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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
Study period:
26 March 2002 to 10 April 2002
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study was performed in compliance with GLP and in accordance with the standardised guidelines OECD 471 and US EPA OPPTS 870.5100. The study was performed to a high standard sufficient to assess the quality of the presented results.

Data source

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

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: crystalline
Details on test material:
- Name of test material (as cited in study report): dioctyloxostannane (dioctyltin oxide)
- Physical state: Powder, white crystalline
- Storage condition of test material: -20 °C in the dark

Method

Target gene:
Histidine requirement in the Salmonella typhimurium strains
Tryptophan requirement in the Escherichia coli strain.
Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Details on mammalian cell type (if applicable):
- Type and identity of media: Stored as nutrient broth cultures with 7.4 % DMSO at <60 °C.
- Properly maintained: yes
- Periodically "cleansed" against high spontaneous background: yes
Species / strain / cell type:
E. coli WP2 uvr A
Details on mammalian cell type (if applicable):
- Type and identity of media: Stored as nutrient broth cultures with 7.4 % DMSO at <60 °C.
- Properly maintained: yes
- Periodically "cleansed" against high spontaneous background: yes
Metabolic activation:
with and without
Metabolic activation system:
S9-mix
Test concentrations with justification for top dose:
Range finding test: 0, 0.3, 0.8, 2.3, 7, 21, 62, 185, 556, 1667 and 5000 µg/plate.
Definitive test: 0, 62, 185, 556, 1667 and 5000 µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: Methanol
Controls
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
9-aminoacridine
2-nitrofluorene
sodium azide
benzo(a)pyrene
other: N-ethyl-N-nitrosourea and 2-aminoanthracene
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium; in agar (plate incorporation)
2 mL molten top agar (0.6 % agar, 0.5 % NaCl and 0.05 mM L-histidine.HCl/0.05 mM biotin for the S. typhimurium stains, and supplemented with 0.05 mM tryptophane for the E.coli WP2 uvrA strain), maintained at 46 °C. 0.1 mL of culture of the appropriate strain, 0.1 mL of the appropriate test substance solution, or the solvent or positive control substance and 0.5 mL S9-mix for the plates with metabolic activation or 0.5 mL sodium phosphate 100 mM (pH 7.4) for without metabolic activation. The ingredients were thoroughly mixed and the mix was immediately poured onto minimal glucose agar plates (1.5 % agar in Vogel and Bonner medium E with 2 % glucose).

DURATION
- Exposure duration: 48-72 hours at 37 °C

NUMBER OF REPLICATIONS: The tests were performed in triplicate

DETERMINATION OF CYTOTOXICITY
- Method: Cytotoxicity was assessed as a reduction in the number of revertant colonies and/or a clearing of the background lawn of bacterial growth.
Evaluation criteria:
The mutagenicity study was considered valid if the mean colony counts of the control values of the strains were within acceptable ranges, if the results of the positive controls met the criteria for a positive response, and if no more than 5 % of the plates were lost through contamination or other unforseen events.
A test substance was considered to be positive if the increase in the mean number of revertant colonies on the test plates was concentration-related or if a reproducible two-fold or more increase was observed compared to that of negative control plates.
A test substance was considered to be negative in the bacterial gene mutation test if it produced neither a dose-related increase in the mean number of revertant colonies nor a reproducible positive response at any of the test points.
Statistics:
No statistical analysis was performed.

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:
no cytotoxicity, but tested up to precipitating concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
Positive controls validity:
valid
Species / strain:
E. coli WP2 uvr A
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity, but tested up to precipitating concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
Positive controls validity:
not valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: The test substance was found to precipitate at all concentrations

RANGE-FINDING/SCREENING STUDIES: A dose range finding test was performed with TA 98 in both the absence and presence of S9-mix with ten different concentrations of the test substance, ranging from 0.3-5000 µg/plate. Dioctyloxostannane (dioctyltin oxide) was not toxic at any concentration both in the absence and presence of S9-mix.

