Registration Dossier

Diss Factsheets

Administrative data

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Bacterial reverse mutation assay: the substance 1,3,5-trimethyl-1,1,3,5,5-pentaphenyltrisiloxane was negative with and without metabolic activation in S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102 (LPT, 2002) (OECD TG 471).


Cytogenicity in mammalian cells: the substance 1,3,5-trimethyl-1,1,3,5,5-pentaphenyltrisiloxane was negative with and without metabolic activation in peripheral human lymphocytes (WIL Research, 2016) (OECD TG 473).


Mammalian mutagenicity study: the substance 1,3,5-trimethyl-1,1,3,5,5-pentaphenyltrisiloxane was negative with and without metabolic activation in mouse lymphoma L5178Y cells (Charles River, 2016) (OECD TG 490).

The studies were conducted according to an appropriate OECD test guideline, and in compliance with GLP.

Link to relevant study records

Referenceopen allclose all

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2002-05-10 to 2002-09-02
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Version / remarks:
1997
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Type of assay:
bacterial reverse mutation assay
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
Metabolic activation:
with and without
Metabolic activation system:
Aroclor induced rat liver S9
Test concentrations with justification for top dose:
See table 1
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO

- Justification for choice of solvent/vehicle: The solvent was chosen because of its solubility properties and its relative nontoxicity to the bacteria
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
TA 1535, TA 100 (without activation)
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
2-nitrofluorene
Remarks:
TA 98 (without activation)
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
9-aminoacridine
Remarks:
TA 1537 (without activation)
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
methylmethanesulfonate
Remarks:
TA 102 (without activation)
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: 2-anthracene amide
Remarks:
TA 98, TA 102, TA 1537 (with activation)
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
cyclophosphamide
Remarks:
TA 100, TA 1535 (with 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 assessment
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 experiments.

Positive results have to be reproducible and the histidine independence of the revertants has to be confirmed by streaking on histidine-free agar plates.

Cytotoxicity is defined as a reduction in the number of colonies by >50% compared with the solvent control and/or a sparse background lawn.

Statistics:
MANN and WHITNEY and Spearman’s rank.
Key result
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:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS

- Precipitation: precipitation noted at 3160 - 5000 μg/plate

COMPARISON WITH HISTORICAL CONTROL DATA: Results were within range of historical control data


Remarks on result:
other: No mutagenic potential

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

TA 100

Concentration (μg/Plate)

Plate 1

Plate 2

Cytotoxic (Yes/No)

0

143

146

No

0.316

154

138

No

1

188

140

No

3.16

171

144

No

10

161

162

No

31.6

144

145

No

100

145

138

No

316

115

130

No

1000

156

135

No

3160

195

154

No

5000

171

146

No

*solvent control with DMSO

Table 3: Experiment 1 Plate incorporation Assay 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*

32

36

No

131

137.3

No

298

279.7

No

10

39.3

43

No

130.7

133.3

No

272

280.3

No

31.6

38.3

39.3

No

122

136.7

No

258.3

260.7

No

100

34.3

31.3

No

131

123.7

No

265.3

283.3

No

316

27.7

36.7

No

113

131

No

276

284.3

No

1000

23.3

24.7

No

120

132.3

No

293.7

285.3

No

Positive control

1255

1295.7

No

1194

1169

No

1130.7

1289

No

*solvent control with DMSO

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

 

TA1535

TA 1537

Conc.
(
µg/plate)

— MA

+ MA

Cytotoxic
(yes/no)

— MA

+ MA

Cytotoxic
(yes/no)

0*

13.7

13.7

No

5.3

6

No

10

12.7

13.7

No

4

5.3

No

31.6

13

13

No

3.7

7.3

No

100

14.3

13.3

No

3

7

No

316

13

12

No

5.3

6

No

1000

12.7

14

No

5

5

No

Positive control

564.3

563

No

570.7

566.7

No

*solvent control with DMSO

Table 4: Experiment 2 Pre incubation test 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*

