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Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Bacterial Mutagenicity: Tungsten disulphide was tested in the Salmonella typhimurium reverse mutation assay with four histidine-requiring strains of Salmonella typhimurium (TA1535, TA1537, TA100 and TA98) and in the Escherichia coli reverse mutation assay with a tryptophan-requiring strain of Escherichia coli WP2uvrA. The test was performed in two independent experiments in the presence and absence of S9-mix (Aroclor-1254 induced rat liver S9-mix). In the dose range finding test, tungsten disulphide was tested up to concentrations of 5000 µg/plate in the absence and presence of S9-mix in the strains TA100 and WP2uvrA. Tungsten disulphide precipitated on the plates at dose levels of 1000 µg/plate and upwards. Tungsten disulphide did not induce a dose-related, two-fold increase in the number of revertant (His+) colonies in each of the four tester strains (TA1535, TA1537, TA98 and TA100) and in the number of revertant (Trp+) colonies in tester strain WP2uvrA both in the absence and presence of S9-metabolic activation.  Based on the results of this study it is concluded that tungsten disulphide is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.

Mammalian Mutagenicity: No mammalian mutagenicity studies of sufficient quality are available for tungsten disulphide (target substance). However, mammalian mutagenicity data is available for tungsten metal (source substance), which will be used for reading across. Because of similar water solubility and toxicity for the target substance compared to the source substance, the resulting read across is appropriate to estimate of potential toxicity for this endpoint. For more details, refer to the attached description of the read-across approach. In anin vitroL5178Y TK +/- Mouse Lymphoma Forward Mutation Assay conducted according to OECD 476, tungsten metal was negative for mutagenicity both in the absence and presence of S9-metabolic activation.

Mammalian Chromosomal Aberration: No mammalian chromosomal studies of sufficient quality are available for tungsten disulphide (target substance). However, mammalian chromosomal aberration data is available for tungsten metal (source substance), which will be used for reading across. Because of similar water solubility and toxicity for the target substance compared to the source substance, the resulting read across is appropriate to estimate of potential toxicity for this endpoint. For more details, refer to the attached description of the read-across approach. In an in vitro chromosome aberration assay conducted according to OECD 473, tungsten metal was negative for mutagenicity both in the absence and presence of S9-metabolic activation.

Link to relevant study records

Referenceopen allclose all

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2002-01-09, 2002-01-24
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Type of assay:
bacterial reverse mutation assay
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: 16031/01
- Expiration date of the lot/batch: 01 April 2002
- Purity test date: >98%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: At room temperature in the dark
- Stability under test conditions: Stable
- Solubility and stability of the test substance in the solvent/vehicle: Dimethyl sulfoxide: Not indicated

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: suspended in dimethyl sulfoxide of spectroscopic quality (Merck). The stock solution was treated with ultrasonic waves to obtain a homogeneous suspension

FORM AS APPLIED IN THE TEST (if different from that of starting material)
stock solution

OTHER SPECIFICS:
Target gene:
histidine-requiring Salmonella typhimurium
of tryptophan-requiring Escherichia coli
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Additional strain / cell type characteristics:
not specified
Species / strain / cell type:
E. coli WP2 uvr A
Metabolic activation:
with and without
Metabolic activation system:
Rat liver microsomal enzymes were routinely prepared from adult male Wistar rats
Test concentrations with justification for top dose:
3, 10, 33, 100, 333, 1000, 3330 and 5000 µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: dimethyl sulfoxide
- Justification for choice of solvent/vehicle:
Untreated negative controls:
yes
Remarks:
dimethyl sulfoxide
Negative solvent / vehicle controls:
yes
Remarks:
0.1 ml dimethyl sulfoxide
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
4-nitroquinoline-N-oxide
9-aminoacridine
sodium azide
benzo(a)pyrene
methylmethanesulfonate
other: daunomycine, 2-aminoanthracene
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium; in agar (plate incorporation)

