Toxicological evaluation of some food
    additives including anticaking agents,
    antimicrobials, antioxidants, emulsifiers
    and thickening agents


    The evaluations contained in this publication
    were prepared by the Joint FAO/WHO Expert
    Committee on Food Additives which met in Geneva,
    25 June - 4 July 19731

    World Health Organization


    1    Seventeenth Report of the Joint FAO/WHO Expert Committee on
    Food Additives, Wld Hlth Org. techn. Rep. Ser., 1974, No. 539;
    FAO Nutrition Meetings Report Series, 1974, No. 53.



         These compounds have been evaluated for acceptable daily intake
    by the Joint FAO/WHO Expert Committee on Food Additives (see Annex 1,
    Refs Nos 6, 8 and 30) in 1961, 1964 and 1972.

         Since the previous evaluation, additional data have become
    available and are summarized and discussed in the following monograph.
    The previously published monograph has been expanded and is reproduced
    in its entirety below.



         The available evidence indicates that the esters are hydrolysed
    in the body. Most of the gallic acid is converted into 4-O-methyl
    gallic acid. Free gallic acid or a conjugated derivative of 4-O-methyl
    gallic acid is excreted in the urine. Conjugation of the 4-O-methyl
    gallic acid with glucuronic acid was demonstrated (Booth et al.,

         Rats fed a low-methionine, low-choline diet containing 1% gallic
    acid developed fatty livers which could be prevented by added choline
    or methionine (Booth et al., 1961).

         The detailed metabolic pathways for propyl gallate have been
    described (Dacre, 1960). There is no evidence to suggest that other
    esters of gallic acid differ greatly in their metabolism from the
    pattern described for propyl gallate.


    Special studies on dermatological effects

         Gallates have been shown to cause contact dermatitis in bakers
    and other workers handling gallates. Patch tests with lauryl gallate
    at 0.2% showed a weak positive response in one sensitized individual.
    Other individuals have suffered recurring episodes of dermatitis
    presumably caused by gallates in food products (Brun, 1970).

         Repeated insult patch test with 0.1% n-octyl gallate solution
    showed an overall incidence of reaction in 13/445 or 2.9% individuals
    (Biotest Lab., 1971c). Oral mucosa irritation/sensitization tests were
    conducted with beer containing 20 ppm (0.002%) n-octyl gallate and
    showed that the incidence and severity of erythema were greater with

    beer containing n-octyl gallate, than with untreated beer. Oedema was
    also greater with treated beer. Individuals that had previously shown
    reactions indicative of sensitization in the patch test were more
    susceptible to irritation (incidence and severity) than other
    individuals) Biotest Lab., 1971a, b, c).

    Acute toxicity

    Animal        Route   LD50 mg/kg bw    References

    Rat           oral    6 500            van Sluis, 1951

    Rat           i.p.    100-120          van Esch & van Genderen, 1954

    Animal        Route   LD50 mg/kg bw    References

    Rat           oral    4 700            Van Sluts, 1951

    Rat           i.p.    60-80            van Esch & van Genderen, 1954

    Rat (male)    oral    2 710            Brun, 1970

    Rat (female)  oral    1 960            Brun, 1970

    Rat (male)    oral    2 710            Brun, 1970

    Rat (female)  oral    2 330            Brun, 1970

    Animal        Route   LD50 mg/kg bw    References

    Rat           oral    3 800            Orten et al., 1948

    Rat           oral    3 600            Lehman et al., 1951

    Rat           i.p.    380              Orten et al., 1948

    Mouse         oral    2 000-3 500      Lehman et al., 1951

    Short-term studies


         Levels of propyl gallate of 1.2% and 2.3% in the diet caused 
    interference with weight gain, the bitter taste of the gallate 
    apparently making the diet unpalatable. The higher dose level caused 
    some deaths (about 40%) during the first month; the survivors 
    continued to eat the diet for 10 to 16 months and showed retarded 
    growth, but no pathological lesions. The animals that died exhibited 
    renal damage (Orten et al., 1948).

         Weanling rats were given diets containing 2.5% and 5% dodecyl
    gallate. All animals fed the smaller quantity were dead within
    10 days, and all animals fed the larger quantity died within seven
    days (Allen & De Eds, 1951).

         Rats fed for 70 days on a diet containing 7% fat and 0.2% dodecyl
    gallate showed no effect on body weight (Tollenaar, 1954).

         Weanling rats were fed diets which contained 20% lard and 0, 0.1,
    0.2, 0.3, 0.4 and 0.5% propyl gallate for six weeks. There was no
    effect on body weight, liver weight, liver weight to body weight
    ratio, left adrenal weight, total liver lipid, composition of liver
    polyunsaturated fatty acids, liver cholesterol, adrenal cholesterol or
    serum sodium (Johnson & Hewgill, 1961).

