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Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

TAOBN did not cause a positive mutagenic response in either the presence or absence of Arclor-inducted rat liver S9. While there was no independent repeat, the supporting study also concluded TAOBN was not mutagenic in the microbial assays either in the presence or absence of a liver microsomal system.

Link to relevant study records

Referenceopen allclose all

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Reference:
Composition 0
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
GLP compliance:
no
Type of assay:
bacterial reverse mutation assay
Test material information:
Composition 1
Specific details on test material used for the study:
Haskel No. 9864-02
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with and without
Metabolic activation system:
rat-liver homogenate activation system (S9)
Species / strain:
S. typhimurium TA 100
Metabolic activation:
with and without
Metabolic activation system:
rat-liver homogenate activation system (S9)
Species / strain:
S. typhimurium TA 1537
Metabolic activation:
with and without
Metabolic activation system:
rat-liver homogenate activation system (S9)
Species / strain:
S. typhimurium TA 1538
Metabolic activation:
with and without
Metabolic activation system:
rat-liver homogenate activation system (S9)
Species / strain:
S. typhimurium TA 98
Metabolic activation:
with and without
Metabolic activation system:
rat-liver homogenate activation system (S9)
Test concentrations with justification for top dose:
With activation: 10, 12, 24, 36, 48, 60 µg/plate
Without activation: 2, 4, 6, 8, 10 µg/plate
Vehicle:
Distilled water
Positive controls:
yes
Positive control substance:
other: 2-aminoanthracene
Remarks:
Positive control for assay in the presence of activation
Positive controls:
yes
Positive control substance:
9-aminoacridine
Remarks:
Positive control for assay in the absence of activation
Positive controls:
yes
Positive control substance:
2-nitrofluorene
Remarks:
Positive control for assay in the absence of activation
Negative controls:
yes
Solvent controls:
yes
Remarks:
Negative control for assay in the absence of activation
Negative controls:
yes
Remarks:
S9 negative control included in the activated assay to reassure any activity of the compound in the absence of the S9 activator mixture
Details on test system and conditions:
METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- All plates were incubated at 37 °C for 48 hours.

NUMBER OF REPLICATIONS: 2 plates
Statistics:
Average number of revertants from two plates
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity:
not specified
Vehicle controls valid:
not specified
Negative controls valid:
not specified
Positive controls valid:
not specified
Species / strain:
S. typhimurium TA 1537
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity:
not specified
Vehicle controls valid:
not specified
Negative controls valid:
not specified
Positive controls valid:
not specified
Species / strain:
S. typhimurium TA 1538
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity:
not specified
Vehicle controls valid:
not specified
Negative controls valid:
not specified
Positive controls valid:
not specified
Species / strain:
S. typhimurium TA 98
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity:
not specified
Vehicle controls valid:
not specified
Negative controls valid:
not specified
Positive controls valid:
not specified
Species / strain:
S. typhimurium TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity:
not specified
Vehicle controls valid:
not specified
Negative controls valid:
not specified
Positive controls valid:
not specified
Conclusions:
The test item was tested in Salmonella typhimurium strains TA1535, TA1537, TA1538, TA98 and TA100. The test item was not mutagenic in the microbial assays either in the presence or absence of a liver microsomal system.
Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study
Reference:
Composition 0
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Version / remarks:
1997
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay
Test material information:
Composition 1
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No. of test material: Haskell Number 26185
- Purity: 99.90%
Species / strain:
S. typhimurium TA 98
Metabolic activation:
with and without
Metabolic activation system:
Aroclor-induced rat liver S9
Species / strain:
S. typhimurium TA 100
Metabolic activation:
with and without
Metabolic activation system:
Aroclor-induced rat liver S9
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with and without
Metabolic activation system:
Aroclor-induced rat liver S9
Species / strain:
S. typhimurium TA 1537
Metabolic activation:
with and without
Metabolic activation system:
Aroclor-induced rat liver S9
Species / strain:
E. coli WP2 uvr A
Metabolic activation:
with and without
Metabolic activation system:
Aroclor-induced rat liver S9
Test concentrations with justification for top dose:
The dose levels tested were 3.3, 10, 33, 100, 333, and 1000 µg per plate with E. coli, TA100 in the presence of S9 activation and with TA1535 in the absence of S9 activation and 1.0, 3.3, 10, 33, 100 and 333 µg per plate with all remaining conditions. In the repeat test, the dose levels tested were 1.5, 5.0, 15, 50, 150, 500, 1500 and 5000 µg per plate.

