Registration Dossier

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
Study period:
March 23, 1983 - April 11, 1983
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Test was conducted similar to OECD guideline 474, and in compliance with GLP Standards
Cross-referenceopen allclose all
Reason / purpose:
reference to same study
Reason / purpose:
reference to other study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1983
Report Date:
1983

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Deviations:
yes
Remarks:
Treatment Schedule
GLP compliance:
yes
Type of assay:
micronucleus assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): 2-Ethylhexyl 4-methoxycinnamate
- Physical state: liquid

Test animals

Species:
mouse
Strain:
other: Füllinsdorf Albino SPF, outbred strain
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Institute of Biological and Medical Research, CH-4414 Füllinsdorf
- Age at study initiation: approx. 6-8 weeks
- Weight at study initiation: female: 29.3 g, male: 31.8 g
- Assigned to test groups randomly: no
- Housing: Maximum number of animals per cage: 6
- Diet: Nafag 850-cubes ad libitum
- Water: Basel tap water ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2
- Humidity (%): 55 ± 10
- Air changes (per hr): conventional air-conditioned rooms
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
- Solvent used: rape oil
- Concentration of test material in vehicle: different concentrations at standard dose volume of 0.1 ml/kg bw applied
Duration of treatment / exposure:
30 hours
Frequency of treatment:
Twofold dosing (over two cell cycles/30 hours)
Doses / concentrations
Remarks:
Doses / Concentrations:
1000, 2500, 5000 mg/kg bw
Basis:
actual ingested
No. of animals per sex per dose:
3
Control animals:
yes, concurrent no treatment
Positive control(s):
Procarbazine hydrochloride (3 male animals used)
- Justification for choice of positive control(s): known to induce micronuclei in erythrocytes of mice
- Route of administration: orally
- Doses / concentrations: 50 mg/kg bw

Examinations

Tissues and cell types examined:
bone-marrow; polychromatic erythrocytes (PCE)
Details of tissue and slide preparation:
CRITERIA FOR DOSE SELECTION: The dose of 5000 mg/kg is the highest applicable one as determined by preliminary experiments.

TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields): subacute repeated dosing over two cell cycles (approximately 30 hours in the mouse); twofold oral application of 1000, 2500 and 5000 mg 2-Ethylhexyl 4-methoxycinnamate per kg bodyweight 30 and 6 hours prior to sacrifice of the mice.

DETAILS OF SLIDE PREPARATION: Four bone-marrow smears per animal were prepared according to the method of Schmid (Schmid W., Mut. Res. 31, 1975, 9). The slides were coded and 24 hours after the preparation they were fixed in absolute methanol and stained with May-Grünwald-Giemsa.

METHOD OF ANALYSIS: For each animal 2000 polychromatic erythrocytes were examined in a double blind fashion. Only cells with clearly identifiable anomalies were recorded as cells containing micronuclei. Micronuclei contained in mature erythrocytes were recorded separately, but not taken into the statistical evaluation.
Evaluation criteria:
A positive result: a statistically significant dose-related increase in the number of micronucleated polychromatic erythrocytes in the bone marrow of treated mice.
Statistics:
The results were evaluated by means of the Jonckheere-test (A.R. Jonckheere, Biometrica 41, 1954, 133) and the U-test (F. Wilcoxon et al., Biometrics 19, 1963, 58).

Results and discussion

Test results
Sex:
male/female
Genotoxicity:
negative
Toxicity:
not specified
Remarks:
The dose of 5000 mg/kg is the highest applicable one as determined by preliminary experiments.
Vehicle controls validity:
not specified
Negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
RESULTS OF RANGE-FINDING STUDY
- Other: The dose of 5000 mg/kg is the highest applicable one as determined by preliminary experiments.

RESULTS OF DEFINITIVE STUDY
- Induction of micronuclei (for Micronucleus assay): No compound-related increase of micronuclei observed

Any other information on results incl. tables

Absence of proportion of immature erythrocytes (PCE) among total erythrocytes.

Absence of data on Preliminary test.

No result data on solvent control (rape oil).

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): negative
After twofold oral application of 1000, 2500 and 5000 mg/kg bodyweight, 2-Ethylhexyl 4-methoxycinnamate induces neither chromosome breaks
nor mitotic non-disjunctions in bone marrow cells of mice. Therefore, the substance does not need to be classified as genotoxic according to the
criteria outlined in Annex I of 1272/2008/EC and Annex VI of 67/548/EEC.
Executive summary:

2-Ethylhexyl 4-methoxycinnamate, a UV-B sunscreen, was evaluated for a potential induction of chromosome breaks and/or mitotic non-disjunctions in vivo by means of the micronucleus test.

After twofold oral application of 1000, 2500 and 5000 mg 2-Ethylhexyl 4-methoxycinnamate per kg bodyweight 30 and 6 hours prior to sacrifice of the mice, no compound related increase of micronuclei could be observed. The dose of 5000 mg/kg is the highest applicable one as determined by preliminary experiments.

It is therefore concluded that under the experimental conditions described in this report, 2-Ethylhexyl 4-methoxycinnamate induces neither chromosome breaks nor mitotic non-disjunctions in mouse bone-marrow cells. Therefore, the substance does not need to be classified as genotoxic according to the criteria outlined in Annex I of 1272/2008/EC and Annex VI of 67/548/EEC.