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Safety & Security

Principles of Asepsis

Terms Defined:

Sepsis: the state of infection.

Infection: the body is invaded microorganisms which grow and causes diseases
Disease: a detectable alteration in normal tissue function
Asepsis (Aseptic): freedom from infection; infectious materials; pathogenic microorganisms. It is the
employment of techniques (such as usage of gloves, air filters, uv-rays etc) to achieve
microbe-free environment.
Medical asepsis: all practices intended to confine a specific microorganism to a specific area, limiting
the number, growth, and spread of microorganisms
Surgical asepsis
(or sterile technique): practices which keep an area or object free of all microorganisms;
Sterile: an object is said to be sterile when it’s free of pathogenic microorganism.
Pathogenic: disease causing
Antiseptics/
Bacteriostatic agents: agents that inhibit the growth of some microorganisms, but may not destroy them.
Antisepsis: is the use of chemicals (antiseptics) to make skin or mucus membranes devoid of pathogenic
microorganisms.
Decontamination: is the process of removal of contaminating pathogenic microorganisms from the articles by a
process of sterilization or disinfection. It is the use of physical or chemical means to remove,
inactivate, or destroy living organisms on a surface so that the organisms are no longer
infectious.
Disinfectants/
Bacterial Agents: agents that destroy pathogens other than spores
Disinfection is the process of elimination of most pathogenic microorganisms (excluding bacterial spores)
on inanimate objects. Disinfection can be achieved by physical or chemical methods.
Chemicals used in disinfection are called disinfectants. Different disinfectants have different
target ranges, not all disinfectants can kill all microorganisms. Some methods of disinfection
such as filtration do not kill bacteria, they separate them out.
Sterilization is an absolute condition while disinfection is not. The two are not synonymous.

Sterilization: a process that destroys all living microorganisms, including bacteria and viruses; is achieved
by physical or chemical means.

PHYSICAL AGENTS OF STERILIZATION

1. Mechanical Means - may be used to achieve a high degree of cleanliness and microbe removal, but not to
accomplish sterilization. Mechanical means include the following:
o Scrubbing - thoroughly with soap and water, and often with an additional chemical, removes
microorganisms mechanically, while the soap acts upon them chemically
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o Filtration - does not kill microbes, it separates them out. It is used to remove microbes from heat
labile liquids such as serum, antibiotic solutions, sugar solutions, urea solution. Various applications
of filtration include removing bacteria from ingredients of culture media, preparing suspensions of
viruses and phages free of bacteria, measuring sizes of viruses, separating toxins from culture
filtrates, counting bacteria, clarifying fluids and purifying hydrated fluid. Filtration is aided by using
either positive or negative pressure using vacuum pumps. The older filters made of earthenware or
asbestos are called depth filters.
o Sedimentation - is the process by which bacteria and other particles suspended in a liquid coagulate
and settle to the bottom. This process finds its practical application in water purification and sewage
treatment

2. Heat - is the most widely used and most effective means of sterilization. All species of microorganisms are
killed by heat. The conditions under which the heat is applied, the temperature achieved, the length of time
the temperature is maintained, and the species of microorganism involved are all factors affecting sterilization
by heat. The temperature that will kill a species of microorganism in 10 minutes is known as the thermal
death point. The time required to kill all microorganisms at a given temperature is known as the thermal death
time

o Dry heat - is used primarily for sterilizing objects made of glass, metal, and substances that are
damaged by moisture. For complete sterilization, a temperature of 160º-170ºC (320º-338ºF) must be
maintained for at least 1 to 3 hours. Most fabrics are damaged by this much dry heat.
 Flaming – is the method of passing an article over a gas (Bunsen flame) or alcohol flame, but
not heating it to redness. Articles such as scalpels, mouth of test tubes, flasks glass slides and cover
slips are passed through the flames a few times. This method is limited to those articles which can be
exposed to flames. However, cracking of glassware may occur.
 Incineration - is the approved method for sterilizing and disposing of contaminated materials
such as soiled dressing by burning them in an incinerator. Care must be exercised that complete
combustion of all fragments occurs before any of the residue (ash) is removed from the incinerator.

