Tributyl-2-methoxypropylphosphonium salt with 4,4'-[2,2,2-trifluoro-1-(trifluoromethyl)ethylidene]bis[phenol]

Genetic toxicity: in vitro

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:

31 May 2005 to 16 June 2005

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study conducted under GLP conditions

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2006

Report date:

2006

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Test guideline 2

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

Test material information

Constituent 1

Reference substance name:

MTDID 3285

IUPAC Name:

MTDID 3285

Details on test material:

- Substance type: Light brown waxy chunks
- Physical state: Solid
- Analytical purity: Minimum 80%
- Lot/batch no.: Lot 30045
- Expiration date of the lot/batch: 01 June 2006
- Stability under test conditions: Stable
- Storage condition of test material: At room temperature in the dark
- Other: Molecular weight: 610; Stable at higher temperatures (max 50 degrees C for maximum of 30 to 60 hours).

Method

Target gene:

histidine requiring Salmonella typhimurium bacterial strains resulting in histidine-independent strains and in tryptophan-requiring Escherichia coli bacterial strain resulting in a tryptophan-independent strain.

Species / strain

Species / strain / cell type:

other: S. typhimurium TA 1535, TA 1537, TA 98 and TA 100 and Escherichia coli WP2uvrA

Additional strain / cell type characteristics:

other: S.typhi strains are histidine dependant and E.coli strain is tryptophan dependant.

Metabolic activation:

with and without

Metabolic activation system:

S9 mix (5% and 10% v/v)

Test concentrations with justification for top dose:

3, 10, 33, 100, 333, 1000, 3330 and 5000 micrograms/plate.

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: Dimethyl sulfoxide
- Justification for choice of solvent/vehicle: The test article completely dissolves in DMSO (when treated with ultrasonic waves)

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

other: The solvent, DMSO, was used as a negative control

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

other: 2-aminoanthracene (2AA) was used as a positive control for all strains with metabolic activation. For further details see Remarks field.

Remarks:

For the strains without metabolic activation, sodium azide, 9-aminacridine, 2-nitrofluorene, methylmethanesulfonate and 4-nitroquinoline N-oxide were used as positive controls for strain TA1535, TA 1537, TA98, TA 100 and WP2uvrA, respectively.

Details on test system and experimental conditions:

METHOD OF APPLICATION
- in agar (plate incorporation)
DURATION
- expression time (cells in growth medium): 48 hours
NUMBER OF CELLS EVALUATED:
- revertant colonies were counted manually if there were less than 40 coloneis present. If more than 40 colonies were present, these could be counted automatically with a Protos model 50000-colony counter.
DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; cloning efficiency; relative total growth; other

Evaluation criteria:

A test article is considered negative in the test if A) the total number of revertants in any tester strain at any concentration is not greather than two times the solvent control value, with or without metabolic activiation. B) The negative response should be reproducible in at elast one independently repeated experiment.
A test article is consdiered mositive in the test is A) it indueced at least a 2-fold, dose related increase in the number of revertant with respect to the number induced by the solvent control in any of the tester strains, either with or without metabolic activiation. However, any mean palet count of elss that 20 is considered not to be biologically relevant. B) In case a postiive response will be repeated, the postiive response should be reproducible in at least one independently repeated experiment.
These criteria are not absolute and other modifying factos might enter into the final evaluation decision.

Statistics:

only number of revertant colonies were noted, no formal hypothesis tested.

Results and discussion

Test results

Species / strain:

other: S. typhimurium TA 1535, TA 1537, TA 98 and TA 100 and WP2uvrA

Metabolic activation:

with and without

Genotoxicity:

negative

Remarks:

In the first and second mutation assays, no increases in the number of revertants were observed upon treatment with the test article under all conditions tested.

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

observed in all tester strains at concentrations at or above 333 micrgorams/plate

Vehicle controls validity:

not applicable

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

RANGE-FINDING/SCREENING STUDIES: The test article was tested in the tester strains TA100 and WP2uvrA with concentrations of 3, 10, 33, 100, 333, 1000 and 5000 micrograms/plate in the absence and presence of 5% (v/v) S9-mix. In the dose range finding test, the test article was tested up to concentrations of 5000 micrograms/plate in the absence and presence of S9-mix in the strains TA100 and WP2uvrA. The test article precipitated on the plates at dose levels of 3330 and 5000 micrograms/plate. In tester strain TA100, toxicity was observed at dose levels of 333 micrgorams per plate and above in the absence and presence of S9-mix. In tester strain WP2uvrA, toxicity was observed at dose levels of 1000 micrograms/plate and upwards in the absence of S9-mix and 3330 and 5000 micrograms/plate in the presence of S9-mix. Based on the dose range finding test, the test article was tested in the first mutation assay at a concentration range of 3 to 333 micrograms/plate in absence of S9--mix and at a concentration range of 666 micrograms/plate in the presence of 5% (v/v) S9-mix in tester strains T1535, TA1537 and TA98. For the second mutation assay, strains TA1535, TA1537 and TA100 , concentrations of 3, 10, 33, 100 and 333 micrograms/plate were used without S9-mix. With S9-mix, 10, 33, 100, 333 and 666 micrograms/plate were tested. In the TA98 tester strain, 10, 33, 100, 333 and 666 micrograms per plate were used. In the WP2uvrA tester strain, 33, 100, 333, 1000 and 3330 micrograms/plate were used.
COMPARISON WITH HISTORICAL CONTROL DATA: The negative and strain specific positive control values were within the laboratory histroical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly.

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 results of this study, it is concluded that the test article is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.

Executive summary:

The mutagenic activity of the test article in the Salmonella typhimurium reverse mutation assay and the Escherichia coli reverse mutation assay was assessed. Histidine-requiring strains of Salmonella typhimurium (TA 1535, TA1536, TA100 and TA98) and tryptophan requiring strain of Escherichia coli (WP2uvrA) were used as the tester strains in this study. The test was performed in two independent experiments in the presence and absence of S9-mix. The study procedure was based on most recent OECD and EEC guidelines. The test substance was dissolved in dimethyl sulfoxide of spectroscopic quality.Based on the results of the dose range finding test, the test article was tested in the first mutation assay at a concentration range of 3 to 333 micrograms/plate in the absence of S9-mix and at a concentration range of 10 to 666 micrograms/plate in the presence of 5% S9 mix in tester strains TA1535, TA1537 and TA98. In an independent repeat of the assay with additional parameters, the test article was tested at a concentration range of 3 to 333 micrograms/plate in the absence of S9-mix in tester strains TA100, TA1535 and TA1537. In tester strain TA98, the test article was tested at concentration of 33 to 3330 micrograms/plate in the absence and presence of 10% S9-mix. The test article precipitated on the plates at the top dose of 3330 micrograms/plate. Cytotoxicity was observed in all tester strains. The test article did not induce a dose-related, two-fold increase in the number of revertant (His+) colonies in each of the four tester strains (TA 1535, TA1537, TA100 and TA98) and in the number of revertant (Trp+) colonies in tester strain WP2uvrA both in the absence and presence of S9-metabolic activation. These results were confirmed in an independently repeated experiment. In this study, 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. Based on the results of this study, it is concluded that the test article is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.