Registration Dossier

Administrative data

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

In a bacterial reverse mutation assay / Ames test, conducted according to OECD 471 and in compliance with GLP, triethoxy(3-thiocyanatopropyl)silane was found to be negative in both the presence and absence of metabolic activation. The tested strains included Salmonella typhimurium: TA98, TA100, TA1535 and TA1537 as well as Escherichia coli WP2 (Evonik Degussa, 2000).

In a cytogenicity study in mammalian cells (chromosome aberration), conducted according to OECD 473 and in compliance with GLP, the potential of triethoxy(3-thiocyanatopropyl)silane to induce structural chromosome aberrations in Chinese hamster V79 cells was evaluated. A dose-related increase in the number of aberrations was observed at all concentrations tested with and without metabolic activation. Appropriate solvent and positive controls were included and gave expected results. It is concluded that the test substance is clastogenic (causes structural aberrations in chromosomes) under the conditions of the test. No evidence of polyploidy was observed (BSL Bioservice, 2012).

In a mutagenicity study, conducted according to OECD 476 and in compliance with GLP, a dose-related increase in the mutant frequency was observed in mouse lymphoma L5178Y cells. Therefore, it is concluded that the test substance is positive with metabolic activation. No clastogenic effects were observed (BSL Bioservice, 2011).

Link to relevant study records

Referenceopen allclose all

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:
2000-05-09 to 2000-05-19
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Version / remarks:
1997
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of assay:
bacterial reverse mutation assay
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
Species / strain / cell type:
E. coli WP2 uvr A
Metabolic activation:
with and without
Metabolic activation system:
Type and composition of metabolic activation system: Male Wistar rats
- source of S9 : RCC (Cytotest Cell Research GmbH), Roßdorf, Germany.
- method of preparation of S9 mix: Wistar male rats received 80 mg/kg bw phenobarbital (i.p) and 80 mg/kg bw beta-naphthoflavone (oral) during 3 consecutive days and were sacrificed 24 h after the last administration. The livers were removed from the animals and homogenised. The S9 mix was prepared freshly before use: one part S9 fraction mixed with 9 parts of a cofactor solution.
- concentration or volume of S9 mix and S9 in the final culture medium: 10%
- quality controls of S9: enzymatic activity, sterility and protein content
Test concentrations with justification for top dose:
50-5000 µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO

- Justification for choice of solvent/vehicle: Not reported
Untreated negative controls:
yes
Remarks:
water
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:
yes
Remarks:
water
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
TA 100 (without activation)
Untreated negative controls:
yes
Remarks:
water
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
mitomycin C
Remarks:
TA 102 (without activation)
Untreated negative controls:
yes
Remarks:
water
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
TA 1535 (without activation)
Untreated negative controls:
yes
Remarks:
water
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:
yes
Remarks:
water
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
methylmethanesulfonate
Remarks:
WP2 uvrA (without activation)
Untreated negative controls:
yes
Remarks:
water
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: 2-Aminoflouorene
Remarks:
All strains (with activation)
Untreated negative controls:
yes
Remarks:
water
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: 2-Aminoanthracene
Remarks:
All strains (with activation)
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation); preincubation

ACTIVATION:

DURATION

- Preincubation period: 30 minutes

- Expression time (cells in growth medium): 72 hours

NUMBER OF REPLICATIONS: 3 plates for each test concentration

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth; background lawn assessment
Evaluation criteria:
For a test compound or a test substance to be considered positive, it must (in two independent experiments) cause at least a doubling in the mean revertants per plate of at least one tester strain. The increase must be accompanied by a dose response towards increasing concentrations of the test article. Single increases in revertant frequencies which are not dose-related and not reproducible in two independent tests are considered non-relevant. However, when increases occur in both tests, this will be taken as an indication of a mutagenic effect.

