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Key value for chemical safety assessment

Additional information

Gene mutation in bacteria

The genetic toxicity of the substance in bacteria was evaluated in a study performed according to OECD Guideline 471 (2002-0234-DGM). S. typhimurium strains TA 1535, TA 1537, TA 98, TA 100 and TA 102 were treated with the substance dissolved in distilled water at concentrations of 313, 625, 1250, 2500 and 5000 µg/plate with and without metabolic activation by S9 mix from livers of rats treated with Phenobarbital and beta-Naphtoflavone. No increase in revertants compared to the vehicle control was observed in any of the strains at all test concentrations using the plate incubation method. Toxicity was noted at the highest test concentration in some tester strains. Based on the test results the substance is not mutagenic in bacteria.

 

Gene mutation in mammalian cells

The substance was tested in an in vitro mammalian cell gene mutation assay under GLP according to OECD Guideline 476 (2014-0248-DGM). In three independent experiments L5178Y mouse lymphoma cells (genetic marker: thymidine kinase) were treated with 6.25, 12.5, 25, 50, 75, 100, 150, 200, 300, 400 and 600 µg/mL without or with metabolic activation by Phenobarbital and beta-Naphtoflavone -induced rat liver S9 mix, respectively. The exposure duration was 3 and 24 h in experiments without S9 mix and 3 h in experiments with S9 mix. The mutation frequency of the cultures treated with the substance in the presence and in the absence of S9 mix was within the range of the negative control values, whereas the positive controls caused a pronounced increase in the mutation frequency. Cytotoxicity was noted starting from the concentration of 400 µg/mL in the absence of metabolic activation (3 h) and 100 µg/mL (24 h) and starting from 600 µg/mL in the presence of metabolic activation (3 h). No change was noted in the ratio of small to large mutant colonies.

Thus, based on the test result the substance was not mutagenic in the mouse lymphoma forward mutation assay and did not exhibit clastogenic potential at the concentration-range investigated.

 

Chromosome aberration:

N-Butyl-TAD was tested for itspotential to induce micronucleus formation in the in vitro micronucleus test with manual scoring on microscope slides. The test system used isolated human lymphocytes as the cell type. The substance was applied to the test system under three treatment schedules in accordance with the OECD Guideline 487 (2010): treatment for 3 hours in both the absence and presence of an in vitro activation system based on S9 fraction obtained from Aroclor 1254-induced rat liver (S9 mix), and a continuous treatment in the absence of S9 mix. Lymphocytes isolated from fresh whole human blood and meeting the requirements outlined in OECD Guideline 487 were used in the study. In all treatments, the solvent used was sterile distilled water. The final dose ranges tested from which micronuclei were analysed, were 279 -2120 µg/L (3h+S9 treatment schedule) , 1400 to 2124 µg/mL (3h-S9 treatment schedule) and 200 to 1000 µg/mL (continuous treatment schedule, 24h-S9). In 3h-S9 and continuous treatment schedules, 55% (±5%) cytotoxicity was achieved at 2124 µg/mL and 700 µg/mL, respectively. No cytotoxicity was observed at the highest test concentration of 2120 µg/mL in the 3h+S9 treatment.

It was concluded that the substance did not induce the formation of micronuclei in human lymphocytes in the absence and presence of S9, and under the test conditions used.


Short description of key information:
In vitro:
Ames test: negative with/without metabolic activation
Mouse lymphoma assay: negative with/without metabolic activation
Micronucleus test: negative with/without metabolic activation

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

Based on the currently available experimental in vitro data, non-classification of the substance for this endpoint is adequate in accordance to the CLP Regulation (EC) No 1272/2008.