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Administrative data

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

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.

MEA Polyborate 1:3 is not clastogenic in human lymphocytes under the experimental conditions of a GLP OECD 473 study.

Link to relevant study records

Referenceopen allclose all

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

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



 

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.
Endpoint:
in vitro cytogenicity / chromosome aberration study in mammalian cells
Remarks:
Type of genotoxicity: chromosome aberration
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2010-06-15 to 2010-06-23
Reliability:
1 (reliable without restriction)
Qualifier:
according to
Guideline:
OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of assay:
in vitro mammalian chromosome aberration test
Species / strain / cell type:
lymphocytes:
Additional strain / cell type characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
Rat liver microsomal enzymes were routinely prepared from adult male Wister rats (6), which wre obtained from Charles Reiver (Sulzfeld, Germany)
Test concentrations with justification for top dose:
First assay
With and without S9-mix: 10, 33, 100, 333, 1000, 3330 and 5000 µg/ml culture medium ( 3h exposure time, 24 h fixation time)
Scoring dose: With and without S9-mix : 1000, 3330 and 5000 µg/ml culture medium ( 3h exposure time, 24 h fixation time)
Second assay
Without S9-mix: 100, 200, 300, 400, 500, 600 and 700 µg/ml culture medium ( 24 and 48h exposure time, 24 and 48h fixation time)
With S9-mix : 1000, 3330 and 5000 µg/ml culture medium ( 3h exposure time, 48 h fixation time)
Scoring dose:
Without S9-mix : 100, 400 and 500 µg/ml culture medium ( 24 and 48h exposure time, 24 and 48h fixation time)
With S9-mix : 1000, 3330 and 5000 µg/ml culture medium ( 3h exposure time, 48 h fixation time)
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: [RPMI 1640 medium]
- Justification for choice of solvent/vehicle:none given
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
cyclophosphamide
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
mitomycin C
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium
DURATION
- Preincubation period:
- Exposure duration:3 hour
- Expression time (cells in growth medium):
- Selection time (if incubation with a selection agent):
- Fixation time (start of exposure up to fixation or harvest of cells): 24 hour

SELECTION AGENT (mutation assays):
SPINDLE INHIBITOR (cytogenetic assays):colchcine (0.5
STAIN (for cytogenetic assays):5% (v/v) Glemsa (Merck) solution in tap water

NUMBER OF REPLICATIONS:2

NUMBER OF CELLS EVALUATED:

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index

OTHER EXAMINATIONS:
- Determination of polyploidy:carried out
- Determination of endoreplication:carried out
- Other:

OTHER:
Evaluation criteria:
A substance was considered positive (clastogenic) in the chromosome aberration test if:
a) It induced a dose-related statistically significant (Chi-square test, one sided , p<0.05) increase in the number of cells with chromosome aberrations.
b) A statistically significant and biologically relevant increase in the frequencies of the number of cells with chromosome aberrations was observed in the absence of a clear dose-response relationship.
Statistics:
The parameter used was the increase in the number of cells with chromosome aberrations. The statisitcal significance was given by Chi-square test, one sided , p<0.05.
Species / strain:
lymphocytes: human
Metabolic activation:
with and without
Genotoxicity:
not determined
Cytotoxicity / choice of top concentrations:
cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH:
- Effects of osmolality:
- Evaporation from medium:
- Water solubility:soluble
- Precipitation: no precipitation
- Other confounding effects:

RANGE-FINDING/SCREENING STUDIES:carried out

COMPARISON WITH HISTORICAL CONTROL DATA: within historical control data range

ADDITIONAL INFORMATION ON CYTOTOXICITY:
Remarks on result:
other: strain/cell type: human lymphocytes
Remarks:
Migrated from field 'Test system'.
Conclusions:
MEA Polyborate 1:3 is not clastogenic in human lymphocytes under the experimental conditions of a GLP OECD 473 study.
Executive summary:

1.     

Evaluation of the ability of MEA Polyborate 1:3 to induce chromosome aberrations in cultured peripheral human lymphocytes (with repeat experiment).

 

This report describes the effect of MEA Polyborate 1:3 on the number of chromosome aberrations in cultured peripheral human lymphocytes in the presence and absence of a metabolic activation system (phenobarbital and ß-naphthoflavone induced rat liver S9-mix). The possible clastogenicity of MEA Polyborate 1:3 was tested in two independent experiments.

 

The study procedures described in this report were based on the most recent OECD and EC guidelines.

 

Batch EU-SMG 01696 of MEA Polyborate 1:3 was a clear colourless liquid. MEA Polyborate 1:3 was dissolved in RPMI 1640 medium.

 

In the first cytogenetic assay, MEA Polyborate 1:3 was tested up to 5000 µg/ml for a 3 h exposure time with a 24 h fixation time in the absence and presence of 1.8% (v/v) S9-fraction. This is the highest concentration recommended in the guidelines.

 

In the second cytogenetic assay, MEA Polyborate 1:3 was tested up to 500 µg/ml for a 24 h and 48 h continuous exposure time with a 24 h and 48 h fixation time in the absence of S9-mix. Appropriate toxicity was reached at this dose level. In the presence of S9-mix MEA Polyborate 1:3 was tested up to 5000 µg/ml for a 3 h exposure time with a 48 h fixation time.

 

The number of cells with chromosome aberrations found in the solvent control cultures was within the laboratory historical control data range. Positive control chemicals, mitomycin C and cyclophosphamide, both produced a statistically significant increase in the incidence of cells with chromosome aberrations, indicating that the test conditions were adequate and that the metabolic activation system (S9-mix) functioned properly.

 

MEA Polyborate 1:3 did not induce a statistically significant or biologically relevant increase in the number of cells with chromosome aberrations in the absence and presence of S9-mix, in either of the two independently repeated experiments.

 

No biologically relevant effects of MEA Polyborate 1:3 on the number of polyploid cells and cells with endoreduplicated chromosomes were observed both in the absence and presence of

S9-mix. Therefore it can be concluded that MEA Polyborate 1:3 does not disturb mitotic processes and cell cycle progression and does not induce numerical chromosome aberrations under the experimental conditions described in this report.

 

Finally, it is concluded that this test is valid and that MEA Polyborate 1:3 is not clastogenic in human lymphocytes under the experimental conditions described in this report.


Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Justification for classification or non-classification

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.

MEA Polyborate 1:3 is not clastogenic in human lymphocytes under the experimental conditions of a GLP OECD 473 study.

Therefore the substance is not classified as mutagenic under GHS.