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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:
19 Sep 2001 - 5 Dec 2001
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP-Guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2002
Report Date:
2002

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
Landesanstalt für Pflanzenbau und Pflanzenschutz Rheinland-Pfalz, Mainz
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Protectol PE
- Physical state: colorless liquid
- Analytical purity: 99.9%
- Purity test date: 18 Sep 2001
- Lot/batch No.: 66-4287
- Stability under test conditions: The stability of the test substance throughout the study period (in the vehicle DMSO over 4 hours and in water over 96 hours) was verified by reanalysis.
- Storage condition of test material: Room temperature (N2 conditions)
- Test Substance No.: 01/0498-1

Method

Target gene:
his-operon (S. typhimurium)
trp-operon (E. coli)
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:
S-9 mix. At least 5 male Sprague-Dawley rats received a single intraperitoneal injection of 500 mg Aroclor 1254 per kg body weight. After 5 days, rats were sacrificed, the livers were prepared, and the S-9 fraction was obtained by standard methods.
Test concentrations with justification for top dose:
0, 20, 100, 500, 2500 and 5000 µg/plate with and without metabolic activation
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: Due to the limited solubility of the test substance in water, DMSO was selected as the vehicle, which had been demonstrated to be suitable in bacterial reverse mutation tests and for which historical control data are available.
Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
other: With S-9 mix: 2-aminoanthracene (2-AA); without S-9 mix: N-methyl-N’-nitro-N-nitrosoguanidine (MNNG), 4-nitro-o-phenylenediamine (NOPD), 9-aminoacridine (AAC), 4-nitroquinoline-N-oxide (4-NQO)
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)


DURATION
- Preincubation period: 48-72 hours in the dark


NUMBER OF REPLICATIONS: 3 culture plates per strain per dose or per control were prepared.


NUMBER OF CELLS EVALUATED: Not applicable, as the number of revertant colonies was calculated.


DETERMINATION OF CYTOTOXICITY
- Method: cloning efficiency; relative total growth
Evaluation criteria:
The test chemical is considered positive in this assay if the following criteria are met:
A dose-related and reproducible increase in the number of revertant colonies, i.e. about doubling the spontaneous mutation rate in at least one tester strain either without S-9 mix or after adding a metabolizing system.

A test substance is 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.
Statistics:
not applicable

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:
cytotoxicity
Remarks:
No bacteriotoxic effect was observed in the standard plate test. In the preincubation assay a slight decrease in the number of revertants and/or slight reduction in the titer was occasionally observed depending on the strain and test conditions.
Species / strain:
E. coli WP2 uvr A
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
No bacteriotoxic effect was observed in the standard plate test. In the preincubation assay a slight decrease in the number of revertants and/or slight reduction in the titer was occasionally observed depending on the strain and test conditions.

Any other information on results incl. tables

Refer to attached background material

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative