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Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information
The assessment of genotoxicity in vitro is based on valid experimental data obtained with both dioctadecyl 3,3’-thiodipropionate and didodecyl 3,3’-thiodipropionate. The analogue substance has a C12 alkyl chain instead of a C18 alkyl chain and this structural difference is considered acceptable for the assessment of genotoxicity. No mutagenicity was observed in GLP compliant studies with didodecyl 3,3’-thiodipropionate in bacteria (OECD 471, GLP) in Salmonella strains TA98, TA100, TA1535, TA1537 and TA102 and in the mouse lymphoma assay (OECD 476 of 1984). No clastogenicity was observed in a GLP compliant study with dioctadecyl 3,3’-thiodipropionate in V79 cells (OECD 473). Dioctadecyl 3,3’-thiodipropionate was not mutagenic in a screening test with Salmonella strains TA 98, TA 100 and TA 1537.Overall, dioctadecyl 3,3’-thiodipropionate is concluded to be non genotoxic in vitro. Experimental data on genotoxicity in vivo is not available.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Additional information

Additional information from genetic toxicity in vitro:

Mutatgenicity in Bacteria

The key study for mutagenicity in bacteria (Hazleton, 1992) was performed and reported according to the requirements of OECD testing guideline 471 and GLP. The test item didodecyl 3,3’-thiodipropionate is an analogue substance that differs in having two C12 alkyl chains instead of two C18 alkyl chains. The shorter alkyl chain is expected to result in slightly more favourable solubility and uptake. Considering the 1.3 fold difference in molecular weight, application of the C12 analogue results in a higher number of molecules so that overall, any hazard identified for didodecyl 3,3’-thiodipropionate is considered relevant for dioctadecyl 3,3’-thiodipropionate. Didodecyl 3,3’-thiodipropionate was assayed for mutagenicity in five histidine-requiring strains (TA98, TA100, TA1535, TA1537 and TA102) of Salmonella typhimurium, both in the absence and presence of metabolic activation by an Aroclor 1254 induced rat liver post-mitochondrial fraction (S-9), in two separate experiments.

An initial toxicity rangefinder experiment was carried out in TA100 only, using final concentrations at 8, 40, 200, 1000 and 5000 μg/plate plus a solvent and positive control. These treatments showed no signs of a toxic effect (as indicated by a thinning of the background bacterial lawn or marked reductions in revertant numbers). Precipitation of test agent was however observed at concentrations of 1000 and 5000 μg/plate. The maximum test concentration was therefore reduced to 1000 μg/plate for all subsequent treatments, as an estimate of the solubility limit in this test system. A narrowed dose range was used for all Experiment 2 treatments, to examine as closely as possible those concentrations of the test agent most likely to exhibit a mutagenic response. In addition, treatments in the presence of S-9 in this experiment also incorporated a pre-incubation step, so as to increase the range of mutagens that might be detected using this assay system. Observations of a toxic effect following these treatments probably reflected the increased exposure of the bacteria to the test agent, attained using this method. Precipitation was noted following all treatments of 500 μg/plate and above. Negative (solvent) and positive control treatments were included for all strains in both experiments. The mean numbers of revertant colonies on negative control plates all fell within acceptable ranges, and were significantly elevated by positive control treatments. No treatment of any of the tester strains, either in the absence or presence of S-9, resulted in an increase in revertant numbers sufficient to be considered as evidence of mutagenic activity.

The findings of the key study are consistent with the results of a screening test with Salmonella strains TA 98, TA 100 and TA 1537 that was performed with up to 5 mg/plate of the actual substance (Ciba-Geigy, 1989). The plate incorporation method was applied both with and without the addition of liver postmitochondrial fraction prepared from Arochlor 1254 induced rats and acetone was used as vehicle. With the limitation of the number of tester strains, the procedure and the reporting details are adequate for hazard assessment.

