart 8 " C . ( P a g e5 8 8 )
We tried to explainthis discordantinfluencebetweenthe inducedand
n a t u r a l t e m p e r a t u r c st ,h r o u g h t h e f u n d a m e n t acl h a r a c t e r so f t h e s et w o
factors.In the influenceexertedby the environment,temperaturewith its
variations,reprcsentsa factor which has acted upon organismswith the
samerhythmic charactersfor many millionsof years,while in the experim e n t s ,t h e c o n s t a n c yo f t h e t e m p e r a t u r e
r e p r e s e n t ist s m a i n c h a r a c t e r .

652

, , ' R E S E A R c Hr N p H y s r o p A T H o L o c y

The influenceexertedby the rhythmic environmentalchangesin air

temperature
is reflectedin the parallelbody tempcrature.The organismstill
tries to control this influenceas exerted upon the lower levels.This is
seenin rats in the oppositerhythmicchangcsof the urinary surfacetension
values.The organismsappearsufficientlysensitiveto changesin temperatureoccurringin the environment.
Thc body responses
opposethesechanges
as shown in the variationsin urinary surface tension. This rhythmic
response,as well as that opposingthe variationsin the environmental

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F t c . 2 6 3 . R e l a t i o n s h i pb e t r v e e nt h e c u r v e so f s e r u m a n d t o t a l b l o o d p o t a s s i u m o f a
g r o u p o f 2 0 s u b j e c t sa n d t h e a t m o s p h e r i ct e m p e r a t u r e .h u m i d i t y a n d b a r o m e t r i c
p r e s s u r eP
. a r a l l e l v a r i a t i o n sa r e s e e nb e t w c e nt h e c u r v e so f t o t a l b l o o d p o t a s s i u ma n d
o f b a r o m e t r i c p r e s s u r e .A n i n v e r s er e l a t i o n s h i pi s s e e n w i t h t h e c u r v e o f h u m i d i t y , .
a n d o n l y p a r t i a l r e l a t i o n s h i pt o t h e t e m p e r a l u r e ,

NorEs

653

temperature,
are broken when a continuouslyunchangedhigh or low temp e r a t u r ei s a p p l i e d .W i t h t h e r e a c t i o n arle s p o n s e x h a u s t e d
a f t e ra c e r t a i n
t i m e . t h e i n f l u e n c e x e r t e da p p e a r sa d i r e c to n e , t h e h i g h t e m p e r a t u r e
ind u c i n ga h i g h u r i n a r ys u r f a c et e n s i o na n d t h e l o w t e m p e r a t u r ae l o w o n e .
However, a differenceis seen betweenthe influenceexerted by the
h i g h a n d l o w t e m p e r a t u r eA. f t e r s o m e t i m e , t h e a n i m a l sk e p t i n a r e frigeratorusuallyrecoverthe capacityto fightthe persistently
low temperat u r e ,a n d t h e s u r f a c et e n s i o nr e t u r n st o n o r m a lv a l u e s T
. h i s d o e sn o t o c c u r
f o r t h e p e r s i s t i n g lhyi g h t e m p e r a t u r eT.h e a n i m a l sd i e w i t h a p r o g r e s s i v e l y
h i c h s u r f a c et e n s i o na s i f t h e d e f e n s i vree s p o n s see e nf o r l o w t e m p e r a t u r e s
w o u l d n o t i n t e r v e n ef o r h i g h t e m p e r a t u r e s .
T h i s d i f f e r e n c ien t h c r e s p o n soc f a n a n i m a lt o w a r da p e r s i s t e n t lhyi g h
o r l o w t e m p e r a t u rcea n b e e x p l a i n e db y t h e f u n d a m e n t adl i f f e r e n c ew h i i h
existsbetweenthesetwo factorsfrom the point of view of homo- or heterot r o p y . w h i l e t h c l o w t e m p c r a t u r eh a s a h o m o t r o p i cc h a r a c t e r t, h e h i g h
t e m p e r a t u rhea sa h e t e r o t r o p iocn e .T h e b o d i c sa r e b a s i c a l l ym o r c p r e p a r e d
t o r e a c ts u c c e s s f u ltl oy w a r dh o m o t r o p i ci n f l u e n c eass t h e y h a v ed o n e i t f o r
m i l l i o n so f y e a r si n t h c p a s t ,t h a n t o a n e n t i r c l yu n u s u a lh e t e r o t r o p i icn flucnce.
Chapter 6, Note 28. Barometric Influence
F o r s e v e r ayl e a r s .w e s t u d i e dt h e p o t a s s i u m
c o n t e n to f b l o o d r e d c e l l s
in subjectswith various abnormal conditions,usually following it daily
for the samepatientfor weeksor months.Besidesother informationswhich
this studyfurnishedand which are discussed
above,we want to emphasize
now the relationshipwith barometricpressurewhich we were able to establishduring repeatedanalyseson the samegroup of subjects.we were
impressed
b y t h e p a r a l l e lv a r i a t i o n ss e e ni n t h e a m o u n t o f p o t a s s i u mi n
t h e t o t a l b l o o do f d i f l e r e n p
t a t i e n t so n d i f f e r e ndt a y sr e g a r d l e sosf m e d i c a t i o n . T h e v a r i a t i o n sc o u l d b e c o r r e l a t e dw i t h c h a n g e si n t h c b a r o m e t r i c
p r c s s u r c sW
. i t h l o w e r p r c s s u r c sa, d c c r e a s ci n t h e a m o u n t o f p o t a s s i u m
i n t o t a l b l o o d ( F i g . 2 6 3 1 w a sn o t c d .L e s sm a n i f e sct h a n g e sw e r e s e e nw i t h
t h c o p p o s i t cv a r i a t i o n si n a t n t o s p h e r ihcu n r i d i t y .
Chapter 6. Note 29. Age, Lipoids and Tumor Transplants
The influenceof ageof a host upon the differentnranifestations
through
the interventionof sterolsand lipoacidscan be scenin the followingexperiments.
Transplantsfrom the samewalker tumor weregraftedat the sametime
i n a n i m a l so f d i f f e r e n ta g e s ,s u c h a s n e w b o r n ,w e a n l i n g sy, o u n g a n i m a l s ,
adultsand aged.The differencebetweentransplants
was evidenteven from
b u t w a s s t i l l m o r ee n h a n c e d
thebeginning
i f f u r t h e rt r a n s p l a n tw
s e r em a d e
i n a n i m a l su n d e r s i m i l a r c o n d i t i o n sA
. ftcr one transplana
t nd especially
after severaltransplants,the followingchangcscould be seen.In the newborn, the tumor took on the aspectof an hemangiomatous
lesion,Therewas

654

nESE^RcH tN pHyslopATHoLocy

no massivetumor and the amount of blood present gave the tumor the
appearance
of a pieceof liver. In weanlings,the characterwas oppositc.
Massive tumors without necrosisand with the aspect of fish meat were
seen.In youngstersthe samecharacterwas obtained.In adult animals,the
tumor had a large portion of necrosiswith predominanceof hemorrhagic
fluid. In very old animals,this characterwas still more accentuated,and
the tumor showedbig cavitieswith hemorrhagicfluid and very little tumor
substancebetween
Similar changeswere obtainedby changingthe site of the graft. By
graftinga portion of the sametumor,'intram-usJularly,
we obtaineda massive whitish,nonnecrotictumor, while subcutaneous
injectioneven in the
samcanimal led to the appearanceof a tumor with multiple nccrotic areas
and caviticsfilled with fluid.
We tried to correlatethesechangeswith the nature of differentlipids
predominantat diflcrent ages.The administrationof fatty acids tcnds to
promotenccrosis,edema,and formationof cavitiesfilled with fluid. while
administrationof insaponifiablefractions,espcciallyfrom placentaor liver,
tendsto produce a type of tumor with whitish, nonnecroticmasses.
Chapter 6, Note 30. Temperature, Lipids and Viral Infection
The relationshipbetweenepidemicsand seasonsis a well-established
concept.In an attemptto explainthis correlation,we considered,as one of
the interveningfactors,the seasonal
changcsin lipids,in view of the influence excrtedby thc two oppositegroupsof lipids upon the receptivity and
manifestations
of infectiousdiseasc.
we
As
havc mentioned,viral infectionsare especiallyinfluencedby the
predominanceof one or the other group of lipids in the body. Among
other factors, temperaturechangeswere found related to this predominance.A relativelydirect correlationwas found for polio, for instance.
The appearanceof neurologicalsymptomssuch as paralysiswas seen to
increaseon days with high tempcrature.
Sucha corrclationcould bc establishedexpcrimentally.Among micc injectedsubcutaneously
with smallpox
v a c c i n ea n d k e p t i n a n i n c u b a t o ra t 3 5 ' C , t h e i n c i d e n c eo f e n c e p h a l i t i s
rose to more than 90% as comparedto 10Vo in controlskept in an airconditionedroom. This correlationrvasfurther explainedby the predominance of sterols in the organism undcr the influenceexerted by high
'l'his
predominance
wasfurther seento inducea higher receptempcrature.
viral
infection,
organism
to
the
and a changein the virus virulence
tivity of
itself, both of which increasewith temperaturcand with the richnessof
sterols.
Chapter6, Note 3l . Youth and Vinrses
g s p e c ot f t h e c h a n g e si n d u c c di n v i r u s e sb y l i p o i d s
A n o t h e ri n t e r c s t i n a
with an alcoholicpolar group,especiallysterols,is the relationshipto age
of the host.Thc greatamountof insaponifiable
fractionpresentin youth-

NOTES

//

655

which appearsto be capable,in itself, of increasingvirus virulence-has

been discussed
previously.
In an experiment,smallpox vaccine was inoculatedin groups of very
yomg, adult and old rabbits.The young animalsreactedmuch more intensivelythan did the adults,with conuuentpustulae;in the old animals,
only a few small pustulaeappearcd.After severalpassagcs
in animalsof
the sameage, we testedthe virus on mouscskin and found the virulence
increasedby each passagein yclungrabbits,on the other hand, virulence
was decreased
by passage
in old animals,becomingnegativewith the fourth
passage.
After the third passage,virus obtainedfrom young rabbits induced
a strongresponsein mice, while no pustulaewere obtainedwith virus from
old animals.The latter was still able to induce some responsein young
mice, but only few small pustulae.The virus obtainedfrom the third passage in young rabbits induced a frank responseeven in old mice. This
indicatesthat oppositechangesin virus virulenceare induced simply by
passagethroughyoung and old animals.
This experimcntappearsespeciallyinterestingin connectionwith childhood colds.It is known that childrencatch colds frequentlyand rhat this
susceptibilitydisappearsas thcy approachpuberty.From a pathogenicpoint
of view, it is an interestingfact, however,that older siblings,parents,and
even grandparents
are still infectedif the cold comesfrom a child in the
family. The virus itself seemsto be changedby passagethrough the child
so that it becomesvirulentnot only for teenagers
and parentsbut for older
people,all of whom previouslymay havc beenfree of coldsfor years.
Chaptcr 6, Notc 32. Changesin the Vimses Induced by Lipids
The effectof lipids upon virusesseemsnot to consistcxclusivelyin an
alterationof the host'sresponsebut also includesa changein the viruses'
virulence.We have noted previouslythe big differencein the responseof
the skin of an animal when, prior to inoculationwith smallpoxvirus, a
lipoid with a positiveor negativepolar group was injectedsubcutaneously.
The lipoid with a positivepolar group inducesan exaggerated
response;
the lipoid with a negativepolar group,a reducedone.
We used viruscsobtainedfrom both types of lesionsfor inoculations
in new animals.Virusesfrom lesionsin which the responsehad been exaggeratedproduced again an exaggeratedresponse,while those obtained
Thc eftect,in one or the
from a small pustuleinduceda reducedresponse.
passages
on skin pretreatedby
other direction,was enhancedby successive
were confirmedin
in
virulence
injcctions.
The
changes
also
subcutaneous
mice tests.
The importanceof this experimentlies in the fact that, by treatmentof
the host with membersof one or the other group of lipoids,we can obtain
a desiredincreaseor decreasein virus virulence.In severalother experiments,we found the method applicableto other viruses,generallymaking
the virus more or lessvirulent evenfor intracerebralinoculation.Interesting
changesare producedby the polyunsaturatedfatty acids and the alcohols

656

nESEARcHrN pHysropATHoLocy

obtainedfrom theseacidsby changingthe carboxyl into a primary alcohol

throughtreatmentwith lithium aluminumhydride.Especiallyeffectiveinfluencesare obtained using acid lipidic fractions of refractive speciesin
order to reducethe virulence,or insaponifiablefractionsof especiallysensitive speciesor of chicken embryosto enhancevirulence.Treatment of the
host with lipoacid preparations,repeatedfor severalpassagesof the virus,
has reducedvirulenceto the point where the virus no longer is pathogenic.
This treatment seems to provide a method for reducing virulence and
obtainingnonpathogeniclive virus vaccines.
Chapter6, Note 33. Microbes,Phagesand Lipids
We have investigatedthe influenceof various insaponifiablefractions
upon the relationshipbetweenmicrobesand bacteriophages.
Coli bacilli, of
grown in
a strain which has shown considerable
phage,
were
to
resistance
broth to which insaponifiablefractionsfrom different sources-such as human placenta,eggs,butter-were added in colloidal suspensions.
While in
thesemedia the microbesshoweda higher susceptibilityto being attacked
by phagethan the controls grown in simple broth, only very few microbes
if any remained resistant.In another experiment,this same strain of coli
resistantto phage was grown in successivepassagesin broth containing
insaponifiablefractions.After severalsuchpassages,
the microbe was grown
in simplebroth and its sensitivityto phagewas testedin this medium. While
the phage used appearedunable to attack the microbes of the unreated
cultures,lysis was manifestin the treatedcoli. Thus, insaponifiablefractions
increasedthe sensitivityof the microbesto bacteriophage,not only as an
immediateeftect, but also as a transmissiblecharacter.
Chapter 6, Note 34. Lipids and Survival Time of Tetrahymena
Various fatty acids have shown a specialinfluenceon survival time of
tetrahymenapyriformis.Survivaltime was determinedby keeping a few
drops of culture in a capillary closedat one end and by daily examination
of the mobility of the tetrahymena.By adding progressiveamounts of an
agent and withdrawing such a capillary after each addition, we also were
able to determinethe influenceexertedby different concentrations.
In general,additionof very small amountsof fatty acidswas found to
prolong survival. Similar effectswere obtained with hydropersulfide,and
even with a solution of saponineswhich are known to bound sterols in
insolublecombinations.The most marked effect was obtained with a fatty
acid having a relativelylow number of carbons.The addition of heptanoic
acid in progressiveamountsgreatly increasedlongevity.The longe"ity increasedas the concentrationincreasedup to a point, after which it declined
rapidly when greater concentrationswere used. Of all the substances
studied, heptanoic acid appearedto be most effective in increasing the
longevityof tetrahymenapyriformis.

NOTES

6s7

Chapter 6, Note 35. Lipids, Temperatureand Tetrahymena

We have investigatedthe influenceexertedby difterent lipids upon the
capacityof tetrahymenapyriformisto resistincreasedtemperature.In general, exposureof a cultureof tetrahymcna
pyriformisto increascdtemperature induces rapid death. Kept in capillariesso a temperatureof 37'C
could be quickly attained,tetrahymenadied in about l5 minutes.By growing tetrahymenafor severalsuccessive
generationsin a medium containing
insaponifiablefractions,increasedsensitivityto temperaturewas obtained.
In some experiments,death occurredwithin five minutes after exposureto
37'C. On the other hand,gowth in mediato which fatty acidswere added
markedly increasedresistance,and survival after more than 7z hour of
e x p o s u r et o 3 7 " C w a s s e e n .
Chapter 6, Note 36. Pain Induced by Lipids
During treatmentof cancer patientswith lipids, especiallyin exceptionally large amounts,some who had beenpain-freebefore,experiencedpain.
Correlation could be establishedbetweenthe appearanceof pain and the
administrationof lipids since pain increasedwith each new injection.
Furthermore, a differencewas seento exist bctween the pain induced by
the administrationof fatty acids and insaponifiablefractions. While the
former had an alkaline pattern, the second had an acid one. In several
cases,pain subsidedwith discontinuationof the medication;in others, it
persisted.The pain always was controlled by the administration of the
opposite lipids, fatty acids for the acid type of pain and sterols for the
alkaline.
Chapter6, Note 37. Lipids and Wound Healing
The influence exerted by lipids upon wounds was followed by the
changesinducedin the healingprocesses.
In order to comparethe wounds,
one squarecentimeterof the skin was exciseddown to the aponeurosis
on
the back of rats and rabbitsafter mechanicalepilationof the skin, and the
surfacemeasureddaily. We usedtransparent
cellophaneon which the exact
wounds
the
were
drawn. The outlineswhich corresponded
dimensionsof
to the surfaceof the woundswerc then passedon paper and cut out. The
paper outlines were then weighed to give us a means of comparing the
changesin the actual surfacesof thesewoundsduring healing.
While lipid acids in generalinduce a retardationin the healing of the
wound, the administrationof lipoids with a positive character were seen
to have an opposite effect. It is worthwhile noting that the naturally ocand insaponifiablefractions-are much more
curring sterols----cholesterol
active than the syntheticin inducingrapid wound healing.
We studiedthe histologicaland cytologicalchangesin parallel incisions
made on the back of rats and rabbits and excisedat intervals.In animals
treatedwith sterol preparations,a difterencewas seenbetweenthe healing

658

n E S E A R C HI N p H y s r o p A T H o L o c y

of epithelialand connectivetissuewounds.While healingof the former was

enhanced,healingof the latter was not influenced.On the other hand, the
higher alcohols,such as polyunsaturatedalcoholsor butanol, were more
a c t i v ei n i n c r c a s i n g
h e a l i n go f c o n n c c t i v et i s s u ew o u n d st h a n o f e p i t h e l i r l .
It is interestingto note also that in the wounds treatcd with the sterol
preparations,
the scar showedan epithcliumwith many more layers than
that of the normal surroundingskin. In rabbits, instead of two or three
layers,there were more than ten,
Chapter6, Note 38. Liver Regeneration
The ability of rats to regeneratealmost 3/nof their liver in a short time
has made them valuable for the study of the factors which intervene in
cellular multiplication and differentiation.A study which we made in collaborationwith E. F. Taskierhas shown thc importanceof biologicalage
of the individual in these processes.
Regenerationis rapidly completed in
young animals;much more time is required in the old. Liver regeneration
has beenseento be relatedto the appearanceof fats in the form of droplets
f,lling up liver cells.Regeneration
follows this first phase.The appearance
of the fatty droplets providesa meansof judgrng the velocity of regeneration. The importanceof age is shown by the fact that fatty droplets appear
early in the liver cellsof very younganimals,filling up the cells,in the first
24 hours.They appearlater-in abouttwo days-in young adults;in three
daysfor middlc-agedrats; after the fourth day in old animals.
The influcnceexertedby administrationof different agentsupon regenerationcould alsobe judgcdthroughthe changesinducedin the appearanceof the fatty droplets,The administration
of insaponifiable
fractionsin
generalhas induced an earlier appearanceof the fatty cells. Injection of
2 c c . o f a I O C / os o l u t i o no f t h e i n s a p o n i f i a b lfer a c t i o no f h u m a n o r e v e n
cow placentawas seento inducean early appearanccof the fatty droplets
and a filling up of the liver cells.Even in old rats such changesoccurred on
the secondday, contrastingmarkedlywith control rats of the same age in
which this would happenon the fourth day or later. Under the influenceof
theseinsaponifiablefractions,the old animalsbehavelike youngsters,from
the point of view of liver regeneration.
The opposite effect was exerted by the administrationof I cc. of a
l0% solutionin oil of the lipoacidsof human placentaor of a 107o solution of cod liver oil fatty acids.Appearanceof fatty droplets was delayed.
In young animals,the droplets were not seen until the third or even the
fourth day. With a high dosesuchas 2 cc. twice a day, of the samepreparat i o n i n a n i m a l so f l - 5 0g r a n ' l sn, o f a t t yd r o p l e t sa p p e a r e d
at all.It is interesting to note that in animals treated with such large doses of lipoacids,
still takesplaceevenwithoutthe appearance
regeneration
of the fatty drop
l c t s . I n t h e s cc a s c st h c l i v c r c c l l s a r e c o m p a r a t i v e lvye r y s m a l l a n d h a v e
compactnuclei,insteadof the reticularaspectof the nucleiin the controls.
It is also to bc noted that a parallelismwas seenbetweenthe appearance of fatty dropletsin liver cells and the richnessof adrenalsin sudano-

NorEs

659

phil granules.In casesin which the administrationof large amountsof

lipoacidswas followed by nonappearance
of fatty droplets in the liver, the
adrenalswere found to be entirelydepletedof fats.
The influenceexertedby the lipids upon livcr regenerationconfirmsthe
antagonisticrole of the two groups of lipids in aging processes.The administrationof insaponifiablefractionsproducesa regenerativeresponsc
c h a r a c t e r i s t iocf y o u n g a n i m a l s ,w h i l c l i p o a c i d sp r o d u c et h e r e s p o n s eo f
aged animals.We have appliedthesefindingsto other processes
in which
age is known to be a major factor-such as in thc healingof wounds,and
especiallyof fractures,in older people,where administrationof insaponifiable fractionshas bccn seen to changean atonic lesion into a rapidly
healingone.
The study of liver regenerationhas also indicatedqualitativedifferences
betweenvariouspreparations.It is thus interestingto note that, of all the
insaponifiable
fractionsused,the most activewere thosefrom placentaand
embryos.The insaponifiablefraction of liver also has shown a special
capacityto induce rapid regcnerationcspeciallyof liver tissue.Higher alcohols have shown much less regenerativeeffect than the insaponifiable
f ractions.
Chapter6, Note 39. Lipids and Convulsions
The administrationof insaponifiable
fractionsof placentaor organs
sometimesproducesno observablemanifestations
in rats and mice, except
an exophthalmia.However,more profoundchangesoccur, sinceinjections
of thiaminein doscsotherwiseharmlessare followcdby lethal convulsions
in theseanimals.80 to 100 milligramsof thiamine/100gr. of body weight
produce lethal convulsionsin rats or mice who received I cc. of. 57o insaponifiablefractions of placentaper 100 gr. of body weight in daily inj e c t i o n sf o r a w e e k .I n c o n t r o l s 1
, 5 0 m i l l i g r a m so f t h i a m i n ep e r 1 0 0 g r . o f
body wciglrtwerc neccssary
to inducefatal convulsions.
High dosesof insaponifiable
fraction of organsalone, also produced
convulsions.
The injcctiontwice a day of a 5% oily solutionof insaponifiable fraction of placentain dosesof.2 cc./100 gr. of body weight was
seento inducelethal convulsionsafter lessthan a week of treatment.
C h a p t c r6 , N o t e 4 0 . L i p o i d s a n d C o m a
The administrationof heptanoleven in larger doscsrvas not seen to
induce somnolenceor coma. Intravenousinjectionof a salinesolutionof
I m i l l i g r a mo f h e p t a n o p
l e r c c . i n d u c e dd c a t hi n m i c e i n d o s e sa b o v e0 . 5
c c . W i t h 0 . 3 c c . , t h e m i c e r e m a i n e di n d e e ps l e e p s, o m e t i m ew
s ith respiratory arrest.Most of the animals,however,recovcred,startingto breathe
in lessthan half a minute and awakeningin about ten minutes.A dose as
high as 10 cc. of the same solution,containing10 milligrams,injected
intravenouslyin rabbits, producedno more than a very short period of
inactivity,without inducingsleep.Intramusculardosesas high as 500 mil-

660

RESE^RCH IN

PHYSIOPATHOLocY

ligrams of heptanol in oil in humansdid not produce somnolence.However,

after severaldays of concomitant administration of heptanol and cortisone,
even in reducedamounts such as 50 milligrams of heptanol and of cortisone
daily, deep somnolencewas seen to appear in some patients, and coma
in two cases. In one, a man of 85, we were unable to overcome the
coma. In the other, administrationof cod liver oil fatty acids, sodium
thiosulfate, and especially Vi cc. of DOCA (desoxycorticosterolacetate),
brought the patient back to normal state.
Chapter 6, Note 4l. Cardiac Rhythm
The antagonisticinfluenceexerted by the two groups of lipoids was
seen to have an especiallyinterestingeffect upon the cardiac cells. The
importance for the pharmacologicalstudy of the lipoids, as well as for the
cardiac physiology and pathology of the changesinduced, has urged us
to study them in more detail.
The principal physiologicalproperty of the cardiac cell is its automatism, that is, its capacity to produce the proper energeticinflux which
when discharged,will inducethe contractionof the myofibrils. Through the
cytoplasmaticbond formations characteristicof the myocardial cells, the
dischargedinflux passesalso into the nearbycells where it acts as an external incitation which, in turn, induces the dischargeof the influxes
produced by these cells. It is through this progressivedischargeof contiguouscardiaccellsthat the contractionprogresses
in a centrifugalmanner
through the heart.
Each cell needsa definitetime to "mature" its own influx, a negative
period following each discharge.During this refractory period, the cell
doesnot respondto any influx, eitherfrom nearbycellsor from any external
excitation.On the other hand, due to the sameprogressivematurationof
its proper energeticinflux, if within a certain time this influx incitation
producedin the cell has not been dischargedby an influx coming from a
nearby cell, the cell itself dischargesit. This automatismis common to
all cardiaccells, lt differshowever.from one cell to another in the time
necessaryfor the influx to mature, that is, in the time necessaryto bring
the cell out of its negativerefractoryperiod or to dischargeits own influx,
if not dischargedby an externalincitationto the cell. A cell has a high
automatismif it has a short rcfractoryperiod, if it rapidly producesits
influx, and if it dischargesit early. A cell has a low automatismif its
negativeperiod is long and if it requiresa long time to dischargeits own
influx if not dischargedby an influx coming from the nearby cells. The
rhythm of the contractionsof the entire heart will be given by the dischargetime of the cellswith the highestauromatism.
If groupsof cellshavean abnormallylow automatismand their negative
period is so long that thesecellswill still be in the refractivenegativeperiod
when the influx from nearby cells arrives to them, they will not be dis-chargedby this flux. If the group of cells representsa part of the heart

NorEs

661

throughivhichthe influx hasto passin orderto attainthe entireheart,it will

block its propagation.
The normal cardiac physiologyresults from the inequality of the
automatismof the differentcardiaccells.Thosewith the highestautomatism
will representthe pacemakerfor the entireheartcontraction.Under normal
conditionsthe cells of the sino-auricularnode show this highestautomatism. Other cells with an automatismlower than that of the pacemaker,
but still sufficientlyhigh to be out of their rcfractiveperiod, will respond
when the influx startedby the sino-auricularnode arrives to them. The
automatismof the other centerspresentin the heart-Aschoff-Tawara's
node, Hiss'sband, its branches,Purkinge'scells-progressivelylower than
that of the sino-auricularnode, will supply an influx if that of the sinoa u r i c u l a rf a i l s t o r e a c ht h e m i n d u e t i m e .
U n d e r a b n o r m a lc o n d i t i o n st,h i s a u t o m a t i s mi s i n f l u e n c e dI.t c a n b e
e i t h e ri n c r e a s e o
d r d e c r e a s e dI n. g e n e r a li,f t h e a u t o m a t i s mo f c e l l so t h e r
than thoseof the sino-auricular
node,is increased
abovethat corresponding
t o t h c r h y t h m o f t h i s n o d c . t h c i r i n f l u x w i l l b e p r e m a t u r e l yd i s c h a r g e d .
I f t h e c e l l s a r o u n d i t a r e o u t o f t h e i r r e f r a c t i v ep e r i o d , t h i s i n f l u x w i l l
propagatcand inducc a contraction.They appearas abnormalpacemaker
c e n t e r sd u e t o t h e i r p r e m a t u r ed i s c h a r g a
e n d a l s ot o t h e i r e c t o p i cp o s i t i o n .
The rcsultingcontractionswill bc manifestedas extrasystoles,
if the abnormal dischargeappearsas an isolatcdevent, or as paroxysmaltachyc a r d i ai f t h e a b n o r m a l i t yp e r s i s t sI.n a u r i c u l a fr i b r i l l a t i o nt,h i s a b n o r m a l i t y
takes place in a larger group of cells.Oppositely,a lowered automatism
affectingan entire group of cclls will result in a blockageof the passage
of the normal influx due to this lengthenednegativeperiod.
The factor which appearsto governthe differences
seenin automatism
of the variouscentersin the heart, is the degreeof differentiationof the
respectivecells. As a gcneralrule. a less differentiatedcell has a higher
automatism,while a more differentiatedcell has a lower automatism.
W e h a v e s e c n t h a t u p t o a c c r t a i n p o i n t . t h e p r o p e r t i e sr e l a t e dt o
t h e d e g r e eo f t h e d i f f e r e n t i a t i oonf t h e s ec e l l sc a n b e c o n n e c t e d
with youth
characters.The changcsseen in hcart ccllular physiology,and especially
t h o s ew h i c h a p p e a ru n d e ra b n o r m acl o n d i t i o n sc,a n b e c o n c e i v e d
as taking
place through changcsin the degrcc of the differentiationof the cells.
We have seenabove,in the study of the influenceexertedby lipids, that
rvhile the unsaponifiablefraction induces a "prolonged youth" with a
degreeof the dedifferentiation
of the cells,the acid lipid fractionsinduce
a processsimilar to a morc rapid aging, respectivelya more advanced
This effect was seen also to be generalfor the respective
differentiation.
p o s i t i v ea n d n e g a t i v el i p o i d s .W h i l e f o r o t h e rc e l l ss u c h a c h a n g em a y b e
uneventful,for the cardiaccell it will be marked by a changein automatism.
From this specificpoint of view, we have studiedthe influenceexerted
by differcntagentsupon the heart,seekingin the changesinduced,modificationcorrespondingto an increasedor decreasedautomatism.Clinical
o b s e r v a t i o nhsa v es h o w ns u c hc o r r e l a t i o nE. x t r a s y s t o l ewse r es e e nt o a p p e a r

662

, , ' R F - s E A R c r {r N p H y s I o p A T H o L o c y

in subjectswho had previousextrasystoles,

when lipoidswith positivepolar
groupswereadministered
in high doses.They disappeared
when the medication was stoppedand reappearedwhen medicationwas resumed.In cases
with previous auricular fibrillation we have seen it reappearwith high
dosesof positivelipoids, disappearwith cessationof the medicationand
reappearwhen medicationwas resumedfor even a short time. This was
fully controlledby the administration
of lipoidswith negativepolar groups.
In hundreds of electrocardiograms
taken of experimentalanimals,
such a correlationbetween the administrationof lipoids and induced
arrhythmiaswas investigated
in collaborationwith I. Eroglu. We studied
thus various substances,
lipoids with positiveor negativecharactersadministeredintraperitoneaUy
or intraveneouslyin rabbits. An extremely
high amountof the agentwas necessary
to influencethe cardiacrhythm in
normal animals.It was usuallynear a lethal dose and in general,proportionately many hundred times that used therapeuticallyin humans. In
repeatedinjectionshowcver, changescould be induced with relatively
smallerdoses.In sufficientdoses,the positivelipoids were seento induce
extrasystoles.
Figurcs292 and 293 show suchchangesobtainedwith huge
dosesof butanoland glyceroladministered
intravenously.(Page 714)
In animals,the negativelipoids induce a dromotropicnegativeeftect.
leadingto auricularcontractionsnot passingto the ventricles.Huge doses
were seento induce a bigeminatedpulse.
The study of the interventionof lipoids has lcd to a new therapeutic
paroxysmaltachycardiaand auricularfibrillation
approach.Extrasystoles,
were seento respondwell to the administrationof lipoacidsand lipoids
with negativepolar groups,while partial blocks were influencedby lipoids
with positivepolar groups.
Chapter 6, Note 42. Some General Considerationsof the Role
of Lipids in Blood Physiology"
Lipids and Red Cells "ln Vitro"
Among the first experimentsconcerningthe influenceexertedby sterols
and polyunsaturatedfatty acids in vitro, were those concerned with the
effectsupon red cells. We have noted that when citrated blood is kept for
two hoursat3l"C in a testtube.the wallsof which have beencoatedwith
crystalsof cholesterolor with nonsaponifiable
fractionsobtainedfrom various natural sources,the red cellsbecomemore swollenand turgescent,and
lesscrenatedthan thosenot treated.Seenunder the dark field microscope.
the cell crown appeareduniformly more refringent.It rvas also noted that
the treatedred cellsfailed to form rouleauxor conglutinatessimilar to those
seenas sludgein vivo. At the sametime, the cells appearedricher in their
sterol content. None of these changeswere observedwhen the red cells
were separatedfrom their plasma and washedwith saline and kept in a
salinesolutionwhen treatedwith sterols,in the mannermentionedabove.
+ Delivered at Gordon ResearchConferences,Kimball Academy, Meriden, New
H a m p s h i r e .1 9 5 5

NorEs

663

Opposite effectswere observedwhen fatty acids were added to blood.

