Sodium bis[3-[[1-(3-chlorophenyl)-4,5-dihydro-3-methyl-5-oxo-1H-pyrazol-4-yl]azo]-4-hydroxybenzenesulphonamidato(2-)]cobaltate(1-)

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

In vitro Gene Mutation study in Bacteria -AMES

The substance exerted a marginal mutagenic effect in this test system.

In vitro mammalian cells Chromosome aberration, OECD473

The substance is considered to be weakly mutagenic in this chromosome aberration test. Due to the strong toxic effects, an indirect mechanism for DNA damaging can not be excluded.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (positive)

Genetic toxicity in vivo

Description of key information

In vivo Mammalian Erythrocytes Micronucleous, OECD474, Micronucleus Assay in Bone Marrow Cells of the Mouse

Negative. The substance is considered to be non-mutagenic in this in vivo micronucleus assay.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Additional information

Studies on bacteria (AMES)

The test has been conducted according to the OECD Guideline 471 and to the EU Method B.13/B.14 in order to evaluate the potential of the test item to induce gene mutations according to the plate incorporation test.The following strains of Salmonella typhimurium were used: TA 98, TA 100, TA 102, TA 1535 and TA 1537.

The test was performed with and without the addition of rat-liver post mitochondrial supernatant (S9 fraction) as an extrinsic metabolic activation system.

In the experiments performed without metabolic activation with strain TA 102, a slight increase in the number of back-mutants was observed at concentrations of 625.0 and 333.3 ug/plate. A similar effect with this strain in the experimental part with metabolic activation was not reproducible. No effect at all occurred with the other strains.

As a conclusion, the substance exerted a marginal mutagenic effect in this test system.

Studies on in vitro mammalian cells

The substance was tested for chromosome aberrations in vitro, according to the OECD473, EU B.10.

The test article, dissolved in DMSO, was assessed for its potential to induce structural chromosome aberrations in V79 cells of the Chinese hamster in vitro. The exposure period was 4 h with and 18 h without metabolic activation.

In the main experiments, reduced mitotic indices were observed in the absence of S9 mix after continuous treatment with 10 ug/mL (32.0 % of control) and in the presence of S9 mix after 4 h treatment with 80 ug/ml (69.2 % of control). No relevant reduction of cell numbers was observed.

In the absence of S9 mix, the number of cells carrying structural chromosome aberrations was significantly increased after treatment with 10 ug/ml (6.5 %) and beyond our historical control data range ( 0 - 4 %). However, at this concentration marked toxicity indicated by reduced mitotic indices was observed. Therefore, it can not be excluded, that the observed effects were caused by non-genotoxic DNA damaging mechanisms. In the presence of S9 mix, a significant increase in the number of cells carrying structural chromosome aberrations (4.0 % aberrant cells exclusive gaps) within our historical control data range ( 0 - 4 %) was observed. Since no strong toxicity was induced, it can not be excluded, that treatment with higher concentrations would lead to higher aberration frequencies.

The occurrence of polyploid metaphases. In this study, no biologically relevant increase in the rate of polyploid metaphases was found after treatment with the test article (4.4 % - 7.4 %) as compared to the rates of the negative and solvent controls (3.9 % - 6.2 %).

In conclusion, it can be stated that in the study described and under the experimental conditions reported, the test article induced structural chromosome aberrations as determined by the chromosome aberration test in V79 cells (Chinese hamster cell line) in vitro. Therefore, the substance is considered to be weakly mutagenic in this chromosome aberration test. Due to the strong toxic effects, an indirect mechanism for DNA damaging can not be excluded.

Studies in vivo

The study was performed according to the in vivo OECD474, EU B.12, in order to investigate the potential of the substance to induce micronuclei in polychromatic erythrocytes (PCE) in the bone marrow of the mouse.

After treatment with the test item the number of NCEs was not substantially increased as compared to the mean value of NCEs of the vehicle control thus indicating that the substance had no cytotoxic effectiveness in the bone marrow.

In comparison to the corresponding vehicle controls there was no statistically significant or biologically relevant enhancement in the frequency of the detected micronuclei at any preparation interval after administration of the test item and with any dose level used.

In conclusion, it can be stated that during the study described and under the experimental conditions reported, the test item did not induce micronuclei as determined by the micronucleus test with bone marrow cells of the mouse. Therefore, the substance is considered to be non-mutagenic in this in vivo micronucleus assay.

Justification for classification or non-classification

This hazard class is primarily concerned with substances that may cause mutations in the germ cells of humans that can be transmitted to the progeny.

Substance that are mutagenic in somatic cells may produce heritable effects if they, or their active metabolites, have the ability to interact with the genetic material of germ cells. Conversely, substances that do not induce mutations in somatic cell in vivo would not be expected to be germ cell mutagens.

However, the results from mutagenicity or genotoxicity tests in vitro and in mammalian somatic and germ cells in vivo are also considered in classifying substances and mixtures within this hazard class.

Category 1: substances known to induce heritable mutations or to be regarded as if they induce heritable mutations in the germ cells of humans. Substances known to induce heritable mutations in the germ cells of humans.

Category 2: substances which cause concern for humans owing to the possibility that they may induce heritable mutations in the germ cells of humans.

Classification for heritable effects in human germ cells is made on the basis of well conducted, sufficiently validated tests as In vitro mutagenicity tests such as these indicated in 3.5.2.3.8:

- in vitro mammalian chromosome aberration test;

- in vitro mammalian cell gene mutation test;

- bacterial reverse mutation tests

In the in vitro bacteria experiments performed without metabolic activation with strain TA 102, a slight increase in the number of back-mutants was observed and the substance exerted a marginal mutagenic effect in this test system.

An in vitro additional study is available were the test article induced structural chromosome aberrations as determined by the chromosome aberration test in V79 cells (Chinese hamster cell line) in vitro. Therefore, the substance is considered to be weakly mutagenic in this chromosome aberration test, according to OECD473. Due to the strong toxic effects, an indirect mechanism for DNA damaging can not be excluded.

However, the in vivo OECD474, EU B.12, was performed in order to investigate the potential of the substance to induce micronuclei in polychromatic erythrocytes (PCE) in the bone marrow of the mouse, and during the study described and under the experimental conditions reported, the test item did not induce micronuclei as determined by the micronucleus test with bone marrow cells of the mouse. Therefore, the substance is considered to be non-mutagenic in this in vivo micronucleus assay.

As conclusion, according to the CLP Regulation n.1272/2008 and the ECHA Guidance R.7a, the substance is not classified as mutagenic.