31

34.7

No

144.7

160.7

No

286.3

280

No

100

37.3

36.7

No

128.3

135

No

288

281

No

316

35

33.3

No

155.7

164.3

No

257.3

261.7

No

1000

31.3

27.3

No

150.7

155

No

271.3

267.7

No

3160

30.3

37.3

No

145.3

127

No

270.3

268.7

No

5000

30.7

33

No

131

129

No

282.7

264.3

No

Positive control

1033.7

1141

No

1186.7

1139.7

No

1163.3

1103.7

No

*solvent control with DMSO

Table 3: Experiment 2 Pre incubation test Number of revertants per plate (mean of 3 plates)

 

TA1535

TA 1537

Conc.
(
µg/plate)

— MA

+ MA

Cytotoxic
(yes/no)

— MA

+ MA

Cytotoxic
(yes/no)

0*

15

14

No

5.7

5.7

No

100

13.7

13.7

No

3.7

5

No

316

13

13.7

No

4.7

6

No

1000

14.3

13

No

5

5.7

No

3160

15.7

12.7

No

5.7

5.7

No

5000

14

13

No

4.7

5

No

Positive control

554.3

548

No

553.3

550

No

*solvent control with DMSO

Conclusions:
In a highly reliable test, conducted under OECD 471, with GLP, no mutagenic effect was observed for the test substance tested up to cytotoxic concentration in any of the test strains in two independent experiments without and with metabolic activation. The test substance is non-mutagenic in test strains used.
Endpoint:
in vitro cytogenicity / chromosome aberration study in mammalian cells
Remarks:
Type of genotoxicity: chromosome aberration
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2016-01-15 to 2016-03-08
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
Version / remarks:
2014
Deviations:
no
GLP compliance:
yes
Type of assay:
in vitro mammalian chromosome aberration test
Species / strain / cell type:
other: Peripheral human lymphocytes
Details on mammalian cell type (if applicable):
- Type and identity of media: RPMI 1640 medium
- Properly maintained: yes
Metabolic activation:
with and without
Metabolic activation system:
phenobarbital (80 mg/kg body weight) and ß-naphthoflavone (100 mg/kg) induced rat liver S9
Test concentrations with justification for top dose:
Experiment 1: 1.7, 5.4, 17, 52, 164 µg/ml, with and without metabolic activation
Experiment 2: 5.4, 17 and 52 µg/ml, without metabolic activation
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: dimethyl sulfoxide (DMSO)
- Justification for choice of solvent/vehicle: The solvent was chosen based on the results from the solubility test.
Untreated negative controls:
yes
Remarks:
DMSO
Negative solvent / vehicle controls:
no
True negative controls:
no
Positive controls:
yes
Positive control substance:
mitomycin C
Remarks:
without metabolic activation, 48 hour exposure period
Untreated negative controls:
yes
Remarks:
DMSO
Negative solvent / vehicle controls:
no
True negative controls:
no
Positive controls:
yes
Positive control substance:
cyclophosphamide
Remarks:
with metabolic activation, 3 hour exposure period
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium

ACTIVATION: To 0.5 ml S9-mix components 0.5 ml S9-fraction was added (50% (v/v) S9-fraction) to complete the S9-mix. The added co-factores were: 1.63 mg MgCl2.6H2O; 2.46 mg KCl; 1.7 mg glucose-6-phosphate; 3.4 mg NADP; 4 µmol HEPES.

