Methanesulphonyl chloride

Genetic toxicity: in vivo

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2003

Report date:

2003

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

EU Method B.12 (Mutagenicity - In Vivo Mammalian Erythrocyte Micronucleus Test)

Version / remarks:

Cited as Directive 2000/32/EC, B.12

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Deviations:

no

GLP compliance:

yes

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

Methanesulphonyl chloride

EC Number:

204-706-1

EC Name:

Methanesulphonyl chloride

Cas Number:

124-63-0

Molecular formula:

CH3ClO2S

IUPAC Name:

methanesulfonyl chloride

Details on test material:

Test article name : Methane sulfonyl chloride
Cas n°: 124-63-0
Origin: Atofina NA, Riverview, USA
Batch: 1491F10CD1
Purity: 99.9 %

Test animals

Species:

mouse

Strain:

Swiss

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST SYSTEM
Animals:
Strain: Swiss Ico: OF1 (IOPS Caw).
Breeder: Charles River Laboratories France, l'Arbresle, France.
Age: 6 weeks old on the day of treatment
Acclimation: at least 5 days

ENVIRONMENTAL CONDITIONS:
- temperature: 22 ± 2°C,
- relative humidity: 30 to 70%,
- light/dark cycle: 12 h/12 h (07:00 - 19:00),
- ventilation: at least 12 cycles/hour of filtered non-recycled fresh air.

Housing:
The animals were housed by groups in polycarbonate cages.

Food and water:
Food: A04 C pelleted maintenance diet (SAFE, Villemoisson-sur-Orge, France), ad libitum.
Water: drinking water filtered by a FG Millipore membrane (0.22 micron), ad libitum.

Administration / exposure

Route of administration:

intraperitoneal

Vehicle:

- Vehicle(s)/solvent(s) used: paraffin oil
- Justification for choice of solvent/vehicle: good solubility
- Concentration of test material in vehicle: the test item was supplied as preparations at 200.25 mg/mL and 20.03 mg/mL in
paraffin oil. The latter was used directly for treatment or diluted in the vehicle immediately before use, depending on the target dose-level..
- Amount of vehicle (if gavage or dermal): 10 ml/kg

Details on exposure:

EXPERIMENTAL DESIGN
Range-finding toxicity assays:
In order to select the top dose-level for the cytogenetic study, 200.3, 50 or 30 mg/kg/day were administered twice, to three males and three females. The interval between each administration was 24 hours. Clinical signs and any mortality were recorded for a period of 48 hours.

Micronucleus assay:
Administration:
Three groups of five male and five female mice were given intraperitoneal administrations (10 ml/kg) of Methane Sulfonyl Chloride at dose-levels of 0 (paraffin oil), 7.5,15 and 30 mg/kg/day, over a 2-day period. One group of five males and five females received the positive control test item (cyclophosphamide) once by oral route at the dose-level of 50 mg/kg.

Duration of treatment / exposure:

2 administrations at 24 hours interval

Frequency of treatment:

daily

Post exposure period:

24 h

Doses / concentrations

Remarks:

Doses / Concentrations:
7.5, 15 and 30 mg/kg/day
Basis:

No. of animals per sex per dose:

5 or 8

Control animals:

yes, concurrent vehicle

Positive control(s):

The positive control was cyclophosphamide (CPA) dissolved in distilled water at a concentration of 5 mg/mL.

Examinations

Tissues and cell types examined:

Bone marrow

Details of tissue and slide preparation:

Preparation of the bone marrow smears:
The animals of the treated and vehicle control groups were killed 24 hours after the last treatment and the animals of the positive control group were killed 24 hours after the single treatment. The femurs of the animals were removed and the bone marrow was flushed out using fetal calf serum. After centrifugation, the supernatant was removed and the cells in the sediment were resuspended by shaking. A drop of this cell suspension was placed and spread on a slide. The slides were air-dried and stained with Giemsa. The slides were coded so that the scorer is unaware of the treatment group of the slide under evaluation ("blind" scoring).

Microscopic examination of the slides:
For each animal, the number of the micronucleated polychromatic erythrocytes (MPE) was counted in 2000 polychromatic erythrocytes; the polychromatic (PE) and normochromatic (NE) erythrocyte ratio was established by scoring a total of 1000 erythrocytes (PE + NE).

