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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
September 13 - November 11, 2011
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2011
Report Date:
2011

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Qualifier:
according to
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
GLP compliance:
yes (incl. certificate)
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Specific details on test material used for the study:
Name of test material: Art. 139222
Synonym: HTM-081
Chemical name: biphenyl-2-yl-(9,9-dimethyl-9H-fluoren-2-yl)-(9,9’-spirobifluoren-2-yl)-amine
Batch No.: OS11008922
Purity (HPLC): > 99,95 % (Area %)
Appearance: pale yellow powder without visible impurities
Supplier: Merck KGaA, Darmstadt
Analytical Report: Merck KGaA, Darmstadt, WL-AC6, Dr. Ulrich Engel
Released until: August 15, 2016
Solvent: dimethyl sulfoxide (DMSO) purity ≥ 99.9 % grade: for analysis

Method

Target gene:
Salmonella: histidine operon
E coli. tryptophane operon
Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
Details on mammalian cell type (if applicable):
- Type and identity of media:
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: yes
- Periodically checked for karyotype stability: yes
- Periodically "cleansed" against high spontaneous background: yes
Species / strain / cell type:
S. typhimurium TA 102
Details on mammalian cell type (if applicable):
- Type and identity of media:
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: yes
Metabolic activation:
with and without
Metabolic activation system:
S9 mix from Aroclor 1254 pretreated rats
Test concentrations with justification for top dose:
1st series: 5.00, 15.8, 28.1, 50.0, 158, 500, 1580 µg/plate
2nd series: 15.8, 28.1, 50.0, 88.9, 158 µg/plate
Vehicle / solvent:
DMSO
Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
other: Sodium azide, 2-Aminoanthracene, Cumene hydroperoxide, 9-Aminoacridine, Daunomycin, 4-Nitroquinoline-N-oxide
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Preincubation period: 3-5 hours
- Exposure duration: about 2 days


SELECTION AGENT (mutation assays): Histidine, Tryptophane

NUMBER OF REPLICATIONS:
Negative controls 6
Test material 3
Positive controls 3


NUMBER OF CELLS EVALUATED: about 1e9


DETERMINATION OF CYTOTOXICITY
- Method: other: background cytotox
Rationale for test conditions:
According to guideline
Evaluation criteria:
- valid assay (cf. laboratory historical data)
- no or weak increase in revertant colonies = negative
- clear, dose dependent (over at least two concentrations), and reproducible increase in revertant colonies= positve
Statistics:
not applied

Results and discussion

Test results
Key result
Species / strain:
other: all strains/cell types tested
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no 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
- Effects of osmolality: no
- Evaporation from medium: no
- Water solubility: ok
- Precipitation:yes ( ≥ 158 µg/plate)
- Other confounding effects: no

Applicant's summary and conclusion

Conclusions:
With and without addition of S9 mix as the external metabolizing system, the test material was not mutagenic under the experimental conditions described.
Executive summary:

With and without addition of S9 mix as the external metabolizing system, the test material was not mutagenic under the experimental conditions described.