C.I. Reactive Yellow 161

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1983

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1983

Report date:

1983

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

EU Method B.12 (Mutagenicity - In Vivo Mammalian Erythrocyte Micronucleus Test)

GLP compliance:

yes (incl. QA statement)

Type of assay:

mammalian erythrocyte micronucleus test

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

400-640-4

EC Name:

-

Cas Number:

198153-89-8

Molecular formula:

C32ClH25N10Na2O9S3

IUPAC Name:

disodium 2-(4-{[4-({3-[2-(5-carbamoyl-1-ethyl-2-hydroxy-4-methyl-6-oxo-1,6-dihydropyridin-3-yl)diazen-1-yl]-4-sulfonatophenyl}amino)-6-chloro-1,3,5-triazin-2-yl]amino}phenyl)-6-methyl-1,3-benzothiazole-7-sulfonate

Test material form:

solid: particulate/powder

Details on test material:

Reactive Yellow 161

Test animals

Species:

mouse

Strain:

CF-1

Remarks:

non-consanguineous OF-1 albino mice

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: IFFA-CREDO, L'Arbresle, France (Charles River)
- Age at study initiation:
- Weight at study initiation: ca 25 g
- Assigned to test groups randomly: yes
- Housing: in groups of 5 separated by sex
- Diet (ad libitum): Aliment Rats-Souris Charles River, produced by U.A.R., Villemoisson, France
- Water (ad libitum): tap water
- Acclimation period: 1 week

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

water

Details on exposure:

We first carried out a preliminary range finding test by treating three groups of three male mice with doses of 500, 1000 and 2000 mg/kg given by intra-gastric intubation. As no death occurred, the dose of 2000 mg/kg was chosen for the micronucleus test. This concentration was the maximum quantity of the test substance which can be suspended in water.
For the micronucleus test, each treatment group was comprised of five male and five female mice receiving one intragastric intubation using 0.5 ml per 25 g body weight (20 mL/kg bw). Animals were sacrificed at regular intervals after treatment.

Duration of treatment / exposure:

1. Control (distilled water): 0.5 ml - sacrificed at 44 hours
2. Positive control (Thio-TEPA): 20 mg/kg - sacrificed at 44 hours
3. FAT 40162/B: 2000 mg/kg - Sacrificed at 20 hours
4. FAT 40162/B: 2000 mg/kg - Sacrificed at 44 hours
5. FAT 40162/B: 2000 mg/kg - Sacrificed at 68 hours.

Frequency of treatment:

single

Post exposure period:

20, 44, 68 h

Doses / concentrations

Dose / conc.:

2 000 mg/kg bw/day (nominal)

No. of animals per sex per dose:

5

Control animals:

yes, concurrent vehicle

Positive control(s):

Thio-TEPA (N, N', N"-triethylenethiophosphoramide), reference N° 509-227, made by Lederle Laboratories Ltd.

Examinations

Tissues and cell types examined:

bone marrow - erythrocytes
Two types of erythrocytes were observed in the bone marrow smears: normo-chromatic (mature red blood cells about to pass into the blood stream) and polychromatic (immature red blood cells).
The proportion of polychromatic erythrocytes containing one or more micronuclei was compared with the total number of polychromatic erythrocytes, and statistical comparisons were made between these ratios for the different groups. A minimum of 500 polychromatic erythrocytes were counted per smear (two smears per animal).

Details of tissue and slide preparation:

The method of preparation of mouse bone marrow smears for use in the micro-nucleus test was described in detail by Schmid (1977).
After sacrifice of the animals the femurs were taken and broken open at one end. Bone marrow cells were suspended in foetal calf serum using a small syringe, and the cells were centrifuged at 120 x g for 5 minutes. The super-natant was removed with a Pasteur pipette, cells were spread on a microscope slide and the smears allowed to dry in air. The following day smears were stained with Giemsa (1:6 in water) and mounted with a coverslip after drying.

Statistics:

The proportion of polychromatic erythrocytes containing one or more micronuclei was compared with the total number of polychromatic erythrocytes, and statistical comparisons were made between these ratios for the different groups using BMPD computer programme 7D.

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

negative

Toxicity:

no effects

Vehicle controls validity:

valid

Positive controls validity:

valid

Additional information on results:

There was no increase in the number of micronucleated polychromatic erythrocytes in animals exposed to 2000 mg/kg bw compared to negative control animals. In animals treated with Thio-TEPA there is a statistically significant increased number of micronucleated cells.
The ratio of polychromatic to normochromatic erythrocytes is markedly decreased in mice treated with Thio-TEPA. There is no statistical difference between the animals treated with the test substance and the negative control for this ratio.

Applicant's summary and conclusion

Conclusions:

The test substance did not exhibit any sign of cytogenicity in the mouse micronucleus test.

Executive summary:

Reactive Yellow 161 was assayed for mutagenicity using the micronucleus test (EEC protocol). The compound was administered orally to mice at a concentration of 2000 mg/kg body weight. No mutagenic effect was observed in bone marrow smears taken 20, 44 and 68 h after administration of the test substance. A positive control (Thio-TEPA) administered at a concentration of 20 mg/kg showed pronounced evidence of mutagenicity 44 h after administration.