Registration Dossier

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:
2005
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Study performed according to OECD guideline 474 in compliance to GLP.

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
2005
Report Date:
2005
Reference Type:
other: Published secondary source
Title:
Unnamed
Year:
2006
Report Date:
2006

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
GLP compliance:
yes
Type of assay:
micronucleus assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Test item : HC Blue 2 No 2
EC number : 251-410-3
Batch number : 114B5
Content : 98.7%

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
0.5% aqueous carboxymethylcellulose
Duration of treatment / exposure:
The animals in the positive control, 500 and 1000 mg/kg groups were killed 24 hours after treatment.
5 males and 5 females were killed 24 hours after administration of the vehicle and 2000 mg/kg groups.
The remaining 5 males and 5 females were killed 48 hours after treatment.
Frequency of treatment:
Single dose
Post exposure period:
The animals in the positive control, 500 and 1000 mg/kg groups were killed 24 hours after treatment.
5 males and 5 females were killed 24 hours after administration of the vehicle and 2000 mg/kg groups.
The remaining 5 males and 5 females were killed 48 hours after treatment.
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
500 mg/kg
Basis:
actual ingested
Remarks:
Doses / Concentrations:
1000 mg/kg
Basis:
actual ingested
Remarks:
Doses / Concentrations:
2000 mg/kg
Basis:
actual ingested
No. of animals per sex per dose:
5 males and 5 females for the positive control, 500, 1000 mg/kg groups
10 males and 10 females for the control, 2000 mg/kg groups
Control animals:
yes, concurrent vehicle
Positive control(s):
Positive control : cyclophosphamide
Route of administration : Oral
Dose/concentration : 60 mg/kg administered once

Examinations

Tissues and cell types examined:
Bone marrow
Details of tissue and slide preparation:
For each animal, bone marrow smears were prepared and the micronucleated polychromatic erythrocytes were counted in 2000 polychromatic erythrocytes. The polychromatic/normochromatic erythrocyte ratio was established by scoring a total of 500 erythrocytes per animal.

Results and discussion

Test results
Sex:
male/female
Genotoxicity:
negative
Toxicity:
yes
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
No mortality occurred during the study. All animals in all dose groups showed purple urine and purple staining of one or more areas of the body on day 1 and day 2. This finding was not considered to be a sign of toxicity but was attributed to the colour of the test item. The maximum tolerated dose of 2000 mg/kg bw was achieved.

Statistically significant increases in micronucleus frequencies were observed in male rats at 500 and 2000 mg/kg bw. The increases were found to be within the laboratory historical control range and the micronucleus frequency in the vehicle control males was relatively low. The increased micronucleus frequency was not considered to be biologically relevant. No statistically significant differences were observed in the PCE/NCE ratio at any dose level. Analysis showed mean plasma levels of 18.9 ug/mL and 10.7 ug/mL 1 hour and 4 hours after oral administration at 2000 mg/kg bw respectively and confirmed the systemic exposure of the animals to the test item. Positive controls yielded significant increases in micronucleus frequencies in PCEs.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): negative
Under the experimental conditions reported, HC Blue No 2 did not induce biologically relevant micronucleus formation in the bone marrow cells of Sprague-Dawley rats treated orally up to the maximum recommended dose of 2000 mg/kg. The test substance was considered to be non-mutagenic in this micronucleus assay.