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The genotoxic potential of Fatty acids C18 unsat, reaction products with pentaethylenehexamine, acetate salts in bacterial cells in vitro was investigated in a study conducted according to OECD Test Guideline 471 (Schreib, 2012). In the study, negative results were obtained in Salmonella typhimurium strains TA98, TA100, TA102, TA1535 and TA1537, in the presence and in the absence of metabolic activation (S9 mix). Under the conditions of the study, the test article did not cause gene mutations by base pair changes or frameshifts in the genome of the tester strains tested. 

Studies on the genotoxicity of Fatty acids C18 unsat, reaction products with pentaethylenehexamine, acetate salts in mammalian cells in vitro are not available. Based on existing datasets and structural and chemical considerations, read-across from this substance to genotoxicity studies using Fatty acid C18 unsat reaction products with diethylenetriamine is appropriate to meet the REACH Annex VII-IX data requirements. Read-across is scientifically justified and also enables the REACH requirements to be adequately addressed, while avoiding unnecessary animal testing in accordance with EU Directive 86/609/EEC. The available data within the group of Amidoamine imidazoline (AAI) substances indicate that for substances based on shorter polyethyleneamines (EA), higher toxicity is observed compared to AAI based on longer EA. The forming of imidazoline itself does not seem to play a significant role. For cross-reading in general Fatty acid C18 unsat reaction product with diethylenetriamine therefore represents the worst case.

The potential for Fatty acids, C18 unsaturated, reaction products with diethylenetriamine to induce cytogenic effects in mammalian cellsin vitrowas investigated in a study conducted according to OECD Test Guideline 473 (and GLP) using the chromosome aberration test (Notox, 2009a).In the study, the test substance was studied for its effect on the number of chromosome aberrations in cultured peripheral human lymphocytes in the presence and absence of a metabolic activation system (phenobarbital and ß-naphthoflavone induced rat liver S9-mix), in two independent experiments.In the first cytogenetic assay, the test substance was tested up to 22 and 35 µg/ml for a 3 hour exposure time with a 24 hour fixation time in the absence and presence of 1.8% (v/v) S9-fraction, respectively. Appropriate toxicity was reached at these dose levels. In the second cytogenetic assay, the test substance was tested up to 12 µg/ml for a 24 hours continuous exposure time with a 24 hour fixation time and up to 10 µg/ml for a 48 hour continuous exposure time with a 48 hour fixation time in the absence of S9-mix. In the presence of S9-mix, the substance was tested up to 50 µg/ml for a 3 h exposure time with a 48 h fixation time. Appropriate toxicity was reached at these dose levels.

The number of cells with chromosome aberrations found in the solvent control cultures was within the laboratory historical control data range. Positive control chemicals, mitomycin C and cyclophosphamide, both produced a statistically significant increase in the incidence of cells with chromosome aberrations, indicating that the test conditions were adequate and that the metabolic activation system (S9 -mix) functioned properly.

Fatty acids, C18 unsaturated, reaction products with diethylenetriaminedid not induce a statistically significant or biologically relevant increase in the number of cells with chromosome aberrations in the absence and presence of S9-mix, in either of the two independently repeated experiments. It was noted that the substance increased the number of polyploid cells both in the absence and presence of S9-mix in the first cytogenetic assay and in the presence of S9- mix in the second cytogenetic assay. This may indicate that the substance has the potential to inhibit mitotic processes.

No effects ofFatty acids, C18 unsaturated, reaction products with diethylenetriamineon the number of cells with endoreduplicated chromosomes were observed both in the absence and presence of S9-mix in both cytogenetic assays. The substance was not found to be clastogenic in human lymphocytes. Based on these findings,Fatty acids C18 unsat, reaction products with pentaethylenehexamine, acetate salts is not predicted to be clastogenic in human lymphocytes.

