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Administrative data

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

In a first Ames test, metabolites of the test substance formed as a result of microsomal activation exerted a weak mutagenic action in this test system. In a second Ames test, no evidence of the induction of mutagenic effects by the test substance or by its metabolites formed as a result of microsomal activation was detectable in this experiment. This result was confirmed by an additional Ames test conducted using a Prival-modification. In a HPRT test, no genotoxicity was observed. A micronucleus assay yielded also negative results for genotoxicity.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Additional information

Bacterial reverse mutation assay

In an Ames test performed comparable to OECD guideline 471, 4 Salmonella typhimurium strains (TA 98, TA 100, TA 1535 and TA 1537) were used to test the mutagenic potential of the test substance (25, 75, 225, 675 and 2025 µg/0.1 mL) both with and without metabolic activation (CIBA-GEIGY, 1980a). In the experiments without microsomal activation, comparison of the number of back-mutant colonies in the controls and the test substance treated cultures revealed no marked differences. In the experiments whith metabolic activation, treatment with test substance led to a slight increase in the number of back-mutant colonies of Strain TA 98. In order to confirm this result, additional experiments were performed on Strains TA 98 and TA 1538 with the test substance (1000, 2000, 4000, 8000 µg/0.1 mL). In the experiments with microsomal activation treatment with test substance resulted in a slight increase of back-mutant colonies with both strains. The metabolites of the test substance formed as a result of microsomal activation thus exerted a weak mutagenic action in this test system.

In a study comparable to OECD guideline 471 the mutagenic potential of the test substance was tested in 5 Salmonella typhimurium Strains (TA 98, TA 100, TA 1535, TA 1537 and TA 1538) and in E. coli WP2 uvrA strain (CIBA-GEIGY, 1980b). The investigations were performed with the following concentrations of the test substance without and with microsomal activation: 5, 10, 50, 100, 500, 1000 and 5000 µg/0.1 mL. Since the results obtained in these experiments were equivocal, the experiments with Strains TA 98, TA 1537, TA 1538 and E. coli were repeated. In the experiments carried out without microsomal activation on Strains TA 1537 and TA 1538 a reduction in the colony count due to a growth-inhibiting effect of the compound was observed at the concentration of 5000 µg/0.1 mL. No evidence of the induction of point mutations by the test substance or by the metabolites of the substance formed as a result of microsomal activation was detectable in the strains of S. typhimurium and E. coli used in these experiments.

The test substance was tested for its mutagenic potential based on the ability to induce point mutations in selected loci of several bacterial strains, i.e. Salmonella typhimurium and Escherichia coli, in a reverse mutation assay (Ames standard plate test, Preincubation test and Prival preincubation test) at concentration of 33 - 5200 µg/plate in presence and absence of a metabolic activation system. The modified Bacterial Reverse Mutation Test according to Prival facilitates azo reduction and is therefore the most approriate method for the investigation of azo-dyes and diazo compounds. Precipitation of the test substance was found from about 100 μg/plate onward with and without S9 mix. A weak bacteriotoxic effect was occasionally observed depending on the strain and test conditions from about 1 000 μg/plate onward. A biologically relevant increase in the number of his+ or trp+ revertants was not observed in the standard plate test, the preincubation test or in the prival preincubation test either without S9 mix or after the addition of a metabolizing system. Thus, under the experimental conditions of this study, the test substance is not mutagenic in the Salmonella typhimurium/Escherichia coli reverse mutation assay in the absence and the presence of metabolic activation.

In vitro gene mutation test in mammalian cells

In a mammalian cell cytogenetics assay CHO cell cultures were exposed to the test substance at concentrations of 0;1.56;3.13; 6.25; 12.50; 25.00; 50.00μg/mL, (1. Experiment) and 0; 2.50; 5.00; 10.00; 20.00; 40.00; 80.00 μg/mL (2. Experiment) with and without metabolic activation (S9 mix,phenobarbital and β-naphthoflavoneinduced). The study was conducted according to GLP and OECD Guideline 476. The exposure duration was 4 hours. The test substance was tested up to clearly precipitating concentrations in culture medium due to missing cytotoxicity in the pretest. Although, in the 1st Experiment after 4 hours exposure in the absence of metabolic activation clear cytotoxicity was observed at a strongly precipitating concentration. Positive and vehicle controls induced the appropriate response. Both positive control substances, EMS and DMBA, led to the expected increase in the frequencies of forward mutations. The test substance did not cause any biologically relevant increase in the mutant frequencies either without S9 mix or after the addition of a metabolizing system in two experiments performed independently of each other. The test substance was therefore concluded to be non-mutagenic in the HPRT locus assay.

In vivo genotoxicity

The test item was administered by gavage. Treatment consisted of one daily dose of 750, 1500 or 3000 mg/kg on two consecutive days. The animals were sacrificed 24 h after the second application. From the bone marrow smears were made. The experiment was performed to evaluate any mutagenic effect on somatic interphase cells in vivo. Mutagenic effects present themselves in interphase cells in form of nucleus anomalies of bone marrow cells. These anomalies occur in interphase cells as a consequence of damage during the mitotic process. The increase in anomalies shows a clear dose dependency, comparable to the occurrence of chromosome aberrations in metaphase preparations . The bone marrow smears from animals treated with various doses of the test material showed no significant difference from the control. The incidence of bone marrow cells with anomalies of nuclei corresponds to the frequency observed in the control group. It is concluded that under the conditions of this experiment, no evidence of mutagenic effects was obtained in Chinese hamsters treated with the test substance.


Justification for selection of genetic toxicity endpoint
There are several studies available addressing the genetic toxicity of the test substance. As the genetic toxicity is assessed using more than one study type, all of the available data were used for the assessment and no endpoint selection was made.

Justification for classification or non-classification

Dangerous Substance Directive (67/548/EEC)

The available studies are considered reliable and suitable for classification purposes under 67/548/EEC. As a result the substance is not considered to be classified for genotoxicity under Directive 67/548/EEC.

 

Classification, Labeling, and Packaging Regulation (EC) No. 1272/2008

The available experimental test data are reliable and suitable for classification purposes under Regulation 1272/2008. As a result the substance is not considered to be classified for genotoxicity under Regulation (EC) No. 1272/2008.