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    INTERNATIONAL PROGRAMME ON CHEMICAL SAFETY

    WORLD HEALTH ORGANIZATION



    TOXICOLOGICAL EVALUATION OF SOME
    FOOD COLOURS, EMULSIFIERS, STABILIZERS,
    ANTI-CAKING AGENTS AND CERTAIN
    OTHER SUBSTANCES



    FAO Nutrition Meetings Report Series 
    No. 46A WHO/FOOD ADD/70.36




    The content of this document is the result of the deliberations of the
    Joint FAO/WHO Expert Committee on Food Additives which met in Rome,
    27 May - 4 June 19691





    Food and Agriculture Organization of the United Nations

    World Health Organization



                   
    1 Thirteenth report of the Joint FAO/WHO Expert Committee on Food
    Additives, FAO Nutrition Meetings Report Series, in press;
    Wld Hlth Org. techn.  Rep. Ser., in press.


    POLYGLYCEROL ESTERS OF INTERESTERIFIED RICINOLEIC ACID

    Biological Data

    Biochemical aspects

    Twelve rats fed for 10 weeks a diet containing 1 per cent. or 10 per
    cent. groundnut oil or 1 per cent groundnut oil + 9 per cent. of the
    substance were divided into two groups. One was fed a fat-free diet
    for six days, the other group was fed test or control diets. No
    acetone or acetoacetic acid was detected in any urine, pointing to
    absence of interference with fat metabolism.

    Three groups of 10 male and six female rats fed 45 weeks on diets
    containing either 1 per cent. or 10 per cent. groundnut oil or 1 per
    cent. groundnut oil + 9 per cent. of the substance ware given three
    days later an additional 9 per cent. groundnut oil for seven days. No
    differences between tests and control groups were detected regarding
    oil absorption, protein digestibility or diet digestibility.

    Three groups of 10 male and six female rats were fed diets containing
    9 per cent. or 18 per cent. groundnut oil or 9 per cent. groundnut oil
    + 9 per cent. of the substance. Clearing of lipaemic plasma by heparin
    was similar in all three groups and plasma turbidity was not
    increased. Chylomicron concentration in rats before and after feeding
    the substance in their diet showed absence of alimentary
    chylomicronaemia. Absorption does therefore not occur into intestinal
    lymphatics although a specific test to confirm absorption into portal
    capillaries was not done.

    Three groups of rats were fed four weeks on diets containing 1 per
    cent. or 10 per cent. groundnut oil or 1 per cent. groundnut oil + 9
    per cent. of the substance. Digestibility of the substance was 98 per
    cent.

    Another three groups of two male and two female rats were fed eleven
    weeks on diets containing either 1 per cent. or 10 per cent. groundnut
    oil or 1 per cent. groundnut oil + 9 per cent. of the substance. No
    adverse influence was noted on fat storage, body composition and
    vitamin A content of the liver; the substance was not stored
    significantly in the body fat.

    The in vitro action of lipase on groundnut oil was not affected by
    the presence of the substance (Unilever Research Laboratories, 1966).

    Acute toxicity

                                                                     

    Animal    Route         LD50            Reference
                        mg/kg body-weight
                                                                     

    rat       oral        > 18 500          Unilever Research
                                            Labs, 1966

    rat       i.p.        > 12 000                "
                                                                     

    No mortality could be obtained, though animals received up to 47.5
    g/kg daily for 5 days (Unilever Research Laboratories, 1966).

    Short-term studies

    Rat. Three groups of eight male and eight female rats were fed a
    diet containing either 1 per cent. or 10 per cent. groundnut oil or 1
    per cent. groundnut oil + 9 per cent. of no substance for 30 weeks. No
    adverse effects on growth, food intake and faecal appearance were
    noted. During this test all three groups were restricted to 5 g fat
    free diet for 17 days and then realimented. No difference in weight
    gain was noticeable, nor in carcass and liver fat amounts or
    composition. The test group on the substance had a shorter
    Bromosulphophthalein retention time. The specific gravity of the urine
    was comparable in all groups. Organ weights of liver, kidney, adrenal,
    testes and spleen were similar in all groups. In view of the special
    design of the study, the statement on absence of significant changes
    in liver weight applies only to three groups of selected animals not
    participating in the food restriction experiment, i.e. two female and
    four male rats on 1 per cent. groundnut oil, four female and three
    male rats on 10 per cent. groundnut oil and two female and five male
    rats on 1 per cent. groundnut oil + 9 per cent. of the substance.
    Despite the small numbers the test may be taken to indicate that 9 per
    cent. of the substance was not causing liver enlargement when fed for
    30 weeks.

    In another study three groups of 10 male and six female rats were kept
    on diets containing either 1 per cent. or 10 per cent. groundnut oil
    or 1 per cent. groundnut oil + 9 per cent. of the substance for 45
    weeks. No adverse effect on growth or food intake was noted, The
    groups were comparable as regards Bromosulphophthalein retention,
    specific gravity of urine, haematological indices and red cell
    fragility. Organ weights of kidney, adrenal, testes and spleen were
    similar in all groups but the liver weights of rats fed of the
    substance (9 per cent. of the diet) were significantly higher than the
    controls. Similar effects were seen in rats fed castor oil. This was
    not due to an increased blood volume of the liver (Unilever Research
    Laboratories, 1966).

