1-(2,6-bis(4-tolyl)-1,3-dioxano(5,4-d)-1,3-dioxan-4-yl)ethane-1,2-diol

Endpoint summary

Additional information

Bacterial reverse mutation assay (AMES):

In this in vitro assessment of the mutagenic potential of technical GEL-ALL-MD, histidine-dependent auxotrophic mutants of Salmonella typhirium (strains TA 1535, TA 1537, TA 1538, TA 98 and TA 100) were exposed to test material diluted in acetone which was also used as a negative control.

Two independent mutation tests were performed using agar plates, in the presence and absence of liver preparations from Aroclor 1254-induced rats.

In the preliminary dose range-finding study no toxicity was observed. A top dose level of 5000 µg/plate was chosen for the subsequent mutation study. Other dose levels used in the mutation assays were: 1500, 500, 150, 50 µg/plate

The concurrent positive control compounds demonstrated the sensitivity of the assay and the metabolising activity of the metabolic activation system.

No evidence of mutagenic activity was seen at any dose level of technical GEL-ALL-MD in either mutation test. Precipitation was observed at the highest dose level tested, 5000 µg/plate

It is concluded that, when tested to the limits of solubility in acetone, technical GEL-ALL-MD was not mutagenic in either the presence or absence of metabolic activation.

Chromosome Aberration study:

There were two in-vitro chromosome aberration studies conducted, in CHL cells and human lymphocytes. Both studies showed no evidence of chromosome damaging activity in either the presence or absence of metabolic activation and showed the substance to be non-clastogenic. Both studies were conducted according to OECD guidelines (No.473) and GLP, however the study conducted by NP Wright in 1996 was chosen as the key study as it is considered slightly more reliable and adequately reported.

Key Study: NP Wright (1996):

The study was conducted to assess the potential chromosomal mutagenicity of the test material on the metaphase chromosomes of the Chinese Hamster Lung cell line according to OECD Guideline 473. 

Duplicate cultures of Chinese hamster lung (CHL) cells, treated with the test material were evaluated for chromosome aberrations at a minimum of four dose levels together with vehicle and positive controls. Four treatment conditions were used: 4 hours exposure with the addition of an induced rat liver homogenate metabolising system at 10 % in standard co-factors with cell harvest at 20 h and 30 h after exposure. Exposure in the absence of metabolic activation was continuous with cell harvest at 20 and 30 h after culture initiation.

The dose range for metaphase analysis was selected from a series of at least four dose levels chosen on the basis of the results of a preliminary toxicity test. The test material was evaluated at doubling dose levels between 241.25 and 1930 µg/mL for the with metabolic activation treatments and 3.75 to 30 µg/mL for the without metabolic activation cases. The continuous exposures were more toxic than the pulse exposures.

All vehicle (solvent) controls gave frequencies of cells with aberrations within the range expected for the CHL cell line. Both of the positive control treatments gave highly significant increases in the frequency of aberrations, indicating the satisfactory performance of the test and of the activity of the metabolising system.

The test material demonstrated no toxicologically significant, dose-related increases in the frequency of cells with aberrations either with or without metabolic activation. The test material was shown to be toxic in CHL cells in vitro in the without metabolic activation treatment groups. 

The test substance was shown to be non-clastogenic to CHL cells in vitro.

Supporting study:

When dissolved in dimethylsulphoxide and tested to the maximum achievable concentration in tissue culture medium (30 µg/ml)

, technical Gel-ALL-MD showed no evidence of chromosome damaging activity in either the presence or absence of metabolic activation.

Mouse Lymphoma assay (mammalian cell gene mutation test):

The study was conducted according to a method that was designed to assess the potential mutagenicity of the test material on the thymidine kinase, TK +/- locus of the L5178Y mouse lymphoma cell line.

L5178Y TK +/- 3.7.2c mouse lymphoma cells (heterozygous at the thymidine kinase locus) were treated with the test material at up to eight dose levels, in duplicate, together with vehicle (solvent) and positive controls. The entire experiment was repeated to confirm the result of the first experiment. Four-hour exposures were used both with and without activation in Experiment 1. In Experiment 2, the exposure time without activation was increased to 24 hours.

The dose range of test material, plated for expression of mutant colonies, was selected based on the results and observations of a preliminary toxicity test and was 3.91 to 500 µg/ml in the absence of metabolic activation, and 3.91 to 62.5 µg/ml in the presence of metabolic activation for the first experiment. For the second experiment the dose range was 3.91 to 500 µg/ml both with and without activation.

The maximum dose level used was limited by the presence and nature of the precipitate observed. In the preliminary toxicity test a precipitate of test material was observed at and above 15.63 µg/ml, increasing with intensity with increase in dose concentration until it aggregated at 1000 µg/ml. It was considered that the aggregation of the precipitate effectively reduced exposure to the cells and, therefore, this dose level was not selected for the mutagenicity tests. The vehicle (solvent) controls had acceptable mutant frequency values that that were within the normal range for the L5178Y cell line at the TK +/- locus. The positive control materials induced marked increases in the mutant frequency indicating the satisfactory performance of the test and of the activity of the metabolising system.

The test material did not induce a statistically significant or dose-related increase in the mutant frequency at any dose level, either with or without metabolic activation, in either the first or the second experiment.

The test material was considered to be non-mutagenic to L5178Y cells under the conditions of the test.

Short description of key information:
Four in-vitro studies are available on the substance:
- Reverse mutation assay 'Ames Test' using S. typhimurium and E. coli
- Two Chromosome aberration test in CHL cells and human lymphocytes
- Mouse lymphoma assay
All 3 studies gave negative results.

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

Based on negative results in the four following in-vitro studies, the substance is not classified for germ cell mutagenicity.

 

- Reverse mutation assay 'Ames Test' using S. typhimurium and E. coli:

The tested substance was considered to be non-mutagenic under the conditions of this test.

- Chromosome aberration test in CHL and human lymphocytes:

The test substance is considered to be non-clastogenic.

- Mouse lymphoma assay:

The tested substance was considered to be non-mutagenic to L5178Y cells under the conditions of the test.