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Toxicological information

Genetic toxicity: in vivo

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Administrative data

Endpoint:
in vivo mammalian cell study: DNA damage and/or repair
Remarks:
Type of genotoxicity: DNA damage and/or repair
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP study, study design according to an internationally accepted protocol for the comet assay in vivo

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2005

Materials and methods

Principles of method if other than guideline:
Study was performed according to an internationally accepted protocol for the comet assay in vivo (Hartmann et al., Mutagenesis vol. 18, no. 1, 45-51, 2003).
GLP compliance:
yes
Type of assay:
mammalian comet assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Purity: 96.2% (analytical result dated August 19, 2005), white powder

Test animals

Species:
rat
Strain:
Wistar
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River, Sulzfeld, Germany
- Age at study initiation: 6 to 8 weeks
- Weight at study initiation: 218-272 g
- Housing: animals were kept individually in type IIA cages
- Water:ad libitum
- Acclimation period: at least 5 d


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 1.5°C
- Humidity (%): 40% to 70%
- Air changes (per hr): 10 times per hour
- Photoperiod: 12 hrs dark / 12 hrs light


Administration / exposure

Route of administration:
oral: gavage
Vehicle:
- Vehicle(s)/solvent(s) used:
Details on exposure:
The test substance was administered in a volume of 20 ml/kg.
Duration of treatment / exposure:
Number of application: 1, rats were sacrificed 3 h after administration.
Frequency of treatment:
Once
Post exposure period:
3 h
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 2000 mg/kg
Basis:
nominal conc.
No. of animals per sex per dose:
5 males/dose
Control animals:
yes, concurrent vehicle
Positive control(s):
400 mg/kg EMS (administration volume: 10 ml/kg)

Examinations

Tissues and cell types examined:
Liver and stomach
Details of tissue and slide preparation:
Liver perfusion in situ, preparation of hepatocytes
preparation of stomach cells
Evaluation criteria:
An assay is considered acceptable if the following criteria are met:
1. Tail length data obtained for a given treatment are acceptable as part of the evaluation if obtained from at least two slides and 100 cells per animal.
2. The positive control EMS should induce a mean tail length increase of at least 30 % compared to the mean tail length value of the vehicle control group.
A test was considered positive if there was a biologically relevant and statistically significant increase of the evaluated parameter(s) in tissue cells of treated animals in comparision to the negative control.
Statistics:
One-sided Wilcoxon rank test

Results and discussion

Test results
Sex:
male
Genotoxicity:
negative
Toxicity:
no effects
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
Results dose rage finding study:
on day 2 post administration 1 male showed piloerection. There were no gross pathological findings at necropsy. Histopathologically neither necrotic nor apoptotic cells were seen in stomach and liver cells; based on these results the limit dose of 2000 mg/kg was chosen for the comet assay in male rats.

Any other information on results incl. tables

Clinical Observations: no clinical findings after treatment with the test substance in any rat of the 2000 mg/kg treatment group Tissue Tissue Cytotoxicity:

   Liver    Stomach  
 Dose Group  Absolute Viability* [%]   Relative Viability# [%]   Absolute Viability* [%]  Relative Viability# [%] 
 Vehicle Control  75.4 100  80.5  100 
Vulcuren VP KA 9188200 mg/kg  77.7 101.7  77.5  96.3 
 Positive Control EMS400 mg/kg 75.2  98.4  68.4  85.0 

*= mean viability of cell preparation per dose group after perfusion #= relative to vehicle control animals The cell viability of vehicle controls was well within the range of defined cell quality criteria. The viability of liver, and stomach cells of rats exposed to Vulcuren VP KA 9188 was comparable to the respective control value. The same applied to the cells of the evaluated tissues of the positive control animals. Comet assay: Mean tail length per dose group

 Dose Group  Liver Mean Tail Length [µm] +/- SD Stomach  Mean Tail Length [µm] +/- SD
 Vehicle Control  22.23 +/- 1.7  21.12 +/- 0.8
 Vulcuren VP KA 91882000 mg/kg  24.12 +/- 2.2  23.07** +/- 1.5
 Positive ControlEMS 400 mg/kg  34.44** +/-2.3  33.48** +/- 3.0

** p<0.01 (one-sided Mann-Whitney-Wilcoxon)

After administration of 2000 mg/kg Vulcuren VP KA 9188 to male rats the tail lengths of liver cells was comparable to values of the cells of the respective tissue vehicle control animals. The mean comet tail length of stomach cells of the rats treated at 2000 mg/kg Vulcuren VP KA 9188 were statistically significantly different as compared to concurrent control mean value. However, the observed tail length values of stomach cells are comparable to historical control data (see overall remarks). In addition, the individual animal tail length data (see overall remarks) showed that the statistically significant of mean values is not biologically relevant.

In summary, this difference is thus not considered to be biologically relevant.

The tail length of the positive control rats was clearly increased demonstrating the sensitivity of the test system for the detection of genotoxic effects.

Applicant's summary and conclusion

Conclusions:
Interpretation of results: negative
Executive summary:

Male Wistar rats, which were treated with 2000 mg/kg Vulcuren VP KA 9188, were evaluated in the Comet assay. The treated rats showed no signs of toxicity. After single application of 2000 mg/kg test substance the tail lengths of liver cells as well the tail lengths of stomach cells were not biologically relevant increased. Based on these findings, the author concluded that Vulcuren VP KA 9188 is non-gentoxic in the comet assay in vivo to liver and stomach cells of male Wistar rats (Lanxess GmbH, 2005).