Registration Dossier

Administrative data

Key value for chemical safety assessment

Additional information

Justification for grouping of substances and read-across

There are no sufficient data available on the genotoxic properties of Multi constituent ester of pentaerythritol 2-ethylhexanoate. In order to fulfil the standard information requirements set out in Annex VII-IX in accordance with Annex XI, 1.5, of Regulation (EC) No 1907/2006, read-across from structurally related substances was conducted.

In accordance with Article 13 (1) of Regulation (EC) No 1907/2006, "information on intrinsic properties of substances may be generated by means other than tests, provided that the conditions set out in Annex XI are met.” In particular for human toxicity, information shall be generated whenever possible by means other than vertebrate animal tests, which includes the use of information from structurally related substances (grouping or read-across).

Having regard to the general rules for grouping of substances and read-across approach laid down in Annex XI, 1.5, of Regulation (EC) No 1907/2006, whereby physicochemical, toxicological and ecotoxicological properties may be predicted from data for reference substance(s) by interpolation to other substances on the basis of structural similarity, Hexanoic acid, 2-ethyl-, 2,2-bis [ [(2-ethyl-1-oxohexyl)oxy] methyl] -1,3-propanediyl ester, and 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate are selected as source substances for assessment.

Genetic toxicity

CAS

-- (a)

7299-99-2 (b)

28510-23-8 (b)

Chemical name

Multi constituent ester of pentaerythritol 2-ethylhexanoate

Hexanoic acid, 2-ethyl-, 2,2-bis [ [(2-ethyl-1-oxohexyl)oxy] methyl] -1,3-propanediyl ester

2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate

MW

388.5 - 640.9 g/mol

640.9 g/mol

356.5 g/mol

Bacterial mutagenicity

RA: CAS 7299-99-2; CAS 28510-23-8

Experimental result: negative

Experimental result: negative

Mammalian cytogenicity

RA: CAS 7299-99-2; CAS 28510-23-8

Experimental result: negative

Experimental result: negative

Mammalian mutagenicity

RA: CAS 28510-23-8

--

Experimental result: negative

(a) The substance subject to the REACh Phase-in registration deadline of 31 May 2013 is indicated in bold font. Only for this substance a full set of experimental results and/or read-across is given.

(b) Reference (read-across) substances are indicated in normal font. Lack of data for a given endpoint is indicated by “--“.

The above mentioned substances are considered to be similar on the basis of the structural similar properties and/or activities. The available endpoint information is used to predict the same endpoint for Multi constituent ester of pentaerythritol 2-ethylhexanoate.

A detailed analogue approach justification is provided in the technical dossier (see IUCLID Section 13).

Discussion

Genetic toxicity in vitro

Ames test

Genetic toxicity in bacteria was analysed in a GLP study performed according to OECD guideline 471 (Masumori, 2005). Bacteria strains S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2 uvrA were treated with concentrations of 78.1, 156, 313, 625, 1250, 2500 μg/plate (main study; precipitation at ≥ 1250 µg/plate) Hexanoic acid, 2-ethyl-, 2,2-bis [ [(2-ethyl-1-oxohexyl)oxy] methyl] -1,3-propanediyl ester with and without metabolic activation by a cofactor supplemented post-mitochondrial fraction (S9 mix), prepared from the livers of rats treated with phenobarbital and 5,6-benzoflavone, respectively. As no increase in revertants was noted in all strains at all concentration, the test substance was not mutagenic under the conditions applied. Precipitation was observed at concentrations of ≥ 1250 µg/plate at the end of the exposure period.

In another Ames test performed in compliance with GLP and according to OECD guideline 471, the structural analogue 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate was tested in S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102 (Thompson, 1991). In the main experiment bacteria were treated with concentrations of 50, 150, 500, 1500 and 5000 µg/plate with and without metabolic activation by a cofactor supplemented post-mitochondrial fraction (S9 mix). No cytotoxicity was observed and precipitation was found at ≥ 1500 µg/plate. No increase in revertants was found. Thus, under the conditions of the experiment 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate was not mutagenic.