DURATION

- Exposure duration:
Experiment 1: 3 hours, with and without metabolic activation
Experiment 2: 24 and 48 hours, without metabolic activation
- Expression time (cells in growth medium):
Experiment 1: 20-22 hours

- Fixation time (start of exposure up to fixation or harvest of cells):
Experiment 1: 24 hours
Experiment 2: The cells were fixed immediately after exposure

SPINDLE INHIBITOR (cytogenetic assays): colchicine (0.5 µg/ml medium) was added during the last 2.5 - 3 h of the culture period
STAIN (for cytogenetic assays): 5 % v/v Giemsa

NUMBER OF REPLICATIONS: duplicate cell cultures

NUMBER OF CELLS EVALUATED: 1000 cells

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index

OTHER EXAMINATIONS:
- Determination of polyploidy: yes
- Determination of endoreplication: yes
Evaluation criteria:
A test item is considered positive (clastogenic) in the chromosome aberration test if all of the following criteria are met:
a) at least one of the test concentrations exhibits a statistically significant (Fisher’s exact test, one-sided, p < 0.05) increase compared with the concurrent negative control.
b) The increase is dose related when evaluated with an Cochran Armitage trend test.
c) Any of the results are outside the 95% control limits of the historical control data range.
Statistics:
Fisher’s exact test and Cochran Armitage trend test
Key result
Species / strain:
other: Peripheral human lymphocytes
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity, but tested up to precipitating concentrations
Vehicle controls validity:
not applicable
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS

- Precipitation:
Experiment 1: At the 3 h exposure time a concentration of 164 µg/ml the test item precipitated in the culture medium
Experiemnt 2: ). At 24 and 48 hour exposure times a concentration of 52 µg/ml test item precipitated in the culture medium

ADDITIONAL INFORMATION ON CYTOTOXICITY:
- Polyploidy: The test item did not increase the number of polyploid cells.
- Endoreplication: The test item did not increase the number of endoreplicated chromosomes
Remarks on result:
other: No mutagenic potential

Table 1: Mitotic index of human lymphocyte cultures treated with the test item in the first (experiement 1) cytogenetic assay

Test item concentration (µg/ml)

Number of metaphases

 

 

Absolute

Number of cells scored

Percentage of control

3 hour exposure, 24 hour fixation time, without metabolic activation

Control (DMSO)

96 - 110

1027 - 1011

100

17

82 - 89

1018 - 1010

83

52

86 - 95

1008 - 1011

88

164

72 - 93

1015 - 1002

80

MMC-C; 0.5 µg/ml

55 - 63

1020 - 1011

57

MMC-C; 0.75 µg/ml

45 - 43

1022 - 1007

43

3 hour exposure, 24 hour fixation time, with metabolic activation

Control

107 – 98

1007 – 1014

100

17

99 – 85

1009 – 1026

90

52

95 – 101

1016 – 1008

96

164

94 – 82

1002 – 1012

86

CP; 10 µg/ml

62 - 50

1014 - 1004

55

Table 2: Chromosome aberrations in human lymphocyte cultures treated with the test item in the absence of S9-mix in the first (experiment 1) cytogenetic assay (3 h exposure time, 24 h fixation time)

Concentration

DMSO

(1.0% v/v)

17

µg/ml

52

µg/ml

164

µg/ml

MMC-C

0.5µg/ml

Culture

A           B     A+B

A           B     A+B

A           B     A+B

A           B     A+B

A           B     A+B

Mitotic

Index (%)

          100

            83

            88

            80

            57

No. of

Cells scored

150    150     300

150    150     300

150    150     300

150    150     300

150    150     300

No. of

Cells with

aberrations

(+ gaps)a)

1

0

1

0

0

0

0

0

0

0

0

0

40

48

***)

88

 

No. of

Cells with

aberrations

(- gaps)

1

0

1

0

0

0

0

0

0

0

0

0

40

46

***)

86

 

total aberr

(+ gaps)

1

0

 

0

0

 

0

0

 

0

0

 

51

48

 

total aberr

(- gaps)

1

0

 

0

0

 

0

0

 

0

0

 

51

46

 

Table 3: Chromosome aberrations in human lymphocyte cultures treated with the test item in the presence of S9-mix in the first (experiment 1) cytogenetic assay (3 h exposure time, 24 h fixation time)

Conc                                                 

DMSO

(1.0% v/v)

17

µg/ml

52

µg/ml

164

µg/ml

CP

10µg/ml

Culture

A           B     A+B

A           BA+B

A           BA+B

A           BA+B

A           B A+B

Mitotic

Index (%)