As the direct contact with the red cells produceshemolysis,the following
techniquewas used.Fatty acid preparationscspeciallyas mixturesobtained
from blood or cod liver oil, were addedto hcparinizedor citratedplasma,
thoroughlyagitatedand the excessseparatedby centrifugation.The plasma
so treatedwas then added in variousproportionsto citrated or heparinized
b l o o d f r o m t h e s a m c s u b j c c t .T h i s p o r t i o n o f t h i s b l o o d w a s c e n t r i f u g e d
and the treatedplasmaaddcd to the supcrnatantplasmafrom which thc
sameamountwas withdrawn.The addedplasmawas mixed with the supernatantplasmaand thcn this wasmixed with the red cells.In this way hemolysis was prevented.Smail amountsof treatedplasmacausedthe red cells
to shrink in size and frequentlybecomccrenatcd.In addition, a strong
tendencyto conglutinatewhich exceededthat noted in correspondingcontrol specimens,was observed. When the quantity of fatty acid-treated
p l a s m ac x c c e d c da c e r t a i na m o u n t ,h c m o l y s i sw a s i n d u c e d .T h e a d d i t i o n
of thesetwo groups of lipids to red blood cells, have appcaredto exert
frank antagonistic
effects.
SedimentationRate
The two groups of lipids were also found to influenceoppositely the
red cell sedimentationrate in citrated blood. When citrated blood samples
h a v i n g h i g h s e d i n r c n t a t i ovne l o c i t i c sw, c r e t r e a t c dw i t h c h o l e s t e r ool r a n
insaponifiable
fraction in the manner describedabove,the speedof sedimentationwas markedlyreduccd.Tesr-e XXVIII showsresultsobtained
T r n rr , X X V I I I
R r . oC r r . l S r , o r v r x r e r t o xR , r r r ( n t m . h r . ) S , , r v p r . rTsR p _ , r r e o
W t r l r U x s l p o N t F r A B L EF n r c r r o x o r B l o o o L t p t o s

Control
lt0
96

ul
4fi
t8
l2
I
8
6

Treated
t2
l9
l8
l5
l0
6
1

tl
5

mentationratc tendedto increaseto abnormalvalues.This varicd with the

a m o u n to f t r e a t e dp l a s m aa d d e d .( T n n l t X X I X )
in differentblood samplesin which the sedimcntarion
rate during one hour,
was measuredby the Westergrenmethod.In gcneral,it can be seenthat
the higherthe sedimentation
rate of the untreatedsample,the greaterwas
the eftectof addingsterols,
on the other hand, whcn polyunsaturated
fatty acidswcre addedin the
manneralreadydescribed,to citratedblood from healthlysubjects,the sedi-

664

xESEARcH rN pHysropATHoLocy

Trnlr XXIX
Reo Cells SeorpreNr,rtroN
Rrrt'-ntm,i'lst hour
Fatty Acid Used
Stearic
Palmitic
Linoleic
Linolenic
Cod Liver Oil

Control
9
9
9
9
9

Quantity of Treated Plasma

Added to 5 cc. Citrated Blood
1/4 cc.
l/2 cc.
I cc.
8
9
9
l0
9
t0
l5
l8
22
l5
2l
25
20
36
Hemolysis

Red Cell Volume

The same opposite effectsof sterols and fatty acids were further observedupon the volume of red cells, as determinedby the hematocrit,or
alsowhen the sedimentation
in tubeswas observedover a 24 hour period.
(TeuLE XXX) Sterol-treated
blood showeda significantincreasein red
T r s L EX X X
CHrNcesrN Volupre on REoCells rN CrrR,rreoBlooo TRrlreo rN VlrR<t
(Sedimentation
After 24 Hours)
SubstanceUsed
Unsaponifiablefraction of blood
Stearic Acid
Saponifiablefraction of blood

Control

53
53
53

Treated
66
54
50

cell volume,while on the other hand,with the additionof polyunsaturated

fatty acids,the red ceil volume decreased.This agreeswith a retention of
water by the cells in generalwhen richer in sterols,which Schaefferhas
describedas the lipocytic index.
On further analysis,theseeffectsof fatty acidsupon red cellsmentioned
abovecould be relatedto the polyunsaturationof theseacids,sinceby treating the blood undcr the sameconditionswith saturatcdmembers,such as
palmidg or stearic,thesechangeswere not obtained.
The treatmentof red cellswith conjugatedfatty acids,especiallytrienes.
inducesa marked vacuolization.This is seen to occur through an accumulationof part of the contentof the red cell in droplets,stronglystained
w i t h e o s i n . ( F i S . 2 6 4 a )S i m i l a rc h a n g e sa r c s e e nt o o c c u r i n v i v o . I n
lesionscharacterized
by a predominance
of fatty acids or induced by the
of conjugatedfatty acids,suchvacuolatedred cellsare often
administration
togetherwith othercharacters,
seen.We usedtheir presence,
for the pathol o g i c a ld i a g n o s ios f t h e t y p e D p r e s e n itn a l c s i o n . ( F i S . 2 6 4 b )
Red Cells, Plasma and Lipids
As most of thesechangesdid not occur with the red cellsin suspension
in differentisotonicsalinesolutions,we have attemptedto explore the re-

NorEs

665

l a t i o n s h i po f p l a s m at o r e d c e l l sa n d l i p i d s ,T h i s w a s d o n e i n t h e f o l l o w i n g
manner. The cholesterolcontent of red cells was seen to be progressively
lowered by repeatedwashingswith isotonicsaline.when the amount of
cholesterolis reducedbelow a certainlevel,hemolysisensues.Standardizing thesewashingsby replacingthe plasmawith an equalamountof saline,
hemolysisis usuallyobtainedin some bloodsafter I or 2 washingswhile
i n m o s ta f t e rm o r et h a n l 0 w a s h i n g sT.h i s o c c u r sw h e nt h e c h o l e s t e r oclo n -

oc2; &

o Co
(a)

(b)

F t c . 2 6 4 . T ' h e t r e a t m c n to f r e d c e l l s i n v i t r o w i t h c o n j u g a t e df a t t y a c i d s ( t r i e n e s ) i n d u c e st h e a p p e a r a n c eo f v a c u o l e s .( a ) I n s p o n t a n e o u Ise s i o n sc h a r a c t e r i z e db y a n o f f b a l a n c e t y p e D w i t h a p r e d o m i n a n c eo f f a t t y a c i d s o r i n l e s i o n s i n d u c e d b y t h e
a d m i n i s t r a t i o no f c o n j u g a t e dt r i e n i c f a t t y a c i d s , v a c u o l a t e dr e d c e l l s a r e s e c n . ( b )

tent of the red cellsfalls below 58 mgm. Vo. When the separatedplasmais
again added to these repeatedlywashed and consequentlycholesterolimpoverishedblood cells, their cholesterolrises. (Trnle XXXI) By
further additionsof new portionsof the sameplasma,the cholesterolcontent of the red cells can be progressively
elevatedto the original values.
Apparentlycholesterolpassesreadilyfrom the plasmato the red cells.This
Trsle XXXI
I N r . r - u E r c eo r C t r n a r e o P l , r s u l U p o x R E o C e l l C H o l e s r e R o l
BeforeTreatnrent
After l0 Washings
After First Treatnrent
with Plasma
After Second Treatment
with Plasma

186 mgr.9t
62
lil

t42

666

nEsEARcH rN pHysropATHoLocy

was confirmed by measuring the cholesterolcontent of the plasma before

and after it was mixed with cholesterol-impoverishedcells. After treating
a portion of citrated plasmaseveraltimes with other portions of cholesterolimpoverished red cells, the amount of cholesterol in the plasma was
markedly reduced. The addition of the unwashedred cells to this cholesterol-impoverishedplasmaraisedback its cholesterolcontent. The content
could be increasedstill further almost back to the previous values by repeating this procedure. (Tnnre XXXII) It was clear that a balance in
Trslp XXXII
INn'r-ur'NcE
on Rro Cells Upox Pl,rsu,r CHolestenoL
Before Treatment
After Fifth Treatmentwith
Washed Red Cells
After First Addition of
Unwashed Red Cells
After Second Addition of
Unwashed Red Cells

226 mgr.7o
96
163
180

the cholesterolcontent seemsto be realized between plasma and red cells

through possiblepassagesin both directions.This fact would imply that the
red cells may serve as a bufter reservefor rapid changesoccurring in the
plasmasterols.
The role of red cells in the transportationand distribution of fatty acids
throughthe blood not only appearsmore evident,but also indicatesa selective intervention. When plasma was treated as mentioned above with fatty
acids that were easilyidentifiable,and then mixed with the red cells, an
unequaldistribution betweenplasma and red cells was seen.The difterent
influenceof the different fatty acids becameobvious. Saturatedfatty acids
could not be found in the red cells when theseacids were used, while unsaturatedfatty acids seemedto be selectivelyretained.Fatty acids such as
oleic, linolenic, eleostearic,or norbixine were found convenientto be used
for this pulpose. They were easily identified, the first through its chemical
character,the second after conjugation,and spectral analysis,the third
through its characteristicabsorptionin ultravioletlight, and the last through
its color in chromatographiccolumn. After mixing with the red cells, they
were found unequallyand selectively
fixed to the red cells: leastfor oleic,
fixationwas found to increasewith the degreeof desaturation.
The same
fixation was seen to occur in vivo. When animals were treated with saturated fatty acids, these substancesdid not appear in the red cells. When
treated with oleic, linolenic, eleostearicacid or norbixine, the content of
theseacids in the red cells was found to be for the last acids as much as
five times higher than in plasma.A selectivefixation of those fatty acids on
the red cells could thus be recognized.This appearedstill mst. striking
when compared with that of cholesterol.In animals treated with cholesterol, the relative proportions between the proportion in plasma and cells

NorEs

667

was not seento be altered by a total increasein cholesterol.It appearsthat

the red cells have the capacity of selectivelyfixing from the plasma certain
fatty acids,particularlythe more unsaturated
ones.
Lipids and Blood Oxygen Transporl
one of the most interesting observationsand one of the simplest in
vitro experiments that indicates these opposite eftects of sterols and fatty
acids is their influence upon the oxidation processesin which red cclls
intervene.when a sampleof ordinary venousblood is treatedwith a preparation of cholesterol or nonsaponifiablefractions, using the above mentioned techniqueand after its separationfrom the cholesterolit is agitated
with air or oxygenwhich is passedthroughthesesamples,the color becomes
a bright vermilion red, and this persistsfor a long time. when the same
venous blood is treated with a preparation of polyunsaturatedfatty acids
as mentioned above, the color becomes very dark, almost black purple.
when air or oxygen is passedthrough these samples,the blood becomes
lighter in color for only a short time, the darker color reappearingwithin
a few minutes.one is immediately impressedby the similarity of the cholesterol-treated blood to arterial blood, while the fatty acid-treated blood is
similar to venous blood, and especially to the color of venous blood in
casesof shock.
We tried to tie in thesefindingswith the observationof Binet concerning the changesin blood fatty acids when passingthrough the lungs. He
has been able to show that the amount of the polyunsaturated
members
appearsto be reducedby the passageof blood through the lungs. We could
show that the red cells leavingthe pulmonaryvascularbed are somewhat
richer in the unbound cholesterol than they have been in the blood which
entered the lungs. The lipid content is altered in an oppositeway as the
blood travels through the general circulation. That is, the polyunsaturated
fatty acid content is increasedin the red cells while the quantity of free
cholesterolseemsto be diminished.The sterol-richerred cells appearcapable of retaining for a longer period of time, the amount of oxygen which
hemoglobin has fixed, while a rapid reduction of oxyhemoglobin is seen in
the red cells when the polyunsaturatedfatty acidsintervene.This led us to
consider an interventionof these two groups of lipids in relation to the
oxygen transportationby the red cells.Bearing in mind the fact that while
cholesterolreducescell permeability and polyunsaturatedfatty acids increaseit, an alternatinginterventionof theselipids seemsto play a role in
a better distribution of oxygen. The oxygen which is fixed by hemoglobin
rvhenthe red cclls have passedthe lungs, is largely retainedas such by
the intervention of the sterolsuntil they reach the point in the tissueswhere
liberation of oxygen is necessary,this being favored now by the intervention of the fatty acids.
Lipids and State ol Shock
The abnormallydark color of the blood resultingfrom its treatmentin
vitro with polyunsaturatedfatty acidshas suggested
the interventionof such

668

nEsEARcHrN pHysropATHoLocy

substances
in thoseclinical conditionsin which similar color changesare
notedin the blood as in shock.We will presentour studiesin shock belou'.
For the momentwe will only note that in the stateof shockexperimentally
inducedby trauma,burns or irradiation,or found in terminallyill adrenalectomized
animals,theseanimalshavenot only a high fatty acid content,
but that the kind of fatty acidsencountered
are not the sameas in normal
animals.We have discussed
theseabnormalfatty acidsabove.The existing
differences
have beenshown by measuringthe quantity of oxalic acid that
is producedwhen thesefatty acidsare submittedto a careful standardized
oxidativefission.The oxidativefissionof the fatty acids not only from their
entirebody but even from their blood has shown that for normal animals,
no oxalic acid could be found, leadingto the assumptionthat no conjugatedfatty acidsare present.On the other hand, oxalic acid appearcdwhen
fatty acidsobtainedfrom animalsin shockor from their blood were broken
down with the analyticalmethod utilized.
Of particularsignificance
for the pathogenicrole of thesefatty acids is
the fact which we will discussagain below, that death appearsto ensue
when the conjugationof fatty acids reachesa certain value, which is ap
proximatelythe same whetherthe animal has been traumatized,burned,
irradiatedor adrenalectomized,
and independentof the fact that death occurs in a short time or severa-l
days.It corresponds
to l4-17 mgm. of oxalic
acid per gram of fatty acids.It is also interestingto note that these abnormal fatty acids were found to be more abundantin the red cells than
in the plasma.
Eflect upon Leucocytes
The biologicalantagonismbetweensterolsand fatty acids has appeared
in the influenceupon other blood constituents.We have observcdthat the
administrationof sterolstends to elevatethe total white blood cell count
and especiallythe number of neutrophilic granulocytes.Polyunsaturated
fatty acids,on the other hand, producerapid leucopenia,and again it is
the neutrophileelementsthat are first affected.A hyperleucocytosis
often
was seenfollowingthe neutropeniainducedby polyunsaturated
fatty acids
if small amountsare administered.This effectcould be consideredas being
reactionalto thc first leucopenia,since it is retardedor even preventedif
large dosesof thesefatty acidsare injected.(Tnnr-e xxxIII)
It is also
interestingto note that a deviationto the right, in Arneth's formula, was
seenafter the treatmentwith fatty acids;and to the left after treatment
with sterols.Thus, this concordswcll with the antagonisticeffectsupon
the agingprocessseenfor theselipids and which is discussedbelow.
Lipids and Blood Serum Cholesterol
Further study of the relationshipbetweenblood and lipids has permitted the recognitionof severalpeculiaritiesconcerningthe blood serum
which when relatedto abnormalconditions,acquiresa specialsignificance.
Policard has observedthat when crystalsof cholesterolare added to blood
sera,two oppositechangescan ensue.In one, a precipitateappearswhile

N O T E S

,,'

669

TrsLE XXXIII
E r r e c r o r L r p r o sA o v t x r s t e R E D
l N V l v o U p o Nr H e T o r , r l N u t ' l s r R
or Leucocytes
Fractionof Blood-lO% Solution-S cc. I.P.
Unsaponifiable
BeforeAdministration
14,600
12,000
2 Hours Later
18,400
19,000
77r Hours Later
26,000
22600
Fractionof Blood-10% Solution-S cc. I.P.
Saponifiable
BeforeAdministration
13,200
16,200
2 Hours Later
I 1.000
6,800
7 V 2 H o u r sL a r e r
5,000
5,100
S t e a r i cA c i d - 1 0 7 o S o l u t i o n - Sc c . I . P .
B e f o r eA d m i n i s t r a t i o n
I6.100
14,200
2 H o u r sL a t e r
12,800
15,100
7 7z Hours Later
I 5,000
I 2,000
the serumcholesterol
contentdecreases.
On the contraryin the other,a part
of the addedcholesterolpassesin solutioninto the serum,therebycausing
an increasein cholesterolcontent. When animals were treated with large
amountsof sterolsfor a long time, their sera showedthis tendencyto precipitationwhen in contactwith cholesterolin vitro, while the sera of animals treated with large amounts of fatty acids showed the capacity of
dissolvingmore cholesterol.We believethat this capacityof sera to precipitate in the presenceof cholesterolmay be correlatedwith the clinical conditions presentin arteriosclerosis
when acute episodesoccur.
Thesestudiesof the role of lipids in blood physiologysuggestthat the
general antagonismbetween sterols and polyunsaturatedfatty acids also
intervenesin other important processes
of blood physiology.It has thus
raisedthe questionof the role theselipids may play, through their opposite
eftects, in different metabolic balancesof the body governed by blood
changes.
Chapter7, Note l. AnalysesUsed for the Study of Hemoshock
Besides body temperature and blood pressure,the following blood
were made: completeblood count;coagulationtime; clot retractaanalyses
bility; values of albumin and globulin, total and free cholesterol.free
fatty acids,degradatedproteins,antitrypticpower of the serum, esterase
amylase,potassium,sodium,calcium and glucose.Most of theseanalyses
were made in venousblood samplesobtaincdevery fivc minutes,during
the fifteenminutesprecedingand the half hour following the noxious intervention.

67t)

RESEARCH IN

PHYSIOPATHOI-OCY

Chapter 7, Note 2. blorphine and Shock

T'hc possibilityof inducing hemoshocksimply through an intravenous
injection o[ colloid;rl metals has provided a useful method to study the
conditions under which hemoshockcan be induced clr suppressed.We
have investigateda seriesof agentsto determinetheir influenceupon shockAdrenalin, quinine, ephedrineand atropinc havc not changedthe coursr
of clinical and hematicmanifestations.
On the other hand, nrorphine,as
well as other opium derivativcs,completelypreventedthc developmentof
thesemanifestations,Subcutancousinjection of 2 centigramsof morphine

2A
630
subjectssubjects

a
- t q
'.4

lb

(J

wi thout r i t h
i se
exerci se exerc
F t c . l 6 - i . ' I ' h e p r o p r : r t ' t i o no f c h i l l s a p p e a r i n g a f t e r d i r e c t t r a n s f u s i o n ( w i t h t h e
Jubi syringc, injecling 500 cc in less than l0 minutes) increasesmanifestly after cxe r c i s e( w u l k i n g i n t h e r o o m ) .

sulfate lifteen minutes bcfore thc intravenousinjection of tle metal al\\'ayssuppressed

the ntanifestations
as did an intravenousinjection of 5
oi
morphine
milligrarns
sulfatc.Analyseshave shown no leucopcnia,to
occur. Similar efTectswere obtained,with none of the changescharacteristic of henroshock,for instlncc in lransfusionswhere 500 cc. were injected
in iessthan tcn nrinutesand rvheremorphineprcventedthe frequentlyob'l'hc
served chills.
inlluence of morphine upon the leucocyteshas been
con{irmed in thc follorving cxperintcnt.
In rahbits.a pleuralexudatewasobtainedby an intrapleuralinjectionof
broth, 16 hours later, pleural puncturesfurnished fluid rich in leucocytes.
As we have noted above, the adclitionof a colloidal suspensionof silver

NorEs

671

proteinate (collargol) was followed by the appearanceof rapidly gowing

vacuoleswhich led to bursting of leucocytes.(Fig. 76)
The addition of even minimal amounts of morphine or other opium
derivativesentirely prevented these changesin leucocytes.No lysis nor
vacuoleswere seen.
Chapter 7, Note 3. Physical Exercise and Shock
The study of hemoshockhas shownthe influenceexercedby physical
exerciseupon shock.A leucopeniawas observedin normal subjectsafter
an intenselysustainedphysicaleffort, suchas after running for five minutes,
a fact which led us to try to see what influence exercise would have
on the shock induced by the intravenousinjection of colloidal metal.
The shock was seen to be much strongerthan usual. The chill which followed 25 minutes later was also proportionatelysevere.We have since
correlatedthe appearanceof hemoshockwith exercisein patientshaving
direct transfusion.If the patientexercisedimmediatelyafter the transfusion,
a chill consistently
followedabout a half hour later.(Fig.265)
Chapter 7, Note 4. Lymphocytes and Effects in Vitro
The capacity of lymphocytesto hydrolyze even higher esterscan be
demonstratedby having lymphocytesseparatedand their activity tested.
Fluid obtainedfrom tuberculouspleural effusionrich in lymphocyteswas
centrifugedand the fluid decanted.The centrifugatewas then put on a plate
of beeswax,covered with a cup and left for several hours at 37"C. A
clearly visible depressionappearedwhere the lymphocytepreparationhad
been added.
Chapter 7, Note 5. Lipids and Immunity
Three groups of five rabbits each, of the same sex and weight, were
injectedintravenouslyon two consecutivedays with the same amount of a
suspensionof killed Eb. Thyphi. One group was kept as control, receiving
daily injectionsof I cc. of cottonseedoil. Of the other groups, one was
injected subcutaneouslydaily with 1 cc. of a SVo solution in cottonseed
oil of the acid lipids mixture obtained from human placenta. The third
group receiveddaily I cc. of a 5l/c solutionin cottonseedoil of the insaponifiablefraction of the same origin. Every secondday, venous blood
was obtainedand the agglutinatingpower of the serum determined.Figure
265A. showsthe averagevaluesfor each group.
Chapter 7, Note 6. Microbes Treated with Lipids
The injectionof microbeskilled by heat and treatedin vitro with various lipids and lipoids had an interestingeffecton the appearanceof antibodies.Eb. typhi, cultivatedon agar and suspendedin salineso as to give
' : i

.
.1

1' :,.

j.

l.l
, rir:
...1

6'72 .'

R E S E A R c Hr N P H Y S T o P A T H o L o c Y

nephelometricalvaluescorrespondingto 30 mil. pr ., were killed by heatwere treatedby

ingfor I hour at62oC. Differentportionsof this suspension
mixing them with preparationsof the acid lipidic or insaponifiablefractions
of various origins,such as human placenta,cow, carp or rabbit organs;
entire bodies of guinea pigs or rats; entire bodies of squid; seedsof Bixa
orellana;microbessuchas Esch.coli, B. subtilis,and tuberclebacilli. Fatty
acidssuch as oleic, linoleic, eleostearicor mixturesof acids obtained from

tf50f.

H-. Control
o-H I nsaponifiablefraction
.-r'| Lipoacids

1 ?3 4 s 6 ? 8 I 1 0 n
D a v s
Flc. 265A. Lipids and ugglutinirres.lnfluence exerted by the administration of unsaponifiable lipid fraction and the acid lipid fraction of human placenta upon the appearanceof ugglutinine.ragainst Eb. typhi vaccinesinjected intravenously. Each curve
c o r r e s p o n d st o t h e a v e r a g ev a l u e o f 5 r a b b i t s .T h e a g g l u t i n i n e sa p p e a r e a r l i e r a n d
t h e i r a m o u n t i n c r e a s e sm o r e r a p i d l y i n t h e a n i m a l s t r e a t e d w i t h l i p i d s t h a n i n t h c
controls.

cod liver oil or butanol or heptanolalso were used.5 cc. of a 2Vo alcoholic
solutionof the lipoids were introducedin I l0 cc. distilled water and rhe
solventeliminatedby boiling the mixture to bring the amount to l0O cc.
l-5 cc. of thesemilky preparationswere added to 5 cc. of the microbe
suspensionand incubatedat 37oC for 24 hours. The treated microbes
were then separatedby centrifugation,the fluid decantedand the microbes
in the same amount of saline.They were used in doses from
resuspended
l/10-l cc. for subcutaneous
injectionsin rabbis, repeatedevery third day
for two weeks.As controls,animalswere injectedwith the same amounrs
of suspensionof untreatedmicrobes.

NorES

7'

673

For the first two weekssmall amountsof blood were obtainedevery

third day from the ear veins of the treatedanimals.Blood was obtained
from the heartevery secondweek for 8 weeks.The agglutininswere determined for all the samples.In thc rwo-weeksamples,the presenceof immune antibodieswas determinedby the capacityof differentamountsof
t h e s e r at o p r e v e n a
t l e t h a li n f e c t i o ni n m i c eg i v c ni n t r a p e r i t o n e a
i nl j e c t i o n s
o f s t a n d a r da m o u n t so f l i v i n gm i c r o b c sT. h e a g g l u t i n i nw
s e r es e e nt o a p p e a r
earlierthan in s6n11615-ina mannersimilarto that seenwhen lipids were
i n j e c t e di n t h e a n i m a la s r e l a t e di n C h a p t e r7 N o t e 5 . I t w a s e s p e c i a l l iyn
the immuneprotectiveantibodieswherethe differencewas manifest.It was
n o t o n l y a n e a r l i e r a p p c a r a n c eo f t h c s c a n t i b o d i e sb, u t t h e p r o t e c t i o n
a g a i n sat l e t h a li n j e c t i o nw a so b t a i n e dw i t h n r u c hs m a l l e ra n r o u n t o
s f serum.
Chapter7, Note 7. Skin Allergy
An aqueousextract of squid body was preparedby blending it and
extractingwith salinein a proportionof 1720.After filtration,the mixture
was centrifugedand the supernatantfluid put in ampuleswith mcthiolate
added as a preservative.
Someof the ampuleswere tyndallizedby heating
them at 56'C one hour daily for four consecutive
days.
l / 1 0 c c . o f t h c s ep r c p a r a t i o n w
s a s i n j e c t e di n t r a d e r m i c a l liyn v a r i o u s
subjectswho also receivedcontrol injectionsof saline.Immediate and 24
hour reactionswere noted.An indurationpresentthe secondday was consideredas a positivecellularresponse,
while the immediateappearance
of a
hive was consideredas a reactiontaking place in the metazoiccompartment.
Twelve days after a first injectionin exactlythe sameplace,a second
injectionwasgivenwith the samentaterial.The immediateand the 24 hour
reactionswere judged. If the reactionwas negative,a third injectionwas
made, ten days after the second,in the sameplace.It could be seenthat
in normal individuals,the secondsomctimes,
and the third injectionalways
inducedsecondday indurationwhich persistedfor severaldays. In cancer
patients,includingthosein terminalcondition,the injectionof this antigen
inducedvirtually the samereactionas in normal individuals.
Chapter 9, Note l. Hemoglobintrriaa figore (244)
Certaininformationabout shockwhich has emergedfrom the study of
a rare condition is worth noting here, When a patient with paroxysmal
hemoglobinuria-alsoknown as both hcmoglobinuriaa frigore and cold
immerseshis hand in icy water, he experiences
hemoglobinuria,
a chill a
half-hourlater which is followed by the appearance
of hcmoglobinin the
urine. Classically,this phenomenonwas consideredto result from the interventionof a hemoshock.We investigated
such"attacks"of hemoglobinuria in three cases,inducingand studyingthe phenomenonseveraltimes
i n e a c hs u b j e c tU
. s u a l l y ,o b s e r v a t i o nwse r ec a r r i c do u t d u r i n ga t h r c c h o u r

674

n E S E A R c Hr N p H y s r o r , ^ r H O L o G y

period after the immersion of the patient's hands for ten minutes in icy
water and includedthe followingprocedures:
1) Measurements
of blood pressureand temperatureevery 5 minutes;
2) Determinationsat ten minute intervals,of coagulationtime, clot retraction,white cell count and differential;serum hemoglobincontent,
serumproteins,antitrypticpowerand esterase-allmeasu:ements
being
made on venousblood.
3 ) White cell count and differentialmeasuredon capillary blood obtained
every l0 minutesby finger punctureat 5 rninuteintervalsafter withdrawalsof venoussamples.
4) Tests for the presenceand amount of hemoglobinin the urine at 15minute intervals.
Coagulationtimc was established
in the centrifugetube and was related
to the moment when blood ccasedto flow if the vertical positionof the tube
waschanged.CIot retractionwasdeterminedby centrifugingthe coagulated
blood after 2 hours at room temperatureand measuringthe amount of
serum obtainedfrom l5 cc. of blood. The serum and urine hemoglobin
contentwas determinedphotometrically.
For total proteincontent,we used
both the refractomctricindcx of the serum and gravimetricmeasurements
after adequateprccipitation.For antitryptic power we determinedthe inhibitory effectof the scrum upon the digestionof a solutionof caseinby
trypsin.The quantity of esterasepresentwas determincdby the changes
upon cthyl-butyratc.
The data obtaincdwereplottedas curveswith time as commonabscissa.
Parallelvariationsrvereobservedin all three patientsduring repeatedly
inducedattacks.
For almostall analyses,
exceptfor the presence
of hemoglobinin urine.
t h e v a r i a t i o n si n d i c a t c d: r d i p h a s i cp h c n o m c n o n( .F i g . 2 6 6 ) T h e f i r s t p h a s c
was characterized
clinically by hypotensionand slight hypothermia.The
characteristicanalyticalchangeswere lcucopenia,prolongedcoagrlation
time, reductionin clot retraction,lower refractometricserum value. lou,er
antitrypticserumpowerand increased
scrumesterase.
During this first phase
of the diphasicphenomenon,hemoglobinalso appearedin the serum and.
when abundantin the serum,also was found in the urine. The first phase
was followed by a second5 to l0 minuteslater. The clinical manifestations were a sensationof chill, varyingfrom very slight to severe,followed
by temperatureelevationand slight hypertension.Analytical changesin
the opposite direction from those noted during the first phase could be
seen.Hyperleucocytosis,
reducedcoagulationtime, higher retractionof the
clots, elevatedrefractometricvalue and antitryptic power, and reduced
estcrase
contentwere charactcristic
of the secondphase.Hemoglobinpresphase
ent in the serumin the previous
disappeared
at this time.
The most intercstingfinding in paroxysmalhemoglobinuriawas that
two or three such distinctdiphasicepisodesfollowed each immersion.In
all cases,the first diphasiccomplex appearedin about 10 minutes after

NOrEs

675

immersion.It was relatively mild and lasted in gcneral about l0 minutes,

after which all valuesrcturned to prc-attack.
About a half-hour later, however, a seconddiphasic conrplex, much
more intensein its manifestations,
was noted.Hypothermiaand hypotension
were more marked. Leucopenia was more intense,the number of leucocytes
falling to as low as 200 pcr cubic mm. The quantity of hemoglobinin thc
serum w&s very high. with hemoglobinspilling ovcr into the urine in large
amounts.Scrum antitryptic powcr dccreascdto nruch lowcr lcvels than
during the first diphasic complex. In sonrecasesthe coagulationtimc increasedto l5 minutes and the clot almostfailcd to rctract. Thesc chanscs

Gi -c

:L ( . )
aDt

(n*c
(J
C.a

at)
. o lt
I

+ l

E
5
tG'
vl

eo,
:tc
l- .(uL
.ta
c c

-o.o

+ +
r(f.rr

r$g ,rO llt

tlo

lLt

aE

I'linutes
F t c . 2 6 6 . T ' h c c l i n i c a l d a t a a n d a n a l y t i c a lc h a n g e so f h l L x r da n d u r i n c f o l l o w i n g t h e
immersion in icy water clf the hand of a subiect with henroglobinuria a frigore rev e a i t h r e e d i s t i n c t d i p h a s i c p h e n o m e n ac o r r e s p o n d i n gr c s p c c t i v c l yt'o t h r c e s c p a r a t e
hemo shocks. Their intensity appe{irs correlated to the clegree of the occurring
leucopenia.

were followed by the secondphaseof the complex, with scvcrc chill and
manifestchangesin the analyscsin the oppositedirection. lt is the second
phaseof the diphasiccomplcx that is recognizcdclinically as the "attack"
of hemoglobinuria.l'he chill was often very intense,followed by a temperature above 39"C, and hypertension.Leucocytesincreasr:dto as much as
20,000 per cmm., blood coagulationtimc rvas abnormllly shortenedand
clot retractionincreased.Hemoglobindisappearedrapidly from the serunl.
The albumin contcnt as well as the antitryptic porvcr of thc serum increased,whilc the esterasccontent fcll. In about 30 minutes,horvever,all
these changeswere dampened and the blocld slowly rcsained its nornral
This period, with almost all manifestationsslorvlyreturning
characteristics.
to normal often rvas followed by a third diphasicconrplcx.not clinically
evidentbut revealedby hcmatologicalfindings.In most cases,it appcared
about two hours after immersion.While it was much milder than the frrst
two, its diphasiccharacterwas quite clear.Occasionallythc patient rcported
a slight sensationof cold. Thc amountof hemoglobinin the serum was less

676

. . ' R E , s E A R C Ht N p H y s t o p A T t { o t - o c y

than during the first complex, and hemoglobinuriawas never seen.Figurc

266 showsthescfindingsin a typical succession
of thesecomplexes.
An analysisof thesc casesindicatedtwo striking characteristics.One
was the timc sequenceof the three diphasicchangeswhich was uniform for
all attacks in all subjects.The secondwas that although they differed in
intensity,all thc attackswere qualitativclyalike.
In furthcr studieswc ried to undcrstandthe mcaningof thesechanges.
similar changcs,but without henroglobin
in serumand urine, and with only
slight chill as a clinical manifestation,could be observedrvhcn the hands
of normal individualswcre placectin icy water for l0 minutes.Trvo similar
diphasicphenonrcna
werc secnto appear.while their intensitywas greatly
reduced,the time of appearanceof the two diphasicphenomenaobservcd
r,1 c u,Jr

I
I

' i, *"t-:

I
/ a !ri)nl
t

io /';"'1
1
I

o
:

O,
J

i
I

,i

f,
\ 1

l
-

l
l

.rl

, ir

* l'

r*1

*.-.J-

{1, .rl
l

F t c . 2 6 7 . I n t h c p a t i c n t w i t h h e m o g l o b i n u r i aa f r i g o r e . e x e r c i s ea f t e r i m m e r s i o n o f
t h e h a n d i n i c y w a t e r i n d u c e sa v i o l e n t c h i l l w i t h a b u n d a n l h e m o g l o b i n u r i a .

was basicallythe sameas in paticntsrvithcold hemoglobinuria.

This would
indicatcthat thc leucolysisin hcmoglobinuriaa frigore corrcspondsto a
physiological
responsewhich induceshcmolysisbecausethe red cells have
been scnsitizcdby cold. This scnsitizationis rccognizedin thc DonathLandsteinerreaction.It appcnrsprobablethat leucolysisliberatesthe complementnccessaryto induce the hemolysisof the sensitizedred cells.
We have tricd to correlatcthcscchangeswith other processesencountered in nornral and abnormalphysiology.ln dilTerentialstudiesof bltxnJ
smearsobtaincdduring an induccdattackof hcmoglobinuria,
nuclearshadows were obscrvedin the smearsat the tinre of the leucopeniaof the first
phasc.This was exccptionallymanifestfor the seconddiphasiccomplex.
whena markcdlcucopeniaoccurs.We haveseenabovethat this leucopenia
has beencorrelatedto the lysisof the lcucocytes.
In patients with cold hemoglobinuria,rve were able to show that. if
with any physicalexerciseafter the handswere out of the icy water, such
as even walking in the room, the scverityof the attack induced was much
greater than rvhen they werc allowed to rcst quietly. Not only was the

NorEs

677

severityof the chill and thc degreeof hcmoglobinuria

corresponding
to the
seconddiphasicphenomenongreatlyincreased,
but all other changeswere
similarlyintensified.The numbcr of leucocytes
decreascdto lcss than 200
per cubic mm. Blood coagulationtime went to valucsas high as over Vz
hour with almostno retractionof the clots. The antitrypticpower of the
serum, obtainedonly after centrifugation,reachedthe lowest values observed.Figs.266 and 267 show the changesfor the samesubjectwith and
withoutphysicalexercise.
Thesefindingscould explainobservations
indicating the importanceof rest,after blood transfusions.
With the direct transfusion method,where 500 cc. of blood was administeredin less than 10
minutes,chill was seldomseenin patientsrestingquietly,while it was constantlyseento appearin subjectstakingany exerciseimmcdiatelyfollowing
the transfusion.(Fig. 265)

v,
(u
>\
()
o

()
=
q,

t
I
g

o
cl

o
E
|9
CL

R
f

Min:tes
F r c . 2 6 E . T h e a d m i n i s t r a t i o n o f 0 . 0 3 8 m P a n t o p o n p r e v e n t s t h e a p p e a r a n c eo f
m a n i f e s t a t i o n sa f t e r l h e i m m e r s i o n o f t h e h a n d i n i c y w a t e r , i n t h e s u b j e c t w i t h
hemoglobinuria a frigore.

