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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:
2000
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2000
Report date:
2000

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)
Deviations:
no
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
2-ethoxybenzenecarboximidamide hydrochloride
EC Number:
606-080-9
Cas Number:
18637-00-8
Molecular formula:
C9 H12 N2 O . Cl H
IUPAC Name:
2-ethoxybenzenecarboximidamide hydrochloride

Method

Target gene:
Histidine gene locus
Species / strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
Additional strain / cell type characteristics:
not applicable
Metabolic activation:
with and without
Metabolic activation system:
Aroclor 1254 induced male rat liver S9 mix
Test concentrations with justification for top dose:
0, 50, 158, 500, 1581, 5000 µg/plate and µg/tube (+/-S9 mix, all strains)








Vehicle / solvent:
deionized water
Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
no
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: sodium azide (only TA 1535), nitrofurantoin (only TA 100), 4-nitro-1,2-phenylene diamine (TA 1537 and TA 98), cumene hydroperoxide (only TA 102), 2-aminoanthracene.
Remarks:
The positive controls sodium azide, nitrofurantoin, 4-nitro-1,2-phenylene diamine and cumene hydroperoxide were only used without S9 mix; the positive control 2-aminoanthracene was only used with S9 mix.
Details on test system and experimental conditions:
METHOD: Standard plate test and preincubation test; each concentration including the controls was tested in triplicate.
Evaluation criteria:
A reproducible and dose-related increase in mutant counts of at least one strain is considered to be a positive result. For TA 1535, TA 100 and TA 98 this increase should be about twice that of negative controls, whereas for TA 1537 at least a threefold increase should be reached. For TA 102 an increase of about 100 mutants should be reached. Otherwise, the result is evaluated as negative. However, these criteria may be overruled by good scientific judgment. In case of questionable results, investigations should continue, possibly with modifications, until a final evaluation is possible.
Statistics:
not specified

Results and discussion

Test results
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
weak strain-specific bacteriotoxic effect at 5000 µg/plate, however, this dose could nevertheless be used for assessment purposes
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
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

Table 1: Summary of results from the Salmonella mutagenicity assay with Ethoxyamidin without S9 mix (mean values of revertants per plate)

 Dose (µg per plate)

Without metabolic activation

 

TA 1535

 TA 100

 TA 1537

 TA 98

 TA 102

0

8

81

8

31

182

50

8

72

8

25

180

158

9

74

9

32

172

500

10

80

9

30

170

1581

7

75

8

34

154

5000

9

76

6

33

145

 Positive control

704

198

145

173

346 

 Dose ( µg per tube)

Without metabolic activation

 

TA 1535

 TA 100

 TA 1537

 TA 98

TA 102

0

12

95

9

23

228

50

14

105

8

30

274

158

14

104

9

26

265

500

11

112

8

24

267

1581

12

99

7

31

237

5000

7

71

6

17

157

 Positive control

818

255

163

189

592

Table 2: Summary of results from the Salmonella mutagenicity assay with Ethoxyamidin with S9 mix (mean values of revertants per plate)

Dose (µg per plate)

With metabolic activation (liver S9 mix)

 

TA 1535

 TA 100

 TA 1537

 TA 98

 TA 102

0

8

81

8

31

182

50

8

72

8

25

180

158

9

74

9

32

162

500

10

80

9

30

170

1580

7

75

9

34

154

5000

9

76

8

33

145

 Positive control

704

198

145

173

346

 Dose ( µg per tube)

With metabolic activation (liver S9 mix)

 

TA 1535

 TA 100

 TA 1537

 TA 98

TA 102

0

12

95

9

23

258

50

14

105

8

30

274

158

14

104

9

26

265

500

11

112

8

24

267

1580

12

99

7

31

237

5000

7

71

6

17

157

 Positive control

818

255

163

189

592 

 

 

Doses up to and including 1580 µg per plate did not cause any bacteriotoxic effects. Total bacteria counts remained unchanged and no inhibition of growth was observed. At 5000 µg the substance had a weak strain-specific bacteriotoxic effect, not limiting evaluation.

No evidence of mutagenic activity of Ethoxyamidin was seen in any tester strain with and without metabolic activation.

The positive controls sodium azide, nitrofurantoin, 4-nitro-1,2-phenylene diamine, cumene hydroperoxide and 2-aminoanthracene had a marked mutagenic effect, as was seen by a biologically relevant increase in mutant colonies compared to the corresponding negative controls.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative
Executive summary:

The mutagenic potential of Ethoxyamidin was evaluated in a Salmonella/microsome test with the S. typhimurium strains TA 98, TA 100, TA 102, TA 1535 and TA 1537 in the presence and absence of S9 mix according to OECD TG 471. Doses up to and including 1580 µg per plate did not cause any bacteriotoxic effects. At 5000 µg/plate a weak strain-specific bacteriotoxic effect occurred, which however did not affect assessment purposes. No evidence of mutagenic activity of Ethoxyamindin was seen in the treated cultures with and without S9 -mix. Thus, the test item can be considered as non- mutagenic in the Ames Test.

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