DURATION
- Preincubation period: until the cultures reached an optical density of 1.0 ± 0.1 at 700 nm (109 cells/ml}.
- Exposure duration: 48h
- Expression time (cells in growth medium): 20 min in medium (PIT) and incubation at 37C for 48-72 hours
- Selection time (if incubation with a selection agent):
- Fixation time (start of exposure up to fixation or harvest of cells):

SELECTION AGENT (mutation assays):
based on a dose range finding test with strain TA100 and the WP2uvrA strain, both with and without S9-mix


NUMBER OF CELLS EVALUATED: 109cells/ml

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth
Rationale for test conditions:
Recommended test system in international guidelines (e.g. EPA, OECD, EEC).
Evaluation criteria:
The test substance is considered positive in this assay if the following criteria are met:
• A dose-related and reproducible increase in the number of revertant colonies, i.e. about doubling of the spontaneous mutation rate in at least one tester strain either without S9 mix or after adding a metabolizing system.
A test substance is generally considered non-mutagenic in this test if:
• The number of revertants for all tester strains were within the historical negative control range under all experimental conditions in at least two experiments carried out independently of each other.
Statistics:
not specified
Key result
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity, but tested up to precipitating concentrations
Remarks:
?
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Key result
Species / strain:
E. coli WP2 uvr A
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity, but tested up to precipitating concentrations
Remarks:
?
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
?
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field "Test System"

According to the results of the present study, the test substance did not lead to a relevant increase in the number of revertant colonies either without S9 mix or after adding a metabolizing system in two experiments carried out independently of each other (standard plate test and preincubation assay).

Besides, the results of the negative as well as the positive controls performed in parallel corroborated the validity of this study, since the values fulfilled the acceptance criteria of this study.

In this study with and without S9 mix, the number of revertant colonies in the negative controls was within the range of the historical negative control data for each tester strain.

In addition, the positive control substances both with and without S9 mix induced a significant increase in the number of revertant colonies within the range of the historical positive control data or above.

Conclusions:
Based on the results of this study it is concluded that tungsten disulphide is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay
Executive summary:

Tungsten disulphide was tested in theSalmonella typhimuriumreverse mutation assay with four histidine-requiring strains of Salmonella typhimurium (TA1535, TA1537, TA100 and TA98) and in theEscherichia colireverse mutation assay with a tryptophan-requiring strain of Escherichia coliWP2uvrA. The test was performed in two independent experiments in the presence and absence of S9-mix (Aroclor-1254 induced rat liver S9-mix). In the dose range finding test, tungsten disulphide was tested up to concentrations of 5000 µg/plate in the absence and presence of S9-mix in the strains TA100 and WP2uvrA. Tungsten disulphide precipitated on the plates at dose levels of 1000 µg/plate and upwards. The bacterial background lawn was not reduced at any of the concentrations tested and no biologically relevant decrease in the number of revertants was observed.

In the first and second mutation assay, tungsten disulphide was tested up to concentrations of 1000 µg/plate in the absence and presence of S9-mix. tungstn disulphide precipitated on the plates at this dose level. The bacterial background lawn was not reduced at any of the concentrations tested and no decrease in the number of revertants was observed. The presence of 5 and 10% (v/v) liver microsomal activation did not influence these findings.

Tungstn disulphide did not induce a dose-related, two-fold increase in the number of revertant (His+) colonies in each of the four tester strains (TA1535, TA1537, TA98 and TA100) and in the number of revertant (Trp+) colonies in tester strain WP2uvrA both in the absence and presence of S9-metabolic activation.  Based on the results of this study it is concluded that tungsten disulphide  is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay

Endpoint:
in vitro cytogenicity / chromosome aberration study in mammalian cells
Remarks:
Type of genotoxicity: chromosome aberration
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2004-04-08 to 2004-04-08
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
Only an unsigned draft report available for review., however the study was well documented study conducted according to OECD guideline 473 and GLP.
Justification for type of information:
REPORTING FORMAT FOR THE CATEGORY APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH: The hypothesis is that properties are likely to be similar or follow a similar pattern because of the presence of a common metal ion, in this case tungstate.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES):
Source: Tungsten Metal
Target: Tungsten Disulphide