          100

            90

            96

            86

            55

No. of

Cells scored

150    150     300

150    150300

150    150300

150    150300

150    150 300

No. of

Cells with

aberrations

(+ gaps)a)

0

1

1

0

0

0

0

0

0

0

0

0

36

30

***)

66

 

No. of

Cells with

aberrations

(- gaps)

0

1

1

0

0

0

0

0

0

0

0

0

36

30

***)

66

 

total aberr

(+ gaps)

0

1

 

0

0

 

0

0

 

0

0

 

42

37

 

total aberr

(- gaps)

0

1

 

0

0

 

0

0

 

0

0

 

42

37

 

 

Table 4: Mitotic index of human lymphocyte cultures treated with the test item in the second (experiement 2) cytogenetic assay

Test item concentration (µg/ml)

Number of metaphases

 

 

Absolute

Number of cells scored

Percentage of control

24 hour exposure, 24 hour fixation time, without metabolic activation

Control

69 - 78

1021 - 1013

100

5.4

68 - 69

1013 - 1016

93

17

76 - 66

1033 -1002

97

52

73 - 69

1030 - 1018

97

MMC-C; 0.2 µg/ml

39 - 45

1029 - 1005

57

MMC-C; 0.3 µg/ml

18 - 23

1017 - 1010

28

48 hour exposure, 48 hour fixation time, without metabolic activation

Control

44 - 44

1014 - 1013

100

5.4

47 – 43

1013 - 1021

102

17

37 - 37

1003 - 1004

84

52

38 - 37

1013 - 1006

85

MMC-C; 0.1 µg/ml

34 - 36

1020 – 1016

80

MMC-C; 0.15 µg/ml

29 - 31

1003 - 1021

68

Table 5: Chromosome aberrations in human lymphocyte cultures treated with the test item in the absence of S9-mix in the second (experiment 2) cytogenetic assay (24 h exposure time, 24 h fixation time)

Conc

DMSO

(1.0% v/v)

5.4

µg/ml

17

µg/ml

52

µg/ml

MMC-C

0.2µg/ml

Culture

A           B  A+B

A           B  A+B

A           B  A+B

A           B  A+B

A           B    A+B

Mitotic

Index (%)

          100

            93

            97

            97

            57

No. of

Cells scored

150    150   300

150    150   300

150    150   300

150    150   300

150    150     300

No. of

Cells with

aberrations

(+ gaps)a)

0

1

1

1

1

2

2

0

2

1

0

1

53

43

***)

96

 

No. of

Cells with

aberrations

(- gaps)

0

1

1

1

1

2

2

0

2

1

0

1

52

42

***)

94

 

total aberr

(+ gaps)

0

1

 

1

1

 

2

0

 

1

0

 

60

56

 

total aberr

(- gaps)

0

1

 

1

1

 

2

0

 

1

0

 

59

53

 

Table 6: Chromosome aberrations in human lymphocyte cultures treated with the test item in the absence of S9-mix in the second (experiement 2) cytogenetic assay (48 h exposure time, 48 h fixation time)

Conc

DMSO

(1.0% v/v)

5.4

µg/ml

17

µg/ml

52

µg/ml

MMC-C

0.1µg/ml

Culture

A           B     A+B

A           B     A+B

A           B     A+B

A           B     A+B

A           B     A+B

Mitotic

Index (%)

          100

          102

            84

            85

            80

No. of

Cells scored

150    150     300

150    150     300

150    150     300

150    150     300

150    150     300

No. of

Cells with

aberrations

(+ gaps)a)

0

1

1

0

0

0

0

1

1

0

0

0

42

53

***)

95

 

No. of

Cells with

aberrations

(- gaps)

0

1

1

0

0

0

0

0

0

0

0

0

42

52

***)

94

 

total aberr

(+ gaps)

0

1

 

0

0

 

0

1

 

0

0

 