A reduction in the number of spontaneous revertants, and / or a thinning of the background lawn is indicative of toxicity.
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
Species / strain:
S. typhimurium 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:
valid
Positive controls validity:
valid
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:
TEST-SPECIFIC CONFOUNDING FACTORS

- Precipitation: precipitation noted at 1600 and 5000 µg/plate

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

Table 2: Experiment 1 Plate incorporation Number of revertants per plate

 

TA 98

TA 100

TA 102

Conc.
(
µl/ml)

MA

+

MA

Cytotoxic
(yes/no)

MA

+

MA

Cytotoxic
(yes/no)

MA

+

MA

Cytotoxic
(yes/no)

Negative Control

13

13

No

129

237

No

380

434

No

0*

15

9

No

104

2219

No

329

351

No

50

18

2

No

103

237

No

398

312

No

160

16

7

No

111

233

No

388

308

No

500

16

5

No

130

225

No

350

347

No

1600

22

7

No

135

223

No

345

298

No

5000

13

7

No

138

199

No

349

268

No

Positive control

100

2216

No

587

1637

No

1205

837

No

Positive control 2

-

1606

No

-

1524

No

-

522

No

*solvent control with DMSO

Table 2: Experiment 1 Plate incorporation Number of revertants per plate

 

TA 1535

TA 1537

WP2 uvrA

Conc.
(
µl/ml)

MA

+

MA

Cytotoxic
(yes/no)

MA

+

MA

Cytotoxic
(yes/no)

MA

+

MA

Cytotoxic
(yes/no)

Negative Control

12

15

No

13

16

No

12

12

No

0*

14

8

No

12

15

No

9

11

No

50

13

7

No

14

17

No

11

15

No

160

11

7

No

9

12

No

9

14

No

500

13

9

No

11

16

No

8

12

No

1600

13

10

No

17

11

No

11

14

No

5000

10

10

No

11

9

No

9

12

No

Positive control

576

31

No

43

69

No

185

40

No

Positive control 2

-

142

No

-

116

No

-

20

No

*solvent control with DMSO

Table 3: Experiment 2 Pre incubation Number of revertants per plate

 

TA 98

TA 100

TA 102

Conc.
(
µl/ml)

MA

+

MA

Cytotoxic
(yes/no)

MA

+

MA

Cytotoxic
(yes/no)

MA

+

MA

Cytotoxic
(yes/no)

Negative Control

26

30

No

196

197

No

406

365

No

0*

23

33

No

181

182

No

350

361

No

50

31

34

No

182

185

No

391

360

No

160

30

31

No

195

179

No

361

336

No

500

37

31

No

196

174

No

374

359

No

1600

27

35

No

200

175

No

399

334

No

5000

27

28

No

191

200

No

385

277

No

Positive control

137

1966

No

680

1331

No

1390

1173

No

Positive control 2

-

1113

No

-

1064

No

-

471

No

*solvent control with DMSO

Table 3: Experiment 2 Pre incubation Number of revertants per plate

 

TA 1535

TA 1537

WP2 uvrA

Conc.
(
µl/ml)

MA

+

MA

Cytotoxic
(yes/no)

MA

+

MA

Cytotoxic
(yes/no)

MA

+

MA

Cytotoxic
(yes/no)

Negative Control

15

11

No

6

10

No

13

11

No

0*

16

9

No

9

9

No

10

13

No

50

13

10

No

8

7

No

13

17

No

160

16

11

No

9

15

No

12

18

No

500

11

10

No

6

10

No

14

16

No

1600

12

11

No

11

13

No

8

16

No

5000

12

12

No

9

14

No

12

12

No

Positive control

555

24

No

39

78

No

222

52

No

Positive control 2

-

128

No

-

71

No

-

22

No

*solvent control with DMSO

Conclusions:
Triethoxy(3-thiocyanatopropyl)silane has been tested in a reliable study conducted in accordance with OECD 471 (1997) and in compliance with GLP using Salmonella typhimurium strains TA 98, 100, 1535, 1537 and 1538, and E.coli WP2uvrA. No evidence of test substance induced increase in the number of revertants was observed up to limit concentrations in any of the bacteria tested without and with metabolic activation in either the initial plate incorporation assay or the repeat experiment using the preincubation method. Appropriate untreated, solvent and positive controls were included and gave expected results. In conclusion, the test substance is non-mutagenic under the conditions of the test.
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:
2011-10-19 to 2011-12-29
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.5375 - In vitro Mammalian Chromosome Aberration Test
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.10 (Mutagenicity - In Vitro Mammalian Chromosome Aberration Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
Bayerisches Landesamt für Gesundheit und Lebensmittelsicherheit, München, Germany
Type of assay:
in vitro mammalian chromosome aberration test
Species / strain / cell type:
Chinese hamster lung fibroblasts (V79)
Details on mammalian cell type (if applicable):
- Type and identity of media: MEM
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: yes
Metabolic activation:
with and without
Metabolic activation system:
Phenobarbital and ß-naphthoflavone-induced rat liver S9 mix containing 8 mM MgCl2; 22 mM KCl 5 mM Glucose-6-phosphate and 4 mM NADP
Test concentrations with justification for top dose:
Pre-experiment:
with and without metabolic activation:
0.020, 0.039, 0.078, 0.16, 0.31, 0.63, 1.25, 2.50, 5.0 and 10.0 mM

