Registration Dossier

Administrative data

Key value for chemical safety assessment

Additional information

Justification for grouping of substances and read-across

The PFAE fumarates (Polyfunctional Aliphatic Ester) category consists of seven members, which are either well-defined mono-constituent substances or related UVCB substances, with varying fatty alcohol chain lengths. The distinguishing feature of this category of chemicals is that its members are diester derivatives of fumaric acid (CAS 110-17-8). The alcohol moiety of the dicarboxylic esters generally falls in the C12-C22 carbon number range, including linear, even and odd numbered alcohols. 

In order to avoid the need to test every substance for every endpoint, the category concept is applied for the assessment of environmental fate, environmental toxicity and human health hazards. Thus where applicable, environmental and human health effects are predicted from adequate and reliable data for source substance(s) within the group by inter- or extrapolation to the target substances in the group (read-across approach) applying the group concept in accordance with Annex XI, Item 1.5, of Regulation (EC) No 1907/2006. Structural similarities and similarities in properties and/or activities of the source and target substances in the category are the basis of read-across.

The available studies providing information on the human health hazard assessment within the PFAE fumarates category were conducted with the category member Didodecyl fumarate (CAS 2402-58-6). This substance represents the category member with the shortest fatty alcohol side chain, and consequently with the lowest molecular weight, which is regarded as worst-case approach in terms of hazard assessment of the PFAE fumarates for the local as well as for systemic effects. Additionally, experimental data is available from the category member Di-C12-15 Alkyl Fumarate.

Furthermore, the category is supported by another polyfunctional aliphatic ester, namely Bis(2-ethylhexyl) adipate (CAS 103-23-1). This supporting chemical is used to cover toxicological endpoints, exclusively. The read across of Bis(2-ethylhexyl) adipate (CAS 103-23-1) to the PFAE fumarate category is justified due to the similar structural and physico-chemical properties, as well as their toxicological, and ecotoxicological profiles.

A detailed justification for the grouping of chemicals and read-across is provided in the technical dossier (see IUCLID Sections 7.1 and 13) and within Chapter 5.1 of the CSR.

Endpoint specific data matrix:

Overview of genetic toxicity

CAS

Chemical name

Genetic toxicity (mutagenicity) in vitro, bacteria

Genetic toxicity (cytogenicity) in vitro, mammalian cells

Genetic toxicity (mutagenicity) in vitro, mammalian cells

1

2402-58-6

Experimental result: not mutagenic

Experimental result:
not clastogenic

Experimental result:
not mutagenic

2

10341-03-4

RA: CAS 2402-58-6

RA: CAS 2402-58-6

RA: CAS 2402-58-6

3

no CAS / List No. 938-575-3

RA: CAS 2402-58-6

RA: Di-C12-15 Alkyl Fumarate

RA: CAS 2402-58-6

RA: CAS 2402-58-6

4

no CAS / Di-C12-15 Alkyl Fumarate

Experimental result: not mutagenic

--

--

5

no CAS / List No. 938-576-9

RA: CAS 2402-58-6

RA: Di-C12-15 Alkyl Fumarate

RA: CAS 2402-58-6

RA: CAS 2402-58-6

6

1187576-41-5

RA: CAS 2402-58-6

RA: Di-C12-15 Alkyl Fumarate

RA: CAS 2402-58-6

RA: CAS 2402-58-6

7

68921-53-9

RA: CAS 2402-58-6

RA: CAS 2402-58-6

RA: CAS 2402-58-6

 

Discussion:

Genetic toxicity in vitro

Gene mutation in bacteria in vitro

CAS 2402-58-6

The potential mutagenicity of Didodecyl fumarate (CAS 2402-58-6) was tested in a bacterial gene mutation assay according to OECD 471 (Woitkowiak, 2013). In two independent experiments, the S. typhimurium strains TA 1535, TA 1537, TA 98 and TA 100 and E. coli WP2 uvr A were exposed to the test substance at concentrations of 33, 100, 333, 1000, 2700 and 5400 µg/plate. Both experiments were performed in the absence or presence of a liver microsomal activation system (S9 mix) using either the plate incorporation or the pre-incubation method (20 min pre-incubation time). No cytotoxicity was noted in the treated bacteria strains up to precipitating concentrations when compared to controls. Precipitation of the test substance was observed at 333 µg/plate onward with and without S9 mix. The mean number of revertant colonies per plate was not significantly increased at any test concentration. The solvent and the positive control substances proved the validity of the study for each tester strain. Under the conditions of this assay, Didodecyl fumarate was found to be non-mutagenic in S. typhimurium strains (TA 1535, TA 1537, TA 98 and TA 100) and E. coli WP2 uvr A, both in the presence and absence of S9 mix.

