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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:
19 Jun 2017 - 17 Jul 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report Date:
2017

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Test material form:
liquid
Remarks:
clear colourless
Details on test material:
UVCB
At room temperature protected from light
31 March 2018 (expiry date)

Method

Species / strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
Metabolic activation:
with and without
Metabolic activation system:
Rat liver microsomal enzymes (S9 homogenate), prepared from male Sprague Dawley rats that had been injected intraperitoneally with Aroclor 1254 (500 mg/kg body weight).
Test concentrations with justification for top dose:
Dose-range Finding Test
Selection of an adequate range of doses was based on a dose-range finding test with the strains TA100 and the WP2uvrA, both with and without S9-mix. Eight concentrations, 1.7, 5.4, 17, 52, 164, 512, 1600 and 5000 µg/plate were tested in triplicate.

The highest concentration of the test item used in the subsequent mutation assays was 5000 μg/plate. At least five different doses (increasing with approximately half-log steps) of the test item were tested in triplicate in each strain in the absence and presence of S9-mix.
Vehicle / solvent:
The vehicle of the test item was Milli-Q water (Millipore Corp., Bedford, MA., USA).
Controlsopen allclose all
Untreated negative controls:
yes
Remarks:
Milli-Q water
Positive controls:
yes
Positive control substance:
4-nitroquinoline-N-oxide
2-nitrofluorene
sodium azide
methylmethanesulfonate
other: ICR-191 (Sigma), 2-aminoanthracene (2AA)
Details on test system and experimental conditions:
Salmonella typhimurium bacteria and Escherichia coli bacteria
Rationale for test conditions:
Recommended test system in international guidelines (e.g. OECD, EC).

Results and discussion

Test resultsopen allclose all
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 1537
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 98
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium 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
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 nor precipitates, but tested up to recommended limit concentrations
Untreated negative controls validity:
valid
Positive controls validity:
valid

Any other information on results incl. tables

Dose-Range Finding Test: Mutagenic Response of MEA Polyborate 1:3 in the Salmonella typhimurium Reverse Mutation Assay and in the Escherichia coli Reverse Mutation Assay

Direct plate assay


Dose

(µg/plate)


Mean number of revertant colonies/3 replicate plates (
±S.D.) with one Salmonella typhimuriumand one Escherichia colistrain.

 


TA100


WP2uvrA

 



 

                                                     Without S9-mix

 

Positive control

1033

±

46

 

1120

±

71

Solvent control

129

±

3

 

36

±

7

1.7

113

±

16

 

27

±

6

5.4

144

±

10

 

30

±

8

17

150

±

10

 

29

±

10

52

141

±

18

 

33

±

9

164

152

±

10

 

29

±

6

512

132

±

16

 

31

±

8

1600

162

±

3

 

29

±

11

5000

192

±

7

n NP

39

±

3 

n NP

 

                                                      With S9-mix

 

Positive control

975

±

16

 

378

±

30

Solvent control

136

±

24

 

40

±

14

1.7

167

±

9

 

44

±

4

5.4

165

±

9

 

37

±

3

17

126

±

5

 

32

±

7

52

149

±

15

 

40

±

4

164

141

±

22

 

35

±

7

512

153

±

7

 

38

±

8

1600

158

±

16

 

46

±

10

5000

197

±

11

n NP

58

±

7n NP

 

NP

No precipitate

n

Normal bacterial background lawn


Experiment 1: Mutagenic Response of MEA Polyborate 1:3 in the Salmonella typhimurium Reverse Mutation Assay


Direct plate assay


Dose

(µg/plate)


Mean number of revertant colonies/3 replicate plates (
±S.D.) with
different strains of Salmonella typhimurium.

 


TA1535


TA1537

 


TA98

 

                                                      Without S9-mix

 

Positive control

927

±

65

 

350

±

59

 

1026

±

82

 

Solvent control

16

±

2

 

9

±

2

 

15

±

4

 

52

13

±

1

 

9

±

2

 

13

±

2

 

164

8

±

3

 

4

±

1

 

12

±

5

 

512

16

±

6

 

5

±

5

 

18

±

4

 

1600

15

±

4

 

5

±

3

 

17

±

2

 

5000

14

±

3

n NP

8

±

4

n NP

17

±

1

n NP

 

 

 

 

 

 

 

 

 

 

 

 

 

                                                       With S9-mix

 

Positive control

329

±

23

 

839

±

48

 

675

±

86

 

Solvent control

10

±

3

 

8

±

3

 

28

±

2

 

52

12

±

4

 

8

±

4

 

42

±

21

 

164

18

±

3

 

6

±

7

 

16

±

5

 

512

15

±

1

 

6

±

3

 

19

±

8

 

1600

15

±

3

 

6

±

6

 

13

±

4

 

5000

18

±

4

n NP

9

±

1

n NP

17

±

8

n NP

 

 

 

 

 

 

 

 

 

 

 

 

 

 

NP

No precipitate

n

Normal bacterial background lawn

 

Experiment 2: Mutagenic Response of MEA Polyborate 1:3 in the Salmonella typhimurium Reverse Mutation Assay and in the Escherichia coli Reverse Mutation Assay

Pre-incubation assay


Dose

(µg/plate)


Mean number of revertant colonies/3 replicate plates (
±S.D.) with
different strains of Salmonella typhimurium and one Escherichia coli strain.

 


TA1535


TA1537

 


TA98


TA100


WP2uvrA

                                                            Without S9-mix

 

Positive control

962

±

34

 

177

±

20

 

1282

±

46

 

800

±

131

 

125

±

19

 

Solvent control

12

±

2

 

5

±

1

 

13

±

4

 

132

±

10

 

30

±

1

 

52

11

±

4

 

7

±

2

 

15

±

7

 

136

±

17

 

29

±

8

 

164

11

±

6

 

6

±

3

 

14

±

3

 

122

±

16

 

25

±

7

 

512

12

±

3

 

6

±

2

 

17

±

2

 

131

±

20

 

25

±

7

 

1600

8

±

2

 

8

±

3

 

15

±

8

 

138

±

8

 

33

±

15

 

5000

8

±

3

n NP

6

±

3

n NP

12

±

7

n NP

209

±

6

n NP

43

±

9

n NP

 

With S9-mix

 

Positive control

262

±

5

 

221

±

18

 

619

±

21

 

1219

±

115

 

403

±

14

 

Solvent control

12

±

6

 

8

±

3

 

19

±

8

 

141

±

8

 

46

±

3

 

52

16

±

2

 

7

±

2

 

25

±

4

 

117

±

17

 

46

±

10

 

164

13

±

7

 

5

±

2

 

22

±

3

 

133

±

19

 

54

±

9

 

512

14

±

2

 

7

±

2

 

27

±

8

 

132

±

7

 

55

±

9

 

1600

13

±

4

 

5

±

5

 

21

±

6

 

158

±

7

 

54

±

2

 

5000

13

±

6

n NP

7

±

3

n NP

25

±

2

n NP

204

±

17

n NP

65

±

14

n NP

 

 

NP

No precipitate

n

Normal bacterial background lawn



 

Applicant's summary and conclusion

Conclusions:
Based on the results of a GLP OECD 471 study it is concluded that MEA Polyborate 1:3 is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.