Registration Dossier

Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
12 Oct 2017 - 09 Jan 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2018
Report Date:
2018

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Version / remarks:
July 1997
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Version / remarks:
May 2008
Qualifier:
according to
Guideline:
EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)
GLP compliance:
yes (incl. certificate)
Remarks:
GLP-Landesleitstelle Bayer, Bayerrisches Landesamt für Gesundheit und Lebensmittelsicherheit
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid

Method

Target gene:
His operon
Species / strain
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
Metabolic activation:
with and without
Metabolic activation system:
Post mitochondrial supernatant (S9) prepared from livers of Phenobarbital/β-naphthoflavone-induced male rats
Test concentrations with justification for top dose:
Experiment I (the incorporation test)
0.01, 0.0316, 0.1, 0.316, 1.0, 2.5 and 5.0 μL/plate

Experiment II (the pre-incubation test)
0.00316, 0.01, 0.0316, 0.1, 0.316, 1.0, 2.5 and 5.0 μL/plate
Vehicle:
- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: the solvent was compatible with the survival of the bacteria and the S9 activity.
Controls
Negative controls:
yes
Remarks:
A. dest.
Solvent controls:
yes
Remarks:
DMSO
True negative controls:
no
Positive controls:
yes
Positive control substance:
sodium azide
methylmethanesulfonate
other: 4-nitro-o-phenylene-diamine, 10 µg/plate for TA 98, 40 µg/plate for TA 1537, without metabolic activation; 2-aminoanthracene, 2.5 µg/plate; 10 µg/plate for TA 102, with metabolic activation
Details on test system and conditions:
METHOD OF APPLICATION: in agar (plate incorporation); preincubation

DURATION
- Preincubation period: 60 min
- Exposure duration: 48 – 72 h

NUMBER OF REPLICATIONS: 3

DETERMINATION OF CYTOTOXICITY
- Method: the diminution of the background lawn or a significant reduction in the number of revertants in solvent control.
Evaluation criteria:
Negative control data within historical range. Positive controls show a distinct enhancement of revertant rates over the control plate. At least five different concentrations of each tester strain are analyzable.
Positive response: a clear and dose-related increase in the number of revertants occurs and/or a biologically relevant positive response for at least one of the dose groups occurs in at least one tester strain with or without metabolic activation.
A biologically relevant increase is described as:
- if in tester strains TA 98 and TA 100 the number of reversions is at least and twice as high
- if in tester strains TA 102 the number of reversions is at least 1.5-fold higher
- if in tester strains TA 1535 and TA 1537 the number of reversions is at least three times higher than the reversion rate of the solvent control.
Negative response: neither a dose related increase in the number of revertants nor a reproducible biologically relevant increase.

Results and discussion

Test results
Key result
Species / strain:
other: TA 98, TA 100, TA 1535, TA 1537, TA 102
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity:
yes
Vehicle controls valid:
yes
Negative controls valid:
yes
Positive controls valid:
yes
Additional information on results:
RANGE-FINDING/SCREENING STUDIES: A pre-experiment for toxicity was performed at 0.00316, 0.0100, 0.0316, 0.100, 0.316, 1.0, 2.5 and 5.0 µL/plate with tester strains TA 98 and TA 100. The test substance were observed toxic effects at the concentration of 0.316 µL/plate or higher.

HISTORICAL CONTROL DATA
- Positive historical control data: Positive controls were met the criteria of validity
- Negative (solvent/vehicle) historical control data: Negative controls were within historical norms

Any other information on results incl. tables

Table 1: Results Experiment I

 

TesterStrain:TA98                                                                                    Experiment:1

 

Treatment

 

Dose/plate

REVERTANT COLONIES PER PLATE

MUTATION FACTOR

 

Without activation (-S9)

With activation (+S9)

 

Counts

Mean

SD

Counts

Mean

SD

-S9

+S9

 

 

27

 

 

24

 

A. dest.

 

29

26

3.1

17

25

8.0

1.1

1.1

 

 

23

 

 

33

 

 

 

 

 

 

23

 

 

21

 

 

 

 

DMSO

 

31

24

7.0

26

23

2.5

1.0

1.0

 

 

17

 

 

