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Genetic toxicity: in vivo

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Administrative data

Endpoint:
genetic toxicity in vivo
Remarks:
Type of genotoxicity: other: Micronucleus assay
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
data from handbook or collection of data
Justification for type of information:
Data is from peer reviewed journal

Data source

Reference
Reference Type:
publication
Title:
In vivo micronucleus test in mice with 1-phenylethanol
Author:
Günter Engelhardt
Year:
2006
Bibliographic source:
Arch Toxicol (2006) 80:868–872

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
other: OECD No. 474 (July 21, 1997) and EC Directive 2000/32, B. 12 (May 19, 2000)
Principles of method if other than guideline:
In vivo micronucleus test in mice was performed to evaluate the mutagenic nature of the test compound 1-phenylethanol
GLP compliance:
yes
Type of assay:
other: Mouse bone marrow micronucleus assay

Test material

Constituent 1
Chemical structure
Reference substance name:
1-phenylethanol
EC Number:
202-707-1
EC Name:
1-phenylethanol
Cas Number:
98-85-1
Molecular formula:
C8H10O
IUPAC Name:
1-phenylethanol
Test material form:
other: Liquid
Details on test material:
Details on test material
- Name of test material (as cited in study report): 1-phenylethanol
- Molecular formula (if other than submission substance): C8H10O
- Molecular weight (if other than submission substance): 122.166 g/mol
- Substance type: Organic
- Physical state: Liquid
Purity: 98.2%
- Impurities (identity and concentrations): 1.8 %

Test animals

Species:
mouse
Strain:
other: NMRI
Details on species / strain selection:
No data
Sex:
male
Details on test animals or test system and environmental conditions:
Details on test animals and env conditions
TEST ANIMALS
- Source: Charles River Deutschland GmbH (Sulzfeld, Germany)
- Age at study initiation: 5–8 weeks old
- Weight at study initiation: 30g
- Assigned to test groups randomly: [no/yes, under following basis: ] Random
- Fasting period before study: No data available
- Housing: Makrolon cages type MI in fully air-conditioned rooms
- Diet (e.g. ad libitum): standard rat/mouse laboratory feed ad libitum
- Water (e.g. ad libitum): drinking-quality tap water ad libitum
- Acclimation period: Minimum 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20–24⁰C
- Humidity (%): 30–70%
- Air changes (per hr): No data available
- Photoperiod (hrs dark / hrs light): 12/12-h light/dark cycle (light on 6:00–18:00 h)

IN-LIFE DATES: From: To: No data available

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
Vehicles
- Vehicle(s)/solvent(s) used: Olive oil
- Justification for choice of solvent/vehicle: No data available
- Concentration of test material in vehicle: 0, 187.5, 375 or 700 mg/kg body weight
- Amount of vehicle (if gavage or dermal): 10 ml/ kg body weight
- Type and concentration of dispersant aid (if powder): No data available
- Lot/batch no. (if required): No data available
- Purity: No data available
Details on exposure:
- Vehicle(s)/solvent(s) used: Olive oil
- Justification for choice of solvent/vehicle: No data available
- Concentration of test material in vehicle: 0, 187.5, 375 or 700 mg/kg body weight
- Amount of vehicle (if gavage or dermal): 10 ml/ kg body weight
- Type and concentration of dispersant aid (if powder): No data available
- Lot/batch no. (if required): No data available
- Purity: No data available
Duration of treatment / exposure:
Duration of exposure: 24 or 48 hrs
Frequency of treatment:
Once
Post exposure period:
No data available
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 187.5, 375 or 700 mg/kg body weight
Basis:
actual ingested
No. of animals per sex per dose:
Total: 40
0 mg/Kg bw: 10 (5/dose at 24 and 48 hrs)
187.5 mg/Kg bw: 5/dose at 24 hrs
375 mg/Kg bw: 5/dose at 24 hrs
750 mg/Kg bw: 10 (5/dose at 24 and 48 hrs)

Positive control 1: 5/dose at 24 hrs
Positive control 2: 5/dose at 24 hrs
Control animals:
yes, concurrent vehicle
Positive control(s):
Positive controls
Cyclophosphamide- for clastogenicity
Vincristine sulfate- for aneugenic activity
- Justification for choice of positive control(s): No data available
- Route of administration: Oral
Cyclophosphamide- Oral
Vincristine sulfate- intraperitoneally

- Doses / concentrations:
Cyclophosphamide- 20 mg/kg body weight
Vincristine sulfate- 0.15 mg/kg body weight

Examinations

Tissues and cell types examined:
PCEs and NCEs isolated from bone marrow cells
Details of tissue and slide preparation:
Details of tissue and slide preparation
CRITERIA FOR DOSE SELECTION: In a range finding study for the determination of the
acute oral toxicity using male and female mice, deaths were observed at doses of 800 mg/kg body weight and above. At 750 mg/kg body weight, all animals survived, but clinical signs of toxicity such as piloerection, irregular respiration and staggering were observed.

