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    FAST GREEN FCF

    Explanation

         Fast Green FCF was evaluated by JECFA in 1969 (FAO, 1970) and was
    allocated an ADI of 0-12.5 mg/kg bw. Since the previous review, new
    data have become available and are included in this monograph.

    BIOLOGICAL DATA

    BIOCHEMICAL ASPECTS

         Rats and dogs were given orally 200 mg of the colour. In the rats
    the urine and faeces were collected for 36 hours. In the dogs, a bile
    fistula was made for bile analysis. Almost all the administered colour
    was excreted unchanged in the faeces of rats. No colour was found in
    the urine. In the bile of the dogs, the amount of colour never
    exceeded 5% of the given dose. After feeding, the colour was found in
    the bile of rats and rabbits, but not in their urine. It was concluded
    that the quantity found in the bile provides a reasonable estimate of
    the amount absorbed from the gastrointestinal tract (Hess & Fitzhugh,
    1953, 1954 and 1955). Following i.v. injection in rats, over 90% of
    the colour was excreted in the bile within four hours (Iga et al.,
    1971).

         Fast Green FCF was found to have a high binding affinity for
    plasma protein (Gangolli et al., 1967 and 1972; Iga et al., 1971).

    TOXICOLOGICAL STUDIES

    Special studies on carcinogenicity (see also Long-term studies)

    Rat

         Eighteen weanling Osborne-Mendel rats of both sexes received
    weekly s.c. injections of approximately 30 mg (1 ml of a 3% aqueous
    solution) of the colour for 94-99 weeks. Subcutaneous fibrosarcomas
    appeared at the site of injection in 15 animals (Nelson & Hagan, 1953;
    Hansen et al., 1966).

         Two groups of 16 female rats (control group of 10 rats) were
    given subcutaneous injections of 0.5 ml of a 3% and 6% solution (the
    rats received with each injection respectively 15 and 30 mg). The used
    colour was certified as 92% pure and was supplied as the disodium
    sulfonate salt. The 10 control rats were given distilled water
    injections. At first, injections of 6% were given three times a week;
    after 17 weeks it became necessary to reduce the dose to 3%.
    Thereafter, both groups were given injections of 3% twice weekly for
    nine weeks. The rest of the time, 22 weeks, both groups were injected
    usually once a week, occasionally two injections were tolerated.

    Growth inhibition was found. Thirteen out of 16 animals receiving 6%
    of the colour had fibrosarcomas. The animals given 3% showed also
    fibrosarcomas (10 out of 12). The controls did not show neoplastic
    tissue at the site of injection (Hesselbach & O'Gara, 1960).

         Subcutaneous injection of 1 ml of a 0.8% solution twice weekly
    produced histological changes suggestive of subsequent sarcoma
    formation unassociated with chemical carcinogenic potential (Grasso &
    Golberg, 1966).

         No tumours were produced in 11 hamsters injected with 1 mg of the
    dye in 0.1 ml water (Price et al., 1978).

    Special studies on mutagenicity

         Fast Green FCF was non-mutagenic in the Salmonella/microsome
    assay (Brown et al., 1978) and negative results were also obtained in
    bacterial DNA repair tests (Kada et al., 1972; Rosenkranz & Leifer,
    1980). The colour was inactive in a gene conversion assay in diploid
    yeast (Sankaranarayanan & Murthy, 1979).

         In one of two experiments, the colour-induced cell transformation
    in cultured Fisher rat embryo cells at a concentration of 1 g/ml
    (Price et al., 1978) and chromosome damage was reported in an in
    vitro test using Chinese hamster ovary cells (Au & Hsu, 1979).

    Acute toxicity
                                                             

                              LD50
    Animal      Route     per mg/kg bw       Reference
                                                             

    Rat         Oral         > 2000      Lu & Lavallee, 1964
                                                             

    Dog

         The colour given in single 200 mg doses to dogs did not produce
    catharsis (Radomski et al., 1956).

