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Additional information

The substance revealed uniformly negative results when tested in in vitro gene mutation assays and in the in vitro micronucleus assay. All studies were conducted without and with a metabolic activating system.

 

The respective Ames test was conducted according to OECD TG 471 with the following tester strains: Salmonella typhimurium TA 1535, TA 100, TA, 1537, and TA 98, and Escherichia coli WP2 uvrA. The assay was performed in two independent experiments, the initial plate incorporation and the subsequent preincubation test with doses up to and including 5000 µg/plate. Toxic effects occurred in all strains with and without metabolic activation.

No substantial increase in revertant colony numbers of any of the five tester strains was observed following treatment with the test substance at any dose level, neither in the presence nor absence of S9 mix. Appropriate reference mutagens were used as positive controls and showed a distinct increase of induced revertant colonies.

 

As further genotoxicity test a HPRT according to OECD TG 476 was conducted in order to investigate the potential of the substance to induce gene mutations at the HPRT locus in mammalian cells. V79 cells were exposed to the test item in the first experiment for 4 hours with and without metabolic activation and in the second experiment for 4 hours with and for 24 hours without metabolic activation. The maximum concentration of the pre-experiment (2440 µg/mL) was based on the solubility properties of the test item. The concentration range of the main experiments was limited by cytotoxicity.

No substantial and reproducible dose dependent increase of the mutation frequency was observed in both main experiments. Appropriate reference mutagens, used as positive controls, induced a distinct increase in mutant colonies and thus, showed the sensitivity of the test system and the activity of the metabolic activation system.

 

An in vitro MNT assessed the potential to induce micronuclei in human lymphocytes in accordance to OECD Guideline 487. In three independent experiments lymphocytes were exposed either for 4 hours (pulse exposure) with or without S9 mix, or for 20 hours (continuous exposure) without S9 mix. The highest treatment concentration in this study, 2400 μg/mL, was chosen with regard to the solubility properties of the test item.

In all experimental parts, concentrations showing clear cytotoxicity were not evaluable for cytogenetic damage. However, moderate cytotoxicity of 40.0 or 43.3 % cytostasis was observed in Experiment I with S9 mix and Experiment IB without S9 mix at the highest evaluated concentrations. In Experiment II in the presence of S9 mix clear cytotoxicity (52.5 % cytostasis) was observed at 447.8 μg/mL.

No biologically relevant increase in the number of cells carrying micronuclei was observed, neither in the absence nor in the presence of S9 mix. The positive controls used showed distinct increases in cells with micronuclei.

 

Summarizing, the available tests on different genotoxicity endpoints (gene mutations in bacterial and mammalian cells and clastogenicity) give no indication for a potential for genetic toxicity of the substance.


Justification for selection of genetic toxicity endpoint
None of the available studies is selected here, since all are relevant for the assessment of mutagenicity.

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

According to Regulation (EC) No 1272/2008, Annex I, no classification is warranted for genetic toxicity.