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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:
20 to 28 October 2011
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study undertaken at a GLP accredited laboratory, to internationally accepted guidelines.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2011
Report date:
2011

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)
Deviations:
no
Qualifier:
according to guideline
Guideline:
JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Reference substance name:
LumiNova Blue Green
IUPAC Name:
LumiNova Blue Green
Details on test material:
- Name of test material (as cited in study report): LumiNova Blue Green
- Substance type: Fluorescent additive
- Physical state: Light yellowish-green powder
- Analytical purity: > 99%
- Purity test date: no data
- Lot/batch No.: PBM-029
- Expiration date of the lot/batch: 04 February 2012
- Stability under test conditions:
- Storage condition of test material: Room temperature in the dark
- Other: Date received 14 February 2011

Method

Target gene:
base change mutagens: S. typhimurium TA1535 and TA100, and E. coli WP2 uvrA
(pKM101).
frameshift mutagens: S. typhimurium TA1537 and TA98.
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 pKM 101
Metabolic activation:
with and without
Metabolic activation system:
S-9 mix
Test concentrations with justification for top dose:
5, 15, 50, 150, 500, 1500 and 5000 µg
Vehicle / solvent:
- Vehicle used: agar

- Justification for choice of vehicle: inert and commonly accepted.
Controlsopen allclose all
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
In absence of S-9 mix
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
9-aminoacridine
Remarks:
In absence of S-9 mix
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
2-nitrofluorene
Remarks:
In absence of S-9 mix
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
4-nitroquinoline-N-oxide
Remarks:
In absence of S-9 mix
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: 2-Aminoanthracene
Remarks:
In presence of S-9 mix
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
benzo(a)pyrene
Remarks:
In presence of S-9 mix
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Preincubation period: 10 hours at 37°C
- Exposure duration:
- Expression time (cells in growth medium): 24 hours at 37°C
- Selection time (if incubation with a selection agent):
- Fixation time (start of exposure up to fixation or harvest of cells):

NUMBER OF REPLICATIONS: 1

NUMBER OF CELLS EVALUATED: 10E+09 / ml

DETERMINATION OF CYTOTOXICITY
- Method: mutagenic sensitivity
Evaluation criteria:
For a test to be considered valid, the mean of the vehicle control revertant colony numbers for each strain should lie within or close to the current historical control range of the laboratory unless otherwise justified by the Study Director. The historical range is maintained as a rolling record over a maximum of five years. Also, the positive control compounds must induce an increase in mean revertant colony numbers of at least twice (three times in the case of strains TA1535 and TA1537) the concurrent vehicle controls. Mean viable cell counts in the 10-hour bacterial cultures must be at least 10E+09 / mL.

If exposure to a test substance produces a reproducible increase in revertant colony numbers of at least twice (three times in the case of strains TA1535 and TA1537) the concurrent vehicle controls, with some evidence of a positive dose-response relationship, it is considered
to exhibit mutagenic activity in this test system. No statistical analysis is performed. If exposure to a test substance does not produce a reproducible increase in revertant colony numbers, it is considered to show no evidence of mutagenic activity in this test system. No statistical analysis is performed.

If the results obtained fail to satisfy the criteria for a clear “positive” or “negative” response, even after additional testing, the test data may be subjected to analysis to determine the statistical significance of any increases in revertant colony numbers. The statistical procedures used are those described by Mahon et al (1989) and are usually Dunnett’s test followed, if appropriate, by trend analysis. Biological importance should always be considered along with statistical significance. In general, treatment-associated increases in revertant colony numbers below two or three times the vehicle controls (as described above) are not considered biologically important. It should be noted that it is acceptable to conclude an equivocal response if no clear results can be obtained.

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:
not determined
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Species / strain:
E. coli WP2 uvr A pKM 101
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not determined
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative

It is concluded that LumiNova Blue Green showed no evidence of mutagenic activity in this bacterial system under the test conditions employed.