This relationshipof leucolysisto the pathogenesis

of the diphasiccomplex phenomenonin hemoglobinuriaa frigore was confirmedwhen the attack could be preventedby a pretreatmentwhich influencedthe leucocytes.
The study of the influenceexercisedby various agentsin vitro upon the
Ieucocytesof a pleural effusion preparationfrom rabbits, when collargol
solutionswere added, has shown that only morphine and other opium alkaloids were able to prevent lysis. Adrenalineand quinine-among others
-appeared inactive. Administered to patients with paroxysmal hemoglobinuria,theselast substanccs
did not preventattacks.However, morphine in minimal doses,such as 5 mgr. (I / 12 g,) or less,by intravenous
of any clinical manifestations
injection,entirelypreventedthe development
as well as analytic changes.Minimal or no leucopeniawas secn,and there
were no changesin coagulationtime, retractionof the clot, antitryptic
power, refractionindex of the serum.esterase,etc, (FiS. 268)
In addition to confirmingthe hypothesisthat leucolysisintervenesin the

678

,'

RESE^RcH

IN

PHYSIoPATHoLoGy

pathogenesisof the diphasic complex, the influence of morphine has furnisheda meansof preventingthe attacks.
The presenceof two or three successivediphasic phenomena in the
case of paroxystic hemoglobinuriaattack has revealedanother important
relationship.As mentionedabove, hemolysishas been seen to occur only
in the first phaseof the diphasicphenomenon.Hemoglobindisappearsfrom
the serum betweenthe complexes.It is classicallyacceptedthat in cold
hemoglobinuriatwo factors must interveneto produce hemolysis.One is
sensitizationof the red cells which resultsfrom the influenceof cold; the
other is the presenceof the complement.It appearsthat while sensitization
persistsfor a while after exposureto cold, hemolysiswitl occur when the
secondcondition is also fulfilled. Complementis releasedin the first phase
of the diphasicphenomenasinceit is during the first phasethat hemolysis
occurs through the changesinduced by the lysis of the leucocytes.The
correlationbetweenthese two processesis clear from the fact that, under
the influenceof morphine,leucolysisdoes not take place and consequently
hemolysisis prevented.
Chapter 9, Note 2. Lipids and Rouleaux and Sludge Formation
Human blood obtainedthroughvenouspuncturewas mixed in the syri n g e w i t h l / 1 0 o f i t s v o l u m eo f 1 . 5 7 os o d i u mc i t r a t e ,p a s s e di n t o s c v e r a l
centrifugetubes and separatedinto plasmaand cells. The plasma from the
various centrifuge tubes was placed in separatctest tubes. One of these
samplesof plasmawas treatedwith a mixture of conjugatedfatty acids of
cod liver oi.l, another with a lipoacid preparation of human placenta, a
third with a preparationof the insaponifiablefraction of human placenta.
As control, the plasmawas treatedwith liquid paraffin. The mixtures, frequently shaken,were kept in a warm bath at 37'C for one hour, after
which they were centrifugedand the oiJy material separated.The plasma
treatedwith the lipoacidswas then added in proportion of l07o to untreatedplasmaswhich then were reunitedwith their red cells. The plasma
treatedwith the insaponifiablefraction was mixed directly with its respective red cells. Plasma and red cells were shakenfor five minutes and left
at room temperaturefor anotherten minutes.One small drop of this blood
obtained with a platinum loop or a capillary pipette was mixed with two
dropsof salineon a slidecoveredby glassand examinedunder the microscope. While the controls showed few short rouleaux, the blood treated
with insaponifiablefraction showedisolatedcells. In the blood treated with
placenta lipoacids,almost all of the red cells formed rouleaux; in blood
treatedwith conjugatedfatty acid,almostall cellsformed sludges.
Chapter 9, Note 3. Dark Color of the Blood in Shock
In order to determinewhy dark-coloredblood is seenin shock and the
role of fatty acids,the followingexperimentswere performed.
Venousblood of patientsin severestateof shock was drawn and mixed

with 1/10 of its volume of a citrate solution. Th, h..:,**

:::
"r".normal
determinedand salineadded in order to bring it to the value of the
blood. Through these blood samples,oxygen was passedfor five minutes
at the rate of 50 cc. per minute. At the same time, blood from a normal
person was obtained and similarly treated. The changesin color of the
two samplesafter cessationof oxygenationwere compared.While the normal blood neededalmost ten minutes to return to the previous color, the
blood from the patientin shockwas back to the deepdark color in lessthan
tlree minutes.
Blood samplesfrom subjcctsin shock werc treatedin vitro with unsaponifiablefractions and subsequentlywith oxygenfor five minutes at the
rate of 50 cc. per minute. While the color became red immediately,the
time requiredfor it to return to the previousdark color was entirely different from that of controls. As againsta few minutesfor controls, more
than twelveminutcswerc requiredfor the blood trcatedwith insaponifiable
fraction.From theseexperimentsit appearsthat the dark color of blood
in shock resultsfrom changesin the red cells and not becauseof impaired
circulation, and that these changescan be related to the intervention of
fatty acids.This was confirmed by the fact that the dark blood of shock
patients,if treated in vitro with insaponifiablefractions of placenta,for
instance,losesits characteristiccolor.
Chapter9, Note 4. Induction of Acute Shock
Acute shock can be induced by intraperitonealinjectionsof mixtures
of conjugatedfatty acids.The preparationlargelyusedwas a l07o solution
in oil of fatty acid of cod liver oil conjugatcdthroughtreatmentwith KOH
in ethyleneglycol or in ethyl alcohol.For a rat of 200 grams,8 cc. of this
preparationinjectedat once was ableto inducean acuteshock.
Chapter9, Note 5. Induction of State of Shock
Stateof shockwas inducedby the repeatedadministration
of a mixture
of conjugatedfatty acids obtainedfrom cod liver oil. To insure a progressive systemicabsorption,the preparationwas injectedsubcutaneously.
The
injection in rats of I cc. per 100 gram of body weight of the l07o solution
of thesefatty acidsin oil, repeatedevery hour was seento induce after 3-5
injectionsa stateof shock.The addition of 4% of,sodium thiosulfatein
a doseof 5 cc. per 100 gram of body weightin rats, was seento favorize
the appearanceof this stateof shock.
Chapter 9, Note 6. Influence of Fatty Acids Upon Traumatic Shock
Rats of 250 grams were introducedin the Collip-Noble Drum with
their forepawstapedwith adhesiveand submittedto 500 falls at a rate of 40
per minute.50% died of acuteshockin lessthan two hours.If 2 cc. of a
preparationof I OVo cod liver oil fatty acids in oil per 100 grams of body

680

p . E S E A R c Hr N p H y s t o p A T H o L o c y

weight was injected intraperitoneallyor even subcutaneouslyth hour before the animalswere placedin the drum, more than 5OVoof the animals
died during the trauma itself and the fatality rate in some experiments
approached100%. Bleedingfrom the nose and mouth was dark in color
and smallerin quantity than in untreatedanimals.If the same amount of
the lipoacid preparationwas injected immediately after the animal came
out of the drum, it also increasedmortality within the first two hours. For
some animals,death occurredin a few minutesafter the injection.
Chapter 9, Note 7. Infuence of UnsaponiftableFractions Upon
Traumatic Shock
The influenceexertedby the unsaponifiable
fractionsupon traumatic
shockappearedevidentin rats submittedto 5-700 falls in the Collip-Noble
drum. l07o solutionsin sesameoil of the unsaponifiable
fractionof human
or cow placenta,of eggsor of butter,were usedin theseexperiments.I to
5 cc. of thesesolutionswere injectedintraperitoneally
at differentintervals
before or after trauma. The injection of 2 cc., Vz hour before trauma, rvas
seen to entirely preventlethal shock (0/20) in a group of experiments
wherethe mortalityof controlswas 18/20. The sameresultswere obtained
with 2-3 cc. of the preparations
injectedimmediatelyafter the animalswere
takenout of the drum. Dosesas high as 5 cc. injectedone hour after the animal wasremovedfrom the drum, protectedonly a fewanimals(l l/20) and
generallyonly those without symptomsof shock. Once the symptomsof
shock were present,the effectof the unsaponifiable
fractionswas greatly
reduced.(From 2/20 to 5/20 in diffcrentexperiments.)
Chapter 10, Notc I . Oxalic Index
The need to havc quantitativeinformation about the amount of double
bonds presentin the organismor in its lipoacidshas led to a method of
analysisbasedon the fact that molecularbreakdownor fissionwill furnish
characteristiccomponents.With fissionoccurringat the level of the double
bonds,the fraction correspondingto a conjugateddouble bond will appear
as oxalic acid. The problem was to obtain this fissionwith a carboxyl correspondingto each carbon and without having artificially induced displacementsof the doublebond which is a frequentresultof treatment.
We employedthe following technique.Fatty acids from an organism
or any other preparationwere neutralizedwith the exactamountof sodium
carbonatenecessary.
This amount was established
through the neutralization indexof the substances
to be treated.After sufficientdilution,an excess
of sodium carbonatewas added with the aim of obtaining an alkaline
medium.After bringingthe solutionto 4oC potassiumpermanganatewas
addeduntil further discolorationof the permanganate
stopped,after which
ZOVomore of the amount alreadyused was added.The mixture was kept
refrigeratedat 4oC for 16 hours, after which the excessof permanganate
was reducedby sodium bisulfide.The liquid obtained was filtered and the

': "l i!t

NorEs

681

precipitatewashed.The liquid was extractedfirst with ether to eliminate

the higher fatty acids,after which it was submittedto distillationin order
t o e l i m i n a t et h e v o l a t i l ef a t t y a c i d s .I n t h e r e m a i n i n gp a r t , t h e o x a l i c a c i d
was precipitatcdwith calcium chloride. From the precipitate,the part
correspondingto calcium malonatewas separatedfrom calcium oxalate,
by usingthe differencein solubilityat the boiling temperature.The oxalic
acid was then titrated in the usual manner.The amount of oxalic acid
divided by the quantity of lipoacidsused representsthe oxalic index of the
preparation.Carlos HuescaMejia and Daisy Franco have widely studied
the changesof this oxalic acid index in our laboratories.
Pure nonconjugated
fatty acidstreatedin this manner yield no oxalic
acid. When linoleicacid is conjugated(e.9.by treatingwith KOH in ethylene glycol) oxa.licacid is found in the fissionproductsin amountsthat
graduallychangeas the treatmentcontinues.(Tlnr-e XXXIV)
Trsr-E XXXIV
THr Qurnrtry oF Oxnltc Acro Pnrsexr ARrrn Oxlo,rrtve Frssrol.r,rxp IoolNe
Nur'{san oF S,rvples or LrNor-ercAcro CoN:ucr,tE'DFoR Dlrrenr.Nt PeRroos
or TIlre. (LrNolerc Acro Mlxeo Wrrx Equll Qu,rxrrrresor KOH: Drss o l v e o 5 9 t l N E t H y r - r . N eG l y c r _ r r _Cl o N r r N u o u sT n r . , r r v r l r l N R r ' r r - u x . )
Time
Before treatment
After 30 minutes
"
I hour
"
2 hours
"
4 hours
"
8 hours
"
12 hours
"
24 hours
"
36 hours
"
48 hours
" 144 hours

Oxalic Acid
nrg gnr of Fatty_Acid
0
tt7
205
| 14.2
l 19.4
99.9
92

u5
80
77.3
40.8

Iodine
Number

180
I19.8
l l5.l
94
96
9l
86
8l,7
76
76.5
57.3

The quantitativerelationshipbetweenknown proportionsof conjugated

fatty acidsand the oxalic acid obtainedthroughtheir oxidativefissionhas
beenstudied.
Autolytic changeshave becn found to influencethe natureof the fatty
acids extractedfrom tissues.Studieshave shown that formalin fixation
doesnot significantly
changethe fatty acidspresent.(TnrlE XXXV)
It hasbeennotedthat relativelystrongermethodsof conjugation,utilizing KOH at higherconcentration
and highertcmperature,are necessary
to
conjugatethe fatty acids of normal tissues,than are neededfor the fatty
acids extractedfrom pathologicaltissucs (burn, shock, adrenalectomy,
tumor necrosis)which can readily be conjugatedby much milder procedures.

682

RESEARCH IN

PHYSIOPATHOLOGY

T,rsLeXXXV
Ernecr on FoRveltx UpoN rxr, QulNrtry oF Oxllrc Acro Pnoouce,oBy
Oxro,rtrve Ftssrox on A Mrxrune on CoN.luc,rrep F,rrry Acns. (5 cc.
Alrquors or A MrxruRL, oF CoN.luclrro F,rrry Acros Wenr, MxEo Ix A
Sropprnrp Cyr-rxorn Wrrn l0 cc. oF A 20Vo Fonvol Solurlox. THE CyLrNoen Wrs Snlxen ,rNn S,rprpr-rsWrnr Rrproveo Ar FneeuENT INTERVATJ
Fon DerenMrNArroNon Oxruc Acro.)
Time

Mgm Oxalic Acid/gm

Fatty Acids

Before mixing
After
I hour
"
24 hours
"
48 hours
"
72 hours
"
144 hours

tzt.7
121.8
120.5
120.9
I 18.9
t2t.9

Chapter 10, Note 2, Irradiation and Oxalic Index

Forty male albino rats separatedin groupsof l0 were irradiatedwith
nonfilteredX-rays from a 200,000v. machine,receivingin one sessiona
doseof 1500 r. which is considered
a lethaldose.Similarexperimentswere
repeatedscveraltimes,somewith the radiationdose obtainedfrom radioactivecobalt. The four groups of animalswere first mixed togetherand
thcn separatedin four big cagcs.The animalsand controlswere kept on
PurinaChow and water as libitum.Two of the controls,two of the treated
animals,and any others approachingdcath were sacrificeddaily.
Each dead animal was saponifiedseparately.The total amount of acid
lipids extractedwas analyzedfor conjugatedfatty acids, using the method
indicatedpreviouslyin which the amountof oxalic acid which appearsas
the result of oxidativefissionis measured.The valueswere expressedin
oxalicindex,which corresponds
to the amountof oxalic acid in milligrams
gram
pcr
of fatty acids.Figure [i5 showsthe resultsof two such cxperinlents.
Values found for normailswcre zcro or less than 0.5, but a constant
increaseof the oxalic indcx was noted after irradiation.While irregular
valueswere still seenduring the first three days, all were above 3 on thc
fourth day and continuedto increaseconstantlyafterward.Death occurrcd
when the oxalic indcx reacheda critical point which was found to correspondto l4-17 mgr. of oxalicacid per gram of thc fatty acidsof the entire
body.
Chapter 10, Note 3. Oxalic Index in SublethalInadiation
We studiedthe changesin oxalicacid in animalstreatedwith an amount
of radiationbelow the lethal doses.The oxalic acid rose after treatment
with valuesmuch lower than thoseseenwhen lethaldoseswere used.Three
groupsof ten animalseach were radiatedwith 600 r., and every few days

NorEs

683

two of the animalswere sacrificedalong with two of the controls.Figure 91

showsthe values obtained for these animals.It could be seenthat although
indicesof 6 and 7 were found, thesenever reachedthe critical point of
14, and that the indiceswent down in an irregularfashion.While some animals had values of around 4 and 5 after the tenth day, others had high
values at the same time.
The administrationof polyunsaturatedconjugatedfatty acids (l-2 cc.
daily of a 5Vo oily solution of conjugatedcod liver oil fatty acid) induced
death in a high proportion (16/20) of animals irradiated with otherwise
nonlethaldosessuch as 800 r. In theseanimalsthe oxalic index was high,
often showing values above 17.
Chapter 10, Note 4. Radiation-induced Offbalances
Through the routine analyticaltechniqueused for the study of the
oftbalances,
we studiedthe changesoccurringin a goup of 56 subjectswith
differentconditions,submittedto radiotherapyalone,we tried to correlate
the clinical noxious effectsof radiation to theseoffbalances.

ir-1;-_z-r7

:---'wr==^.<r'i;-zv-Ul^.

\,/-

Frc. 270. L.N., 59 yearsold. Cancerof tbe breast.Mastectomytwo monthsearlier.

No recunence.Treated with radiation, l25r per seanc.Entirely uneventful except
moderateerythema.Minimal or no changesin the analyses.

In this part of the study, we limited ourselvesto the considerationof

the immediatechanges.Figs. 270 and 271 show the analysesin two cases
in which no clinical, local or systemicchangeswere observedduring and
after the radiation. It can also be seen that no changesoccurred in the
analyses;they remained within normal limits during the entire period of
observation.It is especiallynoted that the peroxideswere present,almost
constantlyin high amountsin the urine.
Fig.272 showsa casewho died during the radiotherapeutic
treatment,
probably directly influencedby it. All the analysesshow that a changeto-

684

c , E s E A R c Ht N p H y s r o p A T H o L o c y

ward offbalancetype D had occurred. Among these, we want to single

out the urinary surfacetensionand the urinary chloride index. both evidencingvery manifestchangestoward the patternscorrespondingto type
D. The peroxideswere on the contraryalmostcompletelymissingfrom the
urine.
Anothercase(Fig. 273) had severeclinicalreactionto radiationwhich
persisteduntil the deathof the patient.Again, we judged rheseeffectsof
radiationmainly throughthe changesoccurringin the two analyses,
urinary
surfacetensionand chloride index, which show the same shift toward a
strongoffbalancetypeD. The pcroxidereactionbecamenegativeat the end.

_-^---^
. -Vf

.,^t--V\--:

1r.\

F t c . 2 7 l . K . L . 5 2 y e a r so l d . C a n c e ro f t h e h y p o p h a r y n xT. r e a t e dw i t h 3 0 0 r f o r e a c h
s e s s i o n .T h e h i g h s p e c i f i cg r a v i t y s h o w s a t e n d e n c y l o w a r d t h e t y p e A . a m o n t h
a f t e r t h e c o m p l e t i o no f t h e t r e a t m e n t .
W h i l e s p c c i f i cg r a v i t y a n d S T s h o w a s l i g h t o f f b a l a n c c[ ) d u r i n g t h e t r e a t m e n ta n d
t h e p H a m a n i f e s tc h a n g et o w a r d t h e t y p e D , f o l l o w i n g t h e r a d i a t i o n a l l t h e a n a l y s e s
show a changetoward the type A, one month after completion of rhe radiation.
P e r o x i d e sp e r s i s t e di n t h e u r i n e . C l i n i c a l l y t h e e n t i r e e v o l u t i o n w a s e v e n t l e s s .

When the differenttestswere discussed,

we mentionedthat each one
of the analyscu
s s e df u r n i s h c di n f o r m a t i o nc o n c e r n i n g
c h a n g e sw h i c h t a k e
place at a specificlevel of the organization.This would explain rvhy the
noxiouseffectsare seento be seriouswhen the changestake place conc o m i t a n t l yi n d i f f e r e n ta n a l y s e st h
. a t i s . a t d i f f e r e n tl e v e l s ,i n d i c a t i n gr h u s
a more completeoffbalance.When this concomitance
does not exist,rvhen
t h e a b n o r m a lp a t t c r n sc o n c c r no n l y o n e a n a l y s i st,h e c l i n i c a l m a n i f e s t a t i o n sa r e s e e nt o b c l e s ss c r i o u sT. h i s w a s s e e nt r u e i n t h e c a s es h o w ni n
F i g . 2 7 4 .T h e i m p o r t a n c o
e f t h i sc o n c o n r i t a n ci ne t h ec h a n g e p
s r e s e nits s e e n
i n F i g . 2 7 5 . l n t h i s c a s e ,a l t h o u g hm a n i f e sct h a n g e sc o r r e s p o n d i ntgo t y p e
D are seenfor someanalysis,they do not coincide.The pcroxidereaction
in the urine is constantlypositive.This seemsto permit the patientto rvithstand the noxious effect of radiation.The evolution of the chansesinducedby radium applicationis shownin Fig, 276.

.:JJ.

J-^v

The possibitityto evaluatethrough urine analyr., ;t:t-:"r'.r.'.1:

o f r a d i a t i o n sh, a s a p p e a r e de s p e c i a l l yi m p o r t a n tf o r t h c p r e v c n t i o na n d
even treatmentof the serious inconveniences
during radiation therapy.
B y o b s e r v i n gt h e c h a n g e si n t h c a n a l y s e sp.a r t i c u l a r l yo f t h e u r i n a r y s u r face tensionand chloride index, valuableinformationcan be obtained.
p e r m i t t i n go n e t o g u i d e t h e a p p l i c a t i o no f t h e s et h e r a p e u t i ca g c n t s .B y
being easily and continuouslyinformed about the occurringchanges,we
need no longer considerthc amountsof radiationto be administeredas
standardvaluesfor each patient.Throughanalyticallyguidedradiotherapy
o n e c a n r e p l a c et h c c o m m o n p a t t c r np r c s c n t l yu s c d f o r a l l p a t i c n t sb y

L .r .

t r anao Ltt2Jtr r tl 6:/l dZ 2 t+S

"iF:
l

-/Y

^ L : .
u t t ; -

f---\'-^r

\-

\2---=^-f

zr'r

al'.1
la
8

len0x.

F r c . 2 7 2 . L . W . , 5 8 y e a r s o l d . C a n c e r o f t h e l u n g . A f t e r r e c e i v i n go n l y 1 0 0 0 r ,v c r y
w e a k . M u c h w o r s e a f t e r , w i t h v e r y r a p i d d o w n h i l l c o u r s e .A f t e r t b e t r e a t m e n t o n
3 / 5 , t h e p a t i e n t e n t e r e di n t o c o m a a n d d i e d 3 / 7 . T h e e x t r e m e l yh i g h v a l u e s f o r C l I
a n d h i g h s p e c i f i cg r a v i t y c h a r a c t e r i z et h e a n a l y t i c a lc h a n g e s .I t i s i n t e r e s t i n gt o n o t e
tbe negative reaction for peroxidesfor almost the entire time.

IiIIEiI
ESEEES

i:
E5$ E

I:;Ei!F
l = &E ; g ;
I E fr U;T 5

i : 3gE ;
s : ; E!:

E 8B : EFE
H Ers5s s

l6 c: I : i 0

F . E . :E t

i;r!r::

;igEigfi

NorE s

697

individualizedtreatmentsadapted to the need and the responseof each

subject.
Continuouslyfollowed analysespermit their utilization as a guide for
more general application of radiotherapy.When the surface teniion and
chloride index remain within normal vilues, treatment can be continued
with administrationof dosesabove those originally intended,without any
danger of serious noxious effects.A change towird low surface tension

1iJ0r.

F r c . 2 7 4 . C a n c e r o f t h e c e r v i x i n t e n s i v e l yi r r a d i a t e d . A l t h o u g h c h a n g e s o c c u r i n
the analyses,the p""ssges in offbalance D do not coincide in the difterent tests. With
the continuation of the treatment, the patient passcs in offbalance A. No clinical,
noxious manifestations were seen.

below 65 dynes/cm. or high chloride index above 5, representsa warning

which should not be ignored. The treatmentshould be discontinued,the
dosagereduced or the sessionsmore widely spaced,even if the desired
radiationdosehas not yet been attained.Concomitantchangesof the analyses should constitute a serious warning even when the general clinical
conditiondoes not indicateany abnormality.The bad prognosisof persistent strong "D" offbalanceduring radiationis relatedto the progressionof
the anomaly as describedin the experimentalstudies.For this reason,a
persistentstrong offbalanceD seen for the urinary surface tension and
chloride index indicatesthe need for the administrationof lipoids with
positivecharacter,even if the clinical manifestationsare not too serious.

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lJr

NOTES

689

F t c . 2 7 6 . F . 8 . , 6 4 y e a r s o l d . C a n c e r o f t h e f u n d u s o f t h e u t e r u s ,t r e a t e d w i t h 4 5
mgr. radium for 122 hours in situ. with a total of 5490 mgr. hours.Felt subjectively
w e l l a f t e r r a d i a t i o n w i t h o u t a n y c o m p l a i n t .T h e a n a l y s e ss h o w a m a n i f e s t c h a n g ef o r
almost one week from the type A to type D. For the S.T. it starts 2 days after the
i n s e r t i o no f r a d i u m w i t h t h e v a l u e sd r o p p i n g f r o m 7 0 t o 5 5 . F o r t h e p H , t h e c h a n g e
s t a r t e da f e w d a y s l a t e r w i t h t h e v a l u e sp a s s i n gf r o m 7 . 8 t o 4 . 6 . T h e c b a n g e so f C l I
s h o w v a l u e sa s h i g h a s 7 w i t h o u t h o w e v e r ,h a v i n g t h e c h a n g e sc o i n c i d ew i t h t h e p H
a n d S . T . t 6 d a y s a f t e r r a d i u m w a s t a k e n o u t , t h e a n a l y s e sw e n t t o p r e v i o u s v a l u e s ,
cxcept the peroxides,which remain present.

Chapter I l, Note 1. CarcinogenicActivity of Urethane

The interestingrcsearchof Bercnblumhas broughtan importantcontributionnot only for the largelydebatedrole of urethaneas carcinogen,
but alsofor the problemof carcinogenesis
in general.The fact that croton
oil, applied to the skin, inducesthe appearanceof malignanttumors in
animalspreviouslyfed with urethane,concordslargelywith the conceptof
plural changestaking placein carcinogenesis.
The analysisof the influence
exertedby carbamicacid upon amino-acidswould place the intervention
of this agentat the first membersof the biologicalrealm. It can thus be
seenthat the bond betweenthe amino-acidgroup and the carboxyl and
aminegroupsof carbamicacid occur in a way similarto that which occurs
betweentwo amino-acidswith the big differencethat in the first case it
w o u l d r e s u l ti n t h e a p p e a r a n c o
e f t h c C N C N f o r m a t i o n .( F i g . 2 7 7 ) A s
a
b
o
v
e
,
t
h
i
s
mentioned
C N C N f o r m a t i o n r e p r e s e n t st h e g r o u p w h i c h
the first biologicalentity.Thc placc of this CNCN group, not
charcterizes
at theend of the moleculeopposedto the carboxylas in the alkalineaminoto thc bond which resultsin polymcrs,rcpreacids,but as corresponding
sentsthe anomaly,which accordingto the work hypothesiswe advance.
would correspondto the first cancerousentity.The fact that the carcino-

690

nesEARcH rN PHYSIOPATHOLOGY

0
I

NHr
I

C - S l+ H 0 - C - C - R +
t

ltl2

-C-M

HNA

groupleadsto the
Frc. 277.The bondbetween
acidandan amino-acid
thecarbamic
group.
formation
of theNCNC
genicactivityof urethanetakesplace at the lowestlevelsof organization,
explainsthe necessitythat a certain time separatesits interventionfrom
that of croton oil, which would act only at the higher levels,probably
inducing the passagefrom noninvasiveto invasivephase. This timc is
necessaryfor the first cancerouschangesto build up the seriesof cancerous
hierarchicentitiessincc the cocarcinogcn,
croton oil, would act only in
those more evoluted cancerousentities.In experimentsin course, the
passageof the noninvasiveurethane-induced
carcinogenesis
entitics into
invasivecancer,is successfully
inducedby preparationsof unsaponifiable
fractions.
Chapter 11, Note 2. Constitution of Viruses
It is superfluous
to emphasize
the interestwith which virusesare being
studiedfrom all points of view. Their role in carcinogenesis
has placed
them in the limelightof cancerresearch,and any contributionconcerning
their constitutionor activity is of great interest.A much debatedinitial
problem concernsthe nature of viruscsand their place among the other
entities.(293)
Two fundamentalgroupsof constituents-DNA and proteins-(301.
289) havebeenrecognized
to takepart in theformationof theviruses.These
two groups could be separatedand reunited, reproducingthe original
virus with all its characters.Futhermore,new virusescould be created
when fractionsresultingfrom differentviruseswere bound together.(289)
However,the fact that a part of the virus, the DNA fraction,was seento
be furnishedby the constituents
of the host cell, and the protein directly
by it ( 312), has raisedthe <;uestion
of the natureof the virus itself. Some
workers have gone so far as to seethe virusesas parts of the constituents
of the cells. By consideringthe virusesin the conceptof the hierarchic
organizationpresentedabovc, a new aspectemerges.
Accordingto this organizational
concept,a virus representsan entity
that has reacheda certainstep in the hierarchicevolution,and remained
Like all entities,a virus can be contherethroughoutits individualization.
ceivedto be formed by a principal part bound to a secondarypart. the
ensuingentity limited by a boundaryformation.The principal part would
be formed by a group of immediatelyinferior entities in the hierarchic

NorES

691

scale.In thc caseof viruscs,suchinferiorentitieswould be formed by what

we could call "proviruses,"which correspondto characteristic
DNA formations. The principal part of the virus would be formed by the grouping
togetherof provirusentities.proper to each virus. The secondarypart is
conceivedto resultfrom the immediateenvironmentof theseentities,taken
directly from the host's own protoplasmaticor nuclear formation in which
the principal parts are presentas free entities.This secondarypart is
represented
by the protein fraction furnishedas such by the invadednucleus or cell protoplasmaticformations.This protein fraction conserving
its characteristics
can be recognizedand identified.
Having nuclear formations,nuclei and protoplasmaticformationsas
their environment,the principalpart of the virus, the proviruses,multiply
as proper hierarchiccntities.These proviruseswill leave the host usually
when the cell bursts. bound this timc to the proper secondaryparts
directlyfurnishedby the host entity. Under theseconditions,the principal
part multiplied in the protoplasmaticformationsor in the nucleusand
the secondaryparts, furnishedas such by the host, would form an imm e d i a t e l yh i g h e re n t i t y ,t h e v i r u s .I n t h e m u l t i p l i c a t i o on f t h e v i r u s ( 2 9 9 ) ,
the patternfollowed is the same as that of other hierarchicentities.This
has to occur in the proper environmentwhich, for the viruses,is the
immediatelyhigherhierarchicentity,the nuclearlevel.This is represented
in microbesby the individualitself,and in cellsby the nucleusor by the
protoplasmatic
formations,which we considerto belongto the nuclearlevel,
due to their ribonucleo-proteins.It is in these nuclei or protoplasmatic
formationsthat the virusesmultiply. This explainsthe developmentof
virusesin cellsin compactgroups,which would correspondto parasitated
protoplasmaticformations and not in a diffuse form in the cytoplasma.
The virus losesits secondarypart upon entcringthc entity where it will
multiply, but will take it back when it leavesits host, becomingagain the
entity, virus. The parasitatcdcntity has contributedtwo parts to the multiplicationof the virus----one,
indirectly,by furnishingmaterialwhich will
be utilized by the provirus and transformedinto its spccificDNA, and
two, the secondarypart, directlyformed by its own proteins.Like all the
secondaryparts, that of the virus directlyfurnishedby the host will surround the group of provirusesforming the principal part.
Chapter 12, Note 1. Lipids and Cytolytic Activity of Sera (245,246)
The fact that cancer cells are found circulating in the blood without
throughoutthe body, as might be expected,has made
inducingmetastases
the meansby which the organismrids itself of
variousworkersinvestigate
these cells and which might representpart of a general processthrougb
which the organismcould fight cancer.Researchto confirm the capacityof
blood sera for influencingthe lysis of cancer cells has been carried on in
our laboratoriesby Robcrt Willhcim and co-workers.Sera of both normal individualsand cancerpatientswere found to induce lysis of cancer
cells in vitro. Ehrlich, Krebs and Sa 180 ascitesin mice were used,as well

692

R I:Sl:ARC ll

I N

Pll I'Si()l,A THOt,O{iY

as cclls obtitincdfronr solid turnorsuith cnccphaloidcharactcr.Tumors

obtainedfront biopsiesor autopsies
of hunranbeingsalso werc used.Lytic
activitywils dcterminedthroughthe dilution of scra u,hichluter rvasmixed
r v i t ha s u s p c n s i oonf a k n o r v nn u m b e 'or f c c l l s .A f t c r i n c u b t t i o nf o r o n c h o u r

;S,tS.

"iil

aD.
I

- ?

Sr

Fu; l l l { . I r t i e r r c t r o no f h u r n a ns e r u n tu p ( ) nS u 1 t i Oa s c i t c sc c l l s .
{I)
t1t
(3l
( : 1)
[ - \)
( ( r)

N o r n r a l S . ' \ l t t ( } ( ' c l l r X J t } ( 1n. o t s t u i n c d .

L l t i c e t l c c t o f S c r u n ro n S A 1 8 0 X 1 0 0 . n o t s t a i n e d .
N o r n r r r iS A l l { { ) ( ' c l l s . X l ( X ) ( } G
, i c n r s as r a i n .
I . l l i e c f i e c t o f S c r r r n ro n S . , \ l l { t } ( ' e l l r X l 0 0 t ) . C l e r n \ i i \ l a i n
N o r r r r i r lK r c h r ( c l l r X l ( X X ] .( i i c n r r a r t r r r n .
I . 1 t i c e f f c c to f S en r n r r r n K r e l r s( e l l s \ 1 0 0 0 .C i e n r s as t a i n .

. llli

693

NOTES

{fG

'

t-:,:
ti

''
f
4$

;s
d'

s*

.:'rfi."

;fi

,lr' l
t,

.i

"r i1r

&il'_

l,

$*

i4

*' 6 ;S.n

'-

ilW

Frc. l?9.
( 7|
(ttl
(9)

' , i

f2.',

it

Con t inuatic'rn-'.
N o r m a l S A l l { 0 ( c l l r X l 8 { ) 0 .( i i e n r r i t \ t i l n .
C a r c i n o l y ' t rcct l c c t o f S e r u mo n S , { c c l l s X l t { 0 1 }(.i i c r n s l r t ; r i n .
N o r r n a l l - 1 r i s o f S , \ l E { ) ( e l l r ; r s c o n l r i . r il n P o l r r i r c c h . i r r d el ns h r b r t i o n S t u d i e so n C ' l r c r n o l ir t s , X . - l ( X )n. t r t s l i i i n e d .
( l 0 ) E l l c c l o n C ' a r c i n o ls\ i r h y t h e P o l l : a c c h a r i d eD e. ' t t r a nX 1 0 0 , n o t * t a i n e d .
i l l r E f f e c t o f l - c v a n o n ( a r c i n o l v s i r .N t ; t e t h c c l c a r l y d i s t i n c t o u l l i n c o f u n a f f e c t e dc e l l s . X l ( X ) . n { ) t r t a i n e d .
i l l ) E f f e c l o f i l u c i n o n ( - u r c i n o l y r i rX 3 0 0 , n o t s t i i i n c d ( e v i d e n c eo f i n h i h i t e d
Lysis.

694

s , E s E A R c Hr N p H y s r o p A T H o L o c y

at 37"C the product was examinedand the cells countedin an adequate

chamber.(Fie.278)Relativelybig variationswere seenin differentindividuals, the resultsbeing negativefor I :1 dilution in some casesand positive
in 1:32 and even higher for other individuals.No correlationcould be
establishedbetween this lytic power and the clinical condition of an individual.
With R. Willheim and M. Auber (247 ) we showedthat the addirion
of a suspensionof unsaturatedfatty acids is able to induce such a lytic
property in sera without this capacity,while the addition of insaponifiable
fractionsor of cholesterolinhibitcd lyric activiry.With R. Willheim and
M. Auber we (248) have investigatedthe problem of the correlation
betweenthe lytic activity and the structureof differentfatry acids and their
sodium soaps.The higher membersof the fatty acid series have shown
intenselytic activity. It could be shown that the lipoidic characterof the
fatty acidsincreasestheir lytic capacity.The value of the lipoidic character
of the fatty acidsusedhas appearedwhen memberswith the same number
of carbons but having an hydroxyl or carboxyl in their molecule, were
tested.With disappearance
of lipoidic character,lytic property disappeared.
The researchcoverednot only cancercellsbut also liver, red blood cells and
lymphocytes.It could be seen that a correlationexists between the rapid
growth characterof thesecells and the capacityof the fatty acids to attack
them. It is interestingto note that cancer cells treated in vitro with fatry
acids having lytic activity no longer produced cancerousgrowths when
transplanted
to animals.(2a9)
Chapter 12, Notc 2. Fatty Acids Transportationin the Blood
It appearcdespeciallyinteresting
to studythe distributionand transport
of polyunsaturated
fatty acidsin the body in view of the fact that parenteral
administration
of the acid lipidic fractionof variousorgansto subjectswit-h
acutepain had an effectwithin only a few minutes.The rapid action was
independent
of the nature of the inducedchange,that is, decreaseof the
intensityof pain of acidpattern,and increase
of pain of an alkalinepattern.
This effect in opposite directions,occurring after the same short interval
and seenin hundredsof cases,eliminatedthe possibilityof a psychological
factor, as suspectedat the beginning.
The fact that the changeoccursat the level of the painful lesionitself
raisedthe questionof rapid transportbetweenthe injection site and the
lesion.In order to investigate
it, we usedtwo fatty acid preparationscontaining easily identifiablesubstances,norbixine and polyconjugatedfatty
acids.Norbixinc could be identifiedby its characteristic
color while the
polyconjugatedfatty acidswere identifiedby their specificcurvesin spectral
analysisin ultraviolctlight.
Adult New Zealandrabbitswere injectedintraperitoneallywith 8 cc. of
0.3% solutionof bixine in sesameoil. The injectedanimalswere bled at
differentintervalsby heart puncturc.The red cellswere separatedfrom the
plasmaby immediaterapid centrifugation.
Each fraction,plasma and red

NorEs

695

cells,was hydrolyzedseparatelywith 5 o/aKOH, The acid lipidic fractions

obtainedas a solutionin benzcncwas passcdthrough a chromatographic
column with alumina,Bixine was casilyrecognizcd
becauseof its red color.
After elution with chloroformthe amount presentwas determinedphotometrically. Similar experimentswere made by using a l}Vo solution of
eleostearicacid in oil, The acid lipidic fractionsobtaincd separatelyfrom
the red cells and the plasmawere submittedto spectralanalysisand the
presenoeand amount determinedby the characteristicpeaks.Both norbixine and eleostearic
acid were seento appearin the rcd cellsin lessthan
two minutes,the amountincreasingrapidly.
A marked diflerencewas found betweenthe amount of thesefatty acids
in the red cells and in the plasmaof the same blood, for all the samples.
The red cellscontained5 to 6 timcs as much of the injectedlipids as the
plasma.This unequaldistribution,also secn for other highly unsaturated
fatty acids,indicatesan importantphysiological
rolc for the rcd cellswhich
has not been recognizedbefore.Red cellsappearto bc preferredvehicles
for transportingpolyunsaturatcd
fatty acids throughthe blood.
Chapter 12, Note 3. Coniugationl\{ethod
Spectralanalysisof a mixture of fatty acidssuch as obtainedfrom cod
liver oil has shownthat prolongedconjugationsometimesis detrimentalfor
somemembers.Prolongationof conjugationwas found necessary,
however,
sinccthe memberswith a lower numberof conjugateddoublebondsneeded
more time to appear.We investigated
the factorswhich would intervenein
thesechanges.High temperaturcwas seen to affect the polyconjugated
formations.With ethylcneglycol or glycerolas solvent,conjugationtook
p l a c e r a p i d l y , b u t t h e p e a k s o f t e t r a e n e sa n d e s p e c i a l l yp e n t a e n e sa n d
'fhis
were sccn to go down rapidly.
hexaenes
was not sccn to occur if the
temperatureof conjugationwas lower. In this last case, the conjugation
was seento take much more time. This study led us to use ethyl alcohol
a s a s o l v e n t .M a x i m u m c o n j u g a t i o nh o w c v e r ,r c q u i r e d a l o n g c r t i m c ,
usually around 100 hours. With this method we could obtain from the
same preparationsmuch highcr amountsnot only of conjugatedpentane
a n d h e x a n eb u t a l s od i e n ca n d t r i e n e (. F i g . 2 8 0 )W e a l s ou t i l i z e dt h e s a m e
methodfor analyticalpurposeswith the samegood results.
Chaptcr 12, Note 4. QuenchingAction and Anti-CarcinogenicEffect
of Conjugated Fatty Acids
We haveinvestigated
the influenceexertedby differentfatty acids,conjugatedand nonconjugatcd,
upon variouscarcinogens,
In a first group of
experimentswe studiedthis influencein vitro, and chosethe quenchingof
of the carcinogenas criterionof activity.
the fluorescence
n ith C. HuescaT h i s p a r t o f t h c r c s e a r c hw a s m a d ei n c o l l a b o r a t i o w
Mejia and P. Teitelbaum.
The quenchingeffecthas beenstudiedas follows:

696

x E S E A R C HI N P H Y s r o P A T H o L o c y

The fluorescence
of carcinogenichydrocarbonsis measuredby meansof
the fluorcscentattachmentto the Beckman spectrophotometer
using a
wavelengtharound365 mp. The sensitivityof the apparatusis adjustedto
show a value of 100 for the fluorescentlight utilizing the concentrationof
the hydrocarbonhaving the maximum fluorescence.
The carcinogenichydrocarbonsare dissolvedin alcohol,iso-octans
or cyclohexane,
the last two
being purified and thus renderedoptically inactive by passagethrough a
silicacolumn.
The fatty acidsare dissolvedin varyingdilutionsin the samesolvents
and addedto the solutionof hydrocarboncarcinogenwhich has been previously chosento give a fluorescentvalue of 100. The fluorescenceof the
mixture is immediatelydetermined.The quenchingeflect of the fatty acid
is shownby the percentof the residualfluorescence
of the carcinogenwhen
mixed with differentsolutionsof a mixture of conjugatedfish oil fatty acids.
Tesle xxxvl
showsthe quenchingeffectof mixturesof fatty acidsconjugatedby treatmentwith KoH upon differcntcarcinogenicand related
hydrocarbons.

