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Genetic toxicity in vitro

Description of key information

Two in vitro genetic toxicity studies have been conducted on the test article.  The results of the studies are:

Bacterial Reverse Mutation Assay: Positive when tested according to OECD 471 (1997).

Mammalian Micronucleus Assay: Positive when tested according to OECD 487 (2016).

Additional information

The mutagenic potential of the test article was evaluated in the bacterial reverse mutation assay (Ames assay) with S. typhimurium strains TA98, TA100, TA1535, and TA1537 and E.coli strain WP2 uvrA in the presence and absence of metabolic activation. The study was conducted according to OECD 471 in compliance with OECD GLP. Based on the results of a dose-range finding test, the test article was tested in the mutation assay at a concentration range of 52 to 5000 μg/plate in the absence and presence of S9-mix in the tester strains TA1535, TA1537 and TA98. The bacterial background lawn was not reduced at any of the concentrations tested and no biologically relevant decrease in the number of revertants was observed. In the absence of S9-mix, the test item induced dose-related increases in the number of revertant colonies compared to the solvent control in three tester strains (TA100, WP2uvrA and TA1535). The

increases observed were above the laboratory historical control data range, and were up to 7.9-

3.0-, and 16-fold the concurrent vehicle controls in the tester strains TA100, WP2uvrA and TA1535, respectively. In the presence of S9-mix, the test item induced dose-related increases in the number of revertant colonies compared to the solvent control in three tester strains (TA100, WP2uvrA and TA1535). The increases observed were above the laboratory historical control data range, and were up to 11-, 2.6-, and 112-fold the concurrent vehicle controls in the tester strains TA100, WP2uvrA and TA1535, respectively. The two other bacterial strains showed negative responses over the entire dose range, i.e. no dose related increase in the number of revertants. The negative and strain-specific positive control values were within the laboratory historical control data ranges indicating that the test conditions were adequate

and that the metabolic activation system functioned properly. Since 2.6- to 112-fold, dose related increases were observed in three tester strains, both in the absence and presence of S9-mix, these increases are considered biologically relevant and the test article is mutagenic in the absence and presence of S9-mix. Based on the results of the study, the test article is positive in the Ames assay in the presence and absence of metabolic activation.

The genotoxic potential of the test article was evaluated in the mammalian cell micronucleus test in Human Peripheral Blood Lymphocytes (HPBL). The study was conducted according to OECD 487 (2016) in compliance with OECD GLP regulations. The test article was dissolved in DMSO and HPBLs were exposed to the test article in medium at concentrations up to 60 ug/mL without metabolic activation and 125 ug/mL with metabolic activation (S9 mix). HPBLs were exposed to the test article for 4 and 24 hrs without activation and 4 hrs with activation. Cytotoxicity was observed in groups without metabolic activation at doses greater than or equal to 40 ug/mL (4 hr), greater than or equal to 30 ug/mL (24 hr) and in groups with metabolic activation at doses greater than or equal to 80 ug/mL (4 hr). In the 4-hour group without metabolic activation, statistically significant dose-dependent increases in micronuclei induction were observed at doses 20 and 40 ug/mL. In the 24-hour group without metabolic activation, a statistically significant increase in micronuclei induction was observed at 10 ug/mL but the response was not dose-dependent. In the 4-hour group with metabolic activation, a statistically significant and dose-dependent increase in micronuclei induction was observed at 80 ug/mL. Based on the results of the study, the test article is positive for the induction of micronuclei with and without metabolic activation in the in vitro mammalian micronucleus test using human peripheral blood lymphocytes.

Justification for classification or non-classification

Based on the available in vitro data and out of an abundance of caution, the test article should be classified as a GHS Category 2 germ cell mutagen (CLP: Muta. 2).

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