COMPARISON WITH HISTORICAL CONTROL DATA:

ADDITIONAL INFORMATION ON CYTOTOXICITY: A slightly more dense background lawn of the bacterial growth was observed at the dose levels at which the slight increases were observed; however, these occured alongside precipitation of the test substance.
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

Table 2: Results of range finding test

Dose µg/plate

TA 98

Without S9-mix

With S9-mix

0

38

52

36

46

29

48

Mean

34

49

SD

5

3

0.3

46

57

48

61

40

46

Mean

45

55

SD

4

8

0.8

46

69

44

57

26

60

Mean

39

62

SD

11

6

2.3

52

52

42

55

34

57

Mean

43

55

SD

9

3

7

27

59

42

59

46

60

Mean

38

59

SD

10

1

21

31

53

27

42

36

47

Mean

31

47

SD

5

6

62

42

56

33

47

40

60

Mean

38

54

SD

5

7

185

45

58

36

63

39

46

Mean

40

56

SD

5

9

556

34

38

39

45

35

40

Mean

36

41

SD

3

4

1667

45

59

44

57

39

68

Mean

43

61

SD

3

6

5000

37

57

41

52

38

50

Mean

39

53

SD

2

4

Positive control

1170

1090

1225

1051

1201

1074

Mean

1199

1072

SD

28

20

*The positive controls employed are detail in table 1.

Table 2: Results of the definitive test

Dose µg/plate

TA 1535

TA 1537

TA 98

TA 100

E. coli

Without S9-mix

With S9-mix

Without S9-mix

With S9-mix

Without S9-mix

With S9-mix

Without S9-mix

With S9-mix

Without S9-mix

With S9-mix

0

15

17

8

22

29

77

165

156

**

31

21

28

13

6

32

51

186

189

23

26

16

15

13

15

27

36

163

175

37

25

Mean

17

20

11

14

29

55

171

173

30

27

SD

3

7

3

8

3

21

13

17

10

3

62

15

20

5

15

16

60

151

170

40

46

18

24

15

14

27

40

148

166

46

39

20

22

12

8

24

51

173

198

40

42

Mean

18P

22

11

12

22

50

153

178

42*

42

SD

3

2

5

4

6

10

14

17

3

4

185

8

23

11

16

25

57

159

181

35

41

23

24

6

4

28

55

149

161

36

34

20

13

4

9

28

56

211

182

31

47

Mean

17P

20

7

10

27

56

173

175

34*

41

SD

8

6

4

6

2

1

33

12

3

7

556

17

20

9

9

31

38

183

144

49

37

30

23

9

8

26

53

168

155

36

33

14

25

8

11

29

35

170

159

30

49

Mean

20P

23

9

9

29

42

174

153

38*

40

SD

9

3

1

2

3

10

8

8

10

8

1667

20

11

5

6

48

31

146

162

33

50

28

20

18

8

35

37

146

152

41

38

22

31

18

12

36

85

150

170

67

27

Mean

23P

21

14

9

40

51

147

161

47*

38

SD

4

10

8

3

7

30

2

9

18

12

5000

31

20

8

11

31

40

153

166

46

49

34

19

7

14

51

42

137

161

46

39

30

28

11

15

26

48

131

161

42

41

Mean

32*

22

9

13

36

43

140

163

45*

43

SD

2

5

2

2

13

4

11

3

2

5

Positive control

442

365

1053

211

1105

634

744

1245

287

698

444

345

863

212

1185

738

704

940

221

677

427

374

913

216

1186

555

708

1012

242

715

Mean

438

361

943

213

1159

642

719

1066

250

697

SD

9

15

98

3

46

92

22

159

34

19

P: Precipitation of test substance

* : Precipitation of test substance and slightly more dense background lawn of bacterial growth than in concomitant control plates.

**: Not counted; no bacteria on the agar plate

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative

Under the conditions of the test, the results obtained with the test substance in Salmonella typhimurium strains TA 1535, TA 1537, TA 98 and TA 100 and in the Eschericia coli strain WP2 uvrA, in both the absence and presence of metabolic activation (S9-mix), the test substance was not mutagenic.
Executive summary:

The mutagenicity of the test substance was assessed in a bacterial reverse mutation assay (Ames Test) in accordance with the OECD guideline 471 and EPA OPPTS 870.5100 and to GLP. The cultures were exposed to 0, 62, 185, 556, 1667 and 5000 µg/plate. Precipitation of the test substance was observed at a number of the concentrations tested. In some of the plates a slight dose-dependant increase in the number of revertants were noticed, however this was accompanied by an increased background lawn and precipitation. This was therefore not attributed to the mutagenic potential of the test substance. Under the conditions of the test, the results obtained with the test substance in Salmonella typhimurium strains TA 1535, TA 1537, TA 98 and TA 100 and in the Eschericia coli strain WP2 uvrA, in both the absence and presence of metabolic activation (S9-mix), the test substance was not mutagenic.