Animal Feed Science and Technology, 3(1978) 15--22 15

Elsevier Scientific Publishing Company, Amsterdam -- Printed in The Netherlands

PROXIMATE CHEMICAL ANALYSIS OF FINAL CANE MOLASSES AND

EFFECT OF FEEDING 30% MOLASSES ON INTESTINAL SUCRASE AND
MALTASE ACTIVITIES IN THE RAT

J. LE DIVIDICH, R. CHRISTON, J. PEINIAU and A. AUMAITRE

Institut National de la Recherche Agronomique, Station de Recherches sur l'Elevage des
Porcs, 78350 Jouy-en-Josas (France)
(Received 4 March 1977)

ABSTRACT

Le Dividich, J., Christon, R., Peiniau, J. and Aumaitre, A., 1978. Proximate chemical
analysis of final cane molasses and effect of feeding 30% molasses on intestinal sucrase
and maltase activities in the rat. Anita. Feed Sci. TechnoI., 3: 15--22.

The final cane molasses used contained, on an air dry basis, 18.5% moisture, 62.1%
alcohol-soluble sugars, 32.2% sucrose, 8.6% glucose and 15% fructose. The nitrogen content
was particularly low {0.58%, out of which scarcely half was present as amino acids). The
effect of molasses on growth rate and on the activities of intestinal disaccharidases was
investigated using three groups of nine weanling rats fed for 21 days on diets containing 30%
of either final molasses, sucrose, or maize starch, respectively. Growth rate and food intake
were similar in all groups; feed/gain was similar for molasses and maize starch based diets
and significantly lower for sucrose. Molasses significantly increased the weight of intestinal
tissue and the protein content of intestine. No difference in the total and specific activity
of intestinal maltase was found between the treatments. On the contrary, a considerable
increase (P < 0.01) in both total and specific activity of sucrase occurred in rats fed on
molasses compared with those fed on sucrose or maize starch. Present data suggest that
sucrose comprising 30% of the diet does not induce intestinal sucrase adaptation, in contrast
to a similar level of molasses. Conclusions about the use of molasses for feeding monogastric
animals are drawn.

INTRODUCTION

Molasses w o u l d be an i m p o r t a n t source of c a r b o h y d r a t e s for m o n o g a s t r i c

a n i m a l f e e d i n g ( M c L e o d e t al., 1 9 6 8 ; M a r r e r o a n d L y , 1 9 7 6 ) , b u t a h i g h level
o f f i n a l c a n e m o l a s s e s i n t h e d i e t g e n e r a l l y i n d u c e s a severe d i a r r h e a i n t h e pig
( C o m b s , 1 9 7 3 ; Le D i v i d i c h e t al., 1 9 7 4 ) , i n c o n t r a s t t o t h e g o o d t o l e r a n c e o f a
h i g h level o f s u c r o s e ( B r o o k s , 1 9 7 2 ) . F o r m a n y y e a r s , a h i g h m i n e r a l c o n t e n t
(potassium in particular) was considered to be responsible for this disturbance,
b u t r e c e n t s t u d i e s w i t h pigs ( M a n e r e t al., 1 9 6 9 ) a n d c h i c k e n s ( C u e r v o e t al.,
1972) disproved this hypothesis.
S i n c e s u c r o s e is t h e m a i n c a r b o h y d r a t e i n m o l a s s e s , t h e d i g e s t i o n o f m o l a s s e s
16

involves breakdown of sucrose by the intestinal sucrase, but very little is known
about sucrase activity in the animal fed on a high level of molasses. As in man
(Weijers et al., 1961; Pink, 1967), a deficiency in this enzyme might induce
digestive disturbances and could explain the low digestibility of molasses energy
(Le Dividich et al., 1974). Besides, the fact that an appreciable a m o u n t of
soluble sugars was found in the caecum of pigs fed on "high-test molasses"
(Ly, 1975) supported the idea of a difficulty for the animal in the hydrolysis
of the carbohydrates present in final molasses.
The present study was set up to determine the composition of final cane
molasses and to study the effect of 30% molasses in the diet on the intestinal
disaccharidase activities in the rat.

