Fatty acids, sunflower-oil, conjugated, maleated,reation products with diethanolamine, maleated tall-oil fatty acids and triethanolamine

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Ames: neg (BASF 2017)

HPRT: neg (BASF 2017)

MNT in vitro: neg (BASF 2018)

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Additional information

AMES

The registered substance was tested for its ability to induce point mutations in several bacterial strains, i.e. Salmonella typhimurium TA 1535, TA 100, TA 1537, TA 98 and E. coli WP2 uvrA. Doses ranged from 33 - 10000µg/plate in a standard plate incorporation assay and a pre-incubation assay. The maximum dose was selected to consider the water content of app. 50% in the supplied sample. Rat liver S9 was used as a metabolic activation system. A bacteriotoxic effect was occasionally observed depending on the strain and test conditions from about 1000 μg/plate onward. No increase in the number of his+ or trp+ revertants (factor ≥ 2: TA 100, TA 98 and E.coli WP2 uvrA or factor ≥ 3: TA 1535 and TA 1537) was observed in either assay with or without S9. Consequently, the test substance is not mutagenic in the Salmonella typhimurium/Escherichia coli reverse mutation assay in the absence and the presence of metabolic activation.

HPRT

The ability of the registered substance to induce gene mutation in mammalian cells was analyzed in an HPRT assay according to OECD guideline 476 and GLP (BASF 2017). In this study, no relevant increase in the number of mutant colonies was observed with or without S9 mix. In both experiments after 4 hours treatment with the test substance the values for the corrected mutation frequencies (0.00 – 13.27 per 10^6 cells) were close to or within the respective vehicle control values (1.01 – 8.91 per 10^6 cells).

In the 1st Experiment in the absence of S9 mix statistically significant increases in the mutant frequencies were observed at 31.25; 62.50; 250.00 and 500.00 μg/mL (5.98 – 13.27 per 10^6 cells). All values were however, within the historical control range (0.00 – 9.93 per 10^6 cells), except the value obtained at 500.00 μg/mL(13.27 per 10^6 cells). Precipitation was already observed at 125.00 μg/mL. Thus, it cannot be excluded that the obtained value was due to test substance precipitation. For this purpose this experimental part was repeated in two independent experiments designated experiment 2 and 3.

In the second experiment in the absence of S9 mix test substance precipitation was observed only at 1000.00 μg/mL, which, however, was not scorable due to strong cytotoxicity. The evaluation of all other test groups in this experiment did not confirm the data from the first experiment. The corrected mutation frequencies ranged between 0.33 – 4.59 per 10^6 cells and were lower than the values obtained from the respective vehicle control group.

In the third experiment in the absence of S9 mix test substance precipitation was observed at 600.00 μg/mL and above. At 600.00 μg/mL (the lowest precipitating test concentration) the corrected mutant frequency was 0.00 per 10^6 cells. The evaluation of all other test groups in this experiment also did not confirm the data from the first experiment. The corrected mutation frequencies ranged between 0.55 – 3.83 per 10^6 cells. These values were close to the value obtained for the corresponding vehicle control group (2.03 per 10^6 cells) and within the historical negative control data.

In the presence of S9 mix no biologically relevant increase in the mutant frequencies were observed in both experiments performed. In the 1st Experiment the values for the corrected mutation frequencies in test groups 7.81 μg/mL (9.15 per 10^6 cells) and 15.63 μg/mL (8.11 per 10^6 cells) were slightly above the range of the 95% control limit and close to or within the concurrent negative control (8.91 per 10^6 cells). Nevertheless, the values were neither statistically significant nor dose-related increased. In addition, these results were not confirmed in the 2nd Experiment.

The positive control substances induced a clear increase in mutation frequencies.

Cytotoxic effects, as indicated by clearly reduced cloning efficiencies of about or below 20% of the respective negative control values were observed in the absence of S9 mix, at 1000.00 μg/mL onward in all experiments. With S9 mix, there was a decrease in the number of colonies from about 500.00 μg/mL. With and without S9, cell morphology and attachment was adversely influenced (grade > 2) in at least the

highest applied concentration.

All in all, in the absence and the presence of metabolic activation, Efka FA 4671 is not a mutagenic substance in the HPRT locus assay using CHO cells under the experimental conditions chosen. The single value in the first experiment at 500 μg/mL, which was higher than the historical control data and well in precipitating levels could not be reproduced under identical conditions in two separate experiments, thus is considered as artifactual.

MNT in vitro

The registered substance was assessed for its potential to induce micronuclei in TK6 cells in vitro (clastogenic or aneugenic activity). Two independent experiments were carried out with and without the addition of liver S9 mix from phenobarbital- and β-naphthoflavone induced rats. According to an initial range-finding cytotoxicity test and taking into account the cytotoxicity actually found in the main experiment, the cells were exposed to up to 250 - 1000µg/mL. In the absence of a metabolic activiation system exposure lasted for 4 or 24 hours, and the cells were prepared after 24h. In the presence of S9, treatment lasted for 4h with two preparation intervals at 24 and 44 hours. 1000 cells were scored for each of the two replicates per group, i.e., 2000 cells per test group. In one Experiment (44h preparation interval with S9 mix) a non-concentration related and not statistically significant increase in the micronucleus frequencies (2.1 and 1.8% micronucleated cells) were observed at the two lower concentrations (125.00 and 250.00 µg/mL). These values were above the concurrent negative control value (1.7% micronucleated cells) and above the historical control data range. In order to confirm the relevance of the obtained data, additional cells were scored for the dose test groups and the negative control group. Thus, for these groups a total of 6000 binucleated cells were scored instead of 2000. For the highest test substance concentration (500.00 µg/mL test group) only 5000 cells could be scored due to lack of sufficient number of binucleated cells. The results from the extended evaluation of the test groups could not verify the originally obtained results. Negative and positive controls gave the expected results. Since no relevant increase in the number of micronuclei was observed after treatment with the registered substance, it is considered non-clastogenic in mammalian cells.

Justification for classification or non-classification

The registered substance did not induce gene mutations in bacteria or mammalian cells and was not-clastigenic in a mammalian micronucleus assay. Consequently, no classification for mutagenicity is required according to EU GHS, Regulation (EC) 1272/2008.