         Propyl gallate was added to the dietary fat of weanling rats at
    levels of 0.02% in the fat for 13 weeks. The fat content of the diet
    accounted for 30% of its calorific value. There was a very slight
    inhibition of growth. The same rats were then placed on a partial
    starvation diet and kept until they died. The survival time of the
    animals which had received the propyl gallate was considerably reduced
    and the reduction in their total body protein was greater (Buckman,

         Groups each of 20 rats (equally divided by sex) were maintained
    on diets containing 0, 1000, 2500 and 5000 ppm (0%, 0.1%, 0.25% and
    0.5%) of octyl gallate for 13 weeks. All groups showed normal weight
    gains, food consumption. Haematologic and blood chemistry and urine
    analyses were normal. A complete gross and histopathologic examination
    showed no compound-related effects (Hazleton Lab. Inc., 1969a).


         Propyl gallate fed to guinea-pigs in groups of 20 at a level of
    0.0117% in the diet for 14 months caused no detectable ill effects
    (Orten et al., 1948).


         Groups each of eight dogs (equally divided by sex) were fed diets
    containing 0, 0.1, 0.3%n-octyl gallate for 90 days, and 1.0% for four
    weeks; the 1.0% level has been then reduced to 0.6% for the rest of
    the study. All groups showed normal weight gains, food consumption
    (except 1.0% group). Haematologic and blood chemistry and urine
    analyses were normal. A complete gross and histopathologic examination
    showed no compound-related effects (Industrial Biotest Lab., 1970). In
    another study groups each of eight dogs (equally divided by sex) were
    maintained on diets containing 0, 0.1, 0.25 or 0.5% octyl gallate for
    13 weeks. All animals showed normal food consumption and weight gain.
    Haematologic and urine analyses were similar for test and control
    animals. SGOT was slightly elevated in the 0.5% group. Gross and
    histopathologic examination of tissues and organs showed no compound-
    related effects (Hazleton Lab. Inc., 1969b).

         A level of 0.0117% of propyl gallate in the diet was well
    tolerated by a group of seven dogs over a period of 14 months (Orten
    et al., 1948).


         Diets containing 0.2% of propyl, octyl or dodecyl gallate were
    fed to pigs without demonstrable ill effect; no anaemia was observed
    (van Esch & van Genderen, 1954).

    Long-term studies

    Mouse and rat

         A level of 5% propyl gallate in the diet in a two-year chronic
    toxicity test on rats and mice gave rise to patchy hyperplasia in the
    proventiculus. At a level of 1% no difference was noted between test
    and control animals (Lehman et al., 1951).


         Rats in groups of 10 males and 10 females were fed for two years
    on diets containing 0%, 0.00117%, 0.0117%, 0.117%, 1.17% and 2.34% of
    propyl gallate. The groups receiving 1.17% and 2.34% of propyl gallate
    showed stunted growth and evidence of renal damage. In the other
    groups, there was no detectable effect on haemoglobin, erythrocyte or
    leucocyte levels in the blood, or on the histological appearance of
    the organs examined (Orten et al., 1948).

         Propyl, octyl and dodecyl gallate were fed to rats at
    concentrations of 0.035%, 0.2% and 0.5% in the diet. Growth was
    affected only at the 0.5% level of dodecyl gallate; there was
    significant retardation, particularly in the second generation. At

    this level of dodecyl gallate, some litters were lost in the second
    generation because they were not sufficiently fed by the mothers. A
    slight hypochromic anaemia was noticed in the groups on diets
    containing 0.2% octyl and dodecyl gallate. No abnormalities were
    observed in the organs or tissues of the rats at autopsy (Van Esch &
    van Genderen, 1954).

         Young rats in groups of 12 males and 12 females were fed diets
    containing 7% fat and 0.2% octyl or dodecyl gallate. There was no
    significant difference between test and control animals over three
    generations (van Sluis, 1951).

         n-Octyl gallate was fed in the diet to groups of eight male and
    16 female rats for two successive generations at levels of 0, 0.1 or
    0.3% (and 0.6% for one generation). Rats were mated to produce two
    litters per generation with next generation selected from weanlings of
    the second litter. A dietary level of 0.1% (1000 ppm) had no effect on
    reproduction performance or the offspring. At 0.3 and 0.6% dietary
    octyl gallate, there was no significant effect on the rat fetuses
    during pregnancy, but a marked effect was observed on survival through
    weaning. In the case of the 0.6% group, return to normal diet for six
    weeks, prior to a third breeding, did not result in increased survival
    of offspring through weaning (Industrial Biotest Lab., 1970a).

         Groups each of 10 male and 20 female rats were maintained on
    diets containing 0, 1000 or 5000 ppm (0%, 0.1% or 0.5%) of octyl
    gallate. The animals were bred twice for the first generation, and
    three times for the second generation. At the time of weaning of the
    B1B litters the 5000 ppm (0.5%) level was replaced by a 2500 ppm
    (0.25%) level for the second generation. In the case of the second
    generation approximately 24 hours after birth, selected litters were
    redistributed to female parents so that control females nursed pups
    from test animals, and test animals nursed pups from control and other
    test groups. One half P2 females bred for the third time (F2c) were
    examined by caesarean section at time of delivery and the number of
    implantation sites, corpora lutea and fetuses determined. One half of
    the pups from each litter were examined for skeletal abnormalities,
    and the other half for visceral abnormalities. The other parameters
    measured in this study were appearance, behaviour, growth of pups
    during the nursing and weaning process, fertility index, gestation
    index, live birth index, weaning survival index. Autopsies were
    carried out on F2b weanling pups, (control suckled by control, each
    groups suckled by respective group parent), as well as a microscopic
    examination of pituitary, thyroid, liver, spleen, kidneys, adrenals,
    stomach, pancreas, small intestine, large intestine and any unusual
    lesion of five males and five females, high level and control groups.
    Weaning survival index, and body weight at weaning was considerably
    reduced in the 5000 ppm (0.5%) group of the F1A and F1B generation.
    Reduction of indices was still apparent in the F2A and F2B
    generations, when the dietary level was reduced to 2500 ppm (0.25%).