51

66

 

total aberr

(- gaps)

0

1

 

0

0

 

0

0

 

0

0

 

51

65

 

*) Significantly different from control group (Fisher’s exact test), * P < 0.05, ** P < 0.01 or *** P < 0.001

Conclusions:
1,3,5-Trimethyl-1,1,3,5,5-pentaphenyltrisiloxane has been tested in a valid study for ability to induce chromosome aberrations in Peripheral human lymphocytes according to OECD TG 473 (2014), and in compliance with GLP. No statistically significant or biologically relevant increase in the number of cells with chromosome aberrations was observed either with or without metabolic activation when tested up to precipitating concentrations. Appropriate positive and negative controls were used and gave the expected results. It is concluded that the test item is negative for the induction of chromosome aberrations under the conditions of the study.
Endpoint:
in vitro gene mutation study in mammalian cells
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2016-06-07 to 2016-08-22
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 490 (In Vitro Mammalian Cell Gene Mutation Tests Using the Thymidine Kinase Gene)
Version / remarks:
2015
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Type of assay:
other: in vitro mammalian cell gene mutation assay
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: 0006760363
- Expiration date of the lot/batch: 8th November 2016
- Purity test date: 8th October 2015

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: at room temperature
- Stability under test conditions: stable
- Solubility and stability of the test substance in the solvent/vehicle: dimethyl sulfoxide
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: stable in dimethyl sulfoxide

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: no specific handling conditions required
- Preliminary purification step (if any): no correction was made for the purity/composition of the test item
Target gene:
thymidine kinase locus
Species / strain / cell type:
mouse lymphoma L5178Y cells
Details on mammalian cell type (if applicable):
CELLS USED
- Source of cells: American Type Culture Collection, USA
- Suitability of cells: recommended test system in international guidelines
- Cell cycle length, doubling time or proliferation index: no data
- Sex, age and number of blood donors if applicable: no data
- Whether whole blood or separated lymphocytes were used if applicable: no data
- Number of passages if applicable: no data
- Methods for maintenance in cell culture if applicable: no data
- Modal number of chromosomes: no data
- Normal (negative control) cell cycle time: no data

MEDIA USED
- Type and identity of media including CO2 concentration if applicable:
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: yes
- Periodically checked for karyotype stability: yes
- Periodically 'cleansed' against high spontaneous background: yes
Metabolic activation:
with and without
Metabolic activation system:
of phenobarbital (80 mg/kg body weight) and ß-naphthoflavone (100 mg/kg body weight) induced rat liver S9
Test concentrations with justification for top dose:
Dose range finding test: 1.7, 5.4, 17, 52, 164, 512 μg/ml (with and without S9 mix)
Experiment I: 0.056, 0.18, 0.55, 1.7, 5.4, 17, 52 and 164 μg/ml (with and without S9 mix)
Experiment II: 0.018, 0.056, 0.18, 0.55, 1.7, 5.4, 17, 52 and 164 μg/ml (without S9 mix)
Doses were selected based on cytotoxicity results from dose-range finding test
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: dimethyl sulfoxide
- Justification for choice of solvent/vehicle: based on solubility in dimethyl sulfoxide observed during a solubility test.
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
Remarks:
dimethyl sulfoxide
True negative controls:
no
Positive controls:
yes
Positive control substance:
methylmethanesulfonate
Remarks:
without metabolic activation
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
Remarks:
dimethyl sulfoxide
True negative controls:
no
Positive controls:
yes
Positive control substance:
cyclophosphamide
Remarks:
with metabolic activation
Details on test system and experimental conditions:
METHOD OF APPLICATION: in medium
- Cell density at seeding: 9.6 x 10⁵ cells/ concentration

ACTIVATION: S9 mix was prepared immediately before use and kept on ice. S9-mix components contains per ml physiological saline: 1.63 mg MgCl2 H2O; 2.46 mg KCl; 1.7 mg glucose-6-phosphate; 3.4 mg NADP; 4 µmol HEPES. The final concentration of the S9-fraction in the exposure medium was 4%.

