Hexadecyl acetate

Endpoint summary

Additional information

Mammalian Cell Gene Mutation 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 in alignment with the Japanese MITI/MHW guidelines for testing of new chemical substances.

Method

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 (acetone) and positive controls using 4-hour exposure groups both in the absence and presence of metabolic activation (2% S9). 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) and a 24 hour exposure group in the absence of metabolic activation. The dose range of test item used in the main test was selected following the results of a preliminary toxicity test. The maximum dose level used in the main test was limited by test item induced toxicity.

Results

The maximum dose levels used in the Mutagenicity Test were limited by test item-induced toxicity and the onset of precipitate. Overall a precipitate of the test item was observed at and above 67.5 µg/mL. The vehicle controls (acetone) had mutant frequency values that were considered acceptable for the L5178Y cell line at the TK +/— locus. The positive control treatment induced marked increases in the mutant frequency indicating the satisfactory performance of the test and of the activity of the metabolizing system. The test item did not induce any toxicologically significant increases in the mutant frequency at any of the dose levels, either with or without metabolic activation, in either the first or the second experiment.

Conclusion

The test item did not induce any toxicologically significant increases in the mutant frequency at the TK +/- locus in L5178Y cells.

Mammalian Cell Chromosome Aberration Assay

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.

Methods

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

Results

All vehicle (acetone) 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 that the sensitivity of the assay and the efficacy of the S9-mix were validated. The test item was slightly toxic and exposure was limited by the solubility of the test substance. The test substance did not induce any statistically significant increases in the frequency of cells with aberrations, in either of two separate experiments.

Conclusion.

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

In vitro gene mutation study in bacteria

An in vitro gene mutation study in bacteria is not available for the test item, is predicted to be negative, and testing is considered unnecessary for a substance that isused exclusively as a cosmetic ingredient in an industrial environment, has been declared safe for use in cosmetics by the Cosmetic Ingredient Review Expert Panel, and for which other relevant results are available (negative chromosome aberration test and negative mouse lymphoma assay). The non-mutagenic prediction for the test item is supported by literature in which the parent alcohol and acid were both reported to give negative results in the Ames test (Blevins, RD & Taylor, DE (1982) Mutagenicity screening of twenty‐five cosmetic ingredients with the salmonella/microsome test, Journal of Environmental Science and Health, Part A: Environmental Science and Engineering: Toxic/Hazardous Substances and Environmental Engineering, 17:2, 217-239; Morita T, Takeda K & Okumura K (1990) Evaluation of clastogenicity of formic acid, acetic acid and lactic acid on cultured mammalian cells, Mutation Research, 240: 195-202).

Justification for selection of genetic toxicity endpoint
Two in vitro mammalian cell assays (gene mutation and chromosome aberration) were both negative for genotoxicity. Furthermore, the parent alcohol and acid are both reported in literature to give negative results during in vitro gene mutation studies in bacteria.

Short description of key information:
Two Klimisch grade 1, GLP compliant mammalian cell assays performed according to OECD 476 (gene mutation assay) and OECD 473 (chromosome aberration test).

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

Harmonized classification:

The substance has no harmonized classification for mutagenicity according to the Regulation (EC) No. 1272/2008.

Self classification:

Based on the available data, no additional classification for mutagenicity is proposed according to the Annex VI of the Regulation (EC) No. 1272/2008 (CLP).