INTERNATIONAL PROGRAMME ON CHEMICAL SAFETY
WORLD HEALTH ORGANIZATION
TOXICOLOGICAL EVALUATION OF SOME
FOOD COLOURS, ENZYMES, FLAVOUR
ENHANCERS, THICKENING AGENTS, AND
CERTAIN FOOD ADDITIVES
WHO FOOD ADDITIVES SERIES 6
The evaluations contained in this publication were prepared by the
Joint FAO/WHO Expert Committee on Food Additives which met in Rome,
4-13 June 19741
World Health Organization Geneva 1975
1 Eighteenth Report of the Joint FAO/WHO Expert Committee on
Food Additives, Wld Hlth Org. techn. Rep. Ser., 1974, No. 557.
FAO Nutrition Meetings Report Series, 1974, No. 54.
AZORUBINE
BIOLOGICAL DATA
BIOCHEMICAL ASPECTS
Rats were injected intravenously with approximately 1 mg of the
dye. The bile was collected for six hours and analysed. The recovery
of the dye was an average of 38% (30-40%) of the administered quantity
(Ryan & Wright, 1961). This like any other azo dye is probably reduced
in the gut by bacterial azo reductases (Walker, 1970).
TOXICOLOGICAL STUDIES
Special studies on Heinz bodies
Cat
Four cats were given 5% aqueous solution in doses of 1g on the
first day and 0.1 g on the ninth and eighteenth day. A negative test
for Heinz bodies was obtained (Deutsche Forsch., 1957).
Special studies on mutagenicity
The colour was tested for mutagenic action in a concentration of
0.5 g/100 ml in cultures of Escherichia coli. No mutagenic effect
was found (Lück & Rickerl, 1960).
Special studies on reproduction
Rat
Twenty-five male and 25 female rats received 1% azorubine in
their drinking water for 180 days, giving approximately 55 g dye per
animal. A similar group of 50 rats acted as controls. Weight gain,
mortality and general condition were similar in both groups. After
seven months animals were mated and an F1 generation produced. After
weaning the pups were put again on 1% azorubine and after four months
mated to produce an F2 generation. No abnormalities regarding litter
number or fertility were noted. After 200 days on 1% azorubine the F2
generation was kept on normal diet and water for two years. No adverse
effects were seen on mortality or tumour incidence (Hecht, 1966).
Special studies on sensitizing effects
Guinea-pig
In experiments on guinea-pigs, it was found that this colour had
no sensitizing activity (Bär & Griepentrog, 1960).
Acute toxicity
LD50
Animal Route (mg/kg bw) Reference
Mouse oral 1 000 Deutsche Forsch., 1957
oral 8 000 Gaunt et al., 1967
i.p. 900 Gaunt et al., 1967
i.v. 800 Deutsche Forsch., 1957
Rat oral 10 000 Gaunt et al., 1967
oral 1 000 Deutsche Forsch., 1957
i.p. 1 000 Deutsche Forsch., 1957
i.p. 900-1 100 Gaunt et al., 1967
i.v. 800 Deutsche Forsch., 1957
Short-term studies
Rat
Three weanling rats were given a 0.1% solution of carmoisine to
drink for 28 days (daily consumption approximately 15 mg). No toxic
effects were noted (Goldblatt & Frodsham, 1952).
Five groups of 16 male and 16 female rats were fed diets
containing 0, 0.05, 0.1, 0.5 and 1.0% carmoisine for three months.
There were no adverse effects on general health, growth or food
consumption and no departure from normality was seen in the
haematological studies and in the liver and kidney function tests. The
principal organ weight finding was an elevated relative kidney weight
at 1% in females. The types and incidence of histological changes were
comparable in control and test groups (Gaunt et al., 1967).
Pig
Four groups of three male and three female pigs received
carmoisine at dose levels of 0, 250, 500 or 1000 mg/kg/day for three
months. There were no adverse effects on growth and no differences
between treated and control groups were evident in the haematological
examination, serum chemistry, urine analyses, organ weights or
histological examination (Gaunt et al., 1969).