Di-C12-15 Alkyl Fumarate

Di-C12-15 Alkyl Fumarate was tested for its potential to cause mutation at the histidine operon of S. typhimurium strains TA98. TA100, TA1535, TA1537 and TA1538 and at the tryptophan operon of E. coli strain WP2uvrA. The test material, dissolved in acetone, was tested for toxicity to strains TA 100 and WP2uvrA in a range finding test at a concentration ranging from 5.0 to 5000 µg/plate. The tester strains were exposed to the test material in the absence and in the presence of S9 mix. The toxicity was evaluated based on the reversion frequency, viability and integrity of the background lawn. Based on the results of the range finding test the first mutation assay was performed using concentrations of 100, 250, 500, 750 and 1000 µg/plate in the presence and absence of S9 activation. The second mutation test was performed with the preincubation method to confirm the first assay, using the same concentrations. The results of both mutation assays indicated that the test material did not induce any positive increase on the number of revertant colonies for any of the tester strains in the presence or absence of S9 mix. Under the conditions of the study, Di-C12-15 Alkyl Fumarate was negative in the S. typhimurium/E. coli plate incorporation / preincubation mutation assay.

Cytogenicity in vitro

CAS 2402-58-6

The potential cytogenicity of Didodecyl fumarate was tested in the in vitro mammalian cell micronucleus test (Bohnenberger, 2013), conducted according to OECD 487, and in compliance with GLP. Chinese hamster lung fibroblasts (V79) were treated with the test item dissolved in tetra hydro furan (THF) both in the presence and absence of a metabolic activation system (S9 mix). Two independent experiments were conducted with concentrations of 0.6-2200.0 µg/mL: in Experiment I the cells were exposed to the test substance for 4 h with and without S9 mix; in Experiment 2 exposure was conducted for 24 h without S9 mix and for 4 h with S9 mix. In every case, duplicate cultures were prepared. After fixation and staining with Giemsa, 1000 cells/culture were scored and examined for micronucleated cells. Cytotoxicity was determined by means of proliferation index. There were no statistically significant cytotoxic and genotoxic properties noted at any dose level tested up to precipitating concentrations. Appropriate solvent and positive controls were included into this test and gave the expected results: the solvent control data were within the range of the historical control data, and a significant increase in micronucleated cells treated with the positive control substances proved the sensitivity of the test. The test item Didodecyl fumarate is therefore concluded to be not genotoxic under the conditions of this test. In conclusion, Didodecyl fumarate was not clastogenic to Chinese hamster lung fibroblasts (V79) with or without exogenous metabolic activation system.

Gene mutation in mammalian cells in vitro

CAS 2402-58-6

An in vitro mammalian cell gene mutation assay (HPRT assay) with Didodecyl fumarate (CAS 2402-58-6) was performed according to OECD 476 and in compliance with GLP (Wollny, 2013). Mutations at the HPRT locus of Chinese hamster lung fibroblasts (V79) were investigated in two independent experiments at test substance concentrations of 1, 3, 9, 27 and 81 µg/mL. The cells were exposed both with and without metabolic activation (S9-mix) for 4 h each in the first experiment and for 4 and 24 h with or without S9-mix in the second experiment, respectively. The cells were allowed to express their genotype for 7-9 days, followed by a selection period of 8 days in the presence of 6-TG. The test substance did not induce a significant increase in the mutant frequency at any test substance concentration with or without metabolic activation. There was no cytotoxicity noted at any concentration tested. Test substance precipitation was noted at 27 µg/mL and above in all experimental parts; in addition small droplets were noted as phase separation. Appropriate positive and solvent controls were included into the test and gave the expected results. In conclusion, the test substance was not mutagenic to Chinese hamster lung fibroblasts (V79) in the HPRT assay with or without metabolic activation up to precipitating concentrations.

Conclusions for genetic toxicity in vitro

Data investigating genetic toxicity in vitro within the fumarate category are available for Didodecyl fumarate (CAS 2402-58-6). In summary, Didodecyl fumarate (CAS 2402-58-6) was tested negative for mutagenicity to bacteria in the Amest test, negative for cytogenicity in the in vitro mammalian cell micronucleus test and negative for mutagenicity to mammalian cells in the HPRT test. Further, a negative Ames test result is also available for Di-C12-15 Alkyl Fumarate.

Therefore, based on the available data the members of the PFAE fumarates category are considered to be non genotoxic.


Justification for selection of genetic toxicity endpoint
Hazard assessment is conducted by means of read-across. No study was selected, since all available in vitro genetic toxicity studies were negative. All available studies are adequate and reliable based on the identified similarities in structure and intrinsic properties and overall quality assessment (refer to the endpoint discussion for further details).

Short description of key information:
Based on the available data the members of the PFAE fumarates category are considered to be non genotoxic.

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

Based on read-across from category members, the available data on genetic toxicity do not meet the classification criteria according to Regulation (EC) 1272/2008 or Directive 67/548/EEC, and are therefore conclusive but not sufficient for classification.