Sodium 1H-benzotriazolide

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in mammalian cells

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: OECD Guidance study with minor deviations

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1986

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)

Deviations:

yes

Remarks:

test article treatment is 4 hours instead of 5; whole fetal bovine serum is used instead of dialyzed calf serum

GLP compliance:

yes

Type of assay:

mammalian cell gene mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Benzotriazole

EC Number:

202-394-1

EC Name:

Benzotriazole

Cas Number:

95-14-7

Molecular formula:

C6H5N3

IUPAC Name:

1H-Benzotriazole

Constituent 2

Reference substance name:

1H-1,2,3-Benzotriazole

IUPAC Name:

1H-1,2,3-Benzotriazole

Test material form:

other: granules

Details on test material:

- Name of test material (as cited in study report): Benzotriazol Granulat
- Molecular formula (if other than submission substance): C6H5N3
- Molecular weight (if other than submission substance): 119.14 g/mol
- Smiles notation (if other than submission substance): c12c(cccc1)N=NN2
- InChl (if other than submission substance): 1S/C6H5N3/c1-2-4-6-5(3-1)7-9-8-6/h1-4H,(H,7,8,9)
- Structural formula attached as image file (if other than submission substance):
- Substance type: yellow granulat
- Physical state: solid
- Analytical purity: 99.83 %
- Lot/batch No.: 909

Method

Target gene:

HGPRT (Hypoxanthine guanine phophoribosyl transferase

Species / strain

Species / strain / cell type:

Chinese hamster Ovary (CHO)

Details on mammalian cell type (if applicable):

- Clone: CHO-K1BH4
- Type and identity of media: Ham's Nutrient Mixture F10 supplemented with L-glutamine, penicillin G, strptomycin sulfate, fungizone fetal bovine serum
- Periodically checked for Mycoplasma contamination: yes
- Periodically checked for karyotype stability: yes
- Periodically "cleansed" against high spontaneous background: yes

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with and without

Metabolic activation system:

S9 rat liver homogenate

Test concentrations with justification for top dose:

50 to 1000 µg/ml see tables

Vehicle / solvent:

DMSO

Controls

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

3-methylcholanthrene

other: 5-Bromo-2'-deoxyuridine

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium;

DURATION
- Exposure duration: 4 h
- Expression time (cells in growth medium): 6 to 7 days
- Selection time (if incubation with a selection agent): 10 days

SELECTION AGENT (mutation assays): 6-thioguanine
STAIN: Giemsa

NUMBER OF REPLICATIONS:

NUMBER OF CELLS EVALUATED:

Evaluation criteria:

Relative survival
Relative Population Growth
Absolute cloning efficiency
Mutant Frequency

Statistics:

mean values and standard deviation; significance calculated

Results and discussion

Test results

Test results 1

Species / strain:

Chinese hamster Ovary (CHO)

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

Chinese hamster Ovary (CHO)

Metabolic activation:

with

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Additional information on results:

RANGE-FINDING/SCREENING STUDIES:
concentrations ranging from 2 to 1000µg/ml were tested for cytotoxic effects.
Without activation, only in the highest concentration a decrease in the relative survival to 80 % was obtained.
With activation, a concentration of 125 µg/ml lead to a survival of 70 % and at the highest concentration the rate was 0%.

MUTAGENICITY STUDY:
Non activation
Three trials were performed under nonactivation conditions, but the first trial was not used for evaluation, because the cloning efficiency of the vehicle controls was too low. In the second trial the assayed cultures showed a low toxicity up to 1000 µg/ml. The lowest two concentrations were not assayed, because enough concentrations were left for cloning. None of the ten treated cultrues showed increases in mutant frequency that were statistically significantly elevated over the concurrent vehicle controls. Therefore, this nonactivation assay was evaluated as negative. In the independent repeat test concentrations from 400 to 1000 µg/ml were cloned, inducing low toxicities. The mutatnt frequencies varied for concentrations up to the maximum applied concentration of 1000µg/ml. The test material was therefore evaluated as nonmutagenic in this trial.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

In this study the toxicity and mutagenicity of Benzotriazole was assayed.
Under non-activating conditions only low toxicity could be induced up to the maximum applied concentration (1000 µg/ml).
With S9 metabolic activation the substance is more toxic to the tested cells as seen by decreases in relative survival and relative population growth.
In all assays the mutant frequencies were within or near the range that is typical of vehicle control variation between trials (1 * 10E6 - 15 * 10E6), while no dose-response relationships were evident.
Based on these conclusions, the substance 1H-benzotriazole is considered to be non-mutagenic.

Executive summary:

For Benzotriazole a well-conducted in vitro study is available showing no genetic toxicity for gene mutation in mammalian cells. This means that a similar result for Sodium Benzotriazolate can be anticipated.

Sodium Benzotriazolate is not expected to be mutagenic in vitro to mammalian cells based on a read across approach.

 

A DNEL for oral, dermal and/or inhalation route can be based on this information.

Classification and labelling are / are not needed for this endpoint.

A risk characterisation will be performed because the substance is classified for oral toxicity.