2-ethylhexanal

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was conducted according to OECD guideline 474 (Mammalian Erythrocyte Micronucleus Test) under GLP conditions. There were only minor deviations. The study is fully sufficient for endpoint evaluation.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2004

Report date:

2004

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

GLP compliance:

yes

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

2-ethylhexanal

EC Number:

204-596-5

EC Name:

2-ethylhexanal

Cas Number:

123-05-7

Molecular formula:

C8H16O

IUPAC Name:

2-ethylhexanal

Details on test material:

- Name of test material (as cited in study report): 2-ethylhexanal
- Physical state: colourless to yellow liquid
- Analytical purity: 99.5%
- Impurities (identity and concentrations): not reported
- Purity test date: not reported
- Lot/batch No.: 00STD77L0
- Stability under test conditions: in corn oil, stability analytically verfied with a comparable batch (9147-105)
- Storage condition of test material: at RT (light protected, under nitrogen)

Test animals

Species:

mouse

Strain:

NMRI

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: RCC Ltd., Animal breeding services, CH-4414 Füllinsdorf
- Age at study initiation: 8-10 weeks
- Weight at study initiation: M 33.4g (SD +-0.7g), F 29.8g (SD +-1.3g)
- Fasting period before study: starvation over night, but water ad libitum
- Housing: Makrolon Type I, with wire mesh top (EHRET GmbH, D-79302 Emmendingen)
- Diet: pellet standard diet ad libitum (Harlan Winkelmann GmbH, D-33178 Borchen)
- Water: tap water ad libitum
- Acclimation period: minimum 5 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22+-3°C
- Humidity (%): 30-70%
- Air changes (per hr): not reported
- Photoperiod (hrs dark / hrs light): 6/6 (6.00a.m. - 6.00p.m.)

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

- Vehicle(s)/solvent(s) used: corn oil
- Justification for choice of solvent/vehicle: corn oil is relatively non-toxic for the animals
- Concentration of test material in vehicle: 200 mg/ml
- Amount of vehicle (if gavage or dermal): 10 ml/kg bw

Duration of treatment / exposure:

animals received the test substance orally via gavage

Frequency of treatment:

single administration

Post exposure period:

24h

Doses / concentrations

Remarks:

Doses / Concentrations:
2000 mg/kg bw
Basis:
nominal conc.
Limit test

No. of animals per sex per dose:

6

Control animals:

yes, concurrent vehicle

yes, historical

Positive control(s):

cyclophosphamide (CPA)
- Justification for choice of positive control(s):
- Route of administration: orally, once
- Doses / concentrations: 40mg/kg bw (Volume 10 ml/kg bw)

Examinations

Tissues and cell types examined:

- bone marrow
- animals were examined 1h, 2-4h, 6h and 24h after treatment

Details of tissue and slide preparation:

CRITERIA FOR DOSE SELECTION: maximum dose - limit test (2000 mg/kg bw)

TREATMENT AND SAMPLING TIMES (in addition to information in specific fields): orally once vie gavage, sampling after 24h

DETAILS OF SLIDE PREPARATION:
- bone marrow was flushed out with fetal calf serum after femora were removed and epiphysis were cut off
- cell suspension was then centrifuged at 390 x g for 10 min, supernatant was discarded
- a small drop of the suspension was spread on a slide and the smear was air dried
- staining with MAY-GRÜNWALD (Merck, D-64293 Darmstadt) / GIEMSA (Gurr, BDH Limited Poole, Great Britain)
- cover slips were mounted with EUKITT (Kindler, D-79110 Freiburg)

METHOD OF ANALYSIS:
- evaluation was performed with an NIKON microscope with 100 x oil immersion objectives
- analysis was performed in coded slides

Evaluation criteria:

- cytotoxic effect description: ratio of polychromatic erythrocytes (PCE) and total erythrocytes
- 5M and 5F were analyzed
- at least 2000 polychromatic erythrocytes (PCE) were analyzed for micronuclei

Statistics:

non-parametric Mann-Whitney Test

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

negative

Toxicity:

yes

Remarks:

Mortality: 1M and 1F died after treatment

Vehicle controls validity:

valid

Negative controls validity:

not specified

Positive controls validity:

valid

Any other information on results incl. tables

Table shows a summary of the micronucleus test:

Test group	Dose [mg/kg bw]	Sampling time (h)	PCEs with micronuclei (%)	Range	PCE per 2000 erythrocytes
Vehicle	0	24	0.035	0-2	985
Test substance	2000	24	0.100	1-4	1012
Positive control	40	24	1.935	14-61	936

PCE - Polychromatic erythrocytes

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): negative

Executive summary:

The study was conducted according to OECD guideline 474 (Mammalian Erythrocyte Micronucleus Test) under GLP conditions. There were only minor deviations, the study is fully sufficient for endpoint evaluation. 2-ethylhexanal was tested for micronucleus induction in mouse after single administration of 2000 mg/kg bw (Limit test). A vehicle control and a positive control (cyclophosophamid) CPA for micronucleus formation were run in parallel and were both valid. Twelve animals (6M/6F) were treated per test group and analyzed after 24h (5M/5F). A cytotoxic effect was described as the ratio between polychromatic and total erythrocytes. 1M and 1F died after treatment with this dose. In a prior study (supporting study in this endpoint section) a weak positiv effect was detected, but only males were tested and effect were slightly higher than historical controls. Therefore the study was repeated (this study) in both sexes at the limit dose.

No statistically significant enhancement in the frequency of micronuclei after 2-ethylhexanal exposure was observed in this study. The test substance does not induce micronuclei in mouse erythrocytes in vivo.