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:
1999
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline Study (OECD), GLP.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2000
Report Date:
2000

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Version / remarks:
(July 21, 1997)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Deviations:
no
GLP compliance:
yes
Remarks:
Department of toxicolgy, BASF AG
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name as cited in report: Adipic acid, cyclic ester with diethyleneglycol
- Analytical purity: ca. 90%
- Physical state: Solid/white crystals
- Storage: refrigerated
- Lot/batch No.: B 74
- Stablity: verified by reanalysis

Method

Target gene:
S. typhimurium: his-
E. coli: trp -
Species / strain
Species / strain / cell type:
other: TA 1535, TA 100, TA 1537, TA 98 and E . coli WP2 uvrA
Metabolic activation:
with and without
Metabolic activation system:
S9 mix from Aroclor-induced liver of male Sprague-Dawley rat (200 - 300g).
Test concentrations with justification for top dose:
STANDARD PLATE TEST (SPT): 0; 20; 100; 500; 2,500 and 5,000 µg/plate
PREINCUBATION TEST (PIT): 0; 4; 20; 100; 500 and 2,500 µg/plate
Vehicle / solvent:
- DMSO
- Justification for choice: Due to the limited solubility of the test substance in water, DMSO, which had been demonstrated to be suitable in bacterial reverse mutation tests and for which historical control data were available, was selected as the vehicle.
Controls
Untreated negative controls:
yes
Remarks:
(sterility control)
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
other: With S9 Mix; 2-aminoanthracene (2-AA) and without S9 mix; N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), 4-nitro-o-phenylendiamine (NOPD), 9- aminoacridine (AAC), 4-nitroquinoline-N-oxide (4-NQO)
Remarks:
The stability of the selected positive controls was well-defined under the selected culture conditions, since they are well-established reference mutagens.
Details on test system and experimental conditions:
1). STANDARD PLATE TEST (SPT) (in agar incorporation method)
- Exposure duration: 48 - 72 h (in the dark) @ 37°C

2). PREINCUBATION TEST (PIT)
- Preincubation period: 30 min
- Exposure duration: 48 - 72 h (in the dark) @ 37°C


DETERMINATION OF CYTOTOXICITY
Toxicity was detected by a
- decrease in the number of reve rtants
- clearing or diminution of the background lawn (= reduced his" or trp" background growth)
- reduction in the titer
Evaluation criteria:
The test chemical was considered positive in this assay if a dose-related and reproducible increase in the number of revertant colonies, i .e. about doubling of the spontaneous mutation rate in at least one tester strain either without S-9 mix or after adding a metabolizing system, was seen. A test substance was generally considered nonmutagenic in this test if the number of revertants for all tester strains were within the historical negative control range under all experimental conditions in two experiments carried out independently of each other.

Results and discussion

Test results
Species / strain:
other: TA 1535, TA 100, TA 1537, TA 98 and E . coli WP2 uvrA
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
> 500 µg
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
An increase in the number of his+ or trp+ revertants was not observed in the standard plate test or in the preincubation test either without S-9 mix or after the addition of a metabolizing system.

ADDITIONAL INFORMATION ON CYTOTOXICITY:
A slight decrease in the number of revertants was observed in the standard plate test depending on the strain and test conditions from about 500 µg - 2,500 µg/plate onward. In the preincubtion assay, a weak bacteriotoxicity (reduced background growth, slighty decrease in the number of revertants and/or slight reduction in the titer) was observed depending on the strain and test conditions from about 500 pg/plate onward.

Any other information on results incl. tables

Standard Plate Test

Strain

Metabolic activation system

Replicates

Revertant factor range

dose dependency

Assessment

TA 98

no

3

0.5 - 0.9

no

negative

yes

3

0.5 - 0.8

no

negative

TA 100

no

3

0.7 - 0.9

no

negative

yes

3

0.5 - 0.9

no

negative

TA 1535

no

3

0.4 - 0.9

no

negative

yes

3

0.4 - 0.9

no

negative

TA 1537

no

3

0.4 - 1.0

no

negative

yes

3

0.4 - 0.7

no

negative

WP2uvrA

no

3

0.7 - 0.8

no

negative

yes

3

0.8 - 1.0

no

negative

Preincubation Test

Strain

Metabolic activation system

Replicates

Revertant factor range

dose dependency

Assessment

TA 98

no

3

0.6 - 0.9

no

negative

yes

3

0.6 - 0.8

no

negative

TA 100

no

3

0.9 - 1.0

no

negative

yes

3

0.8 - 1.0

no

negative

TA 1535

no

3

0.6 - 0.9

no

negative

yes

3

0.5 - 0.9

no

negative

TA 1537

no

3

0.6 - 0.9

no

negative

yes

3

0.6 - 0.9

no

negative

WP2uvrA

no

3

0.9 - 1.0

no

negative

yes

3

0.7 - 0.9

no

negative

Applicant's summary and conclusion