3-amino-2-chlor-6-methylphenol

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Type of information:

experimental study

Adequacy of study:

key study

Study period:

09.08.2004 - 07.12.2004

Reliability:

1 (reliable without restriction)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2005

Report date:

2005

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

mammalian germ cell cytogenetic assay

Test material

Test material information

Constituent 1

Reference substance name:

3-amino-2-chlor-6-methylphenol

EC Number:

283-144-9

EC Name:

3-amino-2-chlor-6-methylphenol

Cas Number:

84540-50-1

Molecular formula:

C7H8ClNO

IUPAC Name:

3-amino-2-chloro-6-methylphenol

Test material form:

other: beige powder

Details on test material:

molecular weight: 157.5

Test animals

Species:

mouse

Strain:

NMRI

Details on species / strain selection:

The mouse is an animal which has been used for many years as suitable experimental animal in cytogenetic investigations. There are many data available from such investigations which may be helpful in the interpretation of results from the micronucleus test. In addition, the mouse is an experimental animal in many physiological, pharmacological and toxicological studies. Data from such experiments also may be useful for the design and the performance of the micronucleus test (1,2,3,4,5,6).

Sex:

male/female

Details on test animals or test system and environmental conditions:

Age:
males: 5-8 weeks
females: 7-10 weeks
Acclimatisation: minimum 5 days
Body weight:
males mean value 38.8 g (SD ± 3.3 g)
females mean value 31.9 g (SD ± 3.0 g)
Housing: single
Feed: pelleted standard diet, ad libitum
Water: tap water, ad libitum,

Administration / exposure

Route of administration:

intraperitoneal

Vehicle:

aqueous DMSO (30%)

Details on exposure:

dose volume: 10 mL/kg b.w.
On the day of the experiment, the test item was formulated in aqueous DMSO (30%). The vehicle was chosen to its relative non-toxicity for the animals.

Duration of treatment / exposure:

single administration

Frequency of treatment:

single administration

Post exposure period:

24 and 48 hours

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

100 mg/kg bw/day

Remarks:

24 h preparation interval

Doses / concentrations 2

Dose / conc.:

200 mg/kg bw/day

Remarks:

24 h preparation interval

Doses / concentrations 3

Dose / conc.:

400 mg/kg bw/day

Remarks:

24 h preparation interval

Doses / concentrations 4

Dose / conc.:

400 mg/kg bw/day

Remarks:

48 h preparation interval

No. of animals per sex per dose:

5

Control animals:

yes

Positive control(s):

CPA; Cyclophosphamide
Purity: >98%
Dissolved in: deionised water

Examinations

Tissues and cell types examined:

Blood cells - erythrocytes

Evaluation criteria:

A test item is classified as mutagenic if it induces either a dose-related increase or a clear increase in the number of micronucleated polychromatic erythrocytes in a single dose group. Statistical methods (nonparametric Mann-Whitney test will be used as an aid in evaluating the results. However, the primary point of consideratrion is the biological relevance of the results.
A test item that fails to produce a biological relevant increase in the number of micronucleated polychromatic erythrocytes is considered non-mutagenic in this system.

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

negative

Toxicity:

yes

Any other information on results incl. tables

Summary of Micronucleus Test Results

test group	dose mg/kg b.w.	sampling time (h)	PCEs with micronuclei (%)	range	PCE per 2000 erythocytes
vehicle	0	24	0.120	0 -5	1056
test item	100	24	0.150	1 -5	1081
test item	200	24	0.080	0 -3	1081
test item	400	24	0.105	0 -6	1116
positive control	40	24	2.670	22 -86	1078
test item	400	48	0.105	0 -5	1061

Applicant's summary and conclusion

Conclusions:

In conclusion, it can be stated that under the experimental conditions reported, the test item did not induce micronuclei as determined by the micronucleus test with bone marrow cells of the mouse.

Therefore, A 094 is considered to be non-mutagenic in this micronucleus assay.

Executive summary:

This study was performed to investigate the potential of A 094 to induce micronuclei in polychromatic erythrocytes (PCE) in the bone marrow of the mouse.

The test item was formulated in aqueous DMSO (30%), which was also used as vehicle control. The volume administered intraperitoneally (i.p.) was 10 mL/kg b.w.. 24 h and 48 h after a single administration of the test item the bone marrow cells were collected for micronuclei analysis.  

Ten animals (5 males, 5 females) per test group were evaluated for the occurrence of micronuclei. At least 2000 polychromatic erythrocytes (PCEs) per animal were scored for micronuclei.

To describe a cytotoxic effect due to the treatment with the test item the ratio between polychromatic and total erythrocytes was determined in the same sample and reported as the number of PCEs per 2000 erythrocytes.

The following dose levels of the test item were investigated:

24 h preparation interval: 100, 200 and 400 mg/kg b.w.

48 h preparation interval: 400 mg/kg b.w.

As estimated by pre-experiments 400 mg A 094 per kg b.w. was the highest applicable dose without significant effects on the survival rates, but with clear signs of toxicity. At a higher dose (500 mg/kg) one male out of 4 animals (2 males, 2 females) died.

After treatment with the test item the number of PCEs was not substantially decreased as compared to the mean value of PCEs of the vehicle control thus indicating that A 094 did not exert any cytotoxic effects in the bone marrow. The bioavailability of the test item was, however, confirmed by chemical analysis of the blood of the treated animals.

In comparison to the corresponding vehicle controls there was no biologically relevant or statistically significant enhancement in the frequency of the detected micronuclei at any preparation interval after administration of the test item and with any dose level used.

40 mg/kg b.w. cyclophosphamide administered i.p. was used as positive control which showed a substantial increase of induced micronucleus frequency.