23

 

 

 

 

 

 

24

 

 

28

 

 

 

 

Test Item

0.0100 µL

22

24

1.5

27

38

17.6

1.0

1.6

 

 

25

 

 

58

 

 

 

 

 

 

23

 

 

23

 

 

 

 

Test Item

0.0316 µL

20

18

6.8

23

24

1.7

0.7

1.0

 

 

10

 

 

26

 

 

 

 

 

 

16

 

 

19

 

 

 

 

Test Item

0.100 µL

34

23

9.9

28

26

5.9

1.0

1.1

 

 

18

 

 

30

 

 

 

 

 

 

46

 

 

18

 

 

 

 

Test Item

0.316 µL

26

33

11.0

22

20

2.1

1.4

0.9

 

 

28

 

 

21

 

 

 

 

 

 

17

 

 

24

 

 

 

 

Test Item

1.0 µL

13

17

4.5

27

25

2.1

0.7

1.1

 

 

22

 

 

23

 

 

 

 

 

 

9

 

 

27

 

 

 

 

Test Item

2.5 µL

15

12

3.0

24

23

5.1

0.5

1.0

 

 

12

 

 

17

 

 

 

 

 

 

6

 

 

21

 

 

 

 

Test Item

5.0 µL

11

8

2.6

20

22

2.1

0.3

0.9

 

 

7

 

 

24

 

 

 

 

 

 

367

 

 

/

 

 

 

 

4-NOPD

10 µg

489

451

73.2

/

/

/

19.1

/

 

 

498

 

 

/

 

 

 

 

 

 

/

 

 

3261

 

 

 

 

2-AA

2.5 µg

/

/

/

2784

2986

246.9

/

128.0

 

 

/

 

 

2912

 

 

 

 

TesterStrain:TA100                                                                                      Experiment:1

 

Treatment

 

Dose/plate

REVERTANT COLONIES PER PLATE

MUTATION FACTOR

 

Without activation (-S9)

With activation (+S9)

 

Counts

Mean

SD

Counts

Mean

SD

-S9

+S9

 

 

103

 

 

95

 

A. dest.

 

95

94

9.5

87

93

5.3

1.1

1.0

 

 

84

 

 

97

 

 

 

 

 

 

75

 

 

93

 

 

 

 

DMSO

 

103

89

14.0

81

92

10.5

1.0

1.0

 

 

90

 

 

102

 

 

 

 

 

 

87

 

 

63

 

 

 

 

Test Item

0.0100 µL

69

82

11.0

80

71

8.5

0.9

0.8

 

 

89

 

 

71

 

 

 

 

 

 

74

 

 

92

 

 

 

 

Test Item

0.0316 µL

60

66

7.1

104

91

13.5

0.7

1.0

 

 

65

 

 

77

 

 

 

 

 

 

52

 

 

92

 

 

 

 

Test Item

0.100 µL

112

73

34.1

71

75

15.0

0.8

0.8

 

 

54

 

 

63

 

 

 

 

 

 

35

 

 

64

 

 

 

 

Test Item

0.316 µL

44

39

4.5

61

67

7.4

0.4

0.7

 

 

39

 

 

75

 

 

 

 

 

 

44

 

 

31

 

 

 

 

Test Item

1.0 µL

53

44

9.5

66

51

18.2

0.5

0.6

 

 

34

 

 

57

 

 

 

 

 

 

46 B

 

 

39

 

 

 

 

Test Item

2.5 µL

48 B

46

2.5

51

49

9.6

0.5

0.5

 

 

43 B

 

 

58

 

 

 

 

 

 

39 B

 

 

54 B

 

 

 

 

Test Item

5.0 µL

64 B

49

13.4

57 B

59

5.7

0.5

0.6

 

 

43 B

 

 

65 B

 

 

 

 

 

 

726

 

 

/

 

 

 

 

NaN3

10 µg

732

726

6.0

/

/

/

8.1

/

 

 

720

 

 

/

 

 

 

 

 

 

/

 

 

1900

 

 

 

 

2-AA

2.5 µg

/

/

/

2754

2233

457.0

/

24.3

 

 

/

 

 

2045

 

 

 

 

