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The test item was investigated for its potential to induce gene mutations according to the plate incorporation test (experiment I) and the pre-incubation test (experiment II) using S. typhimurium strains TA98, TA 100, TA 1535, TA 1537 and tester strain E. coli WP2 uvrA. Each assay was conducted with and without metabolic activation. The following concentrations were used: 31.6, 100, 316, 1000, 2500 and 5000 ug/plate. Precipitation of the test item was observed in any tester strain used in experiment I and II (with and without metabolic activation). In experiment II toxic effects of the test item were noted at concentrations of 316 ug/plate and higher (with metabolic activation) in tester strain TA 1535. No biologically relevant increases in revertant colony numbers of any of the five tester strains were observed following treatment at any concentration level, neither in the presence nor absence of metabolic activation in experiment I and II. In conclusion, it can be stated that during the described mutagenicity test and under the experimental conditions reported, the test material did not cause gene mutations by base pair changes or frameshifts in the genome of the tester strains used.

In order to investigate the potential of of the test item for its ability to induce structural chromosome aberrations in V79 cells of the Chinese hamster in vitro a chromosome aberration assay was carried out. The chromosomes were prepared 20 h after start of treatment. The treatment interval was 4 h with and without metabolic activation at concentrations of 62.5 - 500 ug/ml in experiment I. In experiment II, the treatment interval was 4 h with and 20 h without metabolic activation at concentrations of 62.5 - 600 ug/ml. In experiment I and II, precipitation of the test item was noted with and without metabolic activation in all dose groups evaluated. Toxic effects were observed in experiment I and II in presence and absence of S9 mix at different concentrations. No biologically relevant increase of the aberration rates was noted after treatment with the test item. In both experiments with and without metabolic activation no biologically relevant increase in the frequencies of polyploid cells was found after treatment with the test item as compared to the controls. In conclusion, it can be stated that during the described in vitro chromosomal aberration test and under the experimental conditions reported, the test item did not induce structural chromosomal aberrations in the V79 Chinese hamster cell line.

There are also reliable studies of an analogue substance (final product) available to complete information on genotoxicity of the test item (intermediate). The compounds share high similaritiy in structure, are of low water solubility and have a log Pow > 8. It is, therefore acceptable to derive information on mutagenicity and clastogenicity from experimental data of this read across substance.

A detailed risk assessment and read across justification was send to the german chemical agency BAuA in 2007 with the purpose of a national substance notification (VIIA). The read across justification was accepted by the national agency.

In a GLP conform study according to OECD guideline 471 the potential of the analogue (purity: >98 weight-%) to induce gene mutations according to the plate incorporation test (experiment I) and the preincubation test (experiment II) using the Salmonella typhimurium strains TA 1535, TA 1537, TA 98, and TA 100, and the Escherichia coli strain WP2 uvrA (RCC Ltd., 2002). The assay was performed in two independent experiments both with and without liver microsomal activation. Each concentration (10 - 5000 µg/plate), including the controls, was tested in triplicate. No visible reduction of the background growth was observed up to the highest concentration. Cytotoxicity was observed in experiment I and II at the two highest concentrations in presence and absence of S9 mix. 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 metabolic activation (S9 mix). In conclusion, it can be stated that during the described mutagenicity test and under the experimental conditions reported, the test item did not induce gene mutations by base pair changes or frameshifts in the genome of the strains used. Therefore, the test substance is considered to be non-mutagenic in this Salmonella typhimurium and Escherichia coli reverse mutation assay.

 

In a GLP conform study according to the OECD guideline 476 the potential of the analogue (purity: >98.8 weight-%) to induce mutations V 79 cells was assessed. The assay was performed in two independent experiments, using two parallel cultures each. The first main experiment was performed with and without liver microsomal activation and a treatment period of 4 hours. The second experiment was performed with a treatment time of 4 hours with and 24 hours without metabolic activation. Due to limited solubility of the test item in a suitable solvent the highest concentration of the test item in the pre-experiment was 2666.6 mg/L resulting in strong test item induced precipitation at the end of the treatment period. As no cytotoxicity was observed the highest concentration applied in the main experiments was far above the limit of solubility of the test item in culture medium. No substantial and reproducible dose dependent increase of the mutation frequency was observed in the main experiments. Appropriate reference mutagens, used as positive controls, induced a distinct increase in mutant colonies and thus, showed the sensitivity of the test item and the activity of the metabolic activation system

In a GLP conform study according to OECD guideline 473, the analogue, dissolved in THF, was assessed for its potential to induce structural chromosome aberrations in V79 cells of the Chinese hamster in vitro in two independent experiments (RCC Ltd., 2002).

Since no relevant toxicity was observed in the pre-test on toxicity, the test item was tested up to a concentration exhibiting clear test item precipitation (2000 µg/ml). No toxic effects indicated by reduced mitotic indices or reduced cell numbers were observed after treatment with the test item. In both independent experiments, no biologically relevant increase in the number of cells carrying structural chromosomal aberrations was observed after treatment with the test item. In conclusion, it can be stated that under the experimental conditions reported, the test item did not induce structural chromosome aberrations as determined by the chromosome aberration test in V79 cells (Chinese hamster cell line) in vitro. Therefore, the test substance is considered to be non-clastogenic in this chromosome aberration test neither with nor without S9 mix.


Short description of key information:
Two studies according GLP and OECD guideline (471 and 473) were conducted to examine the genotoxic potential of the test item. The test material is of low solubility and preicipitates at low concentrations. Mutagenicity in bacteria or clastogenicity in mammalian cells was not observed. Information on mutagenicity and clastogenicity are completed by data of a strucutral analogue. Three GLP and OECD guideline conform in vitro assays were performed to examine the genotoxic potential of the test item. The test item was negative for mutagenicity in Ames test (OECD 471) and in an HPRT assay in V79 cells (OECD 476). The test material did also not induce clastogenicity in mammalian cells (OECD 473).

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 genotoxicity under Directive 67/548/EEC, as amended for the 30th time in Directive 2008/58/EC.

 

 

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, as amended for the second time in Directive (EC 286/2011).