,t-tr-./"'

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lave Lemth!rt)

tb

'I'he
F t c . 2 8 0 . C ' o n j u g a t i o nm e t h o d .
use of ethylic alcohol as solventfor conjugation
c h a n g e st h e a m o u n t o f p o l y c o n j u g a t e dm e m b e r s o b t a i n e d f o r c o d liver oil fatty
a c i d s ( a ) a s c o m p a r e d w i t h t h o s e o b t a i n e dw h e n t h e c o n j u g a t i o ni s made at higbcr
t e m p e r a t u r e su s i n g e t h y l e n e g l y c o l(,b ) g l y c e r o l o r o t h e r s o l v e n t s .
Dilution 0.002t; in ethvl alcohol.

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N O

>\t.C.

NOTES

699

ConjugatedFatty Acids and Quenching

Nonconjugatedfatty acids such as linoleic acid, linolenic acid, arachidonic acid, mixed fatty acids from body liver oil and cod liver oil have a
limited quenching action. Conjugateddienes such as isomers of linoleic
acid or conjugatedtrienes such as eleostearicacid obtained through conjugation of linolenic acid or extractedfrom China wood oil also have a
Iimited quenchingeffect upon hydrocarboncarcinogensand related compounds.(TlnI-r XXXVII) The sameis true for mixturesof conjugated
dienesand trienes.

0 . 0 0 2 ii n e t h y l a l c o h o l , / e t h y l

c
o

c
I

ra

d a v eI e n o t h( r y )
F t c . 2 E l . S p e c t r a la n a l y s i so f c o n j u g a t e dc o d l i v e r o i l f a t t y a c i d s s h o w s t h e p r e s e n c e
o f c o n j u g a t e dd i - , t r i - , t e t r a - ,p e n t a - a n d h e x a e n e s .

Fatty acid mixtures having conjugateddi-, tri-, tctra-, penta- and

h e x a e n eass s h o w nb y s p e c t r aal n a l y s i (sF i g . 2 8 1 ) h a v e b e e nf o u n d t o e x h i b i t a h i g h d e g r e co f q u c n c h i n ga c t i v r t y( F i s . 2 8 2 ) ( T n s l e X X X V I I )
when mixed with hydrocarboncarcinogens.
The quenchingaction of fatty acids upon the fluorescenceof hydrocarbon carcinogensappearsto be nonadditive,When the incident ray is
passedfirst through an 0.2Vo solution of conjugatedfish oil fatty acids in
alcohol and then through an 0.012% solutionof methylcholanthrene
in
is 8lVo. When the
alcohol in separatevessels,the residualfluorescence
sametwo solutionsare mixed tosetherin one cell. the residualfluorescence
is lL.2%.

700

xESEARcH rN pHysropArHoLocy

The relationship
of the differentconjugated
membersto the quenching
effecthasbeenstudied.Fatty acidmixtureshavingdiflerentproportionsof
isomerswith 2, 3, 4, 5 and 6 conjugated
doublebondswere obtainedby
conjugation
or by treatmentof conjugated
mixtureswith heat, oxygen,
0uenchino
effect

o
u
c
O
IJ
vl

a,
L

o
t
l!

bt

C o n c e n t r a t i oonf t h e C o n . i u o a t F
atty Acids
ed
F I c . 2 8 2 . Q u e n c h i n go f f l u o r e s c e n c e
o f a m e t h y l c h o l a n t h r e n e( . 0 0 6 2 9 b) s o l u r i o n i n
alcohol by different concentrationsof conjugated fish oil fatty acids.

chlorine or sulfur. Changesin the proportionsof the di-, tri-, tetra-, pentawere followedby meansof spectralanalyses.
and hexaenes
The changesin
the heightof the peaksin thesecurvescorresponding
to the differentconjugated polyeneswere then comparedwith the changesin the quenching
fatty acid mixtures.Figure 283 showsthe speceffectof the corresponding

N o r Es

701

tral analysisof samplesobtained at various intervalsduring the action of

oxygenupon a mixture of conjugatedfatty acids.As seen,oxygeninduces
unequalchangesin the heightof the peaksin the curvesof which correspondto di-, tri-, tetra-,penta-and hexaenes.
Fig. 284 showsthe quenching
activityof the mixtures.It can be seenthat a parallelismexistsbetweenthe
relativeproportionsof the tetraeniccomponentand the quenchingactivityof

:
6

2
6

e
F

ta

lave

Length

Crt't')

F t c . 2 8 3 . C h a n g e si n t h e a b s o r p t i o ns p e c t r ao f a m i x t u r e o f c o n j u g a t e df i s h o i l f a t t y
a c i d s i n d u c e d b y t r e a t m e n t w i t h o x y g e n . T h e t r e a t m e n th a s a g r e a t e r e f f e c t o n t h e
h i g h e r u n s a t u r a t e dm e m b e r s ,w i t h t h e p r o p o r t i o n o f t r i - , t e t r a - , p e n t a - ,a n d h e x a e n e s
d e c r e a s i n ga s t r e a t m e n tc o n t i n u e s a
. s s e e nb y t h e r e d u c t i o ni n t h e h e i g h t o r e v e n d i s a p p e a r a n c eo f t h e p e a k s .A f t e r l 2 h o u r s o f t r e a t m e n t ,t h e c o n j u g a t e dd i e n e sa r e t h e
only ones not yet influenced.
D i l u t i o n0 . 0 0 2 9 i i n e t h v l a l c o h o l .

t h e m i x t u r e s ,I n t h i s e x p e r i m e n ti t a p p e a r st h a t t h e q u e n c h i n ge f f e c t
could also be relatedto the presenceof conjugatedpentaenes.Evidence
availablefrom othcr cxpcrimentsdo not, howcvcr,suffhciently
supportthis.
We studiedin a similar way the effectinducedby the treatment-with
sulfuricacid-of a mixture of conjugatedfatty acidsof cod liver oil. Fig.
285 shows part of the occurringchangesand Fig. 286, the quenching
effect.
Similarly,wc studiedthc changesin the quenchingeffect during the
conjugationwith KOH of cod liver oil fatty acidsin ethyl alcohol.Fig,287

702

RESEARCH IN

PHYSIOPATHOLOGY

R e s i d u aF
l luorescence

'-"'
----- ---

total mixture
234dienes
2 7 0t r i e n e s
3I4 tetraenes
c46pentaenes
37iihexaenes

c
o

E
c
f!
L

;a

n 0u r s
F r c . 2 8 4 . T h e r e l a t i o n s h i pb e l w e e nc h a n g e so f t h e d i - , t r i - , t e t r a - ,p e n t a -a n d h e x a e n i c
peaks as found on spectral analysis of samples of conjugated fish oil fatty acids
treated for different lengths of time with oxygen and the quenching effect of tbe
same samples.A close parallelism exists between the decreascin the proportion of
t e t r a - a n d p e n t a e n i cp e a k sa n d t h e q u e n c h i n ga c t i v i t y o f t h e m i x t u r e .

showsthe conjugationeffectand 288, the quenchingeffectof the preparation at differentmonlents,in variousdilutions.

The entire problem was simplified by studying a pure conjugated
tetraene.We have obtainedpure tetraenicparinaric acid from akariton fat

NOTES

703

of Parinarium laurinum seeds.In addition, we have preparedalmost pure

tetraenesutilizing the techniquedescribedby Maury, Brode and Brown.
Unfortunately,with the last method, the results weie less favorable, the
proportion of tetraenesbeginningto decreaselong before the conjugated
pure tetraenicconjugated
dienesand trieneshave disappeared.
has
shown that the quenchingaction is relatedalmost entirely to the "iia
tetraenic
componentalone, and in a mixture it is largely parallel to the content in
conjugatedtetraenicfatty acids,Fig. 289 showsthe quenchingcurve inducedby parinaricacid.

f a v e L e n g t h( 1 )

F l c . 2 8 5 . T h e c h a n g e si n t h e s p e c t r a la n a l y s i so f a m i x t u r e o f c o n j u g a t e dc o d l i v e r
o i l f a t t y a c i d s , i n d u c e d b y t h e t r e a t m e n tw i t h s u l f u r i c a c i d . T h e t r e a t m e n t l e a d s t o
u n e q u a l d e c r e a s ei n t h e a m o u n t o f d i f f e r e n t c o n j u g a t e dm e m b e r s .O n l y t w o c u r v e s
a r e s h o w n : a t t h e b e g i n n i n go f t h e t r e a t m e n ta n d a t 2 6 0 m i n u t e s .
Conjugated Fatty Acids and Induced Carcinogenesis

We have investigatedthe influenccexertedby the fatty acids---<onjugatedor not, and their mixture,upon the inductionof tumors by carcinogens.From the variousexperiments,
somewere eliminated,either because
the dose of methylcholanthreneemployed did not produce tumors in a
sufficientnumber in control animalsto permit any conclusivecomparison,
or the deathrate from intercurrentcauseswas abnorrnallyhigh so that the
entire experimentshad to be discarded.
The experimentsthat were satisfactorilycompleted are summarized

'104

nESEARcHrN pHysropATHoLocy

in the followingthree tables.In the first group of experiments(Teau

X X X V III),4 g ro u p se achcom posed
of 40 adultSwissmice( 20 maleand
20 femalesin eachgroup) wereemployed.
Each animalreceivedin the
ri g h t fl a n k a si n g l esubcutaneous
injection0.2 mg. of methylchola n-

g
o
o
o

i\ ..

trlene

Li
c(rJ

r ng
Cuench

ie

c
c

.D
f
cl
t
f

.o

l ' ' in u te s
F r o . 2 8 6 . T h e c h a n g e si n t h e q u e n c h i n go f t h e t r e a t e d m i x t u r e p a r a l l e l t b e c h a n g e s
ioduccd in the amounts of conjugatedtetra-, penta- and bexaeoes.

NorEs

705

threne as a 0.2vo solution in tricaprylin.These animals also received

subcutaneous
injectionsof a mixture of fatty acidsextractedfrom cod liver
oil, or a mixture of cod liver oil fatty acids conjugatedby treating them
with KOH. The fatty acids were administeredas a 57o solution in cotton-

/1,

ir[

l^

, i
/ \'.^,
f\

,i..'\.

\.

,,

lr^tr,,/
t
t
r .
i r

,,

,r',/
\
i

"-'...j'

ll

,i ti

tr '\.

,',

".,'

'

\ \

\ \ "(a)
\

\r(b)

t
I
I
I
I
I
I

'\f
t

'r,'

l'-'

,!

have

Lenolh (np1

F t c . 2 8 7 . S p e c t r a la n a l y s e so f f a t t y a c i d s o f c o d l i v e r o i l , t r e a t e dw i t h K O H i n c t h y l
a l c o h o l .T h e y s h o w t h e a p p e a r a n c eo f h i g h a m o u n t so f t c t r a - , p c n t a - a n d h e x a e n e s .

seedoil. Animals trcatedwith fatty acidsreceived0.3 cc. of this solution

in the contralateralside twice a week for three months.The control animals receivedthe same volume of cottonseedoil in the same number of
injections.In addition,one group of animalstreatedwith the conjugated
fatty acids receivedfour injections during the two weeks preceding the
Trst-e XXXVIII
Treatment

Died
Without Tumors

Tumors

l0
3

t2/3O
t8/37

40
48

7/34

20

3,/26

II

Cottonseed oil-controls
Fatty acids from cod liver oil
Conjugatedfatty acids from
cod liver oil
Conjugatedfatty acids from
cod liver oil ( + )

1 A

%
With Tumors

+ Received4 injectionsof fatty acids before methylcholanthrenewas administered.

706

RESEARCH IN

PHYSIOPATHOLOGY

\\.\
\

q,
(J
c
qJ
a

4o

L
o
t!

5'o

q,
-ol

o,
E

}{

C o n c e n t r a t roof nt n e f a t t y a c r ds o l s .
F r c . 2 8 8 . C h a n g e si n t h e t o t a l q u e n c h i n gc a p a c i t yo f s a m p l e so f c o d l i v e r o i l d u r i n g
i s o m e r i z a t i o nw i t h K O H i n e t h y l a l c o b o l . W h i l e t h e q u e n c h i n ge f f e c t i s r e d u c e d even for high concentration-for the sample having only 15 hour of conjugation, it is
high for that obtainedafter 24 hours. It remainsalmost thc same for the samplc
after E4 hours of conjugation. The quenching appears related to the presence of
c o n j u g a t e di s o m e r s ,w i t h 4 o r m o r e d o u b l e b o n d s .

:,

:,

I.

.;,

:t,tti

NOTES

C
C'
v,
o

707

&

E
6
c

.rt
L
F

la

.12

,62

.ott

,.b-

.@t

.@

,<b.t

!{)ol'

Concentrat
ion
Flc. 289. Quenching effect of parinaric acid upon the ffuorescence of methylcholanthrene. The relationship between tbe quenching eftect and the prescnce of
conjugatedletraenesis seen in the fact that parinaric acid has a quenching eftect of
96.2 for a dilution of 0.O06Voand still one of 62% for a dilution of. O.AOO2%.

708

nESEARcHtN pHysropATHoLocy

methylcholanthrene
injection.Thirty-threeanimals died without tumors
during the courseof the experimental
period,The number of animalssurviving for five months plus the number in which tumors developedduring
this period of time are listed for each group.
In a secondgroup of experiments(Tnnle XXXIX) six groupsof 40
mice each receivedone subcutaneousinjection of 0.25 mgm. of methylcholanthreneof an 0.ZVosolutionin tricaprylin and a secondsimilar injection one week later. Groups were treated twice weekly for three months
with 0.3 cc. of 57o solutionsof the following fatty acids in cottonseedoil:
Fatty acids from cod liver oil, conjugatedfatty acids from cod liver oil,
eleostearicacid, linoleic acid and conjugatedlinoleic acid. A control goup
of animalsreceived0.3 cc. of cottonseedoil in the contralateralflank twice
weeklv for three months.
T,rsLr XXXIX
Died
Without Tumors

Treatment

Cottonseedoil--<ontrols
Fatty acids from cod liver oil
Conjugatedfatty acidsfrom cod
liver oil
Eleostearicacid
Linoleic acid
Conjugatedlinoleic acid

-/o

Tumors

With Tunrors

4
3

3t.36
33i37

86
89

6
l0
o
6

1 5 ,3t 4
24,30
29//40
25,,34

44
80
72
73

In the third group of experiments(TnalE XL), four groupsof 3O mice

each were employed.Mixtures of the methylcholanthrene
and of the fatty
acidsusedwere preparedby adding0.5 cc. of the 5Vo f.attyacids in cottonseedoil solutionsto 0.25 mg. of methylcholanthrene
of O.25% solution
in tricaprylin. The injectionswere made subcutaneously
immediately after
mixing. Each animal receivedthree injectionsat intervalsof one week and
T,tsLP XL
Treatment

% Residual Died WithFluorescenceout Tumors

Methylcholanthrene
f
cottonseedoil
Methylcholanthrene-lfatty acidsfrom
fish oil
r
Methylcholanthrene
conjugatedfatty acids
from fish liver oil
MethylcholanthreneL
eleostearicand
conjugatedlinoleic
acid

Tumors

% With
Tunrors

95

33i44

85

20./46

43

t9

6i46

l3

24,/39

6l

92

ll

NorE s

709

the observationson tumor incidencewere followed for 5 months. The

fatty acidsemployedwere fatty acidsfrom fish oil, conjugatedfatty acids
from fish oil, a mixtureof equalpartsof eleostearic
and conjugatedlinoleic
acids,and cottonseed
oil as a control.The quenchingeffectis shownas the
percentof residualfluorescence
of methylcholanthrene
when mixed with
the fatty acid mixtures.
These results indicate a certain relationshipbetweenthe quenching
action of the conjugatedfatty acids upon hydrocarboncarcinogensand the
ability of fatty acids to reduce the carcinogenicityof these hydrocarbons.
It is not sufficientto have a conjugatedfatty acid present,in order to have
the effectupon carcinogenesis.
Eleostearicacid did not significantlyreduce
the incidenceof tumors and conjugatedlinoleic acid was no more active
than its nonconjugatedisomer.
Conjugatedfish oil fatty acids (which contain di-, tri-, rctra-, pentaand hexanes) when mixed with methylcholanthrenereduced the tumor
incidenceto l3 Vo, whl/'ea mixture of eleostearicand conjugatedlinoleic
acid (di-, and triene conjugatedacids) which have a limited quenching
action gave an incidenceof 6l % . Although the incidenceof tumors was
much lower in thc group receivingconjugatedfish oil fatty acids,the nonconjugatedfatty acidsfrom the samesourcehas a limited influenceupon
the cancerinducingproperty of the hydrocarbon.Whcn fatty acids were
not mixed with the carcinogen,but were injected separately,the nonconjugated acids appearedwithout eftect.
Statisticalanalysisof the data from thesethree experimentsshow the
following: thc resultsare significantfor the group treatedwith conjugated
fatty acids from cod liver oil bcfore and after methylcholanthrene
was
as comparedwith controlgroup treatedwith cottonseed
administered
oil in
E x p e r i m e nIt ( x ' : 6 . 6 5 o n b a s i so f t u m o r / n o t u m o r ) . I n E x p e r i m e n It I ,
TrsLr XLI
Q u r - N c H r n r co r M e r H y r . c H o L ^ N T H R n N e0 . 0 6 2 V o l N E r u y l
sy SussreNcrs OrHrn TH,rN Ferry Acros

Substance
Glycerol
n-Butanol
Butyl mercaptan
Hexyl mercaptan
Dodecyl mercaptan
Hexadecyl nrercaptan
Na thiosulfate
Ethyl sulfate
N itrogen mustard
Allyl K xanthate
Nitromethane
Ethylene trithiocarbanrate
Cholesterol

Alconol_

% Dilution Used
5.0
4.5
1.0
2.0
2.0
2.0
.0-5cc. fronr
50% solution
1.0
0.1
1.0
1.0
1.0
1.0

Fluorescence
106.8
96.4

ro2.5
92.O
82.0
70.0
97.0
95.0
79.9
3.8
7.4
,7
93.0

lllElFFl-ffi*iEgiE-

710

xESEARcHIN PHYSIopATHoLocY

the resultsare very significantfor the goup treated with conjugatedfatty

acidsof cod liver oil as comparedwith the control group (xr: 13.09).
In Experiment III, the resultsare very significantfor all tfuee groups in
w h i c hf a t t y a c i d sw e r ea d d e dt o t h e m e t h y l c h o l a n t h r e n
( xe, : 1 3 . 3 , 4 1 . 5 6
and 8.32 respectively).
When comparisonis made on the basisof tumors/no tumors between
groups receivingnonconjugatedand conjugatedisomersof the same fatty
acid mixtures,the resultswere significantin all three experimenb(x. : 8
in ExperimentI, 22 inExperimentII, and l2 in ExperimentIII).
In the light of the relationshipbetweenquenchingactiviry and the
reductionof the carcinogenicactivity,we are investigatingdifferentother
agents.Table XLI showsthe valuesof this effect.
Chapter 12, Note 5. Lipids and Tumor Chlorides
We submitted groups of mice grafted with DBA mammary adenocarcinoma,to treatmentwith variouslipoacidsor positive lipoids preparations. After ten days of treatmentthe tumors were removed and analyzed
for their contentof chlorides,usingthe Volhard technic,in which the titration of silver nitrate was made electrometrically.As lipoacids,we used for
experimentcod liver oil fatty acids,lipoacidsfrom human placenta and
butyl-mercaptan;
for lipids with a positivecharacter,we usedcholesterol,
insaponifiable
fractionsof humanplacentaand butanol.In all casestreated
with lipoacids,the amount of chlorideswas higher than in untreatedcontrols. With cod liver oil fatty acids,valuesas high as 1357o above those
of controlswere found. With the lipoacidsof human placenta,the average
valuewas ll4o/o abovethat of controls;with mercaptans,
78Vo above.The
influence exerted by the opposite lipids was much less manifest. With
cholesteroland insaponifiablefractions,chloride values were ZOVobelow
thoseof controls;with butanol,33Vo below.
Chapter 12, Note 6. n-OH Fatty Acid and Experimental Tumors
We studiedthe influenceexertedby the seriesof alpha OH fatty acids,
saturatedand unsaturated,upon the evolution of different tumors in mice
and rats, to find that only one memberhas a manifesteffect which is limgraftsof 6C3HED lymphosarcomain
ited to a singletumor. Subcutaneous
C3H mice grew with abnormalrapidity.48 hours after the transplant,the
tumor could be felt. A very soft, highly edematousand, for this reason,
diffusetumor developedrapidly so that deathoccurredusuallyaround the
tenth day. This tumor was especiallyresistantto most of the chemotherapeutic agentstested.Daily administrationof a 5% solutionof alpha OH
caprylic acid in a dose of 0.2-0.5 cc., started evcn the fifth day after
the graft when the tumor was alreadywell-developed,
was followed by its
in a high proportionof cases(55/60).
rapid involutionand disappearance
In the few casesin which the tumor persisted,
its evolutionwas very much
changed.The animalsremainedalivefor more than a month. If, after tlree

7ll

Control

# t
Start treataent

rith
Treated

o(- 0H
caprylicaci

tjto. 290. The administraiion of alpha oH c:aprylic acid in micc bcnring 6c.HED
lympbosarcoma tumor, induccs the dirappcaranccof the tumor in a high proportion
of cases.

or four days,when the tumor was alreadyhighly reduced,the administration of the preparertionwas discontinucd.the tumor began to grow again,
but much more slowly than usual.
Chapter 12, Note 7. Hydropersulftdes
The existenceof different bonds betweenunsaturatedfatty acids and
oxygenled us to study difTcrentbondssinrilarlyoccurringbctwecnthe same
fatty acidsand sulfur, the secondmenrberof the oxygen series.The treatment of polyunsaturatcdfatty acids or their triglycerideswith sulfur has
shown that two different formationscan bc obtained,By hcating the mixtures above ll0' but below 125'C. precipitatedsulfur is incorporated
without a manifestchangcin color or other properties.The iodine numbcr
is not changed.When conjugatcdfatty acids or thcir triglyceridesare
treated, no changesare seen in thc spectral analysis.By heating abovc
130uC, the color of thc prcparationchangcdprogrcssively
reachingdeep
prolonged.
rcd-brown if the trcatment is suflicicntly
Concomitantly, the
iodine number decreascsprogressivelyand cventually reacheszero. The
spectralanalysisof the conjugatedfatty acids shows the peaks going progressivelydown until no more conjugatedformationsare present,indicating
that thesechangesaffect the double bond.
The analogy between the fixation effectsof oxygcn and sulfur has sug-

712

REsEARcH rN pHysropATHoLocy

gested that the first bond correspondsto a hydropersulfide similar to a

hydroperoxide.The secondbond would representa fixation of the sulfur
at the level of the doublebond itself, similar to a peroxide.Studiesof similar bondsof sulfur were made in tetralinwhere hydropersulfides
were obtained.The study of the proprtiesof all thesepreparationsseemsto confirm the hypothesisthat the compoundsobtainedare hydropersulfides.

F I c . 2 9 l . Q u e n c h i n go f m e t h y l c h o l a n t h r e n e
f l u o r e s c e n c ei n s a m p l e so b t a i n e d w h e n
0 . 5 % s u l f u r i n c o t t o n s e e do i l w a s h e a t e d f r o m 1 2 0 " C t o 2 8 0 ' C . T h e s a m p l e sw e r e
d i s s o l v e di n e t h e r - a l c o b o lm i x t u r e a n d m i x e d i n e q u a l p a r t w i t h . O l 2 S % m e t h y l cholanthrene.The existenceof concomitant changes in the oil with sulfur and in
the oil alone, indicates that the variations correspond primarily to cbangcs which
t a k e p l a c ei n t h e o i l i t s e l f w h e n h e a t e d .

We studiedthe changesin thc qucnchingeffectupon the fluorescence

which occur when sulfur added to a mixture of
methylcholanthrene
of
is heated.This was comparedwith the effectof heatingupon
triglycerides
cottonseedoil alone.Fic. 291 showsthe resultsof this analvsis.
Chapter 12, Note 9. I\{agnesiumand Adrenalectomy
The biological antagonismbetween homotropic magnesiumand heterotropicsodium,both acting at the metazoiclevel, has led us to the study
of the specificinfluenceexertedby magnesiumupon the recoveryprocesses
in adrenalectomized
rats. It is known that, while an adrenalectomyis not
alwaysfatal in old rats--{eath occursuniformly in younger animalsweigh-

NorEs

713

ing less than 150 grams.The administrationof l7o sodium chloride as

drinkingwatcr is known to protectthe adrenalectomized
animaland, if administercdfor a sufficientlengthof time, to preventdeath.The administration of mapmcsium
sulfatcby repcatcdinjcctionsof .5 cc. of a lOVo solution
grams
per 100
of body weight or even orally as .5-l Vo in drnking water
hasan antagonistic
effectto that of sodiumchloride.A75% mortalityrate
in olderanimalsreceivingmagnesium
sulfateas comparedto a 20Vo ratein
thc untreatcdwas seen.Similarly,in young animalsreceivingmagnesium
sulfatein additionto salty drinking water, the mortality rate in some experimentswas over 80Vo.
C h a p t e r1 3 , N o t e l . G l y c e r o l a n d C h i l l s
In one of a group of severelyburned subjects,who were at that time
underour care and had beenexperiencing
severalchills a day, an injection
just at the momentwhen
of glycerolsolutionhad beengivenby coincidence
a chill was starting.WhiJc such a chill alwayspreviouslyhad lastedfor
more than ten minutesin this patient,it stoppedalmostimmediatelyafter
the glycerolinjection.
An experiment was set up to confirm or negate this correlation. As
soon as any patientof this group felt the sensationor premonitionof a chill,
he was given either an intramuscularinjectionof 3-5 cc. of a ZOVosolution of glycerolin salineor 3 cc. of salinealone as placebo.In almost
every case,the chill was cut short by the glycerolwhile the placebohad
no effect. Less striking but stiU interestingeffectswere obtained when 20
to 30 drops of glycerol werc given orally in 50 cc. of water againstan oral
solutionof.lVo sugarin wateras a placebo.
Not one of thc other substanccs
used at this time, such as adrenalin,
quinine,pilocarpineor pantopon,orally or parenterallyinfluenceda chill
once it had begun.Later, butanolalso was found to have an effectsimilar
to that of glycerolalthoughlessmanifest.
Sincethe first experimentwith glyccrol,we have tried it in many patientssubjectto repeatedchills and have frequentlyobtainedthe sameresults.we havetried to explainglycerol'seffectuponchillsby considering
the
role of chills in the defensemechanism.
would
Chill
mark the beginningof
the secondphaseof the diphasicdefensephcnomenon.(See Chapter 5.)
I t b r i n g s v a r i o u s c o n s t i t u e n t sc,s p c c i a l l yt h o s e w h i c h h a v e t o r e p l a c e
constituentsalteredin the first hydrolyticphase,Among them are agents
especiallyable to influencethe free fatty acidsliberatedin the first phase.
Apparently,the fact that it takessomc time for the anti-fattyacid agents
to pass into the circulation,most of them coming from the RES cells,
makcsthe chill last so long.The immediatepresence
in the blood of a sufficient amount of glycerol, which is a relativelyefficient anti-fatty acid
agent,climinatesthe need for liberationof body anti-fatty acid agents.
Thus, with glycerol,the chill would no longer be requiredto induce liberationof suchagentsand would stop.

714 I

R E s r , A R c HI N p H y s r o p A T H o L o c y

Chapter13, Note 2. Influenceof Glycerol Upon the Cardisc Rhythm

Figure292 showsthe electrocardiogram
of a rabbitreceivinga solution
of 20% glycerolintravenously.
Frequentextrasystolcs
appeared.[t rvas
interesting
to notethat,at thc sametime,the animalbecamcsomnolent.

Glycerol

Eefone

After

Ftc. 292. Electrocardiogram of a rahbit receiving intravcinous injections of a rolulion of glycerol ?0f1. characterizedby the appearanceof extrasystoles.(a) bcfore
treatment. (b) after 30 cc.

Butarrcl

F 1 o . 2 9 3 . E . r t r a s y s t o l eas p p c a r a f t e r t h c i n t r a p c r i t o n c a li n j e c t i o n o f b u t $ n o l i n v e r y
h i g h d o s e s .( a ) b e f o r e t r e a t m e n t .( b ) a f t e r 1 . 6 g r a n r / 1 0 0 0g r o f a n i m s l .

Chaptcr 13, Note 3. Glycerol Induccd Convtrlsions

Repeatedinjcctionsof glycerolin rats wcre seento inducc convulsions.
Using rats weighing200-250 grams, 5 to l0 cc. of the ?Afo solution of
glycerol in saline were injected intraperitoneally.The injections were repeatedoncc or twice a day. After sevcraldays of treatment,usually from
3 to 5 days, onc of the injectionswas followed within a few minutes by a
severeconvulsion,lethal for most of the animals.In the survivinganimals.
the ncxt injectionwas alwaysfollorvedby a lethalscizure.
Chapter 13, Note 4, Suspensionsof Lipoids
variouslipoids were dissolved
In ordcr to obtain colloidal suspensions,
in alcohol and a certain amount of the alcohol solution mixed with water.
saline or isotonic solutions,From the resulting milky suspensions,the al-

NorEs

715

cohol was eliminatedby boiling under reducedpressure.To insure almost

completeeliminationof the solvent,an excessof water was addedand the
excesswas then eliminatedthroughboiling.The useof acetoneor ether as
solventgave much lessfavorableresults.
R e l a t i v e l ys t a b l e s u s p e n s i o nw
s e r e o b t a i n e db y m i x i n g s o m e l i p i d
p r e p a r a t i o nssu c ha s m i x t u r e so f u n s a t u r a t efda t t y a c i d sw i t h a 0 . 5 7 o s o l u tion of cellulosegum. Such stablesuspensions
could not be obtainedwith
preparationsof positivelipids.
Chapter 13, Note 5. CholesterolInduced Convulsions
When relatively larger doses,such as 5 cc. of 2Vo cholesteroldaily,
were administered
repeatedlyto rats of around250 gr. of weight,convulsionsappearedafter 4 to 8 days.They were inducedearlier in femalesthan
males.The first convulsionalwayswas lethal.Convulsionsalso occurred
in humansaftcr repeatedinjcctionsof cholesterol
in dosesas high as 20 cc.
of the ZVo solutionin oil. Even small doses,such as 2 or 3 cc. of the same
solution,inducedconvulsionsin paticntswith brain metastases
or h those
who had had previousconvulsions.
Chapter 13, Note 6. TreatmentsIn SuccessiveGenerations
The relativelyshort survivaltime of animalsbearingtransplantedtumors has been a handicapfor the study of the influenceexertedby many
agents.Effectsrequiringsome time before they can be inducedare thus
missed.Changeswhich occur in tumors-such as the tendencyto ulcerate
after treatmentwith fatty acids-have been found to be transmittedin
generations
successive
of the tumors.This has led us to carry on treatment
beyondthe survivaltime of one individualhost in order to study the influcnceof variousagents.In one groupof experiments,
this was done through
treatmentof the successive
hosts of serial transplants.In anothergroup
of experiments,
the treatmentwas appliedto the transplantsthemselves
in
hosts.
successive
Mice with graftedtumors were treatedwith the chosenagents.When
the tumor in a treatcdhost. or in a control,had grown to lt/z centimeter
diameter,it was removed.Part of it was usedfor further transplants,part
for microscopicstudies.The rest of the animalswere kept until death and
the survivaltime was noted.Transplantsof the tumor from treatedanimals
as weU as from controls were graftedin new animalsand the treatment
continuedfor the new hosts.This procedurewas repeatedfor successive
generations.
In other experiments,
the successive
transplantswere dipped,
prior to grafting,in an oily solutionor in a suspension
in salineof the agent
being tested.Thc procedurewas repeatedcontinuouslyfor both treated
animalsand controls,and growth and survivaltime were noted. The followingexperiments
are characteristic.
Usingthe insaponifiable
fractionsof humanplacentain an oily solution

n E s E A R c Hr N p H y s r o p A T H o L o c y

716 /

of. 5vo, or in a saline suspensioncorrespondingto two milligrams of the

material per cubic centimeter,the following resultswere observedin the
caseof Ehrlich mammary carcinomain mice. No changesin survival time,
evolution of the tumor, gross or microscopiccharacter were seen in the
fint and in some experimentseven in the secondgeneration.Usually with
the third generation,the survival time was reduced, the tumor growing
much more rapidly and killing the animal in around 20 days. The malignant characterof the tumor was seento increasein the subsequenttransplants and in the fifth generationin someexperiments,killed the animal in
0.ll

C
o

i n e t h v l a l c o h o l / e t h yal l c o h o l

Lyrphnodes
Adrenalgland
Sonetumrs
t y p e0

Io
L

La

f a v e L e n o t h( m l ' )

Ftc. 294. Spectral analysis of the insaponifiablefraction of various origins. It shows

a c h a r a c t e r i s t i cp e a k a t 4 5 0 m p , a n o t h e r a t 3 6 0 a n d a n o l h e r a t 2 7 2 . T h e o r g a n s i n
which tbey are especiallypresent are indicated.

less than a week. The morphologicalchangeobservedin these successive

transplantswere also characteristic.The tumor was seen to change from
characterwas thus altered
solid to encephaloid.The adenocarcinomatous
was
by passingthrough the
increased
and the degreeof undifterentiation
generation
in some experithird, fourth and sixth generation.At the sixth
ments-and the fifth or eighth in others-microscopic examinationshowed
that sarcomatoidportions were presentin the tumor. The malignancyappearedto be at its maximum in thesetumors. Transplantsof tumors with
sarcomatoidmicroscopiccharacter,if treated in the same manner, gave
negativegrafts.Thus, it appearsthat the treatmentwith the insaponifiable

NorEs

717

fractions has progressivelyincreasedthe malignancy until the moment

when sarcomatouscharacterappearedafter which negativetransplants
were observed.
The treatmentof a tumor with lipoacidpreparations
of human placenta
has producedoppositechangesmanifesteven in the first transplants.These
0 . 2 Zs o l u t i o n

t J . 4 Xs o l , r rt o n

c
o

E
6

c
4'
t-

}{

l'

J@

fraveLenoth(m r.)
Frc. 295. Details of the spectral analysis of rat colostrum indicates the existenceof
a formation with three peaks in the region 290-250 mp with some similarity to tbe
c o n j u g a t e dt r i e D e s .

