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:
From 18-MAY-1995 to 05-OCT-1995
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study performed similarly to OECD guidelines and according to GLP

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1995
Report Date:
1995

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Deviations:
yes
Remarks:
only 1000 cells scored per animal
GLP compliance:
yes
Type of assay:
micronucleus assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): dimethyl sulfide
- Substance type: pure active substance
- Physical state: clear, colorless liquid
- Stability under test conditions: data not available
- Storage condition of test material: room temperature, protected from exposure to light

Test animals

Species:
mouse
Strain:
ICR
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Sprague-Dawley, Inc., Frederick, MD
- Age at study initiation: 6 to 8 weeks old
- Weight at study initiation: males were 27.4 to 35.8 grams and females were 24.5 to 30.7 grams
- Assigned to test groups randomly: yes, according to a computer-generated program which is based on distribution according to body weight
- Fasting period before study: data not available
- Housing: mice of the same sex were housed up to five per cage in plastic autoclavable cages with filter tops. Heat-treated hardwood chips were used for bedding
- Diet: certificed laboratory rodent chow, ad libitum
- Water: tap water, ad libitum
- Acclimation period: animals were quarantined for no less than 5 days after receipt


ENVIRONMENTAL CONDITIONS
- Temperature: 74 +/- 6°F (23.3 °C)
- Humidity: 50 +/- 20%
- Air changes: data not available
- Photoperiod: 12 hour light/dark cycle


IN-LIFE DATES: data not available

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
- Vehicle(s)/solvent(s) used: corn oil
- Justification for choice of solvent/vehicle: the test article was soluble in corn oil at a concentration of 250 mg/mL, the maximum concentration tested
- Concentration of test material in vehicle: the maximum concentration was 250 mg/mL
- Amount of vehicle: 20 mL/kg
- Lot/batch no.: data not available
- Purity: data not available
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: data not available
Duration of treatment / exposure:
Test substance was given once and bone marrow cells were collected 24, 48 and 72 hours after treatment
Frequency of treatment:
once
Post exposure period:
bone marrow cells were collected 24, 48 and 72 hours after treatment
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 1250, 2500 and 5000 mg/kg
Basis:
nominal conc.
No. of animals per sex per dose:
5 animals per sex and per collection time
Control animals:
yes, concurrent vehicle
Positive control(s):
cyclophosphamide
- Justification for choice of positive control(s): cited in the OECD guideline
- Route of administration: oral gavage
- Doses / concentrations: 60 mg/kg

Examinations

Tissues and cell types examined:
Using oil immersion, 1000 polychromatic erythrocytes were scored for the presence of micronuclei (for each mouse and treatment group).
The proportion of polychromatic erythrocytes to total erythrocytes was also recorded per 1000 erythrocytes.
Details of tissue and slide preparation:
CRITERIA FOR DOSE SELECTION: based on a preliminary toxicity assay (in which no mortality was observed at 5000 mg/kg)


DETAILS OF SLIDE PREPARATION:
the mice were sacrificed by CO2 asphyxiation. The bone marrow cells were extracted and prepared. The slides were fixed in methanol, stained with May-Gruenwald-Giemsa and permanently mounted.
Evaluation criteria:
The test article was considered to induce a positive response if a treatment-related increase in micronucleated polychromatic erythrocytes was observed and one or more doses were statistically elevated relative to the vehicle control at any sampling time.
If a single treatment group was significantly elevated at one sacrifice time with no evidence of a dose-response, the assay was considered a suspect or unconfirmed positive and a repeat-assay recommended.
The test article was considered negative if no statistically significant increase in micronucleated polychromatic erythrocytes above the concurrent vehicle control was observed at any sampling time.
Statistics:
Statistically significance was determined using the Kastenbaum-Bowman tables which are based on the binomial distribution. All analyses were performed separately for each sex and sampling time.

