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

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2003
Report Date:
2003

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

Method

Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Species / strain / cell type:
E. coli WP2 uvr A
Metabolic activation:
with and without
Metabolic activation system:
Phenobarbital beta-naphthoflavone induced rat liver S9
Test concentrations with justification for top dose:
33, 100, 333, 1000, 2500 and 5000 μg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: Ethanol

- Justification for choice of solvent/vehicle: the solvent was chosen because of its solubility properties and relative non-toxicity to bacteria, and on request of the sponsor.
Controlsopen allclose all
Untreated negative controls:
yes
Remarks:
untreated
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: 2-aminoanthracene
Remarks:
All strains (with activation)
Untreated negative controls:
yes
Remarks:
untreated
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: 4-nitro-o-phenylene-diamine
Remarks:
TA 98, TA 1537 (without activation)
Untreated negative controls:
yes
Remarks:
untreated
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
TA 100, TA 1535 (without activation)
Untreated negative controls:
yes
Remarks:
untreated
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
methylmethanesulfonate
Remarks:
WP2uvrA (without activation)
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation); preincubation

DURATION

- Preincubation period: 60 minutes

- Expression time (cells in growth medium): 48 hours

NUMBER OF REPLICATIONS: 3 plates for each test concentration

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth; background lawn
Evaluation criteria:
The test article is considered a mutagen if a biologically relevant increase in the number of revertants exceeding the threshold of twice (strains TA-98, TA-100 and WP2 uvrA) or thrice (strains TA-1535, TA-1537) the colony count of the corresponding solvent control is observed. An increase exceeding the threshold at only one concentration is judged as biologically relevant if produced in an independent second experiment. A dose dependent increase is considered biologically relevant if the threshold is exceeded at more than one concentration. A dose dependent increase in the number of revertant colonies below the threshold is regarded as an indication of a mutagenic potential if reproduced in an independent second experiment. However, whenever the colony counts remain within the historical range of the negative and solvent controls such an increase is not considered biologically relevant.
Statistics:
No statistical evaluation is required. Responses (number of revertants) to the test article were compared to concurrent
negative and positive controls.

Results and discussion

Test resultsopen allclose all
Key result
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98 and 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
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Key result
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
Additional information on results:
No substantial increase in revertants colony numbers of any the five tester strains was observed following treatment with D5 at any dose level, either in the presence or absence of metabolic activation. Appropriate reference mutagens were used as positive controls and showed a distinct increase of induced revertant colonies. The controls were within historical values with the following exceptions: Strain TA 1535 without activation: number of revertants greater than historical values; small deviations were observed in the negative controls of two strains in the second experiment. These deviations do not affect the validity of the experiment.

Any other information on results incl. tables

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

 

TA 1535

TA 1537

TA 98

Conc.
µg/plate

- MA

+ MA

Cytotoxic
(yes/no)

- MA

+ MA

Cytotoxic
(yes/no)

- MA

+ MA

Cytotoxic
(yes/no)

Untreated

 16

 17

no

 6

 21

no

 27

32

no

0*

 13

 27

no

 8

 15

no

 28

 44

no

33

 13

 14

no

 5

 13

no

 30

 41

no

100

 14

 17

no

 5

 11

no

 32

 42

no

333

 10

 18

no

 7

 16

no

 34

 42

no

1000

 15

 14

no

 9

 15

no

 30

 40

no

2500

 14

 16

no

 7

 11

no

 33

 39

no

5000

 16

 13

no

 8

 13

no

 36

 33

no

Positive control

 884

 161

 -

 58

 67

 -

 273

 658

-

*solvent control with ethanol

 

 Table 2 Experiment 1 plate incorporation: Number of revertants per plate (mean of 3 plates)

 

TA 100

E coli WP2 uvrA

Conc.
µg/plate

- MA

+ MA

Cytotoxic
(yes/no)

- MA

+ MA

Cytotoxic
(yes/no)

Untreated

 162

178

no

 35

 42

no

0*

 152

164

no

 40

 47

no

33

 166

194

no

 38

36

no

100

 156

190

no

 45

 39

no

333

 151

215

no

 35

 43

no

1000

 148

207

no

 35

 47

no

2500

 164

206

no

 26

 46

no

5000

 144

190

no

 31

 47

no

Positive control

 773

 970

 -

 768

 180

 -

*solvent control with ethanol

 

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

 

TA 1535

TA 1537

TA 98

Conc.
µg/plate

- MA

+ MA

Cytotoxic
(yes/no)

- MA

+ MA

Cytotoxic
(yes/no)

- MA

+ MA

Cytotoxic
(yes/no)

Untreated

 18

 16

no

 8

 15

no

 33

 40

no

0*

 15

 21

no

 9

 12

no

 38

 48

no

33

 11

17

no

 11

 14

no

 32

 39

no

100

 16

 20

no

 10

 9

no

 34

 46

no

333

21

 19

no

 8

 15

no

 40

 48

no

1000

 18

 17

no

 11

 13

no

 41

 47

no

2500

 15

 15

no

 9

 10

no

 40

 48

no

5000

 16

 14

no

 8

 11

no

 43

 55

no

Positive control

 1319

 147

-

 72

 64

no

 229

 507

 -

*solvent control with ethanol

 

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

 

TA 100

E coli WP2 uvrA

Conc.
µg/plate

- MA

+ MA

Cytotoxic
(yes/no)

- MA

+ MA

Cytotoxic
(yes/no)

Untreated

178

248

no

38

31

no

0*

206

287

no

44

37

no

33

191

265

no

32

49

no

100

187

251

no

38

51

no

333

213

244

no

41

46

no

1000

185

239

no

36

38

no

2500

217

269

no

39

35

no

5000

229

318

no

47

43

no

Positive control

581

624

-

284

166

-

*solvent control with ethanol

Applicant's summary and conclusion

Conclusions:
Decamethylcyclopentasiloxane has been tested according to OECD 471 and in compliance with GLP using Salmonella typhimurium strains TA 1535, TA 1537, TA 98 and TA 100 and Escherichia coli WP2 uvrA. No test-substance related increase in the number of revertants was detected in the presence or absence of metabolic activation in either the initial (RCC Cytotest cell research, 2003) plate incorporation assay or the repeat experiment using pre-incubation, up to limit concentrations. Appropriate positive, solvent and untreated controls were included and gave expected results. The test substance is therefore considered to be negative for mutagenicity to bacteria under the conditions of the test.