718

xESEARcH tN pHysropATHoLocy

increasedin the secondgeneration.After the secondand very rarely after

the third grafts, negativetransplantswere obtained.We used this method
of treating tumors through successivegenerationsroutinely. The results
obtained for difierent agentsare discussedin the text of this publication.
Chapter 13, Note 7. Conjugated Trienic Alcohols
Spectralanalysishas permittedus to recognizethe presence,in certain
mixturesof the insaponifiable
fraction.of severalpeaks,someespeciallyinteresting.
peakswereseenat 450, 360 and 272 my.,as shorvn
Characteristic
in Figure294. ln the first analysis,
one was identifiedas correspondinc
to a
peakof 2720 Angstroms.
In morecompletefurtherspectralanalyses,
it could
be seento correspondto a conjugatcdtriene with its characteristicthree
peaks.The fact that it correspondsto a substancewith positive polar group
explainswhy, comparedwith conjugatedacids,the curve showsa marked
displacement
of the peakstoward higherwave lengths.(Fig. 295l This can
bc related to the different influenceexerted by the electrically opposite
polar groups.This compoundwas first found in the colostrumobtained
from thc stomachof newborn rats on the first day. In smaller amounts,it
has been seenin other samplesof milk or butter, and in pork kidneys. It
has been found less frequently in growing tumors and is even rarer in
growing animals.
The samespectralanalysishas permittedus to recognizeother peaks
and relate them to the differentsourcesfrom which the unsaponifiable
fraction was obtained.Fig. 294 shows these peaks and indicatestheir
correlationwith the orisin of the material.
Chapter 13, Note 8. Toxicity of Butanol in Humans
A group of advancedschizophrenic
patients(221) were given 500 cc.
of a 6Vo solutionof n-butanolin salineintravenously,
the entire amount
being injcctedin 30 minutes.The only manifestation
which could be consideredto parallel the toxic eflect in animals was a very short period of
which, in only one or two cases,could be consideredas sleep.
somnolence
Usually,even with dosesof 500 cc. of a 60/osolutionadministeredintravenouslyin less than 25 minutes,it was not possibleto obtain even this
transitorysomnolence.No toxic effectwas noted when the same dose was
again administered24 to 48 hours latcr, and repeatedseveraltimes. Except for an inflammationof the vein which appearsonly if hundredsof cc.
of a solutionabove6Vo is injected,no othernoticeableeffectsare observed.
The intravenousadministration,
in postoperative
cases,even of l5 gm.
of butanoldiluted in about 2-3 liters of salineper day, repeatedfor four
days,has beenentirelyfree of any toxic effect.
and even five consecutive
Intramuscularadministrationwas observedto be well toleratedeven
of butanol.we obtainedconcentratcdaqueous
for higher concentrations
solutionsby dissolvingbutanol in a 357o solutionof sodium benzoatein
containingmore than 30o/obutanol seemedto induce
water.Preparations

NorEs

'l19

necrosiswhen administeredintramuscularly
in animalsand to inducepain
at the site of injectionh humans.A 307o solutionof butanol,however,
was well tolerated.Daily administrationof subnarcoticdosesfor long periods to micecausedno toxic effects.On the other hand, repeated injections
with narcotizingdoseswere toxic and even led to death of the animals
after severaldays.
Chapter 13, Note 9. Butanol and Leucocytes
T h e a d m i n i s t r a t i oonf b u t a n o li n s o l u t i o n so f 6 . 5 % i n s a l i n ei n t r a p e r itoneallyin rats was seento inducea hyperleucocytosis.
5 cc. injectedat
once was seento double the prcviousamountof leucocytes.
This increase

6l
o
>
o
I
J
o

J2

6() :to

Jo

oo

so

lLo

,r"

tuo )ra

rar

rj,

160 te

tzo ,60

i,rr?rtff
F t c . 2 9 6 . T h e a d m i n i s t r a t i o no f 5 c c . o f a s o l u t i o no f 6 . 5 % n - B u t a n o l i n t r a - p e r i t o n e c l l y t o r a t s i n d u c e sa n i n c r e a s ei n t h e l e u c o c y t en u m b e r .

s t a r t e dt w o h o u r s a f t e r t h e b e g i n n i n go f t h e i n j e c t i o n sa n d c o n t i n u e d
prog5cssively,
to reach the valuc of 34,000, seven hours after the bcginning of the injections.The hyperleucocytosis
was seen to persistfor
hours.
was
more than 24
The numberof leucocytes increasedin the animal
s h o w ni n F i g . 2 9 6 .
A s t i l l m o r e m a n i f e set f f e c tw a so b t a i n e dw i t h i n j e c t i o n o
sf I cc.of the
e v e r yh o u r d u r i n gt h e d a y . I t i s i n t e r e s t i ntgo n o t e
s a m es o l u t i o nr, e p e a t e d
that this effect was manifestedalmost 6 hours after thc injection with
b u t a n o lF
. i g . 2 9 7 s h o w sa n e x a m p l eo f t h e s ee x p e r i m e n ti sn w h i c ht h e n u m arrivedat 42,500.
ber of leucocytes

'720

nesEARcH rN pHystopATHoLocy

4{w

o
>
(,

CI
.J
J

3 4,no

60

Jo

Jo

90

/2o

/t0

lto

2to

2(o

27o

J@

JJO

J6O

J9O

.aO

Hinutes
Ftc. 297. The administration of I cc. of n-Butanol, repeated every hour, induces a
sizable increaseof leucocvtesin rats.

Chapter 13, Note 10, Butanol-SodiumLactate in Burns

In collaborationwith R. Ravich and P. Teitelbaumwe studied the
effectof variousagentsupon the survivaltime of mice to which a severe
caloricburn was inflicted.Under etheranesthesia,
adult white femalemice
were scaldeduntil the xyphoid,in water maintainedat 90'C. When the
durationof this treatmentwas 4 or morc seconds.the animalsdied in a
few minutesin superacute
shock.With 3 secondsthe animalssurvivedthe
immediateeffectof the burning,
They startedto die severalhourslater,in 6 hours4O% of theseanimals
h a d d i e d , a n d a t t h e l 8 t h h o u r , 9 O V ow e r e d e a d .
The influenceexerted by various agentswas studied by injecting the
respective
solutions2-3 timcsa day accordingto the experiment.The effect
was judged accordingto thc survivaltime. As the animalsdid not eat or
drink and especiallydue to the local burns did not urinate or defecate,we
consideredthe effectsobtained during the first l8 hours, after which the
animals were sacrifiedand used for the study of the chemical changes.
Fig. 138 shows the resultsof such an cxpcrimentwith sodium chloride,
isotonicsolution-sodium lactate6 M solution,butanol 6.5Vo in salineand butanol 6.5% in the sodium lactate solution. While sodium lactate

;ir:it:!

NorEs

721

alonc seemedcven to incrcasethe mortality.and butanol in salinealone

influencelittle this mortality,the effectof the butanol-sodium lactatesolution was manifest.The mortalitywas reducedfrom beginningto end of the
experiment.At the 18th hour it was of 30% insteadof 90% for the controls.And of.95% for the animalstreatedwith sodiumlactatealone.
Chapter 13, Note I l. Effect of Heptanol
Adult rats and mice were injected subcutaneouslyin the back with
20 cc. and 3 cc. respectivelyof nitrogenwhich had been sterilizedby being
passedthrough steri.lecotton plugs.Into the pouchesso formed, a suspnsion of living coli bacilli-2 cc. for rats and Yq cc. for misg-\'/35 injected.
This suspensionwas obtained from a 24-hour culture on agar and was
diluted to provide l0 millon microbesper cc. One group of animals was
treated by intraperitonealinjection of I cc. (for rats) and Vq cc. (for
mice) of sterile sesameoil. The other group receivedinjectionsof similar
dosesof.2% heptanol in oil. In someexperimcnts,only one injection with
heptanolwas given, while in othersthis was repeateddaily or every second
day. In the controls no specialreactionwas seen.In the heptanol-injected
animalsan exudateappearedin the infectedpouch and led to rapid necrosis of the skin. A characteristicof the exudatewas the prescnceof a small
number of leucocytes.
Chapter 14, Note 1. Observationsof Dr. E. Stoopen
From a seriesof observationspublishedby E. Stoopen (184), we
chosethe following:
"Right trigeminal neuralgia for the past l0 years, with short, sharp
pains. Neither food intake, nor time of day have ever influencedthe pain.
The patient was submitted to various treatmentssuch as ultraviolet rays,
quinine, neosalvarsan,cobra poison and vitamin B. First alcohol nerve
block calmed the pain for 15 months. Secondalcohol injection brought
no relief. Third block calmed the pain for one year. Fourth alcohol injection calmcd pain for three months. The last injection caused, however,
trophic ulcerationsof the throat and cornealulcerationswith ultimate loss
of the sight.
In June 1942, a treatmentwith glycerinand the insaponifiablelipid
fraction was begun. The pain which proved to be of an alkaline type,
ceasedin 3 days.
In July 1943,the patienthad a lumbagoattack,a conditionfrom which
she had often suffcredand which had beenboth long-lastingand resistant
to classicalmedication. Treatment with the insaponifiablelipid fraction
made the pain subsidewithin a few days.
In an attempt to modify the ulcerationsin the throat, though the pain
had not reappeared,the patient was continuouslytreatedwith the insaponifiable fraction and cholesterol,and with large dosesof vitamin A. However, there was no effect on the ulcerations.

'122

nEsEARcH rN pHysropATHoLocy

In February 1944, the pain reappeared.Study of the pain curve revealedit to be of the acid type sincethe paroxysmalpain correspondsto a
very low pH. The patient was given ammonium acetate and lipoesters.
Four days later the pain had considerablydecreased.and fifteendays later,
completelysubsided."
"Mrs. W. For elevenyearsthis patienthas sufferedof a left trigeminal
neuralgia.Each year the crisislastedfour to five weeks,during which time
the pain always appearedbetweenfour and seven in the morning, lasted
for one to two hours and then disappeared.
The pain was so severe,being
almost unbearable.For the rest of the day, the patient only felt a slight
sensitivity.Barbituratestaken even in large amountshad no influenceon
the pain. Removal of the Gassergangliawas suggestedas the sole possible
cure by numerousdoctorsconsultedin Mexico and the United States.
The patient came under our care on November 12, 1943. Her most
recentcrisishad startedon October3l . Studyof hcr pain showedit to be
of an alkalinecharacter,sincepain is quite intenscwhen the urinary pH is
glycerine.The patientprotested,feelingthat a few
high. we recommended
dropsof glycerinewould not be ableto help her, pointingout that intensive
treatment,had given no results.However,on November l5th, when pain
startedat 5:00 A.M.,the patienttook the glycerinedropsand to her amazement, the pain disappearedwithin two minutes.She assertedthat no medicine had ever beenable to stop the pain once it had started.At 7:30,r.rrr.
the pain returned but again decreasedafter the patient had taken several
glycerinedrops.
O n N o v e m b c rl 6 t h , t h e p a t i e n te x p e r i e n c epda i n a t l : 3 0 , 5 : 3 0 a n d
7:30 n.t"r.During the first two periodsof pain, the pain was instantaneously calmedwith glycerineand phosphoricacid; the third period of pain
was decreased
in intensitybut lastedfor 40 minutes.
on NovemberlTth: pain appearedat 9:00 l.r',r.but disappeared
three
minutesafter medicationof glycerineand phosphoricacid.
O n N o v e m b e rl 8 t h : p a i n w h i c h s t a r t e da t 8 : 0 0 L v . c o u l d n o t b e
calmedwith medication.Coramine,Cholesterol
and insaponifiable
fractions
rverethen prescribcd.
o n N o v e m b e rl 9 t h : p a i n w a s e x p e r i e n c eadt 4 : 0 0 a n d 8 : 0 0 p . r r r .b u t
subsidedwithin three minutes after medicationof glycerine,phosphoric
acid and coraminewas taken.
on Novcmber20th: from this day throughNovember25th, the medication was unableto influenccthe pain, and consequently
the patient becamedisheartened.
A study of her pain patternat this point indicatedthar
it had changedto the oppositetype-from alkaline to a defrniteacid pattern.
on November 26th: bicarbonatewas given at the onset of pain resulting in a considerabledecrcasein its severity.No further pain was felt on
the following days and the crisis was consideredended.In this instancc.
the crisishad lastedfor 27 days (the lengthusuallyvaried from 27 to 35

ror"l,
days). However, the treatmentachievedwhat had been;;::rr.',
previouslytried treatments-the cessationof pain once it had started.
These observationsled to the following pertinentconclusions:1 ) they
showedthe existenceof typical acid or alkaline pain;2) the possibilityof
changingpain from one type to the oppositeone, either during the course
of the diseaseor due to medication;and 3) the possibilityof eliminating
pain with appropriatetreatment."
Chapter 14, Note 2. Dr. Welt's Publicationon Butanol-Conclusions
"n-Butanolwas administered
to a largenumberof patientswith pain duc
to the trauma of variouscommon otorhinolaryngological
and ophthalmologic surgicalprocedures.Pain was relievedin approximately90 per cent
of the patientsso treated.
Theseclinical resultswere consideredin the light of studiesby Revici
and his co-workersregardingthe physiopathologyof wounds. The results
indicatethat the proposedconceptof pain has significantpracticalclinical
applications."
Chapter 14, Note 3. Dr. A. Ravich'sConclusions(189)
In his articleconcerningthe post-opcrative
A.
care in prostatectomies,
Ravich arrivesat the followingconclusions
298.
exemplifiedin Figure
SUMMARY

" A n e w c o n c c p to f t h e l o c a l p h y s i c o c h c m i ccahl a n g e so c c u r r i n gw i t h i n
pathologicalfoci as introducedby Revici, has beenbriefly described.Accordingto this view, pain is the resultof local pH changesbrought about
by the accumulation
within disturbedtissues.
of acid or alkalinesubstances
Changesin the lipid balanceare associated
with and may accountfor these
alterations.
"Thc possibilityof correctingor neutralizing
suchlipid changeshasbccn
exploredclinically in severalseriesof urologicalcases.Thc favorableeffects
upon pain as well as upon bleeding,wound healingand other important
postoperativeproblems and complicationsindicate the need for further
study along theselines."
Chapter 14, Note 4. Treatment of Post-traumaticConditions
Of spccialtheoreticaland practicalintercsthas becn the treatmentof
traumaticconditions,especiallythose following surgicalprocedures.The
recognitionof the role of fatty acids acting at differentlevelsof the organizationand inducing severaldifferent manifestations,
has led to the
concomitantuse of variousagentsproper to the levels.From the various
agentsstudied,heptanolwas thus chosenas acting at the cellular level,
glycerol,polyunsaturated
alcoholsand alkaline amino acids and butanol

724 /

RESEARCH IN

I)ayof

PHYSIOPATHOLOGY

r. lst. Post-op.2nd.Post-op.3rd. Post-op.

severe
!
I-'l moderate

C o n tro l n _ C ontnol n_ Contr ol n_ . Contr ol n_

E u ta n o l
Butanol
Butanol
ilutdnol
Ftc, 298. Tbe administration of n-Butanol after prostatectomy markedly reduccs
pain. (J. Urol. 62: 629, 1949.)

at the tissular level, glycerophosphoricand organic acids at the systemic

level.
The various preparations,obtained by combining or mixing these
agents,were administeredby intravenousinfusions together with glucose
and saline or glucoseand sodium lactate,in the more severecases,and
intramuscularlyor orally in the milder cases.The results obtained with
thesepreparationsin hundredsof subjectshave been highly satisfactory.
Chapter 14, Note 5. Dr. B. Welt's Conclusionson Hearing
In his studieson hearing,Welt arrives at the following conclusions,
communicatedat the Brooklyn Eye and Ear Society.
"l) The ideas,methodsand substances
devisedby Revici have been
applied to the problem of impaired hearing, and have shown significant
resultsin improving that function.
2) The study has confirmed the dualistic concept about pathological
foci, namely that a pathological focus may exist in two states of metabolic
imbalance,leading either to a local alkaline or acid change.
3) The substances
utilizsd in this study have beeneffectivein influenc-

NorEs

725

ing the symptom of impaired hearing,and it is reasonableto believe that

the pathologicalstructurehas alsobeeninfluenccdto somedegree.
4) Revici's ideas of the fatty acid-stcrolimbalancehave been confirmedby showingthat other instances
havingsimilarbiologicactivities,act
in the samemanner.Their clinicalapplicationconfirmsthis.
5) This studyshowsthat both air conductionand boneconductionmay
be benefited.
No methodhas beenfound to improveboneconductionup to
the prcsenttime.
6) From the biochemicaland chemotherapeutic
point of view, this
study indicatesthat the vestibularlabyrinth and the cochlea should be
viewedas one organ. Phylogcnetic,
histological,and this clinical study all
tend to support this idea.
7) The study indicatesthat peopleup to 60 yearsof age may obtain
normal audiometrichearing if trcated early enough. Children and the
youngerage groupswere thosethat had the highestincidenceof good results.
8) Finally, this study has indicateda dual therapeuticattack on a
hithertoinsolubleproblem.It can help us in this otologicproblem at any
age in life, the formativeyears,when hearingis vital to the educationof
the children.
9 ) I t s h o u l dn o t b e c o n s t r u e do r i n f e r r e d ,f r o m t h i s c o m m u n i c a t i o n ,
that a cure for hearingis implicd.Thc only conclusionto be drawn is that
thc author has beneficiallyinfluencedthe impaired hcaring function, or
induceda remissionfor varyingperiodsof time at an improvedor normal
f u n c t i o n al c v e l . "
Chaptcr 14, Note 6. Butanol in PlasticSrrrgery
F o l l o w i n go u r i n d i c a t i o n sS. . S h e rh a s u t i l i z c db u t a n o li n p o s t - o p e r a t i v e c a s e s .O n e o f t h c m o s t d i s a g r c c l b l cc o m p l i c a t i o n s e e n i n p l a s t i c
s u r g e r yo f t h e n o s ci s s e v c n t hd a y b l e c d i n uw
, h i c hw h i l eu s u a l l yn o t s e v e r e ,
hasbecnknown to cndangcrthc livcsof sevcralpatients.Usc of antibiotics
has reducedremarkablyboth incidenceand severityof the hemorrhage.
However,the preventionof seventhday bleedinghas remaineda problem
f o r t h e p l a s t i cs u r g c o ns. . S h e rh a s a p p l i c do u r t r e a t m e nw
t i t h b u t a n o li n
almost2,000 cases.Immediatelyfollowingsurgcry.10 cc. of.a 6.5Vo solution of butanol is injectedintramuscularly,thc injection being repeated
everysix hours for the first day. After 24 to 48 hours,the butanol is administeredorally in dosesof one tablespoonful
everyfour to six hours,and
this is continueduntil after the eighth day following the operation.With
this treatment,no scverebleedinghas beenseen.[n severalcaseswhen the
patientfailed to follow instructionsand did not continuetaking butanol,
hemorrhageresultcd.In two cases,blceding was relatively severe,the
hemorrhagewas brought under control by the intravenousinjectionof l0
to 20 cc. of the butanolsolution.Administrationof butanol afterwardpreventedsubsequent
bleeding.The valueof butanolas a preventiveof seventh
d a y h e m o r r h a g teh u . sh a s b c e n c o n f i r m e d (. l l i g )

726

xESEARcH rN pHysropATHoLocy

Chapter 15, Note l. Radio and ChemotherapeuticalEssays

Though carriedout only at the beginningof thesestudies,the application
of this method,conductedby LeonardB. Goldman,M.D. in 1950 and 1951
appearsinteresting.As part of this investigation,lipids were used in associationwith radiotherapy.In the sameresearch,a group of patientswere
also treated with lipids alone. A report on this serieswas presentedby
L. Goldman as part of a symposiumon the therapy of advancedcancer
patients, before the Radiological Section of the American Medical Associationat its annualconventionin AtlanticCity on June 14, 1951 (327).
His results with patients treated with X-ray and lipids and with lipid
therapy alone are summarizedin Table XVIA alrd XVIB.
Trsln XVIA
Resulrs op Ltpro Turn,rpyCorunlxeowrrn InnrDrATroN
(In excess
of thoseexpected
with irradiationalone)
Total
Type of
NumMalignancy
ber
Breast
8
Lymphoblastoma 7
Lung
6
Head and Neck
il
Gastro-intestinal 6
Gynecological
3
Genito-urinary
Sarconra
2
Miscellaneous
4
Total
5
0

Reliefof Symptoms
NumMod.a r K ber
Slight erate
ed
z
l
44
2
l
I
5
2
0
J
4
2
0
2
4
0
l
3
J

0
I

2
3
0
( 6 0 %)

l
0
0
l
1

l
0
0
0
0

I
0
I
I

Temporary

l Aresl'

I
I

s -

Regression
2
0

i ') l
I

i
i
6

0
0
I

0
0
0
I

1
2
6
(24%) (t2%)

T,rsr-pXVIB
Resurrsor LtproTHenrpyAloxr,
TemTotal
Type of
NumMalignancy
ber
Breast
t4
Lymphoblastoma 8
Lung
3
Head and Neck
3
Gastro-intestinal 6
Gynecological
)
t
Sarcoma
I
Miscellaneous
2
Total

porary
Reliefof Symptoms
NumMod- MarkArber
Slight erate
rest
ed
-2
1
0
6
5
0
0
5
3
3
0
0
3
0
2
l
0
l
0
3
l
l
l
0
I
0
0
r
I
0
0
0
0
0
2
0
0
2
I
23
l9
7
(58c/c)

Regression

2
2
0
0

I
0
I
6
(t8vo)(ts%)

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318. WERTLAKE, P. T., WILCOX, A. A., HALEY, M. I., and PETERSON,
J, E.: Variation on serum lipids during mental and emotional stress.
Am. Heart Assoc.Mtng. Oct., 1958.
319. AYLWARD, F.: Excretionof cholesterol.Lancet. October. 1958.
320. KINSELL, L. W., MICHAEL, G. D., FRISKEY, R. W., and SPLITTER,
S.: Essentialfatty acids,lipid metabolismand atherosclerosis.
Lancet.
Feb., 1958.
321. MOLMROS, H., and WICAND, G.: Atherosclerosisand deficiencyof
essentialfatty acids.Lancet. Nov. 7, 1959.
322. AHRENS, E. H,, INSULL, W., HIRSCH, J., STOFFEL, W.. PETERSON, M., FARQUHAR, J., MILLER, T., and THOMASSON. H.:
The effecton human serum lipids of a dietary fat, highly unsaturated
but poor in essentialfatty acids.Lancet.Jan. 17, 1959.
323. NOTHMAN, M., and PROGER, S.: On the effectof arachidonicacid on
serum lipids in ntan. Fed. Proc. 19:Part I, 1960.
324. RAVICH, A., and RAVICH, A, R.: Prophylaxis of cancer of the prostate, penis,and cervix by circumcision.N. Y. StateJ. ol Med.5l:
1519.1951.
RAVICH, A.: l. Urol. 48:298, 1942.
325. B A R O N , 8 . , a n d A N G R I S T , A . : A r c h . P a t h . 3 2 ' 7 8 7 , 1 9 4 1 .
326. REVICI, E.: The role of biological imbalancesin physiopathologyand
therapy. Rev. Med. Pasteur,l5:.133, 1942.
3 2 7 . GOLDMAN, L. B.: Use of lipids to enhancethe effect of roentgen therapy in the treatmentof pain from advancedcancer. Presentedbelore
Amer. Medical Association,Sectionol Radiology,Atlantic Ciry, June
14,1951.
3ll.

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748 /

rxoex

Alcohols and blood clot retractibiliry,3E4

A l i p h a t i c c h a i n f l e x i b i l i t y ,1 3 7 , 6 3 3
A l k a l i n e a m i n o a c i d s ,5 4 6
and eosinophiles,569
i n o r g a n i z a t i o n ,2 l
- hydrolyticfraction, 192
A l k a l i n e p a t t e r na n d t r a u m a ,4 2 3
- - w o u n d s ,5 0 , 4 2 3 , 5 9 7
A l k a l i z i n g a g e n t s ,5 3
A l k a l o i d s ,6 0 4
A l k y l a t i n g a g e n t s ,3 4 5
Allergy, 180, 193, 195, 2lO, 2lS, 267,
673
A l l e r g i c a n t i b o d i e si n d e f e n s e ,t 9 l
--and
cancer,2l5
- c o n d i t i o n s ,4 5 1
- d e f e n s e .1 8 0 .2 t 5
- i n c u b a t i o n ,1 8 0
- p r e c i p i t a t e s 1, 9 3
- reaction as test in cancer, 267
- 7th day manifestations,210, 7Zs
- s k i n ,6 7 3
A l l o p r e g n a n e ,1 4 4
- a n d a r a c h i d o n i ca c i d , 6 3 1
Allotropic resonanccforms, 302
Allyalcohol, 389
- K x a n t h a t e ,q u e n c h i n ge f f e c t ,7 0 9
- mercaptan, 332
A l p h a h y d r o x y f a t t y a c i d s ,6 3 9 , 7 1 0
- and microbes. 375
- a n d t u m o r s , 3 2 6 , 5 0 9 ,7 l 0
A l p h a - t h i o f a t t y a c i d s ,3 3 8
A l p h a t o c o p h e r o l ,4 1 0
A l t e r n a t eo x i d a t i o no f f a t t y a c i d s ,l 3 l
Alternate polarity, 272
- - i n f a t t y a c i d s ,l 3 l , 6 2 7
- - a n d t w i n f o r m a t i o n , 2 T O ,6 2 7
A l u m i n u m .4 0 3
A m i n o a c i d s ,4 1 , 1 5 5 , 1 6 9 ,3 9 1 , i 4 9 , 5 5 1
- - dextrorotatory, 55 I
Aminofluorene, 274
Aminophyllin, 604
4 - a m i n o s t i l b e n ed e r i v a t i v e s ,2 7 5
A m i n o s u S a r s ,1 4 5 , 4 3 5 , 4 3 8 ,6 3 8
A m m o n i o s e l i c ,5 9 6
A m m o n i u m , 1 0 7 ,3 9 5 , 3 9 E
- c h l o r i d e .5 4 . 4 0 3
- molybdate, 399
- m o n o p h o s p h a t e4, 3 4
- a n d S . d . c .p H , 6 0 0
A m y l a s e ,1 7 2
n - A m y l a l c o h o l ,1 2 2
A m y l m e r c a p t a n ,3 3 2
A n a b o l i c p r o c e s s eas n d o x y g e n , 4 0 7
Analyses, see specific substances and
conditions
- seefatty acids

Analytical dualism,86
- patterns,90
A n a p h y l a t i cs h o c k ,l 8 l
A n d r o s t e r o n e l,4 I
A n e s t h e t i c si ,n d e x o f r e p a r t i t i o n ,1 2 5
A n g i n a p a i n a n d p r o c a i n e ,3 9 2
A n i m a l s , p l a c c i n o r g a n i z a t i o n ,3 0
A n i o n s i n S . d . c .p H , 5 9 9
A n o x y b i o s i s ,9 5
Antagonismagents,seepharmacodynamic
activity
- b e t w e e ne l e m e n t s ,1 0 4 - 1 0 7
- - falty acids-antifatty acids, I4g-

r65,362

- - positive and negative lipoids, 362

- - p a t t e r n s ,4 3 - 9 9
Anthrax. 197
A n t i - a c i dr o l e o f e o s i n o p h i l e s5, 6 7 , S 7 O
A n t i - a n e m i cl i v e r e x t r a c t ,6 0 4
Antibiotics of fungal origin, 604
A n t i b o d i e s ,a l l e r g i c , 1 7 9
- i n c a n c e r ,2 1 2 , 2 1 8
- i n i n f e c t i o u sd i s e a s e s1, 9 5
- l i p i d o - p r o t e i c ,l 8 l
- n e u t r a l i z i n gp r o t e i c , l E 2
A n t i - d i p h t h e r i as e r u m , 1 9 9
A n t i - f a t t ya c i d s ,I 3 5 , 1 4 5 - i , 6 5j .6 Z
- - a c t i o n o f c o r t i c o i d s ,3 7 2
of elements3
, 94
of glycerol,363
ccid, 368
o f g l y c e r o p h o s p h o r ia
o f s t e r o l s ,3 6 9
--and
c a r d i a cr h y t h m , 1 6 4
- - a n d c o m p l e x o r g a n i s m s ,l 5 E
-a n d c o n v u l s i o n s1. 6 3
- - a n d i n t e s t i n e sk i d n e y , 1 6 2
- - i n m i c r o b e s ,1 5 7
-i n o e s l r a lc y c l e , 1 6 4
- - in pain, 160
- - p h a r m a c o l o g y ,3 6 2
- - I n p r o t o z o a ,1 5 8
r a t i o n ,l 6 l
- - s y s t e m i cp a t t e r n st e m p e r a t u r e ,1 6 5
- - a n d v i r u s e s ,1 5 4 , 2 9 6
- - a n d w o u n d h e a l i n g ,l 6 l
Antigen antibody complex, 179
A n t i g e n s ,a f f i n i t y , l E 9
A n t i h e m o r r h a g i ca g e n t s .4 4 1, 4 4 8
A n t i h e t e r o g e n e o ur se a c t i o n ,1 7 7 , l 8 E
A n t i o x i d a n t s ,4 0 9
A n t i p n e u m o c o c c i ci m m u n e s e r a , 1 9 8
A n t i p y r e t i c s ,6 0 4
A n t i r a b i e ss e r u m , 2 0 3
A n t i t h r o m b o c y t i ca g e n t , 3 5 5
Antitryptic power in serum,674
Antityphoid serum, 201

INDEX

Anuria..ll7
Arrchitlunic acid and rdrenals.316. 373
* - - * a n d u l l o p r e g n a n e6, 3 |
- - - - - a n c lb l o t i l r e t l c e l l r . 3 1 6
* - and corticoid\, | "19
* - a \ c r s e n t i a lf a t t y a c i d , 6 ? I
--- - irnd qucnching tif llut>resccncc,
697
- * - * a n d S . d . c .p H . t r Ol
- - ' . a n d r m a l l p o xv i r u s .I l 6
- * a n d r l e r o l s ,I 1 6 . 6 l t
,\rginine, t 69
- l n d e o r i n o p h i l e r .- s 6 9
' i n r ' r ; l r r n i r ; i l i r '1r S
r.
. \ r n c l l r . l u i - r t t r r l l irl n i i I r [ . r \ l \ ,( r t ) S
,.\r.tnir: ,rnrl lrirc. ii,rlijr,tiil. J{l:
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r a r f ! ' i l l r r g u t 1t .{ t t )
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l l t i ' t , . l , r ' L :' ,

B c r r r ' l l i u n r ,J - 1 4
[ 3 e t r g k r h u l i n s ,l l l l
B e t l n r c r e i r p t o p r o p a n ot ci ci d . , 1 3 9
B e t ao r i d a t i o n .l : t t . 1 3 1 .6 1 6 . 6 l l t
- - * * . i n r a n c i d i { y ,l l t {
B et l n r d i a t i o n .l . l I
[ ] i c i i r h o n a t ci o n i i n d S . d . c .p H . 6 0 0
1 3 i o l o g i c af al c l o r s a n d S . d . c .p H . 6 0 8
- p f r l p c r l i c Joi f l i p i d s . 1 2 . 5
- r e i l l m .l 7
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l i ' , t . , i : . l ri ' t
I i . ' r t . jt ' r 1 ' . r ' l r. r , r , . { ...f r i r , r i r l r , ' Iq,.I l . J { 1 . 1
t , ' r

gllrrlrrl. lir. ,i1;'1

! l 1 L ' L 'r1r l r l 1 1 1 ' . r l t r r :; t t . i t l
I f,l

li-

750

rNDEX

Bond to fatty acids, unsaponifiablefract i o n s ,3 7 1

-sterols, 136,369
Boundary formalions, 126
- - i n b i o l o g i c a lr e a l m ,2 3
B r a d y c a r d i a ,8 3 , 1 6 5
B r a i n a n d s t e r o l s ,3 6 9
B r e a s t c a r c i n o m a ,i n f l a m a t o r yf o r m , 2 1 5
Broad scale viruses, 292
Bromine. 104
B r o t h i n j e c t i o n s ,p l e u r a l , 1 7 4
Brucellosisand copper, 359
" B u l k h e a d ' sd i s e a s e , "5 5 I
Bullet wounds and shock, 226
Burns and acid-basebalance, 50
-anuria, 427
- b u t a n o l - s o d i u ml a c t a t e ,3 t 0 , 4 2 7 , 7 2 0 ,
723
- r a d i a t i o n ,2 5 2 , 4 2 7
Butanedine,622
B u t a n o l a n d a n u r i a ,4 2 7
- and arteriosclerosis,458
- and burns. 426
- i n c a n c e r t h e r a p y ,4 9 3
- g l y c e r o p h o s p h o r iac c i d , 3 8 0
- and growth, 379
- and hemorrhage4
, 4I
- a n d i n d u c e dw o u n d s , 3 7 9
- a n d i t c h i n S ,4 3 2
- a n d l e u c o c y t e s3, 7 9 , 7 1 9
- mixed with blood,449
-_ narcotizing dose, 7 I 9
- and pain, 380, 422, 723
- a n d p l a s t i cs u r g e r y ,4 4 3 , 7 2 5
- in postoperativecare, 427, 426,723
- q u e n c h i n ge f f e c t o f , 7 0 9
- a n d r a d i a t i o nb u r n s , 3 8 0
- a n d s c h i z o p h r e n i a7, I 8
- S . d . c .p H , 6 0 2
- a n d s h o c k ,2 3 4 . 3 8 0
-5ediurn benzoate,720
- s o d i u m l a c t a t e ,3 8 0 , 4 2 7 , 7 2 0 , 7 2 3
- s u S n s p s e t i z i ndgo s e ,7 1 9
- t h e r a p y ,7 2 1 . 7 2 2
- t o x i c i t y ,3 7 8 . 7 l 8
- i n t r a u m a ,4 2 4 ,7 2 3
- tumors.493
- a n d t u m o r s c h l o r i d e s7, 1 0
B u t t e r , u n s a p o n i f i a b l ef r a c t i o n s ,3 7
B u t y l a l c o h o l s .s e e b u t a n o l
B u t y l a m i d e ,1 2 3
B u t y l k e t o n e ,l 2 3
B u t y l - m e r c a p t a n3
, 32, 709
B u t y r i ca c i d , l 2 |
C , r c o o v L e r ei o n a n d S . d , c .p H , 6 0 0

Caffeine, 604
C a l c i f e r o la n d c a l c i u m , 3 5 8
Calcium in blood, 86
- and calciferol,358
- andcancer3
, 5 6 , 3 6 1 ,3 9 5
- a n d c a r c i n o g e n s3, 5 7
- a n d c c l l u l a rm e m b r a n e .1 2 6
- c h l o r i d e ,4 0 3
- and copper,359
- a n d d u a l i s m ,E 6 , 1 0 2
- e l e m e n t s ,3 5 6
- excretionof, I 66. 595
- a n d f a t t y a c i d s ,1 2 6 , 3 5 8
- and grass tetany, 355
-index, 575, 595
- i n d u c e dc h a n g e s 3
, 57
- a n d l i v e r c e l l r e g e n e r a t i o n3, 5 6
- a n d p a t t e r n s8, 5
- a n d p h o s p h o r u s ,3 5 9
- r e t e n t i o n ,1 6 6 , 3 5 9 , 5 7 5 . 5 9 5
- a n d S . d . c .p H , 3 5 7 , 6 0 0
- and testosterone,358
-and zinc.40l
C a l o r i c m e t a b o l i s mo f f a t t y a c i d s , 1 2 9 .
I 35.361
- - r o l e o f g l y c e r o l ,3 6 3
Cancer, sce also canccrous
- seealso tumors
- a b n o r m a lg r o w t h i n , 4 0
- allergic reaction as test in, 267
- allotropic resonanceforms, 302
- a n d c a l c i u m .3 5 7 . 3 5 9
- a n d c a r b a m i ca c i d , 3 0 7
- of the cervix and circumcision3
. 0i
- and chloridecontent, 312, 710, 716
- c i r c u l a t i n gc e l l s , 2 6 6
- and copper,l l l, 360
- d i a g n o s t i ct e s t s ,2 6 5
- and divorce, 308
- d u a l i s m i n p h y s i o p a t h o l o g yo f , 9 9
- e v a l u a t i o no f r e s u l t s ,4 1 7
- a n d e m o t i o n a l r e l a t i o n s h i p ,3 0 8 , 3 0 9
- and fatty acids. 312, 323, 326, 466.
4 7 3 , 5 0 1 ,5 3 3 , 7 0 3 , 7 t 0 , 7 t 5
- s e ea l s o f a t t y a c i d s
- and genetics3
, O 2 ,3 0 7
- a s h i e r a r c h i cc o n d i t i o n , 3 8 . 2 6 8
- immunological problems, 212-Z2O
- induction, see carcinogenesis
- i n s i r u ,3 9 , 4 0 , 3 0 5 , 5 5 0
- i n v a s i v e p h a s e ,4 0 , 3 0 5
- and lipids. see specific lipids
- and lipoids, see specific lipoids
- and magnesium3
, 55
- a n d m a r i t a l s t a t u s ,3 0 8
- a n d m e r c a p t a n ,3 3 2 . 4 7 7
- ngn-inyssive phase, 39

