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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-compliant guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2009
Report date:
2009

Materials and methods

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

Test material

Constituent 1
Chemical structure
Reference substance name:
methyl 5-(dimethylamino)-2-methyl-5-oxopentanoate
EC Number:
700-204-6
Cas Number:
1174627-68-9
Molecular formula:
C9H17NO3
IUPAC Name:
methyl 5-(dimethylamino)-2-methyl-5-oxopentanoate
Test material form:
other: liquid
Details on test material:
- Name of test material (as cited in study report): DV-SOLV 1059

Method

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
Metabolic activation:
with and without
Metabolic activation system:
Phenobarbital/Beta-Napthoflavone induced rat liver S9
Test concentrations with justification for top dose:
Experiments 1 & 2: 5, 15.81, 50, 158.1, 500, 1581 and 5000; µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO; water
- Justification for choice of solvent/vehicle: Solubility of test material and positive controls
Controlsopen allclose all
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
Salmonella TA 100, TA 1535: without activation
Positive control substance:
9-aminoacridine
Remarks:
Salmonella TA 1537: without activation
Positive control substance:
other: 4-nitro-1,2-phenylene-diamine (NPD)
Remarks:
Salmonella TA 98: without activation
Positive control substance:
methylmethanesulfonate
Remarks:
E coli: without activation
Positive control substance:
other: 2-aminoanthracene
Remarks:
Salmonella all strains, E coli: with activation
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (experiment 1: plate incorporation); experiment 2: preincubation

DURATION - Preincubation period: 48 hours
Evaluation criteria:
Criteria for positive result:
- a dose-related increase in the number of revertants and/or
- a reproducible biologically relevant positive result for at least one of the dose groups in at least one strain with or without metabolic activation
An increase is considered biologically relevant if the number of reversions is twice that of controls for S. TA 100, or three times higher for other tested strains.

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
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
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
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

There were no toxic effects. Plates incubated with the test item showed normal background growth up to 5000µg/plate with and without S9 mix in both experiments.
No substantial increase in revertant colony numbers of any of the 5 tester strains was observed following treatment with
DV-SOLV 1059at any dose level, neither in the presence nor absence of metabolic activation (S9 mix). There was also no tendency of higher mutation rates with increasing concentrations in the range below the generally acknowledged border of biological relevance.

Appropriate reference mutagens were used as positive controls and these showed a distinct increase of induced revertant colonies.

Results of Gene Mutation Assay: Experiment 1 (Plate incorporation):

Metabolic

Activation

Test

Group

Dose Level

(µg/plate)

 

Revertant Colony Counts (Mean ±SD)

 

 

 

 

 

 

 

 

 

 

 

 

 

TA 1535

TA 1537

TA 98

TA 100

WP2 uvrA

 

 

 

 

 

 

 

 

 

Without Activation

Untreated

 

 

7 ± 3

3 ± 1

21 ± 6

90 ± 23

31 ± 2

DMSO

 

 

11 ± 5

6 ± 2

20 ± 6

90 ± 20

26 ± 4

Distilled water

 

 

12 ± 2

6 ± 3

19 ± 1

86 ± 24

32 ± 6

DV-SOLV 1059

5 µg

 

10 ± 1

4 ± 2

17 ± 6

93 ± 14

27 ± 6

 

15.81 µg

 

7 ± 3

8 ± 4

28 ± 6

87 ± 8

30 ± 10

 

50 µg

 

11 ± 2

5 ± 1

18 ± 1

104 ± 13

33 ± 10

 

158.1 µg

 

10 ± 3

6 ± 2

15 ± 2

106 ± 9

35 ± 3

 

500 µg

 

8 ± 1

8 ± 3

21 ± 7

99 ± 8

36 ± 3

 

1581 µg

 

8 ± 1

7 ± 3

19 ± 5

93 ± 25

29 ± 4

 

5000 µg

 

8 ± 3

6 ± 2

16 ± 5

101 ± 10

34 ± 4

NaAZ

2 µg

 

635 ± 129

 

 

990 ± 56

 

2AA

2 µg

 

 

 

 

 

 

9AA

50 µg

 

 

287 ± 23

 

 

 

NPD

4 µg

 

 

 

292 ± 03

 

 

MMS

2 µL

 

 

 

 

 

651 ± 57

 

 

 

 

 

 

 

 

 

With Activation

Untreated

 

 

8 ± 3

5 ± 0

29 ± 5

88 ± 5

42 ± 6

DMSO

 

 

8 ± 1

8 ± 1

20 ± 6

95 ± 6

37 ± 7

Distilled water

 

 

11 ± 5

9 ± 1

19 ± 1

86 ± 7

44 ± 16

DV-SOLV 1059

5 µg

 

10 ± 3

8 ± 2

26 ± 5

84 ± 7

37 ± 12

 

15.81 µg

 

8 ± 6

6 ± 3

24 ± 2

86 ± 8

42 ± 3

 

50 µg

 

11 ± 2

9 ± 1

24 ± 7

86 ± 5

32 ± 7

 

158.1 µg

 

11 ± 2

5 ± 2

19 ± 3

78 ± 1

41 ± 5

 

500 µg

 

12 ± 2

7 ± 2

26 ± 11

84 ± 5

44 ± 2

 

1581 µg

 

8 ± 3

9 ± 2

26 ± 5

85 ± 11

41 ± 8

 

5000 µg

 

9 ± 5

10 ± 2

27 ± 12

72 ± 15

38 ± 8

2-AA

2 µg

 

