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Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

In vitro Gene Mutation study in Bacteria -AMES

Positive

Genetic toxicity, in vitro, OECD490 screening

Positive

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (positive)

Genetic toxicity in vivo

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

Additional information

Studies on bacteria (AMES)

The test item was examined for the ability to induce gene mutations in tester strains of Salmonella typhimurium and Escherichia coli, as measured by reversion of auxotrophic strains to prototrophy. The five tester strains TA1535, TA1537, TA98, TA100 and WP2 uvrA were used.

The test item was used as a solution in sterile water for injection and, as requested by the Sponsor, concentrations were expressed in terms of active ingredient.

Toxicity test

The test item was assayed in the toxicity test at a maximum concentration of 5000 µg/plate and at four lower concentrations spaced at approximately half-log intervals: 1580, 500, 158 and 50.0 µg/plate.

At the end of the incubation period, no precipitation of the test item was observed with any tester strain, at any concentration tested, in the absence or presence of S9 metabolism. Dose-related toxicity, particularly severe with WP2 uvrA, was observed with all tester strains, in the absence and presence of S9 metabolism. Large increases in revertant numbers were observed with TA1537, TA98 and TA100 tester strains, at all concentrations tested, even at moderate toxic levels, in the absence and presence of S9 metabolic activation.

On the basis of toxicity test results, in Main Assay, using the plate incorporation method, the test item was assayed at specific doses.

No precipitation of the test item was observed, at the end of the incubation period, with any tester strain, at any dose level, in the absence or presence of S9 metabolism.

Moderate to slight toxicity was observed with TA1535, TA1537 and TA100 tester strains, at the two or three highest concentrations tested, in the absence and presence of S9 metabolism. Dose-related increases in the number of revertant colonies were noticed with TA1537, TA98 and TA100, in the absence and presence of S9 metabolic activation, and with TA1535, only in the presence of S9 metabolism. Since a clear positive response was observed, no further experiment was undertaken.

Results

It is concluded that the test item induces reverse mutation in Salmonella typhimurium strains, both in the absence and presence of S9 metabolism, under the reported experimental conditions.

In vitro study-screening

The test item was examined for mutagenic activity by assaying for the induction of 5 trifluorothymidine resistant mutants in mouse lymphoma L5178Y cells after in vitro treatment, in the absence of S9 metabolic activation, using a fluctuation method (OECD490).

The mutant frequencies in the solvent control cultures fell within the normal range. Marked increases were obtained with the positive control treatments indicating the correct functioning of the assay system. It is concluded that the substance induced mutation at the TK locus of L5178Y mouse lymphoma cells cultured in vitro, both in the absence and presence of S9 metabolic activation, under the reported experimental conditions.

Justification for classification or non-classification

This hazard class is primarily concerned with substances that may cause mutations in the germ cells of humans that can be transmitted to the progeny.

Substance that are mutagenic in somatic cells may produce heritable effects if they, or their active metabolites, have the ability to interact with the genetic material of germ cells. Conversely, substances that do not induce mutations in somatic cell in vivo would not be expected to be germ cell mutagens.

However, the results from mutagenicity or genotoxicity tests in vitro and in mammalian somatic and germ cells in vivo are also considered in classifying substances and mixtures within this hazard class.

Category 1: substances known to induce heritable mutations or to be regarded as if they induce heritable mutations in the germ cells of humans. Substances known to induce heritable mutations in the germ cells of humans.

Category 2: substances which cause concern for humans owing to the possibility that they may induce heritable mutations in the germ cells of humans.

Classification for heritable effects in human germ cells is made on the basis of well conducted, sufficiently validated tests as In vitro mutagenicity tests such as these indicated in 3.5.2.3.8:

- in vitro mammalian chromosome aberration test;

- in vitro mammalian cell gene mutation test;

- bacterial reverse mutation tests

During the OECD471 test, the substance induces reverse mutation in Salmonella typhimurium, both in the absence and presence of S9 metabolism, under the reported experimental conditions.

In addition, a screening on in vitro mammalian cell was performed, according to OECD490.

The substance induced mutation at the TK locus of L5178Y mouse lymphoma cells cultured in vitro, both in the absence and presence of S9 metabolic activation, under the reported experimental conditions.

As conclusion, according to the CLP Regulation n.1272/2008 and the ECHA Guidance R.7a, the substance is classified as mutagenic, Muta. 2, H341: Suspected of causing genetic defects.