3. CATEGORY APPROACH JUSTIFICATION: See Annex 1 in CSR

4. DATA MATRIX: See Annex 1 in CSR
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
Species / strain / cell type:
Chinese hamster Ovary (CHO)
Details on mammalian cell type (if applicable):
- Type and identity of media: The CHO cells were grown in McCoy's 5a culture medium supplemented with ~10 % heat-inactivated fetal bovine serum (FBS), L-glutamine (2mM), penicillin G (100 units/mL), and streptomycin (100 ug/mL).
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: yes
- Periodically checked for karyotype stability: yes
Additional strain / cell type characteristics:
other: The CHO-WBL subclone is a permanent cell line with an average cycle time of 12 to 14 hours and a modal chromosome number of 21.
Metabolic activation:
with and without
Metabolic activation system:
Aroclor 1254 induced rat liver S9
Test concentrations with justification for top dose:
Initial assay with and without metabolic activation (3-hour treatment)- 13.6, 19.4, 27.7, 39.5, 56.5, 80.7, 115, 165, 235, 336, 480, 686, 980, 1400 and 2000 ug/mL
Confirmatory assay without metabolic activation (20-hour treatment)- 3.13, 6.25, 12.5, 25.0, 50.0, 100, 200, 300, 400 and 500 ug/mL
Confirmatory assay with metabolic activation (3-hour treatment)- 50.0, 100, 200, 300, 400 and 500 ug/mL
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: 0.9 % NaCl USP Saline
- Justification for the choice of solvent/vehicle: In saline, the test substance formed a homogeneous, opaque, dark grey suspension with the test substance settling down slowly at a concentration of ~20.0 mg/mL. At a dose concentration of 2000 ug/mL (dosed in the absence of cells using a 10 % dosing volume in McCoy's 5a culture medium), the test substance formed a homogenous, slightly translucent, light red suspension with a precipitate settling down slowly in the dilution tube, and pH was 8.5 (pH of the culture medium was 8.5).
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
mitomycin C
Remarks:
Non-activation assay Migrated to IUCLID6: 0.750 and 1.50 ug/mL, for the 3-hour treatment; 0.200 and 0.400, for the 20-hour treatment
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
cyclophosphamide
Remarks:
Activation assay Migrated to IUCLID6: 7.50 and 12.5 ug/mL
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium

DURATION
- Exposure duration: Initial assay with and without S9- 3 hours; Confirmatory assay- 20 hours without S9, 3 hours with S9
- Expression time (cells in growth medium): Initial assay with and without S9- approximately 15 hours; Confirmatory assay- approximately 15 hours for cultures treated in the presence of S9 and no expression period for cultures treated in the absence of S9
- Fixation time (start of exposure up to fixation or harvest of cells): approximately 20 hours, with Colcemid present during the last 2 +/-0.5 hours


SPINDLE INHIBITOR (cytogenetic assays): Colcemid
STAIN (for cytogenetic assays): 5 % Giemsa


NUMBER OF REPLICATIONS: 2


NUMBER OF CELLS EVALUATED: 100 cells, if possible, from each replicate culture were analyzed for the different types of chromosomal aberrations. At least 25 cells were analyzed from those cultures that had greater than 25 % of cells with one or more aberrations.


DETERMINATION OF CYTOTOXICITY
- Method: mitotic index, visual assessment of the percent confluence of the cell monolayer as compared to the vehicle control.


