Registration Dossier

Administrative data

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Based on the results of in vitro read across studies, the test substance is considered to be non-genotoxic.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Study 1. Anin vitrobacterial reverse mutation assay was conducted to determine the genetic toxicity of the read across substance, mono- and di- C12 PSE, K+, according to OECD Guideline 471, in compliance with GLP. The potential to induce gene mutations in the plate incorporation test (experiment I) and the pre-incubation test (experiment II) usingSalmonella typhimuriumstrains TA 1535, TA 1537, TA 98, and TA 100, and theEscherichia colistrain WP2 uvrA. The assay was performed in two independent experiments both with and without rat liver microsomal activation (S-9 mix). Each concentration and the controls were tested in triplicate. The test substance was tested at concentrations ranging from 0 to 5000 µg/plate. The plates incubated with the test substance showed reduced background growth. No substantial increase in revertant colony numbers of any of the five tester strains was observed following treatment with the test substance at any dose level, neither in the presence nor absence of metabolic activation (S9 mix). There was also no tendency of higher mutation rates with increasing concentrations in the range below the generally acknowledged border of biological relevance. Appropriate reference mutagens were used as positive controls. They showed a distinct increase of induced revertant colonies. Based on the results of the read across study, the test substance is considered to be non-mutagenic inS. typhymuriumandE. coli(Sokolowski, 2008).

Study 2. Anin vitrobacterial reverse mutation assay was conducted to determine thein vitrogenetic toxicity of the read across substance, mono- and di- C12 -18 PSE, Na+, according to OECD Guideline 471 and EU Method B.13/14 (mutagenicity - reverse mutation test using bacteria), in compliance with GLP. Dose range finding tests as well as direct plate and pre-incubation assays both in the absence and presence of S9-mix were performed.Salmonella typhimuriumstrains TA1535, TA1537, TA100 and TA98 and Escherichia coli strain WP2uvrA were exposed to the test substance at concentration levels of 0.55, 1.7, 5.4, 17, 52, 164, 512, 1600 and 5000 µg/plate, to negative or positive control substances for 48 ± 4h (plus a pre-incubation of 30 min if needed). In the dose range finding study, the test substance was initially tested up to concentrations of 5000 µg/plate in the tester strains TA100 and WP2uvrA in the direct plate assay. The test substance precipitated on the plates at the dose level of 5000 μg/plate. Cytotoxicity, as evidenced by a decrease in the number of revertants was observed in tester strain TA100 in the absence and presence of S9-mix. In tester strain WP2uvrA, no toxicity was observed at any of the dose levels tested. In the first mutation experiment, the test substance was tested up to concentrations of 1600 and 5000 µg/plate (absence and presence of S9-mix, respectively) in the tester strains TA1535, TA1537 and TA98. The test substance precipitated on the plates at the dose level of 5000 μg/plate. Cytotoxicity, as evidenced by a decrease in the number of revertants, reduction of the bacterial background lawn and/or the presence of microcolonies, was observed in all tester strains in the absence and presence of S9-mix. Since the test substance was severely cytotoxic in the first mutation experiment, an additional dose range finding test was performed with strains TA100 and WP2uvrA, both with and without S9-mix according to the pre-incubation method. In this dose range finding study, the test substance was initially tested up to concentrations of 512 and 5000 µg/plate in the tester strains TA100 and WP2uvrA, respectively. The test substance precipitated on the plates at dose levels of 1600 and 5000 μg/plate. Since the test substance precipitated heavily on the plates at the concentration of 5000 μg/plate, the number of revertants of this dose level could not be determined. Cytotoxicity was observed in tester strain TA100 in the absence and presence of S9-mix. In tester strain WP2uvrA, no toxicity was observed up to the dose level of 1600 μg/plate. In the second mutation experiment, the test substance was tested up to concentrations of 164 and 512 µg/plate (absence and presence of S9-mix, respectively) in the tester strains TA1535, TA1537 and TA98 in the pre-incubation assay. The test substance did not precipitate on the plates at this dose level. Cytotoxicity was observed in all three tester strains in the absence and presence of S9-mix. The test substance did not induce a significant dose-related increase in the number of revertant (His+) colonies in each of the four tester strains (TA1535, TA1537, TA98 and TA100) and in the number of revertant (Trp+) colonies in tester strain WP2uvrA both in the absence and presence of S9-metabolic activation. These results were confirmed in an independent repeated experiment. The negative and strain-specific positive control values were within the laboratory historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly. Based on the results of the read across study, the test substance is considered to be non-mutagenic in bacteria (Verspeek-Rip, 2006).

