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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:
1992-08-19 to 1992-08-28
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Well performed guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1993
Report date:
1993

Materials and methods

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

Test material

Constituent 1
Chemical structure
Reference substance name:
Sodium 2-hydroxyethanesulphonate
EC Number:
216-343-6
EC Name:
Sodium 2-hydroxyethanesulphonate
Cas Number:
1562-00-1
Molecular formula:
C2H6O4S.Na
IUPAC Name:
sodium 2-hydroxyethane-1-sulfonate
Details on test material:
- Name of test material (as cited in study report): Ethansalz 97/100 %
- Molecular weight: 148.05 g/mol
- Chemical nomenclature: Ethansulfonacid, 2-Hydroxy-, Monosodiumsalt
- Substance type: solid
- Physical state: white powder
- Analytical purity: 99.6%
- Purity test date: 1992-01-21
- Lot/batch No.: 95
- Expiration date of the lot/batch: 1994-01-21
- Stability under test conditions: stable
- Storage condition of test material: dark, at room temperature
- melting point: appr. 230°C

Method

Species / strainopen allclose all
Species / strain / cell type:
other: Salmonella typhimurium TA 98, TA 100, TA 1535, TA 1537, TA 1538
Additional strain / cell type characteristics:
not specified
Species / strain / cell type:
E. coli WP2 uvr A
Additional strain / cell type characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
S9 mix
Test concentrations with justification for top dose:
Test item:
- Toxicity test: 4 - 10000 µg/plate in aqua bidest.
- Mutagenicity test: 4-5000 µg/plate in aqua bidest.

Positive controls:
- sodium azide: 1 µg/plate
- 9-Aminoacridine: 50 µg/plate
- 2-Nitrofluorene: 2.5 µg/plate
- N-Methyl-N-nitro-N-nitrosoguanidine: 2.5 µg/plate
- 2-Aminoanthracen: 0.5-10 µg/plate
Vehicle / solvent:
Aqua bidest.
Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: for details on controls please see below
Details on test system and experimental conditions:
The assay was performed in two independent experiments (3 plates/dose, with and without metabolic activation):

1. Experiment: 0-10000 µg test item/plate
2. Experiment: 0-5000 µg test item/plate

DURATION:
After mixing the overnight culture and test compound the mixture is poured into a petridish with minimal agar and incubated for approx. 48h at 37°C in the dark.

The first experiment was performed with all tester strains using three plates per dose to get information on mutagenicity and cytotoxicity for calculation of an appropriate dose range. A reduced rate of spontanously occuring colonies as well as visible thinning of the bacterial lawn were used as indicator for toxicity.

POSITIVE CONTROL SUBSTANCES:
without metabolic activation: Sodium azide (TA 100, TA 1535), 9-Aminoacridine (TA 1537), 2-Nitrofluorene (TA 98, TA 1538), N-Methyl-N-nitro-N-nitrosoguanidine (WP2uvrA)
with metabolic activiation: 2-Aminoanthracene (TA 98, TA 100, TA 1535, TA 1537, TA 1538, WP2uvrA)
Evaluation criteria:
A test article is classified mutagenic if either of the following conditions under a) and b) is achieved:
a) a test article produced at least a 2-fold increase in the mean number of revertants per plate of at least one of the tester strains over the mean number of revertants per plate of the appropriate vehicle control at complete bacterial background lawn.
b) a test article induces a dose-related increase in the mean number of revertants per plate of at least one of the tester strains over the mean numberof revertants per plate of the appropriate verhicle control in at least two to three concentrations of the test article at complete bacterial background lawn.
The tests must be reproducible.

Results and discussion

Test resultsopen allclose all
Species / strain:
other: Salmonella typhimurium TA 98, TA 100, TA 1535, TA 1537, TA 1538
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
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 nor precipitates, but tested up to recommended limit concentrations
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

Ames Test: Ethansalz 97/100 (Study # 92.0810)

 

 

Table: Summary of mean number of revertants (mean of 3 plates) in Salmonella typhimurium andE. coli strains with and without metabolic activation

Experiment I

Concentration

TA 98

TA 100

TA1535

TA 1537

[µg/plate]

- S9 mix

+ S9 mix

- S9 mix

+ S9 mix

- S9 mix

+ S9 mix

- S9 mix

+ S9 mix

0

24

31

151

187

13

13

9

12

4

27

33

153

185

10

14

9

9

20

25

35

156

189

10

15

9

9

100

26

34

155

184

12

13

9

11

500

26

33

150

187

15

12

9

11

2500

29

30

161

181

12

13

9

12

10000

31

34

161

184

12

16

7

10

Experiment I

 

TA 1538

WP2uvrA

 

 

 

- S9 mix

+ S9 mix

- S9 mix

+ S9 mix

 

 

 

 

0

20

27

29

31

 

 

 

 

4

19

23

28

31

 

 

 

 

20

19

25

30

26

 

 

 

 

100

21

27

29

28

 

 

 

 

500

20

27

32

31

 

 

 

 

2500

21

24

30

30

 

 

 

 

10000

18

23

29

30

 

 

 

 

Experiment II

 

TA 98

TA 100

TA1535

TA 1537

 

- S9 mix

+ S9 mix

- S9 mix

+ S9 mix

- S9 mix

+ S9 mix

- S9 mix

+ S9 mix

0

24

25

134

134

13

14

13

12

4

26

22

139

140

10

13

13

14

20

26

24

133

137

8

15

12

12

100

21

23

148

141

8

12

15

15

500

22

25

153

155

10

11

14

14

2500

18

31

151

153

10

10

14

11

5000

25

24

156

156

12

14

15

12

Experiment II

 

TA 1538

WP2uvrA

 

 

 

- S9 mix

+ S9 mix

- S9 mix

+ S9 mix

 

 

 

 

0

22

22

29

31

 

 

 

 

4

20

19

32

37

 

 

 

 

20

17

24

35

39

 

 

 

 

100

19

24

30

36

 

 

 

 

500

19

20

25

33

 

 

 

 

2500

19

21

28

39

 

 

 

 

5000

19

17

29

34

 

 

 

 

 

The sensitivity of the test system used was demonstrated by the induction of an increased number of revertants by the positive controls.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative

The test compound did not cause a significant increase in the number of revertant colonies with any of the tester strains either in the absence or presence of S9 mix. No dose dependent effect was obtained.

it is cocluded that the test substance is not mutagenic in these bacterial test systems either in the absence or in the presesnce of an exogenous metabolizing system.
Executive summary:

Ethansalz 97/100 % was tested for mutagenicity with the strains TA 100, TA 1535, TA 1537, TA 1538, TA 98 of Salmonella typhimurium and E. coli WP2uvrA.

The mutagenicity studies were conducted in the absence and presence of a metabolizing system derived from rat liver homogenate. A dose range of 6 different doses from 4 µg/plate to 10000 µg/plate was used.

Control plates without mutagen showed that the number of spontaneous revertant colonies was similar to that described in the literature. All the positve control compounds gave the expected increase in the number of revertant colonies.

Toxicity: The test compound proved to be not toxic to the bacterial strains.

Mutagenicity: In the absence of the metabolic activation system the test compound did not show a dose dependent increase in the number of revertants in any of the bacterial strains. Also in the presence of a metabolic activation system, treatment of the cells with Ethansalz 97/100 % did not result in relevant increases in the number of revertant colonies.

Summarizing, it can be stated that Ethansalz 97/100 % is not mutagenic in these bacterial test sytems either with or without exogenous metabolic activation at the dose levels investigated.