Tellurium

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:

key study

Study period:

2012-09-17 to 2012-11-28

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.17 (Mutagenicity - In Vitro Mammalian Cell Gene Mutation Test)

GLP compliance:

yes (incl. QA statement)

Type of assay:

mammalian cell gene mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Tellurium dioxide

EC Number:

231-193-1

EC Name:

Tellurium dioxide

Cas Number:

7446-07-3

Molecular formula:

O2Te

IUPAC Name:

oxotellane oxide

Constituent 2

Reference substance name:

oxotellan oxide

IUPAC Name:

oxotellan oxide

Test material form:

solid: particulate/powder

Details on test material:

- Name of test material (as cited in study report): Tellurium dioxide
- Substance type: inorganic

Method

Species / strain

Species / strain / cell type:

mouse lymphoma L5178Y cells

Details on mammalian cell type (if applicable):

The original L5178Y TK+/- 3.7.2 C mouse lymphoma cell line was obtained from the American Type Culture Collection. Cells were stored as frozen stocks in liquid nitrogen. Each batch of frozen cells was purged of TK-/--mutants and checked for the absence of mycoplasma.

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with and without

Metabolic activation system:

rat liver S9-mix (Phenobarbital and ß-naphthoflavone induced).

Test concentrations with justification for top dose:

Assay 1: 3-hour treatment with metabolic activation: 100; 75; 50; 25; 20; 15; 10; 7.5; 5; 2.5; 1.25 and 0.625 µg/mL
Assay 1: 3-hour treatment without metabolic activation: 80; 70; 60; 50; 40; 30; 20; 10; 5; 2.5; 1.25 and 0.625 µg/mL
Assay 2: 3-hour treatment with metabolic activation: 20; 17.5; 15; 12.5; 10; 7.5; 5; 2.5; 1.25 and 0,625 µg/mL
Assay 2: 24-hour treatment without metabolic activation: 15; 12.5; 10; 9; 8; 7; 6; 5; 4; 2; 1; 0.5 and 0.25 µg/mL

Vehicle / solvent:

1 % (w/v) methyl cellulose solution

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

cyclophosphamide

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium

DURATION

- Exposure duration: 3 and 24 hours
- Expression time (cells in growth medium): 3 days
- Selection time (if incubation with a selection agent): two weeks


SELECTION AGENT (mutation assays): 5-trifluorothymidine (TFT)

NUMBER OF REPLICATIONS: duplicate cultures

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth

OTHER EXAMINATIONS:
- Other: smal and large colonies

Evaluation criteria:

The assay was considered valid if all of the following criteria were met:
1. The mutant frequency in the negative (vehicle) control cultures fall within the normal range (50-170 mutants per 106 viable cells).
2. The positive control chemicals induce a statistically significant increase in the mutant frequency.
3. The plating efficiency (PEviability) of the negative (vehicle) controls is within the range of 65% to 120% at the end of the expression period.
4. At least four test concentrations are present, where the highest concentration produces approximately 80-90% toxicity (measured by %RS or RTG), results in precipitation, or it is 5 mg/mL, 5 µL/mL or 0.01 M (whichever is the lowest), or it is the highest practical concentration.

The test item was considered to be mutagenic in this assay if all the following criteria were met (based on M. Moore 2006 [6]):
1. The assay is valid.
2. Statistically significant (p < 0.05) and biologically relevant increases in mutation frequency are observed in treated cultures compared to the
corresponding negative (vehicle) control values at one or more concentrations.
3. The increases in mutation frequency are reproducible between replicate cultures and/or between tests (under the same treatment conditions).
4. There is a significant concentration-relationship as indicated by the linear trend analysis (p < 0.05).
5. The mutation frequency at the test concentration showing the largest increase is at least 126 mutants per 106 viable cells
(GEF = the Global Evaluation Factor) higher than the corresponding negative (vehicle) control value.
Results, which only partially satisfied the acceptance and evaluation criteria, were evaluated on a case-by-case basis.

Statistics:

Statistical significance of mutant frequencies (total wells with clones) was performed using Microsoft Excel software. The control log mutant frequency (LMF) was compared to the LMF from each treatment dose, based on Dunnett's test for multiple comparisons and the data checked for a linear trend in mutant frequency with treatment dose using weighted regression. The test for linear trend was one-tailed, therefore negative trend was not considered significant. These tests required the calculation of the heterogeneity factor to obtain a modified estimate of variance.