Study 3. Anin vitrochromosomal aberration assay was conducted to determine the genetic toxicity of the read across substance, mono- and di- C12 PSE, K+, according to OECD Guideline 473, EU Method B.10 and EPA OPPTS 870.5375 (in vitrocytogenicity / chromosome aberration test in mammalian cells), in compliance with GLP. Chinese hamster lung fibroblasts (V79 cells; 1.0 - 5.0E04 cells) were exposed to the test substance at concentrations ranging from 0 to 1800 with or without metabolic activation (liver S9 of Wistar phenobarbital and ß-naphthoflavone-induced rat liver S9 mix). The chromosomes were prepared 20 h after start of treatment with the test substance. The treatment interval was 4 h with and without metabolic activation in experiment I. In experiment II, the treatment interval was 4 h with and 20 h without metabolic activation. In both experiments, no biologically relevant increase of the aberration rates was noted after treatment with the test substance with and without metabolic activation. The aberration rates of all dose groups treated with the test substance were within the historical control data of the negative control. In the experiments I and II with and without metabolic activation no biologically relevant increase in the frequencies of polyploid cells was found after treatment with the test substance as compared to the controls. Toxic effects of the test substance were observed in experiment I without metabolic activation at concentrations of 500 µg/mL and higher, with metabolic activation at concentrations of 1000 µg/mL and higher. In experiment II without metabolic activation (long time exposure), toxic effects of the test substance were observed at concentrations of 62.5 µg/mL and higher, with metabolic activation, at concentrations of 1600 µg/mL and higher. EMS (400 and 900 µg/mL) and CPA (0.83 µg/mL) were used as positive controls and induced distinct and biologically relevant increases in cells with structural chromosomal aberrations. The positive controls induced the appropriate responses. There was no evidence of test substance-induced over background. Precipitation of the test substance was observed with and without metabolic activation in both experiments. Based on the results of the read across study, the test substance is considered to be non-clastogenic in V79 cells (Oppong-Nketiah, 2012).

Study 4. Anin vitromammalian cell gene mutation assay was conducted to determine the genetic toxicity of the read across substance, mono- and di- C12 PSE, K+, according to OECD Guideline , EU Method B. and EPA OPPTS 870.5300 (in vitrogene mutation test in mammalian cells), in compliance with GLP. Chinese hamster lung fibroblasts V79 cells were exposed to the test substance suspended in cell culture medium (MEM + 0% FBS 4h treatment; MEM + 10% FBS 20h treatment) at concentrations of 50, 100, 250, 500, 750, 1000, 1250, 1500 and 2000 µg/mL (without metabolic activation, Experiment I); 50, 100, 250, 500, 750, 1000, 1250 and 1500 µg/mL (with metabolic activation, Experiment I), 7.5, 10, 25, 50, 75, 100, 125, 150 and 175 µg/mL (without metabolic activation, Experiment II), and 150, 200, 300, 600, 800, 1000, 1200 and 1400 µg/mL (with metabolic activation, Experiment II). The test substance was tested up to cytotoxic concentrations. Metabolic activation consisted in liver S9 of Wistar Phenobarbital and ß-Naphthoflavone-induced rat liver S9 mix. Biologically relevant growth inhibition was observed in experiment I and II with and without metabolic activation. In experiment I without metabolic activation, the relative growth was 24.6% for the highest concentration evaluated (2000 µg/mL). The highest biologically relevant concentration evaluated with metabolic activation was 1500 µg/mL with a relative growth of 23.9%. In experiment II without metabolic activation, the relative growth was 15.7% for the highest concentration (175 µg/mL) evaluated. The highest concentration evaluated with metabolic activation was 1400 µg/mL with a relative growth of 20.0%. In both the experiments no biologically relevant increase of mutants was found after treatment with the test substance (with and without metabolic activation). No dose response was observed. EMS and DBA were used as positive controls and showed distinct and biologically relevant effects in mutation frequency. In conclusion, based on the results of the read across study, the test substance can considered to be non-mutagenic in the HPRT locus using V79 cells of the Chinese Hamster (Wallner, 2012).

Justification for classification or non-classification

Based on the results of read across studies, the test substance does not meet the criteria for classification for this endpoint according to CLP (Regulation 1272/2008/EC) criteria.


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