OTHER EXAMINATIONS:
- Determination of polyploidy: Yes, by evaluating at least 100 metaphases, if available.
- Determination of endoreplication: Yes, by evaluating at least 100 metaphases, if available.
Evaluation criteria:
The following factors were taken into account in evaluation of the data:
- The number and percentages of aberrant cells excluding and including gaps
- Evidence of a dose-response relationship.
Evaluation of a Positive Response- The test substance was considered positive for inducing chromosomal aberrations if a significant increase (the difference was considered significant when p<= 0.01) in the number of cells with chromosomal aberrations was observed at one or more concentrations. The linear trend test evaluated the dose responsiveness. A dose-response should be observed if a significant increase was seen at one or more concentrations.
Evaluation of a Negative Response-The test substance was considered negative for inducing chromosomal aberration if no significant increase was observed in the number of cells with chromosomal aberrations at any of the concentrations.
Equivocal Evaluation- Most assays give clearly positive or negative results, but in rare cases the data would preclude making a definitive judgment about the activity of the test substance. Results might remain equivocal or questionable regardless of the number of times the assay was repeated.
-In certain circumstances the Study Director might use additional considerations to obtain a final evaluation of the test substance based upon the Study Director's scientific judgment.
Statistics:
Statistical analysis employed a Cochran-Armitage test for linear trend and Fisher's Exact Test to compare the percentage of cells with aberrations in treated cells to the results obtained for the vehicle controls.
Statistical analysis was also performed for cells exhibiting polyploidy and/or endoreduplication in order to indicate significant (p<= 0.01) increases in these events as indicators of possible induction of numerical aberrations; however, the groups were evaluated only for structural aberrations and not for numerical aberrations by this protocol.
Species / strain:
Chinese hamster Ovary (CHO)
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity, but tested up to precipitating concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
Precipitation and pH: In the initial assay without metabolic activation with a 3-hour treatment, a precipitate was observed after dosing and prior to washing of the cultures treated with >/=980 ug/mL. A precipitate was observed prior to harvest of the cultures treated with >/= 336 ug/mL. In the initial assay with metabolic activation with a 3 hour treatment, a precipitate was observed after dosing and prior to wash of the cultures treated with >/= 480 ug/mL. A precipitate was observed prior to harvest of the cultures treated with >/= 336 ug/mL. In the confirmatory assay without metabolic activation with a 20-hour treatment, a precipitate was observed after dosing and prior to harvest of the cultures treated with >/=300 ug/mL. In the confirmatory assay with metabolic activation with a 3-hour treatment, a precipitate was observed after dosing and prior to harvest of the cultures treated with >/=300 ug/mL.


COMPARISON WITH HISTORICAL CONTROL DATA: Vehicle control and positive control values for % of cells with chromosome aberrations, % of cells with chromosome aberrations + % of cells with gaps, % of polyploidy cells and % endoreduplication cells were all within historical data ranges, except for the % of cells with chromosome aberrations + % of cells with gaps in the vehicle control group of the initial assay in the absence of metabolic activation. The average value for this group was 4.5 % which was slightly above the historical data range of 0-4 %.


ADDITIONAL INFORMATION ON CYTOTOXICITY:
1. Initial assay (3-hour treatment): In the assay without metabolic activation, the monolayer confluence as compared to vehicle control was 86 % at 2000 ug/mL; at all other concentrations it was 100 %. Reductions of 0, 0, 0, 21, 0 and 4 % were observed in the mitotic indices of the cultures treated with 115, 165, 235, 336, 686 and 2000 ug/mL, respectively, as compared with the vehicle control cultures. In the assay with metabolic activation, the monolayer confluence as compared to the vehicle control was 86 % at 1400 and 2000 ug/mL; at all other concentrations it was 100 %. Reductions of 0, 2, 0, 0, 0, 0, and 0 % were observed in the mitotic indices of the cultures treated with 115, 165, 235, 336, 686, 1400 and 2000 ug/mL, respectively, as compared with the vehicle control cultures. Since there was no toxicity exhibited at any of the doses tested, the lowest dose with a precipitate at the time of harvest was selected as the highest dose selected for chromosome aberration analysis as recommended by OECD Testing Guidelines. Chromosome aberrations were analyzed from the cultures treated with 115, 165, 235 and 336 ug/mL.
2. Confirmatory assay: In the assay without metabolic activation, reductions of 3, 3, 3 and 17 % were observed in the mitotic indices of the cultures treated with 50, 200, 300 and 500 ug/mL, respectively, as compared with the vehicle control cultures. In the assay with metabolic activation, reductions of 0, 0, and 1 % were observed in the mitotic indices of the cultures treated with 200, 300 and 500 ug/mL, respectively, as compared with the vehicle control cultures. Since there was no toxicity exhibited at any of the doses tested in the absence or presence of metabolic activation, the lowest dose with a precipitate at the time of harvest was selected as the highest dose selected for analysis as recommended by the OECD Testing Guidelines. In the absence of S9, chromosomal aberrations were analyzed from the cultures treated with 25, 50, 200 and 300 ug/mL, and in the presence of S9, chromosomal aberrations were analyzed from the cultures treated with 50, 100, 200 and 300 ug/mL
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

The sensitivity of the cell cultures for induction of chromosomal aberrations was shown by the increase frequency of aberrations in the cells treated with both positive controls.

