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Toxicological information

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:
22 Mar - 30 May 2002
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2002
Report Date:
2002

Materials and methods

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

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Silan P-Triethoxy
- Substance type: Colourless liquid
- Physical state: Liquid at room temperature
- Lot/batch No.: GE 03287
- Expiration date of the lot/batch: 2004-01-01
- Storage condition of test material: At room temperature, no direct sun light

Method

Target gene:
his operon
Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Species / strain / cell type:
S. typhimurium TA 102
Metabolic activation:
with and without
Metabolic activation system:
cofactor supplemented post-mitochondrial fraction (S9 mix), prepared from the livers of rats treated with Aroclor 1254
Test concentrations with justification for top dose:
See table 1
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: abs. ethanol
- Justification for choice of solvent/vehicle: Solvent chosen due to solubility properties and relative nontoxicity to bacteria.
Controls
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: Sodium azide (TA 1535, TA 100); 2-nitrofluorene (TA 98); 9-aminoacridine (TA 1537); ethyl methanesulphonate (TA 102); +S9: 2-anthracene amide (TA 98, TA 102, TA 1537); cyclophosphamide (TA 100, TA 1535)
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation); preincubation

DURATION
- Preincubation period: 60 minutes
- Exposure duration: 48 hours

NUMBER OF REPLICATIONS: 3 plates for each test concentration

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth

POSITIVE CONTROLS
Plate incorporation
-S9: Sodium azide (TA 1535: 770.3 µg/plate, TA 100: 1142.3 µg/plate); 2-nitrofluorene (TA 98: 648.3 µg/plate); 9-aminoacridine (TA 1537: 527 µg/plate); ethyl methanesulphonate (TA 102: 1171 µg/plate); +S9: 2-anthracene amide (TA 98: 618.7 µg/plate, TA 102: 1200 µg/plate, TA 1537: 557.7 µg/plate); cyclophosphamide (TA 100: 1094.7 µg/plate, TA 1535:728.7 µg/plate)

Preincubation
-S9: Sodium azide (TA 1535: 808.3 µg/plate, TA 100: 1145 µg/plate); 2-nitrofluorene (TA 98: 618.7 µg/plate); 9-aminoacridine (TA 1537: 653.3 µg/plate); methyl methanesulphonate (TA 102: 789.3 µg/plate); +S9: 2-anthracene amide (TA 98: 731.7 µg/plate, TA 102: 1125.3 µg/plate, TA 1537: 793.3 µg/plate); cyclophosphamide (TA 100: 1133 µg/plate, TA 1535: 891 µg/plate)


Evaluation criteria:
A result is positive if the number of revertants is significantly increased compared with the solvent control to at least 2-fold of the solvent control for TA 98, TA 100 and TA 102 and 3-fold of the solvent control for TA 1535 and TA 1537 in both independant experiments. Positive results have to be reproducible and the histidine independence of the revertants has to be confirmed by streaking random samples on histidine-free agar plates.
Statistics:
MANN and WHITNEY and Spearman's rank correlation.

Results and discussion

Test resultsopen allclose all
Species / strain:
S. typhimurium, other: TA 1535, TA 1537, TA 98 and TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
Scarce background lawn from 100 µg. >50% reduction in revertants from 316 µg.
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 102
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
Scarce background lawn from 100 µg. >50% reduction in revertants from 316 µg.
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
Positive controls validity:
valid
Additional information on results:
See tables 1-4

COMPARISON WITH HISTORICAL CONTROL DATA: Controls were within range of historical data.