Main Experiment :
without metabolic activation:
0.40, 0.60 and 0.80 mM

with metabolic activation:
0.15, 0.30 and 0.50 mM
Vehicle / solvent:
-Vehicle (s)/solvent(s) used: DMSO (final concentration of 1% solvent v/v)
- Justification for choice of solvent/vehicle: The test item could be dissolved in DMSO and was compatible with the survival of the cells.
Untreated negative controls:
yes
Remarks:
900 µg/mL
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
ethylmethanesulphonate
Remarks:
without metabolic activation
Untreated negative controls:
yes
Remarks:
0.83 µg/mL
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
cyclophosphamide
Remarks:
with metabolic activation
Details on test system and experimental conditions:
TREATMENT TIME:
4 hours (Main Experiment with and without metabolic activation)

FIXATION INTERVAL: 20 hours (Main Experiment with and without metabolic activation)
NUMBER OF REPLICATIONS: 1 experiment
NUMBER OF CELLS SEEDED: 1 x 10^4 - 5 x 10^4 cells
NUMBER OF CULTURES: two cultures per concentration
NUMBER OF CELLS SCORED: 200 cells per concentration (100 cells per culture)
DETERMINATION OF CYTOTOXICITY: Mitotic index, cell density
Evaluation criteria:
There are several criteria for determining a positive result:
- a clear and dose-related increase in the number of cells with aberrations,
- a biologically relevant response for at least one of the dose groups, which is higher than the laboratory negative control range (0.0% - 4.0% aberrant cells (with and without metabolic activation)).


Statistics:
According to OECD guidelines, the biological relevance of the results is the criterion for the interpretation of results, a statistical evaluation of the results is not regarded as necessary.
Species / strain:
Chinese hamster lung fibroblasts (V79)
Metabolic activation:
with and without
Genotoxicity:
positive
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
0.6 mM (without metabolic activation)
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid

Results of chromosome analysis                         without metabolic activation                          
     Cytotoxicity Chromatid aberrations         Isochromatid aberrations       rel. Mitotic index (%) rel. Cell density (%)  Poly-ploidy mean % aberrant cells
Scored cells  gaps breaks  inter-changes  other  gaps breaks  inter-changes  other incl. Gaps excl. Gaps
Main Experiment 
negative control 200 - 2 3 0 0 0 0 0 0 87 100 1 2.5 1.5
solvent control 200 - 2 0 0 0 0 0 0 0 100 100 1 1.0 0.0
0.02 mM - no n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. 101 n.d. n.d. n.d. n.d.
0.03 mM - no n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. 95 n.d. n.d. n.d. n.d.
0.06 mM - no n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. 97 n.d. n.d. n.d. n.d.
0.08 mM - no n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. 85 n.d. n.d. n.d. n.d.
0.10 mM - no n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. 91 n.d. n.d. n.d. n.d.
0.20 mM - no n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. 80 n.d. n.d. n.d. n.d.
0.40 mM 200 no 5 12 4 2 1 0 0 0 73 89 2 10.0 7.5
0.60 mM 200 yes 7 22 1 1 1 0 0 1 48 77 1 11.5 8.5
0.80 mM 200 yes 6 28 4 1 0 1 0 0 32 48 0 13.5 12.0
EMS 900 µg/mL 200 - 7 6 8 3 1 0 6 0 105 84 1 12.5 9.5
Results of chromosome analysis
with metabolic activation
     Cytotoxicity Chromatid aberrations         Isochromatid aberrations       rel. Mitotic index (%) rel. Cell density (%)  Poly-ploidy mean % aberrant cells
Scored cells  gaps breaks  inter-changes  other  gaps breaks  inter-changes  other incl. Gaps excl. Gaps
Main Experiment 
negative control 200 - 3 3 2 0 0 0 1 0 101 101 1 4.5 3.0
solvent control 200 - 6 4 0 0 1 0 0 0 100 100 1 5.0 2.0
0.02 mM - no n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. 98 n.d. n.d. n.d. n.d.
0.05 mM - no n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. 96 n.d. n.d. n.d. n.d.
0.08 mM - no n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. 89 n.d. n.d. n.d. n.d.
0.15 mM 200 no 8 10 0 0 0 0 0 2 86 95 0 8.5 6.0
0.30 mM 200 no 3 15 3 1 0 1 0 2 87 94 2 10.0 9.0
0.50 mM 200 no 4 31 6 1 0 0 0 1 83 82 0 15.0 14.5
0.75 mM - yes n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. 63 n.d. n.d. n.d. n.d.
1.00 mM - yes n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. 45 n.d. n.d. n.d. n.d.
1.25 mM - yes n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. 12 n.d. n.d. n.d. n.d.
1.50 mM - yes n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. 28 n.d. n.d. n.d. n.d.
CPA 0.83 µg/mL 200 - 7 13 6 0 0 1 0 1 106 99 2 11.0 9.5

