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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1999-11-18 to 2000-01-24
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
The test substance was similar to the registered substance but contained a higher proportion of an S3 isomer.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2000
Report Date:
2000

Materials and methods

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

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Type:
Constituent

Method

Species / strain
Species / strain / cell type:
E. coli WP2 uvr A
Metabolic activation:
with and without
Metabolic activation system:
Aroclor induced rat liver S9
Test concentrations with justification for top dose:
75-5000 µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO

- Justification for choice of solvent/vehicle: Sponsor's request and compatibility with target cells.
Controlsopen allclose all
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: 2-aminoanthracene
Remarks:
All Salmonella Strains, WP2 uvrA (with activation)
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
2-nitrofluorene
Remarks:
TA 98 (without activation)
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
TA 100, TA 1535 (without activation)
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
9-aminoacridine
Remarks:
TA 1537 (without activation)
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
methylmethanesulfonate
Remarks:
WP2 uvrA (without activation)
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar, preincubation


DURATION
- Preincubation period: 60 minutes

- Expression time (cells in growth medium): 48 to 72 hours

- Fixation time (start of exposure up to fixation or harvest of cells): 12 hours

NUMBER OF REPLICATIONS: 3 plates for each test concentration; study repeated

DETERMINATION OF CYTOTOXICITY
- Method: Background lawn monitoring
Evaluation criteria:
For the test article to be positive, it must cause a dose-related increase in mean revertants per plate of at least one tester strain with a minimum of two increasing concentrations per test article.

A result is positive if the number of revertants is significantly increased compared with the solvent control to at least 2-fold of the solvent control for TA 98, TA 100 and WP2 uvrA, and 3-fold of the solvent control for TA 1535 and TA 1537.

Cytotoxicity is defined as a reduction in the number of colonies by >50% compared with the solvent control and/or a sparse background lawn.

Results and discussion

Test resultsopen allclose all
Key result
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:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
Positive controls validity:
valid
Key result
Species / strain:
E. coli WP2 uvr A
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
Positive controls validity:
valid
Additional information on results:
COMPARISON WITH HISTORICAL CONTROL DATA: Results were within range of historical control data

Any other information on results incl. tables

Table 2: Experiment 1 Preliminary toxicity assay Number of revertants per plate

 

TA98

TA100

TA1535

Conc.
(
µg/plate)

— MA

+ MA

Cytotoxic
(yes/no)

 MA

+ MA

Cytotoxic
(yes/no)

MA

+ MA

Cytotoxic
(yes/no)

0*

NC***

30

No

128

172

No

15

8

No

6.7

14

29

No

145

124

No

19

14

No

10

18

22

No

150

171

No

19

11

No

33

12

19

No

142

177

No

12

13

No

67

21

28

No

170

193

No

16

10

No

100

9

19

No

136

189

No

19

13

No

333

17

24

No

138

181

No

23

23

No

667

23

23

No

147

162

No

15

15

No

1000

14**

24**

No

132**

169**

No

19**

25**

No

3333

15**

17**

No

167**

184**

No

24**

16**

No

5000

21**

26**

No

139**

199**

No

23**

15**

No

*solvent control with DMSO

**Non-Interfering Precipitate

***No count due to procedural error in which plate did not receive an aliquot of tester strain

Table 2: Experiment 1 Preliminary toxicity assay Number of revertants per plate

 

TA1537

WP2 uvrA

Conc.
(
µg/plate)

— MA

+ MA

Cytotoxic
(yes/no)

— MA

+ MA

Cytotoxic
(yes/no)

0*

56

45

No

14

16

No

6.7

49

42

No

19

15

No

10

37

41

No

12

19

No

33

49

43

No

12

17

No

67

47

40

No

10

10

No

100

47

46

No

13

17

No

333

37

48

No

9

12

No

667

48

39

No

10

16

No

1000

37**

67**

No

11**

12**

No

3333

45**

52**

No

8**

18**

No

5000

44**

55**

No

15**

14**

No

*solvent control with DMSO

**Non-Interfering Precipitate

***No count due to procedural error in which plate did not receive an aliquot of tester strain

Table 3: Experiment 2 Preincubation mutagenicity assay, Number of revertants per plate (mean of 3 plates)

 

TA98

TA100

TA1535

Conc.
(
µg/plate)

— MA

+ MA

Cytotoxic
(yes/no)

MA

+ MA

Cytotoxic
(yes/no)

MA

+ MA

Cytotoxic
(yes/no)

0*

16

21

No

174

203

No

13

15

No

75

9

25

No

161

217

No

13

16

No

200

14

24

No

166

213

No

17

16

No

600

10

18

No

159

212

No

17

18

No

1800

13

17

No

169

232

No

14

16

No

5000

9

17

No

140

209

No

16

18

No

Positive control

981

1350

No

626

1197

No

370

140

No

*solvent control with DMSO

Table 3: Experiment 2 Preincubation mutagenicity assay, Number of revertants per plate (mean of 3 plates)

 

TA1537

WP2 uvrA

Conc.
(
µg/plate)

— MA

+ MA

Cytotoxic
(yes/no)

— MA

+ MA

Cytotoxic
(yes/no)

0*

8

9

No

12

14

No

75

4

6

No

9

12

No

200

5

5

No

8

13

No

600

4

9

No

9

11

No

1800

4

5

No

9

15

No

5000

5

8

No

9

6

No

Positive control

827

149

No

495

462

No

*solvent control with DMSO

Table 4: Experiment 2 Preincubation (Repeat) mutagenicity assay, Number of revertants per plate (mean of 3 plates)

 

TA98

TA100

TA1535

Conc.
(
µg/plate)

— MA

+ MA

Cytotoxic
(yes/no)

 MA

+ MA

Cytotoxic
(yes/no)

 MA

+ MA

Cytotoxic
(yes/no)

0*

17

16

No

152

160

No

11

10

No

75

14

14

No

145

171

No

7

13

No

200

15

19

No

135

189

No

7

15

No

600

14

10

No

137

159

No

10

9

No

1800

16

14

No

143

156

No

9

12

No

5000

15

12

No

115

173

No

8

7

No

Positive control

409

785

No

677

759

No

207

68

No

*solvent control with DMSO

Table 4: Experiment 2 Preincubation (Repeat) mutagenicity assay, Number of revertants per plate (mean of 3 plates)

 

TA1537

WP2 uvrA

Conc.
(
µg/plate)

— MA

+ MA

Cytotoxic
(yes/no)

— MA

+ MA

Cytotoxic
(yes/no)

0*

5

6

No

10

10

No

75

4

4

No

12

6

No

200

4

6

No

9

8

No

600

5

5

No

12

10

No

1800

2

7

No

9

12

No

5000

4

5

No

11

8

No

Positive control

719

71

No

317

75

No

*solvent control with DMSO

Applicant's summary and conclusion

Conclusions:
S2 has been tested a reliable assay conducted according to OECD TG 471 and in compliance with GLP. The test substance did not cause a positive response in S. typhimurium strains TA 1535, TA 1537, TA 98 and TA 100 and E. coli WP2 uvrA with or without metabolic activation in either the initial or repeat tests using the preincubation method. Appropriate solvent and positive controls were included and gave expected results. The test substance is negative for mutagenicity to bacteria under the conditions of the test.