MATERIAL AND METHODS

Analysis of molasses

Total alcohol-soluble sugars were determined by the anthrone m e t h o d

(Loewus, 1952) after two extractions with hot 80% ethanol, two washings
and purification of the extract. Fructose was determined by the anthrone
method at 50C (Johnson et al., 1964). True free glucose was measured by
glucose-oxidase reagent according to Huggett and Nixon (1957). Finally,
sucrose was determined, after invertase hydrolysis, by the glucose-oxidase
m e t h o d (Cerning, 1970). Phosphorus was determined by means of a m m o n i u m
phosphovanamolybdate p h o t o m e t r y , sodium and potassium by flame spectro-
metry, calcium and magnesium by atomic absorption spectrometry. Deter-
mination of amino-acids was carried out after acid hydrolysis by the standard
technique using a JEOL amino-acid analyser. Tryptophane totally destroyed
during hydrolysis was not measured.

Diets

The composition of the diets is shown in Table I. The control diet contained
77.66% maize starch. In the experimental diets 30% starch was replaced by
either 30% final cane molasses or 30% sucrose. The protein source was a
herring meal supplemented with synthetic DL methionine and L tryptophane
at a sufficient level to supply 8% crude protein in the air-dry diets.

Animals

Twenty-seven male weanling Wistar C.F. rats were fed for seven days on a
commercial stock diet, after which they were assigned individually to three
lots of nine rats each. The average body weight of each lot was 80.5 +9.1 g
at the beginning of the test. The rats were then individually fed ad libitum on
the experimental diets for 21 days. At the end of the experiment they were
starved for 12 h with free access to water and subsequently euthanized with
 17

TABLE I

C o m p o s i t i o n o f t h e diets (% as fed)

Diet

Molasses Sucrose Maize s t a r c h

Components
Molasses 30.00 -- --
Sucrose -- 30.00 --
Cornstarch 47.66 47.66 77.66
Supplement* 22.34 22.34 22.34

Analysis
Dry m a t t e r 86.7 94.1 90.4
Crude protein 9.6 8.5 8.7

* C o m p o s i t i o n (%): N o r w a y h e r r i n g ' m e a l , 11.2; arachis oil, 4.0; p u r i f i e d w o o d cellulose,

2.0; D L m e t h i o n i n e , 0.10; L t r y p t o p h a n e , 0.04; m i n e r a l m i x , 4.0; v i t a m i n m i x , 1.0
( t o t a l = 22.34).

T h e m i n e r a l m i x s u p p l i e d t h e f o l l o w i n g p e r 100 g o f diet: Ca2H2(PO,)2, 1.50 g; KH2PO+,

0.8 g; CaCo 3 , 1.05 g; NaCI, 0.35 g; M g C O 3 , 0 . 2 6 g; F e S O 4 .7 H 2 O, 2 6 . 4 0 mg; C u S O 4 . 5 -
H 2 O, 4.0 rag; M n S O 4 . H~ O, 1 0 . 8 0 rag; Z n S o 4 , 6.80 rag; CoSO 4 , 0 . 0 4 rag; KI, 0.04 mg.

T h e v i t a m i n m i x s u p p l i e d t h e f o l l o w i n g p e r 100 g of diet: r e t i n o l a c e t a t e , 1 6 0 0 I.U. ;

ergocalciferol, 320 I.U.; riboflavin, 1.5 rag; c a l c i u m p a n t o t h e n a t e , 3.0 rag; folic acid, 0.1
rag; vit K, 0.5 mg; t h i a m i n e , 0.8 mg; n i c o t i n i c acid, 3.0 rag; p y r i d o x i n e , 0.8 m g ; inositol,
50 rag; b i o t i n , 0.02 rag; vit B 1 2 , 0 . 0 3 rag; a m i n o - b e n z o i c acid, 50 rag; c h o l i n e , 100 mg.

ether. The entire small intestine was excised immediately. After the adhering
fat and mesentery had been removed, the small intestine was emptied through
gentle pressure, weighed and kept frozen ( - 1 5 C) until analysed.