    At the 1000 ppm (0.1%), the indices were similar to control.
    Redistribution of F2B pups to females of control groups, resulted in
    similar growth of all pups to weaning. Allowing pups from high level
    group to be nursed by control dams resulted in a marked increase in
    survival indices, whereas when control pups were nursed by high level
    dams, there was a marked decrease in survival indices. Examination of
    P2 parents following third breeding indicated a dose-dependent
    reduction in implantation sites, as well as a reduction in number of
    corpora lutea. Fertility index of high level P2 females was depressed
    at the F2C stage. Skeletal evaluation of F2C litters showed
    incomplete skull ossification in some pups in the test groups, but
    this was not considered remarkable for the size of the fetuses.
    Necropsy of the pups indicated a higher incidence of gross kidney
    alterations than that observed in controls. Ne compound-related
    histology was reported (Hazleton Lab. Inc., 1970b).


         Reproduction studies in the rat indicate that the mortality of
    suckled young produced by the higher levels was due to a factor
    entering the mother's milk and was therefore a perinatal rather than a
    prenatal effect. It has not been established whether the reported
    decrease in implantation sites and number of corpora lutea that
    occurred at the third breeding of the F2 parents was due to an effect
    on viability of ova or sperm. The "no-effect" level in rats, based on
    reproduction studies, is 1000 ppm (0.1%) of the diet, i.e. 50 mg/kg bw
    per day. The n-octyl gallate can cause reactions indicative of
    allergic sensitization in the buccal mucosa of individuals previously
    sensitized by exposure to n-octyl gallate in beer or other beverages
    consumed in large amounts, therefore, its use in such beverages is not

         Those occupationally exposed to gallate esters should be made
    aware of the sensitizing potential of these substances.

         With one exception, in long-term studies in rats gallates
    caused no demonstrable ill effects when fed at a level of 0.2% in
    the diet; in one investigation, however, this level resulted in
    hypochromic anaemia. It seems likely that this may have been due to
    interference with iron absorption, but the cause was not established.
    Haematological effects were carefully examined in a number of other
    investigations and no abnormalities were seen. It seems possible,
    therefore, that this effect was related to the particular
    circumstances of one study and can be properly disregarded in arriving
    at the level that causes no significant effects in the rat. However,
    the no-effect level in the reproduction study is used for this


    Estimate of acceptable daily intake for man

         0-0.2* mg/kg bw.**


         Required by 1976. Studies on the effect on reproduction of
    mixtures of BHA, BHT and propyl gallate.


    Allen, C. S. & De Eds, F. D. (1951) J. Amer. Oil Chem. Soc., 28, 304

    Booth, A. N. et al. (1959) J. biol. Chem., 234, 3014

    Booth, A. N., Robbins, D. J. & De Eds, F. (1961) J. Nutr., 75, 104

    Brun, R. (1970) Dermatologica, 140, 390

    Buckman, N. D. (1962) Vop. Pitan., 21, 68

    Dacre, J. C. (1960) J. N.Z. Inst. Chem., 24, 161

    Esch, G. J. van & Genderen, H. van (1954) Netherlands Institute of
         Public Health, Report No. 481

    Hazleton Lab. Inc. (1969a) Unpublished report. Project No. 458-117

    Hazleton Lab. Inc. (1969b) Unpublished report. Project No. 458-115

    Hazleton Lab. Inc. (1970a) Unpublished report. Project No. 458-116

    Industrial Biotest Lab. (1970b) Unpublished report. IBT No. C8472

    Industrial Biotest Lab. (1971a) Unpublished report. IBT No. P8473

    Industrial Biotest Lab. (1971b) Unpublished report. IBT 8473

    Industrial Biotest Lab. (1971c) Unpublished report. IBT No. F9655


    *    As sum of dodecyl, octyl, and propyl gallate n-octyl gallate
    should not be used in beverages.

    **   Temporary.

    Johnson, A. R. & Hewgill, F. R. (1961) Aust. J. exp. Biol. med. Sci.,
         39, 353

    Lehman, A. J. et al. (1951) Advanc. Food Res., 3, 197

    Orten, J. M., Kuyper, A. C. & Smith, A. H. (1948) Food Techn., 2, 308

    Sluis, K. J. H. van (1951) Food Manuf., 26, 99

    Tollenaar, F. D. (1954) Fette u. Seifen, 56, 41

    See Also:
       Toxicological Abbreviations