Chromosome aberration assay

In an in vitro chromosome aberration assay, Hexanoic acid, 2-ethyl-, 2,2-bis [ [(2-ethyl-1-oxohexyl)oxy] methyl] -1,3-propanediyl ester was tested in Chinese Hamster Lung cells in accordance to OECD guideline 473 (Masumori, 2005). Concentrations of 1250, 2500, 5000 μg/mL were applied to the cultured cells (main test) with and without metabolic activation by a rat liver S9-mix. Both in the absence and presence of S9-mix Hexanoic acid, 2-ethyl-, 2,2-bis [ [(2-ethyl-1-oxohexyl)oxy] methyl] -1,3-propanediyl ester did not induce a statistically significant or biologically relevant increase in the number of cells with chromosome aberrations in two independent experiments. No cytotoxicity was observed, whereas precipitation was found at concentrations including and higher than 625 µg/mL. Thus, Hexanoic acid, 2-ethyl-, 2,2-bis [ [(2-ethyl-1-oxohexyl)oxy] methyl] -1,3-propanediyl ester did not disturb mitotic processes and cell cycle progression and did not induce numerical chromosome aberrations under the experimental conditions described.

The potential of 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate to induce chromosomal damage in mammalian cells was evaluated in an in vitro mammalian chromosome aberration test performed according to GLP and OECD guideline 473, and EU method B.10 (Buskens, 2010). In a dose range finding study, properly maintained cultured peripheral human lymphocytes were treated with doses of 3, 10, 33, 100 and 333 µg/mL with and without metabolic activation for 3 h and with doses of 3, 10, 33, 100, 333, 1000 and 3333 µg/mL with and without metabolic activation for 24 and 48 h, respectively. Since the substance precipitated at 333 µg/plate and higher, concentrations of 3, 10, 33, 100 and 333 µg/mL were used for a 3 h treatment in the first cytogenetic assay, with and without metabolic activation by a cofactor supplemented post-mitochondrial fraction (S9 mix). A second cytogenetic assay was performed, where the cells were treated for 3 h with 50, 100 and 350 µg/mL with metabolic activation and for 24 and 48 h with doses of 5, 10, 50, 150 and 200 µg/mL only without metabolic activation, respectively. Additionally, a repeat of the 48 h treatment was performed with doses of 10, 30, 50, 60, 70, 80, 90 and 100 µg/mL without metabolic activation. Ethanol was the vehicle in all experiments. After the treatment, the cells were treated with colchicine and stained for analysis, where 1000 cells per dose were analysed. Cytotoxicity was observed starting at the dose of 100 µg/mL at the 48 h exposure without metabolic activation. However, no increase in number of aberrant cells with or without gaps was found for any concentration and any exposure time used. Thus, 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate did not induce chromosome aberration in mammalian cells.

 

Mouse Lymphoma Assay

Gene mutation in mammalian cells is assessed in a study with the structural analogue 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate, which was tested in a GLP study performed according to OECD guideline 476 (Verbaan, 2010). Mouse lymphoma L5178Y cells were treated with concentrations of 0.03, 0.1, 0.3, 1, 3, 10, 33, 100 µg/mL (with (8% v/v S9-mix) and without metabolic activation) for 3 h. Metabolic activation was performed with rat liver S9-mix (S9-fraction from Wistar rats induced by a combination of phenobarbital and ß-naphthoflavone). In the second experiment the cells were incubated with 0.03, 0.1, 0.3, 1, 3, 10, 33, 100 µg/ml (with metabolic activation (12%, v/v) and 0.3, 1, 3, 10, 33, 100, 200, 300, 400, 500 and 600 µg/ml (without metabolic activation) for 3 and 24 h, respectively. As result, no significant increase in the mutation frequency in the presence or absence of S9-mix was found, independent of incubation time or composition of the S9 mix. No cytotoxicity was observed, whereas precipitation was observed at and above 33 µg/mL test substance. Thus, 2,2-dimethylpropane-1,3-diyl 2-ethylhexanoate did not induce gene mutations in mammalian cells under the conditions of the test.

Conclusion 

In conclusion, assessment of the available data from analogue substances together with the lack of functional groups associated with genotoxicity indicates that there is no mutagenic or clastogenic potential of Multi constituent ester of pentaerythritol 2-ethylhexanoate.

 


Justification for selection of genetic toxicity endpoint
Hazard assessment is conducted by means of read-across from structural analogues/surrogates. 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 between source and target substances and overall quality assessment (refer to the endpoint discussion for further details).

Short description of key information:
In vitro:
Gene mutation (Ames test): S. typhimurium TA 1535, TA 1537, TA 98, TA 100, and E.coli WP2 uvrA: negative with and without metabolic activation (according to OECD TG 471)
Mammalian cytogenicity (chromosome aberration): Chinese Hamster Lung cells: negative (OECD TG 473)
Mammalian mutagenicity (MLA): negative with and without metabolic activation (OECD TG 476)

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

The available data on genetic toxicity of the substance do not meet the criteria for classification according to Regulation (EC) 1272/2008 or Directive 67/548/EEC, and are therefore conclusive but not sufficient for classification.