The data contained in this document were examined by the
    Joint FAO/WHO Expert Committee on Food Additives*
    Geneva, 18-27 April 1977

    Food and Agriculture Organization of the United Nations
    World Health Organization

    * Twenty-first Report of the Joint FAO/WHO Expert Committee on Food
    Additives, Geneva, 1977, WHO Technical Report Series No. 617





         When ponceau SX was given orally in doses of 100 mg to rats, only
    2% of unchanged dye was found in the faeces, but when it was
    administered slowly by intrasplenic infusion to rats with cannulated
    bile ducts 82% of the administered doses was excreted in the bile and
    4% in the urine within eight hours. Products of reductive cleavage of
    the colour, 2-amino-l-hydroxy-4-naphthalenesulfonic acid, 1-amino-2,
    4-dimethyl-5-benzenesulfonic acid and 2-acetamino-l-hydroxy-4-
    naphthalenesulfonic acid were found in the urine of rats fed the
    colour (Radomski and Mellinger, 1962).

         These results are in agreement with the findings that ponceau SX
    is rapidly reduced by bacteria obtained from the intestines of rats.
    Three amines were detected by thin-layer chromatographs in the
    incubation mixture (Roxon et al., 1967).

         After intravenous injection of ponceau SX into rats with bile
    fistulae, 66-71% of the dye was excreted in bile within four hours.
    The rate of excretion of ponceau SX (and of amaranth under the same
    conditions) varied with the dose (Iga et al., 1970).

    Special studies on carcinogenicity

    Oral administration

         Groups of 50 male and 50 female C57BL/He mice and similar groups
    of C3Heb/Jax mice were fed a diet containing 10 000 or 20 000 mg
    commercial ponceau SX per kg of diet for two years. Two groups of
    100 male and 100 female mice of each strain were fed a control diet.
    In C57BL/He mice fed the 10 000 mg/kg or 20 000 mg/kg levels, tumours
    occurred in 8/47 (six hepatomas, one reticulum-cell sarcoma and one
    other) and 12/56 (one mammary tumour, nine hepatomas and two others),
    respectively, compared with 13/91 (12 hepatomas and one reticulum-cell
    sarcoma) in controls. No tumours were reported in 50 and 28 C3Heb/Jax
    mice fed ponceau SX at each level, respectively, compared with 5/66 in
    controls (the numbers of animals given are those examined for
    pathology). In treated animals, 56-67% survived 52 weeks, compared
    with 91% of control C57BL/He and 58% of control C3Heb/Jax mice (Davis
    et al., 1966).

         A group of five male and five female Wistar rats was fed a diet
    containing 40 000 mg ponceau SX per kg of diet for up to 18 months. In
    1/7 rats living to a tumour-bearing age, a mesenteric lymphosarcoma
    was observed. No tumours occurred in 50 controls surviving for
    20 months or more (Willheim and Ivy, 1953). (The limited number of
    animals used was noted by the Working Group.)

         Five groups, each containing 12 male and 12 female Osborne-Mendel
    rats, were fed a diet containing 0 (control), 5000, 10 000, 20 000 or
    50 000 mg commercial ponceau SX per kg of diet for two years. Growth
    effect was negligible and there was no effect on survival, haematology
    or on weights of heart, liver, kidneys, spleen and testes at autopsy.
    Benign and malignant tumours, mainly pulmonary lymphosarcomas, mammary
    fibroadenomas and mammary adenocarcinomas, occurred in 10/19, 8/23,
    10/22 and 5/24 rats in the respective treatment groups, compared with
    7/16 controls (the numbers of animals given are those whose organs
    were examined microscopically) (Davis et al., 1966).

         Additional experiments, using groups of 200, 100 and 100 Osborne-
    Mendel rats of both sexes and 200, 100 and 100 Sprague-Dawley rats of
    both sexes fed a diet containing 0, 10 000 or 20 000 mg ponceau SX per
    kg of diet, resulted in tumour incidences of 67/171 (39%), 23/89 (26%)
    and 32/89 (36%) in Osborne-Mendel rats and 38/147 (26%), 16/83 (19%)
    and 14/74 (19%) in Sprague-Dawley rats in the respective groups (the
    numbers of animals given are those examined for pathology). Survival
    rates at 80 weeks were 79, 75 and 78%, respectively, in Osborne-Mendel
    rats and 61, 66 and 54%, respectively, in Sprague-Dawley rats (Davis
    et al., 1966).

         In a group of 50 non-inbred rats given ponceau SX in the diet at
    a concentration of 20 000 mg/kg of diet on six days a week for 33
    months (total dose, 107-139 g), 4/38 rats surviving at the appearance
    of the first tumour developed tumours, including three subcutaneous
    sarcomas and one hepatoma. No tumours were reported to have occurred
    in 50 controls surviving up to 33 months (Andrianova, 1970).

