Diethyl oxalate

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

The test substance was not mutagenic in a Bacterial Reverse Mutation assay in Salmonella typhimurium and E.coli strains (OECD 471; GLP-conform).

The test substance was non-mutagenic in the in vitro mouse lymphoma (L5178Y TK+/-) forward mutation assay (MLA) (OECD 476; GLP-conform).

The test substance was non-clastogenic in the in vitro Micronucleus Test (OECD 487; GLP-conform).

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Bacterial Reverse Mutation Test

The test was performed according to EU method B.13/14 Mutagenicity – Reverse mutation test using bacteria, which is analogous to the OECD Test Guideline No. 471. Four indicator Salmonella typhimurium strains TA 98, TA 100,  TA 1535 and TA 1537 and one indicator Escherichia coli WP2 uvrA strain were used. The test substance was suspended in DMSO and assayed in doses of 50-5000 µg/plate. Two series of experiments were performed with each strain - without metabolic activation and with a supernatant of rat liver and a mixture of cofactors (S9 -mix). In the arrangement given above, the test substance was not mutagenic in all bacterial strains with and without metabolic activation. This study is classified as acceptable and satisfies the requirements for Test Guideline OECD 471 for in vitro mutagenicity (bacterial reverse gene mutation) data.

 

Gene Mutation Test on Mouse Lymphoma Cells

The test substance was evaluated in the in vitro mouse lymphoma (L5178Y TK+/-) forward mutation assay (MLA). The genotoxic potential of the test substance was assessed in two independent assays in the absence and presence of an externally supplied metabolic activation system (S9-mix). Influence of the test substance on density and viability of L5178Y TK+/- cells was preliminary assessed in a Trypan blue exclusion assay. The cells were exposed to 8 concentrations ranging from 10 to 0.078 mM, for 3 and 24 hrs. Following 3 hrs-exposure (with or without metabolic activation system) no differences in density and viability were observed between the control and test cultures. However, about 50% decrease in the number of exposed L5178Y TK+/- cells in comparison to control group was observed at the highest concentration of the test substance tested (10 mM) after exposure for 24 hrs (without metabolic activation system). The concentrations of 5 and 2.5 mM (for 24-hrs exposure) induced only a weak decrease in the cell number (about 15%) and showed no effect on viability. In the main study, L5178Y TK+/- cells were exposed to the test substance in a concentration range of 10 – 0.313 mM for 3 hrs, with or without metabolic activation, and as well as for 24 hrs without metabolic activation. As a negative control group, RPMI 1640 culture medium was used. As positive controls methylmethanesulfonate (MMS; 10 μg/mL for the 3-hrs treatment and 5 µg/mL for the 24-hrs treatment, without metabolic activation) and benzo[a]pyrene (B[a]P; 3 μg/mL, with metabolic activation) were used in the tests. The measurements performed did not reveal any significant mutagenic effect of the test substance after exposure to the cells for 3 hrs (with or without metabolic activation system), or for 24 hrs without metabolic activation. This study is classified as acceptable and satisfies the requirements for Test Guideline OECD 476 for in vitro mutagenicity (in vitro mouse lymphoma (L5178Y TK+/-) forward mutation assay (MLA)) data.

 

Micronucleus Test in vitro on Human Lymphocytes

The test substance was tested for potential mutagenic properties on human peripheral blood lymphocytes in a micronucleus test in vitro (MNT). The test was carried out on lymphocytes isolated from two healthy, non-smoking donors. The cells were exposed to the test substance at selected concentrations for 3 (with or without exogenous metabolic activation, S9-mix) or 24 hours (without S9 -mix). In the preliminary study, lymphocyte cultures were exposed to the test substance at 5 concentrations where the highest concentration (10 mM) was applied according to OECD TG 487 adopted on 22 July 2010 “In Vitro Mammalian Cell Micronucleus”. The concentration range (10 mM, 3 mM, 1 mM, 0.3 mM, 0.1 mM) was prepared using a dilution factor of 3.16 (√10). The cells were exposed for 3 hrs, with or without S9-mix, and for 24-hrs, without S9 -mix. As a result of the study, in lymphocytes from donor 1 exposed to the test substance at concentrations 10 – 0.3 mM for 3 and 24 hours no significant decrease in the Cytokinesis-Block Proliferation Index (CBPI), Replicative Index (RI) and the percentage of the cytostasis (0%) was observed. In donor 2 only a small decrease (not exceeding 20%) was detected after exposure for 3 hrs in the presence of S9-mix, however the decrease was not concentration-dependent, i.e. the top concentration of 10 mM did not differ from the control value. In the positive (Mitomycin C, 0.15 or 0.05 µg/mL; Vinblastine, 10 or 0.6 ng/mL; Cyclophospamide, 2.5 µg/mL) and negative control cultures no significant changes in the parameters studied were observed. In the main study, the test substance tested at concentrations of 10 mM, 3 mM and 1 mM, both after 3-hrs exposure (with or without metabolic activation system) and after 24-hrs exposure (without metabolic activation system) did not induce any statistically significant increase in the micronuclei frequency in exposed cell cultures compared to the control cultures (Student’s t-test at p<0.05). The analysis of a linear trend did not indicate any significant association between the concentrations of the test substance and the MNT frequency. Reference mutagens without S9-mix, i.e. Mitomycin C and Vinblastin as well as with S9-mix, i.e. Cyclophosphamide, induced statistically significant increases in the MNT frequency comparing to the control values. These results meet the acceptance criteria of the test. The results obtained indicate that under the experimental conditions used, the test substance does not induce a mutagenic effect in the micronucleus test on human peripheral blood lymphocytes. This study is classified as acceptable and satisfies the requirements for Test Guideline OECD 487 for in vitro mutagenicity ( In vitro Mammalian Cell Micronucleus Test) data.

Justification for classification or non-classification

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, as amended for the tenth time in Regulation (EU) No 2017/776.