WORLD HEALTH ORGANIZATION
WHO Food Additives Series 1972, No. 1
TOXICOLOGICAL EVALUATION OF SOME
ENZYMES, MODIFIED STARCHES AND
CERTAIN OTHER SUBSTANCES
The evaluations contained in this publication were prepared by the
Joint FAO/WHO Expert Committee on Food Additives which met in Rome,
16-24 June 19711
World Health Organization
1 Fifteenth Report of the Joint FAO/WHO Expert Committee on Food
Additives, Wld Hlth Org. techn. Rep. Ser., 1972, No. 488; FAO
Nutrition Meetings Report Series, 1972, No. 50.
The monographs contained in the present volume are also issued by the
Food and Agriculture Organization of the United Nations, Rome, as FAO
Nutrition Meetings Report Series, No. 50A
(c) FAO and WHO 1972
ESTERS OF GLYCEROL AND THERMALLY OXIDIZED SOY BEAN FATTY ACIDS
Groups of 5 male and 5 female rats received in their diet either 20
per cent. soya bean oil or thermally oxidized material. Highly
modified material reduced the absorption of dietary fat in proportion
to the degree of modification introduced. Modified material delays
absorption of oil from the gastrointestinal tract as measured by
residuals found in the gut. Similar delaying effects have been
demonstrated on the presence of chylomicrons and fat in the intestinal
lymph fluid. The passage of intestinal contents is delayed if
modified oil is administered. Compared with normal oil there is an
early increase followed by only slight decrease in bile flow following
oral administration of modified oil. Intraperitoneal administration
of modified oil increases the diuretic effect of normal soya bean oil
when given intraperitoneally. Study of the liver function after 8
weeks feeding of modified oil showed retention of the
bromosulphophthalein used as indicator compared with normal oil
(Kieckebusch et al., 1962). The in vitro effect of modified oil on
the kinetics of various cellular enzyme systems showed generally no
inhibition of oxidative metabolism (Kieckebusch et al., 1962).
Groups of 20 male and 20 female rats received over 12-16 weeks in
their diet either 20 per cent. untreated soy bean oil of 5 per cent.
variously modified soy bean oil plus 15 per cent. olive oil or 2.5 per
cent. modified oil plus 17.5 per cent. olive oil. Only highly
modified oils showed significant reduction in growth, food intake and
increased mortality. No definite effect was noted at the 2.5 per
cent. level. Motor activity of animals on high doses of highly
modified oil was increased compared with controls. The weights of
major organs were similarly increased for the groups on the more
highly modified oils. Gross and histopathology showed some
pathological changes in the thyroid and kidney of the group receiving
the most highly modified material (Kieckebusch et al., 1962).
Three groups each of 6 rats were treated for 17 weeks with an
esterified product at 0, 0.084 and 0.84 per cent. of their basic diet.
There was no significant effect on weight gain and macroscopic
appearance. Liver and stomach were examined histologically and found
to be normal (Dam, 1952).
Two groups of 9 male rats received for 36 weeks feeds containing
margarine made with an esterified product at 0.3 and 3 per cent.
levels. No controls were used. No weight differences or gross
pathological changes were noted. The histology of kidneys, liver and
small intestine was normal (Aaes-Jorgensen et al., 1954).
Four groups of 4 female pigs received an esterified product for 98
days at dietary levels of 0, 0.4, 2 and 10 per cent. No significant
effects were discovered on growth rate, food consumption, blood
picture, liver and kidney function, serum chemistry, organ weights,
gross and histopathology due to administration of the test compound
(Gyrd-Hansen & Rasmussen, 1968).
A three generation study was performed using an average level of 15
per cent. esterified product in the diet. The P generation (57
females, 15 males) was observed for at least 24 months. Growth,
body-weight gain and appearance were similar to controls (7 male, 7
female) receiving 15 per cent. soy bean oil. Five animals in the test
group developed tumours but none in the control group. There were 16
survivors in the test group and 2 in the control group.
The F1 generation (37 male, 37 female) was also observed for 24
months. Thirty-seven males and females received soya bean oil as
controls. Twenty-four test and 10 control animals survived 2 years.
Two test-animals and 3 controls developed tumours. The F2 generation
(57 female, 27 male) was observed for over 2 years. Seventy males and
females acted as controls on soya bean oil. Four test and 19 control
animals survived 24 months. No animal in the test group but 3 in the
control group developed tumours. Both the F1 and F2 generation
showed no significant differences from controls as regards growth,
body-weight gain, gross and histopathology.
In another experiment an esterified product was fed orally at the rate
of 3 g/day and injected s.c. to 29 rats at 1 ml weekly for 5 months
and 2 ml bi-weekly for a further 3 months. Animals were observed for
27 months. No tumours developed. A control series of 30 rats treated
similarly with oral esterified product and injected s.c. with refined
soya bean oil showed 4 tumours after 24 months observation, none at
the site of injection. The F1 test generation (9 male, 9 female)
received 3 g esterified product orally and 1 ml i.p. for 8 weeks,
followed by a further 1 ml i.p. for 4 months. After 11 months 6
survived without any tumours being noted. The control group (14 male,
4 female) received 1 ml soya bean oil s.c. for 5 months and 2 ml s.c.
for 3 months. During 29 months observation, 1 rat developed a tumour
at the site of injection (Harmsen, 1959; 1960, 1961).
Oxidized and heated fats have been shown to produce toxic symptoms
when fed in large amounts to animals but the effects are not readily
correlated with the physical characteristics or the various polymeric
and peroxide-type of compounds formed during this treatment. The
significance of many of the observed changes is doubtful when
assessing the hazard to man from ingestion of minute amounts of these
compounds. The specific compound studied is produced by thermal
oxidation and steaming. Short-term studies in two species are
available for evaluation, as well as an adequate, combined life-span
and reproduction study in the rat.
Level causing no significant toxicological effect in the rat
Fifteen per cent. (150 000 ppm) in the diet equivalent to 7500 mg/kg
Estimate of acceptable daily intake for man mg/kg body-weight
Temporary acceptance 0-75
Further work required by 1974
Life-span studies in a second species and more adequate work on the
metabolism of these compounds.
Aaes-Jorgensen, E., Punch, J. P., Engel, P. P. & Dam, H. (1954)
Unpublished report supplied by Grindstedvaerket Laboratoriet
Dam, H. (1952) Unpublished report submitted by Grindstedvaerket
Gyrd-Hansen, N. & Rasmussen, F. (1968) Fd. Cosmet. Toxicol. 6, 163
Harmsen, H. (1959) Unpublished report submitted by Grindstedvaerket
Harmsen, H. (1960) Unpublished report submitted by Grindstedvaerket
Harmsen, H. (1961) Unpublished report submitted by Grindstedvaerket
Kieckebusch, K., Jahr, K., Czok, G., Griem, W., Bissler, K.-H.,
Hammar, D. C.-H. & Lang, K. (1962) Fette, Seifen, Anstrichmittel, 64,