TesterStrain:TA1535                                                                                    Experiment:1

 

Treatment

 

Dose/plate

REVERTANT COLONIES PER PLATE

MUTATIONFACTOR

 

Withoutactivation(-S9)

With activation (+S9)

 

Counts

Mean

SD

Counts

Mean

SD

-S9

+S9

 

 

24

 

 

32

 

A. dest.

 

26

25

1.2

19

23

7.5

1.0

1.0

 

 

24

 

 

19

 

 

 

 

 

 

28

 

 

20

 

 

 

 

DMSO

 

24

25

2.3

32

23

7.6

1.0

1.0

 

 

24

 

 

18

 

 

 

 

 

 

21

 

 

25

 

 

 

 

Test Item

0.0100 µL

25

23

2.1

12

23

10.1

0.9

1.0

 

 

24

 

 

32

 

 

 

 

 

 

33

 

 

12

 

 

 

 

Test Item

0.0316 µL

21

27

6.0

25

23

10.1

1.1

1.0

 

 

26

 

 

32

 

 

 

 

 

 

32

 

 

17

 

 

 

 

Test Item

0.100 µL

20

26

6.0

22

19

2.9

1.0

0.8

 

 

26

 

 

17

 

 

 

 

 

 

16

 

 

21

 

 

 

 

Test Item

0.316 µL

25

22

4.9

17

19

2.1

0.9

0.8

 

 

24

 

 

18

 

 

 

 

 

 

23

 

 

17

 

 

 

 

Test Item

1.0 µL

18

23

5.0

21

19

2.1

0.9

0.8

 

 

28

 

 

18

 

 

 

 

 

 

27

 

 

16

 

 

 

 

Test Item

2.5 µL

25

27

2.0

19

18

1.7

1.1

0.8

 

 

29

 

 

19

 

 

 

 

 

 

10 B

 

 

2 B

 

 

 

 

Test Item

5.0 µL

8 B

7

3.1

8 B

5

3.1

0.3

0.2

 

 

4 B

 

 

4 B

 

 

 

 

 

 

928

 

 

/

 

 

 

 

NaN3

10 µg

933

942

20.1

/

/

/

37.2

/

 

 

965

 

 

/

 

 

 

 

 

 

/

 

 

199

 

 

 

 

2-AA

2.5 µg

/

/

/

202

201

2.1

/

8.6

 

 

/

 

 

203

 

 

 

 

TesterStrain:TA1537                                                                                  Experiment:1

 

Treatment

 

Dose/plate

REVERTANT COLONIES PER PLATE

MUTATION FACTOR

 

Without activation (-S9)

With activation (+S9)

 

Counts

Mean

SD

Counts

Mean

SD

-S9

+S9

 

 

15

 

 

11

 

A. dest.

 

17

15

1.5

15

13

2.1

1.1

0.9

 

 

14

 

 

12

 

 

 

 

 

 

18

 

 

12

 

 

 

 

DMSO

 

12

14

3.2

17

14

2.5

1.0

1.0

 

 

13

 

 

14

 

 

 

 

 

 

13

 

 

12

 

 

 

 

Test Item

0.0100 µL

15

14

1.0

15

14

2.1

1.0

1.0

 

 

14

 

 

16

 

 

 

 

 

 

13

 

 

14

 

 

 

 

Test Item

0.0316 µL

12

14

2.1

13

14

1.5

1.0

1.0

 

 

16

 

 

16

 

 

 

 

 

 

18

 

 

17

 

 

 

 

Test Item

0.100 µL

11

14

3.6

12

13

3.2

1.0

0.9

 

 

13

 

 

11

 

 

 

 

 

 

15

 

 

15

 

 

 

 

Test Item

0.316 µL

14

14

0.6

14

14

1.5

1.0

1.0

 

 

14

 

 

12

 

 

 

 

 

 

13

 

 

16

 

 

 

 

Test Item

1.0 µL

19

14

4.6

14

15

1.0

1.0

1.0

 

 

10

 

 

15

 

 

 

 

 

 

15

 

 

12

 

 

 

 

Test Item

2.5 µL

18

15

3.0

14

14

1.5

1.0

1.0

 

 

12

 

 

15

 

 

 

 

