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:
25.9.-17.10.2000
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: This study was carried out in accordance with internationally valid GLP principles.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2000
Report Date:
2000

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Deviations:
not specified
GLP compliance:
yes
Type of assay:
micronucleus assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid: viscous
Details on test material:
Details on test materials:
Name of test material (as cited in study report): Solvent Yellow 124
Substance type: organic
Physical state: liquid
Batch No.: S0455/2000

Test animals

Species:
mouse
Strain:
NMRI
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: M & B A/S, DK-8680 Ry
- Age at study initiation: 7 weeks
- Weight at study initiation: 28-33g
- Assigned to test groups randomly: yes, using a randomisation scheme
- Fasting period before study: no data
- Housing: groups of 2 or 5 in transparent polycarbonate (macrolone type III) cages ( floor area: 810 cm2)
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum, domestic drinking quality acified with HCl to pH 2.5 in order to prevent microbial growth
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21°C±3°C
- Humidity (%): 55% ±15%
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
sezame oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
Solutions were prepared in warm sezame oil aprox. 37°C at concentrations 50, 100, and 200 mg/ml fresly before use. These solutions were used to dose the mice at 500, 1000, and 2000 mg/kg using a fixed dose volume of 10 ml/kg.
Duration of treatment / exposure:
24,48 hours
Doses / concentrations
Remarks:
Doses / Concentrations:
2000 mg/kg
Basis:
actual ingested
No. of animals per sex per dose:
5 males (24 hours) and 5 males (48 hours)
Control animals:
yes, concurrent vehicle
Positive control(s):
cyclophosphamide
- Route of administration: oral (gavage)
- Doses / concentrations: 20 mg/kg

Examinations

Tissues and cell types examined:
• A bluish mauve strongly coloured uniform circular particle in the cell
• The particle should have a certain size (not being punctiform) and it should be located in the same plane as the cell (cell and the micronucleus should be in focus at the same time)
• During focusing, the particle should stay uniform in colour, light refraction and shape within a relatively large interval
• Cells with 2 or more micronuclei were counted as single micronucleated cells
Details of tissue and slide preparation:
CRITERIA FOR DOSE SELECTION: In the preliminary toxicity test, no adverse reactions to treatment was observed at 500, 1000, and 2000 mg/kg body weight. Accordingly 2000 mg/kg was selected for the main test as the maximum dose level required by OECD quideline for materials of low toxicity.

DETAILS OF SLIDE PREPARATION:
The cell suspension bone marow from each femur in 2.5 ml of foetal calf serumwas centrifuged for 10 minutes at 1000 rpm and most of the supernatant was removed. The cells were resuspesed in the remainder and smeared on clean glass slides. The slide prepartion was fixed in methanol and stained with Giemsa. The slides were dried and coverslips were aplied using Dammarxylen® mountant
Evaluation criteria:
• increases in the frequency of micronucleated polychromatic erythrocytes were observed in treated animals compared to the corresponding negative controls
• the increases were reproducible between the animals of each group
• the increases were statistically significant
• the increases exceeded historical control range for this laboratory
Statistics:
SAS® procedures (v 6.12) described in “SAS®/STAT User´s Guide, Version 6, Fourth Edition, Vol. 1+2”, 1989, SAS Institute Inc., Cary, North Carolina 27513, USA

Results and discussion

Test results
Sex:
male
Genotoxicity:
negative
Toxicity:
no effects
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): negative
It is concluded that Solvent Yellow 124 showed no evidence of mutagenic or clastogenic activity in this mouse micronucleus test after administration by oral gavage.
Executive summary:

Solvent Yellow 124 was tested in the Mouse Micronucleus Test performed in accordance with the OECD quideline “Mammalian Erythrocyte Micronucleus Test”, No 474 (1997).

 

In the main test, a group of male mice was treated with a solution of the test article in sesame oil at a dose level of 2000 mg/kg. A negative control group was dosed with sesame oil, and a positive control group was dosed with Cyclophosphamide at 20mg/kg. All the mice were dosed by oral gavage on one occasion using a dose volume of 10 ml/kg body weight.

 

Bone marrow samples were taken from five mice from each of the three groups 24 hours after dosing and from five mice of the test article treatment and negative control groups 48 hours after dosing. Bone marrow smears were prepared on glass slides, stained, and scored using a microscope.

 

No adverse clinical sings were observed in the nice. All mice treated with the test article lost weight during the treatment period. The urine of mice treated with the test article was dark yellow two hours after dosing. The internal organs of these mice were also coloured yellow after 48 hours after they had been dosed. These observations demonstrate systemic distribution of the test article and/or its metabolite s, and hence, exposure of the bone marrow. No reduction in the frequency of polychromatic erythrocytes among total erythrocytes was observed after treatment with Solvent Yellow 124, indicating that there was no toxic effect on the bone marrow.

 

No biologically or statistically significant increases in the frequency of micronucleated polychromatic erythrocytes were seen in mice treated with Solvent Yellow 124.

 

A large statistically significant increase in the frequency of micronucleated polychromatic erythrocytes was observed in the positive control group, demonstrating the sensitivity of the test system.

 

It is concluded that Solvent Yellow 124 showed no evidence of mutagenic or clastogenic activity in this mouse micronucleus test after administration by oral gavage.