n.d. not determined

Conclusions:
Triethoxy(3-thiocyanatopropyl)silane has been tested in a reliable in vitro chromosomal aberration test conducted according to OECD 473 and in compliance with GLP. A dose-related increase in the number of aberrations was observed at all concentrations tested with and without metabolic activation. Appropriate solvent and positive controls were included and gave expected results. It is concluded that the test substance is clastogenic (causes structural aberrations in chromosomes) under the conditions of the test. No evidence of polyploidy was observed.
Executive summary:

To investigate the potential of triethoxy(3-thiocyanatopropyl)silane to induce structural chromosome aberrations in Chinese hamster V79 cells, an in vitro chromosome aberration assay was carried out.

The chromosomes were prepared 20 h after start of treatment with the test item. The treatment interval was 4 h with and without metabolic activation in the main experiment. Duplicate cultures were treated at each concentration. 100 metaphases per culture were scored for structural chromosomal aberrations.

The test item could be dissolved in DMSO and was compatible with the survival of the cells.

The following concentrations were evaluated for the microscopic analysis of chromosomal aberrations:

without metabolic activation: 0.40, 0.60 and 0.80 mM

with metabolic activation: 0.15, 0.30 and 0.50 mM

In the main experiment precipitation of the test item was noted after incubation with the test item without metabolic activation at a concentration of 0.80 mM, with metabolic activation no precipitation was observed.

Toxic effects of the test item were observed in the main experiment without metabolic activation at concentrations of 0.60 mM and higher, with metabolic activation no toxic effects were noted at the concentrations evaluated.

A clear increase of aberrant cells was found in the main experiment with and without metabolic activation at all concentrations evaluated. In addition, a dose response relationship was observed.

In the main experiment with and without metabolic activation no biologically relevant increase in the frequencies of polyploid cells was observed after treatment with the test item as compared to the controls.

EMS (900 µg/mL) and CPA (0.83 µg/mL) were used as positive controls and induced distinct and biologically relevant increases in cells with structural chromosomal aberrations.

The positive controls induced the appropriate responses.

There was a concentration-related positive response of test item induced over background.

This study is classified as acceptable. This study satisfies the requirement for Test Guideline OPPTS 870.5375; OECD 473 for in vitro cytogenetic mutagenicity data. 

Endpoint:
in vitro gene mutation study in mammalian cells
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2011-09-26 to 2011-11-15
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.17 (Mutagenicity - In Vitro Mammalian Cell Gene Mutation Test)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.5300 - In vitro Mammalian Cell Gene Mutation Test
Deviations:
no
Qualifier:
according to
Guideline:
other: IWGT Recommendations
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
(Bayerisches Landesamt für Gesundheit und Lebensmittelsicherheit, Germany)
Type of assay:
mammalian cell gene mutation assay
Target gene:
Thymidine kinase
Species / strain / cell type:
mouse lymphoma L5178Y cells
Details on mammalian cell type (if applicable):
- Type and identity of media: RPMI
- Properly maintained: Yes
- Periodically checked for Mycoplasma contamination: yes
- Periodically "cleansed" against high spontaneous background: yes
Metabolic activation:
with and without
Metabolic activation system:
Phenobarbital and ß-Naphthoflavone-induced rat liver S9 mix containing 8 mM MgCl2; 33 mM KCl; 5 mM Glucose-6-phosphate and 4 mM NADP.
Test concentrations with justification for top dose:
Pre-experiment with [0.2, 0.5 and 2.5 mM] and without metabolic activation [0.2, 0.5, 1 and 2 mM]

