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Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Methacrylates behave as a chemical family when studied for genotoxicity potential and, with few exceptions based on alerting chemical structures, new compounds in this family may be considered for waiver from acutal testing based on structure-activity relationships. Therefore the genotoxic behaviour of a similar chemical can be predicted with confidence by inclusion within this chemical class, thus avoiding unnessary testing. (Johannsen FR et al.(2008) Regulatory Toxicology and Pharmacology 50: 322 - 335)

There is only one in vitro mutagenicity study concerning the mutagenic potential for n-Octyl methacrylate available where n-Octyl methacrylate was negative in vitro (Ames test, Zeiger et al., (1987)). The results of genetic toxicity studies of the structurally closely related 2-Ethylhexyl methacrylate (also C8-Ester) is considered to be representative for the genetic toxicity of n-Octyl methacrylate (n-C8 -Ester).

The absence of a mutagenic potential was demonstrated for gene mutations as well as chromosome aberrations for the structurally closely related
2 -Ethylhexyl methacrylate. So we expect n-Octyl methacrylate to be none mutagenic as well. This is also supported by the in vitro Ames test with n-Octyl methacrylate.

Endpoint Conclusion: No adverse effect observed (negative)

Genetic toxicity in vivo

Description of key information

Data availability

For 2-EHMA the entire set of screening studies is available, including two gene mutation assays in bacteria, two chromosome mutation assays in hamster cells and human lymphocytes and a gene mutation test in mammalian cells. Therefore, further testing in vivo is not required.

Additional information

Data availability

For 2-EHMA the entire set of screening studies is available, including two gene mutation assays in bacteria, two chromosome mutation assays in hamster cells and human lymphocytes and a gene mutation test in mammalian cells. Therefore, further testing in vivo is not required.

In vitro gene mutation assay on bacteria

A Salmonella typhimurium reverse mutation assay was performed with 2-ethylhexyl methacrylate according to OECD Guideline 471 and GLP (Molinier, 1994). S. typhimurium strains, TA98, TA100, TA102, TA1535 and TA1537 were exposed to five concentrations of 2-ethylhexyl methacrylate in the presence and absence of a metabolic activation system using the plate incorporation and the preincubation methods. All doses of 2-ethylhexyl methacrylate, vehicle control and positive controls were plated in triplicate and incubated for 48-72 hours at 37°C. The positive controls gave expected responses. 2-ethylhexyl methacrylate was cytotoxic at >= 100 µg/plate without S9, >= 5000 µg/plate with S9 (plate incorporation assay) and >= 250 µg/plate (preincubation assay). Treatment of the S. Typhimurium strains with 2-ethylhexyl methacrylate in the presence or absence of metabolic activation did not induce any significant increases in the revertant numbers.

A second, equally valid bacterial mutation assay in S. Typhimurium strains, TA98, TA100, TA1535 and TA1537 and Escherichia coli WP2 uvr A according to OECD 471/472 also produced a negative result (Miyagawa et al., 1998). In this study the range of tested concentrations was between 10 and 5000 µg/plate without and with metabolic activation by benzoflavone/phenobarbital induced S9 using both, the plate incorporation and the preincubation protocol.

In vitro chromosomal aberration assay on mammalian cells

In an OECD guideline 473and GLP in vitro cytogenetics assay, 2-Ethylhexyl methacrylate was tested using duplicate human lymphocyte cultures from a male human donor (1997). Treatments covering a broad range of doses, separated by narrow intervals, were performed both in the absence and presence of metabolic activation by a rat liver post-mitochondrial fraction (S-9) from Aroclor 1254 induced animals. The highest dose level used, 1980 µg/mL was equivalent to a 10 mM concentration and exceeded the limit of solubility. Appropriate negative (solvent) control cultures and untreated cultures were included in the test system under each treatment condition. The proportion of cells with structural aberrations in negative control cultures fell within historical solvent control ranges. Untreated cultures were not analysed. 4-Nitroquinoline 1-oxide and cyclophosphamide were employed as positive control chemicals in the absence and presence of liver S-9 respectively and both compounds induced statistically significant increases in the proportion of cells with structural aberrations.

Treatment of cultures with 2-ethylhexyl methacrylate in the absence of S-9 resulted in frequencies of cells with aberrations which were similar to and not significantly different from those seen in concurrent negative control ranges. Numbers of cells with aberrations in all treated cultures fell within the historical negative control (normal) range at both sampling times.

Treatment of cultures with 2-ethylhexyl methacrylate in the presence of S-9 resulted in frequencies of cells with aberrations which were significantly higher than those in concurrent controls. Numbers of aberrant cells, however, exceeded the normal range in only a single replicate at the highest dose level at each sampling time. Insofar as increases at this concentration were very small and not reproduced in both replicates the observation was not considered biologically significant.

A second, equally valid assay in Chinese hamster lung (CHL/IU) cells was also reported with a negative result (Ohta et al., 1998). The CHL cells were tested at concentrations in a range of 10 to 5000 µg/ml without and with metabolic activation by benzoflavone/phenobarbital induced S9. No increase in structural or numerical aberrations was reported.

It is concluded that 2-Ethylhexyl methacrylate did not induce chromosome aberrations in cultured human peripheral blood lymphocytes and Chinese hamster lung cells when tested to its limit of toxicity in both the absence and presence of S-9.

In vitro gene mutation assay on mammalian cells

The potential of 2-Ethylhexyl methacrylate to induce gene mutations at the HPRT locus in V79 tells of the Chinese hamster was investigated in an OECD guideline 476 and GLP study (Harlan, 2008). The assay was performed in two independent experiments. The cells were exposed to the test item for 4 hours in the first experiment with and without metabolic activation. The second experiment was performed with a treatment period of 24 hours in the absence and 4 hours in the presence of metabolic activation. The maximum dose of the pre-test was 2000 µg/mL corresponding to a molar concentration of about 10 mM. The concentration range of the main experiments was limited by cytotoxic effects and was 0.1 - 16.0 (1stexperiment) and 3.8 – 60.0 (2ndexperiment) µg/ml without S9 and 62.5 – 2000 µg/ml with S9 (1stand 2ndexperiments). No substantial and reproducible dose dependent increase of the mutation frequency was observed in the main experiments up to the maximum concentration. Appropriate reference mutagens were used as positive controls and showed a distinct increase in induced mutant colonies and thus showed the sensitivity of the test system and the activity of the S9 mix. In conclusion, 2-ethylhexyl methacrylate did not induce gene mutations et the HPRT locus in V79 cells.

Summary

EHMA does have a full set of standard mutagenicity screening tests. It is negative in the bacterial gene mutation test, as well as in gene mutation and chromosome mutation tests in mammalian cells. In summary, 2-EHMA is regarded as non-mutagenic.


Short description of key information:
2-ethylhexyl methacrylate is not genotoxic in in vitro gene mutation assays in bacteria (Ames test) and mammalian cells (HGPRT assay) and in an in vitro chromosomal aberration assay in human lymphocytes and Chinese hamster lung cells.

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

According to the available negative in vitro assays and CLP criteria, no classification is warranted for germ cell mutagenicity.