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Ames test

Sodium ethylenesulphonate was tested as a 30 % aqueous solution in the Salmonella typhimurium reverse mutation assay (Hoechst AG (b), 1988) with the strains of Salmonella typhimurium (TA1535, TA1537, TA100 and TA98) and in Escherichia coli reverse mutation assay with a tryptophan-requiring strain of Escherichia coli (WP2uvrA) according to OECD guideline 471 and GLP. Two independent mutagenicity studies were conducted (plate incorporation method), each in the absence and in the presence of a metabolising system derived from a rat liver homogenate. For both studies each bacterial strain was exposed to 6 dose levels. In the first experiment concentrations ranged from 4 to 10000 µg/plate and in the second experiment from 4 to 5000 µg/plate. Control plates without mutagen showed that the number of spontaneous revertant colonies was similiar to that described in the literature. All the positive control compounds gave the expected increase in the number of revertant colonies. The test solution proved to be not toxic to the bacterial strains. In the absence of the metabolic activation system the test compound did not show a dose dependent increase in the number of revertants in any of the bacterial strains. Also in the presence of a metabolic activation system, treatment of the cells with the 30 % Sodium ethylenesulphonate solution did not result in relevant increases in the number of revertant colonies. In conclusion the test solution, containing 30% Sodium ethylenesulphonate, did not demonstrate mutagenic potential. Under the conditions of the present bacterial reverse mutation assay the maximum test concentration of Sodium ethylenesulphonate was 3000 µg/plate.

In vitro mammalian chromosome aberration test

A test solution, containing 25 % Sodium ethylenesulphonate, was assessed to its ability to induce chromosomal aberrations in huma lymphocytes according to OECD guideline 473 and GLP (Huntingdon Life Science Ltd. (a), 1997). In two independent experiments at least 100 metaphase figures were analysed at concentrations of 1250, 2500 and 5000 µg/mL of the 25 % Sodium ethylenesulphonate solution. No toxicity was observed after treatment with the 25% Sodium ethylenesulphonate solution in any of the tests, either in the absence or the presence of S-9 mix. No significant increases in mutant frequency were observed in cultures treated with the 25% Sodium ethylenesulphonate solution in any of the tests either in the absence or the presence of S-9 mix.The positive controls induced highly significant increases in mutant frequency in all of the tests in both the absence and presence of S-9 mix thus demonstrating adequate sensitivity of the test system. In conclusion, the test solution, containing 25% Sodium ethylenesulphonate, did not induce chomosomal aberrations over background when tested up to cytotoxic concentrations. Under the conditions of the present mammalian chromosome aberraton test the maximum test concentration of Sodium ethylenesulphonate was 1250 µg/mL (5000 µg/mL of a 25 % aqueous Sodium ethylenesulphonate solution). This concentration is in the range of the maximum test concentration of 1300 µg/mL (0.01M) recommended according to the guideline.

 

In vitro mammalian cell gene mutation test

A test solution, containing 25 % Sodium ethylenesulphonate, was tested in a mammalian gene mutation test in CHO cells (HPRT assay) according to OECD guideline 476 and GLP (Huntigdon Life Science Ltd. (b), 1997). In two independent experiments concentrations of 1250, 2500 and 5000 µg/mL of the 25 % Sodium ethylenesulphonate solution were tested with and without metabolic activation (S9 -mix). Some increases of pH values and dose related increases in osmolarity were observed in all tests. No significant increases in mutant frequency were observed after treatment with 25 % Sodium ethylenesulphonate in either test in the absence and the presence of S-9 mix. Ethyl methanesulphonate and 20-Methylcholanthrene, the positive controls, induced highly significant increases in mutant frequency in both tests. It is concluded that the test solution, containing 25 % Sodium ethylenesulphonate, did not demonstrate mutagenic potential in this in vitro gene mutation assay. Under the conditions of the present mammalian gene mutation test the maximum test concentration of Sodium ethylenesulphonate was 1250 µg/mL (5000 µg/mL of a 25 % aqueous Sodium ethylenesulphonate solution). This concentration is in the range of the maximum test concentration of 1300 µg/mL (0.01M) recommended according to the guideline.

In the present three in vitro genotoxicity studies (Ames test, HPRT assay and chromosome aberration test) no mutagenic potential was indicated. Under the conditions of the mammalian chromosome aberraton and gene mutation test the maximum test concentration of Sodium ethylenesulphonate was in the range of the maximum test concentration recommended according to the guideline. In the Ames test the maximum test concentration of Sodium ethylenesulphonate was minimally lower (3000 µg/plate instead of 5000 µg/plate) than specified in the guideline. However, since the result of the Ames test was in line with the mammalian chromosome aberration and gene mutation test and overall no signs of genotoxicity were indicated, it is concluded that Sodium ethylenesulphonate will not demonstrate mutagenicity in the Ames test in concentrations up to 5000 µg/plate. Of note, in a QSAR analysis by means of the software toxtree v. 2.5.0 (Benigni/Bossa rulebase for mutagenicity and carcinogenicity) Sodium ethylenesulphonate was no potenital S.typhimurium TA 100 mutagen based on available structure attributes.

 


Justification for selection of genetic toxicity endpoint
No study was selected because all in vitro studies were of toxicological relevance.

Short description of key information:
Ames test: not mutagenic; HPRT test performed with CHO cells: not mutagenic; Chromosome aberration test performed with V79 cells: not clastogenic

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

Based on the available results Sodium ethylenesulphonate was not classified and labelled for genotoxicity according to Directive 67/548/EEC (DSD) and Regulation (EC) No 1272/2008 (CLP).