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
Geneva
1972
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
MICROBIAL RENNET
Biological data
Source
This enzyme preparation is prepared from the species Mucor pusillus.
Acute toxicity
Animal Route LD50 Reference
(mg/kg body-weight)
mouse oral 4 500 Hara et al., 1963
I.V. 358 Ito et al., 1964
rat oral >5 000 Hara et al., 1963
Twenty per cent. aqueous suspension was non-irritant to rabbit
conjunctiva (Ito et al., 1964).
Short-term studies
Rat
A 10-week study in groups of male and female rats receiving 0, 2, 20
and 1000 ppm in their diet showed a dose-related weight increase
compared with controls. No data are available on organ weights or
haematology but LFTs showed no deleterious effect. Histopathology
showed nothing abnormal (Ito et al., 1964).
A 90-day study was carried out on groups of rats using 0, 10, 1000 and
10 000 ppm in their diet. Some reduction in growth was observed which
was not dose-dependent and statistically not significant. However the
WBC showed increases in lymphocytes with increasing dosage and changes
in monocyte and neutrophil counts which were unrelated to dose. Organ
weights and histopathology were normal (Hara et al., 1963). Another
90-day study was carried out in 3 groups of 10 male and 10 female rats
receiving 0, 1250 or 12 500 ppm in their diet. No abnormalities were
detected as regards behaviour, appearance, food consumption, growth,
haematology, organ weights, gross and histopathology (van Logten &
Kroes, 1968a). A further 90-day study is underway.
Dog
Groups each comprising 4 male and 4 female dogs were fed dietary
levels of 0, 0.5, 1.0 or 2.0 per cent. of fungal rennet (from the same
batch used in the rat study described below) for 2 years. There was
only 1 death unrelated to compound administration. General behaviour
and appearance, body-weight gain and food intake were all comparable
between groups. Blood pressures and heart rates were generally within
normal limits and electrocardiograms revealed no gross abnormalities.
Slight ophthalmological abnormalities observed in 2 control dogs, 3
dogs fed 0.5 per cent. and 5 fed 2.0 per cent. were considered not
related to compound administration. Haemograms, clinical chemistry
and qualitative urinary analyses were comparable for all groups.
There were no gross or histopathological abnormalities that could be
related to feeding fungal rennet (Hollingsworth and Woodard, 1968b).
A 53-week study has been carried out, but details are not available
(Woodard Research Co., 1968).
Long-term studies
Rat
A 53-week study has been carried out, details of which are not
available (Woodard Research Co., 1968a). Another 2-year study is
underway in the Netherlands.
Groups each comprising 20 male and 20 female rats were fed dietary
levels of 0, 0.5, 1.0 or 2.0 per cent. of fungal rennet (identified as
microbial rennet from Meito Sangyo Co. Ltd Lot No. R2G7801). General
appearance and behaviour were comparable in test and control groups.
Incidence of mortality appeared unrelated to feeding the test
compound. Body-weight gain and food intake were normal between the
groups. Periodic haemograms including haemoglobin, microhaematocrit,
coagulation time and general clinical chemistry determination as well
as absolute and relative organ weights and gross and histopathology
revealed no abnormalities related to feeding the rennet (Hollingsworth
and Woodard,, 1968a).
Special studies
Six groups of 18-20-day old ducklings were given 0, 2 mg, 10 mg, 50 mg
and 250 mg material by gavage as well as a positive control group
receiving 12.5 mg aflatoxin. Deaths occurred only in the aflatoxin and
250 mg groups. There was some weight loss throughout the test in the
250 mg group and less so at lower test levels. Liver weight was
increased on the 8th day at all levels. Histologically no lesions
were seen which pointed to the presence of aflatoxin (van Logten &
Kroes, 1968).
A multigeneration study in rats over 3 generations is underway.
Comments
This preparation has been tested by short-term studies in rats and a
short-term study in dogs but none exceeded 53 weeks. The highest
level tested was 2.0 per cent. in the diet of both species. None of
these studies are of sufficient length to reveal possible chronic
effects although the levels used would have revealed any deleterious
effects due to mycotoxins. Moreover, this possibility was tested by a
study in ducklings. A further 2-year study in rats is under way.
EVALUATION
Temporarily not limited.*
Further work required by 1974
Results of 2-year study in rats.
REFERENCES
Hara, S., Shibuya, T., Yakazu, K., Horibe, M., & Sato, S., (1963)
Report of Dept. Pharmac. Tokyo Med. Coll. submitted to WHO
Hollingsworth, R. C. & Woodard, G. (1968a) Unpublished report dated 7
November 1968 from Woodard Research Corporation submitted by Noury &
van der Lande N. V.
Hollingsworth, R. C. & Woodard, G. (1968b) Unpublished report dated 6
November 1968 from Woodard Research Corporation submitted by Noury &
van der Lande N. V.
Ito, H., Kond, H., & Tokunaga, Y., (1964) Fd. Hyg. Mag. Japan, 5(1), 1
Nakamura K., (1966) Report submitted by Nat. Inst. Hygienic Sciences,
Tokyo to WHO
van Logten, M. J., & Kroes, R., (1968) Report 82/68 Tox submitted by
Ryksinstituut Utrecht
van Logten, M. J., & Kroes, R., (1968a) Report 88/68 Tox submitted by
Ryksinstituut Utrecht
Woodard Research Co. (1968) Unpublished Report dated 6.11.1968
submitted to WHO
Woodard Research Co. (1968a) Unpublished Report dated 7.11.1968
submitted to WHO
*Except for good manufacturing practice.
See Also:
Toxicological Abbreviations
Microbial rennet (WHO Food Additives Series 1)
Microbial rennet (WHO Food Additives Series 1)