    Short-term studies

    Rat

         Groups of 50 weanling Osborne-Mendel rats, evenly divided by sex,
    were fed diets containing 0, 0.5, 1.0, 2.0 or 5.0% colour for two
    years. No effects on growth or mortality were observed. Microscopic
    examination revealed no lesions that were attributed to the feeding of
    the colour (Hansen et al., 1966).

         The colour was fed at a dietary level of 4.0% to five male and
    five female rats for periods from 18 to 20 months. This procedure
    resulted in gross staining of the forestomach, glandular stomach,
    small intestine and colon. Granular deposits were noted in the
    stomach. No tumours were observed (Willheim & Ivy, 1953).

    Dog

         Four beagles per group, equally divided by sex, were fed at 0,
    1.0 and 2.0% for two years. Histopathology attributable to the colour
    was limited to green blobs of pigment in the renal cortical tubular
    epithelial cytoplasm of a male dog on a high dose level; a female dog
    on a high dose level showed slight interstitial nephritis and slight
    bone marrow hyperplasia (Hansen et al., 1966).

    Long-term studies

    Mouse

         Groups of 50 male and 50 female C3HeB/FeJ mice were fed diets
    containing 1.0 or 2.0% colour for two years and 100 mice of each sex
    served as controls. After 78 weeks, 56 controls and 27 animals in the
    1.0%, 17 animals in the 2.0%, treatment groups still survived.
    Microscopic examination revealed no lesions that were attributed to
    feeding of the colour (Hansen et al., 1966).

    Rat

         Groups of 50 weanling Osborne-Mendel rats, evenly divided by
    sex, were fed diets containing 0, 0.5, 1.0, 2.0 or 5.0% colour for
    two years. No effects on growth or mortality were observed.      
    Microscopic examination revealed no lesions that were attributed to
    the feeding of the colour (Hansen et al., 1966).

         The colour was fed at a dietary level of 4.0% to five male and
    five female rats for periods from 18 to 20 months. This procedure
    resulted in gross staining of the forestomach, glandular stomach,
    small intestine and colon. Granular deposits were noted in the
    stomach. No tumours were observed (Willheim & Ivy, 1953).

    OBSERVATIONS IN MAN

         No data available.

    Comments

         The production of a high percentage of local sarcomata at the
    site of subcutaneous injection in rats is considered to be related to
    the physicochemical properties of the colour and the special
    conditions of the experiments, and does not constitute evidence of
    carcinogenicity by the oral route. However, in reviewing the

    carcinogenicity of Fast Green FCF, an IARC Working Group emphasized
    the inadequacy of the long-term feeding studies in mice and rats
    (IARC, 1978); in the rat study, only 12 animals fed 5% of the dye were
    subjected to detailed microscopical examination.

         Biochemical studies have shown that the colour is poorly absorbed
    and is almost completely excreted in faeces after parenteral
    administration.

         No data were available on reproductive toxicology and teratology.

    EVALUATION

    Level causing no toxicological effect

    Rat: 5% in the diet equivalent to 2500 mg/kg bw.

    Estimate of temporary acceptable daily intake for man

    0-12.5 mg/kg bw.

    FURTHER WORK OR INFORMATION

    Required by 1985

         Adequate long-term feeding studies in the rat and a
    multigeneration reproduction/teratology study.

    REFERENCES

    Au, W. & Hsu, T. C. (1979) Studies on clastogenic effects of biologic
         stains and dyes, Environmental Mutagenesis, 1, 27

    Brown, J.P., Roehm, G. W. & Brown, R. J. (1978) Mutagenicity testing
         of certified food colours and related azo, xanthene and
         triphenylmethane dyes with the Salmonella/microtome system,
         Mutation Res., 56, 249-271

    FAO (1970) Toxicological evaluation of some food colours, emulsifiers,
         stabilizers, anti-caking agents and certain other substances,
         FAO Nutr. Mtgs. Rep. Series No. 46A

    Gangolli, S. D., Grasso, P. & Golberg, L. (1967) Physical factors
         determining the early local tissue reactions produced by food
         colourings and other compounds injected subcutaneously, Fd.
         Cosmet. Toxicol., 5, 601-621

