2,2`-[6-(phenylamino)-heteromonocycle-2,4-diyl]diphenol

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

Study period:

2008

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2008

Report date:

2008

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: Commission Directive 2000/32/EC, Annex 4C

Version / remarks:

dated MAy 19, 2000

GLP compliance:

yes

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

2,2`-[6-(phenylamino)-1,3,5-triazine-2,4-diyl]diphenol

Cas Number:

1248-66-4

Molecular formula:

C21H16N4O2

IUPAC Name:

2,2`-[6-(phenylamino)-1,3,5-triazine-2,4-diyl]diphenol

Test animals

Species:

mouse

Strain:

NMRI

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan Winkelmann GmbH
- Age at Start of Acclimatisation: 8 - 10 weeks
- Weight at study initiation: males mean value 37.8 g (SD ± 2.0 g); females mean value 29.6 g (SD ± 2.0 g)
- Assigned to test groups randomly: yes
- Fasting period before study:
- Housing: single in Makrolon Type I, with wire mesh top
- Diet: pelleted standard diet, ad libitum
- Water: tap water, ad libitum
- Acclimation period: minimum 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3 °C
- Humidity (%): 30 - 70 %
- Photoperiod (hrs dark / hrs light): artificial light 6.00 a.m. - 6.00 p.m.

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

30% DMSO/70% PEG 400

Details on exposure:

substance formulated in vehicle

Duration of treatment / exposure:

animals received the test item once

Frequency of treatment:

once

Post exposure period:

24 h, 48 h

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
2000 mg/kg body weight
Basis:
actual ingested

Doses / concentrations 2

Remarks:

Doses / Concentrations:
1000 mg /Kg body weight
Basis:
actual ingested

Doses / concentrations 3

Remarks:

Doses / Concentrations:
500 mg/kg body weight
Basis:
actual ingested

No. of animals per sex per dose:

low dose: 6 females, 6 males
medium dose: 6 females, 6 males
main experiment: 12 females, 12 males

Control animals:

yes, concurrent vehicle

Positive control(s):

Cyclophosphamide, 40 mg/kg b.w; 24 h;

Examinations

Tissues and cell types examined:

bone marrow cells; polychromatic erythrocytes; normochromatic erythrocytes

Details of tissue and slide preparation:

according guideline

Evaluation criteria:

standard

Statistics:

nonparametric Mann-Whitney test

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

negative

Toxicity:

yes

Remarks:

ruffeled fur within 4 h after treatment with 2000 mg/kg b.w.

Vehicle controls validity:

valid

Negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

pre-experiment:
4 animals (2 males, 2 females) treated with 100 mg/kg b.w. did not express any toxic reactions;
4 animals (2 males, 2 females) treated with 1000 mg/kg b.w: : 1 male and 1 female had ruffeled fur within 2 and 6 h;
4 animals (2 males, 2 females) treated with 2000 mg/kg b.w: :2 males, 1 female had ruffeled fur 1 h post-treatment and 2 males and 2 females within 2 and 24 h post-treatment;

Applicant's summary and conclusion

Conclusions:

The oral application of the substance to mice did not induce increased micronucleated polychromatic erythrocytes in bone marrow.

Executive summary:

The test item was assessed in the micronucleus assay for its potential to induce micronuclei in polychromatic erythrocytes (PCE) in the bone marrow of the mouse. The test item was formulated in 30% DMSO / 70% PEG 400, which was also used as vehicle control. The volume administered orally 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 normochromatic 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: 500, 1000, and 2000 mg/kg b.w..
48 h preparation interval: 2000 mg/kg b.w..
As estimated by a pre-experiment 2000 mg test item per kg b.w. (the maximum guideline-recommended dose) was suitable.
The mean number of polychromatic erythrocytes was not decreased after treatment with the test item as compared to the mean value of PCEs of the vehicle control indicating that the test item did not have any cytotoxic properties in the bone marrow.
In comparison to the corresponding vehicle controls there was no statistically significant or biologically relevant enhancement in the frequency of the detected micronuclei at any preparation interval and dose level after administration of the test item. The mean values of micronuclei observed after treatment with test item were below or near to the value of the vehicle control group. 40 mg/kg b.w. cyclophosphamide administered orally was used as positive control which showed a statistically significant increase of induced micronucleus frequency.
In conclusion, it can be stated that during the study described and under the experimental conditions reported, the test item did not induce micronuclei as determined by the micronucleus test in the bone marrow cells of the mouse.