Registration Dossier

Administrative data

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

3 OECD TG GLP studies:

- OECD TG 471 (Bacterial Reverse Mutation Assay): negative

- OECD TG 476 (In Vitro Mammalian Cell Gene Mutation Test): negative

- OECD TG 487 (In Vitro Mammalian Cell Micronucleus Test): negative

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

OECD TG 471 - Bacterial Reverse Mutation Assay

A bacterial reverse mutation assay was performed equivalent to OECD guideline 471 under GLP conditions. The Salmonella typhimurium strains TA100, TA1535, TA1537, TA98 and Escherichia coli strain E. coli WP2 uvrA were incubated with the test substance at concentrations of 0.1 - 5500 µg/plate (Standard plate test, SPT) and 0.03 - 100 µg/plate (Preincubation test, PIT) without and with metabolic activation (liver S9 mix from induced rats). The PIT was carried out using a pre-incubation of 20 minutes followed by a plate incorporation assay with an expression time of 48 hours. For the SPT no pre-incubtaion step was performed. Appropriate positive controls and a vehicle control were also tested.

No precipitation of the test substance was found with and without S9 mix.

A strong bacteriotoxic effect was observed depending on the strain and test conditions from about 0.3 μg/plate onward.

A relevant increase in the number of his+ or trp+ revertants (factor ≥ 2: TA 100, TA 98 and E.coli WP2 uvrA or factor ≥ 3: TA 1535 and TA 1537) was not observed in the standard plate test or in the preincubation test without S9 mix or after the addition of a metabolizing system.

Under the experimental conditions of this study, the test substance was not mutagenic in the Salmonella typhimurium/Escherichia coli reverse mutation assay in the absence and the presence of metabolic activation.

OECD TG 476 - In Vitro Mammalian Cell Gene Mutation Test

The vitro mammalian cell gene mutation test was performed in two independent experiments, using two parallell cultures of V79 cells each, according to OECD guideline 476 under GLP conditions. The first main experiment was performed with a treatment period of 4 hours with and without microsomal activation. The second main experiment was performed with a 24 hours treatment period without microsomal activation and a 4 hours treatment period with microsomal activation. The maximum test item concentration of the pre-experiment (2208.0 μg/mL) was chosen with respect to the current OECD guideline 476 regarding the content of the test item.

No substantial and reproducible dose dependent increase of the mutation frequency was observed in the main experiments. Therefore, the test substance is considered as non-mutagenic in this HPRT assay.

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.

OECD TG 487 - In Vitro Mammalian Cell Micronucleus Test

To assess its potential to induce micronuclei in human lymphocytes in vitro in the absence and presence of metabolic activation by S9 mix an in vitro mammalian cell micronucleus test was performed according to OECD guideline 487 under GLP conditions.

Two independent experiments were performed. In Experiment I, the exposure period was 4 hours with and without S9 mix. In Experiment II, the exposure period was 20 hours without S9 mix. The cells were prepared 40 hours after start of treatment with the test item.

In each experimental group two parallel cultures were analyzed. 1000 binucleate cells per culture were evaluated for cytogenetic damage on coded slides. Dose selection of the cytogenetic experiment was performed considering the toxicity data and the occurrence of test item phase separation in accordance with OECD guideline 487.

In Experiment I in the absence and presence of S9 mix, clear cytotoxicity was observed to the highest evaluated concentration. In Experiment II in the absence of S9 mix, concentrations showing clear cytotoxicity were not evaluable for cytogenetic damage.

In the absence and presence of S9 mix, no relevant increases in the numbers of micronucleated cells were observed after treatment with the test item. Therefore, the test substance is considered to be non-mutagenic in this in vitro micronucleus test, when tested up to cytotoxic or to the highest evaluable concentration.

Appropriate mutagens were used as positive controls. They induced statistically significant increases in cells with micronuclei.

In conclusion, it can be stated that the test item was not mutagenic in the bacterial reverse mutation assay and in the HPRT assay and did not induce micronuclei as determined by the in vitro micronucleus test in human lymphocytes.

Justification for classification or non-classification

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.