DURATION
- Exposure duration:
Dose-range finding test: 3-hour treatment with and without metabolic activation
Experiment I: 3-hour treatment, with and without metabolic activation
Experiment II: 24-hour treatment, without metabolic activation
- Expression time (cells in growth medium): the remaining cells were cultured for 2 days after the treatment period.
- Selection time (if incubation with a selection agent): 11-12 days
- Fixation time (start of exposure up to fixation or harvest of cells):

SELECTION AGENT (mutation assays): selective medium (TFT-selection agent)

NUMBER OF REPLICATIONS: duplicate plates

METHODS OF SLIDE PREPARATION AND STAINING TECHNIQUE USED: the plates of the TFT-selection were stained for 2 hours by adding 0.5 mg/ml 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) (Sigma) to each well.

NUMBER OF CELLS EVALUATED:

DETERMINATION OF CYTOTOXICITY
- Method: cloning efficiency; relative total growth
- Any supplementary information relevant to cytotoxicity: suspension growth

OTHER EXAMINATIONS:
- Determination of polyploidy: no
- Determination of endoreplication: no

Rationale for test conditions:
The test concentrations in experiment I and II were selected based on cytotoxicity data from a dose-range finding study.
Evaluation criteria:
A test item is considered positive (mutagenic) in the mutation assay if it induces a mutation frequency (MF) of more than MF(controls) + 126 in a dose-dependent manner. Any observed increase should be biologically relevant and will be compared with the historical control data range.
Statistics:
A Cochran Armitage trend test (p < 0.05) was performed to test whether there was a significant trend in the induction of mutations.
Key result
Species / strain:
mouse lymphoma L5178Y cells
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity, but tested up to precipitating concentrations
Vehicle controls validity:
not applicable
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: no data
- Effects of osmolality: no data
- Evaporation from medium: the test item was insoluble in exposure medium and ethanol
- Water solubility: no data
- Precipitation: when mixing with exposure medium the test item precipitated at the concentration of 16.4 mg/ml and higher.
- Definition of acceptable cells for analysis: no data


RANGE-FINDING/SCREENING STUDIES: No toxicity in the suspension growth was observed up to and including the highest test item concentration of 512 μg/ml compared to the suspension growth of the solvent control both in the absence and presence of S9-mix. No toxicity in the relative suspension growth was observed up to the test item concentrations of 512 μg/ml compared to the solvent control.


HISTORICAL CONTROL DATA
- Positive historical control data: within the ranges of historical control data
- Negative (solvent/vehicle) historical control data: within the ranges of historical control data. One of the solvent/negative control cultures in experiment II was just above the 95% control limits of the historical control data range. The observed mutation frequency, however, was within the acceptability criteria of this assay.
ADDITIONAL INFORMATION ON CYTOTOXICITY:
- Measurement of cytotoxicity used: No cytotoxicity was observed in experiment I and II.
- Other observations when applicable: In experiment I the number of small and large colonies in the test item treated cultures were comparable to the number of small and large colonies of the solvent controls. In experiment II the test item precipitated in the exposure medium at concentration of 52 µg/ml after 24-hour treatment.
Remarks on result:
other: No mutagenic potential
Remarks:
Experiment I and II

Table 1: Experiment I, 3 -hour exposure without metabolic activation

Test Group

Concentrations

µg/ml

RCE %

RTG %

RSG %

MF

(mutants/106cells)

GEF exceeded

Statistical Significant Increase

SC1

0

100

100

100

79

-

-

SC2

0

100

100

100

124

-

-

1

0.056

102

101

99

93

-

-

2

0.18

104

92

89

97

-

-

3

0.55

111

102

92

106

-

-

4

1.7

134

126

94

95

-

-

5

5.4

105

103

98

51

-

-

6

17

111

100

90

64

-

-

7

52

104

96

93

76

-

-

8

164*

116

104

90

94

-

-

MMS

15 

63

39

62

632

+

+

Table 2: Experiment I, 3 -hour exposure with metabolic activation

Test Group

Concentrations

µg/ml

RCE %

RTG %

RSG %

MF

(mutants/106cells)