INDEX
Cancer as organized condition, 38
- painful phase, 40
- pathogenesis,264
- physiopathologydualism, 99
- plurality of phases, 38, 40, 264
- a n d p o t a s s i u m ,3 9 6
- preterminal phase, 40
- of prostate and circumcision, 305
- o f p r o s t a t ea n d z i n c , 4 0 l
- psychologicalfactors, 308, 3 t0
- r a d i a t i o n ,2 6 1 , 2 6 2 , 5 2 4 , 6 8 3 , 7 2 6
- a n d s m e g m a ,3 0 5
-and
sterols, 148, 158, 369-373, 466,
7t5
- and sulfur, sec specific agents
- t e r m i n a l p h a s eo f , 4 l
- therapeutic approach to, see cancer
therapy
- and unsaponifiablefractions, 306,372,
468
-- and urethane. 689
- viruses.290-301
- and widows, 308
C a n c e rt h e r a p y ,a c i d l i p i d i c f r a c t i o n ,4 6 8 .
533
-b u t a n o l .4 9 3 . 5 3 3
- - chemotherapy,463, 542
- - c o d l i v e r o i l f a t t y a c i d s ,5 3 3
- - c o n d u c t o f t r e a t m e n t ,5 3 6
--conjugated
fatty acids, 501
- - criteria used, 532
- - g l y c e r o l ,4 9 3 , 5 3 3
- - group of agents in, 473
-g u i d e dt h e r a p y , 4 6 1 , 4 6 3 , 5 2 6 , 5 4 7
- - g u i d e d l e v e l c h e m o t h e r a p y5
, 26
- - m e r c a p t a n s4
, 77
- - p l a c e n t ae x t r a c t s ,4 6 3
- - p r e s e n tf o r m , 5 3 1
- - r a d i a t i o n 2, 6 1, 5 2 4 , 6 E 3 , 7 2 6
- - radiotherapy, 524
- - r e s u l t s ,4 6 3 , 5 2 6 , 5 3 7
- - selenium5
, 1 2 ,5 1 8 , 5 3 3
- - s t e r o l s ,4 6 6
- - s u l f u r i z e do i l , 4 8 1
- - t e t r a l i n ep e r s e l e n i d e5. 1 8 , 5 3 3
persulfides, 49 l , 5 33
- - t h i o s u l f a t e s4, 8 2 , 5 3 3
- - u n s a p o n i f i a b l fer a c t i o n s ,4 6 8 , 5 3 3
C a n c e r o u sa m i n o a c i d s . 3 0 7
- c e l l s ,4 0 . 3 0 3
-chromosomes, 39. 41, 303
- e n t i t i e s a s o n t o g e n i ca l l o t r o p y , 3 0 3
- h i e r a r c h i ce n t i t i c s ,4 1 , 3 0 3
Capillary hemorrhage in shock. 228
C a p r i ca c i d ,l 2 l , 6 3 9
Caproicacid, l2l, 639
C a p r y l i ca c i d , l 2 l , 6 3 9

751

C a r b a m i ca c i d , 1 7 0 , 3 0 7
Carbon, 108,334
C a r b o n a t ei o n s i n s h o c k .2 3 0
Carbonium ion, 279
Carboxyl, resonanceforms, 8
Carcinogenesis,allotropic resonance,302
- a n d a r s e n i c ,4 0 3
- and copper, 360
- and croton oil. 690
- energeticfactors, 270
- epoxides,28 I
- e x p e r i m e n t a l2, 6 8 , 3 0 7
- and genetics,302
- and host, 295
- and lipids, 304
- and lipoids, 296
- a n d n i t r o g e n m u s t a r d ,2 7 E
- a n d s y n j u g a t ef o r m a t i o n s ,2 E 3
- t w i n f o r m a t i o n s ,2 7 1
- unsaponifiablefractions, 690
- a n d u r e t h a n e ,3 0 7
- a n d v i r u s .2 9 0 . 3 0 1
Carcinogenic hydrocarbons, fl uorescence,
695
Carcinogens,see specificsubslances
- and calcium,357
- and energetic spectrum, 288
- a n d f a t t y a c i d s ,7 0 5 , 7 0 8
- m o n s t r o s i t i e s a n d m u t a t i o n s ,2 6 8
C a r d i a cr h y t h m a n d g l y c e r o l ,3 6 6
- - a n d l i p i d s , 1 6 9 ,7 I 4
- - a n d l i p o i d s ,6 6 1 ,7 1 4
C a t a b o l i cp h a s e o f w o u n d s , l 6 l
C a t a l a s e sl ,l l , 3 6 0 , 4 0 9
Cations, see elements
- of different comparlment, 27
- a n d o r g a n i z a t i o n 2, 7
- i n S . d . c .p H , 5 9 9
C e l l s ,p l a c e i n o r g a n i z a l i o n 2
,3
- c a n c e r o u s4, 0 , 3 0 3 , 3 0 5
- - i n p r o s t a t e ,3 0 5
- rapid aging, 82
C e l l u l a r a d h e s i v e n e sasn d c a l c i u m , 3 5 7
- b a s o p h i l c y t o p l a s m ,E 2
- m e m b r a n e s1, 2 7
- o x i d a t i o n ,1 6 6
- vacuolation in shock. 228
-youth, prolonged, lO2
C e l l u l o s eg u m , 7 1 5
Cenapse, lipids, 622
Ceruloplasmin and ammonium molybdate,399
- a n d c o p p e r ,3 5 9
Cervix cancerin Jewishwomen, 305
C h a i n ,r i g i d i t y , 1 3 7
Character of tumors. adenocarcinomatous,716

752

TNDEX

Characterof tumors, hemangiomatus,653

sarcomatoid, Tl6
ulcerated,653
Charcot-Leyden crystals, 567
Chemical factors and S.d.c. pH, 60t
Chemical shock and oxalic index, 248
Chemotherapy and radiotherapy, 726
Chickcnpox, 197, 295, 296, 312, 313,
6 5 4 ,6 5 5
chitf, r75, 226, 427
- and defcnse,185
- and glycerol, 366, 7 13
China wood oil fatty acid, see eleosrearic
Chlorbutanol, 404
Chlorides, bound to fatty acids, I 34, 622
- content of lesions, 102, 166
- - a n d l i p i d s ,7 1 0
-o f t u m o r s ,3 1 2 , 7 1 0
--of
w o u n d s ,2 1 0
- excretion, 166
- and gastric ulcerations,403
- index, 595
- irreversiblefixation. 134, 622
- a n d l i v e r d a m a g e ,4 0 4
- r e t e n t i o n ,1 6 6 , 2 5 4 , 4 1 9
- a n d S . d . c .p H , 6 0 0
-and shock, 231, 396, 404, 419
- in urine,86, 156, 532, 595
Chlorine,105,403
- treatment of fatty acid, 700
Chloroform. 604
Chloropropanediol,348
I C h o l a n t h r e n e - 3M e t h y l , 6 9 5
Cholera,197
C h o l e s t e r o la s a n t i - f a t t y a c i d . | 4 2
- and arteriosclerosis,453
- a n d e x p e r i m e n t a lt u m o r s , 3 7 0
-and
f o o t u l c e r a t i o n s .3 7 1
- a n d g l y c e r o l ,3 6 5
- a n d i s c h e m i ci n f a r c t . 3 7 0
- and Kuppfer cells, 456
- a n d m a n i f e s t a t i o n s3,7 0 , 7 1 5
- and microbes, 369
- and old age.646
- p a r a p l e g i ai n d u c e d b y , 6 4 4
- p h a r m a c o l o g y ,3 6 9
- p r e c i p i t a t i o n4, 5 3 . 6 6 9
- quenching effect, 709
- and RES, 456
- from scalene, 63 I
- a n d v i r u s e s .2 9 7
Choline,368
C h r o m a t o g r a p h y , g a s , f a t t y a c i d s , 627
- u n s a p o n i f i a b l ef r a c t i o n s , 3 7 3
Chromium, 399
Chromomeres,place in organization,l 7

Chromonemata,place in organization, 17,

l8
Chromosomesin cancer, 550
- p l a c e i n o r g a n i z a t i o n ,| 7 , l 8
C h y l o m i c r o n s ,1 3 5
Circulatingcancer cells. 257
C i r c u l a t i o no f f a t t y a c i d s , 1 3 5
Circumcision and cancer of the cervix,
305
- and prostatic cancer, 305
C i t r a c o n i ca c i d . 4 1 0
C i t r i c a c i d a n d S . d . c .p H . 6 0 0
C l i n i c a l m a n i f e s t a t i o n si n d e f e n s e ,1 8 5
C l o n i c c o n v u l s i o n si n s h o c k ,2 2 6
C l o t r e t r a c t i o n ,1 7 2
- - a n d a l c o h o l s .3 8 4 , 4 4 4
--in
hemoglobinuria frigore,674
C l u p a n o d o n i ca c i d s ,3 l 7
c-N-c-N 2t, 545
- a n d u r e t h a n e ,6 8 9
CO as polar group, 120
Co: combining power, blood, 46, 555
C r e a c t i v ep r o t e i n , E 6 , 1 0 2
C o a g u l a b i l i t y ,b l o o d . 1 7 2
C o a g u l a n ta n t i b o d i e s ,l 7 9
Coagulation time in hemoglobinuria a
frigore. 674
C o b a l t a n d S . d . c .p H , 6 0 0
Co-carcinogens,270, 307, 6E9
Cod liver oil fatty acids
- _- and cancer therapy, 466
- - and carcinogens,703
- - a n d c h l o r i d e s ,7 1 0
- - quenching effect, 707, 7O9
- - sedimentation red cells,
664
- - and shock.667
- - l i p o - a l c o h o l s3. 8 8
C o h e s i o nf o r c e s . I 1 4 . 3 7 8
Colds in children.6-55
Coli bacilli infection.721
C o l f o i d a l s i l v e r - p r o t e i n ,1 7 4 . 6 7 0
- s u l f u r ,3 3 4
- s u s p e n s i o n s6,5 6 , 7 | - 5
Colloids, urinary surfacetension,590. 64E
Colon and lipids. 162
Color index. red cells. 102
Colorimetrjc oxidoreduction pot. in urine.
590
Coma and lipids, 163, 6-59
Community, 548
Compartments,organization, 27
Condensationof electronsin carcinogens.

270
Conductof treatmentin cancer,535
red cells, 179
Conglutinated

rNDEx
Conjugated fatty acids and ascitescclls,
't1)

753

C o n v u l s i o n sa n d h e p t o n a l , 3 8 3
- and lipid, 163,370, 659,715
- and magnesium3
i n c a n c e r t h e r a p y ,5 0 1
, 55
- and octanediol, 3E7
a n d c a r c i n o g e n e s i s7 ,0 3
- a n d s t e r o i d ,4 4 8
and carcinogens,705
- i n d u c e d b y t h i a m i n e , 3 l 3 - 31 7 , 3 2 4 ,
from cod liver oil, 335
a n d c o n v u l s i o n s3
, 24
356.44E
and fixation of chlorides, 135
Copper, 358-360
- and calcium,356
a n d l i v e r r e g e n e r a t i o n3, 2 3
- a n d c a n c e r ,I I l , 3 6 0
a n d l y m p h a t i c s y s t e m .3 2 3
- d i s t r i b u t i o n ,1 0 9 , I I I
a n d m a n i f e s t a t i o n s3, 2 2
- - - and neoglucogenic corticoids, - i n l i v e r c e l l s . I I I
- i n p e r i o d i cc h a r t , 1 0 5
362
- a n d S . d . c .p H , 6 0 0
and oxygen,701
p h a r m a c o l o g y ,3 l 9
C o r a m i n e a n d h e m o r r h a g e ,4 4 0
a n d q u e n c h i n g o f f l u o r e s c e n c e , C o r o n a r y o c c l u s i o n ,3 7 0 , 4 5 9
- t h r o m b o s i sa n d m a g n e s i u m 3
699
, 55
and S.d.c. pH,323
C o r p u s c l e s s, u b n u c l e a r ,5 4 3
a n d s h o c k ,2 2 7
C o r t i c o i d s ,1 4 4 , 3 5 5 , 3 6 7 , 3 7 3 , 3 8 3 , 6 3 5 ,
transport of, 694
637
- and aminosugars1
t r e a t m e n t so f , 7 0 1 , 7 0 2
, 45, 437, 639
-and
and tumors,322
a r a c h i d o n i ca c i d , 1 3 9 , 1 4 3 , 6 3 1
- c o m a . 1 6 3 .6 5 9
and viruses, 322
- f o r m a t i o n s ,1 3 5 , 6 2 2
- c o n v u l s i o n s , 1 6 3 .3 8 7 , 6 5 9
- i5sms15,see conjugated fatty acids
- and defense6
, 40
- linoleic acid, 335, 639, 708
- a n d e o s i n o p h i l e s5, 6 9
- pentaenics,701
C o r t i c o s t e r o n e3, 6 5
- polyenes,283
C o r t i s o n e ,1 4 5 , 4 3 5 , 4 3 8 , 6 3 6 , 6 3 7 , 6 3 8 .
- tetraenics,700
640
- t r i e n e sa n d a d r e n a ld e f e n s ei n d e x ,6 9 5 C o t t o n s e e do i l l i p o a l c o h o l s ,3 8 8
- - a n d r a d i a t i o n .2 3 8 , 2 4 E
C r i s i s i n p n e u m o n i a l. 9 E
- - and trauma, 375
C r i t e r i a f o r t r e a t m e n t ,4 1 5 , 5 3 2
- t r i e n i c a l c o h o l s ,7 1 8
Critrcaloxalic acid index,227
C o n j u g a t i o nm e t h o d s ,1 2 9 , 1 3 3 .2 3 8 . 6 7 9 , C r o t o n o i l a n d c a r c i n o g e n e s i s6,9 0
695
C r o t o n i c a l d e h y d e ,3 3 0
Connective tissue,amount of, 102
Crotyl alcohol, 389
- - a n d b i x i n e .3 2 4
Crust of the earth, constitution, 108
- - conditions, 437
C y c l i z a t i o no f a r a c h i d o n i ca c i d , 1 3 9 , 1 4 3 ,
--and
g l u c o s a m i n e1, 4 5 , 4 3 8 , 6 4 6
631
- - healing,161
- of squalene,t 39
- - and radiation, 256
Cyclopentane forces. | 39
- group and twin formations, 273
Constancyand organization,33
C o n s t a n t so f t h e e n t i t y , l 7 l
C y c l o p e n t a n o p h e n a n t h r e noer i,g i n o f , 6 3 |
- role in organization, 28
C y s t e i n e ,3 4 0
C o n s t i p a t i o n ,E 3 , 1 0 2 , 1 6 6
C y t o c h r o m eo x i d a s e ,3 9 9
- - and copper, 3-59,360
Constitucnts, 104, 222
Constitution, crust of the certh, 108
Cytologicaldualism, 82
- of viruses. 690
C y t o l y t i c a c t i v i t y ,6 9 1 - 6 9 4
C o n s t i t u t i o n a lr o l e o f f a t t y a c i d s . 1 3 0
C y t o p l a s mi n c a n c e r ,4 0 , 5 . 5 0
C o n v u l s i o n sa n d c h o l e s t e r o l3, 7 0 . 7 1 5
C y t o p l a s m i cv i r u s . 2 9 2
- and coramine, 448
- and deoxycorticosterone,367, 659
D,ux color of blood. 678
- a n d e t h y l m e r c a p t a n3, 3 2
D e a d v i r u sv a c c i n e , 2 l 9
- and fatty acids, see specificagents
Deafness,see Impaired Hearing
- and glucose, 36E
D e a t h i n c o m a , o r c o n s c i o u s ,1 0 2
- a n d g l y c e r o l ,3 6 6 , 7 1 4
- and organization, 32
- hemorrhagiparousagents,448
- in shock. 226

754

INDEX

Decarboxylation,633
Decyl alcohol, 602
D e f e n s e ,l 7 l - 2 2 4 , 3 1 2 , 6 3 9
- adrenal index. see adrenal
* and allergic reaction, I 80
- a n t i b o d i e sl ,E 0 , l E l , l E 2
- a n t i g e n s ,I 8 9
-antiheterogenous reaction,177
- and cancer, 212
- at cellular level, 206
- role of corticoids.640
- d i p h a s i cp h e n o m e n u m ,1 7 2
- a n d d u a l i s m .2 2 3
- f a t t y a c i d s ,6 3 9
- a n d g r a f t s ,2 1 4
- and hetero- and homotropy, 224
- a n d h i e r a r c h i cl e v e l s ,1 8 4
- and infectious diseasc,195
*and lipids,176
- n o x i o u s a g e n t s ,1 7 l
-oscillatory movement, 223
- prolonged hemoshock, 176
- phylogeneticdevelopment, 222
- a n d r e t i c u l o e n d o t h e l i asl y s t e m ,2 2 3
- t i s s u e s2, 0 6
- and trauma. 640
D e f i n i t i o no f I i p i d s , I l 4
Degradatedproteins, 172
Deoxycorticorticosterol convulsive seizu r e s ,3 6 7 , 6 5 9
- defcnse.640
- and glycerol, 367
- a n d h e p t a n o l ,3 E 3
- as lipid,637
- a n d S . d . c .p H , 6 0 4
D e r m a t o t r o p i cv i r u s e s ,2 9 9 , 3 1 4
D e s a t u r a t i o no f f a t t y a c i d s , 1 2 9 , 1 3 0
- in the liver,619
- a n d r a d i a t i o n ,2 3 E
D e s m o l y s e6, 4 2
Destructionof thrombocytes,magnesium,
3_r5
D e s t r u c t i v ei n f e c t i o n , 2 9 5
D e t e r g e n t sa n d c a n c e r ,5 5 0
D e t o x i f i c a t i o ng, l u c u r o n i ca c i d , 6 4 3
- sulfuric acid, 641
Dewar structures,benzene.E
D e x t r o r o t a t o r ya m i n o a c i d s , 1 7 0
a n d c a n c e r ,5 5 1
D i a c i d f a t t y a c i d s ,6 1 8
Diagnosis of pain pattern, 364
Diagnostictestsin cancer,265
D i a r r h e a ,8 3 , 1 6 6
1 , 2 , 5 , 6D i b e n z a n t h r a c e n e
and quenching
of fluorescence,698
3 : 4 : 5 : 5 d i b e n z c a r b a z o l e2, 7 3
9 - , l 0 - d i c h l o r o s t e a r i c4, 0 4

D i c u m a r o l ,6 0 4
Di-epioxides2
, 78
Diethenoics, 3 l5
Diethylaminoethanol, 393
Diethylcarbinol, 122
Diethylene-glycol,3E7
Diethylstilbestrol, 140, 276
Differentiation of cells and cancer. 102.
291,551
Digitaline, 604
9-, l0-diiodostearic acid, 405
Dimercapto-propanol,333
Dimethanesulfoxonoxyalcanes,
27E
Di methylaminoazobenzete,275
2 : 4 Dinotrophenyl-ethylcneimine,280
Dinucleophily, 144
Diols, 387
D i p h a s i cp h e n o m e n o n ,1 7 2 , 1 7 3
- - in hemoglobinuria a frigore, 674
D i p h t h e r i a ,1 8 7 , 1 9 9
Dipolar lipidic boundaries, 126
Dipolarity of red cells in shock, 233
D i p o s i t i v ep o l a r i t y , 1 3 9
Direct action of radiation, 257
- transfusionsand shock, 226
Discharge,electric,545
D i s p l a c e m e n to f t h e d o u b l e b o n d , 1 3 5 ,
622
D i u r e s i s .1 6 6
- and lipids, 162
- and mercury, 402
DNA in viruses,690
Dodecyl mercaptan,333
Double bonds,l3l
- - c h a n g e s ,6 1 9
--conjugation,
132, 135,695
- - fixation on, 622
- - position of, 618
- - a n d s y n j u g a t i o n ,2 E 3
D u a l i s m ,3 5 , 4 3 - 1 0 3
- a b n o r m a l s u b s t a n c e s6, 0
- acid-basebalance,45
- a c i d i f y i n ga n d a l k a l i z i n ga g e n t s ,5 2
- in blood analyses,86
- a n d c a n c e r p h y s i o l o g y ,9 9
- a n d c a r d i a cr h y t h m , 1 6 4
- a t c e l l u l a r l ev c l , 8 2
- a n d c h l o r i d e e x c r e t i o n .6 l
- for coma and convulsion, 163
- in defense,223
- in dyspnea, 77
- of elements, 105
- a n d e o s i n o p h i l e s8. 6 , 5 6 7
- of fatty acids and aoti-fatty acids. l4E
- of fundamentaloffbalances,9l
- in impaired hearing, 73
- for intestines,152

INDEX

Dualism, in itching,61, 64, 562

- i n k i d n e y p h y s i o l o g y ,1 6 2
-for
lipids and lipoids, 125
- in manic depression,75
- in microbes. 175
- in nature, 35
- i n n s r v o u ss y s t e m ,1 6 2
- at organic level, 83
- oxydoreduct.pot. tissue,59
urine. 86, 590
- pain, 43, 45, 160.552
- a n d p a t t e r n s ,4 5 , 8 2
- and potassium,6 | 2
- a n d r e g e n e r a t i o n l. 6 l
- and S.d.c.pH, 597
- at systemiclevel, E3
- f o r t e m p e r a t u r e 8, 3 , I 6 4
- i n t u m o r s .9 9
- i n u r i n e a n a l y s e s8, 6 , 9 0 , 1 6 5 ,5 5 7 , 5 9 0
- a n d v i r u s e s ,1 5 4
- i n v e r t i g o ,6 7
- a n d w h e a l r e s o r p t i o n ,8 3 , 5 6 6
Duodenal fluid accumulation in shock,
228
D y n a m i c b a l a n c e ,1 7 2
Dysentery, 197
Dysoxybiosis,95
D y s p n e a6, 1 , 7 7 , 8 3 , 1 0 2
- a s a c i d b a s ep a t t e r n s ,7 8 , 7 9

'7
55

Endarterial obliterations, 370

Endodermic formations, defense,223
Endothelial proliferation and radiation,

2s6

E n d o t o x i n ,l 9 7
Energeticcentersof nonpolar group, l3l
- - i n s t c r o i d s ,6 3 5
- factors in carcinogenesis,270
-spectrum of carcinogens,2E8
E n o p h t h a l m i a ,1 0 2 , 2 2 6 , 3 7 1
E n t i t i e s ,l 2
Environmental influencc, 649
- - and surface tension, 65 I
E n z y m a t i ch y d r o l y s i s ,1 7 4
- - of procaine,393
- K n o o p b e t a o x i d a t i o n ,1 2 9
E o s i n o p h i l e s8 ,6 , 1 0 2 ,1 6 6 , 3 2 4 , 5 3 2 ,5 6 7 570
E p h e d r i n e3, 9 1
E p i c h l o r o h y d r i n ,3 4 5 , 3 4 8 , 4 2 2 , 4 3 2
- i n c a n c e r ,5 2 3 , 5 3 3
Epidemiology of poliomyelitis, 298
Epileptic seizures,magnesium, 355
Epinephrine,39l
E p i t h e l i ah e a l i n g , l 6 l
Epoxides,409
- c a r c i n o g e n s2, EI
- of fatty acids, 132, 327
E r y s i p e l a s1, 9 7 , 2 0 4
E r y t h r a l g i a ,5 5 4
E o n u , r a n d h e p t a n o l ,3 8 3
Erythrocytes,see red cells
- in shock,228
E s s e n t i a lf a t t y a c i d s , 1 3 2 , 6 2 1
- and wheal resorption, 566
E s t e r a s e ,1 7 2 , 6 7 4
Ehrlich ascitestumor, 322, 348, 691
E s t r a d i o l .1 4 0
Ehrlich mammary tumor, 715
E s t r o g e n i cp r o p e r t i e s ,1 4 0
E l e c t r i c d i s c h a r g e s5
. 45
E t h y l a l c o h o l i n c o n j u g a t i o n ,1 2 2
- b u t y r a t e ,6 7 4
Electron. 544
- transfer. 5
- m e r c a p t a n3, 3 0 - 3 3 3
E l e c t r o n i c c o n f i g u r a t i o n .e l e m e n t s , 1 0 6 - - i n t h e r a p y , 4 7 6
E l e c t r o p h o r e t i c a n a l y s i s o f a n t i b o d i e s , - s u l f a t e ,q u e n c h i n ge f f e c t o f , 7 0 9
t8l
E t h y l e n et r i t h i o c a r b a m a t eq, u e n c h i n ge f E l e c t r o s t a t i cf o r c e s i n a t o m , 5
fect, 709
- - in molecules.6. 9
Ethyleneglycol,38T
E l e m e n t s 1. 0 5 . I l 2
E t h y l e n i m i n e s 2, 8 0
- anli-A, 105,349-361
E t h y l e n i m o n i u mi o n , 1 7 9
-anri-D. 105.394-404
Evaluation of projective personality, 308
- i n o r g a n i z a t i o n .1 0 7
-of
r e s u l t si n c a n c e rt h e r a p y ,4 1 7
- p r o p e r l e v e l ,3 5 0
E v e n i n gd y s p n e a ,7 7
E l e o s t e a r i ca c i d . 3 2 0 , 3 3 5
E x c r e t i o ni n d e x . 8 6 , 4 1 5 , 5 9 4
- - and carcinogens.708
- o f s u r f a c ea c t i v es u b s t a n c e s1,6 6
--defense against,639
Exercise and blood changes,67 |
- - and oxidation, 129
E x f o l i a t i v ec y t o l o g y ,4 0 , 5 5 0
--and
S.d.cp
. H,601
E x o p h t h a l m i a1, 0 2 , 3 7 0
E l e o s t e a r i ca l c o h o l .3 8 8
Exotoxin,197
Embryos' unsaponifiable fractions, 371
Experimentalcarcinogenesis,see carcinoE m u l s i o n .1 2 4
genesis
E n c e p h a l o i dt u m o r s , 3 7 2
E x p l o r a t o r yp s y c h o t h e r a p y3, 0 E

756 /

lNoex

Extracellularpotassium,547
Extrasystoles
and lipoids,661

Fatty acids, liver regeneration,162, GjE

- - lytic activity, 364, 693
- - melting point, 130
FrorNo response,
325
- - organizational role, 135
Fall and leukoses,298
- - oxalic index, see fission
Family and organization,54g
- - o x i d a t i o n1
, 2 9 , l 3 l , 1 3 3 ,3 5 4 , 5 1 6 ,
Fate of transplants,
2l3
6lE
Fattyacids,abnormal,132,134,ZZ7,2gS, - - a n d
p a t t e r n s ,1 4 8 , 1 5 8
3 1 9 ,5 0 1 ,6 9 5 ,7 0 0
- - p a t h o g e n i cr o l e , 1 3 5
--of
a d r e n a lo r i g i n , 3 7 4 , 4 S z
- - peroxides, 129, 132
- - alphahydroxy,326,375, S09,639, - p h a r m a c o l o g y ,3 1 2
710
--positive
carbon, l3l, 132
-a l p h at h i o l , 3 3 8
- - and quenching of fluorescence,697
- - alternate polariry, l3l, 621.
- - and radiation, 239, 248, 2SB, 32?,
--anafyses,
1 3 0 , 1 3 4 , 2 2 7 , 2 g S , 622,
Sz4
626, 627, 680, 695, 700
- - r a n c i d i ( y .| 2 8 , 1 3 3 ,4 1 0
- - and Arneth's formula, 66g
--and
r e d c e l l s ,3 l S , 3 2 2 , 6 6 3 , 6 U ,
- - i n a r t e r i o s c l e r o s i s4 .5 6 . 4 5 7
665,666.678
--bound
t o g l y c e r o l 1, 3 j . 3 6 2
--role
i n m e m b r a n e .1 2 6
to glycerophosphoricacid. f 3 6
--saturated, l17, lzg,227,3lZ,362,
t o s t e r o l s ,1 3 6 , l 3 i
60t, 639
-b r e a k d o w n 5, 1 6
- - s e m i p o l a rc e n t e r i n , | 3 l
--and
c a l c i u m ,1 2 6 , 3 5 9
- - and serial treatment.7 l5
- * as caloric metabolites,l2g,
129, --and
shock, 227-233, 625. 679
t 3 0 .t 3 5
- - a n d s m a l l p o x ,1 5 4 , 1 5 5 , 3 1 6 , 3 2 2
- - a n d c a n c e r ,3 2 3 , 3 2 6 , 4 6 6 , 4 7 3 ,
501, --spectral
analysis1
, 14,626
533.703, 7t0, 7ts
- - t e t r a e n i c ,6 9 7 , 7 0 0
-a n d c h l o r i d e s1 3 5 , 6 2 2
- - and tetrahymena, 646, 656, 657
- - and chlorine, 403, 700
- - - o f c o d l i v e r o i t , 2 E 5 , 3 4 0 . 4 i 0 . 466' - - transportationin blood. 694
and tumors, see cancer
531, 695. 700
--unsaturated, l17, 130, l3l, 148- - c o n j u g a t e dI,3 3 , 2 2 7 ,2 8 5 ,j 0 l ,
695.
1 6 5 ,2 2 7 , 3 I 3 - 31 7 , 3 6 9 , 4 2 2 , 4 6 6 .
700
4
73, 6t6, 626, 640, 646, 658.
- - c o n s t i t u t i o n arl o l c , t 3 0
663, 678, 679, 680, 695, 700,
- - and cytolytic activity of sera, 694
703,715
--desaturation,
122, 129, 619
- - and viruses, 154, 296, 316, 322
- - d i a c i d ,6 1 8
Fatty infiltration in liver regeneration,
- - a n d e n o p h t h a l m i a ,3 l j
162,658
--epoxides, 132
F
e
m
a
l
e
s
e x h o r m o n e ,1 3 9 ,3 4 7 , 6 0 3
- - essential,132, 621
and cholesterol paraplegia, 371,
- - f i s s i o n ,1 3 0 , I 3 4 , 2 2 7 , 2 3 8 , 2 4 0 , 2 4 3 ,
644
2 4 5 ,2 4 8 ,2 5 1 ,2 5 3 ,3 5 4 ,6 1 6 ,6 8 0 ,
F e n e s t r a t i o n4, 3 0
682
Ferrous sulfate aqd sulfur mustard, 346
--flexibility
of rhe chain, 139,633
Fever, 173, 175, lE5, 399, 713
- - fractionation. 622. 627
- i n h e m o s h o c k .1 7 2
- - free, 134, 172, 622
- in infectious diseases.195
---functional
role, 130, 135, 365
- a n d s t e r o l s ,1 7 5
- - gas chromatography, 627
- - h e a t . o x y g e n o r s u l f u r t r e a t m e n t , F i b e r c e l l s ,5 5 1
Fibroma virus in rabbits. 295
700
- - a n d h e m o l y s i s ,6 6 3
F i c i n e ,4 4 4
- - a n d h e m o r r h a g e4, 5 0
Fish oil. 698
- - h y d r o p e r o x i d eo f , l 3 l
Fission of fatty acids, see fatty acids
- - h y d r o p e r s u l f i d e s4, 8 1 , 7 l l
F i x a t i o n o f c h l o r i d e so n f a t t y a c i d s . 1 3 5 .
- - - a n d i m m u n e a n t i b o d i e s ,1 9 2 ,6 7 |
622
- - i n t e s t i n a la b s o r p t i o n ,1 3 5
Fixed positionsof energetic centen. 137
--leucopenia, 166,316, 568
- virus,202

rNDEx
F l e x i b i l i t yo f f a t t y a c i d c h a i n . 1 3 7
F l o c c u l a t i o n ,1 7 9
F l u o r i n e , 1 0 5 ,4 0 4
F l u o r e s c e n c es, e e q u e n c h i n g o f
F o c i . c o n s t i t u t i o na n d p r o c e s s e isn . 6 0
F o o t u l c e r a t i o n sa n d c h o l e s t e r o l ,3 7 1
F o r m i c a c i d , l 2 - s ,- 5 4 5
Fractionation of f atty acids, 627
F r a c t u r e sa n d a c i d - b a s eb a l a n c e , 5 0
Free lipids, 622
F u n c t i o n a lr o l e o f f a t t y a c i d s ,1 3 0 , 1 3 5 ,
363
.3
F u n d a m e n t a lo f f b a l a n c e s 9
CrLrclosrvrNp, 145
C a l a c t o s e ,1 4 5
G a l l b l a d d e rc o l i c , 5 0 , 4 3 2
G a m m a g l o b u l i n s ,1 8 3
- r a d i a t i o n ,2 4 1
G a s e o u sg a n g r e n e ,1 9 7
G a s t r i c u l c e r a t i o n sa n d c h l o r i d e s ,4 0 3
C e n e s ,p l a c e i n o r g a n i z a t i o n ,2 5
s ,0 2
G e n e t i cf a c t o r i n c a r c i n o g e n e s i 3
Geriatry, see old age
Glioma virus, 295
C l o b u l i n i ca n t i b o d i e s 1
, 83
G l o m e r u l o n e p h r i t iisn i n f e c t i o n ,2 0 4
G l u c a g o na n d z i n c , 4 0 I
Gluco group, 145
G l u c o n a t ei o n a n d S . d . c . p H , 6 0 0
G l u c o n i ca c i d . 1 7 0
G l u c o s a m i n ei n a r t h r i t i s , 4 3 5
- a n d c o n n e c t i v et i s s u e ,4 3 E
- a n d i r i d o c y e l i t i s4, 3 9
- s y n t h e s i s1, 4 5 ,6 3 8
G l u c o s a m i n i ac c i d ,6 1 8
G l u c o s ea n d c o n v u l s i o n s 3
, 6E
- a n d h e m o r r h a g e ,4 4 0
- o x i d a t i o na b n o r m a l , I 3 2
- pH of tumors,368
- a n d p o t a s s i u m3
, 98
- a n d S . d . c .p H , 3 6 E , 6 0 3
G l u c o s i d e s6, 0 4
C l u c u r o n i ca c i d , 1 3 2 , 1 4 7 , 1 7 0
- - c o u p l i n gm e c h a n i s m 6
, 41
- - a n d s u l f u r i cr a d i c a l s5. 4 2
- - in urine,86
G l u t a t h i o n ,3 4 1
C l u t a m i ca c i d , 1 6 9
G l y c e r i ca c i d a n d a l d e h y d e ,3 6 8
s ,5 8
G l y c e r o la n d a r t e r i o s c l e r o s i 4
- c a l o r i c m e t a b o l i s m ,3 6 3
- a n d c a n c e rt h e r a p y ,4 9 3 , 5 3 3
- c a r d i a cr h y t h m , 7 1 4
- c h a n g e si n d u c e s .3 6 6
-and chills,366,713
- a n d c h o l e s t e r o l3, 6 i

