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Administrative data

Key value for chemical safety assessment

Additional information

There are valid in vitro data available for the assessment of the genetic toxicity of the test item.

In vitro

Gene mutation in bacteria

In a non-GLP bacterial reverse mutation assay (BASF, 80/0404, 1981), conducted similar to OECD Guideline 471 (strain e.g. TA 102 or E.coli WP2 to detect crosslinking agents and second independent trial is missing), the test substance (purity > 98%) was analysed for mutagenicity in the Ames test using the plate incorporation test in the presence and absence of mammalian metabolic activation (liver fraction (S9) from Aroclor 1254 induced rat). 

The Salmonella typhimurium strains TA 98, TA 100, TA 1535, TA 1537, and TA 1538 were treated with the test substance dissolved in DMSO at concentrations of 4, 20, 100, 500, and 2500 µg/plate. Each concentration, including the controls (positive and negative), was tested in triplicate.

Incomplete solubility of the test substance was observed at 500 µg/plate and above. 4 replicates of each test group were examined after 48 hours of exposure. An increase in the number of his+ or trp+ revertants was not observed with and without metabolic activation and consistently a negative result was obtained. No cytotoxicity was observed in any of the strains although tested up to precipitation doses. The positive controls yielded the expected results. According to the results of the present study, the test substance is not mutagenic in the bacterial reverse mutation assay under the experimental conditions chosen.

This result is supported by a further Ames test (BASF, 80/0404, 1983), which tested only the Salmonella typhimurium strain TA98 with and without metabolic activation.Using the plate incorporation method the test substance was tested at concentrations of 20, 100, 500, 1000, 2500, and 5000 µg/plate. DMSO was used as vehicle. At 500 µg/plate and above incomplete solubility of the test substance was observed. Four replicates of each test group were examined after 48 hours of exposure. Neither genotoxicity nor cytotoxicity was observed, although the test substance was tested up to precipitation doses. The positive controls yielded the expected results. According to the results of the present study, the test substance is not mutagenic in the bacterial reverse mutation assay under the experimental conditions chosen.

 

Gene mutation in mammalian cells

In a GLP conform in vitro mammalian cell gene mutation test (HPRT-Test) according to OECD Guideline 476 (BASF, 50M0060/11X114, 2012) the test substance was investigated for its potential to induce gene mutations at the HPRT locus in V79 cells. The assay was performed in two independent experiments, using two parallel cultures each. In the first main experiment cells were exposed for 4 hours to the test item at concentrations of 1.3, 2.5, 10, 20, 40 µg/mL with and without liver microsomal activation. The second experiment was performed with a treatment time of 4 hours at concentrations of 2.5, 5.0, 10.0, 20.0, 40.0, 80.0 µg/mL with metabolic activation and a treatment time of 24 hours at concentrations of 1.3, 2.5, 5.0, 10.0, 20.0, 40.0 µg/mL without metabolic activation.

In the first experiment turbidity was observed at 10.0 µg/mL in the absence of metabolic activation.At the two highest concentrations of 20.0 and 40.0 µg/mL precipitation was noted. In the presence of metabolic activation turbidity occurred at 20.0 µg/mL. At 40.0 µg/mL precipitation was noted.

In the second experiment turbidity was observed in the absence of metabolic activation at concentrations of 5.0 - 20.0 µg/mL and precipitation occurred at the maximum concentration of 40.0 µg/mL.In the presence of metabolic activation turbidity was noted from 5.0 to 20.0 µg/mL. At 40.0 and 80.0 µg/mL precipitation occurred.

Appropriate reference mutagens (EMS and DMBA), 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.

No relevant toxic effects occurred up to the maximum concentration with and without metabolic activation following 4 and 24 hours treatment.

No relevant and reproducible increase in mutant colony numbers/106 cells was observed in the main experiments up to the maximum concentration with and without metabolic activation.All mutant frequencies remained well within the historical range of solvent controls.

According to the results of the present study, the test substance is not mutagenic in the in vitro mammalian cell gene mutation test under the experimental conditions chosen.

 

In vitro mammalian chromosome aberration test

In an in vitro mammalian chromosome aberration test (BASF, 32M0060/11X112, 2011), according to OECD Guideline 473 the test substance, (purity 97.7%) dissolved in acetone, was assessed for the ability to induce structural chromosome aberrations in V79 cells of the Chinese hamster in two independent experiments at concentrations of 1.4, 2.7, 5.5, and 10.9, 21.9, 43.8, 87.5, 175.0, 350.0 µg/mL with and without metabolic activation. In the first experiment the exposure period was 4 hours with and without S9 mix and in the second experiment 4 hours with S9 mix and 18 and 28 hours without S9 mix.

Visible precipitation of the test item in the culture medium was observed at 21.9 µg/mL (Experiment I) and 5.5 µg/mL (Experiment II) and above in the absence and presence of S9 mix. No relevant influence on osmolarity or pH value was observed.

No cytotoxicity was observed in the absence and presence of S9 mix up to the highest evaluable concentration. However, evaluation was limited by severe test item precipitation.

No clastogenicity was observed at the concentrations evaluated either with or without metabolic activation. However, in the first experiment in the absence of S9 mix, two statistically significant increases were observed after treatment with 2.7 and 10.9 µg/mL (2.5 and 2.0 % aberrant cells, excluding gaps). The values are in the range of the historical control data (0.0 - 4.0 % aberrant cells, excluding gaps) and therefore biologically irrelevant. No evidence of an increase in polyploid metaphases was found after treatment with the test item as compared to the frequencies of the control cultures.

Appropriate mutagens (EMS and CPA) used as positive controls produced valid responses.

In conclusion, it can be stated that under the experimental conditions reported, the test item did not induce structural chromosomal aberrations in V79 cells of the Chinese hamster in vitro, when tested up to precipitating or the highest evaluable concentration. Therefore, the test item is considered to be non-clastogenic in this chromosome aberration test in the absence and presence of metabolic activation, when tested up to precipitating or the highest evaluable concentration.

 

In vivo

No data were available for the assessment of genetic toxicity in vivo.

Conclusion

The test item gave no indication of a mutagenic effect in bacteria and mammalian cells and was not shown to induce structural chromosome aberrations in mammalian cells in the presence and absence of metabolic activation.


Justification for selection of genetic toxicity endpoint
All studies are considered relevant.

Short description of key information:
In vitro studies:
Ames: negative (BASF, 80/0404, 1981), similar to OECD Guideline 471
Ames: negative (BASF, 80/0404, 1983), no guideline followed
HPRT: negative (BASF, 50M0060/11X114, 2012), according to OECD Guideline 476
CA: negative (BASF, 32M0060/11X112, 2011), according to OECD Guideline 473
In vivo studies:
No data available.

Endpoint Conclusion: No adverse effect observed (negative)

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 mutagenicity under Directive 67/548/EEC, as amended for the 31st time in Directive2009/2/EG.

 

Classification, Labelling, 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 mutagenicity under Regulation (EC) No. 1272/2008, as amended for the second time in Directive (EC 286/2011).