 

 

5 B

 

 

2 B

 

 

 

 

Test Item

5.0 µL

3 B

4

1.0

4 B

4

1.5

0.3

0.3

 

 

4 B

 

 

5 B

 

 

 

 

 

 

111

 

 

/

 

 

 

 

4-NOPD

40 µg

88

95

14.2

/

/

/

6.6

/

 

 

85

 

 

/

 

 

 

 

 

 

/

 

 

312

 

 

 

 

2-AA

2.5 µg

/

/

/

407

361

47.6

/

25.2

 

 

/

 

 

364

 

 

 

 

TesterStrain:TA102                                                                                      Experiment:1

 

Treatment

 

Dose/plate

REVERTANT COLONIES PER PLATE

MUTATIONFACTOR

 

Withoutactivation(-S9)

With activation (+S9)

 

Counts

Mean

SD

Counts

Mean

SD

-S9

+S9

 

 

246

 

 

336

 

A. dest.

 

321

283

37.5

321

337

17.0

1.2

1.1

 

 

282

 

 

355

 

 

 

 

 

 

230

 

 

285

 

 

 

 

DMSO

 

206

230

24.5

302

298

11.9

1.0

1.0

 

 

255

 

 

308

 

 

 

 

 

 

232

 

 

260

 

 

 

 

Test Item

0.0100 µL

297

261

33.0

253

252

8.0

1.1

0.8

 

 

255

 

 

244

 

 

 

 

 

 

362

 

 

309

 

 

 

 

Test Item

0.0316 µL

324

335

23.5

282

293

14.0

1.5

1.0

 

 

319

 

 

289

 

 

 

 

 

 

287

 

 

295

 

 

 

 

Test Item

0.100 µL

282

288

6.0

289

278

25.0

1.2

0.9

 

 

294

 

 

249

 

 

 

 

 

 

334

 

 

230

 

 

 

 

Test Item

0.316 µL

270

304

32.1

264

253

20.2

1.3

0.8

 

 

307

 

 

266

 

 

 

 

 

 

335

 

 

288

 

 

 

 

Test Item

1.0 µL

317

331

12.5

336

339

52.1

1.4

1.1

 

 

341

 

 

392

 

 

 

 

 

 

313

 

 

333

 

 

 

 

Test Item

2.5 µL

289

298

13.3

337

338

5.0

1.3

1.1

 

 

291

 

 

343

 

 

 

 

 

 

77 B

 

 

75 B

 

 

 

 

Test Item

5.0 µL

81 B

77

3.5

64 B

71

6.1

0.3

0.2

 

 

74 B

 

 

74 B

 

 

 

 

 

 

980

 

 

/

 

 

 

 

MMS

1 µL

902

918

55.7

/

/

/

4.0

/

 

 

872

 

 

/

 

 

 

 

 

 

/

 

 

700

 

 

 

 

2-AA

10 µg

/

/

/

729

799

147.1

/

2.7

 

 

/

 

 

968

 

 

 

 

Table 2: Results Experiment II

 

TesterStrain:TA98                                                                                               Experiment:2

 

Treatment

 

Dose/plate

REVERTANT COLONIES PER PLATE

MUTATIONFACTOR

 

Without activation(-S9)

With activation (+S9)

 

Counts

Mean

SD

Counts

Mean

SD

-S9

+S9

 

20

 

 

29

 

A.dest.                                        16

20

4.0

28

30

2.1

0.9

1.4

24

 

 

32

 

 

 

 

22

 

 

22

 

 

 

 

DMSO                                         31

23

7.5

24

22

2.5

1.0

1.0

16

 

 

19

 

 

 

 

26

 

 

30

 

 

 

 

Test Item              0.00316µL           29

29

2.5

29

27

4.9

1.2

1.2

31

 

 

21

 

 

 

 

32

 

 

25

 

 

 

 

TestItem                0.0100µL           25

27

4.4

25

23

3.5

1.2

1.1

24

 

 

19

 

 

 

 

20

 

 

19

 

 

 

 

TestItem                0.0316µL           25

21

4.0

22

25

8.5

0.9

1.2

17

 

 

35

 

 

 

 

21

 

 

26

 

 

 

 

