Hydrocarbon waxes (petroleum), oxidized, Me esters, calcium salts

Endpoint summary

Additional information

In vitro data

Gene mutation in mammalian cells:
A GLP compliant study was performed according to the standardised guidelines OECD 476, Method B.17 of Commission Regulation (EC) No. 440/2008, US EPA OPPTS 870.5300, and would be acceptable to the Japanese METI/MHLW guidelines for testing of new chemical substances.

The study was designed to assess the potential mutagenicity of the test material on the thymidine kinase, TK +/-, locus of the L5178Y mouse lymphoma cell line, both in the absence and presence of metabolic activation (S9 mix).

In Experiment 1, cells were exposed for 4 hours. In the absence of S9 mix the concentrations were 0, 9.77, 19.53, 39.06, 78.13, 156.25, 234.38, 312.5 and 468.75 µg/mL; in the presence of S9 mix the concentrations were 0, 9.77, 19.53, 39.06, 78.13, 156.25, 312.5, 468.75 and 625 µg/mL.

In Experiment 2, cells were exposed for 24 hours in the absence of S9 mix at concentrations of 0, 6.25, 12.5, 25, 50, 100, 150, 200, 250, 300 and 350 µg/mL; in the presence of S9 mix, cells were exposed for 4 hours at concentrations of 0, 50, 100, 200, 300, 350, 400, 450 and 500 µg/mL.

The test material did not induce any toxicologically significant dose-related increases in the mutant frequency at any dose level, with or without metabolic activation, in either the first or the second experiment. Therefore the test material was considered to be non-mutagenic to L5178Y cells under the conditions of the study. 

 

Bacterial reverse mutation

In a GLP compliant study performed to standardised guidelines OECD 471 and EU Method B.13/14, the mutagenic potential of the test material was assayed in an Ames Test. S. typhimurium TA 1535, TA 1537, TA 100, TA 98 and E. coli WP2 uvr A were exposed to the test material in concentrations of 0, 50, 150, 500, 1500 and 5000 µg/plate, with and without metabolic activation.

Under the conditions of the test it was determined that the test material is non-mutagenic as there was no significant increase in revertant colony counts.

Further bacterial reverse mutagenicity studies were available for assessment:
In a non-GLP compliant study performed in part according to ASTM Standard Method (E 1687-98), the Mutagenic Index (MI) was determined in a modified Ames Test. S. typhimurium TA 98 was exposed to the test material at varying concentrations, diluted in DMSO. Under the conditions of the test, the MI was calculated to be 0.16, which according to the criteria set out in the study means that the test material would not be carcinogenic in a mouse skin-painting assay and thus would be classed as non-mutagenic.

 

Chromosome aberration test in vitro

In a GLP compliant chromosome aberration study performed according to the standardised guidelines; OECD 473, EU Method B.10 and UK department of Health Committee on Mutagenicity Guidelines for the Mutagenicity Testing of Chemicals, human lymphocyte cells were exposed to the test material.

In Experiment 1, in a 4 hour exposure both with and without S9 mix cells were exposed to concentrations of 0, 156.25, 312.5, 625, 1250, 1875 and 2500 µg/mL.

In Experiment 2, cells were exposed for 4 hours in the presence of S9 mix to concentrations of 0, 39.06, 78.13, 156.25, 312.5, 625 and 1250 µg/mL; in the absence of S9 mix, cells were exposed for 24 hours at 0, 39.06, 78.13, 156.25, 312.5, 468.75 and 625 µg/mL.

The test material was determined to be non-clastogenic. Under the conditions of the test, there was not a statistically significant increase in the frequency of cell aberrations in the absence or presence of metabolic activation. Therefore the test material is considered to be non-mutagenic.

 

 

All of the available studies were conducted with the structural analogue, Hydrocarbon waxes (petroleum), oxidized. Read across is justified on the basis of similar chemical structures and analogous results from a batch of physico-chemical tests, including water solubility. The key studies were all performed in line with GLP and an accepted standardised guideline with a high level of reporting. As the studies were performed with a structural analogue, they were assigned a reliability score of 2 in accordance with the criteria for assessing data quality as outlined in Klimisch (1997) and considered suitable for assessment as an accurate reflection of the test material.

 

The available data is considered to be complete and the conclusion, non-mutagenic, was taken forward for risk assessment.

Justification for selection of genetic toxicity endpoint
All three key studies are considered as part of a weight of evidence approach.

Short description of key information:
IN VITRO DATA
Gene mutation in mammalian cells: Negative, L5178Y TK+/- with and without metabolic activation, OECD 476, EU Method B.17, EPA OPPTS 870.5300, Japanese METI/MHLW guidelines for testing of new chemical substances - Flanders 2012

Reverse mutation in bacteria: Negative (S. typhimurium strains TA 1537, TA 1538, TA 100 and TA98; E. coli strain WP2 uvr A) with and without metabolic activation, OECD 471, EU method B.13/14, Japanese METI/MHLW guidelines for testing of new chemical substances - Bowles 2003

In vitro chromosome aberration: Negative, human lymphocytes with and without metabolic activation, OECD 473, EU method B.10, UK department of Health Committee on Mutagenicity Guidelines for the Mutagenicity Testing of Chemicals - Wright & Jenkinson 2004

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

In accordance with the criteria for classification as defined in Annex I, Regulation 1272/2008, the test material does not require classification for genetic toxicity based on the overall negative response noted in the available genetic toxicity studies.