Registration Dossier

Administrative data

Key value for chemical safety assessment

Additional information

The test item was assessed for its potential to induce reverse mutations in bacteria in a study according to guidelines OECD 471 and EU B.13/14. The test item did not induce a dose-related increase in the number of revertants in any of the strains in the absence and presence of S9-mix, in either of two independently repeated experiments. Positive control chemicals and solvent controls indicated the validity of the test system. In conclusion it can be stated that under the experimental conditions the test item is not mutagenic in bacteria.

The test item was assessed for its potential to induce chromosomal aberrations in peripheral human lymphocytes in vitro in a study according to guidelines OECD 473 and EU B.10. The test item did not induce a statistically significant and biologically relevant increase in the number of cells with chromosome aberrations in the absence and presence of S9-mix, in either of two independently repeated experiments. Positive control chemicals indicated the validity of the test system. In conclusion it can be stated that under the experimental conditions the test item is not clastogenic in human peripheral lymphocytes.

The test item was assessed for its potential to induce gene mutations in mammalian cells in vitro in a study according to guidelines OECD 476 and EU B.17.

Within the test in the absence of S9-mix after 3 hours treatment, the test item did not induce a significant increase in the mutation frequency in the first experiment. In the repeat experiment with a prolonged treatment period, the test item induced an up to 4.2-fold increase in the mutation frequency. The mutation frequencies were above the GEF + MF (controls) (i.e. 202 per 10EXP6 survivors (global evaluation factor + mutation frequency of controls)). Although the increases were only observed at precipitating dose levels (i.e. 100, 125 and 175 µg/mL), the increases observed were above the GEF, more than three-fold, outside the historical control data range and in a dose dependent manner. Therefore, these increases are considered relevant and the test item is considered mutagenic in the absence of S9-mix by the autorhs of this study. Moreover they state that there were significant increases in the mutation frequency of both the small and large colonies - as compared to the mean mutation frequency of the small and large colonies of the solvent controls - this would indicate increases in both chromosome aberrations and gene mutations.

However, validity of this second without metabolic activation is ambiguous, due to the precipitation observed in the three highest dose levels tested. Indeed, according to the current recommendations of the International Workshops of Genotoxicity testing (IWGT), if precipitation is noted, the highest analysed concentration should be the lowest concentration where precipitation is observed. In the experiment 2, several concentrations where precipitation was observed were tested, starting at 100 µg/mL. In order to follow the IWGT recommendations, concentrations between 66 and 100 µg/mL (the lowest concentration where precipitation is observed) should have been tested.

For this reason experiment 2 is thought to be invalid and no conclusion can be drawn concerning mutagenic properties of the test item based on this experiment.

In the presence of S9-mix, Ni(TTP)4 did not induce a significant increase in the mutation frequency in the first experiment. This result was confirmed in an independent repeat experiment with modifications in the concentration of the S9 for metabolic activation.

 Overall it is concluded that the results obtained for the test item in this mouse lymphoma L5178Y test were ambiguous under the experimental conditions described in this report. No mutagenic effect was observed at a three hour treatment period of the cells with the test item either with or without a metabolic activation system. However a positive mutagenic result was observed after a prolonged treatment period (24 h) in the absence of S9 -mix at precipitating concentrations. The biological significance of this finding is unknown.

Despite the ambigous finding in the MLA study, further testing in an in vivo system is not warranted. In general, it can be reasoned that an effect which only occurs at precipitating concentrations would not be able to achieve blood levels necessary to elicit a positive effect to due poor absorption of the test substance. Additionally, the observed mutagenic response was likely due the presence of marked cytotoxicity and was not a result of any genotoxic potential of the test substance (Section R.7.7.4.1). It is concluded that the positive finding in the MLA study was likely due to cytotoxicity and can only occur at precipitating concentration which are not biologically relevant. Additionally, the facility which produces and uses the test substance is operating under conditions which are already strict enough to deal with CMR substances and no improvement in risk reduction could be achieved regardless of the genotoxicity assessment.


Short description of key information:
The test item was assessed for its potential to induce reverse mutations in bacteria. Under the experimental conditions the test item is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.
The test item was assessed for its potential to induce chromosomal aberrations in peripheral human lymphocytes in vitro. Under the experimental conditions the test item is not clastogenic in human peripheral lymphocytes.
The test item was assessed for its potential to induce gene mutations in mammalian cells in vitro. Under the experimental conditions used the test item revealed no mutagenic activity in cultures with or without metabolic activation system at 3 h exposure time. Without metabolic activation at a prolonged treatment period (24 h) ambiguous results with respect to mutagenicity were obtained at precipitating and cytotoxic dose levels.

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

Under the conditions of these studies there was no mutagenic activity of the test item in bacteria and no clastogenic activity of the test item in human lymphocytes in vitro. However, in the mouse lymphoma L5178Y test system the test item revealed ambiguous results in the absence of metabolic activation at high precipitating and cytotoxic concentrations, whereas a clearly negative result was obtained in the presence of a metabolic activation system. Current findings are not sufficient for classification of germ cell mutagenicity according to the criteria set in the Regulation (EC) No 1272/2008.