The potential for Fatty acids, C18 unsaturated, reaction products with diethylenetriamine to induce gene mutations in mouse lymphoma L5178Y cellsin vitrowas investigated in a gene mutation study conducted according to OECD Test Guideline 476 (Notox 2009b). In the study, the substance was evaluated for its possible induction of forward mutations at the thymidine-kinase locus (TK-locus) in L5178Y mouse lymphoma cells. The test was performed in two independent experiments in the absence and presence of S9-mix.In the first experiment, the test substance was tested up to concentrations of 6.5 and 60 µg/ml in the absence and presence of 8% (v/v) S9-mix, respectively. The incubation time was 3 hours. Toxicity was observed at these dose levels in the absence and presence of S9-mix. The test substance was tested up to cytotoxic levels of 88 and 92% in the absence and presence of S9-mix, respectively. In the second experiment, the test substance was tested up to concentrations of 8.8 and 70 µg/ml, in the absence and presence of 12% (v/v) S9-mix, respectively. The incubation times were 24 hours and 3 hours for incubations in the absence and presence of S9-mix, respectively. The test substance was tested up to cytotoxic levels of 94 and 85% in the absence and presence of S9-mix, respectively.

The spontaneous mutation frequencies in the solvent-treated control cultures were between the minimum and maximum value of the historical control data range and within the acceptability criteria of this assay. Mutation frequencies in cultures treated with positive control chemicals were increased by 14-fold for MMS in the absence of S9-mix, and by 9.2- and 8.5-fold for CP in the presence of S9-mix. It was therefore concluded that the test conditions, both in the absence and presence of S9-mix, were appropriate and that the metabolic activation system (S9-mix) functioned properly.

In the absence of S9-mix,Fatty acids, C18 unsaturated, reaction products with diethylenetriaminedid not induce a significant increase in the mutation frequency in the first experiment. This result was confirmed in an independent repeat experiment with modifications in the duration of treatment time. In the presence of S9-mix, the substance did not induce a significant increase in the mutation frequency in the first experiment. This result was confirmed in an independent repeat experiment with modifications in the concentration of the S9 for metabolic activation.

Fatty acids, C18 unsaturated, reaction products with diethylenetriaminewas not found to be mutagenic in the mouse lymphoma L5178Y test system under the experimental conditions used in the study. Based on these results,Fatty acids C18 unsat, reaction products with pentaethylenehexamine, acetate salts is similarly not predicted to cause gene mutations in mammalian cellsin vitro.


Justification for selection of genetic toxicity endpoint
No study was selected since negative results were obtained in in vitro studies for all end points (weight of evidence approach)

Short description of key information:
No evidence of mutagenicity was seen for Fatty acids C18 unsat, reaction products with pentaethylenehexamine, acetate salts in a bacteria reverse mutation assay (Ames test) conducted according to OECD Test Guideline 471.

Based on existing datasets and structural and chemical considerations, read-across from Fatty acids C18 unsat, reaction products with pentaethylenehexamine, acetate salts to genotoxicity studies using Fatty acid C18 unsat reaction products with diethylenetriamine is appropriate to meet the REACH Annex VII-IX data requirements. Based on read-across to negative results obtained in in vitro mammalian cytogenicity and gene mutation studies using Fatty acid C18 unsat reaction products with diethylenetriamine, Fatty acids C18 unsat, reaction products with pentaethylenehexamine, acetate salts is not predicted to be genotoxic in mammalian cell systems in vitro.

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

Negative results were obtained for Fatty acids C18 unsat, reaction products with pentaethylenehexamine, acetate salts in anin vitrostudy conducted using bacterial cells. Based on read-across to cytogenicity and gene mutation studies using Fatty acids, C18 unsaturated, reaction products with diethylenetriamine,Fatty acids C18 unsat, reaction products with pentaethylenehexamine, acetate salts is not predicted to have any genotoxic potential in mammalian cells in vitro.The substance does not meet the criteria for classification for genetic toxicity according to Directive 67/548/EEC or Regulation No.1272/2008/EC.