Evaluation criteria:

For a result to be considered positive, a statistically significant increase in the frequency of MPE must be demonstrated when compared to the concurrent vehicle control group. Reference to historical data, or other considerations of biological relevance was also taken into account in the evaluation of data obtained.

Statistics:

When there was no significant within-group heterogeneity, using the heterogeneity chi-square test value (Lovell et al., 1989), the frequencies of MPE in each treated group was compared with those in the concurrent vehicle control groups by using a 2 x 2 contingency table to determine the X2 value (Lovell et al., 1989).
When there was significant within-group heterogeneity, then that group was compared with the control group using a non-parametric analysis, the Mann-Whitney test (Schwartz, 1969).
The student "t" test was used for the PE/NE ratio comparison.
Probability values of p <= 0.05 was considered as significant.

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

negative

Toxicity:

yes

Remarks:

Hypoactivity and piloerection were noted in all treated animals at 30 mg/kg/day

Vehicle controls validity:

valid

Negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

PRELIMINARY TOXICITY TEST
At 200.3 mg/kg/day, 3/3 males and 2/3 females dead in the 24 hours following the first treatment.
At 50 mg/kg/day, hypoactivity and piloerection were noted in all treated animals. At the end of the observation period no mortality was noted. However, at the time of sacrifice of the animals 24 hours later, 3/6 treated animals were found dead showing the high toxicity of this
dose.
At 30 mg/kg/day, hypoactivity and piloerection were noted in all treated animals but no mortality was observed.

CYTOGENETIC TEST
No clinical signs and no mortality were observed in the animals of both sexes given 7.5 or 15 mg/kg/day. Hypoactivity and piloerection were noted in all treated animals at 30 mg/kg/day.

For both males and females, the mean values of MPE as well as the PE/NE ratio in the groups treated with the test item, were equivalent to those of the vehicle group.

Cyclophosphamide induced a highly significant increase (p < 0.001) in the frequency of MPE, indicating the sensitivity of the test system under our experimental conditions. The study was therefore considered valid.

Any other information on results incl. tables

Results of the cytogenetic test: data summary

Group	Doses	MPE/1000PE		PE/NE ratio		Time of sacrifice after the last administration
	(mg/kg/day)	mean	(sd)	mean	(sd)	24 h
Males
Vehicle	-	1.7	(0.6)	0.3	(0.1)
Test item	7.5	1.4	(1.0)	0.4	(0.1)
	15	1.1	(0.7)	0.5	(0.2)
	30	0.9	(0.4)	0.3	(0.1)
Cyclophosphamide	50 mg/kg	18.6	(7.1)***	0.5	(0.1)
Females						24 h
Vehicle	-	0.9	(0.5)	0.6	(0.2)
Test item	7.5	0.9	(1.1)	0.8	(0.2)
	15	1.5	(0.6)	0.4	(0.1)
	30	0.6	(0.8)	0.5	(0.1)
Cyclophosphamide	50 mg/kg	17.1	(5.4)***	0.6	(0.1)

MPE: Micronucleated Polychromatic Erythrocytes

PE: Polychromatic Erythrocytes

NE: Normochromatic Erythrocytes

sd: Standard Deviation

*** p < 0.001

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): negative
Methane Sulfonyl Chloride (purity: 99.9%) did not induce damage to the chromosomes or the mitotic apparatus of mice bone marrow cells after two intraperitoneal administrations, with a 24-hour interval, at the dose-levels of 7.5, 15 and 30 mg/kg/day.

Executive summary:

The potential of methanesulfonyl chloride (purity 99.9%) to induce structural or numerical damage in bone marrow cells of mice was evaluated a study performed according to the OECD guideline #474 and GLP.

Three groups of five male and five female mice were given intraperitoneal administrations (10 ml/kg) of methanesulfonyl chloride at dose-levels of 0 (paraffin oil), 7.5,15 and 30 mg/kg/day, over a 2-day period. One group of five males and five females received the positive control test item (cyclophosphamide) once by oral route at the dose-level of 50 mg/kg.

Animal were sacrificed 24 hours after the last administration, and bone marrow smears examined for the presence of micronuclei in 2000 polychromatic erythrocytes per mouse and for the ratio PCE/NCE.

For both males and females, the mean values of MPE as well as the PE/NE ratio in the groups treated with the test item, were equivalent to those of the vehicle group. Methanesulfonyl chloride was concluded to be negative in the mouse micronucleus assay.