TestItem                  0.100µL           22

23

2.6

19

24

4.7

1.0

1.1

26

 

 

28

 

 

 

 

26

 

 

32

 

 

 

 

TestItem                  0.316µL           18

21

4.6

31

31

1.5

0.9

1.4

18

 

 

29

 

 

 

 

21

 

 

20

 

 

 

 

TestItem                       1.0µL           32

25

6.4

29

25

4.5

1.1

1.1

21

 

 

25

 

 

 

 

15 B

 

 

11 B

 

 

 

 

TestItem                       2.5µL           10B

13

2.5

9 B

11

1.5

0.6

0.5

13 B

 

 

12 B

 

 

 

 

12 B

 

 

5 B

 

 

 

 

TestItem                       5.0µL           10B

10

2.0

4 B

4

0.6

0.4

0.2

8 B

 

 

4 B

 

 

 

 

419

 

 

/

 

 

 

 

4-NOPD                 10µg         356

388

31.5

/

/

/

16.9

/

389

 

 

/

 

 

 

 

/

 

 

1169

 

 

 

 

2-AA                       2.5µg              /

/

/

943

1022

127.4

/

47.2

/

 

 

954

 

 

 

 

TesterStrain:TA100                                                                             Experiment:2

 

Treatment

 

Dose/plate

REVERTANT COLONIES PER PLATE

MUTATION FACTOR

Without activation (-S9)

With activation (+S9)

Counts

Mean

SD

Counts

Mean

SD

-S9

+S9

 

96

 

 

89

 

A. dest.

 

63

87

21.0

92

87

5.7

1.2

1.2

 

 

102

 

 

81

 

 

 

 

 

 

66

 

 

68

 

 

 

 

DMSO

 

78

73

6.1

75

70

4.4

1.0

1.0

 

 

74

 

 

67

 

 

 

 

 

 

87

 

 

71

 

 

 

 

Test Item

0.00316 µL

87

89

2.9

70

72

3.2

1.2

1.0

 

 

92

 

 

76

 

 

 

 

 

 

87

 

 

72

 

 

 

 

Test Item

0.0100 µL

82

84

2.5

75

74

1.5

1.2

1.1

 

 

84

 

 

74

 

 

 

 

 

 

92

 

 

71

 

 

 

 

Test Item

0.0316 µL

109

92

16.5

71

73

2.9

1.3

1.0

 

 

76

 

 

76

 

 

 

 

 

 

67

 

 

68

 

 

 

 

Test Item

0.100 µL

89

82

13.0

93

83

13.2

1.1

1.2

 

 

90

 

 

88

 

 

 

 

 

 

75

 

 

91

 

 

 

 

Test Item

0.316 µL

93

83

9.1

64

76

13.7

1.1

1.1

 

 

82

 

 

73

 

 

 

 

 

 

46

 

 

27

 

 

 

 

Test Item

1.0 µL

36

36

9.5

60

49

19.1

0.5

0.7

 

 

27

 

 

60

 

 

 

 

 

 

10 B

 

 

20 B

 

 

 

 

Test Item

2.5 µL

8 B

9

1.0

18 B

17

4.2

0.1

0.2

 

 

9 B

 

 

12 B

 

 

 

 

 

 

2 B

 

 

5 B

 

 

 

 

Test Item

5.0 µL

2 B

2

0.6

1 B

2

2.3

0.0

0.0

 

 

1 B

 

 

1 B

 

 

 

 

 

 

635

 

 

/

 

 

 

 

NaN3

10 µg

770

678

80.0

/

/

/

9.3

/

 

 

628

 

 

/

 

 

 

 

 

 

/

 

 

979

 

 

 

 

2-AA

2.5 µg

/

/

/

740

936

178.9

/

13.4

 

 

/

 

 

1090

 

 

 

 

TesterStrain:TA1535                                                                             Experiment:2

 

Treatment

 

Dose/plate

REVERTANT COLONIES PER PLATE

MUTATION FACTOR

 

Without activation (-S9)

With activation (+S9)

 

Counts

Mean

SD

Counts

Mean

SD

-S9

+S9

 

 

14

 

 

19

 

A. dest.

 

