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Three studies investigating the possible genetic toxicity effects of the test material have been conducted as follows:

Ames study:

Introduction.

The test method was designed to be compatible with the guidelines for bacterial mutagenicity testing published by the major Japanese Regulatory Authorities including METI, MHLW and MAFF, the OECD Guidelines for Testing of Chemicals No. 471 "Bacterial Reverse Mutation Test", Method B13/14 of Commission Regulation (EC) number 440/2008 of 30 May 2008 and the USA, EPA (TSCA) OPPTS harmonised guidelines.

Methods.

Salmonella typhimurium strains TA1535, TA1537, TA98, TA100 and Escherichia coli strain WP2uvrA were treated with the test item, Strontium Titanium Trioxide, using both the Ames plate incorporation and pre-incubation methods at five dose levels, in triplicate, both with and without the addition of a rat liver homogenate metabolising system (10% liver S9 in standard co-factors). The dose range for the first experiment was determined in a preliminary toxicity assay and was 50 to 5000 (µg/plate. The experiment was repeated on a separate day (pre-incubation method) using the same dose range as Experiment 1, fresh cultures of the bacterial strains and fresh test item formulations.

Results.

The vehicle (dimethyl sulphoxide) control plates gave counts of revertant colonies within the normal range. All of the positive controls used in the test induced marked increases in the frequency of revertant colonies, both with or without metabolic activation. Thus, the sensitivity of the assay and the efficacy of the S9-mix were validated.

The test item caused no visible reduction in the growth of the bacterial background lawn at any dose level and was, therefore, tested up to the maximum recommended dose level of 5000 (µg/plate. No test item precipitate was observed on the plates at any of the doses tested in either the presence or absence of S9-mix.

No toxicologically significant increases in the frequency of revertant colonies were recorded for any of the bacterial strains, with any dose of the test item, either with' or without metabolic activation or exposure method. A small, statistically significant increase in TA100 revertant colony frequency was observed in the presence of S9-mix at 50 (µg/plate in Experiment 1. This increase was considered to be of no biological relevance because there was no evidence of a dose-response relationship or reproducibility. Furthermore, the individual revertant counts at 50 µg/plate were within the in-house historical untreated/vehicle control range for the tester strain and the fold increase was only 1.3 times the concurrent vehicle control.

Conclusion.

The test item, Strontium Titanium Trioxide, was considered to be non-mutagenic under the conditions of this test.

Chromosome Aberration study:

Introduction.

This report describes the results of an in vitro study for the detection of structural chromosomal aberrations in cultured mammalian cells. It supplements microbial systems insofar as it identifies potential mutagens that produce chromosomal aberrations rather than gene mutations (Scott et al, 1990). The method used was designed to be compatible with that described in the OECD Guidelines for Testing of Chemicals (1997) No. 473 "Genetic Toxicology: Chromosome Aberration Test", Method B10 of Commission Regulation (EC) No. 440/2008 of 30 May 2008 and the USA EPA OPPTS guideline 40 CFR 799.9537. The study design also meets the requirements of the UK Department of Health Guidelines for Testing of Chemicals for Mutagenicity.

Methods.

Duplicate cultures of human lymphocytes, treated with the test item, were evaluated for chromosome aberrations at three dose levels, together with vehicle and positive controls. Four treatment conditions were used for the study; i.e. In Experiment 1, 4 hours in the presence of an induced rat liver homogenate metabolising system (S9), at a 2% final concentration with cell harvest after a 20-hour expression period and a 4 hours exposure in the absence of metabolic activation (S9) with a 20-hour expression period. In Experiment 2, the 4 hours exposure with addition of S9 was repeated (using a 1% final S9 concentration), whilst in the absence of metabolic activation the exposure time was increased to 24 hours.

The dose levels used in the main experiments were selected using data from the preliminary toxicity test where precipitate was the limiting factor and were as follows:

Group

Final concentration of test item (µg/mL)

4 (20)-hour without S9

28.75, 57.5, 115, 230, 460, 690

4 (20)-hour with S9 (2%)

28.75, 57.5, 115, 230, 460, 690

24-hour without S9

28.75, 57.5, 115, 230, 460, 690

24-hour with S9 (1%)

28.75, 57.5, 115, 230, 460, 690

Results.

All vehicle (solvent) controls had frequencies of cells with aberrations within the range expected for normal human lymphocytes. All the positive control items induced statistically significant increases in the frequency of cells with aberrations indicating the satisfactory performance of the test and of the activity of the metabolising system.

The test item did not induce any statistically significant increases in the frequency of cells with aberrations, in either the absence or presence of a liver enzyme metabolising system in either of two separate experiments. The maximum dose tested in the main experiments was limited by precipitate of the test item occurring on the slides and restricting the ability to accurately score metaphases.

Conclusion.

The test item was considered to be non-clastogenic to human lymphocytes in vitro.

L5178Y TK +/- Mouse Lymphoma Assay

Introduction.

The study was conducted according to a method that was designed to assess the potential mutagenicity of the test item on the thymidine kinase, TK +/-, locus of the L5178Y mouse lymphoma cell line. The method was designed to be compatible with the OECD Guidelines for Testing of Chemicals No.476 "In Vitro Mammalian Cell Gene Mutation Tests", Method B17 of Commission Regulation (EC) No. 440/2008 of 30 May 2008, the US EPA OPPTS 870.5300 Guideline, and be acceptable to the Japanese METI/MHLW guidelines for testing of new chemical substances.

Methods.

Two independent experiments were performed. In Experiment 1, L5178Y TK +/- 3.7.2c mouse lymphoma cells (heterozygous at the thymidine kinase locus) were treated with the test item at eight dose levels, in duplicate, together with vehicle (solvent) and positive controls using 4-hour exposure groups both in the absence and presence of metabolic activation (2% S9 final concentration). In Experiment 2, the cells were treated with the test item at eight dose levels using a 4-hour exposure group in the presence of metabolic activation (1% S9 final concentration) and a 24-hour exposure group in the absence of metabolic activation.

The dose range of test item was selected following the results of a preliminary toxicity test and for Experiments 1 and 2 was 28.67 to 1835 μg/mL in both the absence and presence of metabolic activation.

Results.

The maximum dose levels used in the Mutagenicity Test was the 10 mM limit dose of 1835 μg/mL. Precipitate of test item was observed at and above 28.67 μg/mL in the Mutagenicity Test. The vehicle (solvent) controls had mutant frequency values that were considered acceptable for the L5178Y cell line at the TK +/- locus. The positive control items induced marked increases in the mutant frequency indicating the satisfactory performance of the test and of the activity of the metabolising system.

The test item did not induce any toxicologically significant dose-related increases in the mutant frequency at any dose level, either with or without metabolic activation, in either the first or second experiment.

Conclusion.

The test item was considered to be non-mutagenic to L5178Y cells under the conditions of the test.


Short description of key information:
The test item, Strontium Titanium Trioxide, was considered to be non-mutagenic under the conditions of an Ames test.
The test item did not induce any statistically significant increases in the frequency of cells with chromosome aberrations in either the absence or presence of a liver enzyme metabolising system in either of two separate experiments. The test item was therefore considered to be non-clastogenic to human lymphocytes in vitro.
The test item did not induce any toxicologically significant increases in the mutant frequency at the TK +/- locus in L5178Y cells (in a L5178Y Mouse Lymphoma Assay) and is therefore considered to be non-mutagenic under the conditions of the test.

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

Based on the results of the above studies, the test material can be considered to be non-mutagenic and non-clastogenic. As such the test material can be considered to be non classified for genetic toxicity.

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