Registration Dossier

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:
17-aug-2007 to 08-oct-2007
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study has been performed according to OECD and/or EC guidelines and according to GLP principles.

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)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Deviations:
no
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
tert-butyl N-[(1S,3S,4S)-4-hydroxy-4-[4-methoxy-3-(3-methoxypropoxy)phenyl]-1-[(2S,4S)-5-oxo-4-(propan-2-yl)oxolan-2-yl]-3-(propan-2-yl)butyl]carbamate
EC Number:
700-317-0
Cas Number:
934841-33-5
Molecular formula:
C30H49NO8
IUPAC Name:
tert-butyl N-[(1S,3S,4S)-4-hydroxy-4-[4-methoxy-3-(3-methoxypropoxy)phenyl]-1-[(2S,4S)-5-oxo-4-(propan-2-yl)oxolan-2-yl]-3-(propan-2-yl)butyl]carbamate
Details on test material:
- Name of test material (as cited in study report): SPP100 C6
- Substance type: White, fine-crystallic powder
- Physical state: Solid
- Stability under test conditions: Not indicated
- Storage condition of test material: Stored at ambient temperature
- Analytical purity: 99.2%
Significant impurities: Sum of all impurities by HPLC: 0.8%
Solvent content: Isopropyl acetate (CAS No. 108-21-4): 1.9% (w/w)
Methylcyclohexane (CAS 108-87-2): 5.8% (w/w)
- Lot/batch No.: HH-853.35Kr
- Expiration date of the lot/batch: December 2007

Method

Target gene:
- S. typhimurium: Histidine gene
- E. coli: Tryptophan gene
Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Species / strain / cell type:
E. coli WP2 uvr A
Metabolic activation:
with and without
Metabolic activation system:
Delor 106 (mixture of PCBs)
Test concentrations with justification for top dose:
Preliminary test (without S9) TA100 : 10, 100, 500, 1000, 2500 and 5000 µg/plate
Main study:
Experiment 1:
Without and with S9-mix: 50, 150, 500, 1500 and 5000 µg/plate
Experiment 2
Without and with S9-mix: 15, 50, 150, 500 and 1500 µg/plate.
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: Accepted and approved by authorities and international guidelines
Controlsopen allclose all
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
without S9-mix

Migrated to IUCLID6: 1.5 µg/plate for TA100 and TA1535
Positive control substance:
other: 4-nitro-o-phenylenediamine, 20 µg/plate for TA98
Remarks:
without S9-mix
Positive control substance:
9-aminoacridine
Remarks:
without S9-mix

Migrated to IUCLID6: 100 µg/plate for TA1537
Positive control substance:
other: N-methyl-N-nitro-N-nitrosoguanidine, 20 µg/plate for WP2uvrA
Remarks:
without S9-mix
Positive control substance:
other: 2-aminoanthracene for tester strains TA1535, TA1537 and WP2uvrA
Remarks:
with S9-mix
Positive control substance:
other: 2-aminofluorene for tester strains TA98 and TA100
Remarks:
with S9-mix
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48 - 72 hour

NUMBER OF REPLICATIONS:
- Doses of the test substance were tested in triplicate in each strain. Two independent experiments were conducted.

NUMBER OF CELLS EVALUATED: Not indicated

DETERMINATION OF CYTOTOXICITY
- Method: The reduction of the bacterial background lawn and the reduction of the revertant colonies was determined.

OTHER EXAMINATIONS:
- The presence of precipitation of the test compound on the plates was determined.


Evaluation criteria:
The main criterion for evaluation of results was modified two-fold increase rule. After this rule the result is positive, when reproducible dose-effects and/or doubling of ratio Rt/Rc is reached.

Results and discussion

Test resultsopen allclose all
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
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
Species / strain:
E. coli WP2 uvr A
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
- Precipitation: Slight precipitation was observed at the top dose of 5000 µg/plate

RANGE-FINDING/SCREENING STUDIES:
- No toxicity was observed up to and including the top dose of 5000 µg/plate

COMPARISON WITH HISTORICAL CONTROL DATA:
- The negative and strain-specific positive control values were within our laboratory historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly.

ADDITIONAL INFORMATION ON CYTOTOXICITY:
- No toxicity was observed up to and including the dose levels of 5000 and 1500 µg/plate in the first and second experiment, respectively

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative

The negative and strain-specific positive control values were within the laboratory historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly.

It is concluded that this test is valid and that SP100 C6 is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.