Registration Dossier

Diss Factsheets

Toxicological information

Genetic toxicity: in vitro

Currently viewing:

Administrative data

Endpoint:
in vitro cytogenicity / chromosome aberration study in mammalian cells
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From Dec. 2, 2005 to Jul. 27, 2006
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study (OECD 473)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2006
Report date:
2006

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
Deviations:
no
GLP compliance:
yes
Type of assay:
in vitro mammalian chromosome aberration test

Test material

Constituent 1
Chemical structure
Reference substance name:
1H-1,2,4-triazole, sodium salt
EC Number:
255-280-9
EC Name:
1H-1,2,4-triazole, sodium salt
Cas Number:
41253-21-8
Molecular formula:
C2H3N3.Na
IUPAC Name:
sodium 1,2,4-triazol-1-ide
Details on test material:
- Lot/batch No.: 05.14
- Expiration date of the lot/batch: September 2006
- Storage condition of test material: At room temperature and protected humidity.
- Physical state: White powder
- Analytical purity: 99.2% (w/w)

Method

Species / strain
Species / strain / cell type:
lymphocytes: human lymphocytes
Details on mammalian cell type (if applicable):
- Periodically checked for karyotype stability: yes
Additional strain / cell type characteristics:
not applicable
Metabolic activation:
with and without
Metabolic activation system:
Aroclor 1254-induced rat liver microsomes
Test concentrations with justification for top dose:
1st experiment - S9 (3 hours): 0, 78.13, 156.3, 312.5, 625, 937.5, 1250, 2500, and 5000 µg/mL
2nd experiment -S9 (20 hours): 0, 312.5, 625, 937.5, 1250, 1875, and 2000 µg/mL
2nd experiment - S9 (44 hours): 0, 312.5, 625, 937.5, 1250, 1875, and 2000 µg/mL
1st experiment + S9 (3 hours): 0, 78.13, 156.3, 312.5, 625, 937.5, 1250, 2500, and 5000 µg/mL
2nd experiment + S9 (3 hours): 0, 312.5, 625, 937.5, 1250, 1875, and 2000 µg/mL
2nd experiment + S9 (3 hours): 0, 312.5, 625, 937.5, 1250, 1875, and 2000 µg/mL
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: water for injections
Controlsopen allclose all
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
Remarks:
water for injections
True negative controls:
no
Positive controls:
yes
Positive control substance:
mitomycin C
Remarks:
Migrated to IUCLID6: without S9 mix
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
cyclophosphamide
Remarks:
Migrated to IUCLID6: with S9 mix
Details on test system and experimental conditions:
1st experiment: Lymphocyte cultures were exposed to test article or control for 3 hours both in the absence or presence of S9.
Harvest time was 20 hours after the beginning of treatment.

2nd experiement: Without S9, cells were exposed continuously to the test or control item until harvest. With S9, cells were exposed to the test or control item for 3 hours then rinsed. Harvest time were 20 hours and 44 hours after the beginning of testing.

NUMBER OF REPLICATIONS: 2 independent experiments.

NUMBER OF CELLS EVALUATED: 200 metaphases/dose level.

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index

OTHER EXAMINATIONS:
- Determination of polyploidy: Yes
- Determination of endoreplication: Yes
Evaluation criteria:
The cytotoxicity of the test item was evaluated using the mitotic index. Analysis of 200 metaphases/dose level was made. The following structural aberrations were recorded for each metaphase: gaps, chromatid and chromosome breaks and exchanges, multiple aberrations and pulverizations.
Statistics:
Chi-square test.

Results and discussion

Test results
Species / strain:
lymphocytes: human lymphocytes
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
decrease of mitotic index
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Additional information on results:
-Experiments without S9 :
Cytotoxicity : Following the 3-hour treatment, a slight to strong toxicity was induced at dose-levels > or = 2500 µg/ml, as shown by 33-100% decrease in mitotic index. Following the 20-hour treatment, a slight to moderate toxicity was induced at dose-levels > or = 1875 µg/ml, as shown by 23-45% decrease in mitotic index. Following the 44-hour treatment, no decrease in mitotic index was noted.
Metaphase analysis : The dose-levels selected for metaphase analysis were : 312.3 , 625 and 1250 µg/ml for the 3-hour and 20-hour treatments, higher dose-levels inducing a significant increase in pH value; 1250 µg/ml for the 44-hour treatment, higher dose-levels inducing a significant increase in pH value. No significant increase in the frequency of cells with structural chromosomal aberrations was noted after 3-, 20- as well as 44-hour treatments.

-Experiment with S9 :
Cytotoxicity : At the 20-hour harvest time, a slight to strong toxicity was induced at dose-levels > 2000 µg/ml, as shown by 32-100% decrease in mitotic index. At the 44-hour harvest time, no decrease in mitotic index.
Metaphase analysis : the dose-levels selected for metaphase analysis were 312.5 , 625 and 1250 µg/ml for the 20-hour harvest time in both experiments, higher dose-levels inducing a significant increase in pH value; 1250 µg/ml for the 44-hour harvest time higher dose-levels inducing a significant increase in pH value.
No significant increase in the frequency of cels with structural chromosomal aberrations was noted in either experiment and at either harvest time.
The frequency of cells with structural chromosome aberrations of the vehicle and positive controls was as specified in acceptance criteria. the study was therefore considered valid.
Remarks on result:
other: strain/cell type: human lymphocytes
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

The test item was freely soluble in the vehicle (water for injections) at 100 mg/mL.

No precipitate was noted at the end of the treatement period, at any tested dose.

Due to the increase in pH observed in the treated dose-levels, pH measurements were peformed int reated medium at a time equivalent to the end of the treatment period. At 5000 µg/mL (the highest dose), the pH was approximately 9.7 (7.6 for the vehicle control) and the osmolarity equal to 390 mOsm/kg H2O (292 for the vehicle control).

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative

1,2,4-triazole did not induce chromosome aberrations in cultured human lymphocytes.The frequency of cells with structural chromosome aberrations of the vehicle and positive controls was as specified in acceptance criteria. the study was therefore considered valid.
Executive summary:

The study was performed according to the international guidelines (OECD 473) and in compliance with the principles of GLP regulations. 1,2,4 -triazole sodium salt was tested in two independent experiments, both with and without a liver metabolizing system (S9 mix), obtained from rats previously treated with Aroclor 1254.

The highest dose-level for treatment in the first experiment was selected in the basis of pH, osmolarity and solubility. For selection of the dose-levels for the second experiment, any toxicity indicated by the reduction of mitotic index in the first experiment was also taken into account. In the first experiment, lymphocyte cultures were exposed to the test or control item (with or without S9 mix) for 3 hours then rinsed. Cekks were harvested 20 hours after the beginning of treatment. The second experiment was performed as follow : without S9 mix, cells were exposed continuously to the test or control item until harvest ; and with S9 mix, cells were exposed to the test or control item for 3 hours and then rinsed.Cells were harvested 20 hours and 44 hours after treatment.

One and a half hours before harvest, each cuture was treated with a colcemid solution to block cells at the metaphase-stage of mitosis. 1,2,4 -triazole sodium salt was dissolved in water for injections. Positive controls were used.

No significant increase in the frequency of cells with structural chromosomal aberrations was noted in either experiment and at either harvest time, without and with S9 mix. The frequency of cells with structural chromosome aberrations of the vehicle and positive controls was as specified in acceptance criteria. the study was therefore considered valid.

1,2,4-triazole sodium salt did not induce chromosome aberrations in cultured human lymphocytes.