Mutagenicity in mammalian cells

The key study for mutagenicity in mammalian cells in vitro is was performed in mouse lymphoma L5178Y cells using the C12 analogue (Hazleton, 1992). The study was performed according to OECD testing guideline 476 (1984) and under GLP. Didodecyl 3,3’-thiodipropionate was assayed for its ability to induce mutations at the tk locus (5-trifluorothymidine resistance) in mouse lymphoma cells using a fluctuation protocol. The study consisted of a cytotoxicity range-finder followed by two independent experiments, each conducted in the in the absence and presence of metabolic activation by an Aroclor 1254 induced rat liver post-mitochondrial fraction (S-9). Following a wide range of treatments in the range-finder experiment, separated by two-fold intervals and ranging from 7.813 to 250 μg/ml (limited by solubility in tissue culture medium), cells survived all doses yielding 141.3% relative survival in the absence and 70.8% relative survival in the presence of S-9 at the top dose (250 μg/ml). Accordingly, four doses were chosen for the first experiment, separated by 2-fold intervals and ranging from 31.25 to 250 μg/ml. All doses were plated for viability and 5-trifluorothymidine resistance two days after treatment. The top dose plated yielded 102.9% and 114.4% relative survival in the absence and presence of S9. In the second experiment the same dose range was selected. The top dose plated in this experiment was again 250 /μg/ml in the absence and presence of S9, which yielded 95.8% and 101.5% relative survival respectively. Negative (solvent) and positive control treatments were included in each experiment in the absence and presence of S9. Mutant frequencies in negative control cultures fell within normal ranges, and clear increases in mutation were induced by the positive control chemicals 4-nitrotiuinoline 1-oxide without S9) and benzo(a)pyrene (with S9). Therefore the study was accepted as valid. No statistically significant increases in mutant frequency were observed in the absence or presence of S9 in both experiments.

Chromosomal Damage

The key study for clastogenictiy in mammalian cells in vitro was performed in V79 cells (CCR, 1998). The study was performed according to OECD testing guideline 473 (1983) and under GLP. The test article was formulated in culture medium (MEM). The chromosomes were prepared 18 h and 28 h after start of treatment. The treatment interval was 4 h (exp. I: without and with metabolic activation; exp. II: with metabolic activation) or 18 h and 28 h without metabolic activation (exp. II). In each experimental group two parallel cultures were set up. Per culture 100 metaphase plates were scored for structural chromosome aberrations. The highest applied concentration in the pre-test was chosen with regard to the properties of the formulation of the test article. A homogenous suspension could be prepared at 300 μg/ml in the absence of S9 mix and 285 μg/ml in the presence of S9 mix. Test article concentrations between 1 and 300 μg/ml (without S9 mix) or 1 and 285 μg/ml (with S9 mix) were chosen for the assessment of the cytotoxic potential. In the absence of S9 mix reduced cell numbers to 55.0 % of control were observed after treatment with 300 μg/ml whereas in the presence of S9 mix no toxic effects were observed. In the pre-test precipitation of the test article was observed 4 h after start of treatment at concentrations of 5 μg/ml and above in the absence of S9 mix and 10 μg/ml and above in the presence of S9 mix. In experiment I, test article concentrations within a range of 0.75 - 300 μg/ml (without S9 mix) and 0.6-20 μg/ml (with S9 mix) were applied for the investigation of the potential to induce cytogenetic damage. In experiment II, the applied concentration ranges were 0.75 - 300 μg/ml in the absence of S9 mix and 1.0-40 μg/ml in the presence of S9 mix. In the absence and the presence of S9 mix, in both experiments, no reduction of the mitotic index or the cell number was observed, except in the presence of S9 mix in experiment II at interval 28 h after treatment with 40 μg/ml reduced cell numbers were observed. In both independent experiments, neither a significant nor a biologically relevant increase in the number of cells carrying structural chromosomal aberrations was observed after treatment with the test article. In addition, no increase in the frequencies of polyploid metaphases was found after treatment with the test article as compared to the frequencies of the controls. Appropriate mutagens were used as positive controls. They induced statistically significant increases (p < 0.05) in cells with structural chromosome aberrations.


Justification for selection of genetic toxicity endpoint
GLP-compliant guideline study

Justification for classification or non-classification

Dangerous Substance Directive (67/548/EEC)

The available studies are considered reliable and suitable for classification purposes under 67/548/EEC. As a result the substance is not considered to be classified for genotoxicity under Directive 67/548/EEC,.

  

Classification, Labeling, and Packaging Regulation (EC) No. 1272/2008

The available experimental test data are reliable and suitable for classification purposes under Regulation 1272/2008. As a result the substance is not considered to be classified for genotoxicity under Regulation (EC) No. 1272/2008.