    Man. Nineteen male and female volunteers, aged 19-24 years, took 5 g
    of the substance per day for one week and 10 g per day for another
    week using constant dietary fat intake. Serum protein, albumin and
    globulin levels were unaffected. Thymol turbidity, serum bililubin,
    SGBT, serum cholesterol and serum cholinesterase were normal (Unilever
    Research Laboratories, 1963).

    Long-term studies

    Mouse. Four groups of 25 male and 25 female mice were fed a diet
    with 5 per cent. groundnut oil or the substance with no additional fat
    for 80 weeks. A positive control group received 0.5 mg, 9,
    10-dimethyl-1,2-benzanthracene once per week for 16 weeks. After 50
    weeks there was no difference in growth between test and groundnut oil
    groups but the basic diet and positive control group showed reduced
    growth. Survival was comparable for the three groups, also the
    haematological indices. Liver and kidney of the test group were
    significantly heavier than the controls. No specific histological
    lesion was seen. Carcass fat contained no polyglycerol but only a
    small amount (0.1 per cent.) of ricinoleic acid. Histopathology of all
    other major tissues showed no lesions specifically associated with the
    substance nor was there any significant difference in tumour
    incidence. The strain was sensitive to the carcinogen used in the
    positive control.

    Groups of 20 male and 20 female mice were given repeated s.c.
    injections of 0.5 ml of the substance weekly for five weeks with milk
    or groundnut oil as controls. After 80 weeks no adverse effects were
    noted on growth, survival, haematology, organ weights and
    histopathological findings. Tumour incidence was similar in test and
    control groups.

    Seven groups of 20 male and 20 female mice were painted daily with 20
    mg of the substance for 60 weeks with or without a single preceding
    application of 0.25 mg dimethylbenzanthracene and a positive control
    group was included. After 80 weeks no adverse effects were noted on
    growth, survival or tumour incidence in tests and controls involving
    the substance. There was no promoting or cutaneous carcinogenic effect
    (Unilever Research Laboratories, 1966).

    Rat. A three-generation test was carried out in two groups of 19 and
    28 rats on diets containing 0 per cent. or 1.5 per cent. of the
    substance. No significant differences were seen between the two groups
    as regards fertility, pup weight, pup survival, litter number, etc.
    Each animal was observed for over one year. No consistent
    abnormalities or histopathological changes were seen in the third
    generation.

    In another experiment two groups of 30 male and 60 female rats each
    were fed diets containing five per cent. of the substance or five per
    cent. groundnut oil for 104 weeks. No significant adverse effects were
    seen on growth, food consumption, liver function tests at weeks 89 and
    103, specific gravity of the urine at weeks 89 and 103, or survival.
    The kidneys of males and females at five per cent. of the substance
    and the livers of females at five per cent. of the substance were
    significantly enlarged. There was no polyglycrol or ricinoleic acid
    accumulation in carcass fat. Histopathology of all organs showed no
    abnormalities related to the substance administration nor was there a
    rise in tumour incidence in the test group. The strain was sensitive
    to 20-methylcholanthrene.

    Thirty male and 30 female rats were injected s.c. with 0.5 ml twice
    weekly for 13 weeks at different sites, milk and groundnut oil were
    controls. After two years there were no adverse effects on growth,
    survival, haematological indices, organ weights or histopathology of
    all tissues. No increase in tumour incidence was found but most rats
    treated with the substance developed persistent nodules at injection
    sites.

    Six groups of 10 male and 10 female rats were painted with 50 mg of
    the substance cutaneously daily for 60 weeks with or without a single
    preceding application of 0.25 mg benzanthracene and observed for two
    years. A positive control group was included. No deleterious effects
    were noted on growth and survival. No skin tumours were seen in the
    test group. The strain was sensitive to the carcinogen used as
    positive control (Unilever Research Laboratories, 1966).

    Comments

    The metabolic fate of this material has been studied by a number of
    indirect measurements. In-vivo studies, using labelled material, are
    not available. The long-term studies in rats and mice did not show
    carcinogenic potential. The enlargement of liver and kidneys observed
    in long-term tests was not accompanied by any lesions detectable by
    histopathology. Only the rat study shows a no-effect level for liver
    enlargement. No satisfactory explanation of the observed organ
    enlargements has been offered.

    EVALUATION

    Level causing no toxicological effect in the rat

    1.5 per cent. (= 15 000 ppm) in the diet equivalent to 750 mg/kg
    bodyweight per day.

    Estimate of acceptable daily intake for man

                                            mg/kg body-weight

    Temporary acceptance                         0 - 3 75

    Further work required by June 1972

    In-vivo metabolic studio in several species to determine more
    directly the fate of the major polymers in the material. A 90-day
    study in a non-rodent mammalian species to establish a no-effect level
    for organ enlargement. Investigation of the causes of the observed
    hepatic and renal enlargements.

    REFERENCES

    Unilever Research Laboratories (1966) Unpublished report submitted to
    WHO
    


    See Also:
       Toxicological Abbreviations
       Polyglycerol esters of interesterified ricinoleic acid (WHO Food Additives Series 5)
       POLYGLYCEROL ESTERS OF INTERESTERIFIED RICINOLEIC ACID (JECFA Evaluation)