A preliminary toxicity test was used to establish the dose-range over which the test substance would be tested.
Vehicle:
- Vehicle(s)/solvent(s) used: 100% ethanol
- Justification for choice of solvent/vehicle: selected based on the solubility of the test substance and compatibility with the target cells. Other solvents tested included dimethyl sulfoxide (DMSO) and acetone.
Positive controls:
yes
Positive control substance:
other: 2-aminoanthracene
Remarks:
Positive control for WP2 uvrA strain (for concentrations of 1.0 µg/plate and 1000 µg/plate)
Positive controls:
yes
Positive control substance:
2-nitrofluorene
Remarks:
Positive control for TA98 strain
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
Positive control for TA100 and TA1535 strains
Positive controls:
yes
Positive control substance:
9-aminoacridine
Remarks:
Positive control for TA1537 strain
Positive controls:
yes
Positive control substance:
methylmethanesulfonate
Remarks:
Positive control for WP2 uvrA (for concentration of 1000 µg/plate)
Details on test system and conditions:
METHOD OF APPLICATION:
- in agar (plate incorporation)

DURATION:
- Preincubation period: overnight
- Exposure duration: Not applicable. Cultures harvested via spectrophotometric monitoring of culture turbidity rather than by duration of incubation since overgrowth of cultures can cause loss of sensitivity of some mutagens
- Expression time: Cultures harvested in late log phase
- Fixation time: Each culture was monitoring spectrophotometrically for turbidity and was harvested at a percent transmittance yielding a titer of approximately 109 cells/mL.

NUMBER OF REPLICATIONS: 3 plates/test concentration
Evaluation criteria:
Revertant colonies, except for positive controls, were counted either entirely by automated colony counter or entirely by hand unless the plate exhibited toxicity.

For each replicate plating, the mean and standard deviation of the number of revertants per plate were calculated and are reported. For the test substance to be evaluated positive, it must cause a dose-related increase in the mean revertants per plate of at least one tester strain over a minimum of two increasing concentrations of test substance. Data sets for tester strains TA1535 and TA1537 were judged positive if the increase in mean revertants at the peak of the dose response is equal to or greater than 3.0-times the mean vehicle control value. Data sets for tester strains TA98, TA100, and WP2 uvrA were judged positive if the increase in mean revertants at the peak of the dose response is equal to or greater than 2.0-times the mean vehicle control value.
Key result
Species / strain:
S. typhimurium TA 98
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity:
not specified
Vehicle controls valid:
yes
Negative controls valid:
yes
Positive controls valid:
yes
Species / strain:
S. typhimurium TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity:
not specified
Vehicle controls valid:
yes
Negative controls valid:
yes
Positive controls valid:
yes
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity:
not specified
Vehicle controls valid:
yes
Negative controls valid:
yes
Positive controls valid:
yes
Species / strain:
S. typhimurium TA 1537
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity:
not specified
Vehicle controls valid:
yes
Negative controls valid:
yes
Positive controls valid:
yes
Species / strain:
E. coli WP2 uvr A
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity:
not specified
Vehicle controls valid:
yes
Negative controls valid:
yes
Positive controls valid:
yes
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: No precipitate was observed

RANGE-FINDING/SCREENING STUDIES:
- dose levels tested: 6.7, 10, 33, 67, 100, 333, 667, 1000, 3333, 5000 µg per plate
- no precipitate observed
- toxicity observed at 67, 100, 333, or 667 µg per plate
- based on findings, max doses plated in mutagenicity test were 1000 µg per plate with E. coli, TA100 in the presence of S9 activation and with TA1535 in the absence of S9 activation and 333 µg per plate with all remaining conditions.