Results and discussion

Test results
Sex:
male/female
Genotoxicity:
negative
Toxicity:
yes
Remarks:
lethargy in males and females exposed at 1250, 2500 and 5000 mg/kg
Vehicle controls valid:
yes
Negative controls valid:
not applicable
Positive controls valid:
yes
Additional information on results:
RESULTS OF RANGE-FINDING STUDY
- Dose range: for the toxicity study, dimethyl sulfide was administrated by oral gavage to male and female mice at 3000, 3500, 4000 and 5000 mg/kg
- Solubility: the test article was soluble in corn oil at the maximum test concentration of 250 mg/mL
- Clinical signs of toxicity in test animals: lethargy in males and females was observed at all dose levels on the day following dose administration
- Evidence of cytotoxicity in tissue analyzed: not examined


RESULTS OF DEFINITIVE STUDY
- Induction of micronuclei (for Micronucleus assay): the number of micronucleated polychromatic erythrocytes per 1000 polychromatic erythrocytes in test article treated groups was not statistically increased relative to their respective vehicle control in either male or female mice regardless of dose levels or bone marrow collection time. Cyclophosphamide induced a significant increase in micronucleated polychromatic erythrocytes in both male and female mice
- Appropriateness of dose levels and route: up to 5000 mg/kg

Any other information on results incl. tables

Table 1: summary of bone marrow micronucleus study using dimethyl sulfide

Treatment

Sex

Time (hr)

Number of mice

PCE / total erythrocytes

Micronucleated polychromatic erythrocytes

No. Per 1000 PCE’s (Mean)

No. Per PCE’s scored

Corn oil

M

24

48

72

5

5

5

0.53

0.58

0.57

0.8

1.4

0.8

4 \ 5000

7 \ 5000

4 \ 5000

F

24

48

72

5

5

5

0.70

0.59

0.61

2.0

1.0

1.2

10 \ 5000

5 \ 5000

6 \ 5000

1250 mg/kg

M

24

48

72

5

5

5

0.55

0.62

0.61

0.4

0.8

1.0

2 \ 5000

4 \ 5000

5 \ 5000

F

24

48

72

5

5

5

0.64

0.56

0.63

0.6

1.6

1.4

3 \ 5000

8 \ 5000

7 \ 5000

2500 mg/kg

M

24

48

72

5

5

5

0.56

0.56

0.63

1.0

1.6

0.6

5 \ 5000

8 \ 5000

3 \ 5000

F

24

48

72

5

5

5

0.66

0.55

0.60

0.4

0.6

1.4

2 \ 5000

3 \ 5000

7 \ 5000

5000 mg/kg

M

24

48

72

5

5

5

0.55

0.58

0.51

0.2

0.8

1.2

1 \ 5000

4 \ 5000

6 \ 5000

F

24

48

72

5

5

5

0.67

0.53

0.66

0.8

0.8

1.8

4 \ 5000

4 \ 5000

9 \ 5000

CP 60 mg/kg

M

24

5

0.54

17.4

87 \ 5000 *

F

24

5

0.46

34.4

172 \ 5000 *

* p= 0.05 (Kastenbaum-Bowman Tables)

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): negative
Dimethyl sulfide did not induce a significant increase in the incidence of micronucleated polychromated erythrocytes in bone marrow and was concluded to be negative in the micronucleus test using male and female ICR mice.
Executive summary:

In a ICR mouse bone marrow micronucleus assay performed according to the OECD guideline # c474 and GLP, 5 animals per sex per collection time and per dose level were treated once by oral gavage with dimethyl sulfide at doses of 0, 1250, 2500 and 5000 mg/kg bw.  Bone marrow cells were harvested at 24, 48 and 72 post-treatment.  The vehicle was corn oil.

Lethargy was observed during the study, in males and females at all dose levels. Reduction up to 11% in the ratio of polychromatic erythrocytes to total erythrocytes were observed in some of the test article treated groups relative to their respective vehicle control. Dimethyl sulfide was tested at an adequate dose (up to 5000 mg/kg). The positive control induced the appropriate response.  There was not a significant increase in the frequency of micronucleated polychromatic erythrocytes in bone marrow after any treatment time.