Disodium [2,4-dihydro-4-[(2-hydroxy-5-nitrophenyl)azo]-5-methyl-2-phenyl-3H-pyrazol-3-onato(2-)][3-hydroxy-4-[(2-hydroxy-1-naphthyl)azo]-7-nitronaphthalene-1-sulphonato(3-)]chromate(2-)

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Additional information

Two Ames tests are available on Acid Brown 282; one test (Huntsman, 1995) was performed on strain: TA 1535, TA 1537, TA 98, and TA 100, and one test (R&CKFT, 2020) was performed on strain TA98, TA100, TA102, TA1535, TA1537 and the nitroreductase deficient strains, TA 98NR and TA 100 NR.

The Huntsman test was disrgegarded and the R&C test is considered in a Weight of Evidence approach for the reason discussed in the expert statement.

Therefore, one expert assessments was added as Key study, in order to analyse all the results of the Ames tests, considering also the structural and mechanicistic evaluations.

The final conclusion is that Acid Brown 282 has no concern fo mutagenicity in bacteria.

 

Having the first AMES showing some positivity (Huntsman, 1995), in order to avoid a new study with vertebrate animals, we included in the OECD 422 test (Annex VIII) also a part concerning the mutagenicity potential of the substance, in particular the analysis of the micronuclei.

Same animals used for OECD 422 were investigated and no new animals were killed for assessing this endpoint. Therefore, no testing proposal was inserted in the dossier for in vivo mutagenicity and an in vivo chromosomal aberration test is available on ABr282. The examination of the bone marrow and peripheral blood was performed with females only. At the termination of the Dose Range Finding Experiment with the test substance performed for Combined Study, the bone marrow harvesting and the blood taking was done. Evaluation and interpretation of results was performed according to OECD Test Guideline 474 (1997) Mammalian Erythrocyte Micronucleus Test. Statistically significant increase in the number of micronucleated polycbromatic erythrocytes in the bone marrow and normochromatic erythrocytes in peripheral blood compared to the control was not recorded at any dose level. Negative results in the micronucleus test indicate that under the test conditions, the test substance does not produce micronuclei in polychromatic erythrocytes in bone marrow and normochromatic erythrocytes in peripheral blood of rat. The test substance is considered to be non-genotoxic under the condition of the test (REACH&Colours Kft, 2014).

 

In order to assess also the mammalian cell gene mutation potential, data available on the structural analogue Similar Substance 03 have been taken into account. A study was performed to investigate the potential of test substance to induce gene mutations at the HPRT locus in V79 cells of the Chinese hamster. The assay was conducted in three independent experiments, both with and without liver microsomal activation. According to the pre-test on toxicity the concentration ranges were selected to yield concentration-related toxic effects. The highest concentration produced a low level of survival and the survival at the lowest concentration was approximately in the range of the negative control. Up to the highest investigated concentration no relevant increase in mutant colony numbers was obtained in all independent experiments. Appropriate reference mutagens were used as positive controls and showed a distinct increase in induced mutant colonies. Under the experimental conditions reported the test article did not induce gene mutations at the HPRT locus in V79 cells (Huntsman Textile Effects (Germany) GmbH, 1995).

The read across is discussed in section 13.

 

 

Justification for selection of genetic toxicity endpoint

The assessment  of the endpoint is performed with the integrated evaluation of results for in vitro bacteria gene mutation, in vitro mammalian cell gene mutation assay and in vivo micronucleus assay.

 

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

According to the CLP Regulation (EC 1272/2008), for the purpose of the classification for germ cell mutagenicity, substances are allocated in one of two categories in consideration of the fact that they are:

- substances known to induce heritable mutations or to be regarded as if they induce heritable mutations in the germ cells of humans or substances known to induce heritable mutations in the germ cells of humans or

- substances which cause concern for humans owing to the possibility that they may induce heritable mutations in the germ cells of humans.

The test substance did not show any reasons of concern in the test performed.

In conclusion, the substance is not classified for genetic toxicity according to the CLP Regulation (EC 1272/2008).