Registration Dossier

Administrative data

Endpoint:
in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report date:
2017

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Deviations:
no
GLP compliance:
yes
Type of assay:
mammalian erythrocyte micronucleus test

Test material

Constituent 1
Chemical structure
Reference substance name:
Zinc cyanide
EC Number:
209-162-9
EC Name:
Zinc cyanide
Cas Number:
557-21-1
Molecular formula:
Zn(CN)2
IUPAC Name:
zinc(2+) bis(iminomethanide)
Test material form:
solid: particulate/powder

Test animals

Species:
mouse
Strain:
Crj: CD(SD)
Sex:
male/female
Details on test animals or test system and environmental conditions:
Animal number: Dose range-finding test: male and female, 18 animals / Main test: male, 33 animals
Age: 6 weeks / 7 weeks old
Quarantine and acclimatation: all animals examined for quarantine at the time of receipt and be acclimatized to housing condition. The healthy animals selected and used for the study based on general health conditions.
Identification : the animal labeled on tail with permanent marker pen and identification cards attached to each cage for an individual discrimination
Group assignment: after acclimation periods, animals weighted and randomly assigned according to body weight average and standard deviation.
ENVIRONMENTAL CONDITIONS
Temperature: 22 ± 3°C
Relative humidity: 50 ± 20 %
Air exchange: 10-20 times/h
Light cycle: light 12h / dark 12h
Illumination: 150-300 Lux
Animal per cage: more than 3 animals

Administration / exposure

Route of administration:
oral: gavage
Duration of treatment / exposure:
The test substance was administrated twice to starved animals for 3-4 hours by oral gavage (sonde), and positive control was administrated with a single intraperitoneal injection at the same time as final (2nd) administration day.
Frequency of treatment:
It was injected twice (24 hour interval). However, positive control was administered one time on the last administration day.
No. of animals per sex per dose:
Dose range-finding test 1: 3
Dose range-finding test 2: 3
Dose range-finding test 3: 3
Main test: 5
Positive control(s):
Name of the substance: Cyclophosphamide monohydrate (CPA)
Supplier: Sigma-Aldrich, Inc.
Purity: 99.9%

Examinations

Tissues and cell types examined:
Bone marrow cells
Evaluation criteria:
- Discrimination of polychromatic Erythrocytes: polychromatic erythrocytes are determined by appearence of orange-fluorescent light without nuclei.
- Discrimination of nonchromatic erythrocytes: nonchromatic erythrocytes are determined by appearence of only their black shadows without fluorescent light.
- Criteria of micronucleus: SIZE: from the smallest distinguishable one to the one as large as a half the diameter of erythrocytes. SHAPE: mainly round and includes the form of donut shape, half moon shape and so on. COLOR: same color with near cell nucleus; green fluorescent in acridine orange.
Statistics:
The following statistic analyses were done using a SPSS program (ver. 19.). The result of the statistical evaluation was regarded significantly when the P value was less than 0.5.

Results and discussion

Test results
Key result
Sex:
male/female
Genotoxicity:
negative
Toxicity:
no effects
Negative controls validity:
valid
Positive controls validity:
valid

Applicant's summary and conclusion

Conclusions:
Under the experimental conditions, the results showed that there was no increase of MNPCE at any dose of the test substance compared to the negative control group, and also statistical significance was not observed. In addition, no statistical significance was observed in the value for the ratio of PCE to total erythrocytes between the test substance-dosed group and negative control group.
Therefore, the test substance, Zinc cyanide, was determined not to indice an increased frequency micronuclei in the bone marrow cells of male ICR mice under the present experimental condition.
Executive summary:

For that endpoint, one reliable study to assess in vivo genotoxicity on the registered substance on mammalian erythrocyte was available. The study was performed according to the OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test).

Under the experimental conditions, the registered substance " Zinc cyanide batch n°10185145" does not increase MNPCE at any doses tested compared to the negative control group. In addition, no statistical significance was observed in the value for the ratio of PCE to total erythrocytes between the test substance-dosed group and negative control group. Therefore, the registered substance " Zinc cyanide batch n°10185145" was determined not to induce an increased frequency micronuclei in the bone marrow cells of male ICR mice under these experimental conditions.

The validity criteria were successful and the study was therefore regarded as acceptable for that endpoint.