Conclusions:
The test substance was considered negative for inducing structural chromosomal aberrations in CHO cells with and without metabolic activation under conditions of this assay.
Executive summary:

No in vitro genotoxicity data of sufficient quality are available for tungsten disulphide (target substance). However, in vitro genotoxicity data are available for tungsten metal (source substance), which will be used for reading across. Due to similar water solubility and toxicity for the target substance compared to the source substance, the resulting read across from the source substance to the target substance is appropriate. In addition, read across is appropriate because the classification and labelling is similar for the source substance than the target substance, the PBT/vPvB profile is the same, and the dose descriptors are, or are expected to be, lower for the source substance. For more details, refer to the read-across category approach included in the Category section of this IUCLID submission and/or as an Annex in the CSR.

Endpoint:
in vitro gene mutation study in mammalian cells
Remarks:
Type of genotoxicity: gene mutation
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
key study
Study period:
2004-03-18 to 2004-07-26
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
Well documented study performed according to OECD guideline 476 and GLP.
Justification for type of information:
REPORTING FORMAT FOR THE CATEGORY APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH: The hypothesis is that properties are likely to be similar or follow a similar pattern because of the presence of a common metal ion, in this case tungstate.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES):
Source: Tungsten Metal
Target: Tungsten Disulphide


3. CATEGORY APPROACH JUSTIFICATION: See Annex 1 in CSR

4. DATA MATRIX: See Annex 1 in CSR
Reason / purpose for cross-reference:
read-across: supporting information
Qualifier:
according to guideline
Guideline:
OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)
Deviations:
no
GLP compliance:
yes
Type of assay:
mammalian cell gene mutation assay
Target gene:
thymidine kinase locus
Species / strain / cell type:
mouse lymphoma L5178Y cells
Details on mammalian cell type (if applicable):
- Type and identity of media: The culture medium used in the study was RPMI 1640 supplemented with horse serum (10 % by volume), Pluronic F68, L-glutamine, sodium pyruvate, penicillin and streptomycin. Treatment medium was Fischer's medium with the same medium supplements used in the culture medium except that the horse serum concentration was reduced to 5 % by volume. Cloning medium consisted of the RPMI 1640 culture medium with up to 20 % horse serum, without Pluronic F68 and with the addition of 0.24 % Noble agar to achieve a semisolid state. Selection medium was cloning medium that contained Trifluorothymidine (TFT).
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: yes
- Periodically checked for karyotype stability: yes
Additional strain / cell type characteristics:
other: heterozygous at the TK locus
Metabolic activation:
with and without
Metabolic activation system:
Aroclor 1254 induced rat liver S9
Test concentrations with justification for top dose:
Initial non-activated and activated assays- 9.85, 19.7, 39.3, 78.5, 157, 313, 625, 1250, 2500 and 5000 ug/mL
Confirmatory non-activated and activated assays- 19.7, 39.3, 78.5, 157, 313, 625, 1250, 2500 and 5000 ug/mL
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: 10 % saline
- Justification for choice of solvent/vehicle: The test substance formed an opaque, black, homogeneous suspension in the vehicle at 50 mg/mL, the highest concentration prepared. Upon further dilution, the test substance became a transparent, light-grey solution at concentrations as high as 3.13 mg/mL and remained soluble at all lower concentratoins.
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
methylmethanesulfonate
Remarks:
Non-activation test system Migrated to IUCLID6: 13 ug/mL
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: Methylcholanthrene- 2 and 4 ug/mL
Remarks:
Activated test system
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium

DURATION
- Exposure duration: 4 hours
- Expression time (cells in growth medium): 2days
- Selection time (if incubation with a selection agent): 13 days


SELECTION AGENT (mutation assays): 5-trifluorothymidine


NUMBER OF REPLICATIONS: 3


NUMBER OF CELLS EVALUATED: 3E-6 cells


DETERMINATION OF CYTOTOXICITY
- Method: relative total growth


Evaluation criteria:
see other information on materials and methods
Statistics:
no data
Key result
Species / strain:
mouse lymphoma L5178Y cells
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: The test substance was observed to precipitate from treatment medium by the termination of treatment at concentrations as low as 313 ug/ml.
- Other confounding effects: The test substance formed an opaque, black, homogeneous suspension in the vehicle at 50 mg/ml, the highest concentration prepared. Upon further dilution, the test substance became a transparent, light-grey solution at concentrations as high as 3.13 mg/ml and remained soluble at all lower concentrations.


RANGE-FINDING/SCREENING STUDIES: Cells were treated with the test substance for approximately 4 hours in the presence and absence of metabolic activation at concentrations ranging from 9.85-5000 ug/mL. The test substance induced no cytotoxicity to weak cytotoxicity. Based on these results, 5000 ug/ml was chosen as the top concentration for the definitive assay.


COMPARISON WITH HISTORICAL CONTROL DATA: All vehicle and positive control values were within historical data ranges for mutant frequencies except Methylcholanthrene (MCA) (activation assay) at 4 ug/ml. In the initial activation assay MCA had a mutant frequency 608.1E-6 and in the confirmatory assay MCA had a value of 607.5E-6. The historical range for mutant frequencies is 200.2 to 524.0E-6.
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Initial non-activation and activation assays- Concentrations of 9.85 and 19.7 ug/ml were discarded because a sufficient number of higher concentrations were available.

Confirmatory non-activation and activation assays- The concentration of 19.7 ug/ml was terminated because there were sufficient higher concentrations available for analysis.

Control Values- The average cloning efficiencies for the vehicle controls were 99.3 and 130.6 % without metabolic activation and 104.9 and 116.2 % with metabolic activation, which demonstrated acceptable cloning conditions for the assays. The positive control cultures induced large increases in mutant frequencies that were greatly in excess of the minimum criteria.

Sizing analysis- The L5178Y TK +/- mutation assay produces a bimodal distribution of large and small mutant colonies. The origin of the bimodal of mutant colony sizes is considered to reflect the types of genetic damage, with the large colonies derived from cells with intragenic mutations that affect only the TK gene and the small colonies the result of larger mutations that affect cell growth as well as the TK gene. Mutant colonies from all the cultures showed the expected bimodal distribution, and mutant colonies from the positive control treated cultures showed both small and large colonies.

Conclusions:
The test substance was did not induce forward mutations at the TK locus in L5178Y mouse lymphoma cells, under the activation and non-activation conditions of this study.
Executive summary:

No in vitro genotoxicity data of sufficient quality are available for tungsten disulphide (target substance). However, in vitro genotoxicity data are available for tungsten metal (source substance), which will be used for reading across. Due to similar water solubility and toxicity for the target substance compared to the source substance, the resulting read across from the source substance to the target substance is appropriate. In addition, read across is appropriate because the classification and labelling is similar for the source substance than the target substance, the PBT/vPvB profile is the same, and the dose descriptors are, or are expected to be, lower for the source substance. For more details, refer to the read-across category approach included in the Category section of this IUCLID submission and/or as an Annex in the CSR.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Justification for classification or non-classification

An in vitro reverse gene mutation assays of sufficient quality on tungsten disulphide (target substance) was negative for mutagenicity. In addition, an in vitro L5178Y TK +/- Mouse Lymphoma Forward Mutation Assay and an in and an in vitro chromosome aberration assay on tungsten metal (source substance), used for read-across to tungsten disulphide, were negative for mutagenicity. Therefore, based on the weight-of-evidence from the available data, tungsten disulphide does not warrant classification for genetic toxicity.