18

16

2.1

13

16

3.1

1.1

1.0

 

 

17

 

 

15

 

 

 

 

 

 

20

 

 

15

 

 

 

 

DMSO

 

11

15

4.6

21

15

5.5

1.0

1.0

 

 

14

 

 

10

 

 

 

 

 

 

14

 

 

12

 

 

 

 

Test Item

0.00316 µL

17

16

1.5

12

14

2.9

1.0

0.9

 

 

16

 

 

17

 

 

 

 

 

 

15

 

 

14

 

 

 

 

Test Item

0.0100 µL

14

15

0.6

17

16

1.5

1.0

1.0

 

 

15

 

 

16

 

 

 

 

 

 

21

 

 

14

 

 

 

 

Test Item

0.0316 µL

14

16

4.0

21

18

3.5

1.1

1.2

 

 

14

 

 

18

 

 

 

 

 

 

21

 

 

17

 

 

 

 

Test Item

0.100 µL

12

17

4.5

19

17

2.5

1.1

1.1

 

 

17

 

 

14

 

 

 

 

 

 

12

 

 

14

 

 

 

 

Test Item

0.316 µL

16

15

2.6

14

16

3.5

1.0

1.0

 

 

17

 

 

20

 

 

 

 

 

 

14

 

 

13

 

 

 

 

Test Item

1.0 µL

14

15

1.7

18

17

3.2

1.0

1.1

 

 

17

 

 

19

 

 

 

 

 

 

7 B

 

 

6 B

 

 

 

 

Test Item

2.5 µL

8 B

7

0.6

9 B

7

1.5

0.5

0.5

 

 

7 B

 

 

7 B

 

 

 

 

 

 

3 B

 

 

5 B

 

 

 

 

Test Item

5.0 µL

3 B

2

1.2

1 B

3

2.1

0.2

0.2

 

 

1 B

 

 

2 B

 

 

 

 

 

 

210

 

 

/

 

 

 

 

NaN3

10 µg

213

214

5.1

/

/

/

14.3

/

 

 

220

 

 

/

 

 

 

 

 

 

/

 

 

85

 

 

 

 

2-AA

2.5 µg

/

/

/

116

105

17.6

/

6.9

 

 

/

 

 

115

 

 

 

 

TesterStrain:TA1537                                                                              Experiment:2

 

Treatment

 

Dose/plate

REVERTANT COLONIES PER PLATE

MUTATION FACTOR

 

Without activation (-S9)

With activation (+S9)

 

Counts

Mean

SD

Counts

Mean

SD

-S9

+S9

 

 

22

 

 

15

 

A. dest.

 

20

19

3.6

17

17

1.5

0.8

1.0

 

 

15

 

 

18

 

 

 

 

 

 

29

 

 

15

 

 

 

 

DMSO

 

22

23

6.0

16

16

1.0

1.0

1.0

 

 

17

 

 

17

 

 

 

 

 

 

18

 

 

21

 

 

 

 

Test Item

0.00316 µL

17

18

1.0

18

19

1.7

0.8

1.2

 

 

19

 

 

18

 

 

 

 

 

 

20

 

 

17

 

 

 

 

Test Item

0.0100 µL

15

17

2.5

17

18

1.2

0.8

1.1

 

 

17

 

 

19

 

 

 

 

 

 

18

 

 

21

 

 

 

 

Test Item

0.0316 µL

19

19

1.0

17

18

2.6

0.8

1.1

 

 

20

 

 

16

 

 

 

 

 

 

17

 

 

18

 

 

 

 

Test Item

0.100 µL

16

17

1.0

17

18

0.6

0.8

1.1

 

 

18

 

 

18

 

 

 

 

 

 

20

 

 

15

 

 

 

 

Test Item

0.316 µL

21

19

2.1

18

16

2.1

0.9

1.0

 

 

17

 

 

14

 

 

 

 

 

 

16

 

 

15

 

 

 

 

Test Item

1.0 µL

18

18

1.5

16

15

0.6

0.8

1.0

 

 

19

 

 

15

 

 

 

 

 

 

20

 

 

15

 

 

 

 

Test Item

2.5 µL

20

19

1.7

16

16

1.0

0.8

1.0

 

