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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
Remarks:
Type of genotoxicity: chromosome aberration
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Conducted for the NTP, using a rigorous protocol. Not GLP, no individual data presented.

Data source

Reference
Reference Type:
publication
Title:
Unnamed
Year:
2000

Materials and methods

Principles of method if other than guideline:
The assessment was made on the pheripheral blood of mice used in a 13 week toxicity study (NTP 2000), blood smears assessed for micronuclei
GLP compliance:
no
Type of assay:
micronucleus assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Refer to NTP2000/K2KS/repeat toxicity, mouse

Test animals

Species:
mouse
Strain:
Balb/c
Sex:
male/female
Details on test animals and environmental conditions:
Refer to NTP2000/K2KS/repeat toxicity, mouse

Administration / exposure

Route of administration:
oral: feed
Vehicle:
Placebo capsules
Duration of treatment / exposure:
Continuous exposure for 13 weeks.
Frequency of treatment:
Continuous, diet
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
5000 ppm
Basis:
nominal in diet
Remarks:
Doses / Concentrations:
10000 ppm
Basis:
nominal in diet
Remarks:
Doses / Concentrations:
20000 ppm
Basis:
nominal in diet
Remarks:
Doses / Concentrations:
40000 ppm
Basis:
nominal in diet
Remarks:
Doses / Concentrations:
80000 ppm
Basis:
nominal in diet
No. of animals per sex per dose:
5 males + 5 females
Control animals:
yes, concurrent vehicle
yes, plain diet

Examinations

Tissues and cell types examined:
Pheripheral blood
Details of tissue and slide preparation:
After 13 weeks of dietary treatment with 1,1,1-trichloroethane (NTP 2000) peripheral blood samples were obtained from male and female Balb C mice. Smears were prepared, fixed in methanol and then stained with acridine orange. Slides were scanned to determine the frequency of micronuclei in 2,000 normochromatic erythrocytes (NCEs) in each of 5 animals per exposure group.
Evaluation criteria:
An individual trial was considered positive if the trend test P value was less than or equal to 0.024 or if the P value for any single exposed group is less than or equal to 0.025 divided by the number of exposed groups. A final call of positive for micronucleus induction is preferable based on reproducibility positive trials. Ultimately, the final decision is determined by the scientific staff after considering the results of statistical analysis, the reproducibility of any effects observed and the magnitudes of those effects.
Statistics:
The frequency of micronucleated cells among NCEs was analysed by a statistical software package that tested for increasing trend over exposure groups with a one-tailed Cochran Armitage trend test, followed by pairwise comparisons between each exposure group and the control group. In the presence of excess binomial variation, as detected by a binomial dispersion test, the binomial variance of the Cochran-Armitage test was adjusted upward in proportion to the excess variation.

Results and discussion

Additional information on results:
No individaul data presented.

The results in male mice were considered equivocal because there was a statistically significant (P=0.013) trend across the dose groups, however, none of the individual values were significantly increased compared to the control. In females the frequency of normochromatic erythrocytes ate 2,000 ppm was elevated compared to the controls, but because the increase was small and there was no dose response, the results in females mice were negative.

Although no direct measurement of exposure was made administration of the test material was considered to be associated with a decrease in body weight gain and in the 13 week toxicity study (NTP 2000) the NOAEL was identified as 10,000 ppm in both sexes. This observation and the apparent equivocal effect on the bone marrow of 1,1,1-trichloroethane is considered to support systemic exposure of the mice during this test.

No positive control was used in this test and no discussion regarding the change in the context of background data was made.

Any other information on results incl. tables

 Table #:Results of in vivo micronucleus test

Dose ppm

No. of mice

Micronucleated NCEs/1000 NCEs

P value (vs control)

mean

SD

males

Untreated control

5

0.80

0.34

Vehicle control

5

0.90

0.19

5000

5

1.10

0.19

0.2519

20000

5

1.50

0.22

0.0520

40000

5

1.50

0.32

0.0520

80000

5

0.80

0.2

0.01515

P Value (trend)

0.013

females

Untreated control

5

0.60

0.19

Vehicle control

5

0.80

0.34

5000

5

0.80

0.12

0.3815

10000

5

1.20

0.12

0.0827

20000

5

1.70

0.25

0.0055

40000

5

1.20

0.25

0.0827

80000

5

1.20

0.20

0.0827

P value (trend)

0.116

NCE = normochromatic erythrocyte

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): ambiguous
Following 13 weeks dietary exposure to microencapsulated 1,1,1-trichloroethane, at doses between 5,000 and 80,000 ppm, examination of peripheral blood revealed and equivocal elevation of NCE’s in male mice and a negative response in female mice. The dose levels used were associated with a decrease in body weight gain, supporting systemic exposure to the test material.
Executive summary:

Following 13 weeks dietary exposure to microencapsulated 1,1,1-trichloroethane, at doses between 5,000 and 80,000 ppm, examination of peripheral blood revealed and equivocal elevation of micronucleated NCE’s in male mice and a negative response was recorded in female mice. 2000 normochromatic erythrocytes were exaimined for each aniamal. The dose levels used were associated with a decrease in body weight gain, supporting systemic exposure to the test  material.

In male mice there was a statistically significant rise (dose-related trend) in the number of micronucleated cells up to 40,000ppm.  The mean value at 80,000ppm was the same as the untreated control (0.80).  No group mean values were significantly increased compared to the control (dose-related trend only).  The current OECD guidelines sate that statistical significance should not be the only determining factor for a positive result.  The biological significance of the findings (for example background data) are not discussed in this paper.  It is therefore considered that the results give no clear evidence of mutagenicity.