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Key value for chemical safety assessment

Additional information

Data on genetic toxicity of Amides, C16-C18 (even numbered) is available from a bacterial reverse mutation assay (Ames test) addressing the mutagenicity of the test material in the Salmonella typhimurium strains TA98, TA100, TA1535, TA1537 and TA1538 in the absence and presence of metabolic activation (Jones, 1989). As metabolic activation system rat liver S9-mix, prepared from Aroclor 1254-treated rats, was utilised. Although the strain selection is incomplete according to today's standards, as the S. typhimurium strain TA102 or an equivalent is missing, which serve to detect cross-linking agents, it was appropriate at the time of the study. The substance was tested up to the guideline limit concentration of 5 mg/plate. No cytotoxicity was observed up to the highest tested concentration, and no significant increases in revertant numbers occurred, either in absence or in presence of metabolic activation. Therefore, Amides, C16-C18 (even numbered) is not considered as mutagenic in bacteria.

No further data on genetic toxicity was available for Amides, C16-C18 (even numbered). Therefore, surrogate data from the structurally related substance erucamide (CAS 112-84-5) was included as read-across for regulatory purposes. For this substance a chromosomal aberration test and a mouse lymphoma study, both recently conducted in 2010, are available.

In the in vitro chromosomal aberration test, performed according to OECD Guideline 473, Chinese Hamster V79 cells were treated with erucamide (CAS 112-84-5) in the absence and presence of a metabolic activation system consisting of rat liver S9-mix prepared from phenobarbital / β-naphtoflavone-treated rats (Hall, 2010). Two separate experiments were performed, in the first one, which was originally regarded as preliminary test, the cells were exposed for 4 hours to concentrations ranging from 4.9 to 1250 µg/mL in the absence and presence of metabolic activation, in the second experiment the cells were exposed to 2.3 to 450 µg/mL for 18 hours in the absence and to 4.7 to 300 µg/mL for 4 hours in the presence of S9-mix. Structural chromosomal aberrations were assessed by visual inspection.

In the first experiment precipitation was observed after 4 hours incubation without S9 starting at a concentration of 312.5 µg/mL and with S9 after incubation with 625 µg/mL. Therefore, concentrations were adjusted accordingly in the second experiment. In this experiment cytotoxicity was observed at 300 µg/mL after 18 hours incubation without S9; precipitation was noted at 300 µg/mL after 4 hours incubation with S9. No statistically significant or biologically relevant increase in the number of cells carrying structural chromosomal aberrations was observed in any of the experiments. The aberration rates were close to the values of the solvent control values and within the range of the historical control data. No increase in polyploid metaphases was observed. Therefore, the test substance erucamide was considered not to induce structural chromosomal or numerical aberrations under the conditions of the study. Considering structural and physical chemical similarities to erucamide it is justified to assume that Amides, C16-C18 (even numbered) does not induce an increase in chromosomal aberrations in cultured mammalian cells, either.

The in vitro mutagenicity test in mammalian cells with erucamide (CAS 112-84-5) was performed with mouse lymphoma L5187Y cells, according to OECD Guideline 476 in the absence and presence of a metabolic activation system consisting of rat liver S9-mix prepared from phenobarbital / β-naphtoflavone-treated rats (Wollny, 2010). Here, the ability of the test substance to induce mutations in the thymidine kinase gene in cultured mammalian cells was investigated in two independent experiments. In the first one, the preliminary test, the cells were exposed to test substance concentrations of 9.4 to 300 µg/mL for 4 hours with and without metabolic activation; in the second experiment the cells were exposed to 4.7 to 150 µg/mL for 4 hours with and for 24 hours without metabolic activation. There was no cytotoxic effect observed up to the maximum concentration in presence and absence of metabolic activation following exposure for 4 and 24 hours, respectively. No significant and reproducible dose dependent increase of mutation frequency was observed in both experiments, and no significant dose dependent trend of the mutation frequency was determined in any of the experimental groups. Under the conditions of the experiment erucamide did not induce mutations in the thymidine kinase locus in the L5178Y cell line. Therefore, it is reasonable and justified to anticipate that Amides, C16-C18 (even numbered) does not induce gene mutations in cultured mammalian cells in vitro either, due to the structural and physical chemical similarities to erucamide.

As there is reliable read-across data for a structurally related substance available addressing clastogenicity and mutagenicity in cultured mammalian cells, the conduction of corresponding studies for Amides, C16-C18 (even numbered) would not provide new information relevant for assessment. Therefore, no further studies for Amides, C16-C18 (even numbered) are proposed.

According to Regulation (EC) No 1907/2006, Annex IX, 8.4, column 2, testing for genetic toxicity in vivo is not indicated as the available data for the substance itself and a structurally related substance did not demonstrate any genotoxic activity in bacteria or mammalian cells in vitro.


Short description of key information:
Bacterial reverse mutation assay (Ames), OECD 471: negative
RA-S CAS 112-84-5, Chromosome aberration in mammalian cells in vitro, OECD 473: negative
RA-S CAS 112-84-5, Mammalian cell gene mutation test in vitro (Mouse Lymphoma), OECD 476: negative

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

According to the criteria of EU Directive 67/548/EEC and the criteria of Regulation (EC) No 1272/2008, Amides, C16-C18 (even numbered) does not have to be classified for genetic toxicity.