Any other information on results incl. tables

Table 2: Dose range-finding study Number of revertants per plate (2 plates)

 

TA100

Conc.
(µg/plate)

Plate 1

Plate 2

Cytotoxic
(yes/no)

0*

118

122

No

0.316

108

136

No

1

112

129

No

3.16

127

100

No

10

108

113

No

31.6

120

112

No

100#

135

216

Yes

316#

105

0

Yes

1000#

149

139

Yes

3160

0

0

Yes

5000

0

0

Yes

*solvent control with Abs. ethanol

#colonies may represent pinpoint colonies rather than revertants

Table 3: Experiment 1 Plate incorporation Number of revertants per plate (mean of 3 plates)

 

TA98

TA100

TA102

Conc.
(
µg/plate)

— MA

+ MA

Cytotoxic
(yes/no)

— MA

+ MA

Cytotoxic
(yes/no)

— MA

+ MA

Cytotoxic
(yes/no)

0*

34

40.7

No

126

121.7

No

258

268.3

No

3.16

29

36

No

119.7

123.7

No

238.3

307.7

No

10

31.3

41.7

No

119.7

112.3

No

252.7

290.3

No

31.6

30.7

38.3

No

133.3

113.3

No

255.3

282.3

No

100

30

32.7

No

110

109.3

No

254.3

269.7

No

316

33.7

37.7

No

127.7

127.3

Yes

254.7

304

No

Positive control

618.7

618.7

No

1142.3

1094.7

No

789.3

1200

No

*solvent control with Abs. ethanol

Table 3: Experiment 1 Plate incorporation Number of revertants per plate (mean of 3 plates)

 

TA1535

TA1537

Conc.
(
µg/plate)

— MA

+ MA

Cytotoxic
(yes/no)

— MA

+ MA

Cytotoxic
(yes/no)

0*

15.7

15.3

No

10.7

10.3

No

3.16

18.7

11.3

No

6.7

9

No

10

17.7

15.3

No

7

11.7

No

31.6

16

16

No

8.3

10.3

No

100

15.7

21.7

No

7.7

10

No

316

12.7

12.7

Yes

9.7

11.3

Yes

Positive control

808.3

728.7

No

527

557.7

No

*solvent control with Abs. ethanol

Table 4: Experiment 2 Preincubation Number of revertants per plate (mean of 3 plates)

 

TA98

TA100

TA102

Conc.
(
µg/plate)

— MA

+ MA

Cytotoxic
(yes/no)

— MA

+ MA

Cytotoxic
(yes/no)

— MA

+ MA

Cytotoxic
(yes/no)

0*

28.7

36

No

122.7

131

No

268

251.3

No

3.16

31

34

No

123.3

128

No

259.7

277.3

No

10

27.7

28.3

No

122

134.7

No

247.7

248

No

31.6

26.3

39

No

121

136.7

No

247.3

317.7

No

100

25

0

Yes

145.3

139.3

Yes

207.3

277.3

Yes

316

26

0

Yes

0

0

Yes

258

253.7

Yes

Positive control

648.3

731.7

No

1145

1133

No

1171

1125.3

No

*solvent control with Abs. ethanol

Table 4: Experiment 2 Preincubation Number of revertants per plate (mean of 3 plates)

 

TA1535

TA1537

Conc.
(
µg/plate)

— MA

+ MA

Cytotoxic
(yes/no)

— MA

+ MA

Cytotoxic
(yes/no)

0*

15

14.3

No

11.7

13

No

3.16

11.3

13.7

No

12

12.3

No

10

12

12

No

16.7

12.7

No

31.6

9.7

14.7

No

11

14.3

No

100

0

0

Yes

0

0

Yes

316

0

0

Yes

0

0

Yes

Positive control

770.3

891

No

653.3

793.3

No

*solvent control with Abs. ethanol

Applicant's summary and conclusion

Conclusions:
Interpretation of results: negative

Pronounced cytotoxicity (scarce background lawn) was noted at the concentrations of 100 µg up to 5000 µg of the test substance. Reduction of the number of revertants by more than 50% was noted at concentrations of 316, 3160 and 5000 µg of the test substance.

In a highly reliable test, conducted in accordance with OECD 471, under GLP, no mutagenic effect was observed for the test substance in any of the 5 test strains in two independent experiments without and with metabolic activation (plate incorporation or preincubation test). The test substance is not mutagenic in the test strains used.