Fatty acids, C18-unsatd., dimers, 2-ethylhexyl esters

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / bone marrow chromosome aberration

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Acceptable, well-documented publication meeting basic scientific principles

Data source

Reference

Reference Type:

publication

Title:

Cytogenetic Evaluation of Di-(2-ethylhexyl)phthalate and Its Major Metabolites in Fischer 344 Rats

Author:

Putman, D.L. et al.

Year:

1983

Bibliographic source:

Environmental Mutagenesis 5, 227-231

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 475 (Mammalian Bone Marrow Chromosome Aberration Test)

Deviations:

yes

Remarks:

highest dose did not produce any indication of toxicity, such as reduction in mitotic index, only 50 instead of 100 cells per animals were analysed

Principles of method if other than guideline:

The cytogenetics assay was performed according to a modification of the procedures described by Kilian et al [1977].

GLP compliance:

not specified

Type of assay:

chromosome aberration assay

Test material

Test material information

Constituent 1

Reference substance name:

2-ethylhexan-1-ol

EC Number:

203-234-3

EC Name:

2-ethylhexan-1-ol

Cas Number:

104-76-7

Molecular formula:

C8H18O

IUPAC Name:

2-ethylhexan-1-ol

Details on test material:

- Name of test material (as cited in study report): 2-Ethylhexanol
- CAS No. of test material (as cited in study report): 104-76-7
- Analytical purity: 99.7 %
- Impurities (identity and concentrations): 0.3% 2-ethyl-4-methyl pentanol
- Source: Union Carbide

Test animals

Species:

rat

Strain:

Fischer 344

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Breeding Laboratories, Kingston, NY
- Age at study initiation:
- Assigned to test groups randomly: yes
- Weight at study initiation: 150-190 g
- Housing: three to four per cage during the quarantine period and one per cage during treatment on hardwood chips.
- Diet: free access to certified laboratory chow
- Water: ad libitum
- Acclimation period: 10-14 days

ENVIRONMENTAL CONDITIONS
- Temperature (°F): 74ºF ±6ºF
- Humidity (%): 50 +/-20
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

corn oil

Duration of treatment / exposure:

5 consecutive days

Frequency of treatment:

daily

Post exposure period:

The rats were sacrificed by carbon dioxide asphyxiation 6 hours after the last dose

Doses / concentrations

Remarks:

Doses / Concentrations:
0, 0.21, 0.07 and 0.02 mL/kg bw/day
Basis:
actual ingested

No. of animals per sex per dose:

5

Control animals:

yes, concurrent vehicle

Positive control(s):

A triethylenemelamine (TEM)-positive control group received a single intraperitoneal injection of 0.5 mg/kg TEM one day prior to sacrifice.

Examinations

Tissues and cell types examined:

Bone marrow cells

Details of tissue and slide preparation:

CRITERIA FOR DOSE SELECTION:
The dose levels, the highest of which represented approximately one-tenth of the five-day LD50, were based on a preliminary five-day dose-finding study

TREATMENT
The rats were sacrificed by carbon dioxide asphyxiation six hr after the last dose, the femurs were removed, and the bone marrow flushed into Hanks' balanced salt solution. After centrifugation at 600-800g for 8-10 min, the cells were treated with 0.075 M KCI for 20-30 min at 37°C. The cells were again centrifuged and washed twice with 5 mL of Carnoy's fixative. The cells were re-suspended in 5 mL of Carnoy's fixative and allowed to stand overnight at 4°C. The cells were then centrifuged and re-suspended to opalescence in fresh Carnoy's fixative. Two to five slides were prepared from each animal. Slides were stained with Giemsa and permanently mounted.
A minimum of 50 metaphase spreads from each animal was scored for chromatid and chromosomal gaps, breaks, and fragmentation, structural rearrangements, and ploidy [Cohen and Hirschorn, 1971; Kilian et al, 1977; Legator et al, 1973].
Spreads were selected for evaluation by systematic scanning of slides. Only cells which appeared intact with the chromosomes spread symmetrically, with no other metaphase cells intruding into the vicinity, and which contained no less than 36 chromosomes, were scored. In addition, the mitotic index (mitosis/100 cells) was recorded for each test animal.

Statistics:

chi-square or Student's t-statistics

Results and discussion

Test results

Sex:

male

Genotoxicity:

negative

Toxicity:

no effects

Vehicle controls validity:

valid

Negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

RESULTS OF RANGE-FINDING STUDY
The results of the preliminary five-day oral toxicity studies indicated that the five-day LD50 for 2-ETHYLHEXANOL was 2.1 mL/kg/day.
Based on these data, the dose levels selected for the cytogenetics assay were 0.21, 0.07, and 0.02 ml/kg/day for 2-ETHYLHEXANOL.

RESULTS OF DEFINITIVE STUDY
There were no overt toxicological signs in any test or control animal during the five-day treatment period. The number of cells with chromosomal aberrations for 2-ETHYLHEXANOL-treatment groups was not statistically (P >0.05) increased relative to the corn oil control group. In addition, there were no apparent changes in the percentage of aneuploid cells or mitotic index. The number of chromatid gaps was increased; however, since the genetic significance of chromatid gaps is not clearly understood, they were not included in determination of number of metaphases with aberrations or total number of cells with one or more aberrations.
 
The positive controls, on the other hand, demonstrated severe damage, with approximately 26% of all cells analyzed containing one or more aberrations.
 
Based on the results of this study, it is concluded that under the conditions of the test, 2-ETHYLHEXANOL did not induce detectable chromosomal damage after oral administration.

Any other information on results incl. tables

Table 1: Results of chromosome aberration study in rats with 2-ethylhexanol (EH)

Treatment	No. of animals	No. of metaphases	Mitotic index^a	Gaps^b	Metaphases with aberrations^c		Breaks^d	No. of metaphases with rearrangements	Severely damaged cells
					No.	%
Corn oil (5 mL/kg bw/d)	5	250	4.8	2	0	0	0	0	0
0.2 mL/kg bw/d EH	5	250	3.7	13	1	0.4	1	0	0
0.07 mL/kg bw/d EH	5	250	4.1	13	1	0.4	1	0	0
0.02 mL/kg bw/d EH	5	250	5.0	15	0	0	0	0	0
TEM 0.5 mg/kg bw	5	250	4.2	71	66	26.4	54	19	43

^aNumber of cells in mitosis per 100 cells.

^bThe number of chromatid gaps were recorded for each treatment group; however, since their genetic significance is not clearly understood, they were not included in the assessment of chromosomal damage.

^cIncludes metaphases with breaks and rearrangements.

^dBreaks were of chromatid type only for treated animals.

^eSeverely damaged cells, ie, cells having more than 10 aberrations.

Of the 50 metaphase bone marrow cells examined from each animal, no significant increase in chromatid and chromosome breaks or structural rearrangements were noted. The mitotic index, determined from 100 cells per animal was unaffected.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): negative