2-{[(2-{3-[(4-{Ethyl[3-(vinylsulfonyl)phenyl]amino}-6-fluoro-heteromonocycle-2-yl)amino]-2- (hydroxy-O)phenyl}hydrazono-N2)(phenyl)methyl]diazenyl-N1}benzoato(6-)-O]cuprate(4-) sodium salt, polysulfo

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:

2011-09-16 to 2011-10-21

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2011

Report date:

2011

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

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Bayerisches Landesamt für Gesundheit und Lebensmittelsicherheit, München, Germany

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Method

Target gene:

His

Species / strain

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

experiment I: rat S9 liver microsomal fraction experiment II: hamster S9 liver microsomal fraction (uninduced)

Test concentrations with justification for top dose:

31.6, 100, 316, 1000, 2500 and 5000 µg/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO

Controls

Untreated negative controls:

yes

Remarks:

A. dest., BSL Lot No. 110722, 110825, 111010

Negative solvent / vehicle controls:

yes

Remarks:

DMSO, Applichem Lot No. 1U004467

Positive controls:

yes

Positive control substance:

other:

Remarks:

Without metabolic activation: NaN3 for TA100, TA1535; 4-NOPD for TA98, TA1537; MMS for TA102. With metabolic activation: 2-aminoanthracene for all strains (rat liver) and TA1535, 1537 and 102 (hamster liver), Congo Red for TA98 and 100 (hamster liver)

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation); preincubation

DURATION
- Preincubation period: 30 min; 100 µL of the test item preparation was pre-incubated with the tester strains (100 µL) and sterile buffer or the metabolic activation system (500 µL) for 30 min at 30 °C prior to adding the overlay agar (2000 µL) and pouring onto the surface of a minimal agar plate.
- Exposure duration: at least 48 h; After solidification the plates were inverted and incubated at 37 °C for at least 48 h in the dark.


NUMBER OF REPLICATIONS: For each strain and dose level, including the controls, three plates were used.


DETERMINATION OF CYTOTOXICITY
- Method: clearing or rather diminution of the background lawn or a reduction in the number of revertants down to a mutation factor of approximately ≤ 0.5 in relation to the solvent control

Evaluation criteria:

The Mutation Factor is calculated by dividing the mean value of the revertant counts through the mean values of the solvent control (the exact and not the rounded values are used for calculation).
A test item is considered as mutagenic if:
- a clear and dose-related increase in the number of revertants occurs and/or
- a biologically relevant positive response for at least one of the dose groups occurs
in at least one tester strain with or without metabolic activation.
A biologically relevant increase is described as follows:
- if in tester strains TA 98, TA 100 and TA 102 the number of reversions is at least twice as high
- if in tester strains TA 1535 and TA 1537 the number of reversions is at least three times higher
than the reversion rate of the solvent control.

Statistics:

According to OECD guidelines, the biological relevance of the results is the criterion for the interpretation of results, a statistical evaluation of the results is not regarded as necessary.

Results and discussion

Test results

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Additional information on results:

No precipitation of the test item was observed in any tester strain used in experiment I and II (with and without metabolic activation).

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

No toxic effects of the test item were noted in any of the five tester strains used up to the highest dose group evaluated with and without metabolic activation in experiment I and II. The reduction in the number of revertants down to a mutation factor of 0.4 found in experiment I in tester strain TA 1537 at a concentration of 100 µg/plate (without metabolic activation) was regarded as not biologically relevant due to lack of a dose-response relationship.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

Executive summary:

In a reverse gene mutation assay in bacteria, strains TA98, TA100, TA1535, TA1537 and TA102 of S. typhimurium were exposed to FAT 40853/A TE at concentrations of 31.6, 100, 316, 1000, 2500 and 5000 µg/plate (experiment I and II), in the presence and absence of mammalian metabolic activation according to the plate incorporation method with rat liver S9 (experiment I) and the pre-incubation method with hamster liver S9 (experiment II).

FAT 40853/ A TE was tested up to the limit concentration of 5000 µg/plate in all tester strains used.

The positive controls induced the appropriate responses in the corresponding strains. There was no evidence of induced mutant colonies over background.

This study is classified as acceptable. This study satisfies the requirement for Test Guideline OPPTS 870.5100; OECD 471 for in vitro mutagenicity (bacterial reverse gene mutation) data.