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Administrative data

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Ames assay:

The test chemical did not induce mutation in the Salmonella typhimurium and E. coli strains both in the presence and absence of S9 metabolic activation system and hence is not likely to be mutagenic under the conditions of this study.

Link to relevant study records
Reference
Endpoint:
in vitro gene mutation study in bacteria
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
data from handbook or collection of data
Remarks:
Experimental data from various test chemicals
Justification for type of information:
Data for the target chemical is summarized based on the various test chemicals.
Reason / purpose:
read-across source
Reason / purpose:
read-across source
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Version / remarks:
as per prival modification
Principles of method if other than guideline:
WoE for the target CAS is summarized based on data from various test chemicals.
GLP compliance:
not specified
Type of assay:
bacterial reverse mutation assay
Target gene:
2. Histidine
3. Histidine for S. typhimurium and Tryptophan for E. coli
Species / strain / cell type:
S. typhimurium, other: TA98 and TA100
Remarks:
2
Details on mammalian cell type (if applicable):
not specified
Additional strain / cell type characteristics:
not specified
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Remarks:
3
Details on mammalian cell type (if applicable):
not specified
Additional strain / cell type characteristics:
not specified
Species / strain / cell type:
E. coli WP2 uvr A
Remarks:
3
Details on mammalian cell type (if applicable):
not specified
Additional strain / cell type characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
2. Type and composition of metabolic activation system: cofactors (FMN, NADH, glucose-6-phosphate dehydrogenase, and glucose-6-phosphate), and test chemical were added, mixed, and incubated at 30 °C for 30 min without shaking.
- source of S9: uninduced hamster liver S9
- method of preparation of S9 mix: No data
- concentration or volume of S9 mix and S9 in the final culture medium: 30% v/v
- quality controls of S9 (e.g., enzymatic activity, sterility, metabolic capability) - Nitrogen was blown over the preincubation tube to keep the atmosphere reduced.
3. S9 mix
Test concentrations with justification for top dose:
2. 0, 333, 1000, 3333, 6666, 10000 ug/plate
3. 0, 156*, 31*, 625*, 1250*, 2500*, 5000* microg/plate (*: precipitation of sediment)
Vehicle / solvent:
2. - Vehicle(s)/solvent(s) used: Water
- Justification for choice of solvent/vehicle: Test chemical was soluble in water.
3. - Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: Test chemical was soluble in DMSO.
Untreated negative controls:
not specified
Negative solvent / vehicle controls:
yes
Remarks:
water
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
congo red
Remarks:
2
Untreated negative controls:
not specified
Negative solvent / vehicle controls:
yes
Remarks:
DMSO
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
sodium azide
benzo(a)pyrene
other: S9mix(-); AF-2 (TA98, TA100, WP2uvrA), ICR-191 (TA1537); S9mix(+): 2-AA (TA1535, WP2uvrA)
Remarks:
3
Details on test system and experimental conditions:
2. METHOD OF TREATMENT/ EXPOSURE:
- Cell density at seeding (if applicable): (1-2) x10^9 cells/mL
- Test substance added - preincubation

TREATMENT AND HARVEST SCHEDULE:
- Preincubation period, if applicable:30 min
- Exposure duration/duration of treatment: 48 h
- Harvest time after the end of treatment (sampling/recovery times): No data

METHODS FOR MEASUREMENT OF CYTOTOXICITY
- Method.: Preincubation Method - All plates were counted with an Artek automated colony counter (Artek 880, DynaTech, Chantilly, VA) or Minicount colony counter (Imaging Products International, Inc., Chantilly, VA), which was calibrated prior to use.
3. NUMBER OF REPLICATIONS:
- Number of cultures per concentration - duplicate
- Number of independent experiments - 3

METHOD OF TREATMENT/ EXPOSURE:
- Cell density at seeding (if applicable): No data
- Test substance added - in agar (plate incorporation)
Evaluation criteria:
2. All plates were observed for increase in mutant frequencies.
3. No. of revertants per plates were evaluated.
Statistics:
No data
Species / strain:
S. typhimurium, other: TA98 and TA100
Remarks:
2
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Vehicle controls validity:
valid
Untreated negative controls validity:
not specified
True negative controls validity:
not specified
Positive controls validity:
valid
Species / strain:
S. typhimurium, other: TA 1535, TA 1537, TA 98 and TA 100
Remarks:
3
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Vehicle controls validity:
valid
Untreated negative controls validity:
not specified
True negative controls validity:
not specified
Positive controls validity:
valid
Species / strain:
E. coli WP2 uvr A
Remarks:
3
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Vehicle controls validity:
valid
Untreated negative controls validity:
not specified
True negative controls validity:
not specified
Positive controls validity:
valid
Additional information on results:
2. RANGE-FINDING/SCREENING STUDIES (if applicable): The doses that were tested in the mutagenicity assay were selected based on the levels of cytotoxicity observed in a preliminary dose range-finding study using strain TA100. Ten dose levels of the chemical, one plate per dose, were tested in both the presence and the absence of induced hamster S9. If no toxicity was observed, a total maximum dose of 10 mg of test chemical per plate was used.
3. No data
Remarks on result:
other: No mutagenic potential
Conclusions:
The test chemical did not induce mutation in the Salmonella typhimurium and E. coli strains both in the presence and absence of S9 metabolic activation system and hence is not likely to be mutagenic under the conditions of this study.
Executive summary:

In different studies, the given test chemical has been investigated for the mutagenic nature. The studies are as mentioned below:

 

Bacterial Reverse Mutation Assay was conducted by using the given test chemical as per OECD Guideline 471 (Bacterial Reverse Mutation Assay) and Prival modification by Prival and Mitchell (1982). For tests using the FMN-modified assay, strains TA98 and TA100 were used at doses 0, 333, 1000, 3333, 6666, 10000 ug/plate dissolved in water. The bacteria, uninduced hamster liver S9 (30% v/v), cofactors (FMN, NADH, glucose-6-phosphate dehydrogenase, and glucose-6-phosphate), and test chemical was added, mixed, and incubated at 30 °C for 30 min without shaking. Nitrogen was blown over the preincubation tube to keep the atmosphere reduced. At the end of the incubation period, 2 mL of molten top agar was added to each sample tube and the mixture was poured on a minimal agar plate containing 0.5% glucose rather than the 2% glucose specified by Ames et al. The plates were then incubated at 37 °C for 48 h. The positive control in FMN experiment was Congo red. All plates were counted with an Artek automated colony counter or Minicount Colony counter, which was calibrated prior to use. The given test chemical failed to induce mutation in Salmonella typhimurium TA98 and TA100 in the presence and absence of S9 metabolic activation and hence it is not likely to be mutagenic by in-vitro test.

 

Another gene mutation toxicity study was performed to determine the mutagenic nature of the test chemical as per OECD Guideline 471 (Bacterial Reverse Mutation Assay) on S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2 uvr A at doses 0, 156*, 31*, 625*, 1250*, 2500*, 5000* microg/plate (*: precipitation of sediment). DMSO was used as a solvent. Positive controls used were S9mix(-); AF-2 (TA98, TA100, WP2uvrA), SA (TA1535), ICR-191 (TA1537) / S9mix(+); B[a]P (TA98, TA100, TA1537), 2-AA (TA1535, WP2uvrA). The study was performed in duplicate with 3 plates per test by plate incorporation method. No. of revertants per plates were evaluated. The given test chemical failed to induce mutation in S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2 uvr A in the presence and absence of S9 metabolic activation system and hence it is not likely to be mutagenic by in-vitro test.

 

Thus, based on the above summarized studies on test chemical, it can be concluded that the given test chemical did not induce mutation in the Salmonella typhimurium and E. coli strains both in the presence and absence of S9 metabolic activation system and hence is not likely to be mutagenic under the conditions of this study.

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

Genetic toxicity in vivo

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Ames assay:

In different studies, the given test chemical has been investigated for the mutagenic nature. The studies are as mentioned below:

 

Bacterial Reverse Mutation Assay was conducted by using the given test chemical as per OECD Guideline 471 (Bacterial Reverse Mutation Assay) and Prival modification by Prival and Mitchell (1982). For tests using the FMN-modified assay, strains TA98 and TA100 were used at doses 0, 333, 1000, 3333, 6666, 10000 ug/plate dissolved in water. The bacteria, uninduced hamster liver S9 (30% v/v), cofactors (FMN, NADH, glucose-6-phosphate dehydrogenase, and glucose-6-phosphate), and test chemical was added, mixed, and incubated at 30 °C for 30 min without shaking. Nitrogen was blown over the preincubation tube to keep the atmosphere reduced. At the end of the incubation period, 2 mL of molten top agar was added to each sample tube and the mixture was poured on a minimal agar plate containing 0.5% glucose rather than the 2% glucose specified by Ames et al. The plates were then incubated at 37 °C for 48 h. The positive control in FMN experiment was Congo red. All plates were counted with an Artek automated colony counter or Minicount Colony counter, which was calibrated prior to use. The given test chemical failed to induce mutation in Salmonella typhimurium TA98 and TA100 in the presence and absence of S9 metabolic activation and hence it is not likely to be mutagenic by in-vitro test.

 

Another gene mutation toxicity study was performed to determine the mutagenic nature of the test chemical as per OECD Guideline 471 (Bacterial Reverse Mutation Assay) on S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2 uvr A at doses 0, 156*, 31*, 625*, 1250*, 2500*, 5000* microg/plate (*: precipitation of sediment). DMSO was used as a solvent. Positive controls used were S9mix(-); AF-2 (TA98, TA100, WP2uvrA), SA (TA1535), ICR-191 (TA1537) / S9mix(+); B[a]P (TA98, TA100, TA1537), 2-AA (TA1535, WP2uvrA). The study was performed in duplicate with 3 plates per test by plate incorporation method. No. of revertants per plates were evaluated. The given test chemical failed to induce mutation in S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2 uvr A in the presence and absence of S9 metabolic activation system and hence it is not likely to be mutagenic by in-vitro test.

 

Thus, based on the above summarized studies on test chemical, it can be concluded that the given test chemical did not induce mutation in the Salmonella typhimurium and E. coli strains both in the presence and absence of S9 metabolic activation system and hence is not likely to be mutagenic under the conditions of this study.

Justification for classification or non-classification

Based on the data available and applying weight of evidence approach, the given test chemical does not exhibit gene mutation in vitro by Ames assay. Hence, the test chemical is not likely to classify as a gene mutant as per the criteria mentioned in CLP regulation.