 

17

 

 

17

 

 

 

 

 

 

2 B

 

 

5 B

 

 

 

 

Test Item

5.0 µL

3 B

2

0.6

4 B

4

0.6

0.1

0.3

 

 

2 B

 

 

4 B

 

 

 

 

 

 

178

 

 

/

 

 

 

 

4-NOPD

40 µg

191

185

6.6

/

/

/

8.2

/

 

 

186

 

 

/

 

 

 

 

 

 

/

 

 

157

 

 

 

 

2-AA

2.5 µg

/

/

/

129

153

21.8

/

9.5

 

 

/

 

 

172

 

 

 

 

TesterStrain:TA102                                                                                  Experiment:2

 

Treatment

 

Dose/plate

REVERTANT COLONIES PER PLATE

MUTATIONFACTOR

 

Withoutactivation(-S9)

With activation (+S9)

 

Counts

Mean

SD

Counts

Mean

SD

-S9

+S9

 

 

245

 

 

228

 

A. dest.

 

272

260

13.7

293

264

33.1

1.2

1.1

 

 

263

 

 

271

 

 

 

 

 

 

234

 

 

245

 

 

 

 

DMSO

 

175

216

35.9

242

250

11.9

1.0

1.0

 

 

240

 

 

264

 

 

 

 

 

 

198

 

 

274

 

 

 

 

Test Item

0.00316 µL

222

214

14.2

267

273

5.1

1.0

1.1

 

 

223

 

 

277

 

 

 

 

 

 

258

 

 

280

 

 

 

 

Test Item

0.0100 µL

237

245

11.6

309

310

30.5

1.1

1.2

 

 

239

 

 

341

 

 

 

 

 

 

218

 

 

321

 

 

 

 

Test Item

0.0316 µL

200

202

14.6

322

312

15.9

0.9

1.2

 

 

189

 

 

294

 

 

 

 

 

 

198

 

 

319

 

 

 

 

Test Item

0.100 µL

194

195

2.6

290

314

22.4

0.9

1.3

 

 

193

 

 

334

 

 

 

 

 

 

204

 

 

301

 

 

 

 

Test Item

0.316 µL

202

203

1.2

295

285

22.1

0.9

1.1

 

 

204

 

 

260

 

 

 

 

 

 

226

 

 

279

 

 

 

 

Test Item

1.0 µL

288

264

33.3

304

284

17.6

1.2

1.1

 

 

278

 

 

270

 

 

 

 

 

 

273

 

 

331

 

 

 

 

Test Item

2.5 µL

246

255

15.9

337

328

10.8

1.2

1.3

 

 

245

 

 

316

 

 

 

 

 

 

25 B

 

 

15 B

 

 

 

 

Test Item

5.0 µL

24 B

23

2.6

10 B

12

2.5

0.1

0.0

 

 

20 B

 

 

12 B

 

 

 

 

 

 

1656

 

 

/

 

 

 

 

MMS

1 µL

2276

2100

387.6

/

/

/

9.7

/

 

 

2369

 

 

/

 

 

 

 

 

 

/

 

 

817

 

 

 

 

2-AA

10 µg

/

/

/

914

865

48.5

/

3.5

 

 

/

 

 

865

 

 

 

 

Applicant's summary and conclusion

Conclusions:
Interpretation of results: negative. The substance is considered to be non-mutagenic in this bacterial reverse mutation assay.
Executive summary:

A mutagenicity study with two experiments (used incorporation and pre-incubation method respectively) was performed on the test substance using S. typhimurium in the presence and absence of metabolic activation. Concentrations of 0.0100, 0.0316, 0.100,0.316, 1.0, 2.5 and 5.0 µL/plate in incorporation experiment and the same as pre-incubation experiment but added the 0.00316 µL/plate, each was tested in triplicate. Cytotoxicity was observed from concentrations of 0.316µL/plate (without metabolic activation) and from 2.5 µL/plate (with metabolic activation). All criteria of validity were met. No biologically relevant increases in revertant colony numbers of any of the five tester strains were observed at any concentration level, neither in the presence nor absence of metabolic activation in two experiments. The substance is considered to be non-mutagenic in this bacterial reverse mutation assay.