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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
08 Aug. - 06 Sep. 2007
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2007
Report date:
2007

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Qualifier:
according to guideline
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Principles of method if other than guideline:
Two replicate experiments: plate incorporation assay and preincubation test
GLP compliance:
yes (incl. QA statement)
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
Anthracene oil
EC Number:
292-602-7
Cas Number:
90640-80-5
Molecular formula:
Not applicabel
IUPAC Name:
Anthracene oil
Test material form:
other: depending on composition (UVCB) solid or viscous liquid
Details on test material:
anthracene oil; anthracene oil ≤ 50 ppm BaP, AOL (anthracene oil low)
- substance type: organic
Specific details on test material used for the study:
- Name of test material (as cited in study report): anthracene oil (< 50 ppm BaP)
- Lot/batch No. of test material: ATE No. 10443
- Stability under test conditions: no measured data; based on chemical structure assumed to be stable
- Storage condition of test material: room temperature, exclusion of light

Method

Species / strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
Metabolic activation:
with and without
Metabolic activation system:
Microsomal fraction prepared from induced livers of male Wistar rats, induced with phenobarbital (80 mg/kg bw) and ß-naphthoflavone (100 mg/kg bw) orally (3x)
Test concentrations with justification for top dose:
1st experiment: 3.16, 10, 31.6, 100, 316, and 1000 µg/plate (TA 98, TA 100, -S9)
1.0, 3.16, 10, 31.6, 100, 316, and 1000 µg/plate (TA 1535, TA 1537, TA 102, - S9)
3.16, 10, 31.6, 100, 316, 1000 and 2500 µg/plate (+ S9)
2nd experiment: with variations based on results of 1st experiment (Report, p. 11, p. 19)
3rd experiment: 200, 400, 500, 600, 750, and 1000 (only with TA 100, +S9)
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: compatible with survival of bacteria and S9 activity

Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
other: see Report p. 15
Details on test system and experimental conditions:
METHOD OF APPLICATION:
1st experiment: in agar (plate incorporation)
2nd and 3rd experiment: pre-incubation variant

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth/colony formation
Evaluation criteria:
Considered as mutagenic
- if a clear and dose-related increase in the number of revertants occurs in at least one tester strain with or without metabolic activationand/or
- if a biologically relevant positive response for at least one of the dose groups occurs in at least one tester strain with or without metabolic activation.

An increase is considered relevant
- if in TA 100 and TA 102 mutation rate is at least twice as high as the rate of the solvent control;
- if in TA 98, TA 1535, and TA 1537 the mutation rate is at least 3 fold higher than that of the solvent control.
Statistics:
According to the OECD guidelines, the biological relevance is the criterion for the interpretation of the results: a statistical evaluation was not considered necessary under this premise (report p. 21).

Results and discussion

Test resultsopen allclose all
Key result
Species / strain:
S. typhimurium TA 100
Metabolic activation:
with
Genotoxicity:
positive
Remarks:
in 2nd and 3rd test: reproducible at cytotoxic concentrations (MF = 2.2 and 2.3)
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
at ≥ 200 µg/pl. (+S9)
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 100
Metabolic activation:
without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
cytotoxic at higher concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium, other: TA 98, TA 1535, TA 1537, TA 102
Metabolic activation:
with and without
Genotoxicity:
negative
Remarks:
reproducible in both tests
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
cytotoxic at higher concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid

Any other information on results incl. tables

No mutagenic activity was observed in any test (plate incorporation and pre-incubation assay) at non-cytotoxic concentrations. In the pre-incubation assay, a mutagenic response was only observed in tester strain TA 100 with metabolic activation and at cytotoxic concentrations.

Experiment I (plate incorporation assay): No mutagenic activity was observed at all concentrations with and without metabolic activation.

Experiment II (pre-incubation assay): Relevant increase in the number of revertants in TA 100 but only at cytotoxic concentrations (500 µg/pl., cytotoxic, +S9) with MF = 2.0

Experiment III (pre-incubation assay): Relevant increases in revertants in TA 100 but only at cytotoxic concentrations (≥ 200 µg/plate, cytotoxic, +S9) with MF = 2.2 and 2.3.

Applicant's summary and conclusion

Conclusions:
From the five tester strains tested, only one (TA 100) showed a weak positive response with metabolic activation at already cytotoxic concentrations.