223 ± 10

153 ± 19

1889 ± 157

1937 ± 107

 

2-AA

50 µg

 

 

 

 

 

183 ± 5

 

 

 

 

 

 

 

 

 

 

Key to Positive Controls

 

NaAZ

2-AA

9AA

NPD

MMS

sodium azide

2-aminoanthracene

9-aminoacridine

4-nitro-1,2-phenylene-diamine

methyl methane sulfonate

 

 

 

Results of Gene Mutation Assay: Experiment 2 (Pre-incubation):

Metabolic

Activation

Test

Group

Dose Level

(µg/plate)

 

Revertant Colony Counts (Mean ±SD)

 

 

 

 

 

 

 

 

 

 

 

 

 

TA 1535

TA 1537

TA 98

TA 100

WP2 uvrA

 

 

 

 

 

 

 

 

 

Without Activation

Untreated

 

 

14 ± 5

9 ± 2

19 ± 5

98 ± 3

40 ± 4

DMSO

 

 

10 ± 4

8 ± 3

22 ± 2

100 ± 11

30 ± 0

Distilled water

 

 

16 ± 4

6 ± 2

19 ± 1

105 ± 2

38 ± 5

DV-SOLV 1059

5 µg

 

16 ± 1

6 ± 1

16 ± 6

107 ± 8

37 ± 9

 

15.81 µg

 

16 ± 1

5 ± 1

20 ± 4

103 ± 6

38 ± 9

 

50 µg

 

14 ± 3

5 ± 1

16 ± 4

102 ± 10

39 ± 7

 

158.1 µg

 

17 ± 1

9 ± 5

12 ± 6

98 ± 2

46 ± 4

 

500 µg

 

15 ± 4

6 ± 3

13 ± 1

101 ± 16

36 ± 4

 

1581 µg

 

11 ± 1

6 ± 3

13 ± 1

96 ± 21

33 ± 6

 

5000 µg

 

13 ± 4

4 ± 3

14 ± 3

92 ± 5

28 ± 5

NaAZ

2 µg

 

801 ± 39

 

 

1087 ± 133

 

2AA

2 µg

 

 

 

 

 

 

9AA

50 µg

 

 

220 ± 30

 

 

 

NPD

4 µg

 

 

 

230 ± 35

 

 

MMS

2 µL

 

 

 

 

 

803 ± 21

 

 

 

 

 

 

 

 

 

With Activation

Untreated

 

 

18 ± 6

2 ± 0

30 ± 3

115 ± 20

43 ± 8

DMSO

 

 

16 ± 4

9 ± 1

27 ± 6

102 ± 3

43 ± 8

Distilled water

 

 

14 ± 3

16 ± 6

26 ± 4

113 ± 7

49 ± 4

DV-SOLV 1059

5 µg

 

17 ± 2

8 ± 4

19 ± 6

112 ± 6

58 ± 9

 

15.81 µg

 

16 ± 4

12 ± 5

25 ± 3

105 ± 4

49 ± 3

 

50 µg

 

13 ± 1

9 ± 3

24 ± 3

100 ± 9

54 ± 3

 

158.1 µg

 

14 ± 1

11 ± 3

16 ± 2

98 ± 1

53 ± 10

 

500 µg

 

16 ± 2

10 ± 5

18 ± 3

84 ± 6

63 ± 2

 

1581 µg

 

17 ± 1

7 ± 2

22 ± 4

91 ± 5

58 ± 2

 

5000 µg

 

15 ± 4

8 ± 3

24 ± 3

85 ± 12

47 ± 5

2-AA

2 µg

 

156 ± 13

148 ± 7

1677 ± 198

1887 ± 201

 

2-AA

50 µg

 

 

 

 

 

188 ± 16

 

 

 

 

 

 

 

 

 

 

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative

The test item did not induce gene mutations by base pair changes or frameshifts in the genome of the strains used.
Therefore, DV-SOLV 1059 is considered to be non-mutagenic in this Salmonella typhimurium and Escherichia coli reverse mutation assay.
Executive summary:

This study was performed to investigate the potential of DV-SOLV 1059 to induce gene mutations in the plate incorporation test (experiment I) and the pre-incubation test (experiment II) using Salmonella typhimurium strains TA 1535, TA 1537, TA 98, and TA 100, and Escherichia coli strain WP2 uvrA.

The assay was performed in two independent experiments both with and without liver microsomal activation. Each concentration, including the controls, was tested in triplicate. The test item was tested at the following concentrations of the active ingredient:

Pre-Experiment/Experiment I/Experiment II: 5, 15.81, 50, 158.1, 500, 1581 and 5000 µg/plate

The plates incubated with the test item showed normal background growth up to 5000 µg/plate with and without metabolic activation in both independent experiments.

No toxic effects, evident as a reduction in the number of revertants (below the induction factor of 0.5), occurred in the test groups with or without metabolic activation.

No substantial increase in revertant colony numbers of any of the five tester strains was observed following treatment with DV-SOLV 1059 at any dose level, either in the presence or absence of metabolic activation (S9 mix). There was also no tendency of higher mutation rates with increasing concentrations in the range below the generally acknowledged border of biological relevance.

Appropriate reference mutagens were used as positive controls and showed a distinct increase of induced revertant colonies.

In conclusion, in the mutagenicity test described and under the experimental conditions reported, the test item did not induce gene mutations by base pair changes or frameshifts in the genome of the strains used.

Therefore, DV-SOLV 1059 is considered to be non-mutagenic in this Salmonella typhimurium and Escherichia coli reverse mutation assay.