Enzyme activities and units

Enzyme measurements were made on the supernatant fraction from the

whole intestine homogenized in cooled (+4C) saline with an Ultraturrax
Staufen type 18.2 for 3 minutes, and centrifuged. Sucrase and maltase
activities were determined by the m e t h o d of Dalquist (1964), using pure sucrose
and pure m o n o h y d r a t e maltose respectively in 0.1 M maleate buffer at pH 6.5.
Enzyme activities were expressed in p-moles of corresponding disaccharide
hydrolyzed per 60 minutes at 37C either by total small intestine (total
activity) or by 1 mg of intestine protein (specific activity). Protein in the en-
zyme solutions was determined b y the procedure of Lowry et al. (1951), bo-
vine serum albumin being used as a standard.
18

RESULTS AND DISCUSSION

Chemical composition of molasses

Molasses was characterized by its high soluble sugar c o n t e n t (Table II)

(L.A.T.F.C., 1974). Expressed as glucose, the sugars represented approxi m at el y
86% o f total organic matter. Sucrose was the main c a r b o h y d r a t e and the
c o n t e n t o f free fructose was twice as high as that of glucose. However, it
appeared that there was about 6% of u n k n o w n alcohol-soluble sugars. Molasses
was also rich in minerals and potassium in particular. The calcium c o n t e n t
was high p r o b ab ly because of the calcium h y d r o x i d e added during the
clarification process of cane juice; but the level of phosphorus was very low
(0.06%). From a nutritional point of view, an i m p o r t a n t characteristic was

TABLE II

Percentage composition of molasses (as fed)

Water 18.5
Alcohol-soluble sugars 62.1
Sucrose 32.2 t
Glucose 8.6 56.3
Fructose 15.5
Crude protein (N x 6.25) 3.6
Ash 9.1
Ca 0.85
P 0.06
K 2.67
Na 0.10

the very low level of crude protein (3.6%). However, higher levels (from 6 to
9%) were f o u n d in molasses from cane grown on organic soils (Combs, 1973).
Nevertheless, less than 50% of nitrogen was present in the form of amino-acids.
Thus, the biological value of molasses protein estimated from the sum of
essential and semi-essential amino acids was very low as compared to that of
whole eggs or maize (Table III). Consequently, one might omit the protein
c o n t e n t o f molasses when formulating diets based on this product.

Animal growth rate and the weight of the small intestine

Data on daily growth rate, gain per unit of ingested dry food and small
intestine weight and protein c o n t e n t are shown in Table IV. The highest rate
o f gain was on the starch diet and the lowest on the sucrose diet, but no
significant differences were f ound between the treatments. Daily dry m a t t e r
intakes were also very similar. Gain/feed was significantly higher (P < 0.05)
with maize starch than with sucrose. Thus, as in pigs (Christon and Le
 19

T A B L E III

A m i n o - a c i d c o m p o s i t i o n s (g/16 g N) o f molasses c o m p a r e d t o t h a t o f eggs and m a i z e

Molasses Whole egg Maize

Lysine* 0.6 7.1 2.9

Histidine 0.3 2.6 2.7
Arginine 0.5 6.4 4.4
Aspartic acid 22.4 9.9 6.1
Threonine 1.5 4.8 3.4
Serine 2.0 7.1 4.6
G l u t a m i c acid 10.6 12.7 17.7
Proline 1.3 4.0 10.0
Glycine 1.6 3.2 3.5
Alanine 4.6 5.6 7.2
Valine 2.3 7.0 5.0
Isoleucine 1.1 5.5 3.8
Leucine 1.3 8.5 11.8
Tyrosine 0.7 3.8 4.0
Phenylalanine 0.8 4.8 4.4
Cystine 0.6 2.8 2.4
Methionine 0.5 3.3 2.0

Total essential
and semi-essential
amino-acids m i n u s
leucine 10.2 48.1 35.0

*Essential a m i n o - a c i d s are italicized.