HISTORICAL CONTROL DATA (with ranges, means and standard deviation and confidence interval (e.g. 95%)
- Positive historical control data:
Strain TA98:
Without activation:
Mean: 239 revertants/plate
SD: 158
Range: 30 - 1581 revertants/plate

With activation:
Mean: 679 revertants/plate
SD: 383
Range: 40 - 2294 revertants/plate

Strain TA100:
Without activation:
Mean: 572 revertants/plate
SD: 149
Range: 240 - 1945 revertants/plate

With activation:
Mean: 904 revertants/plate
SD: 437
Range: 163 – 2922 revertants/plate

Strain TA1535:
Without activation:
Mean: 319 revertants/plate
SD: 131
Range: 16 - 978 revertants/plate

With activation:
Mean: 137 revertants/plate
SD: 80
Range: 11 – 2246 revertants/plate

Strain TA1537:
Without activation:
Mean: 634 revertants/plate
SD: 375
Range: 13 - 2389 revertants/plate

With activation:
Mean: 127 revertants/plate
SD: 133
Range: 12 – 2060 revertants/plate

Strain WP2 uvrA:
Without activation:
Mean: 164 revertants/plate
SD: 147
Range: 14 - 1809 revertants/plate

With activation:
Mean: 390 revertants/plate
SD: 255
Range: 22 – 1493 revertants/plate



- Negative (solvent/vehicle) historical control data:
Strain TA98:
Without activation:
Mean: 16 revertants/plate
SD: 6
Range: 5 - 59 revertants/plate

With activation:
Mean: 21 revertants/plate
SD: 7
Range: 5 - 58 revertants/plate

Strain TA100:
Without activation:
Mean: 148 revertants/plate
SD: 36
Range: 68 - 262 revertants/plate

With activation:
Mean: 152 revertants/plate
SD: 38
Range: 74 - 271 revertants/plate

Strain TA1535:
Without activation:
Mean: 15 revertants/plate
SD: 6
Range: 3 - 46 revertants/plate

With activation:
Mean: 13 revertants/plate
SD: 5
Range: 2 – 50 revertants/plate

Strain TA1537:
Without activation:
Mean: 7 revertants/plate
SD: 3
Range: 1 - 22 revertants/plate

With activation:
Mean: 8 revertants/plate
SD: 3
Range: 1 – 28 revertants/plate

Strain WP2 uvrA:
Without activation:
Mean: 14 revertants/plate
SD: 4
Range: 5 - 52 revertants/plate

With activation:
Mean: 14 revertants/plate
SD: 4
Range: 4 – 52 revertants/plate

In the mutagenicity test, no positive mutagenic response was observed. The dose levels tested were 3.3, 10, 33, 100, 333, and 100 µg per plate with E. coli, TA100 in the presence of S9 activation and with TA1535 in the absence of S9 activation and 1.0, 3.3, 10, 33, 100 and 333 µg per plate with all remaining conditions. In the repeat test, the dose levels tested were 1.5, 5.0, 15, 50, 150, 500, 1500 and 5000 µg per plate. No precipitate was observed. Toxicity was observed beginning at 100, 150, 333 or at 1000 µg per plate.

Conclusions:
TAOBN did not cause a positive mutagenic response in either the presence or absence of Arclor-inducted rat liver S9. The study was concluded to be negative without conducting a confirmatory (independent repeat) test because no unique metabolism requirements were known about the test substance and because no equivocal responses were observed in the test that would suggest further testing is warranted.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Additional information

Justification for classification or non-classification

As two independent AMES studies were negative for gene mutation, the substance does not meet the criteria for CLP (EC 1272/2008 as amended) classification for germ cell mutagenicity.