o Moist heat - provides complete sterilization at lower temperatures and in a shorter time than does dry
heat. Whereas dry heat kills by oxidation, a relatively slow process, moist heat kills by coagulation of
the protein in the microorganism--a relatively fast process. Moist heat may be applied in three ways:
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 Boiling - is the most common means of sterilization. Boiling will kill vegetative forms of
pathogenic microorganisms in 5 minutes or less. Spores are much more heat resistant, but they can be
destroyed by boiling for 15 minutes. Boiling is not considered a completely effective sterilizing agent,
since the spores of certain thermophiles (heat-loving
organisms) can survive prolonged boiling.
 Free flowing steam - provides about the same degree of effectiveness as boiling, if
applied in sufficient volume to maintain a temperature of 100ºC (212ºF).
 Steam under pressure – can attain temperatures much higher than boiling water. It is the most
effective means of sterilization - by means of the autoclave. The autoclave is an airtight cylinder in
which steam may be introduced under pressure. Whereas free-flowing steam has a temperature of 100ºC,
steam under 15 pounds of pressure per square inch has a temperature of 121.5ºC (251ºF). Steam under 15-
An 20 pounds of pressure per square inch will kill all organisms and spores in 15-45 minutes.
Autoclave

3. Cold. At very low temperatures most microorganisms lose their viability and multiply very slowly, if at all;
but most are not killed. Refrigeration has become an indispensable adjunct to the food industry, the medical
laboratory, and many other areas in which growth of microorganisms must be inhibited. Some organisms may
be killed at low temperatures--a point of importance in the storage, handling, and shipment of laboratory
specimens. On the other hand, many microorganisms can be frozen for long periods (as spores) and regain
their viability when the temperature is again favorable.

4. Desiccation. Natural or artificial drying results in the destruction of most microorganisms. However,
bacterial spores resist drying for long periods. In dehydrated foods, bacterial growth does not take place;
however, microbial action begins after the foods are reconstituted with water.

5. Radiation. Sunlight and artificially produced ultraviolet radiation have a germicidal effect on
microorganisms. However, neither one is rapid in its effect nor complete in its action. Both sunlight and
ultraviolet lamps are beneficial when used along with other sound procedures for disinfections. X-rays and
other ionizing radiation are known to be lethal to microorganisms, but their use for this purpose has not been
completely developed.

Two types of radiation are used, ionizing and non-ionizing.

Non-ionizing rays: Rays of wavelength longer than the visible light are non-ionizing. Microbicidal
wavelength of UV rays lie in the range of 200-280 nm, with 260 nm being most effective. UV rays are
generated using a high-pressure mercury vapor lamp. It is at this wavelength that the absorption by the
microorganisms is at its maximum, which results in the germicidal effect. UV rays induce formation of
thymine-thymine dimers, which ultimately inhibits DNA replication. UV readily induces mutations in cells

Ionizing rays: Ionizing rays are of two types, particulate and electromagnetic rays. Electron beams are
particulate in nature while gamma rays are electromagnetic in nature. Highspeed electrons are produced by a
linear accelerator from a heated cathode. Electron beams are employed to sterilize articles like syringes,
gloves, dressing packs, foods and pharmaceuticals. Sterilization is accomplished in few seconds. Unlike
electromagnetic rays, the instruments can be switched off.

Disadvantage includes poor penetrative power and requirement of sophisticated equipment.

CHEMICAL AGENTS
1. General. Numerous chemical agents are available as germicides, disinfectants, and antiseptics. Only a few of
the principal ones will be discussed here. Antiseptics and disinfectants may act on microorganisms in anyone
of or a combination of several ways:
 Oxidation.
 Hydrolysis.
 Combination with cell proteins to form salts.
 Coagulation of proteins
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 Disruption of the cytoplasmic membrane.

 Inhibition or inactivation of enzymes.

