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Toxicological information

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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP study according to guideline

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2001
Report Date:
2002

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Version / remarks:
1997
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent

Method

Species / strain
Species / strain / cell type:
other: TA98, TA100, TA1535, TA1537, TA102
Details on mammalian cell type (if applicable):
not applicable
Metabolic activation:
with and without
Metabolic activation system:
S9 mix from Phenobarbital (i.p.) and beta-Naphthoflavone (oral) induced rat liver
Test concentrations with justification for top dose:
TA98: 50, 160, 500, 750, 1000 µg/plate
TA100, TA102, TA1535, TA1537: 50, 160, 500, 1600, 5000 µg/plate
Vehicle / solvent:
DMSO
Controlsopen allclose all
Untreated negative controls:
yes
Remarks:
water
Negative solvent / vehicle controls:
yes
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
2-nitrofluorene
Remarks:
TA98

Migrated to IUCLID6: 2.5 µg/plate
Untreated negative controls:
yes
Remarks:
water
Negative solvent / vehicle controls:
yes
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
TA100, TA 1535

Migrated to IUCLID6: 5 µg/plate (TA100), 2.5 µg/plate TA1535
Untreated negative controls:
yes
Remarks:
water
Negative solvent / vehicle controls:
yes
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
mitomycin C
Remarks:
TA102

Migrated to IUCLID6: 2.5 µg/plate
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
9-aminoacridine
Remarks:
TA1537

Migrated to IUCLID6: 40 µg/plate
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
other: 100 µg/plate 2-Aminofluorene and 2.5 µg/plate 2-Aminoanthracene
Remarks:
all tester strains
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium; in agar (plate incorporation)


DURATION
- Exposure duration: 72h
-Titers of overnight cultures: > 10^8
- Selection time (if incubation with a selection agent): 72h



SELECTION AGENT (mutation assays): histidine deficient agar


NUMBER OF REPLICATIONS:triplicates, two assays


DETERMINATION OF CYTOTOXICITY
- Method: relative total growth with strains TA98 and TA100

Evaluation criteria:
- negative control in the laboratory historical range for each tester strain
- positive control chemicals produces responses in all tester strains within laboratory historical range
- Mean plate count should be at least two times the concurrent vehicle control group mean in both assays and in a dose dependent response
- selected dose range should include a clearly toxic concentration or should exhibit limited solubility as demonstrated by the preliminary toxicity range-finding test or should extend to 5 mg/plate
- To ensure that appropriate numbers of bacteria are plated, overnight culture titers must be in excess of 10^8 bacteria/ml
Statistics:
not applicable

Results and discussion

Test resultsopen allclose all
Species / strain:
other: TA1535, TA100
Metabolic activation:
with and without
Genotoxicity:
positive
Remarks:
with and without metabolic activation
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
5000 µg/plate
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
other: TA 98, TA 1537, TA102
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
5000 µg/plate
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:

COMPARISON WITH HISTORICAL CONTROL DATA: some values were insignificantly above historical control data, but they are regarded as acceptable


ADDITIONAL INFORMATION ON CYTOTOXICITY: In the pretest with the TA98 tester strain, toxicity and/or reduced background lawn was observed at 1600 and 5000 µg/plate
Remarks on result:
other: strain/cell type: TA 1535
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

Assay 1:

Conc (µg/plate)

TA1535

TA1537

TA 100

TA102

 

- S9

+ S9

- S9

+ S9

- S9

+ S9

- S9

+ S9

Control (water)

0.9

1.1

1.2

0.9

1.0

1.0

1.0

1.0

50

1.6

2.4

0.7

0.8

0.9

1.0

1.2

1.1

160

2.0

2.7

0.8

0.9

1.0

1.1

1.2

1.2

500

2.8

3.1

0.8

0.7

1.1

1.4

1.3

1.5

1600

3.9

6.3

0.8

0.7

1.6

1.6

1.4

1.9

5000

2.0#

5.0#

0.5*

0.8

1.6#

1.3#

0.7#

1.1#

Positive control

30.7

 

7.2

 

3.9

 

4.1

 

2-Aminofluorene

 

2.2

 

3.3

 

9.2

 

2.2

2- Aminoanthracene

 

7.2

 

5.6

 

7.5

 

1.5

 

Assay 2

Conc (µg/plate)

TA1535

TA1537

TA 100

TA102

 

- S9

+ S9

- S9

+ S9

- S9

+ S9

- S9

+ S9

Control (water)

0.8

1.5

1.2

1.1

1.0

1.1

1.1

1.0

25

1.2

1.5

1.1

1.0

1.0

1.1

1.0

1.1

160

1.4

2.2

1.1

1.0

1.2

1.0

1.0

1.4

250

1.6

2.3

0.9

1.0

1.4

1.2

1.0

1.4

1600

2.9

5.2

0.9

0.9

2.6

1.7

1.2

1.6

2500

4.5

6.4

0.6#*

0.8

2.9

2.2

0.2

1.4

Positive control

32.6

 

6.3

 

5.5

 

3.4

 

2-Aminofluorene

 

3.5

 

3.2

 

10.9

 

2.6

2- Aminoanthracene

 

10.6

 

4.8

 

7.7

 

1.8

 

TA98

Conc (µg/plate)

Assay 1

Assay2

 

 

 

- S9

+ S9

Control (water)

+ S9

- S9

Control (water)

1.4

1.2

50

1.2

0.9

50

1.0

0.9

160

1.1

0.9

160

1.0

1.1

250

1.0

0.8

500

1.3

1.2

500

1.0

0.8

750

1.2

1.1

750

0.7#

0.7

1000

1.1

0.8

2-nitrofluorene

0.9#

0.9

2-nitrofluorene

5.8

 

2-Aminofluorene

4.5

 

2-Aminofluorene

 

56.6

2- Aminoanthracene

 

41.0

2- Aminoanthracene

 

20.8

Control (water)

 

21.7

 

#: reduced background lawn

*: no background lawn, toxicity

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
positive

Test substance induce a mutagenic effect in S. typhimurium TA 100 and TA1535 with and without metabolic activation. It is therefore considered to be a bacterial mutagen.