Long-term studies
Mouse
Five groups of 30 male and 30 female mice were fed diets
containing 0, 300, 900, 2700 or 8100 ppm of the colour for 80 weeks. A
high death rate was noted in all test groups and in the controls. A
further test was therefore undertaken with the same number of mice but
using dosage levels of 0, 100, 500, 2500 and 12 500 ppm. There were no
adverse effects on mortality, weight gain, organ weights or incidence
of histopathological findings including tumours. There was a mild
anaemia in mice given the highest level (BIBRA, 1973).
A group of 30 mice, comprising equal numbers of both sexes were
given 0.1 ml of a 3% suspension of the colour in arachis oil; after
six months the strength of the suspension was increased to 6%. The
colour was given subcutaneously for 360 days. After this period the
animals were kept for 630 days. The weekly dose was respectively 6 and
12 mg and the total dose was 468 mg/mouse. Seven lymphomas and one
intestinal tumour were observed. In 60 control animals four lymphomas
and an intestinal tumour were present (Bonser et al., 1956).
Rat
Ten rats were given the colour in the drinking water in a
concentration of 1% for 209 days. The daily intake was 1.2 g/kg bw and
the total amount administered was 51 g per animal. The observation
period was 919 days. No tumours were found (Deutsche Forsch., 1957).
Ten rats were given 1% of the colour in the drinking water for
250 days. The daily intake was 7.94 g/kg bw and the total intake
52 g/animal. The observation period was 545 days. No tumours were
found (Deutsche Forsch., 1947).
Ten rats were given a diet containing 0.2% of the colour for 417
days. The daily intake was approximately 0.1 g/kg bw and the total
intake was 11 g per animal. The observation period was 838 days. No
tumours were found (Deutsche Forsch., 1957).
A group of 10 rats was given twice weekly subcutaneous injections
of 0.5 ml of a 1% solution (= 5 mg) of the colour for one year. The
animals were kept under observation for over 938 days. One axillary
tumour was observed in one animal (Deutsche Forsch., 1957).
In a repeat experiment another group of 10 rats was given twice
weekly subcutaneously 0.5 ml of a 1% solution (= 5 mg) of the colour
for one year. No tumours were formed after 521 days each animal having
received 0.5 g (Deutsche Forsch., 1957).
Comments:
Little information is available on the metabolism of this colour.
It has been studied adequately long-term only in the mouse. The long-
term studies in the rat recorded only tumour incidence and survival
while many other essential observations have not been reported. The
reproduction study did not reveal any compound-related adverse
effects. There are no studies available on embryotoxicity including
teratology.
EVALUATION
Level causing no toxicological effect
Rat: 0.2% (= 2000 ppm) in the diet equivalent to 100 mg/kg bw.
Estimate of acceptable daily intake for man
0-0.5 mg/kg bw*
FURTHER WORK OR INFORMATION
Required by June 1978
(a) adequate long-term studies in another species;
(b) investigation of embryotoxicity including teratogenicity;
(c) adequate metabolic studies in several species preferably
including man.
REFERENCES
Bär, F. & Griepentrog, F. (1960) Med. u. Ernährung, 1, 99
BIBRA (1973) Research Report No. 8/1973
Bonser, G. M., Clayson, D. B. & Jull, J. W. (1956) Brit. J. Cancer,
10, 653
Deutsche Forschungsgemeinschaft, Bad Godesberg, Federal Republic of
Germany, Farbstoff Kommission (1957) Mitteilung 6
Gaunt, I. F. et al. (1967) Fd. Cosmet Toxicol., 5, 179
Gaunt, I. F. et al. (1969) Fd. Cosmet Toxicol., 7, 1
* Temporary.
Lück, H. & Rickerl, E. (1960) Z. Lebensmitt.-Untersuch., 112, 157
Ryan, A. J. & Wright, S. E. (1961) J. Pharm. Pharmacol., 13, 493
Walker, R. (1970) Fd. Cosmet Toxicol., 8, 659
Goldblatt & Frodsham (1952) Unpublished information from ICI
Hecht, G. (1966) Report to WHO
See Also:
Toxicological Abbreviations
Azorubine (WHO Food Additives Series 13)
Azorubine (WHO Food Additives Series 18)
AZORUBINE (JECFA Evaluation)