T A B L E IV

E f f e c t o f molasses, sucrose a n d maize starch o n w e i g h t gain, feed intake, g a i n / f e e d and

e m p t i e d small i n t e s t i n e w e i g h t *

Diet S.E.

Molasses Sucrose Maize starch

Daily gain (g) 2.13 a 1.91 a 2.32 a 0.150

Daily dry m a t t e r intake (g) 12.00 a 12.35 a 12.28 a 0.445
Gain/feed 0.18 a 0.155 0.19 a 0.007**
Weight o f e m p t i e d small
intestine
A b s o l u t e (g) 4.19 a 3.22 b 3.365 0.116"**
Relative (% o f liveweight) 3.35 a 2.71 b 2.65 b 0.096***
Total intestinal p r o t e i n (mg) 650 a 557 b 604 ab 15.347"**

*Here a n d in Table V, values in t h e same r o w n o t having t h e same s u p e r s c r i p t l e t t e r are

significantly d i f f e r e n t f r o m each o t h e r .
* * P < 0.05.
* * * P < 0.01.
20

Dividich, 1977), the present results show that 30% molasses in the diet does
n o t depress the growth performances of rats.
The absolute weight of t he e m p t y small intestine o f rats fed on molasses
was increased by 30 and 35% (P < 0.01) compared to that of rats on sucrose
and starch, respectively. Expressed as a percentage of liveweight, the increase
in weight averaged 25% (P < 0.01). The increase in protein intestine cont ent s
indicated an effective devel opm e nt of the intestinal tissue in animals fed on
molasses. The results were consistent with those of Babatunde et al. (1975),
who also f o u n d an increase in weight o f e m p y gut of pigs fed on molasses-
based diets as compared with those fed on a control, molasses-free diet.
Measurements made on only some of the animals showed that the small in-
testine o f rats fed on molasses was 8% longer than that of rats fed on sucrose
or maize starch diets. However, this was n o t sufficient to account for the in-
crease in weight and one might expect an increase in thickness of the intestinal
wall.

Sucrase and maltase activities

No significant difference was f ound in either total or specific activity o f

maltase between dietary treatments (Table V). These results were in good
agreement with those of Deren et al. (1967), who showed t hat intestinal maltase
level did n o t vary according to the kind of disaccharide (sucrose or maltose)
in the diet. Total and specific activities of sucrase were greater by 33 and 22%
respectively (P < 0.05) in rats fed on molasses than in those fed on sucrose
and were greater by 32 and 27% than with maize. Rats fed on sucrose showed

TABLE V

E f f e c t o f molasses, sucrose and m a i z e starch diets on total and specific activities o f sucrase
and maltase

Diet S.E.

Molasses Sucrose Maize starch

Total activity 1
Sucrase 1,357 a 902 b 918 b 81.5"**
Maltase 4,510 a 4,120 a 4,381 a 253.0

Specific activity ~
Sucrase 2.08 a 1.62 b 1.51 b 0.108"*
Maltase 6.93 7.40 7.26 0.376

i u-moles o f disaccharide h y d r o l y z e d in 60 rain by t h e w h o l e small i n t e s t i n e .

