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:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Study in accordance to guideline requirements. Non GLP-Study but sufficiently documented.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2005

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Principles of method if other than guideline:
The test substance was investigated in a modified bacterial reverse mutation test as described by Ames et al.
(1975). The traditional Salmonella microsome mutation assay (Ames test) is used extensively as a routine test
for mutagenicity for more than 25 years. The high throughput microtitre-based version, called Ames II, is based
on the same genetic principle (base-pair substitution and frameshift mutations in the his operon of S. typhimurium)
as the traditional Ames test but combined with the fluctuation method (Gee et al., 1998). Because of its high
concordance (80%) compared with the standard assay, the Ames II procedure seems an effective screen for identifying
bacterial mutagens (Flueckiger et al., 2004). Due to its explorative character and the lack of regulatory acceptance these
data are, however, intended for supporting use only.
GLP compliance:
no
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Reference substance name:
BIBR 1048 Oxa-Ester
IUPAC Name:
BIBR 1048 Oxa-Ester

Method

Species / strain
Species / strain / cell type:
other: S. typhimurium TA Mix (TA 7001, TA 7002, TA 7003, TA 7004, TA 7005 and TA 7006) and TA 98
Metabolic activation:
with and without
Metabolic activation system:
Rat liver (Aroclor 1254)
Test concentrations with justification for top dose:
7 concentration levels from 1 to 5000 μg/mL
Vehicle / solvent:
DMSO
Controls
Untreated negative controls:
yes
Remarks:
DMSO
Negative solvent / vehicle controls:
yes
Remarks:
DMSO
True negative controls:
no
Positive controls:
yes
Positive control substance:
4-nitroquinoline-N-oxide
2-nitrofluorene
other: 2-aminoanthracene
Details on test system and experimental conditions:
The Ames II test was conducted with and without addition of microsomal liver enzymes from rats (Aroclor 1254-induced).
The following base-pair and frameshift-specific tester strains were used: S. typhimurium TA Mix (TA 7001, TA 7002, TA 7003, TA 7004,
TA 7005 and TA 7006) and TA 98, respectively. The Salmonella strains were described by Gee et al. (1998).
Evaluation criteria:
The pH indicator bromocresol purple turns the colour of the cultures from blue to yellow
as the pH drops due to the accumulation of catabolites from the metabolic activity of
revertant cells. The number of positive wells (yellow) out of a total of 48 wells is an
indication of the frequency of reversion per replicate per dose and was compared to the
number of spontaneous revertant wells of the solvent control. Each test point contains
48 wells of a 384-well plate. In each 48-well section, the wells were scored for the number
of revertant wells (yellow) and the mean value of the triplicates was calculated.
The experiment is regarded valid, if the vehicle control showed the normal spontaneous
revertant frequency and the diagnostic mutagens caused the expected increase in the
mutation rate. The individual test chemicals were classified according to the following
criteria:
Negative: ≤8/48 wells Equivocal: 9-12/48 wells Positive: ≥13/48 wells
Historical control range: 0-7/48 wells in ca 300 experiments (1999-up to date)
A concentration-dependent increase of revertant wells (mean of triplicate) over the vehicle
control is indicative for a genotoxic activity of the test substance.

Results and discussion

Test results
Species / strain:
other: S. typhimurium TA Mix (TA 7001, TA 7002, TA 7003, TA 7004, TA 7005 and TA 7006) and TA 98
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
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative

Based on the described results it is concluded, that BIBR 1048 Oxa-Ester, when tested up to recommended or insoluble concentrations,
caused neither base-pair substitutions nor frameshift mutations in bacteria. No evidence of genotoxic activity was observed in a series
of S. typhimurium tester strains (TA Mix and TA 98) in the absence and presence of metabolic activation. The test article is, therefore,
classified as "Ames II negative".