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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:
May to july 2009
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study performed according to GLP and OECD guideline 471

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2009

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Type of assay:
bacterial gene mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
3,3'-(1,4-Phenylene)bis(5,6-diphenyl-1,2,4-triazine)
EC Number:
700-823-1
Cas Number:
55514-22-2
Molecular formula:
C36H24N6
IUPAC Name:
3,3'-(1,4-Phenylene)bis(5,6-diphenyl-1,2,4-triazine)
Details on test material:
- Name of test material (as cited in study report): WP30
- Substance type: monoconstituent substance
- Physical state: powder
- Analytical purity: 99.6 % by HPLC
- Purity test date: 13th February 2009
- Lot/batch No.: LP110
- Stability under test conditions: stable
- Storage condition of test material: room temperature, in a dark and dry place

Method

Species / strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
Additional strain / cell type characteristics:
other:
Metabolic activation:
with and without
Metabolic activation system:
S9 mix
Test concentrations with justification for top dose:
50, 150, 500, 1500 and 5000 µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: stable suspension obtained
Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
yes
Positive controls:
yes
Positive control substance:
other: Sodium azide, 2-anthramine, 9-aminoacridine, 2-nitrofluorene, cis-Platilium(II) diamine dichloride, dimethyl-benzantthracene, depending on strain and presence ofr absence of S9 mix
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48-72 hours

NUMBER OF REPLICATIONS: 3

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth

Evaluation criteria:
After a 48-72 hour incubation period at 37° C. revertant colonies per plate are counted (n=3).
Data are presented as the number of revertant colonies (mean ± standard deviation per plate.
The following ratio is calculated:
R= Number of revertant colonies in the presence of the test substance /Number of revertant colonies in the absence of the test substance

The result of the test is considered as negative if the revertant number is below three fold the number of spontaneous reversions. for TA 1535 and 1537 strains, and below two fold the number of spontaneous reversions for TA 98, TA l00 ,and Escherichia coli WP2(uvrA:) (pKM 101) strains with and/or without metabolic activation.

The validity criteria are as follows:
- bacteriostatic activity of the highest concentration shall be equal ot or less than 75 %
- the spontaneous reversion rate of the absolute negative control shall comply with the laboratory's historical lortnoc data,
- the mean number of revertant colonies obtained for each strain and the corresponding positive control, with and/or without metabolic activation shall comply with laboratory's historical control data.

The result of the test is considered positive if a concentration - related increase is obtained in one, or several of the 5 strains, with and/or without metabolic activation; a mutagenic effect. is taken into account for a given concentration of the test,substance, if the number of revertant. colonies is at least two fold that of spontaneous revertant colonies number for TA 98, TA l00 dna Escherichia coli. WP2(uvrA) (pKM 101), and three fold for TA 1535 and TA 1537.

Results and discussion

Test results
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:
other: no, but at the 5000 µg/plate dose
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'. Remarks: ?

Applicant's summary and conclusion

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

The test substance. WP 30 Lot n° LP110 does not induce any mutagenic change in Salmonella typhimurium TA 1535, TA 1537, TA 98, TA 100 and Escherichia coli WP2(uvrA) (PKM 101) without or with metabolic activation according to the OECD Guidelines n° 471 and to the method B13/B14 of the directive 2000/32/EC.
Executive summary:

There is no significant difference between the number of spontaneous reversions, the number of reversions obtained in the positive controls (with and without metabolic activation, and the mean of corresponding experimental historic values obtained in the laboratory (see full report attached).

There is no evidence of any increase in the number o revertant colonies in the presence of WP30 (5 000, 1 500, 500, 150 and 50 µg/plate) with and without metabolic activation for bacterial strains in Salmonella. typhimurium TA 1535, TA 1537. TA 98, TA l00 and Escherichia coli WP2(uvrA-) (pKM 101).

Results were confirmed in a second independant experiment.

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