2. Phenol - (carbolic acid) Joseph Lister used it to prevent infection of surgical wounds. Phenols are coal-tar
derivatives. They act as disinfectants at high concentration and as antiseptics at low concentrations. The
corrosive phenolics are used for disinfection of ward floors, in discarding jars in laboratories and disinfection
of bedpans. Chlorhexidine can be used in an isopropanol solution for skin disinfection, or as an aqueous
solution for wound irrigation. It is often used as an antiseptic hand wash. 20% Chlorhexidine gluconate
solution is used for pre-operative hand and skin preparation and for general skin disinfection. Chlorhexidine
gluconate is also mixed with quaternary ammonium compounds such as cetrimide to get stronger and broader
antimicrobial effects (eg. Savlon). Chloroxylenols are less irritant and can be used for topical purposes and
are more effective against gram positive bacteria than gram negative bacteria. Hexachlorophene is
chlorinated diphenyl and is much less irritant. Triclosan is an organic phenyl ether with good activity against
gram positive bacteria and effective to some extent against many gram negative bacteria including
Pseudomonas. It also has fair activity on fungi and viruses.

Disadvantages: It is toxic, corrosive and skin irritant. Chlorhexidine is inactivated by anionic soaps.
Chloroxylenol is inactivated by hard water.

1. Alcohol - is one of the most effective and extensively used antiseptics. It is interesting to note, however, that
it is most effective at strength of 70 percent. A 70% aqueous solution is more effective at killing microbes
than absolute alcohols. At concentrations above 90 percent or below 50 percent, it is distinctively less
effective. Alcohols dehydrate cells, disrupt membranes and cause coagulation of protein. Isopropyl alcohol
is preferred to ethanol. It can also be used to disinfect surfaces. It is used to disinfect clinical thermometers.
Methyl alcohol kills fungal spores, hence is useful in disinfecting inoculation hoods.

Disadvantages: Skin irritant, volatile (evaporates rapidly), inflammable

2. Halogens. Iodine and chlorine are two of the best known and widely known disinfectants. Tincture of iodine
(two percent in alcohol solution) is used to disinfect cuts and wounds. Strong or old solutions will burn the
skin. Chlorine, in the form of sodium or calcium hypochlorite, is used in water purification, in sewage
treatment, and in disinfecting food and dairy equipment.

3. Aldehyde s- such as formaldehyde or gluteraldehyde are used for surface disinfection and fumigation of
rooms, chambers,operation theatres, biological safety cabinets, wards, sick rooms etc. Fumigation is achieved
by boiling formalin, heating paraformaldehyde or treating formalin with potassium permanganate. It also
sterilizes bedding, furniture and books. 10% formalin with 0.5% tetraborate sterilizes clean metal instruments.
2% gluteraldehyde is used to sterilize thermometers, cystoscopes, bronchoscopes, centrifuges, anasethetic
equipments etc. An exposure of at least 3 hours at alkaline pH is required for action by gluteraldehyde.

Disadvantages: Vapors are irritating (must be neutralized by ammonia), has poor penetration, leaves non-
volatile residue, activity is reduced in the presence of protein.

4. Ethylene oxide – is a gas which destroys microorganisms by interfering with their metabolic processes.
Unfortunately spores and some viruses are not killed by this method and very toxic to humans.

5. Detergents. Soap and numerous other detergents act upon microorganisms by reducing the surface tension of
the water in which they are used, thereby reducing the ability of the microbial cell membrane to control the
passage of fluids in and out of the cell by osmosis. Soap tends to neutralize the effect of disinfectants;
therefore, it should be thoroughly rinsed off before an antiseptic or disinfectant is applied.
Hexachlorophene, a phenol derivative, is not affected by soap; therefore, it is incorporated
into soaps and detergents used in scrubbing hands of surgical personnel and the skin of
surgical patients. Detergents are not true disinfectants, but are used as adjuncts to other agents
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in order to remove organic matter that might otherwise interfere with the action of the
antiseptic or disinfectant. Detergents are also the principal agents employed in sanitizing food
service equipment, eating utensils, and patient care items not requiring sterility (bed pans,
water pitchers, and so forth)

6. Antibiotics. Numerous chemotherapeutic agents, including penicillin, streptomycin, tetracycline, the

sulfonamides, and many others, are used to inhibit or kill microorganisms which have already gained entrance
into the body of a human or animal host. This usage, however, falls within the realm of therapy, rather than
disinfection and sterilization.