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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
Study period:
1990-03-14 to 1990
Reliability:
1 (reliable without restriction)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1990
Report date:
1990

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 486 (Unscheduled DNA Synthesis (UDS) Test with Mammalian Liver Cells in vivo)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.39 (Unscheduled DNA Synthesis (UDS) Test with Mammalian Liver Cells In Vivo)
Deviations:
no
GLP compliance:
yes
Type of assay:
unscheduled DNA synthesis

Test material

Constituent 1
Chemical structure
Reference substance name:
Sodium 3-(2H-benzotriazol-2-yl)-5-sec-butyl-4-hydroxybenzenesulfonate
EC Number:
403-080-9
EC Name:
Sodium 3-(2H-benzotriazol-2-yl)-5-sec-butyl-4-hydroxybenzenesulfonate
Cas Number:
92484-48-5
Molecular formula:
C16 H17 N3 O4 S .Na
IUPAC Name:
sodium 3-(2H-1,2,3-benzotriazol-2-yl)-5-(butan-2-yl)-4-hydroxybenzene-1-sulfonate
Details on test material:
- Name of test material: sodium 3-(2H-benzotriazol-2-yl)-5-sec-butyl-4-hydroxybenzenesulfonate (FAT 75'309/A)
- Physical state: solid

Test animals

Species:
rat
Strain:
Wistar
Sex:
male
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: SAVO-Ivanovas, med. Versuchstierzuchten GmbH, D-7964 Kisslegg, Germany
- Weight at study initiation: approximately 200 g
- Housing: single in Makrolon Type I cages, with wire mesh top and granulated soft wood bedding
- Diet (e.g. ad libitum): pelleted standard diet
- Water (e.g. ad libitum): tap water, ad libitum
- Acclimation period: minimum 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 +/- 3 degree C
- Humidity (%): 30 - 70%
- Photoperiod (hrs dark / hrs light): artificial light 6.00 a.m. - 6.00 p.m.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
- Vehicle(s)/solvent(s) used: carboxymethylcellulose suspension, 1%
- Justification for choice of solvent/vehicle: The vehicle is chosen according to its relative nontoxicity for the animals.
Duration of treatment / exposure:
in vivo treatment period: 12 - 14 hrs;
in vitro exposure time: 4 hrs
Frequency of treatment:
in vivo: the animals received the test article once
in vitro: three of the cultures from each animal were used for the UDS assay
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
10 mL/kg bw
Basis:
nominal conc.
in vivo
Remarks:
Doses / Concentrations:
100, 330, 1000 mg/kg bw
Basis:
actual ingested
in vivo
No. of animals per sex per dose:
5 males per dose group
Control animals:
yes, concurrent vehicle
Positive control(s):
2-acetylaminofluorene
- Route of administration: singly, orally
- Dosing: 100 mg/kg bw
- Volume administered: 10 mL/kg bw

Examinations

Tissues and cell types examined:
Hepatocytes
Details of tissue and slide preparation:
The animals were sacrificed by liver perfusion. After anesthetizing the rats with 1.5 ml/kg bw Hypnodil i.p. the liver was perfused through the venaportae with Hank's balanced salt solution (HBSS) supplemented with collagenase (0.05 % w/v) adjusted to pH 7.4 and maintained at 37 degree C. The hepatocytes were isolated from the liver and washed twice with HBSS. The crude cell suspension was filtered through a 96 um nylon mesh to yield a single cell suspension. The quality of the actual performed perfusion was determined by the trypan blue dye exclusion method. In addition the number of the isolated cells was determined.
Evaluation criteria:
A test article is classified as positive if it induces either a statistically significant dose-related increase in radiolabel incorporation expressed as grains per nucleus or a reproducible and statistically significant positive response for at least one of the test points. A test article producing neither a statistically significant dose related increase in radiolabel incorporation expressed as grains per nucleus nor a statistically significant and reproducible positive response at any one of the test points is considered non-effective in this system. However, both statistical and biological significance should be considered together.
Statistics:
Statistical significance can be evaluated by means of the nonparametric Mann-Whitney test.

Results and discussion

Test results
Sex:
male
Genotoxicity:
negative
Toxicity:
no effects
Vehicle controls validity:
valid
Positive controls validity:
valid
Additional information on results:
Up to the highest dose level tested no animal expressed toxic reactions. However, at the highest dose level (1000 mg/kg bw) the viability of the isolated hepatocytes of two animals was slightly decreased. The in vitro attachment of the hepatocytes was not affected due to the in vivo pre-treatment with the test article. The interindividual variations obtained for the numbers of isolated hepatocytes as well as for the attachment-efficiency (NR-assay) are in the range of our historical laboratory control. The NR-value obtained for animal no. 17 was relative low, if compared to the other animals. This was due to a technical error during the cell culture inocculation. In case of hepatotoxicity the NR-values expected would be approximately zero. No dose level of the test article revealed UDS induction in the hepatocytes of the treated animals as compared to the current negative controls. Neither the nuclear grains nor the resulting net grains were enhanced due to the in vivo treatment of the animals with the test article.
An appropriate reference mutagen (2-AAF, 100 mg/kg bw) was used as positive control. In vivo treatment with 2-AAF revealed distinct increases in the number of nuclear and net grain counts.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): negative
Under the conditions of this UDS test, sodium 3-(2H-benzotriazol-2-yl)-5-sec-butyl-4-hydroxybenzenesulfonate did not induce DNA-damage leading to repair synthesis in the hepatocytes of the treated rats.
Executive summary:

The test article sodium 3 -(2H-benzotriazol-2 -yl)-5 -sec-butyl-4 -hydroxybenzenesulfonate was assessed in the in vivo/ in vitro UDS assay for its potential to induce DNA repair (UDS) in the hepatocytes of rats. The assay was performed according to OECD Guideline 486 and EU method B.39. The test article was formulated in 1% carboxymethylcellulose suspension (1% CMC). 1% CMC was used as negative control. The volume administered orally was 10 mL/kg bw. After a treatment period of 12 - 14 hours the animals were narcotized and sacrificed by liver perfusion. Primary hepatocyte cultures were established and exposed for 4 hours to 3HTdR which is incorporated if UDS occurs. The test article was tested at the following dose levels: 100; 330; and 1000 mg/kg b.w. For each dose level, including the controls, hepatocytes from three treated animals were assessed for the occurrence of UDS. Up to the highest dose level tested no animal expressed toxic reactions. No dose level of the test article revealed UDS induction in the hepatocytes of the treated animals as compared to the current negative controls. An appropriate reference mutagen (2-AAF, 100 mg/kg bw) was used as positive control. Treatment with 2-AAF revealed distinct increases in the number of nuclear and net grain counts. Under the conditions of this UDS test, sodium 3-(2H-benzotriazol-2-yl)-5-sec-butyl-4-hydroxybenzenesulfonate did not induce DNA-damage leading to repair synthesis in the hepatocytes of the treated rats.