Freitag, 26. Februar 2016

Genetic toxicity: in vivo.001 to 008



http://www.echa.europa.eu/de/web/guest/registration-dossier/-/registered-dossier/15450/7/7/3/?documentUUID=b0282997-ea53-405f-afb7-1c1f4901056e



Administrative data

Study result type:
experimental result
Study period:
No data
Reliability:
4 (not assignable)
Rationale for reliability incl. deficiencies:
No GLP and no test guideline stated.

Data source

Reference
Reference Type:
publication
Title:
Unnamed
Year:
1985

Materials and methods

Principles of method if other than guideline:
Chlorine dioxide was evaluated for the induction of sperm-head abnormalities in mice following oral administration. When the substance is able to disrupt normal sperm morphology it can be considered as a measure of mutagenic potential to germ cells.
GLP compliance:
no data
Type of genotoxicity:
other:
Type of study:
other: Sperm-head abnormalities

Test materials

Identity of test material same as for substance defined in section 1 (if not read-across):
yes
Test material identityopen allclose all
Details on test material:
- Name of test material (as cited in study report): chlorine Dioxide (ClO2)
- Physical state: no data
- Lot/batch No.: no data
- Expiration date of the lot/batch: no data
- Stability under test conditions: assumed to be stable during the test (sponsor responsibility)
- Storage condition of test material: no data
- Other: Chlorine dioxide solutions were prepared by reacting sodium chlorite (NaClO2) with H2SO4, removing free chlorine with NaClO2 trap, and collecting the generated ClO2 in distilled water. The concentration of ClO2 was determined by absorbance at 359 nm. The concentrations were adjusted to 400 mg/L, 200 mg/L and 100 mg/L Cl equivalent in distilled water.

Test animals

Species:
mouse
Strain:
B6C3F1
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River breeding Laboratories, Inc.
- Age at study initiation: 8-11 week old
- Weight at study initiation: no data
- Assigned to test groups randomly: no data
- Fasting period before study: no data
- Housing: housed per groups, separated by sex and by treatment group
- Diet (e.g. ad libitum): Ad libitum, Purina Laboratory Chow
- Water (e.g. ad libitum): Ad libitum
- Acclimation period: no data

ENVIRONMENTAL CONDITIONS
no data

IN-LIFE DATES: From: To: no data

Administration / exposure

Route of administration:
oral: gavage
Vehicle(s):
Vehicle: Distilled water
Concentration in vehicle: ClO2 was adjusted to 400, 200 and 100 mg/L chlorine equivalents in distilled water (equivalent to 16.0, 8.0 and 3.2 mg/kg/day). 
Sodium chlorite solutions were prepared gravimetrically in distilled water at concentrations of 1000, 500 and 200 mg/L (equivalent to 40, 20 and 8.0 mg chlorite/kg/day).
Details on exposure:
Total volume applied: 1 ml
Duration of treatment / exposure:
No data
Frequency of treatment:
5 daily doses, 24 h apart
Post exposure period:
1, 3 and 5 weeks
Doses / concentrations
Doses / concentrations:
400, 200 and 100 mg/L chlorine equivalents (equivalent to 16.0, 8.0 and 3.2 mg/kg/day).
Basis:
actual ingested
No. of animals per sex per dose:
10 males per treatment group.
Control animals:
yes, concurrent vehicle
Positive control(s):
Positive control was ethyl methanesufonate (EMS) in deionised water at 200 mg/kg, administered IP in five daily doses. A positive control was used for each kill time.

Examinations

Tissues and cell types examined:
Tissue: Caudae Epididymes
Number of animals: 10 males
Number of cells: 1000 (500 by each of 2 readers)
Time points: 1, 3 and 5 weeks after final dose.
Type of cells Sperm
Parameters: Sperm head shape abnormalities using the categories of Wyrobek and Bruce (1975).
Details of tissue and slide preparation:
CRITERIA FOR DOSE SELECTION: no data

TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields):

DETAILS OF SLIDE PREPARATION: The caudae epididymes were dissected and placed into a petri dish containing 0.9% saline. After dicing with a scissors, the suspension was gently pipetted 5 to 6 times in and out of a 5- or 10 mL pipette. The suspension was strained trough a 80 µm silk mesh to remove tissue fragments, and 0.5mL of the filtrate was stained in a centrifuge tube with 0.05 mL 1% eosin Y. Slides were prepared from this suspension by spreading a drop over the slide with 3 passes of the edge of another slide. 

METHOD OF ANALYSIS: the sperm-heads shape abnormalities were determined using the categories of Wytobek and Bruce (1975).

OTHER:
Evaluation criteria:
No data
Statistics:
Student's t test or analysis of variance

Results and discussion

Test results
Sex:
male
Genotoxicity:
negative
Toxicity:
no data
Vehicle controls valid:
no data
Negative controls valid:
no data
Positive controls valid:
no data
Additional information on results:
Chlorine dioxide and chlorite did not induce significant increases in the level of sperm-head abnormalities.
Any other information on results incl. tables:
Table 7.6.2/1: Activity of chlorine dioxide and chlorite in the mouse sperm-head abnormality assay 3 weeks after the final exposure.


% abnormal sperm-heada: doseb
Sample
0
1:4
1:1
Undiluted
ClO2
1.22 ± 0.12
1.51 ± 0.17
1.54 ± 0.14
1.43 ± 0.17
ClO2-
3.24 ± 0.22
2.6 ± 0.22
1.96 ± 0.09
2.27 ± 0.20
ClO3-
3.33± 0.64
2.29± 0.30
3.02± 0.33
1.92± 0.28
EMS
18.36 +/- 4.47
EMS : Ethyl Methane sulfonate was administered as a positive control by IP injection at a dose of 200 mg/kg/day for 5 days
a : Values expressed as the mean percent of abnormal sperm-head per animal +/- SEM.
b: Concentrations of the undiluted samples: 400 mg/L for ClO2; 1000 mg/L for ClO2-and ClO3-

Applicant's summary and conclusion

Conclusions:
Chlorine dioxide did not induce sperm-head abnormalities in mice.
However, the biological significance of sperm-head abnormalities is not clear at present.
Executive summary:
In an in vivo sperm-head abnormalities study, male B6C3F1 mice were exposed to Chlorine Dioxide by gavage, at concentrations of 400, 200 and 100 mg/L of test substance (equivalent to 16.0, 8.0 and 3.2 mg/kg bw/day), for 5 succesive days. Then the animals were sacrificed
1, 3 or 5 weeks after the last administration and the sperm was recolted for each animal.
Concurrent vehicle animals were considered as a negative control and the positive control was ethyl methanesufonate (EMS) in deionised water at 200 mg/kg, administered IP in five daily doses.
Sperm head shape abnormalities using the categories of Wyrobek and Bruce (1975) were observed at 1, 3 and 5 weeks after final dose. The positive control gave acceptable results in terms of sperm-head abnormalities.
Genotoxicity results were negative, Chlorine dioxide did not induce significant increases in the level of sperm-head abnormalities. There were no toxicity data.
Chlorine dioxide and sodium chlorite did not induce sperm-head abnormalities in mice. However, the biological significance of sperm-head abnormalities is not clear at present.
Interpretation of results:
negative





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