2,2'-[(3,3'-dichloro[1,1'-biphenyl]-4,4'-diyl)bis(azo)]bis[N-(4-methoxyphenyl)-3-oxobutyramide]

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in mammalian cells

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 21 MAR 1994 to 14 JUN 1994

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study (OECD 476)

Data source

Reference

Title:

Unnamed

Year:

1994

Report date:

1994

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

in accordance with Directive 88/320/EEC

Type of assay:

mammalian cell gene mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

2,2'-[(3,3'-dichloro[1,1'-biphenyl]-4,4'-diyl)bis(azo)]bis[N-(4-methoxyphenyl)-3-oxobutyramide]

EC Number:

250-797-6

EC Name:

2,2'-[(3,3'-dichloro[1,1'-biphenyl]-4,4'-diyl)bis(azo)]bis[N-(4-methoxyphenyl)-3-oxobutyramide]

Cas Number:

31775-16-3

Molecular formula:

C34H30Cl2N6O6

IUPAC Name:

2,2'-[(3,3'-dichloro[1,1'-biphenyl]-4,4'-diyl)bis(azo)]bis[N-(4-methoxyphenyl)-3-oxobutyramide]

Method

Target gene:

TK

Species / strain

Species / strain / cell type:

mouse lymphoma L5178Y cells

Details on mammalian cell type (if applicable):

- Type and identity of media: RPMI 1640 medium supplemented with 10% donor horse serum and 20 mM HEPES buffer (R10)
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: no data
- Periodically checked for karyotype stability: no data
- Periodically "cleansed" against high spontaneous background: yes

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

rat liver S9 mix from Aroclor 1254 pretreated rats

Test concentrations with justification for top dose:

62.5, 125, 250, 500, 1000 µg/l

Vehicle / solvent:

- dimethyl sulphoxide

Controls

Controls 1

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

cyclophosphamide

Remarks:

with metabolic activation

Controls 2

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

ethylmethanesulphonate

Remarks:

without metabolic activation

Details on test system and experimental conditions:

DURATION
- Exposure duration: 3 hrs with manual shaking at approximately 0.5 hour intervals
- Expression time (cells in growth medium): 24 hrs
- Selection time (if incubation with a selection agent): 10-14 days


SELECTION AGENT (mutation assays): trifluorothymidine (4 µg/ml)

NUMBER OF REPLICATIONS: 2 experiments, treatments performed in duplicate, both with and without metabolic activation

DETERMINATION OF CYTOTOXICITY
- Method: colony formation

Results and discussion

Test results

Species / strain:

mouse lymphoma L5178Y cells

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: a precipitate of the test item was noted at all dose levels

RANGE-FINDING/SCREENING STUDIES:
- the maximum dose level of 1000 µg/ml was selected on the basis that it was the maximum practical dose level that could be achieved

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

- no marked reductions in viability of cells treated with the test item in the presence or absence of metabolic activation

- vehicle controls were within the normal range (10 - 125 x 10E-6)

- positive controls gave marked increases in mutant frequency, demonstrating that the test system was operating satisfactorily

- the test item did not induce any statistically significant or dose related increase in mutant frequency

- experiment 1 and 2 gave concordant results

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative without metabolic activation
negative with metabolic activation

The test substance did not increase the mutant frequency at the TK+/- locus in L5178Y cells and is therefore considered to be non-mutagenic under the conditions of the test.

Executive summary:

Mouse Lymphoma cells (L5178Y TK+/-) were treated with the test substance at five concentrations in the range of 62.5 to 1000 µg/ml. A heavy precipitate of the test substance was noted at all dose levels. The solvent control gave mutant frequencies within the normal range. The positive controls gave marked increases in the mutant frequency, both in the presence or absence of metabolic activation, indicating the satisfactory performance of the test system. The test substance demonstrated no statistically significant or dose related increases in mutant frequency at any dose level, either with or without metabolic activation, thus indicating that the test item is not mutagenic under the conditions tested.