GEF exceeded

Statistical Significant Increase

SC1

0

100

100

100

91

-

-

SC2

0

100

100

100

117

-

-

1

0.056

100

90

90

105

-

-

2

0.18

86

75

87

116

-

-

3

0.55

104

97

93

100

-

-

4

1.7

97

83

86

136

-

-

5

5.4

110

82

75

81

-

-

6

17

110

96

87

85

-

-

7

52

94

82

87

97

-

-

8

164*

118

103

87

92

-

-

CP

 7.5

25

7

27

1938

+

+

Table 3: Experiment II, 24 -hour treatment without metabolic activation

Test Group

Concentrations

µg/ml

RCE %

RTG %

RSG %

MF

(mutants/106cells)

GEF exceeded

Statistical Significant Increase

SC1

0

100

100

100

118

-

-

SC2

0

100

100

100

133

-

-

1

0.018

117

109

93

72

-

-

2

0.056

104

95

91

122

-

-

3

0.18

104

93

89

102

-

-

4

0.55

117

111

95

70

-

-

5

1.7

106

107

101

108

-

-

6

5.4

124

120

97

81

-

-

7

17

114

109

96

85

-

-

8

52*

99

87

88

108

-

-

MMS

 15

88

65

74

583

+

+

* the test item precipitated in the exposure medium

RSG: Relative suspension growth

RCE: Relative cloning efficiency

RTG: Relative total growth

SC: solvent control

MMS: methylmethanesulfonate

CP: cyclophosphamine

Conclusions:
1,3,5-Trimethyl-1,1,3,5,5-pentaphenyltrisiloxane has been tested for mutagenicity in mouse lymphoma L5178Y cells, according to OECD 490 and in compliance with GLP. No test substance induced increase in the number of mutations was observed when tested up to precipitating concentration. Appropriate solvent and positive controls were included and gave the expected results. It is concluded that the test substance is negative for mutagenicity to mammalian cells under the conditions of the study.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

1,3,5-Trimethyl-1,1,3,5,5-pentaphenyltrisiloxane has been tested in a valid bacterial reverse mutation assay, according to the OECD TG 471, and in compliance with GLP, using Salmonella typhimurium strains TA 1535, TA 1537, TA 98, TA 100 and TA 102 (LPT, 2002). No increase in the number of revertants was observed in any test strain, with or without metabolic activation, up to limit concentration. Appropriate positive and solvent controls were added and gave expected results. It is concluded that the test substance is negative for mutagenicity to bacteria under the conditions of the test.

1,3,5-Trimethyl-1,1,3,5,5-pentaphenyltrisiloxane has been tested in a valid study for ability to induce chromosome aberrations in peripheral human lymphocytes according to OECD TG 473 (2014), and in compliance with GLP. No statistically significant or biologically relevant increase in the number of cells with chromosome aberrations was observed either with or without metabolic activation when tested up to precipitating concentrations. Appropriate positive and negative controls were used and gave the expected results. It is concluded that the test item is negative for the induction of chromosome aberrations under the conditions of the study.

1,3,5-Trimethyl-1,1,3,5,5-pentaphenyltrisiloxane has been tested for mutagenicity in Mouse lymphoma L5178Y cells, according to OECD 490 and in compliance with GLP (Charles River, 2016). No test substance induced increase in the number of mutations was observed when tested up to precipitating concentration. Appropriate solvent and positive controls were included and gave the expected results. It is concluded that the test substance is negative for mutagenicity to mammalian cells under the conditions of the study.


Justification for classification or non-classification

Based on the available data for 1,3,5 -trimethyl-1,1,3,5,5-pentaphenyltrisiloxane, no classification is required for genetic toxicity according to Regulation (EC) No 1272/2008.