757

G f y c e r o fc o n v u l s i o n s7,1 4 , 7 l 5
- a n d d e s o x y c o r t i c o s t e r o naec e t a t e ,3 6 7
-and fatty acids,135
- h e a l i n go f w o u n d s ,3 6 5
- and hemorrhage4
, 40
- h y p e r t e n s i o n3, 6 6
- a n d m o n o m o l e c u l alra y e r ,3 6 3
- pain, 364, 422
- a n d p a t t e r n s 3, 6 8
- a n d r a d i a t i o nl e s i o n s .3 6 - 5
- a n d s a t u r a t e df a t t y a c i d s , 3 6 2
- and S.d.cp
. H, 603
- a n d t r a u m a ,4 2 4
- and tumors,365
- a n d v i r u s e s ,3 6 4
G l y c e r o p h o s p h o r i ca c i d a n d c a n c e r , 5 3 3
- - b o u n d t o f a t t y a c i d s , l 3 0 , 1 3 7 ,3 6 2
- - and postoperativepain, 426, 723
- - a n d S . d . c .p H , 6 0 3
- - t h y r o i d e f f e c t s ,3 6 9
G l y c i n e ,- 5 4 5
Clycogen in foci, 60
G r a f t s o f t u m o r s i n h u m a n s .2 1 4
G r a m p o s i t i v i t ya n d l i p i d s , 1 5 7 , 3 6 9
G r a n u l o c y t e sa n d h e m o s h o c k , 1 7 2
- l y s i s ,1 7 4
G r a s s - t e t a nay n d m a g n e s i u m 3. 5 5
G r o w i n g t u m o r a l m a s s ,4 0
C r o w t h a n d b u t a n o l ,3 7 9
- i n d u c i n g f a c t o r a n d p o t a s s i u m ,3 9 6
. 65
G u i l l a u m i ni n d i c a t o r 5
(iumatous and iodine, 404
H r l o c r x l c c o m p o u n d so f f a t t y a c i d s ,3 2 8
Headaches,422, 423
H e a l i n g p r o c e s sa n d b u t a n o l , 4 2 2
- - o f t i s s u e sl,6 I
- - a n d p o l y u n s a t u r a t e da l c o h o l s , 4 2 6
- - postoperative,422, 426
- - r a d i a t i o n l e s i o n s ,3 6 5
-a n d u n s a p o n i f i a b l ef r a c t i o n s , 3 7 t
- - wounds, 365
H e a r i n g i m p a i r m e n t a s a c i d - b a s es y m p tom, 6l
- - p a t t e r n s7
. 3 , 4 3l
- - t h e r a p e u t i ce s s a y s 4, 3 0 , 7 2 4
Heart, see also cardiac system
- b l o c k a n d l i p o i d s ,6 2 2
- a n d u n s a p o n i f i a b l ef r a c t i o n , 3 7 2
H e a t i n a c t i v a t i o no f v i r u s e s ,2 9 4
- t r e a t m e n to f f a t t y a c i d , 7 0 0
H e a v y e l e m e n t s3, 6 1
H e m a n g i o m a t o utsu m o r s ,4 0 1 , 6 5 3
H e m o c l a s i a1. 7 2
H e m o c o n c e n t r a t i o inn s h o c k ,2 2 8
Hemoglobin and coppcr, 359

758

tNoex

Hemoglobinuria "a frigore", 229, 673

- - d i p h a s i cp h e n o m e n o ni n , 6 7 4
- - exercise effect. 676
- - a n d l e u c o l y s i s2
, 29, 675
- - morphine eftect in, 679
H e m o p h i l u si n f l u e n z a ,2 9 5
H e m o r r h a g e ,3 ' l l , 4 3 9 - 4 5 O
Hemorrhagiparous agents, 336, 439-440
- - and convulsions,448
Hemoshock, 172, 669, 670
Hemostypic efr.ect,444
Heparin, 569
Heptanoic acid, 656
H e p t a n o l , a c c u m u l a t i o no f f l u i d , 7 2 1
- and arteriosclerosis.458
- and burns. 426
- coma,660
- a n d i n f e c t i o n ,7 2 1
- a n d i t c h i n g ,4 3 2
- i n d u c e dc h a n g e s ,3 8 3
- and pain, 422
- a n d p o s t o p e r a t i v ep a i n , 4 2 6
- a n d S . d . c .p H , 6 0 2
- and trauma, 424
- and tumors, 384
- t h e r a p y o f c a n c e r ,5 3 3 , 5 3 4
- u r i n e a n a l y s e s ,3 8 5
H e p t y l a m i n e ,3 9 0
Heptyl aldehyde, 422
H e p t y l d i s e l e n i d ei 3
,3
Heredosyphilis,204
Heterogeneityof the transplant,206
Heterogenization of body constituents,
l7l
H e t e r o g e n o u se n t i t i e s ,1 7 9
- fatty acids, 326
- tumors,213
H e t e r o t r o p i ce l e m e n t s ,1 0 5 , 3 4 9
Heterotropy, 2
- in defense, 224
- a n d g r o w i n g t u m o r a l m a s s ,4 0
- opposing the homotropy, 221
H e x a d e c y lm e r c a p t a n ,3 3 3
Hexanol, 122, 382
H e x a v a l e nst u l f u r , 3 4 0
H e x y l a m i n e ,1 2 3 , 3 9 0
H e x y l m e r c a p t a n3, 3 2 , 3 3 3
Hexyldiselenide,533
H e x y l s e l e n o i ca c i d , 3 4 4
H e x y l t h i o n i ca c i d , 3 3 8
H i e r a r c h i co r g a n i z a t i o ,n 1 2 , 2 4 , l B 3 , 2 2 1
H i g h e r a l c o h o l sa n d a r t e r i o s c l e r o s i s4.5 8
H i s t i d i n e ,1 6 9
H i s t o n e s ,4 1 , 5 4 6
- and radiation,236
Homopolar bond, 6

H o m o t r o p i c e l e m e n t s ,1 0 5 , 3 4 9
- processesand ulceration, 224
H o r m o n e s a n d c a r c i n o g e n e s i s3,0 1
Horse radish peroxidase,409
Human placenta,see placenta
H y a l u r o n i d a s e6, 4 9
H y d r a z o d e r i v a t i v e s2
. 75
Hydrogen peroxide, 345
H y d r o i d s ,I l 9
Hydrolytic producls, 192
- d i g e s t i o ni n s h o c k , 2 2 8
- e n z y m e s ,l 9 l
- rancidity, 128
H y d r o p h o b i cg r o u p , I l 9
H y d r o n a p h t h a l e np
e e r s u l f i d e s4.9 l . 5 3 3
H y d r o p e r o x i d e s1, 2 9 , 4 0 8 , 4 0 9 , 6 1 7
- of fatty acids, I 3 I
Hydrophilicgroup, I l9
H y d r o p e r s u l f i d ei n c a n c e r t h e r a p y , 4 8 t ,
533
- a n d m i c r o b e s .3 3 4
- and pain,422
- a n d p a t t e r n s ,3 3 4
H y d r o s p h e r e ,5 4 7
Hydroxonium ion and resonance8
.
Hyperalgesia,554
H y p e r k a l e m i a3, 5 3 , 6 1 4
H y p e r c h r o m i aa n d o f t b a l a n c eA , 3 9 9
H y p e r l e u c o c y t o s i st,6 6
H y p e r t e n s i o n1, 6 6 , 1 7 2 , 1 7 3
- _ a n d a d r e n a lf a t t y a c i d s ,3 7 1
- a n d g l y c e r o l .3 6 6
Hyperthemia,| 66
H y p o c h r o m i aa n d o f f b a l a n c eD , 3 9 9
Hypokalemia,39T
H y p o t e n s i o n ,1 6 6 , 2 2 6
Hyponatremia, 397
Hypothermia,166, t74, 226
H y p n o t i c s ,6 0 4
I p r v u x e a n t i b o d i e s1, 8 2
- s e r a . 1 9 8 , 1 9 9 ,2 0 1 . 2 0 3
- s t a g e o f d e f e n s e1, 8 3
l m m u n i t y a g a i n s tv i r u s e s .2 t 8
Immunological aspectof cancer, Zl?-220
- - a g a i n s tc e l l s a n d t i s s u e s 2, 0 6
-t h e r a p e u t i ca p p r o a c h ,2 l E
I m p a i r m e n to f c i r c u l a t i o na n d s h o c k , 2 2 6
I n c r e a s e dp e r m e a b i l i t yi n s h o c k . 2 3 2
Incubation time in allergy, 185
- - i n f e c t i o u sd i s e a s e s1, 9 . 5 .1 9 6 , 1 9 7
I n d e x o f e x c r e t i o na n d r e t e n t i o n ,5 9 5
- of repartition of anesthetics,125
Indigorubine,89, 593
I n d i g o t i n ,8 9 , 5 9 3
I n d i v i d u a l i z e dt r e a t m e n r .4 l 3
Indoxyl in urine, E6, 89, 132, 591

INDEX

lnduced carcinogenesis,
268, 290, 703
- - and conjugated fatty acids, 703
- convulsions,see convulsions
- shock, 679
- tumors, see carcinogenesis
Infection and abnormal fatty acids, 319
- destructive,295
- neoplastic,295
Infectious disease, allergic and toxic
p a t h o g e n e s i s1, 9 5
- i n c u b a t i o n ,1 9 6
- s y s t e m a t i z a t i o no f , 1 9 6
Inflammatory form of breast carcinoma,
215
I n o r g a n i c a c i d s a n d S . d . c .p H , 6 0 2
I n s o m n i a8, 3 , 1 6 6 ,3 7 3
l n s u l i n a n d p o t a s s i u m ,3 9 E
-and
zinc, 401
Intake of food and itching, 62
and pain, 45
I n t e n s i t yo f s t i m u l u sa n d i t c h i n g ,5 6 2
I n t e r m e d i a r yl y s a t e s ,1 9 4
Internal sea, place in organization, 30
I n t e r s t i t i a lf l u i d s a n d e l e m e n t s ,1 0 7
- - p H a n d p a i n ,5 5
I n t e r v i e w so f c a n c e r p a t i e n t s ,3 0 8
I n t e s t i n a l a b s o r p t i o no f f a t t y a c i d s , 1 3 5
- 5sg1s1i6n5
in shock, 230
I n t e s t i n e sa n d l i p i d s , 1 6 2 , 3 7 2
I n t r a c e l l u l a rp o t a s s i u m ,5 4 7
- v a c u o l e si n s h o c k . 2 3 2
I n t r a c t a b l ed i a r r h e a ,3 7 2
I n t r a c y t o p l a s m a t ivci r u s e s ,2 9 0
I n t r a n u c l e a rv i r u s e s ,2 9 0
- vacuoles in shock, 232
I n t r a v e n o u si n j e c t i o ni n d u c i n gs h o c k ,2 2 6
I n v a s i v ep h a s eo f c a n c e r ,4 0 , 3 9 6
- - a n d p o t a s s i u m ,3 9 6
- - a n d s m e g m a ,3 0 5
- - a n d s t e r o l s ,3 0 6
I n v a s i v e n e sasn d c a l c i u m , 3 5 7
I o d i n e . 1 0 5 .4 0 4
I o n i z i n g r a d i a t i o n ,5 4 5
I r i d o c y c l i t i sa n d g l u c o s a m i n e4, 3 9
I r o n , 1 0 5 ,J 9 9 , 6 0 0 , 6 0 4
l r r a d i a t e dw o u n d , 2 5 1
I r r a d i a t i o n ,s e e R a d i a t i o n
Irreversible fixation of chlorides, 134
I s a m i n eb l u e a n d h e m o r r h a g e ,4 4 0
Ischemic infarct and cholesterol,370
Isotropic resonanceforms, 302
Itching as acid-basesymptoms, 6l
- a n d i n t e n s i t yo f s t i m u l u s ,5 6 2 , 5 6 3
- pathologicaa
l n d p h y s i o l o g i c a l6, 1 , 6 2
Krt-eltrr, 166
K a r y o r r h e x i s ,8 2 , 1 6 0

7s9

Kekulian forms in benzene,8, 3

K n o o p o x i d a t i o n ,1 2 9 , 1 3 1 , 6 1 5 , 6 l E
Kuppfer cells and cholesterol,456
Lrcrtc acid pain and foci, 60, 561
L a n t h a n i u m s e r i e s .1 0 6
Lauric acid, 601, 639
L e a d . 1 0 5 ,3 5 3 , 3 6 1
L e c i t h i n e ,1 3 0 , 3 6 2
Leprosis, 197
L e u c o c y t e sa n d b u t a n o l , 3 7 9 , 7 1 9
- collargol and morphine, 167, 670
- a n d f a t t y a c i d s ,3 l 3 , 3 1 6 , 3 2 8
- a n d m e r c a p t a n s ,3 3 2
- and offbalances,86, 102
- in shock, 174, 228, 669, 670, 671
L e u c o p e n i a ,1 0 2 , 1 5 5 , 1 7 2 , 2 2 9 , 3 1 6 , 6 7 3
- a n d e x e r c i s e ,5 7 I
- a n d l i p i d s ,6 5 8
Leukoses,29E
L e v e l c h e m o t h e r a p y ,5 2 5
I-evels in organization, 12
and arteriosclerosis,453
a n d b i o l o g i c a le n t i t i e s ,2 0 , 2 4
and boundary,16
a n d c a n c e r .3 8 , 3 0 2
a n d d e f e n s e ,1 8 3
dualistic patterns, E2
a n d e l e m e n l s ,1 0 7 , 3 4 9 , 3 9 4
and geneticsin cancer, 302
a n d i m m u n i t y ,2 1 8
immunological approach in cancer, 212
a n d l i p i d s ,1 5 4
and shock,227
a n d v i r u s e s ,6 9 0
a n d w a t e r c i r c u l a t i o n ,3 0
- s u b n u c l e a r5, 4 3
Liberationof fatty acids
- o f p o t a s s i u mi n s h o c k ,2 3 2
- o f s t e r o l s ,3 l 6
Life and organization, 32
Linoleic acid and blood patterns' 315
- - a n d c a r c i n o g e n s7, 0 E
- - and CO', 354
-a n d t h i a m i n e c o n v u l s i o n s ,3 1 5
- - d e f e n s ea g a i n s t ,6 3 9
- - as essentialfattY acid, 621
- - as lipid, 129
--and
pain,315
- - and quenching of fluorescence,697
- - radiation of. 238
- - a n d S . d . c .p H , 6 0 1
- - a n d s y s t e m i ci n f f u e n c e '3 1 5
- - a n d t u m o r s .3 1 5
L i n o l e . i ca l c o h o l ,3 8 8

760

INDEX

Linolenic acid, defenseagainst,639

Lipids and tumor chlorides, 715
-- a n d v i r u s e s ,1 5 5 . 1 5 6 , 2 9 3 , 2 9 6 , 1 2 2 .
a s e s s e n t i a fl a t t y a c i d , 6 2 1
- - as lipid, 129
372. 65s
--and
q u e n c h i n go f f l u o r e s c e n c e6,9 7 - and Walker tumors, 625
- and wound healing, 657
Lipamine, 390
Lipase in rancidity, 128
Lipid-treated antigens, 208
- - microbes.67 I
Lipidic system, 125
Lipido-proteic antigens,antibodies,l8 I
Lipoacids, I l7
- a n d a d r e n a l s .6 5 8
Lipids and agglutinines, 672
- analyses, 134, 622, 626, 627
- of blood, 3 18
- biological properties,125
- f r a c t i o n s .3 1 7
- and cancr, sec spccific lipids
- m i x t u r e s .5 3 3
- and cancerouscells, 220
- o f p l a c e n t a3
, 18
- a n d c a r c i n o g e n e s i s2,9 3 , 3 0 5
- S . d . c .p H , 6 0 1
-cardiac rhythm, 169, 714
- treatment in successive generations,
- a n d c e l l u l a r c h a n g e s ,1 6 0
717
- in cenapse, 622
L i p o a l c o h o l s ,1 2 2 , 1 2 3 , 3 7 5 . 3 E 8 . 6 0 1
- and convulsions. 163
Lipoaldehydes and pain, 422
- and cytolytic activity sera, 69 I
L i p o b a s e s ,1 1 7
- cytoplasmic, 160
Lipoidic epoxides,409
- a n d d e f e n s e ,1 5 9 , 1 7 6 , l 9 l , 2 0 5
- peroxides, 409
-definition,5t5
- p r o p e r t i e s b, i o l o g i c a l ,1 2 5
- and diuresis, 162
- - c h e m i c a l ,1 2 4
- a n d d r o s o p h i l am e l a n o g a s t c r1, 5 9
--physical, ll9
- and f.ree f.als, 622
I-ipoids, see specificsubstances
- and immunity, 671
- activity, organic level, 162
- and inductionof pain, 657
- and cancer, see specificlipoids
- and leucocytes,668
- cardiacrhythm, 661
- methods investigation, 134, 622, 626, - character. 694
- c h e m i c a l p r o p e r t i e s ,1 2 4
627
- a n d m i c r o b e s ,1 5 7 , 6 7 1
- a n d c o n v u l s i o n sa n d c o m a . 6 5 9
- and nervous systcm, 163
- and cortisone, 660
- and old age, 646
-definition, ll4, 615
-and
- with sefenium3
organs, 162
, 42, 343.344. 524, 5i3
- oestral cycles, 164
- s o l u b i l i t y ,I l 4
- and oxygen transport, 667
- a n d s u r v i v a lt i m e . 6 5 6
- and pain, 150
- and thiamine, 659
- and patterns, 657
- and thiol group, 330-340
- and phages, 656
- and tumors, see specific lipoids
- potassium and sodium blood content, - a n d t u m o r t r a n s p l a n t s 6
, 53
- and viruses, 154. 296
166
-and protozoa,l5E
Lipoid-treated microbes, 205
- in shock pathogenesis,225, 227, 229. L i p o l y t i c e n z y m e s .i n r a n c i d i t y , 1 2 8
233,625, 667
Lipophilic group, I l9
- a n d r a d i a t i o n ,2 3 6 , 2 3 8 , 2 4 2 , 2 5 2 , 2 6 2 , L i p o t h i o l s ,l l 5 , 3 3 0 . 4 7 7 , 7 0 9
625,693
L i t h i u m a n d S . d . c .p H , 6 0 0
- and red cells, 164, 662
L i t h o s p h e r ep
. o t a s s i u m5
. 47
- rouleaux and sludge forma- L i v e r a n t i a n c m i c e x t r a c t a n d S . d . c . p H .
tion, 678
604
- sedimentationrate, 662
- c e l l sa n d c o p p e r , I l I
- and serum cholesterol, 668
- cytochromeoxidaseand copper. 359
- a n d s t a r v a t i o n ,6 2 4
- damage and chlorides, 404
- and sulfur mustard, 625
- d e s a t u r a t i o n o f f a t t y a c i d s ,6 t 9
- and temperature,654
- i n s u f f i c i e n c ya n d u n s a p . f r a c t i o n s ,3 7 2
- a n d t e t r a h y m e n ap y r i f o r m i s ,6 4 6 . 6 5 6 , - r e g n e r a t i o nl 6
, l , 1 6 2 , 6 5 8 .5 5 9
- s a t u r a t i o no f f a t t y a c i d s , 6 1 9
6s7
- and transplants,209, 653
L i v i n g v a c c i n e s ,2 1 9

rNDEx

761

Local acidosisor alkalosis,60, 1 0 2

M e c h a n i c a lt r a u m a a n d s b o c k , 2 2 6
s e e a l s o S . d . c .p H
Melting point of fatty acids, 130
- anesthetics,391
Membraae pcrmeability and radiation.
- eftects of radiation, 250
253
- pH and pain, 55, 58
- - role of calciumion, 127
L o s s o f e m o t i o n a l r e l a t i o n s h i pa n d c a n - - role of lipids, 166
- with two polar faces, 126
cer, 308, 309
L u e t i n r e a c t i o n ,2 0 3
Meningococcicinfection, 197
L u g o l ' s s o l u t i o n ,4 0 5
M e n t a l e x e r c i s ea n d l i p i d s , l 5 l
Luteoids, 143, 144, 633
M e n t h o l e x c r e t i o n ,1 4 7
L y m p h a n d e l e m e n t s ,1 0 7
M e r c a p t a n s ,3 3 0 , 3 3 4
- see also Ethylmercaptan
L y m p h a t i cs y s t e ma n d a d r e n a l s 3
, 09
- - a n d e t h y l m e r c a p t a n3, 3 2
- pharmacology,330
- - in organization,l9
- and specific substances
- - and radiation, 257
Mercury in pcriodic chart. lOj
- - a n d s e l e n i u m ,3 4 3
- pharmacology, 402
- t i s s u e ,1 0 2
- and S.d.c, pH, 600
Lymphocytes,effects in vitro, 67 I
M e t a s t a s eas n d c a l c i u m . 3 5 7
- a n d r a d i a t i o n2, 1 1 , 2 5 7
- i m m u n o l o g i c a la s p e c t ,2 l 7
L y m p h o m a s a n d p s y c h o l o g i c a l c o n d i - M e l a z o i cc o m p a r l m e n ta n d e l e m e n t s ,1 0 7
- - in organization, 27
tions. 30E
L y m p h o m a t o s i sv i r u s . 2 9 5
M e t e o r o l o g i c a il n f l u e n c e .5 6 1
L y m p h o p e n i aa n d r a d i a t i o n ,2 5 7
M e t h o d o f a p p l i c a t i o no f a g e n t s ,4 1 6
Lypocytic index, 664
Methods of investigation of lipias, ttC,
L y p o l y t i c e n z y m e si n r a n c i d i t y , l 2 E
622, 626, 627
Lysine, 169
Methionine,169, 314
- and molybdenum, 399
Lysis of cancer cells, 694
4 M e t h o x y - 3 ,4 B e n z p y r e n e 6
magnesium,355
, 96
- o f g r a n u l o c y t e s1, 7 4
Methyl alcohol,t22. 602. 60E
- k e t o n e si n r a n c i d i t y , l 2 g
- of thrombi, 355
- mercaptan1
Lytic activity and lipids, 364, 694
, 23,330
Methylcholanthrena
end filtrable tumors.
M r c x e s l u u , 1 0 5 ,3 3 7 ,3 5 4 , 3 5 6 , 5 3 3 . 7 1 2
300
- forces in, 273
- a n d a d r e n a l s 1. 7 2
- induced tumors, 703
- c h a n g e si n d u c e db y , 3 5 4 . 3 5 5 , 3 _ 5 6
- q u e n c h i n go f f l u o r e s c e n coef ,
- a n d p e r i o d i cc h a r t , 1 0 5 , 3 5 3
696
- s p e c t r a la n a l y s i s ,2 9 3
- and S.d.c.pH, 600
- i n t r e a t m e n to f c a n c e r ,3 3 7 , 4 8 2 . _ 5 3 1 M e t h y l e n e b l u e , 6 0 4
M a i n t e n a n c eo f c o n s t a n t sa n d o r g a n i z a - M e t h y l e n e i m i n el : 3 : - 5T r i a z i n e ,2 g l
M e t h y l r h i o g l y c o l a t e3, 3 9
tion,33
M e z o n s ,5 4 5
Male hormone, 139
-M i c e l l e s ,l 0
a n d c h o l e s t e r o lp a r a p l e g i a ,3 7 1
- - a n d S . d . c .p H , 6 0 3
Microbes and lipids, see specific sub_
-and sulfur musta.d, 347
siances
- a n d a r a c h i d o n i ca c i d , 3 1 6
M a l e i ca c i d ,4 1 0
- a n d c o n j u g a t e df a t t y a c i d s ,3 2 2
M a l i g n a n c ya n d c a l c i u m , 3 , 5 6
- a n d e t h y l m e r c a p t a n3. 3 0
- and potassium3
, 97
- l i p i d - t r e a t e d ,6 7 1
M a n g a n e s ea n d S . d . c .p H , 6 0 0
- a n d m o l y b d e n u m ,3 9 9
- c o m p o u n d s ,4 1 0
- and phages,656
- and tumors, 360
- place in organization, 26
Manic-depressivecondition, 75
- a n d s a t u r a t e df a t t y a c i d s , 3 1 2
Marital status and cancer, 308
Migraine headaches,423
Masked virus. 294
Massivenecrosisand sulfur mustard, 346 M i l k f a c t o r s ,2 9 3
Milliar gastric ulcerations in shock, ?28,
Mature eosinophile granulae, 569
329
M e a s l e s ,1 9 7

762

INDEX

Neoglucogenic corticoids pbarmacology,

Mitochondria, 126
- m e m b r a n e .6 1 6
- pre-ferments,223
Molecular oriented layers, 123
and glycerol, 363
Molecules, forces, 6, 9
Molybdenum, pharmacological activity,
399
Mono ammonium phosphateand pain' 54
Monocellular organisms, 3 l2
- - place in organization, 26
M o n o e t h e n i cf a t t y a c i d s , 3 1 3
position of double bond, 618'
619
M o n o - g l y c e r i d e s1, 3 5
M o n s t r o s i t i e s ,1 5 9
- and carcinogens,268
- and ontogeneticallotroPY,303
Morphine and hemoglobinuria a frigore,
679
-and
sbock. 1'14,229, 670
Morphological changesas pattern, 159
Motion of particles in organization, I I
Multiple carcinogenesis,270
Multiplication and organization, 3l
- of viruses, 690
Mumps, 197
M u s c u l a re x e r c i s ea n d l i P i d s ,l 5 l
--and
s h o c k ,6 7 1
Mutations and carcinogens,268
- ontogenetic allotroPY, 303
Myocardial infarct, 7th day manifesta'
tion, 2l I
- - and oxygen, 407
- insufhciency and unsaponifiablefractions, 372
Myristic acid, 639
Myxoma virus, 295

- - template formation, 636

Nervous system and lipids, 163
N e u r a l g i a ,4 2 3
Neurolymphomatosisvirus, 295
Neutralizing antibodies,182
N e u t r o n s ,5 4 5
- in therapy, 260
NH' as polar group, 120
N i c k e l . 1 0 5 .4 0 1
N i c k e t h a m i n ea n d h e m o r r h a g e 4
, 40
- and pain, 422
- and trauma. 424
Nitrogen, 108
_. mustard and carcinogenesis,27E
- - quenching effect, 709
N i t r o g e n o u sb a s e s ,I 3 0 , 5 4 5
Nitromethane, quenching effect, 709
Nocifensor nerve system, 554
N o n a n o l ,3 8 1 . 6 0 2
N o n - i n v a s i v ec a n c e r , 3 9 , 5 5 0
Nonketonic fractions, 373
N o n n e c r o t i ct u m o r s . 1 6 0
Nonsaponifiablefractions, see unsaponifiable
Nonsaturatedfatty acids,see Unsaturated
Norbixine transportation, 694
Normality, 36
N o x i o u s s t i m u l u sa n d p a i n , 4 5
- - and shock, 225
Nuclear compartments, 23, 27, lO7
--vacuoles
and lipids, 160
N u c l e i i n n o n - i n v a s i v ec a n c e r , 5 5 0
- p l a c e i n o r g a n i z a t i o n ,1 7 , 1 8 . 2 3
Nucleoli, place in cancer, 550
- _. in organization, 23
Nuts of parinarium laurinum, 320

NrpxrsrLexe perselenide,344
--and
, 1 9 ,5 3 3
C a t h e r a p y5
2-naphthylamine, 273
Narcotics, 604
- and anti-hemorrhagic action, 448
N a s a l p H , 8 3 , 1 0 2 ,5 6 5
Nature, dualism in, in, 35
- o s c i l l a t o r Ym o v e m e n ti n , 3 4
Neck pain, 422
Necrosisof tumors, 160, 3E7
Negative phase in heart physiology, 660
--in
d e f e n s e 1. 7 3
Neoglucogeniccorticoids, 144
--and
coma,659
- - and conjugated fattY acids, 362
- - and convulsions,659
- - and defense. 640
-a n d g l u c o s a m i n e6, 3 7

l. 2. OcrlxeoIor, 387
l . 8 . O c t a n e d i o l1
, 23
Octanol. 380, 382, 602
Oestral cycles and lipids, 164
Offbalances. 93
OffbalanceA and agents,4l I
- - analyses,95
- - and arteriosclerosis,453
-a n d c a n c e r ,9 9 , 5 3 2 , 5 3 5 , 5 3 7 , 5 4 0 ,
546
--and
e l e m e n t s1, 0 5 , 3 4 9 ,3 5 3 , 3 9 4
- - manifestations,102
- - a n d s t e r o l s ,3 7 1
- - trauma. 423
- - w o u n d s .2 1 4
- - seealso specificagentsand analyses
Offbalancc D, agents,4l I
- - analyses,95

71)

rNDEx
O f f b a l a n c eD a n d c a n c e r , 9 9 , 5 3 2 , 5 3 5 .
537.540
--and
e l e m e n r s1, 0 5 .3 4 9 .3 5 3 .3 9 4
- - a n d r a d i a t i o n ,2 6 1 , 6 8 3
- - a n d t r a u m a ,4 2 3
- - seealso specificagentsand analyses
OH as polar group, 120
Old age and lipoids,646
procaine,393
w h e a l r e s o r p t i o nt i m e . 4 1 9
O l e i c a c i d , a d r e n a l d e f e n s e ,6 3 9
--bacteriophages,3l3
- - d e r m o l r o p ev i r u s e s .3 1 4
- - a n d o x i d a t i o n .1 2 9
O l e i c a l c o h o l .3 8 8
O l i g u r i a .8 3 , 1 6 6
Ontogenesis,24
O n t o g e n e t i ca l l o t r o p y , 3 0 3
O p i u m a l k a l o i d s .2 2 9 , 6 0 4
O r g a n i c l e v e l .2 3
- - a n d c a n c e r ,4 0
- - d u a l i s m ,8 3
--elements, ll0
- - m a n i f e s l a t i o n s1, 6 2 , 1 6 3 , 1 6 4
O r g a n i s ml e v e l , s e e s y s t e m i c
O r g a n i z a t i o na n d a n t i e n t r o p y ,3 , 5
- atom, 3
- basic concept, ,l
- b i o l o g i c a lr e a l m , l 7
- i n c a n c e r ,J 8 - 4 2
- a n d c a t i o n s ,2 7
- a n d c o n s t a n t s3
.3
- a n d d e a t h .3 2
- a n d h i e r a r c h i ce n t i t i e s ,1 2 . 2 0
- o f i m m u n i t y ,1 8 6
- a n d l e v e l s ,I 2
- and life, 32
- m i c e l l e s ,l 0
- m o l e c u l e s ,6
- a n d m o t i o n o f p a r t i c l e s ,t I
- a n d m u l t i p l i c a t i o n3, l
- a n d p h y l o g e n e t i cd e v e l o p m e n t
,24,222
- a n d p o l y m o l e c u l a rf o r m a t i o n s ,9
-social, 19,548
- and solar energy, 33
- s u b a t o m i c ,I I
- s u b n u c l e a r5, 4 3
- a n d w a t e r c i r c u l a t i o n .3 0
O r g a n i z a t i o nr o l e , f a t t y a c i d s , | 3 5
-g l y c e r o p h o s p h o r i ico n , 3 6 3
O r g a n i z e db o u n d a r y , l 5
O r i g i n o f a l l o p r e g n a n e3
, 61
- o f a m i n o s u g a r s6, 3 6
- o f c h o l e s t e r o l ,1 3 9
O r i e n t e d ,m o l e c u l a rl a y e r s ,1 2 3
O s c i f l a t o r ym o v e m e n t ,3 4 , l 7 Z
- - a n d d e f e n s e .2 2 3

763

Osteo-arthritis,450
- - pain and acid-basebalance,50
- - t h e r a p y ,4 3 5
O s t e o m a l a c i aa n d c o p p e r , 3 5 9
- a n d m o l y b d e n u m ,3 9 9
O t o f o g i c a lc o n d i t i o n s ,6 7 , 7 3 , 4 2 8 , 4 3 0
Oxalic index, 23E, 6E0
- a n d r a d i a t i o n ,2 4 0 , 2 4 5 , 2 S l , Z S 3 , 6 8 2
- and shock, 227, 248
O x i d a s ea n d a m m o n i u m m o l y b d a t e ,3 9 9
O x i d a t i o n ,a b n o r m a l , 1 3 2 , 1 4 7
- o f f a t t y a c i d s ,1 2 9 ,l 3 l , 1 3 3 ,3 5 4 ,6 1 6 .
6r8
O x i d a t i v ed e s a m i n a t i o n3
, 54
- fission of fatty acids, see oxalic index
O x i d i z i n g s u b s t a n c e ss, e e a l s o p e r o x i d e s
* - a n d r a d i a t i o n ,2 5 4 , S 2 S ,i 9 4 , 6 g 3
- - a n d s e l e n i u m3, 4 3
- - in urine,89, t}Z, l3Z.265-266
Oxidoreductionpotential of foci, 59
- - o f u r i n e , 8 6 , 8 9 , 1 0 2 ,5 9 0
O x y g e n ,l 0 E , 3 4 5
- and anoxybiotic process,407
- and conjugatedfatty acids,700
- fixation of, 156
- and pain pattern, 405
- sedativeeftect of, 407
- t h e r a p y ,4 0 6
- t r a n s p o r t ,6 5 7
P r r x , 4 3 - 6 1 8, 3 , 1 0 2 ,3 1 3 ,3 1 4 ,3 1 6 ,3 t 7 .
380,421,532,552
- see also specificagents
- a n d a c i d - b a s be a l a n c e4. 6 . 5 6 1
- a m m o n i u m c h l o r i d e .5 4
- i n a r r h r i t i s4
, 35
- and bixin, 324
- a n d b u t a n o l ,3 8 0
* c o n i u g a t e df a t t y a c i d s ,3 2 2
- different agents, 422
- d u a l i s m ,4 3 , 4 5
- ethylmercapta3
n3
,1
- a n d g l y c e r o l ,3 6 4
- h e p t a n o l ,4 2 2
- hydroperselenide,344
- h y d r o p e r s u l f i d e3, 3 4
- and intake of food, 45
- l i p i d sa n d l i p o i d s ,1 6 0 , 6 5 7
- and local pH, 55, 5E
- m o n o - a m m o n i u mp h o s p h a t e5, 4
- o s t e o a r t h r i t i s5.0
- a n d o x i d o r e d u c t i o np o t e n t i a l ,5 9
- and oxygen, 4O5, 407
- p a t h o l o S i c a l4, 4 , 4 7 , 5 5 3
- p a t t e r n s ,4 7 , 5 3 2 , 5 6 1
- p e r s e l e n ied, 3 4 4
- physiological,44, 552

764

INDEX

Pain and potassium, 49, 421

- propionic aldehyde, 536
- and radiation, 253
- as sensorialsensation,552
- sodium bicarbonate,54
Painful phase of cancer, 40
Palmiticacid, 601, 639
Pancreasand zinc, 401
Pancreatic secretions,230
Papilloma, 292
Para-amino-benzoic-acid.
393
Paralysis, diphtheria, 199
- of the posterior limbs in shock, 226
- induced by cholesterol,371, 644
Parasyphiliticmanifestations,204
Parinaric acid, 320, 700, 703
- - quenching effect of, 707
Parinarium laurinum secds,700
Paroxysmal hemoglobinuria, 673
Paroxystic tachycardia and lipoids, 652
Partial inactivation of viruses, 294
Pathogenicaction of fatty acids, 135
Pathogenesisof cancer, 264
Pathologicalchangesin shock, 227
- hemorrhage, 439
- i t c h i n g ,6 1 , 5 6 3
- pain, 44, 47, 553
Patterns. blood. E6
- in cancer, 99
- in dyspnea,77
- and fatty acids,148, 168
- o f i m p a i r e d h e a r i n g ,7 3 , 4 3 0
- o f i t c h i n g ,6 1 , 4 3 5
- a n d l e v e l s ,8 2 , 8 3
- a n d m a n i c d e p r e s s i v e7, 5
- of pain, 45, 421
- a n d s t e r o l s ,l 4 E . l 6 E
- i n t r a u m a ,4 2 3
- in urine analyses,86, 90
- in vertigo, 67, 428
P a u l i e x c l u s i o np r i n c i p l e ,4
Pemphigusand glucosamine,438
Pendant drop surface tension, 575
Penetrating rays, 260
P e n t a e n i cf a t t y a c i d s ,3 7 4 , 7 0 0
a n d a d r e n a l s .3 7 4
P e n t a n o l .3 8 3
Per-acids,345
Perfume rancidity, l2E
P e r i o d i cc h a r t , e l e m e n t s ,1 0 5 , 3 4 9 , 3 9 4
P e r i t o n e a lp e t e c h i a e 2, 2 8
Peritumoral fats and mercury, 402
P e r m e a b i l i t y ,c e l l m e m b r a n e a n d f a t t y
acids,129
and sterols, 127
Peroxidases,409
- in rancidity, | 2E

