Registration Dossier

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

Test material

Reference
Name:
Unnamed
Type:
Constituent

Method

Target gene:
Histidine gene locus
Species / strain
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
Additional strain 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 (first and repeat test)








Vehicle:
DMSO
Controls
Negative controls:
yes
Solvent controls:
no
Remarks:
No solvent control was used since sufficient evidence was available in the literature and from testing laboratory experience, indicating that the solvents used had no influence on the spontaneous mutant counts of the used strains.
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 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 1537, TA 100 and TA 98 this increase should be about twice that of negative controls. 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:
yes
Remarks:
weak, strain-specific bacteriotoxic effect at 5000 µg/plate
Vehicle controls valid:
not applicable
Negative controls valid:
yes
Positive controls valid:
yes
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 (first test) with Imidazotriazinon (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

49

8

15

140

50

10

51

5

16

144

158

5

56

6

17

132

500

6

55

6

12

119

1581

8

60

5

9

115

5000

7

53

4

9

88

 Positive control

629

153

108

156

281 

 Dose ( µg per plate )

With metabolic activation (liver S9 mix)

 

 TA 1535

 TA 100

 TA 1537

 TA 98

TA 102

0

9

72

10

27

211

50

7

114

10

23

215

158

8

107 

6

24

225

500

8

72

8

21

194

1581

7

91

7

27

229

5000

6

98

6

23

194

 Positive control

186

946

59

2269

820

Table 2: Summary of results from the Salmonella mutagenicity assay (repeat test) with Imidazotriazinon (mean values of revertants per plate)

 Dose (µg per plate)

Without metabolic activation

 

 TA 1535

 TA 100

 TA 1537

 TA 98

 TA 102

0

9

61

8

16

223

50

11

75

7

15

219

158

10

77

6

16

210

500

7

64

7

16

185

1581

8

80

9

16

196

5000

6

77

-

14

138

 Positive control

518

218

112

127

452 

 Dose ( µg per plate )

With metabolic activation (liver S9 mix)

 

 TA 1535

 TA 100

 TA 1537

 TA 98

TA 102

0

8

75

8

31

299

50

9

92

8

22

315

158

8

101 

10

28

301

500

8

92

9

20

265

1581

8

122

9

23

277

5000

9

116

-

23

199

 Positive control

146

1386

181

1193

526

Doses up to and including 1581 µg per plate did not cause any bacteriotoxic effects. Total bacteria counts remained unchanged and no inhibition of growth was observed. At 5000 µg per plate, the substance had only a weak, strain-specific bacteriotoxic effect. Due to the weakness of this effect this dose could nevertheless be used for assessment purposes. Substance precipitation occurred at the dose 1581 µg per plate and above.

Evidence of mutagenic activity of Imidazotriazinon was not seen. No biologically relevant increase in the mutant count, in comparison with the negative controls, was observed.

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 Imidazotriazinon 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. Evidence of mutagenic activity was not seen up to the maximum recommended dose level of 5000 µg/plate. No biologically relevant increase in the mutant count, in comparison with the negative controls, was observed. Based on this test, the test substance was considered to be non-mutagenic without and with S9 mix in the plate incorporation as well as in the preincubation modification of the Salmonella/microsome test.