2 tt-moles o f disaccharide h y d r o l y z e d in 60 rain per 1 m g o f p r o t e i n .
* * P < 0.05.
* * * P < 0.01.
 21

a higher (but non-significant) sucrase activity than did rats on the starch diet.
Thus, the increased sucrase activity observed in rats fed on molasses might
indicate a possible influence of one or more components of the diet.
Sucrose has been shown to induce either an increase in sucrase activity (Blair
et al., 1963) in the rat intestine, or no improvement at all (Reddy et al., 1968;
Jones et al., 1972). In our experiment, sucrose c o n t e n t could not be implicated
since the diet based on molasses contained about 10% sucrose while the diet
based on sucrose contained 30%. No sin~ple explanation of the mechanisms
governing an increase in sucrase specific activity is available y e t in the literature.
Rosenzweig and Herman (1968) claimed that dietary fructose could control
jejunal sucrase activity in man while Phan Hoang-Huan and Le Van Hung (1975)
did not confirm the evidence of a specific effect of dietary glucose and/or
fructose on sucrase activity in rats fed on 58% carbohydrate-based diets. In
our conditions, there was evidence that the control of sucrase biosynthesis
was independent of that of maltase, which is in good agreement with recent
results for piglets (Aumaitre and Corring, 1977). The inclusion of molasses in
the diet clearly induced intestinal sucrase activity as well as lactose feeding
has been reported to do in other work (Jones et al., 1972). This effect could
not be attributed to the sucrose, glucose or fructose contained in molasses.
But, the presence in the molasses of complex (unknown) carbohydrates issued
from the action of both h o t alkali and heat treatment during sugar processing
(Binkley and Wolfrom, 1953), and which would be very difficult to split by
intestinal disaccharidases, might explain an increase in sucrase synthesis.
However that may be, the results prove an intestinal sucrase adaptation in
the rat fed on a diet based on molasses. This suggests that high levels of
molasses might be given to monogastric animals, but to allow adaptation in the
y o u n g animal, the level of molasses should be increased progressively.

ACKNOWLEDGEMENT

To Janine Jung for her help with the amino-acid determination in molasses.

REFERENCES

Aumaitre, A. and Corring, T., 1977. D e v e l o p m e n t of digestive e n z y m e s in the piglet f r o m

birth till the age of 8 weeks. II. Intestine and intestinal disaccharidases. Nutrition and
Metabolism (in press).
Babatunde, G.M., Fetuga, B.L. and Oyenuga, V.A., 1975. Effect of feeding graded levels
of cane molasses on the p e r f o r m a n c e and carcass characteristics and organ weights of
Yorkshire pigs in a tropical environment. J. Anim. Sci., 40: 632--639.
Binkley, W.W. and Wolfrom, M.L., 1953. C o m p o s i t i o n of cane juice and final molasses.
Adv. Carbohydr. Chem., 8: 291--312.
Blair, D.G., Yakimets, W. and Tuba, J., 1963. Rat intestinal sucrase. II. The effect o f rat
age and sex and diet on sucrase activity. Can. J. Biochem. Physiol., 41 : 917--929.
Brooks, C.C., 1972. Molasses, sugar (sucrose), corn, tallow, s o y b e a n oil and m i x e d fats as
sources of energy for growing swine. J. Anim. Sci., 34: 217--224.
22

Cerning, J., 1970. Contribution ~ l'~tude de l'~volution de la composition glucidique des

