cyproconazole (ISO); (2RS,3RS;2RS,3SR)-2-(4-chlorophenyl)-3-cyclopropyl-1-(1H-1,2,4-triazol-1-yl)butan-2-ol

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:

3 Dec 1984 to 19 Dec 1984

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1985

Report date:

1985

Materials and methods

Test guideline

Test guideline 1

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Version / remarks:

1997

Deviations:

no

Test guideline 2

Qualifier:

equivalent or similar to guideline

Guideline:

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

Version / remarks:

2000

Deviations:

no

Test guideline 3

Qualifier:

equivalent or similar to guideline

Guideline:

EPA OPPTS 870.5395 (In Vivo Mammalian Cytogenetics Tests: Erythrocyte Micronucleus Assay)

Version / remarks:

1998

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

mammalian erythrocyte micronucleus test

Test material

Test material information

Constituent 1

Reference substance name:

2-(4-chloro-phenyl)-3-cyclopropyl-1-[1,2,4]triazol-1-yl-butan-2-ol

Cas Number:

94361-06-5

Molecular formula:

C15H18ClN3O

IUPAC Name:

2-(4-chloro-phenyl)-3-cyclopropyl-1-[1,2,4]triazol-1-yl-butan-2-ol

Test animals

Species:

mouse

Strain:

Swiss

Remarks:

random

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Weight at study initiation: not reported
- Housing: Animals were group-housed up to 20 mice per cage
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: At least 5 days

IN-LIFE DATES: 3 Dec 1984 to 19 Dec 1984

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

- Vehicle used: DMSO

Frequency of treatment:

Single dose

Post exposure period:

Test group: 24, 48 and 72 hours.
Control animals: 24 hours

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

16.7 mg/kg diet

Remarks:

Low dose

Doses / concentrations 2

Dose / conc.:

55.7 mg/kg diet

Remarks:

Mid dose

Doses / concentrations 3

Dose / conc.:

167 mg/kg diet

Remarks:

High dose

No. of animals per sex per dose:

5

Control animals:

yes, concurrent vehicle

Positive control(s):

- Positive control: Cyclophosphamide (CP)
- Route of administration: intraperitoneal injection
- Doses: 100 mg/kg

Examinations

Tissues and cell types examined:

Bone marrow, Polychromatic erythrocytes (PCEs) and the number of mature erythrocytes (RBCs)

Details of tissue and slide preparation:

BONE MARROW EXTRACTION:
Animals were killed with CO2 or by cervical dislocation and the adhering soft tissue and epiphyses of both tibiae were removed. The marrow was flushed from the bone and transferred to centrifuge tubes containing 3 mL fetal calf serum (one tube for each animal).

DETAILS OF SLIDE PREPARATION:
Following centrifugation to pellet the tissue, the supernatant was drawn off, the cells resuspended in a drop of serum, and the suspension spread on slides and air-dried. The slides were then fixed in methanol, stained in May-Gruenwald solution followed by Giemsa, and rinsed in deionized water (Schmid, 1975).

METHOD OF ANALYSIS:
One thousand PCEs per animal were scored. The frequency of micronucleated cells was expressed as percent micronucleated cells based on the total PCEs present in the scored optic field. The frequency of PCEs versus mature RBCs was determined by scoring the number of mature erythrocytes (RBCs) observed in the optic fields while scoring the first 1000 PCEs for micronuclei. For control of bias, all slides were coded prior to scoring.

Evaluation criteria:

The criteria for the identification of micronuclei were those of Schmid (1976). Micronuclei were darkly stained and generally round, although almond and ring-shaped micronuclei occasionally occur. Micronuclei had sharp borders and were generally between 1/20 and 1/5 the size of the PCE. The unit of scoring was the micronucleated cell, not the micronucleus; thus the occasional cell with more than one micronucleus was counted as one micronucleated PCE, not two (or more) micronuclei. The staining procedure permitted the differentiation by colour of polychromatic and normochromatic erythrocytes (bluish-grey and red, respectively).

Statistics:

Two-tailed student's t-test

Results and discussion

Test results

Key result

Sex:

male/female

Genotoxicity:

negative

Toxicity:

no effects

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

Animals exposed to test substance showed no significant increase in micronucleus frequency at any dose level, kill-time or sex. In addition no increase in the numbers of micronuclei was observed when the data for both sexes were pooled. The positive control, cyclophosphamide, induced a large and statistically significant increase in the frequencies of micronuclei (p ≤ 0.02 for males, p ≤ 0.01 females).

Any other information on results incl. tables

Table 1. Mean percentage (±SD) of micronucleated PCEs in mouse bone marrow

Test substance (concentration)	Sex	24 hours	48 hours	72 hours
167 mg/kg	Males Females Mean	0.32 ± 0.12 0.10 ± 0.06 0.21 ± 0.07	0.28 ± 0.11 0.06 ± 0.05 0.16 ± 0.06	0.40 ± 0.40 0.18 ± 0.08 0.24 ± 0.11
55.7 mg/kg	Males Females Mean	0.30 ± 0.14 0.16 ± 0.05 0.23 ± 0.07	0.16 ± 0.11 0.06 ± 0.02 0.11 ± 0.06	0.25 ± 0.07 0.10 ± 0.00 0.17 ± 0.04
16.7 mg/kg	Males Females Mean	0.14 ± 0.05 0.26 ± 0.14 0.20 ± 0.07	0.44 ± 0.20 0.26 ± 0.14 0.35 ± 0.12	0.24 ± 0.14 0.18 ± 0.04 0.21 ± 0.07
Negative control Vehicle (DMSO)	Males Females Mean	0.10 ± 0.05 0.34 ± 0.06 0.22 ± 0.05
Positive control Cyclophosphamide (100 mg/kg)	Males Females Mean	1.94 ± 0.41* 1.56 ± 0.25** 1.75 ± 0.24**

*) p ≤ 0.02 **) p ≤ 0.01

Applicant's summary and conclusion

Conclusions:

The test substance did not induce any significant increase in micronucleus frequencies in bone marrow polychromatic erythrocytes of the mouse. Therefore, the test substance was considered inactive under the conditions of this assay

Executive summary:

The test substance was evaluated for its ability to induce micronuclei in polychromatic erythrocytes (PCE’s) of male and female Swiss mouse bone marrow in a study following GLP principles and performed similar to OECD TG 474. Dose selection was based upon the LD50 (200 mg/kg for male mice and 218 mg/kg for female mice), using 80% of the combined LD50 for both sexes. Groups of 5 animals per sex per dose were received a single dose of 167.0, 55.7 and 16.7 mg/kg bw via oral gavage. The test substance was dissolved in Dimethylsulfoxide (DMSO). The positive control received an intraperitoneal injection of Cyclophosphamide (CP) at 100 mg/kg. Groups of animals were killed 24, 48 and 72 hours after administration of the test substance, the vehicle control and the positive control.

Results showed that animals exposed to the test substance showed no significant increase in micronucleus frequency at any dose level, kill time or sex. Also, no elevation of the numbers of micronuclei was observed if the data for the sexes are pooled. The positive control compound, cyclophosphamide (CP), induced large and statistically significant increases in the frequencies of micronuclei (p 0.02 ≤ males, p ≤ 0.01 females).

The test substance did not induce any significant increase in micronucleus frequencies in bone marrow polychromatic erythrocytes of the mouse. The test substance showed no mutagenicity under the conditions of this assay.