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Toxicological information

Genetic toxicity: in vivo

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in vivo mammalian germ cell study: cytogenicity / chromosome aberration
Type of genotoxicity: chromosome aberration
Type of information:
experimental study
Adequacy of study:
key study
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
limited but acceptable documented publication which meets basic scientific principles; however study design not in line with current guidelines (e.g. sampling time 1.5 normal cell cylce lengh, preparation time after colcemid or colchicine application ca. 3 to 5 h for mice, 5 animals per sex and dose group, measure of cytotoxicity)

Data source

Reference Type:
Assessment of in vivo chromosomal aberrations - potency of zinc mercapto benzo thiazole
Mohanan, P., V.; et al.
Bibliographic source:
Journal of Biomaterials Aplcations, volume 14,224-228

Materials and methods

Principles of method if other than guideline:
other: chromosomal aberration assay in Swiss albino mice
GLP compliance:
Type of assay:
mammalian bone marrow chromosome aberration test

Test material

Details on test material:
zinc mercapto benzo thiazole (ZMBT)
Specific details on test material used for the study:
provider: National Organic Chemical Industries Ltd., Chennai

Test animals

not specified
Details on test animals and environmental conditions:
- Age at study initiation: weaning Swiss albino mice
- Weight at study initiation: 16 to 20 g
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum

- Temperature (°C): 25 ± 2°C

Administration / exposure

Route of administration:
cotton seed oil, 1 mL/kg bw
Details on exposure:
90 minutes before sacrifice the animals were treated with colchicine (20 µg/kg bw) to arrest mitosis
Duration of treatment / exposure:
Frequency of treatment:
Post exposure period:
36 h
Doses / concentrationsopen allclose all
Dose / conc.:
24 mg/kg bw (total dose)
according to 0.480 mg/20 g animal
Dose / conc.:
43 mg/kg bw (total dose)
according to 0.86 mg/20 g animal
Dose / conc.:
96 mg/kg bw (total dose)
according to 1.920 mg/20 g animal
No. of animals per sex per dose:
4 per dose
Control animals:
yes, concurrent vehicle
Positive control(s):
methyl methane sulfonate (200 mg/kg bw)


Tissues and cell types examined:
bone marrow from femur
Details of tissue and slide preparation:
Both femora were removed through the pelvic bone, just below the knee. The bones were freed from muscles, and the proximal ends of the femora were carefully shortened with scissors. Saline (0.5 ml) was aspirated into the disposable syringe, and subsequently, the needle was inserted a few millimeters into the bone marrow canal. The bone marrow was flushed into centrifuge tubes, mixed thoroughly and were centrifuged. The cell button was collected, mixed with hypotonic solution (0.075 M KCl), and incubated for 20 min at 37°C. The samples were centrifuged again, and were than fixed with fixative (3:1 methanol acetic acid). Sample slides were prepared by flame drying and stained with Giemsa stain. Microscopic analysis was performed by bright field in oil under a light microscope. Evidence of chromosomal abnormalities was then evaluated; 100 well-spread metaphases were analyzed per animal per dose group, solvent control and positive control.

Results and discussion

Test results
not specified
not examined
Vehicle controls validity:
Negative controls validity:
not examined
Positive controls validity:

Any other information on results incl. tables

Chromosome number: number of chromosomes did not vary (no additional data)

The incidences of chromosome/chromatide abnormalities was comparable between treamtent groups and solvent control.

The postivie control induced significant number of chromosomal aberrations.

The authors conclude that the administration of ZMBT at different dose levels in Swiss mice did not increase the frequency of structural chromosomal aberrations and thus ZMBT did not induce structural chromosome aberrations in the bone marrow cells of Swiss mice under the experimental conditions used.

Table: Incidence of chromosomal aberrations after single ZMBT administration in the bone marrow of Swiss mice

              Average Abnormalities/100 plates  
        Chromatid*   Chromosome*  
 Group  Dose/animal (µg)  Gaps  Breaks  Gaps  Breaks  Other changes*
 I  1920  2  0  1  0  5
 II  960  1  1  0  0  3
 III  480  1  0  0  0  5
 IV  solvent control 1.0 ml  2  1  1  0  7
 V  positive control 4.0 mg**  17  6  11  8  19

* no additional data recorded

** methyl methane sulfonate (200 mg/kg bw)

Applicant's summary and conclusion

Executive summary:

The genotoxic potential of ZMBT was evaluated in an in vivo bone marrow chromosome aberration assay with limited documentation not in line with current guidelines. Swiss albino mice (4 animals per group) were administered with ca. 24, 43 and 96 mg/kg bw test substance, concurrent solvent control (cotton seed oil) and positive control (methyl methane sulphonate, 200 mg/kg) once via intraperitoneal injection. Clinical signs or other observations of toxicity were not recorded in the publication. Colchicine (20 µg/kg) was administered 90 minutes before scheduled sacrifice. All animals were sacrificed 36 hours after test sample injection. Bone marrow cells from both femora were prepared, fixed and stained. 100 well-spread metaphases were microscopically analysed for chromosomal aberrations (chromatic and chromosome gaps and breaks and other changes were recorded). No cytotoxicity parameters (e.g. mitotic index) were recorded. The incidences of chromatid and chromosome gaps and breaks documented in all treatment groups were comparable to the solvent control. The positive control methyl methane sulfonate (200 mg/kg bw) led to a distinct increase of chromatid and chromosome breaks. ZMBT did not induce structural chromosomal aberrations in bone marrow cells of Swiss mice under the experimental conditions used.