Thus, this dose was considered to be the maximum tolerated dose and the dose levels selected for this study were 750, 375 and 187.5 mg/kg body weight.

TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields):

DETAILS OF SLIDE PREPARATION: Micronucleus suspension was dropped onto clean slides. Smears were prepared, air-dried and stained with May-Gruenwald and Giemsa solutions.

METHOD OF ANALYSIS: Microscopic analysis was performed

OTHER: The bone marrow was prepared.
After having sacrificed the animals by cervical dislocation, the femora were removed and freed from muscles. The epiphyses were cut off and the bone marrow was flushed out of the diaphysis into centrifuge tubes using a cannulated syringe filled with fetal calf serum (FCS; 37C; about 2 ml/femur). After thorough mixing, the suspension was centrifuged at 1,500 rpm for 5 min and the sediment was re-suspended with 50 µL fresh FCS.
Evaluation criteria:
For each animal, 2,000 polychromatic erythrocytes (PCEs) were scored and evaluated for the presence of small micronuclei (d < D/4) indicative of a clastogenic mode of action and large micronuclei (d ‡ D/4) indicative of an aneugenic mode of action

The number of normochromatic erythrocytes (NCEs) both with and without micronuclei in the fields containing 2,000 PCEs were recorded separately.
Statistics:
Wilcoxon’s rank test (one-sided) was performed for comparison of the relative frequencies of micronucleated cells in each animal of the dose groups with those of the negative control groups.

Results and discussion

Test results
Sex:
male
Genotoxicity:
negative
Toxicity:
not specified
Vehicle controls validity:
valid
Negative controls validity:
not specified
Positive controls validity:
valid
Remarks on result:
other: No mutagenic potential
Additional information on results:
RESULTS OF RANGE-FINDING STUDY
- Dose range: Dose range is not properly mentioned but includes study at 750 and 800 mg/Kg bw
- Solubility: No data available
- Clinical signs of toxicity in test animals: Deaths were observed at doses of 800 mg/kg body weight and above. At 750 mg/kg body weight, all animals survived, but clinical signs of toxicity such as piloerection, irregular respiration and staggering were observed.
- Evidence of cytotoxicity in tissue analyzed: No data available
- Rationale for exposure: No data available
- Harvest times: No data available
- High dose with and without activation: No data available
- Other: No data available

RESULTS OF DEFINITIVE STUDY
- Types of structural aberrations for significant dose levels (for Cytogenetic or SCE assay):
- Induction of micronuclei (for Micronucleus assay): PCEs and NCEs were observed
- Ratio of PCE/NCE (for Micronucleus assay): The ratio of PCE:NCE was not affected in treated animals; the slight depression after 48 h and increase after 24 h may reflect the normal variability rather than bone marrow toxicity.
- Appropriateness of dose levels and route:
Dose level: 0, 187.5, 375 or 700 mg/kg body weight
Route: Oral
- Statistical evaluation: Wilcoxon’s rank test (one-sided) was performed for comparison of the relative frequencies of micronucleated cells in each animal of the dose groups with those of the negative control groups.

Applicant's summary and conclusion

Conclusions:
The test compound 1-phenylethanol is not clastogenic in nature when study was performed using mouse bone marrow cells.
Executive summary:

In vivo micronucleus assay in mouse bone marrow was carried out to test the clastogenic nature of the test compound1-phenylethanol. The animals were given 1-phenylethanol in single oral doses up to the maximum tolerated dose of 750 mg/kg body weight (0, 187.5, 375 or 700 mg/kg body weight).Bone marrow was sampled 24 and 48 h after treatment. Under the experimental conditions used, there was no evidence of increased micronuclei frequencies at any dose or sampling time. The number of polychromaticerythrocytes containing either small or large micronuclei at each dose was not significantly increased above the concurrent negative (solvent) control frequencies and was always within the historical negative control range (0.3–3.3% based on > 300 experiments) at each sampling time. The ratio of PCE: NCE was not affected in treated animals; the slight depression after 48 h and increase after 24 h may reflect the normal variability rather than bone marrow toxicity. Oral administration of a single dose of 1-phenylethanol led to evident signs of toxicity such as staggering in the intermediate (375 mg/kg) and top (750 mg/kg) dose animals and to irregular respiration, piloerection, abdominal position and to a narcotic-like state in all animals of the highest dose group, clearly demonstrating the attainment of a maximum tolerated dose. These findings indicate that 1- phenylethanol is not clastogenic in vivo.