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Toxicological information

Genetic toxicity: in vivo

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Administrative data

Endpoint:
in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
4 (not assignable)
Rationale for reliability incl. deficiencies:
secondary literature
Justification for type of information:
Data is from SCCS report

Data source

Reference
Reference Type:
secondary source
Title:
OPINION ON Basic Violet 2 COLIPA n° B115
Author:
European Commission (EC) - Scientific Committee on Consumer Safety (SCCS)
Year:
2011
Bibliographic source:
Scientific Committee on Consumer Safety SCCS, COLIPA n° B115, 13-14 December 2011

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Principles of method if other than guideline:
In vivo Mammalian Erythrocytes Micronucleus Test was performed ot determine the mutagenic nature of Basic violet 2
GLP compliance:
not specified
Type of assay:
other: In vivo Mammalian Erythrocytes Micronucleus Test

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material: Basic Violet 2
- IUPAC name: 4,4'-[(4-imino-3-methylcyclohexa-2,5-dien-1-ylidene)methylene]bis(2-methylaniline) hydrochloride
- Molecular formula: C22H23N3ClH
- Molecular weight: 365.906 g/mol
- Substance type: Organic
- Physical state: No data
Specific details on test material used for the study:
- Name of test material: Basic Violet 2
- IUPAC name: 4,4'-[(4-imino-3-methylcyclohexa-2,5-dien-1-ylidene)methylene]bis(2-methylaniline) hydrochloride
- Molecular formula: C22H23N3ClH
- Molecular weight: 365.906 g/mol
- Substance type: Organic
- Physical state: No data
- Purity: HPLC: 94%
- Impurities (identity and concentrations): No data

Test animals

Species:
mouse
Strain:
Swiss
Remarks:
CD-1
Details on species / strain selection:
No data
Sex:
male/female
Details on test animals and environmental conditions:
No data

Administration / exposure

Route of administration:
intraperitoneal
Vehicle:
- Vehicle(s)/solvent(s) used: Sterile distilled water
- Justification for choice of solvent/vehicle: The test chemical was soluble in sterile distilled water
- Concentration of test material in vehicle: 0, 3, 6 and 12 mg/kg bw
- Amount of vehicle (if gavage or dermal): No data
- Type and concentration of dispersant aid (if powder): No data
- Lot/batch no. (if required): No data
- Purity: No data
Details on exposure:
For oral route
PREPARATION OF DOSING SOLUTIONS: The test chemical was dissolved in sterile distilled water at dose levels of 0, 3, 6 and 12 mg/kg bw

DIET PREPARATION
- Rate of preparation of diet (frequency): No data
- Mixing appropriate amounts with (Type of food): No data
- Storage temperature of food: No data
Duration of treatment / exposure:
24 and 48 hrs
Frequency of treatment:
No data
Post exposure period:
No data
Doses / concentrations
Remarks:
0, 3, 6 and 12 mg/kg bw
No. of animals per sex per dose:
Total: 20 males and 20 females
0 mg/Kg bw: 5 males and 5 females
3 mg/Kg bw: 5 males and 5 females
6 mg/Kg bw: 5 males and 5 females
12 mg/Kg bw: 5 males and 5 females
Control animals:
yes, concurrent vehicle
Positive control(s):
In accordance with the OECD guideline

Examinations

Tissues and cell types examined:
Bone marrow cells
Details of tissue and slide preparation:
CRITERIA FOR DOSE SELECTION: Test doses were based on acute toxicity in a pretest
with 2 animals per sex/group.

TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields): 24 and 48h control and high dose group only

DETAILS OF SLIDE PREPARATION: Bone marrow cells were collected 24 h or 48 h (control and highest dose only) after dosing and stained with May-Gruenwald and Giemsa.

METHOD OF ANALYSIS: Toxicity and thus exposure of the target cells was determined by measuring the ratio between polychromatic and total erythrocytes (PCE/PCE+NCE) over the negative control value.

OTHER: No data
Evaluation criteria:
Toxicity and thus exposure of the target cells was determined by measuring the ratio between polychromatic and total erythrocytes (PCE/PCE+NCE) over the negative control value.
Statistics:
No data

Results and discussion

Test results
Sex:
male/female
Genotoxicity:
negative
Toxicity:
not specified
Vehicle controls valid:
yes
Negative controls valid:
yes
Positive controls valid:
yes
Remarks on result:
other: No mutagenic potential
Additional information on results:
RESULTS OF RANGE-FINDING STUDY
- Dose range: 3.91, 15.6, 125 mg/Kg bw
- Solubility: No data
- Clinical signs of toxicity in test animals: swollen abdomen, ataxia, hunched posture, lethargy, reduced activity, semi closed eyes and piloerection
- Evidence of cytotoxicity in tissue analyzed: No data
- Rationale for exposure: No data
- Harvest times: No data
- High dose with and without activation: >125 mg/Kg bw
- Other:

RESULTS OF DEFINITIVE STUDY
- Types of structural aberrations for significant dose levels (for Cytogenetic or SCE assay): No data
- Induction of micronuclei (for Micronucleus assay): No data
- Ratio of PCE/NCE (for Micronucleus assay): a reduction in the PCE/(PCE+NCE) ratio was observed for both sexes and both time points
- Appropriateness of dose levels and route:
Dose level: 0, 3, 6 and 12 mg/kg bw
Route: Intraperitoneal injection
- Statistical evaluation: No data

Applicant's summary and conclusion

Conclusions:
Basic Violet 2 did not induce an increase in bone marrow cells with micronuclei and, consequently, Basic Violet 2 is not genotoxic (clastogenic and/or aneugenic) in bone marrow cells of mice.
Executive summary:

In vivo mammalian erythrocytes micronucleus test was performed to determine the mutagenic nature of Basic violet 2. The study was performed using 5 males and 5 females Swiss CD-1 Mice at dose levels of 0, 3, 6 or 12 mg/Kg bw for 24 and 48 hrs. Three toxicity studies with decreasing doses were performed in order to identify the doses to be used in the main study. Animals were inspected for signs of reaction to treatment daily throughout the study. Bone marrow cells were collected 24 h or 48 h (control and highest dose only) after dosing and stained with May-Gruenwald and Giemsa. Toxicity and thus exposure of the target cells was determined by measuring the ratio between polychromatic and total erythrocytes (PCE/PCE+NCE) over the negative control value. In the micronucleus test no animals died. Animals from the 6 mg/Kg bw treatment group showed piloerection; those from the high dose group hunched posture, swollen abdomen, closed eyes, piloerection and pink spots in the cage litter indicative for excretion of coloured urine. A reduction in the PCE/(PCE+NCE) ratio was observed for both sexes and both time points. Biological relevant increases in the number of bone marrow cells with micronuclei compared to the concurrent vehicle controls were not found at any dose tested, either 24 or 48 h after treatment or for males and females.Basic Violet 2 did not induce an increase in bone marrow cells with micronuclei and, consequently, Basic Violet 2 is not genotoxic (clastogenic and/or aneugenic) in bone marrow cells of mice.