P e r o x i d e s ,1 4 6 , 4 0 9
-of
fatty acids, 129, 132. 327
- and iodometry, 593
- a n d r a d i a t i o n ,2 5 4 , 5 2 5 , 5 9 4 , 6 E 3 , 7 2 6
- a n d s c h i z o p h r e n i a ,4 1 9 , 5 9 4
- a n d s e l e n i u m ,3 4 3
- in urine, 86, 102, 132, 254, 265-266.
525, 592
P e r s e l e n i d e s3,4 4 , 5 1 9 , 5 3 3
P e n u f f i d e s3
, 3 6 , 4 2 2 ,4 9 1 , 5 3 3
Pertussis,197
P e t e c h i a e2, 2 8
pH, see also S.d.c. pH
-local,55, 166,316
- t u m o r s a n d g l u c o s e ,3 6 E
- urine. 90, 532, 557
Phagesand arachidonic acid, 316
- a n d g l y c e r o l ,3 6 4
- a n d l i p i d s ,6 5 6
- see also specificagenls
Pharmacology, see pharmacodynamic
activity
Pharmacodynamic activity
- - 3lnsrmal fatty acids.3l9
- - a c i d l i p i d i c f r a c t i o n s ,3 1 7
- - agents with negative polar group.
3t2, 329
- p o s i t i v ep o l a r g r o u p , 3 6 ? , 3 1 , s
- - a l c o h o l s ,3 7 5 , 3 8 0 , 3 t 9
- - a l p h a h y d r o x y f a t r y a c i d s ,3 2 6
t h i o f a t t y a c i d s ,3 3 E
- - a n t i f a t t y a c i d s c o n s t i t u e n t s3, 6 2
- - a n t i o x i d a n t s4, 0 9
- - arsenic,bismuth, mercury, 402
- - b i x i n e ,3 2 4
- - b u t a n o l ,3 7 5
- - c a l c i u m ,3 5 4
- - c a t i o n s ,3 9 5
- - chlorine, fluorine, 403
- - c o b a l t ,3 6 0
--compounds
with thiol groups, 33_5
- - copper,358
- - c o r t i c o i d s ,3 7 2
- - diols. 3E7
- - e l e m e n t s3, 4 9 , 3 6 1 , 3 9 1
- - epichlorohydrin, 348
- - fatty acids, 3 12, 326
- - glucose, glycerophosphoric acid.
368
- - g l y c e r o l ,3 5 3
- - halogencompounds fatty acids, 328
- - heavy etements,361
- - heptyl- and bexyldisclenide,343
- - hydropersulfides,334
- - iodine, 405
- - iron, 399
- - Iipoalcohols, 388

INDEX

Pharmacodynamic activity magnesium,

349
- - m a n g a n e s e3, 5 0
- - m e r c a p t a n s ,3 3 0
- - m o n o v a l e n tc a t i o n s .3 9 5
- - o x y g e n ,4 0 5
p e r o x i d a s e s 4, 0 9
polyols, 387
potassium, 399
procaine,39l
s a t u r a t e df a t t y a c i d s ,3 1 2
s e l e n i u ml i p o i d s , 3 4 2
s t e r o l s ,3 6 9
sulfur compounds, 340
- mustard, 346
t e t r a h y d r o n a p h t h a l e np
ee r s e l e n ide.1.11
- p e r s u l f i d e s3, 3 6
thioglycolic series,338
t h i o s u l f a t e s3. 3 6
- - u n s a p o n i f i a b l fer a c t i o n s ,3 7 1
- - u n s a t u r a t e df a t t y a c i d s ,3 I 3
- - zinc, 40I
P h e n o l e x c r e t i o n ,1 4 7
P h l e b i t i sa n d w h e a l r e s o r p t i o n ,5 6 6 , 5 6 1
P h o s p h a t eesx c r e t i o n ,8 6 , 1 0 2 , 1 6 6 ,5 9 5
P h o s p h o l i p i d s1 ,3 6 .3 6 9
- a n d c o p p e r ,3 5 9
P h o s p h o r u sa n d c a n c e r ,3 - 5 9
P h y l o g e n e t i ca l l o t r o p y , 3 0 2
- d e v e l o p m e na
t n d d e f e n s e ,2 2 2
- - and organization2
,4
P h y s i c a le x e r c i s ea n d s h o c k , 2 2 9 , 6 7 1
P h y s i c o m a t h e m a t i c aal p p r o a c h t o c a r cinogenesis,270
- a s p e c to f l i p o i d s ,6 1 5
Physiologicalitching, 562
- pain, 44, 552, 562
Phytosterol,369
P i t u i t a r yg l a n d sa n d c o p p e r ,3 6 0
P l a c ei n o r g a n i z a t i o n a
. n i m a l s ,3 0
b i o l o g i c a lr e a l m , 2 0
chromomeres and chromonemata. 17. l8
8enes, 26
internal sea, 30
m i c e l l e s ,l 0
microbes,26
monocellular organisms,26
n u c l e i ,1 7 , l E
nucleolus.23
p l a n t s ,3 0
p o l y m o l e c u l a rf o r m a t i o n s , 9
p r o t o p l a s m a t i cf o r m a t i o n s ,2 3
ribo-nucleic acid,23
s u b n u c l e a rr e a l m , 5 4 3
v i r u s e s ,2 6 , 6 9 0

765

Placentaextractsand cancer therapy, 463

- f a t t y a c i d s ,4 6 9
- l i p o a c i d s ,6 7 9
- u n s a p o n i f i a b lfer a c t i o n s ,2 5 7, 3 7 | , 4 2 9 ,
469, 656, 679,710, 7t5
P l a g u e ,1 9 7
P l a n t s ,p l a c e i n o r g a n i z a t i o n .3 0
P l a s t i cs u r 8 e r ya n d b u t a n o l , 4 4 3 , ' 1 2 5
P l a s m ab i c a r b o n a t,e5 5 5
P l e u r a l p e t e c h i a e2, 2 8
P l u r a l i t y o f p h a s e si n c a n c e r ,2 6 4
Pneumococcicpneumonia, 198
P n e u m o n i a ,c r i s i s , 1 9 8
Polar groups,120
P o l a r n u c l e o p h i l i cc e n t e r s .1 4 4
P o l a r i t y o f t e t r a h y m e n a ,1 5 8
P o l i o m y e l i t i s ,1 9 7
- p a r a l y s i s6
, -54
P o l y a l c o h o l s3, 8 7
P o l y c o n j u g a t e df a t t y a l c o h o l s , 3 8 8
P o l y c y c l i cc h a i n , 1 3 7
P o l y e t h y l e n ea m i n e s ,2 7 8
Polymolecular formations, place in organization, 9
Polyols,387
P o l y u n s a t u r a t e da l c o h o l s a n d a r t e r i o sclerosis, 458
- - p h a r m a c o l o g y ,3 8 8
- - and trauma, 424, 426
- - a n d t r e a t m e n to f c a n c e r ,5 3 3 , 5 3 4
- fatty acids and pain, 422
a n d s t e r o l s .3 6 9
lransport, 695
t r e a t m e n to f c a n c e r ,4 6 8
tumon. 3l3
P o l y u r i a , 8 31
, 02
P o r p h y r i n i ca c i d s , 1 4 6
P o s i t i v ea m i n o a c i d sa n d c a n c e r ,1 6 9 ,5 5 t
and radiation,236
- g 3 1 $ e n s o f f a t t y a c i d s ,l 3 l , 1 3 2 , 6 2 1
- c h a r g e dl i p o i d s , I l 8
- n e g a t i v ed i p o l a r i t y , l 4 l
- phasein hemoshock, lE3
P o s i t r o n ,5 4 4
Postoperativecare, 442
- healing period, 426
- pain, 424
- t r e a t m e n t .4 2 6
P o t a s s i u ma n d A C T H . 3 9 8
- in blood. E6
- a n d c a l c i u m .3 5 6
- i n s 6 n s g 13, 9 6 ,5 4 7
- a n d c e l l u l a r c o m p a r t m e n t .6 1 0
- content of blood serum and pain. 49
- c o n t e n ta n d l i p i d s , 1 6 6
- d i s t r i b u t i o n ,5 4 7
- and glucose,398

766

INDEX

Potassium and heart. 614

- i n h e m o s h o c k .5 1 2
- in hydrosphereand lithosphere,547
- and insulin, 398
- intracellular, 547
- iodide. 405
- a n d m e t a z o i cc o m p a r t m e n t ,6 1 0
- and meteorologicalinfluences,651
- a n d m u s c l ea c t i v i t y ,6 l l
- and offbalances,357, 613
- and pain, 49, 60
- and periodic chart, 105, I I I
- a n d r e d c e l l s ,6 l I
- S.d.c. pH, 600
- a n d s e l e n i u m ,6 1 5
-in therapy,6l5
- thiosulfate,337
- in total blood,415
Precancerouslesions,268. 550
- phase,39
P r e c i p i t a t i o n1
, 79
Prc-fermentsin mitochondria, 223
Pregnancy,urinary surface tension, 588
P r e s e n tf o r m o f c a n c e rt r e a t m e n t ,5 3 1
Preterminal phase of cancer, 40
Preventive action of elements in cancer.
355,358,360
Previruses.69l
Primary stage of defense, l7E
- s y p h i l i t i cl e s i o n ,2 0 3
- toxic processes,l7E
- tuberculous chancre, 204
Principal parts in organization, l3
Problems in cancer,264
P r o c a i n e2
, 91,604
- and angina pain, 392
- and old age, 393
P r o g e s t e r o n e1, 4 4 , 6 0 4
Proliferation of connectivetissue,407
P r o l o n g a t i o no f t h e l i f e s p a n , 1 5 9 ,6 5 6
P r o l o n g e dc e l l u l a r y o u r h , 8 2 , 1 0 2 , 6 5 8 ,
661
- h e m o s h o c k ,1 7 6
- p h a s e so f s h o c k , 1 7 6
P r o m e z o n s ,5 4 5
P r o p a n o i ca c i d , l 2 l
P r o p e r l e v e l e l e m e n t ,3 5 0
P r o p e r d i ns y s t e m ,2 1 5
- - a n d m a g n e s i u m ,3 5 5
Propionic aldehyde, 123, 329, 422, 533
Propyl alcohol, 122, 602
- isocyanide, 123
- mercaptan, 332
Prostate and zinc, 401
Prostatectomyand butanol, 723
Prostatic cancer and circumcision. 305

Protamines and heparin, 569

- a n d r a d i a t i o n .2 3 6
Protective antibodies,l9l
Protein hydrolysis and shock, 174, 227
Proteolyticenzymesand hemorrhage, 444
--and
hemoshock,174
Proteopathic sensations,itching, 562
Protoplasmatic formations, place in organization,23
Protons, 545
Psychic states,adrenals, 309
Psychologicalfactors and cancer, 308
Pycnosis,82
- and lipids, 160
Pyretogenics,604
Pyruvic acid, in foci, 60
Qur,lrrrrrve abnormality elements, 350,
351
Q u a n t a sa n d o r g a n i z a t i o n ,3 3
Q u a n t u m a n a l y s i so f c a r c i n o g e n e s i s2,7 1
Quantum forces in atom, 4
- - in micelles,l0
- - i n m o l e c u l e s ,6 , 9
- - as organizational forces, 5
- - polymolecular formations, 9
Quencbingof fluorescenceof carcinogcns,
695
- see also specifc agents
- conjugatedfatty acids,697
- conjugated tetraenes,703
- difterent agents, 709
- eleostearicacid. 699
- fish oil fatty acids, 698
- of methylcholanthrene, 695
- parinaric acid, '107
Quinine, 604
Rrnarr brain, 156
Rabies, 197. 202
R a d i a t i o n ,2 3 6 - 2 6 3 ,3 E 0 , 5 4 5
- a n d a d r e n a l .2 5 7
- and biological realm, 26, 545
- b u r n s a n d l i p o i d s ,3 8 0
- a n d c a n c e r ,2 6 1 , 2 6 2 , 5 2 9 , 6 3 8 , 7 2 6
- a n d c h e m o t h e r a p y5
, 24,726
- a n d e r y t h e m a ,2 5 0
- and fatty acids, 236, 238, 24E, 258,
2 5 4 , 3 1 9 , 3 2 2 ,5 ? 4
- induced offbalances,261, 683
- ionizing, 545
- fesions,249, 254, 256, 316
- and lipids, 236, 238, 242, 252, 262,
625, 683
- a n d l y m p h a t i c o r g a n s ,2 5 7
- and membrane permeability, 253

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R a d i l l e t j v e r r r r . l i i t m ,a { 1
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R ; r r l i o r r r i n r e l iic: l [ e t l r r f f . r l l r . ; r c i i l r , ] { g
R ; r t l i o t i r c r , r p i J. : J . ( ) 1 1
. i r i r r i r r g i r : i t l gi ru i r l c r l .l { r I
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R ; r p i r rl r g i n gl r n J c r r l c i L r nir .\ l
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R ; t t r ; r t i . ' o r t r;l rr n r le t h r l r r r c r t , r p l l r n .i 1 , 1
l t * ; r c ti r ' ; r il o n t ' f r i r L r r r s . l ( ) J
Rcclurnlrnd lrPiiir.lfrl
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R ec l c c l l r , f r r t t r r c j r l s . I I i I l t i . f r h l . f r ( r , l
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I t n t l l t p i t ' l . l r , r r i ' r p l rl . { r { r f r
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R c n i r l c . r c r et i o n , 9 { }
R c p c l t t c l p l r \ \ ; r S fu n r l v i r r r r p ; r t l r o g c r r r c i l l
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761

R r r r J r : r r ]r I i i i f ! c . : " { - i
ll i r,rtr,.rCC
t.tCC,fr(lfi
R r r p i r i r l $ r \ ( ) \ ) h i ( ) t i L n h : 1 \ ei r t l t l f r r c i , f r ( l
R r r L lrt : . e r ' , r l t r . r l i c on f , J 1 ?
[ t r l e r ]l J ( ) n1 1 1 1 1 ci )1{.1 .. 5 9 {
R l : \ r r n i l c i t r r l u s l s l t r l".1 5 { r
r n . l c l ' q n r e .l l J
R l t c r r n r i r l i ul r r ' * r . l t l l
R l t c t r r t t , i l o i rul r t h r i t i r , . 1 . 1 - 5
R h i n r r . p h ; r rnr g c l r l i n f r c t i q : r n i". { 11
R l r rt h r r r ,c ; L r t l i ; r ch,5 9
il1 nirlltrt.,1J
R l l r l n r r r ' l r . i ui r e i r l r n ( 1 t g i r n i . 1 a l i o n: l,l
R l c t t t , r l r ' r ;ur l c o h r r l .l N l { . l l { 9
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l l r ' l , r ' l l r t . r .| ( ) l
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r . i j r r L u l p u r r r r r : ; r h i l i nI .l 6
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R u h h er h l r . l r u p er o r , i t l t r . l l 9
l l r r h i r l i r r n rl ,t ) 7 , 3 9 l l
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\ . r l i c . , ' l i rr r l . i c hrrl c r r n r lp l i i r r .J 3 l
\ . r p , r 1 1 1 l i ; l l rl l1gt - l r , r t 1 r ,r r , ' t l i p r r l r C t r , tl r. n t l
: r er r l l i p t r l i , , f ' l] a i t | t t l
\ . r l r , ' r t t t t r "{ ,r { ) " 1
\ . r r i o r i l ; r i i r i r l{ l t l r f i r a t r t 0 f l t r n r t r t l . I l f r
l i , u l \ l ( r f l t l . l t l ( )onl l t t t t t t r t r J. f l
\ . r i t ; r , r l r " tfi, r l l i i i L r r l r _t ; L j r i r i ur o l c . l : \ i
r l r l e n r c . l ! " r t n \ t .6 3 9
' ' ' ' '- . ; r n r lg l l u c r , r l . . l f r l
p l t ; i r n l ; r e o l t i g l .1 l l
.
r l t r i r n r i r i i l , , ,l l N
i r n d S . r , . r .1 1Il . 6 { ) 1
\ . r l l i t . r t r r r inl l ( l r l c \ . r t r i r i r t i r ri rnr l i r r r . f r I r J
i ; { ; r r f r ) r { l } i 1 1 rgl [ r : c t . r ; i d i l l i i : n . ] - ! 6
S u h i z o p h r c n i i r. r n ( l h u t . r n r i l ,7 l E
' - - ; r r r t lr l r i n ( '
I l e t o r j d c s . ^ ll , 4 t 9
\r-lcrotir rcrir in hclring impirirntcnt, ,i)
S c r l en r n g p r ( ) j c { l i n c i r n c c r . - 1l ?
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768

./

tNoEx

Seasonalchangesin viral infections,297

S e a s o n sa n d l i p i d s , 1 5 2
S e c o n dl a w o f t h e r m o d y n a m i c s2,
Secondaryantigens,204
- - a n d e n v i r o n m e n t a il n f l u e n c e s2, 2 |
- p a r t s i n o r g a n i z a t i o n ,l 3
Sedativeeftect of oxygen, 407
Sedimentationrate and lipids, 562
Selective passage through membranes,
126
S e l e n i ca n d s e l e n i o u sa c i d s ,3 4 2
S e l e n i u mc a n c e r t h e r a p y , 5 1 2 , 5 1 8 , 5 2 6 ,
530,533,534,535
- lipoids, 342
- a n d o x i d i z i n gs u b s t a n c e s3,4 3
- p e r i o d i cc h a r t , 1 0 5
- a n d p o t a s s i u m5, 1 5
- a n d S . d . c .p H , 6 0 0
S e l f s t e r i l i z a t i o no f n e u r o - i n f e c t i o n s2, 9 4
S e m i - p o l a rc e n t e r i n f a t t y a c i d s , l 3 l
Sensorialpain, 44
S e p a r a t i n gm e m b r a n e ,6 1 6
Serial treatment of tumors. 7 | -5
S e r u ma l b u m i n , 1 7 2
- a n t i t r y p t i cp o w e r , 1 7 2 ,2 2 7
- c y t o l y t i c a c t i v i t y , 6 9I
- and pain, 49, 42I
- p o t a s s i u mI, I l . 3 5 2 , 4 1 5 , 5 3 2
S e v e n t hd a y m a n i f e s t a t i o n s2, 1 0
S e x a n d h o r m o n e s ,1 3 9
- and lipids,| 49
S e x u a li n t e r c o u r s ea n d l i p i d s , l 5 l
SH. see sulfhydryl
Shared eleclrons. 5
S h e e pp o x v i r u s , 2 9 5
S h o c k , a c u t e .2 2 6 . 6 7 9
- i n z d y s n q e dc a n c e r .4 2 0
- a n d a l k a l i n e s u b s t a n c e s3, 0
- a n d b u t a n o l .3 E 0
- a n d b u t a n o l - s o d i u ml a c t a t e .7 2 3
- a n d c a r b o n a t ei o n s , 2 3 0
- a n d c e l l u l a r v a c u o l a t i o n ,2 2 8
- a n d c h l o r i d e s ,2 3 0 , 3 9 6 , 4 0 4 , 4 1 9
- a n d c h o l e s t e r o l 3. 7 0
- a n d c l i n i c a l m a n i f e s t a l i o n s2, 2 6
- aad dark color of blood, 225. 228
- a n d d i p h a s i cp h e n o m e n o n ,2 2 7
- and fatty acids, 227-262, 319, 322,
667, 679, 680
- i n d u c t i o n ,2 2 6 , 2 3 3 . 6 7 9
- and infectious diseases.195
- and lipids,225, 227, 667
- m c c h a n i s m .2 2 7
- a n d m o r p h i n e ,1 7 4 , 2 2 9 , 6 7 0
- a n d o x a l i c i n d e x ,2 2 7 , 2 4 8 , 6 8 0
- p a t h o g e n e s i s2,2 5
- p a t h o l o g i c a cl h a n g e s 2
, 27,228, 232

Shock and physical exercise,229,671

- a n d r e d c e l l s .2 3 3
- a n d s o d i u m c h l o r i d e .2 2 9 . 2 3 1
-state of,228, 232, 404, 419, 625, 679
- a n d s t e r o l s .2 3 4
- superacute,233, 679
- t h e r a p y ,2 3 4
- types of., 225
- and unsaponifiablefractions, 372
S i l i c o na n d c a l c i u m ,3 5 5
Skin allergy, 673
S k i n w h e a l ,8 3 , 1 0 2 .4 1 9 , 5 6 6
S l e e pa n d l i p i d s , 1 5 0
S l o u g h i n go f w o u n d s ,l 6 l
S l u d g eb l o o d f o r m a t . , 1 6 6
and lipids,678
i n s h o c k .2 3 3
Smallpox and conjugatedfatty acids, 322
- a n d l i p i d s ,3 1 6 , 6 5 4 , 6 5 5
- receptivity and lipoids, 155
- a n d t e m p e r a t u r e ,1 5 6 , 6 5 4
- and unsaponifiablefractions, 372
Smegma and cancer, 305
SO: as polar group, 120
Social hierarchic organization, 548
Sodium, alkaline compounds and pain,
60
- a n d a t h e r o m a s ,3 9 5
- azide, 573
- b e n z o a t eb, u t a n o l , 7 2 0
- b i c a r b o n a t ea n d p a i n , 5 4
- c h l o r i d e a n d a d r e n a l e c t o m y7, 1 2
- - and defense,640
- - and shock.229, 231, 404, 419
- d i s t r i b u t i o n ,5 4 7
- e x c r e t i o n ,1 0 2
-- index, 595
- I a c t i r t ca n d b u t a n o,l3 E 0 , 4 2 7 , 7 2 0
- a n d p e r i o d i cc h a r t ,1 0 5 , l l l , 3 9 4
* a n d r a d i a t i o n .2 5 3
- a n d S . d . c .p H . 6 0 0
- t e t r a t h i o n a t e3, 3 E
- t h i o s u l f a t e3, 3 6 , 4 8 2 , 5 3 3 . 7 0 9
Solar energy and organization,33
S o l u b i l i t yo f l i p o i d s , I 1 4 , I l 9
S o l v e n tf r a c t i o n a t i o no f l i p i d s , 6 2 2
S o m n o l e n c e 8. 3 . 1 0 2 . 1 6 6
Speciesand phylogenetic allotropy, 302
S p e c i f i cg r a v i t y , u r i n e , E 6 , 1 0 2 , 5 3 2
S p e c t r a l a n a l y s i s ,f a t t y a c i d s , 1 3 4 , 2 3 8 .
626
- - o f l i p i d s ,5 2 6
- - m e t h y l c h o l a n t h r e n e2, 8 3
S p e r m a t o z o i das n d z i n c , 4 0 1
Splanchnicvasodilation in shock, 228
S p l e e na n d r a d i a t i o n , 2 5 7
- and RES, 175

rNDIX

.'.' 769

Squalcneand cholesterol,6.1I
S q u i de x t r a c t s ,6 7 3
S t a p e sm o b i l i z a t i o n .4 3 0
S t a r v a t i o n ,l i p i d s , 6 1 4
Stutus epilepticusand magnesium, 3-56
S t e a r i ca c i d . d e f e n s ea g a i n s t .6 3 9
S t er i c c o u p l i n g ,l 4 ? , 6 3 3
- p o s i t i o n .l 4 l
S t c r o i d s l, l 7 . 1 4 6 ,{ f i 6
. * d c r i v i n g f r o m a r a c h i d t l n i ca c i d , 6 3 |
- ' a n d f a t t y a c i d s .3 1 6
- c n c r g e l l cc e n l e r s ,6 J i

Sulfur in treatntenlgf fatty acirjs,70i)

Sulfuricanions.l{7. l"{:
S u l f u r i z c do i l , s e e h y d r o p e n u l f i d e s
- - and cancer thcr:rpy,48I
-- rerahydronaphthrlenr, sce sulfurized
tetraline
- t c r r a l i n e3, 3 6 ,. f2 1 , 4 9 1
Sulfhvtlryd
l e t e r n r i n a r i o n5 ,7 3
c x c r e l i o n ,8 6 , 1 6 6
i n c l e r .l t l . s , - 5 9 5
- - a n d r a d i a t i o n ,l - 5 7
** and iron, 399

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770

INDEX

Tetralin persulfides, 336, 422, 491, 533

- p e r s e l e n i d e? ,4 4 , 5 1 8 , 5 3 3
Tetrathiostearicacid. 335
Therapeutic approach, see Treatment
T h e r m a l s h o c k a n d o x a l i c i n d e x ,2 4 8
T h i a m i n e i n d u c e d c o n v u l s i o n sa n d f a t t y
acids.163
and thiosulfate,33T
see also specific agents
and hemorrhage, 440
Thioglycolic series, 338
Thiophene nucleus, 274
T h i o s u l f a t e sa n d c a n c e rt h e r a p y ,4 8 2 , j 3 3
- pharmacology,336, 337
- secondday wound crust pH, 600
Thorium, alpha particles,241
Thrombopenia and hemorrhagiparous
agents,441
Thrombosis and cholesterol,370
- a n d m a g n e s i u m ,3 5 5
Thymus and radiation, 257
T h y r o i d a n d g l y c e r o p h o s p h o r iacc i d . 3 6 9
Thyroidectomy, 644
Time of day, itching ad pain, 62
- - a n d l i p i d s a n d s u r f a c et e n s i o n .| 5 l
Tissue level, 27, 40, 57, 83. 3 12
Tissues,conjugatedfatty acids, 322
Titrimetric alkalinityof blood, 46, SS7
Tocopherols,410
Toluidine blue. 89
Tongue cancer and iodine. 404
T o t a l b f o o d p o t a s s i u m ,4 1 5 , S 1 - l
T o x i c m e c h a n i s mi n d e f e n s e ,1 9 6
- stage in defense, 178
Toxicity, see specificagents
T r a n s f u s i o na n d s h o c k ,2 2 6
Transitory somnolence and cholesterol.
370
T r a n s p f a n t e dt u m o r s , 6 1 5 , 7 l s
Transport of fatty acids, 695
- of norbixine, 694
- of oxygen, 657
T r a u b e s t a l a c t o m e t e r5 7 5
T r a u m a a n d a c i d - b a s eb a l a n c e .5 0
- a n d a l c o h o l s ,4 2 4
- a l k a l i n ep a t t e r n ,4 2 3
- and cholesterol.370
- a n d c o n j u g a t e dt r i e n e s ,3 7 5
- and defense.640
-- and heterogenization.206
Traumatic injuries and oxygen, 405
- p a i n a n d b u t a n o l ,7 2 3
- s h o c ka n d f a t t y a c i d s ,6 7 9
- and oxalic index, 24E
Treatment of cancer, 461-542
- - acid lipidic fractions, 468

T r e a t m e n to f a g e n t s ,5 3 3
- - butanol, 493
- - biological guided chemorherapy,
526
- - cod liver oil fatty acids,466
- - conduct of treatment, 536
- - conjugatedfatty acids, 501
- - criteria of, 532
- - glycerol, 493
- - heterogeneousagents,508
- - mercaptans,476
- - p r e s e n tf o r m , 5 3 1 - 5 J 7
- - radiation, 524
- - s e l e n i u mp r e p a r a t i o o ,5 1 2
- - sterols, 465
- - sulfurized oil, 4EI
- - tetralin perselenide,5l E
- - persulfides,491
- - thiosulfates,482
- - unsaponifiablefractions, 468
- - see also Other agents
T r e a t m e n t s ,g u i d e d ,4 1 3
- a g e n t s4
, 16
-criteria,4l5,
- c v a l u a t i o n ,4 1 7
- i n d i v i d u a l i z e d4, 1 3
- in successivegeneralions,7 I 5
- t u m o r s i n a n i m a l s ,4 1 7
- of other conditions than cancer. 4/E
- of acid-basemanifcstation, 420
- o f a l l e r g i cc o n d i t i o n s ,4 5 l
- of arteriosclerosis,457
- o f a r t h r i t i s ,4 3 5
- of burns, 426
- of gallbladder colic, 435
- o f h e a r i n g i m p a i r m e n t ,4 3 0
- of hemorrhage,439
- of itching, 435
- of pain, 42 I
- of trauma, 423
- of vertigo, 42E
Trichlorcthyleoe, 404
Trigerminal neuralgia, butanol treatment.
721
Trichloropropane, 340
Triglycerine compounds, 135
T r i p h e n y l e t h y l e n i ca c i d , 2 7 5
Tryptophane oxidation, 132
Tubercle bacilli, lipidic fracrion, 193, 205
Tuberculin, 208
Tuberculosis.203
Tularemia, 197
Tumors, see also cancer, different agents
- a n d c a l c i u m ,3 5 9
- a n d c h l o r i d ec o n t e D t ,3 1 2 , 7 1 0 , 7 1 6
- and cholesterol.370
- and conjugatedfatty acids,322

1 a
I
I t l

I N D E X

'I'umors

and copper, l l l, 360

- nnd cytolytic ocrivity. 691
* s n d d e f e n s e ,2 1 2
* and dualismpain, l0l
- e n c e p h a l o i dt r a n s f . ,3 - r 2
* and ethyl mercaptan, 333
* a n d f a t t y a c i d s .3 t 2 . 3 1 3 , 1 2 6 . 3 3 : .
d 6 6 ,4 7 1 . 5 3 3 .7 0 1 7 t 0 , 7 t 5
- fibrangiomatous,295
- a n d g l y c e r o l .3 6 5
* g r a f t s i n h u r n a n s ,2 1 4
- g r o w i n g .4 0
- hemangiomatolrlifeirm, 401
- a n d l t c p l ; r n o l .i ' l - 1
- a n c li r r r n r r ri lrrr. , t l { - - l - t t . i
- and ir,rn.,1{}(.l
i n d t t c . . . l . r 1 . q . . , r - 1 . , , , 1 1 ( . r i. ., .! .
_ _ a n d l i l r , l r ' r r , , r .r , . j . I I i
\lll'.ill,
lipid' .rltrl lrJri,rr!.. \rf
*
-

S t i tn u t r
a n d 1 1 l . r l . l 1 c \ 1 1 1 .I < i
and nr:rng;rncs*, 1(ii)

massi!c,

a n d n t r r c i l F l i r n \ . . 1l l . " l l ?
f l n d r l l r t l t \ ' l i l ) { , l . r n l l l r en u , ! 0 { ) . l l . 1
and rrrlllirdr:nulrr_ i!)9

mullinlr.:10

UItrir-violet irradiated bacteriophage,299

- r $ y h u r n s ,4 2 7
Unsaponifiable fractions ss antifatty
a c i d s ,3 7 1
- - . i n d a n u r i a ,4 2 7
* * and blood. 662
- - a n d c a n c e r ,3 t ) 6 . 3 7 2
-'. --- and cancer therapy, 468, 513
- - " c l i n i c i r le f f e c t s 3
. 71,372, 373
-..-- c()nvulsions,.{48
-- -- c),tolytic activity of sera, 694
- * * f r a c t i o n a t i o n ,6 2 3
* -'- of liver.-5JJ
r t l n j i n( r f . l r i , l - " 1
.

n e c f ( ) l l \ ' l r t t i . lt t c r r r . 1 1 1 , r 1 1 r ll jf.,,{ }
ilnd ()l I f;rrlr 'i('t'l\. ilh. : I (l
__ pilpiilrrrrrrL. :!ll
-* and
l r t r l i r s r r i r n t .1 ' l f r
*and
r : r r l r l r l i o r tl .l , l , l r ' r 1 , 5 1 1 , ( l t i , : . 1 ( r
and rlctrrl." 1"1)i, | {rt. .iiV,.i7.i. .l(r/r.
I ' \

*and
. ' \ l { , . 1 - 1 6 .' 1 : : . J f i l . l ( t l
",,1;111.
* $Cf iilrrr .pt'r'ili. irrr'rtl'.
- a n d r u l l r . r rl l r r r ' t . , t , | .i J r ,
-UlCCr:ltrrl. ili
ti{'i
* A n d l t t t s t r p r r i t r l t . rltt',l r( . l r , r r t r l, l l r i i - * 1
46li
* i f n d r j r u . r . r . : ( ) t t .i ( i i
- W a l l c r ' r . : 0 9 , , lj 6 . - { l ) l. r r l } N ,{ ) l J , r ' i I
- and /rni, -.1()l
Tung orl. i1{}
T u r p e n t i n c t r i l . i - 1i
T w i n f o r n t ; i t i r r n ,l l l . { r l l
* - i r r c i r r ei r r o t er r r . l i l
- - n r { t h } l e h d l ; i n t h r c n E 'l.l J
T y p e s t t f r r l l h t r l u r r r r r ' , ' e cl f i l . i i l ; r r r c r . \
rirrJ I)
- - g n p gj y l i ) l r l g i ( c l f ej t . J r l { l
T y p h o i d ,1 9 7 . : { } 1 . : l n

Typhus, lql
iir hnrirrrlrop ir proi:r:rr. l l l
Ulcrnlilor
- a n d l i p i r l s . l { r { 1 .3 3 . 1 ,i , 5 l

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L i r l l - ' l t l L l , : , i . . l . r l , r : ,l i { r
, l l t ' t l , , t l . . . r . ! \. i r i \ : l t i , r l ] t | l ! r
| \ ! r r t l t L ' t t , r n r l f a t c J ) 1 1 ( r tr tn t l c f , 5 l ]
i:lrrtttrrrtti;

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f ) ( t r r \ l ( l r : \ . l i ( ) . l 1 - 1. - ' i l .
f r Xi

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, t r t , . il ' i o i , d I i l r i l l ( 1 r i i a l L r r l i n i t l ' , - { 5 J
'
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l5l.

772

INDEX

Urinary surface tension and environmental temperature,652

experimental research,589
h e p t a n o l ,3 8 5
nature of factors, 5E0
, 81
in normal physiology5
and old age,419,646
i n p r e g n a n c y ,5 E 8
a n d r a d i a t i o n ,2 5 7 , 5 2 4 , 6 8 3
and stone formation, 648
technical problem, 575
urotensiometer.578
VrcuoLes in brain cells, 228
Vacuolization of red cells. 664
Valency bridge, 9
Valeric acid, l2l, 123
van der Waals cohesion forces, 24, l19,
377
Vapor fractionation of fatty acids, 134,
627
Vascular headaches,423
Vascular sclerosisand radiation, 256
Vegetative nervous system, 175
Verruga peruviana, 295
Vertigo as an acid-basesymptom, 6l
- patterns, 69, 42E
- t r e a t m e n t ,4 2 8
Virus and cancer, 290-301
- a n d c l u p a n o d o n i ca c i d , 3 1 7
- and conjugatedfatty acids,322
- c o n s t i t u t i o n ,6 9 0
- e n c e p h a l i t i s6
, 54
- f a t t y a c i d s a n d a n t i - f a t t y a c i d s ,2 9 6
- a n d g . l y c e r o l3, 6 4
- a n d h o r m o n e s ,3 0 1
- a n d t h e h o s t .2 9 5
- i n f e c t i o ni n c h i l d r e n . 1 5 6
- l i p i d s a n d t e m p e r a t u r e ,6 5 4
- and lipoids, 296, see also specific
agents

Virus pathogencity
- place in organization,26, 690
- plural activity, 293
- and seasonalchanges,297
- and tumors. 290-301
- two types, 290
V i t a m i n sA , B ' , B r , 8 " , D , E . K a n d S . d . c .
pH, 604
Volcanoes, 108
WurEn tumor, 326
--and
lipids,625
- - a n d S . d . c .p H , 6 0 8
- - variation, 653
-and zinc,401
W a t e r c i r c u l a t i o na n d o r g a n i z a t i o n .3 0
- e x c r e t i o n .1 6 6
- a n d n i t r o g e nm e t a b o l i s m ,5 9 5
W h e a l r e s o r p t i o n ,8 3 , l l 9
- - i n a b n o r m a l c o n d i t i o n .5 6 6
- - a n d o l d a g e ,4 1 9
Widows and cancer, 30E
Wounds, see also S.d.c. pH
- atonic and oxygen, 406
-and cblorides,2ll
- h e a l i n g ,l 6 l , 6 5 8
X-n.rys, see radiation
Y o u r n c h a r a c t e r s ,3 5 7 , 6 5 4
- - effect of lipids, 165
- - persistence,82, 66 I
Y o u t h a n d v i r u s e s ,1 5 6 . 6 5 4
Zrxc. 359. 401
- a n d c a l c i u m ,3 5 6
- and copper, 359
- in periodic chart, 105, 394
- p h a r m a c o l o g y .4 0 I
- and testesteratomas.298
- and various effects.401