         Commercial ponceau SX was fed at a level of 0, 10 000 and
    20 000 ppm in the diet to five female beagle dogs (age six months) for
    up to seven years; three dogs died at six, nine-and-one-half and 64
    months of treatment. A second group of five females received ponceau
    SX at a level of 10 000 mg/kg of diet for seven years, while nine
    females served as controls. Two (20 000 ppm), five (10 000 ppm) and
    nine (control) dogs, respectively, survived the seven years of
    treatment. There was no apparent effect on weight gain during the
    first six months or any change on haematological parameters examined
    at any time during the duration of the experiment. Haemorrhagic
    streaks and/or projections in the urinary bladder of all test animals
    surviving more than six months were found. Microscopically the lesions
    varied from small submucosal haemorrhages to blood-filled mucosal

    projections usually without clots or fibrous stalk projections.  Most
    of the affected bladders also had lymphocytic foci in the muscular
    layer. The most important microscopic lesion attributable to the
    compound was a moderate to marked atrophy of the zone glomerulose in
    the adrenals of all test animals. There was no consistent effect on
    other zones of the adrenal cortex. In some instances there was partial
    replacement of the zone glomerulose by a subcapsular zone of
    pigmented, foamy macrophages. In dogs of both dose levels small foci
    of pigmented hepatic cells were observed. In one dog at the highest
    dose level congestion and haemorrhages in the ileum and mesenteric
    lymph nodes were found. Of dogs fed the 20 000 mg/kg level, two had
    adrenal cortical adenomas and one had a follicular-cell adenoma of the
    ovary. Of the dogs fed the 10 000 mg/kg level, one had an adrenal
    medullary adenoma, one a mammary adenocarcinoma. In the control dogs,
    two had adrenal cortical adenomas, two hyperplastic foci in the
    mammary glands and one nodular hyperplasia in the liver (Davis et al.,

    Subcutaneous and/or intramuscular injection

         Groups of 50 male and 50 female C57BL/He mice and similar groups
    of C3Heb/Jax mice received monthly s.c. injections of either 0.5 ml of
    a 2% aqueous solution of commercial ponceau SX or 0.5 ml of saline.
    About 60% of both control and experimental animals survived 52 weeks
    or more. In C57BL/He mice, 3/59 controls and 1/59 test animals
    developed tumours (two mammary tumours and one hepatoma in controls,
    one other tumour in treated animals), whereas in C3Heb/Jax mice, 0/69
    controls and 2/66 treated mice developed tumours (Davis et al., 1966).

         Groups of 50 male and 50 female Osborne-Mendel rats and similar
    groups of Sprague-Dawley rats received weekly s.c. injections of
    either 1 ml of a 2% aqueous solution of commercial ponceau SX or 1 ml
    of saline. Survivors at 80 weeks were 70% and 82% for control and test
    rats of the Osborne-Mendel strain and 61% and 56% for the Sprague-
    Dawley rats, respectively. No tumours at the site of injection
    occurred in 98 and 99 controls, compared with 1/100 and 2/98 in
    treated animals. The total tumour incidence was 29/98 in controls,
    compared with 33/100 in treated Osborne-Mendel rats, and 18/99
    controls, compared with 20/98 in treated Sprague-Dawley rats. Tumours
    were mainly mammary fibroadenomas, adenocarcinomas and lymphosarcomas
    of the lung. No tumours occurred in controls which were not found in
    treated animals, other than tumours at the site of injection (Davis et
    al., 1966).

    Acute toxicity

         The oral LD50 in male Wistar rats was > 2 g/kg body weight
    (Lu and Lavallée, 1964).

    Short-term studies

         No data available.

    Long-term studies

         (See carcinogenicity studies.)


    Andrianova, M. M. (1970) Carcinogenous properties of red food pigments
    amaranth, SX purple and 4R purple, Vop. Pitan., 29, 61-65

    Davis, K. J., Nelson, A. S., Zwickey, R. E., Hansen, W. H. and
    Fitzhugh, O. G. (1966) Chronic toxicity of ponceau SX to rats, mice
    and dogs, Toxicol. appl. Pharmacol., 8, 306-317

    Iga, T., Awazu, S., Hanano, M. and Nogami, H. (1970) Pharmacokinetic
    studies of biliary excretion. I. Comparison of the excretion behaviour
    in azo dyes and indigo carmine, Chem. Pharm. Bull., 18, 2431-2440

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

    Radomski, J. L. and Mellinger, T. J. (1962) The absorption, fate and
    excretion in rats of the water-soluble azo dyes FD and C Red No. 2, FD
    and C Red No. 4 and FD and C Yellow No. 6, J. Pharmacol. exp.
    Ther ., 136, 259-266

    Roxon, J. J., Ryan, A.D. and Wright, S. E. (1967) Reduction of water-
    soluble dyes by intestinal bacteria, Fd Cosmet. Toxicol., 5,

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

    See Also:
       Toxicological Abbreviations
       PONCEAU SX (JECFA Evaluation)
       Ponceau SX (IARC Summary & Evaluation, Volume 8, 1975)