Registration Dossier

Administrative data

Key value for chemical safety assessment

Additional information

Genetic toxicity in vitro

Mutagenicity in bacterial test systems

The mutagenicity of 2 -EH was tested in bacterial test sytems (S. tyhimurium TA98, TA100, TA1535, TA1537, TA1538, and E. coli WP2 uvrA) according to OECD TG 471 and TG 472 both with and without metabolic activation in a dose range from 1 to 1000 µg/plate (Shimizu et al., 1985). 2 -EH did not increase the number of revertants in any of the test strains. Growth inhibition was seen at 500 and 1000 µg/plate. The negative and positive controls performed as expected. Therefore, 2 -EH was not mutagenic in baterial test systems in-vitro.

In studies with lower reliability (RL 3, because one important tester strain was missing according to current testing guidelines). For example 2-EH was not mutagenic in Salmonella typhimurium TA98, TA100, TA1535, and TA1537, with and without metabolic activation, at concentrations up to cytotoxic concentrations (330 µg/plate). (NTP, 2007).

The mutagenicity of DEHP and related chemicals including 2 -EH was tested in Salmonella according to OECD TG 471, both with and without metabolic activation by Aroclor 1254 -induced male rat and hamster liver 9000g supernatant (S9) (Zeiger et al., 1985). The test strains used were TA98, TA100, TA1535, and TA1537. The concentration of 2 -EH ranged from 3.3 to 220 µg/plate; the dose range selection was based on the results of a preliminary test using TA100.

2 -EH was not mutagenic in this study, as the number of histidine independent revertants was not increased in a dose-related manner in any test strain, either in the absence or presence of metabolic activation. The negative and positive controls performed as expected.

Moreover there are further studies for which mostly only short abstracts are available or which were cited from secondary sources (RL4), which all show negative findings when Sallmonella strains TA 98, TA 100, TA 1535, TA 1537 and TA 1538 were tested with or without metabolic activation for reverse mutations (various sources). 2 -EH was further examined for DNA modifying activity in E. coli strain W3110 (polA+) and its polymerase deficient deruvative p3478 (pol-). When tested with the relevant vehicle (DMSO) no DNA modifying activity was observed after 2 -EH administration (EPA, 1981). At 500 µg/disk, 2-ethyl-1-hexanol was found to be non-mutagenic in the Rec-assay with Bacillus subtilis strains H17 and M45 (Tomita et al., 1982).

The results of one study were disregarded as the weakly positive response of 8 -aza-guanine resistant mutants of the Salmonelle typhymurium strain TA 100 (increase was 2.2 -fold at 1 mM and 2.8 -fold at 1.5 mM compared to control), was correlated with high cytotoxicity at these concentrations (50 and 70%, respectively).

 

Therefore,2 -EH was not mutagenic in bacterial in-vitro test systems, with and without metabolic activation.

Cytogenicity in mammalian cells in-vitro (or MN)

2 -EH at 50, 108, and 233 µg/mL did not cause chromosome aberration in CHO cells in vitro, with or without metabolic activation (NTP, 2007). This finding is supported by a study of lower reliability, due to some methodological deficiencies. Within this study 2-EH over a 1.5–2.8 mM concentration range did not cause a statistically significant increase in frequency of chromosomal aberrations compared to control in Chinese Hamster ovary cells. In the highest dose tested relevant cytotoxic effects were seen (Phillips, 1982).

Mutagenicity in mammalian cells in-vitro

2-ethylhexanol in the range 0.018, 0.024, 0.032, 0.042, 0.056, 0.075, 0.10, 0.13, 0.18, 0.24 µL/ mL did not increase the mutation frequency in L5178Y mouse lymphoma cells, with or without metabolic activation. The growth rate was 10% at the highest dose level. Complete toxicity was seen at 1 µgL/mL and above. The study is considered to be valid, though documentation is limited. It was similar to OECD TG 476. No data were provided regarding GLP compliance (Kirby et al., 1983).

The genotoxic potential of 2 -ethylhexanol was also examined in a HGPRT test using mammalian cells (CHO cells), with and without metabolic activation (microsomes from Aroclor 1254 induced rat liver). The study was conducted similar to OECD TG 476 and under GLP conditions. The concentration range was selected based on the results of a cytotoxicity study, both with and without metabolic activation. No cytotoxicity was seen at 200 nL/mL, complete cytotoxicity was seen at 500 nL/mL.