grains de c~r~ales au cours de leur maturation; mai~, bl~, orge. Th~se Doc. UniversitY,
Lille (France), 119 pp.
Christon, R. and Le Dividich, J., 1977. Utilisation de la m~lasse de canne ~ sucre dans
l'alimentation du porc; essai d'interpr~tation des acquisitions r~centes. Ann. Zootech.
(in press).
Christon, R. and Le Dividich, J., 1977. Effect of feeding cane molasses on nitrogen
digestibility and retention in the rat. Nutr. Rep. Int. (in press).
Combs, G.E., 1973. Cane molasses as a feed for swine. In: Livestock and Poultry in Latin
America. Seventh annual conference edited by the University of Florida, Gainesville,
FL, pp. 30F--41F.
Cuervo, C., Bushman, D.H. and Santos, D., 1972. The effect of deionization and drying
of cane molasses on their laxative action in chickens. Poultry Sci., 51 : 821--824.
Dalquist, A., 1964. Method of assay for intestinal disaccharidases. Anal. Biochem., 7: 18--25.
Deren, J.J., Broitman, S.A. and Zamcheck, N., 1967. Effect of diet upon intestinal dis-
accharidases and disaccharide absorption. J. Clin. Invest., 46: 186--195.
Huggett, A.S.G. and Nixon, D.A., 1957. Enzymatic determination of blood glucose.
Biochem. J., 66: 12P.
Johnson, G., Lambert, C., Johnson, D.K. and Sunderwirth, S.G., 1964. Colorimetric
determination of glucose, fructose and sucrose in plant material using a combination
of enzymatic and chemical methods. J. Agric. Food Chem., 12: 216--219.
Jones, D., Sosa, F.R. and Skromak, E., 1972. Effects of glucose, sucrose and lactose on
intestinal disaccharidases in the rat. J. Lab. Clin. Med., 79: 19--30.
Latin American Tables of Feed Composition (L.A.T.F.C.), 1974. L.R. McDowell, J.H.
Conrad, J.E. Thomas and L.E. Harris (Editors), University of Florida, Gainesville, FL,
509 pp.
Le Dividich, J., S~ve, B., Canope, I. and Kempf, H., 1974. Essai d'utilisation de fortes
proportions de m~lasse de canne h sucre dans les rations du porcelet sevr~ ~ 5 semaines
et du porc en croissance-finition. Journ~es de la Rech. Porcine en France, I.N.R.A.-
I.T.P. ~d., Paris, pp. 199--207.
Loewus, F.A., 1952. Improvement in anthrone method for determination of carbohydrates.
Anal. Chem., 24: 219.
Lowry, O.H., Rosebrough, N.J., Farr, A.L. and Randall, R.J., 1951. Protein measurement
with Folin phenol reagents. J. Biol. Chem., 193 : 265--276.
Ly, J., 1975. Caecal function in the pig: daily variation of caecal parameters in pigs fed
on molasses diets. Cuban J. Agric. Sci., 9: 39--47.
McLeod, N.A., Preston, T.R., Lassota, L., Willis, M.B. and Velasquez, M., 1968. Molasses
and sugar as energy source for pigs. Rev. Cubana Cienc. Agric. (Eng. ed), 2: 205--210.
Maner, J.H., Gallo, J., Corzo, M. and Buitrago, J., 1969. Effect of minerals in cane molasses
on performance and fecal moisture of pigs. J. Anim. Sci., 2 9 : 1 3 9 (abstr.).
Marrero, L. and Ly, J., 1976. Effect of different proportions of high-test and final molasses
in diets for growing pigs. I. Performance and carcass composition. Cuban J. Agric. Sci.,
10: 41--56.
Phan Hoang-Huan and Le Van Hung., 1975. R~ponses de la saccharase intestinale et
modifications physiologiques observ~es chez le rat ayant ing~r~ des r~gimes ~ diff~rents
constituants glucidiques. Ann. Nutr. Alim., 29: 395--410.
Pink, J.J., 1967. Diarrhoea due to sucrose and isomaitase deficiency. Gut, 55: 328--332.
Reddy, B.S., Pleasants, J.R. and Wostman, B.S., 1968. Effect of dietary carbohydrates on
intestinal disaccharidases in germ-free and conventional rats. J. Nutr., 95: 413--419.
Rosenzweig, N.S. and Herman, R.H., 1968. Control of jejunal sucrase and maltase activity
by dietary sucrose or fructose in man. J. Clin. Invest., 47: 2253--2262.
Weijers, H.A., Van de Kamer, J.H., Dicke, W.K. and Ijsseling, J., 1961. Diarrhoea caused by
deficiency of sugar splitting enzymes. Acta Paediat. (Uppsala), 50: 55--61.