    Gangolli, S. D., et al, (1972) Protein binding by food colourings in
         relation to the production of subcutaneous sarcoma, Fd. Cosmet.
         Toxicol., 10, 449-462

    Grasso, P. & Golberg, L. (1966b) Subcutaneous sarcoma as an index of
         carcinogenic potency, Fd. Cosmet. Toxicol., 4, 297

    Hansen, W. H. et al. (1966) Chronic toxicity of three food colourings:
         guinea green B, light green SF yellowish, and fast green FCF in
         rats, dogs and mice, Fd. Cosmet. Toxicol., 4, 389-410

    Hess, S. M. & Fitzhugh, O. G. (1953) Metabolism of coal-tar dyes. I.
         Triphenylmethane dyes (Abstract No. 1090), Fed. Proc., 12,
         330-331

    Hess, S. M. & Fitzhugh, O. G. (1954) Metabolism of coal-tar colours.
         II. Bile studies (Abstract No. 1201), Fed. Proc., 13, 365

    Hess, S. M. & Fitzhugh, O. G. (1955) Absorption and excretion of
         certain triphenylmethane colours in rats and dogs,
         J. Pharmacol. Exp. Ther., 114, 38-42

    Hesselbach, M. L & O'Gara, R. W. (1960) Fast green and light green
         induced tumours: induction, morphology and effect on host,
         J. nat. Cancer Inst., 24, 769-793

    IARC (1978) IARC Monographs on the Evaluation of the Carcinogenesis
         Risk of Chemicals to Man: some aromatic amines and related nitro
         compounds - hair dyes, colouring agents and miscellaneous
         industrial chemicals. Vol. 16, IARC, Lyon

    Iga, T., Awazu, S. & Nogami, H. (1971) Pharmacokinetic study of
         biliary excretion. II. Comparison of excretion behaviour in
         triphenylmethane dyes, Chem. pharm. Bull., 19, 273-281

    Kada, T., Tutikawa, K. & Sadaie, Y. (1972) In vitro and
         host-mediated 'rec-Assay' procedures for screening chemical
         mutagens and phloxine, a mutagenic red dye detected, Mutation
         Res.,16, 165-174

    Lu, F. C. & Lavallee, A. (1964) The acute toxicity of some synthetic
         colours used in drugs and foods, Canad. pharm. J., 97, 30

    Nelson, A. A. & Hagan, E. C. (1953) Production of fibrosarcomas in
         rats at site of subcutaneous injection of various food dyes
         (Abstract No.1307), Fed. Proc., 12, 397-398

    Price, P. J. et al. (1978) In vitro and in vivo indications of the
         carcinogenicity and toxicity of food dyes, Int. J. Cancer,
         21, 361-367

    Radomski, J. L. & Deichman, W. B. (1956) Cathartic action and
         metabolism of certain coal tar food dyes, J. Pharmacol. Exp.
         Ther., 118, 322-327

    Rosenkranz, H. S. & Leifer, Z. (1980) In Chemical Mutagens: Principles
         and Methods for their Detection. In: de Serres, F. J. &
         Hollaender, A., New York and London, Plenum Press, Vol. 6, p. 109

    Sankaranarayanan, M. & Murthy, M. S.S. (1979) Testing of some
         permitted food colours for the induction of gene conversion in
         diploid yeast, Mutation Res., 67, 309-314

    Willheim, R. & Ivy, A. C. (1953) A preliminary study concerning the
         possibility of dietary carcinogenesis, Gastroenterology, 23,
         1-19
    


    See Also:
       Toxicological Abbreviations
       Fast green FCF  (FAO Nutrition Meetings Report Series 46a)
       Fast Green FCF (WHO Food Additives Series 20)
       Fast Green FCF (WHO Food Additives Series 21)
       FAST GREEN FCF (JECFA Evaluation)
       Fast Green FCF (IARC Summary & Evaluation, Volume 16, 1978)