The concentration range was 20 - 400 nL/mL in the mutagenicity study with little toxicity up to 250 nL/mL, but considerable cytotoxicity at 300 to 400 nL/mL. 2 -ethylhexanol did not increase the number of revertants at any dose, with or without metabolic activation (400 nL/mL was excluded due to high cytotoxicity). Vehicle controls and positive controls performed as expected. The study is considered to be valid as it was performed similar to OECD TG 476 and under GLP conditions (Litton Bionetics, 1985).

Therefore,2 -EH was not mutagenic in mammalian cells in-vitro, with and without metabolic activation.

Other in vitro tests performed with 2-EH

In a reliable study 2-EH was tested in-vitro for its potential to induce DNA-damage (sister chromatid exchange, SCE) in mammalian cells in-vitro. The test was conducted in accordance with OECD TG 479, i.e. the test concentrations (1.7, 5, 17, 50, 167, and 500 µg/mL) ranged up to cytotoxic concentrations, and tests were performed with and without metabolic activation using induced rat liver 9000g supernatant (S9).2 -EH did not induce SCE, a marker of DNA damage, in mammalian cells, both with and without metabolic activation. Negative and positive controls perfomed as expected (NTP, 2007).

A diversity of tests with low reliability testify that 2-EH has no cell promoting or transforming activity. For example, when 2-EH was given to Mouse JB6 C141 cells no anchorage independent cell promotion was observed (Ward, 1986). In various tests with Balb/3T3 mouse cells no cell transforming activity was noted under varying test conditions (e.g. with or without metabolic activity; EPA, 1981, 1983). And no unscheduled DNA repair activity was observed in primary rat hepatocytes when 2-EH was applied (Hodgson, 1982).

Genetic toxicity in vivo

Furthermore, there are different in vivo studies of lower reliability (due to limited reporting) supporting the fact that 2-EH has no genotoxic properties.

Male Fischer 344 rats were dosed by oral gavage for five consecutive days with 0.02, 0.07, and 0.21 ml/kg/day of 2-ethyl-1-hexanol. When compared to controls, there was no increase in chromatid and chromosome breaks. The mitotic index was also not affected in bone marrow cells (Putman et al., 1983).

WhenB6C3F1 mice (6/sex/dose) were administered 2 -ethylhexanol at a dose of 456 mg/kg/d, and polychromatic erythrocytes were investigated no increase of micronuclei was observed. Positive and negative controls yielded the expected results. 2-EH was not considered to be clastogenic under the conditions of this assay (EPA, 1986).

In a dominant lethal study - conducted with ICR mice treated orally with 2-EH for 5 consecutive days at doses of 1000, 500, and 250 mg/kg/d - the fertility indices and average numbers of dead and total implants per pregnancy were within the normal range. It was concluded that 2-ethylhexanol did not induce dominant lethal mutations after oral administration (Rushbrook, 1982).

2-EH was not mutagenic in a sex-linked recessive lethality (SLRL) test in Drosopila melanogaster, when fed in a solution of 5% aqueous sucrose or injected with 2-EH in a solution of 0.7% aqueous NaCl (Valencia, 1985).


Justification for selection of genetic toxicity endpoint
weight-of-evidence: all relevant in vitro and in vivo studies are negative.

Short description of key information:
2-EH was not genotoxic in vitro using bacterial and mammalian cell test systems. 2-EH was not mutagenic in bacteria (Salmonella typhimurium strains TA98, TA100, TA1535, TA1537, TA1538, and E. coli WP2 uvrA), with or without metabolic activation) or mammalian cells in vitro (HGPRT and TK), and it did not induce chromosome aberration or sister chromatid exchange in mammalian cells. 2-EH was not mutagenic in various in vivo genotoxicity tests with mice (in vivo MNT, dominant lethal study and cytogenetic assay) or Drosophila melanogaster (SLRL test).

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

2-EH was not mutagenic in bacteria (Salmonella typhimurium strains TA98, TA100, TA1535, TA1537, TA1538, and E. coli WP2 uvrA),

with or without metabolic activation) or in mammalian cells in vitro (HGPRT in CHO cells and TK in mouse lymphoma cells), and it did not induce chromosome aberration or sister chromatid exchange in mammalian cells. 2-EH was not mutagenic in various in vivo genotoxicity tests with mice (in vivo MNT, dominant lethal study and cytogenetic assay) or Drosophila melanogaster (SLRL test).

According to Regulation (EC) no. 1272/2008 no classification for mutagenic properties is therefore required.