N-cyclopropyl-1,3,5-triazine-2,4,6-triamine

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

1986/11 to 1987/07

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1987

Report date:

1987

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

EU Method B.12 (Mutagenicity - In Vivo Mammalian Erythrocyte Micronucleus Test)

Deviations:

yes

Remarks:

Only 2 dose levels (instead of three) were used; Only 1000 polychromatic erythrocytes (instead of 2000) were analysed for the presence of micronuclei.

GLP compliance:

yes

Type of assay:

mammalian erythrocyte micronucleus test

Test material

Test material information

Constituent 1

Reference substance name:

N-cyclopropyl-1,3,5-triazine-2,4,6-triamine

EC Number:

266-257-8

EC Name:

N-cyclopropyl-1,3,5-triazine-2,4,6-triamine

Cas Number:

66215-27-8

Molecular formula:

C6H10N6

IUPAC Name:

N2-cyclopropyl-1,3,5-triazine-2,4,6-triamine

Test animals

Species:

mouse

Strain:

Tif:MAGf

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: female 6-10 weeks, male 4-9 weeks
- Weight at study initiation: female 27-31 g, male 33-35 g (tolerability test), female 22-29 g, male 26-36 g (mutagenicity test)
- Water: Tap water ad libitum
- Acclimation period: 4-6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22-23 °C
- Humidity (%): 41-66 %
- Photoperiod (hrs dark / hrs light): The room was illuminated for 12 hours daily

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

0.5% CMC

Frequency of treatment:

The animals were treated once.

Post exposure period:

maximum of 72 hours

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

1 080 mg/kg bw/day (nominal)

Remarks:

High dose

Doses / concentrations 2

Dose / conc.:

360 mg/kg bw/day (nominal)

Remarks:

Low dose

No. of animals per sex per dose:

8

Control animals:

yes, concurrent vehicle

Positive control(s):

Cyclophosphamide at 64 mg/kg bw.

Examinations

Tissues and cell types examined:

Bone marrow cells

Details of tissue and slide preparation:

Bone marrow was harvested from the shafts of both femurs with fetal calf serum. After centrifugation small drops of the sediment mixture were transferred on the end of a slide, spread out with the aid of a polished cover glass and the preparations were air-dried. Within 24 hours, the slides were stained in undiluted May-Grunwald solution for 3 min then in May-Grunwald solution/water 1/1 for 2 min. After being rinsed in distilled water, the slides were left immersed in diluted Giemsa solution for 10 min. After rinsing with distilled water and air-drying, the slides were cleared in xylene and mounted.

Evaluation criteria:

The test substance was considered to be active in this test system if a statistically significant increase in the number of polychromatic erythrocytes with micronuclei in comparison with the negative control occurred at any dose and sampling time respectively.

Statistics:

The significance of difference was assessed by X^2-test.

Results and discussion

Test results

Key result

Sex:

male/female

Genotoxicity:

negative

Toxicity:

yes

Vehicle controls validity:

valid

Negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

In the mutagenicity test one male animal of the 1080 mg/kg bw dose group died within the treatment period of 72 hours.

In the mutagenicity test, there was no significant increase in the number of micronucleated polychromatic erythrocytes in the animals treated with the dose of 360 mg/kg bw as well as with the dose of 1080 mg/kg bw as compared with the negative control animals.
By contrast, the positive control (cyclophosphamide) yielded a marked increase of the percentage of micronucleated cells. Here, the mean percentage of polychromatic erythrocytes with micronuclei was 1.99. In comparison with the negative control (0.10%) this value is highly significant (p<0.05).

Any other information on results incl. tables

In a preliminary assay 1080 mg/kg bw was determined to be the maximum tolerated dose. 

The results are provided in the table below.

 

Applicant's summary and conclusion

Conclusions:

Cyromazine did not induce any statistically significant increase in MPCE incidence in mice bone marrow cells in vivo up to and including 1080 mg/kg bw (the maximum tolerated dose).

Executive summary:

In an in vivo Micronucleus study, the test item was administered once by gavage at 1080 or 360 mg/kg bw to male and female mice. The animals were sacrificed 24, 48 and 72 hours after treatment. One male in the high dose group died. The bone marrow smears from treated animals showed no statistically significant increase (p > 0.05) in the number of micronucleated polychromatic erythrocytes in comparison with the negative control animals at all three sampling times. The respective "positive control" experiments with cyclophosphamide (64 mg/kg) yielded an average of 1.99% polychromatic erythrocytes with micronuclei. This is significantly different from the controls treated with the vehicle (0.5% CMC) alone.

 

It is concluded that under the conditions of this experiment, no evidence of mutagenic effects was obtained in mice treated with the test item.