Results and discussion

Test results

Species / strain:

mouse lymphoma L5178Y cells

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: no large changes in pH
- Effects of osmolality: no large changes in osmolarity
- Water solubility: insoluble
- Precipitation: insolubility was detected in the final treatment medium at the end of the treatment at some concentrations

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

In Assay 1, following a 3-hour treatment with metabolic activation, excessive cytotoxicity of the test item was observed at 100, 75, 50, 25 and 20 µg/mL concentrations, cells of these samples did not survive the expression period. Marked cytotoxicity was observed at 15 µg/mL concentration (relative total growth of 4%); therefore, this concentration was excluded from the evaluation. An evaluation was made using data of following concentration of 10 µg/mL (relative total growth of 16%) and the next five concentrations (a total of six concentrations). Statistically significant increases in the mutation frequency were observed at 10, 7.5, 5, 2.5, 1.25 µg/mLconcentrations. However, the difference between the mutation frequency of the test item treated sample and the corresponding vehicle control value did not exceed the global evaluation factor in case of the 7.5, 5 and 2.5 µg/mLconcentrations, thus they were considered as biologically non relevant increases. In case of the 10 and 1.25 µg/mLconcentrations, the values were over the limit of the biological relevance (the difference was higher than the global evaluation factor) but the results did not follow a clear dose response and the increases were not reproduced in Assay 2.

In Assay 1, following a 3-hour treatment without metabolic activation, excessive cytotoxicity of the test item was observed at 80, 70, 60, 50, 40 and 30 µg/mL concentrations, cells of these samples did not survive the expression period. Therefore, an evaluation was made using data of the first surviving concentration of 20 µg/mL (relative total growth of 18%) and the next five examined concentrations (a total of six concentrations).

A statistically significant increase in the mutation frequency was observed at 20 µg/mL concentration. However, the difference between the mutation frequency of the test item treated sample and the corresponding vehicle control value did not exceed the global evaluation factor, thus it was considered as biologically non relevant increase.

In Assay 2, following a3-hour treatment with metabolic activation, similarly to the first test, excessive cytotoxicity was observed at 20 µg/mL concentration, no cells of this sample survived the expression period. Marked cytotoxicity was observed at 17.5, 15, 12.5, 10 and 7.5 µg/mL concentrations (relative total growth values of 2, 3, 6, 5, and 5%, respectively); therefore, these concentrations were excluded from the evaluation. An evaluation was made using data of 5 µg/mL concentration (relative total growth of 12%) and additional three treatment concentrations (a total of four concentrations). Statistically significantincrease in the mutation frequency was observed at 5 µg/mL concentration.However, similarly to the other experiments, the difference between the mutation frequency of the test item treated sample and the corresponding vehicle control value did not exceed the global evaluation factor, thus it was considered as biologically non relevant increase.

In Assay 2, following a 24-hour treatment without metabolic activation, excessive cytotoxicity was observed at 15 µg/mL concentration, no cells of this sample survived the expression period. Marked cytotoxicity was observed at 12.5, 10 and 9 µg/mL concentrations (relative total growth values were less than 1%); therefore, these concentrations were excluded from the evaluation. An evaluation was made using data of 8 µg/mL concentration (relative total growth of 27%) and additional seven treatment concentrations (a total of eight concentrations).A statistically significant increase in the mutation frequency was observed at 8 µg/mL concentration. However, similarly to the other experiments, the difference between the mutation frequency of the test item treated sample and the corresponding vehicle control value did not exceed the global evaluation factor, thus it was considered as biologically non relevant increase.

The experiments were performed using appropriate untreated, negative (vehicle) and positive control samples in all cases. The spontaneous mutation frequency of the negative (vehicle) controls was in the recommended range in each test. The positive controls gave the anticipated increases in mutation frequency over the controls. The plating efficiencies for the negative (vehicle) controls at the end of the expression period were within the acceptable range in all assays. The evaluated concentration ranges were considered to be adequate, as they covered the range from cytotoxicity to no or little cytotoxicity. The number of test concentrations met the acceptance criteria. Therefore, the overall study was considered to be valid.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

In conclusion, no clear mutagenic effect of Tellurium dioxide was observed either in the presence or in the absence of metabolic activation system under the conditions of this Mouse Lymphoma Assay. Therefore, it was considered being not positive with regard to classification.

Executive summary:

An in vitro mammalian cell assay was performed in mouse lymphoma L5178Y TK^+/-3.7.2 C cells at the tk locus to test the potential of Tellurium dioxide to cause gene mutation and/or chromosome damage. Treatment was performed for 3 hours with and without metabolic activation (±S9 mix) and for 24 hours without metabolic activation (-S9 mix).

In conclusion, no clear mutagenic effect of Tellurium dioxide was observed either in the presence or in the absence of metabolic activation system under the conditions of this Mouse Lymphoma Assay. Therefore, it was considered being not positive with regard to classification.