Registration Dossier

Administrative data

Key value for chemical safety assessment

Genetic toxicity in vivo

Description of key information
In vitro genotoxicity assay were generally negative (non-genotoxic) with EHD. In vivo studies indicate that the one positive assay (chromosome aberration assay) is not relevant.
Link to relevant study records
in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
Type of genotoxicity: chromosome aberration
Type of information:
experimental study
Adequacy of study:
key study
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: A standard protocol under GLP was used and results were found to be valid by peer review prior to publication.
equivalent or similar to guideline
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Principles of method if other than guideline:
MacGregor JT, Wehr CM and Gould DH. Clastogen-induced micronuclei in peripheral blood erythrocytes: the basis of an improved micronucleus test. Environ. Mutagen. 2: 509, 1980 and Nichols WW, Miller RC and Bradt C, In vitro anaphase and metaphase preparations in mutation testing. in BJ Kilbey, M. Legator, W. Nichols and C. Ramel (Eds), handbook of Mutagenicity Test Procedures, Elsevier Publ. Co. NY, NY, 1979, p. 225.
GLP compliance:
Type of assay:
chromosome aberration assay
Details on test animals or test system and environmental conditions:
Sprague—Dawley rats, 6-8 weeks old, were obtained from Charles River Breeding Laboratories, Inc., Kingston, N.Y., USA. For the cytogenetic studies, males ranged in weight from 191 to 230 g and females ranged from 141 to 170 g. Animals were quarantined for a 10 to 14-day period prior to dosing and evaluated daily for signs of illness or poor health. Rats were housed 3-4 per cage during the quarantine period and singly thereafter in plastic shoe-box type cages with wire tops and hardwood bedding chips. Animals were provided with certified rodent chow and water ad libitum. Ear tags with sequential animal numbers were used for animal identifica¬tion. The animal room had a controlled environment with a temperature range of 23 ± 3°C (74 ± 6°F), 50 ± 20% relative humidity and a 12-h light/dark cycle.
Route of administration:
corn oil
Details on exposure:
Male and female, Sprague—Dawley rats, 6-8 weeks of age, were used for preliminary dose-finding and cytogenetic studies. Five males and females per dosage group were dosed by i.p. injection with test and control materials. One or two extra animals were added to the highest dosage groups to provide sufficient group sizes in case of deaths. The maximum tolerated dose was determined in preliminary tests as the highest concentration which produced weight loss or clinical signs of toxicity without producing death. The corn oil vehicle and EHD-vehicle mixture was given at a constant volume of 5 ml/kg. Colchicine (1.0 mg/kg) was given by i.p. injection at 2-4 h prior to sacrifice of the animals. Chromosomes were prepared by flushing bone marrow from femurs and fixation by standard procedures [22]. All slides were coded and 100 cells/animal were evaluated randomly. Chromosomes were sampled at 12, 24 and 48 h after acute dosing.
Duration of treatment / exposure:
single ip injection, then assayed 12, 24 and 48 h later.
Frequency of treatment:
Post exposure period:
12, 24 and 48 h
Doses / Concentrations:
60, 200 and 600 mg/kg
other: wt/wt
No. of animals per sex per dose:
Control animals:
yes, concurrent vehicle
Positive control(s):
Triethylmelamine, 0.5 mg/kg
Tissues and cell types examined:
bone marrow (femur)
Details of tissue and slide preparation:
Bone marrow cell preparations were performed according to Nichols, Miller and Bradt, 1979. 1000 cell/animal were assayed to assess bone marrow cytotoxicity.
Evaluation criteria:
A positive effect was defined by a dose-related trend in the incidence of mPCEs with at least 1 statistically significant indication of a difference from the vehicle control. Incidences of mPCEs from males and females were pooled for statistical analyses if no significant sex-related differences were indicated by statistical comparisons.
P < 0.01; Fisher's Exact Test — one tailed
up to 80% LD50
Vehicle controls validity:
Positive controls validity:
Additional information on results:
Following administration of EHD, animals appeared sedated at the highest dose and a moderate loss of weight (4 to 6%) was observed on the day of sacrifice. Measurements of radioactivity in bone marrow and plasma samples from animals dosed by intraperitoneal injection of radiolabeled EHD showed rapid penetration into bone marrow by 0.5 h after dosing . Radioactivity was detectable from 0.5 through 8.0 h after dosing. Bone marrow/plasma ratios were the same between 2 and 8 h, indicating that a steady-state condition between radioactive uptake and elimination was established.

The incidence of chromosomal aberrations did not differ between males and females, thus values were pooled for statistical analysis. No significant increases in chromosome damage were observed regardless of sampling time. The positive control, TEM, produced a highly significant increase in chromosome aberrations. The corn oil controls were in a low and acceptable range for a valid test.
Interpretation of results (migrated information): negative
Acute dosing of 2-Ethyl-1,3-hexanediol (EHD) resulted in no increase in chromosomal aberrations in an in vivo bone marrow assay in the rat. The substance is evaluated as negative (nonclastogenic).
Executive summary:

A chromosome aberrations assay was conducted in vivo on 2-ethyl-1,3-hexanediol (EHD), < 99% purity, by the intraperitoneal route in groups of 5 male and female Sprague Dawley rats. The procedure was according to MacGregor, Wehr and Gould, 1980.  Rats were sacrificed 12, 24 and 48 h after single ip dosing of EHD at 60, 200 and 600 mg/kg bw. In parallel studies, a single 600 mg/kg dose of EHD, identical to that used for the bone marrow cytogenetic test, containing 10 pCi/kg of [14C] EHD, indicated rapidly detectable radioactivity in the bone marrow and plasma.  Positive controls were TEM (300µg/ml). There was no increase in the incidence of chromosome aberrations in the bone marrow tissue at any time point, whereas the positive control, TEM, showed a highly significant increase in chromosome aberrations.  EHD is evaluated as negative (nonclastogenic) under conditions of this assay.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Additional information

Additional information from genetic toxicity in vivo:

2-Ethyl-1,3-hexanediol was found to be non-mutagenic in vitro (two Ames assays, HGPRT assay in CHO cells). One in vitro clastogenicity assay (chromosome aberrations, 8 h exposure with metabolic activation) demonstrated a positive result. Chromosome aberrations were not increased at other time points, nor in the absence of metabolic activation. The substance did not increase the incidence of sister chromatid exchanges in vitro. Three experiments in vivo were undertaken to investigate the relevance of the possible clastogenic effect. An in vivo micronucleus assay in mice failed to produce an increase in micronucleated PCEs in Swiss Webster mice, and two in vivo analyses of bone marrow cells in Sprague-Dawley rats showed no evidence of chromosomal damage.

Justification for selection of genetic toxicity endpoint
in vivo experimental data

Justification for classification or non-classification

The activity of the substance does not meet the criteria for classification as a mutagen in Regulation EC No. 1272/2008.