Main Experiment
with metabolic activation: 0.05, 0.1, 0.2, 0.5, 0.7, 0.9, 1.1 and 1.3 mM
without metabolic activation: 0.005, 0.01, 0.02, 0.05, 0.1, 0.2, 0.5 and 1 mM





Vehicle / solvent:
Based on the solubility test DMSO was used as solvent (final concentration 1% DMSO v/v).
Untreated negative controls:
yes
Remarks:
2.5 µg/ml
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
benzo(a)pyrene
Untreated negative controls:
other: 300 µg/ml
Positive control substance:
ethylmethanesulphonate
Untreated negative controls:
other: 10 µg/ml
Positive control substance:
methylmethanesulfonate
Details on test system and experimental conditions:
METHOD OF APPLICATION: suspended in medium
DURATION: 4 h (short-term exposure)
Expression time (cells in growth medium): 48 h
Selection time (if incubation with selection agent): about 14 days

SELECTION AGENT ( mutation assay) 5 µg/ml trifluorothymidine
NUMBER OF REPLICATIONS: one experiment with single exposure; cells were seeded in 4 plates and evaluated
NUMBER OF CELLS SEEDED: 2000 cells per well
DETERMINATION OF CYTOTOXICITY: relative total growth (RTG)
Evaluation criteria:
The test item is considered mutagenic if following criteria are met:
-The induced mutant frequency meets or exceeds the Global Evaluation factor (GEF) of 126 per 106 cells
- A dose-dependent increase in mutant frequency is detected.

Besides, combined with a positive effect in the mutant frequency, an increased occurrence of small colonies (≥40% of total colonies) is an indication for potential clastogenic effects and/or chromosomal aberrations.
According to the OECD guideline, the biological relevance is considered first for the interpretation of results. Statistical methods might be used as an aid in evaluation the test result.
A test item is considered to be negative if the induced mutant frequency is below the GEF and the trend test is negative.

Statistics:
The non-parametric Mann-Whitney test is applied to the mutation data to prove the dose groups for any significant difference in mutant frequency compared to the negative /solvent controls.
Species / strain:
mouse lymphoma L5178Y cells
Metabolic activation:
with
Genotoxicity:
positive
Cytotoxicity / choice of top concentrations:
cytotoxicity
Untreated negative controls validity:
valid
Positive controls validity:
valid

Summary of mutagenicity data with and without metabolic activation

1 Without metabolic activation

Concentration (mM)

RCE %

RTG %

MF (mutants /10x8 cells)

IMF (mutants /10x8 cells)

GEF exceeded

Statistical significance

Negative control 1

103.2

122.9

69.4

/

/

/

Negative control 2

100.4

121.0

/

/

/

Solvent control 1

100.0

100.0

66.4

/

/

/

Solvent control 2

/

/

/

0.005

101.1

110.1

67.1

0.8

-

-

0.01

99.6

104.1

69.1

2.7

-

-

0.02

109.5

119.3

50.8

-15.5

-

-

0.05

103.2

108.4

85.9

19.5

-

-

0.1

112.3

61.3

74.7

8.3

-

-

0.2

92.6

35.8

93.0

26.7

-

-

0.5

96.8

26.1

113.1

46.7

-

+

1

96.8

13.1

143.3

76.9

-

+

Positive control 1

94.0

99.6

622.7

556.3

+

+

Positive control 2

92.6

88.1

456.8

390.5

+

+

 

2 With metabolic activation

Concentration (mM)

RCE %

RTG %

MF (mutants /10x8 cells)

IMF (mutants /10x8 cells)

GEF exceeded

Statistical significance

Negative control 1

100.0

93.4

88.0

/

/

/

Negative control 2

100.0

106.2

/

/

/

Solvent control 1

100.0

100.0

84.6

/

/

/

Solvent control 2

/

/

/

0.05

103.5

97.3

72.7

-11.9

-

-

0.1

90.2

91.3

107.2

22.6

-

-

0.2

100.7

65.2

127.5

42.9

-

+

0.5

93.7

34.8

161.0

76.4

-

+

0.7

101.4

35.8

130.0

45.5

-

+

0.9

95.8

30.1

186.3

101.8

-

+

1.1

95.8

22.5

226.7

142.1

+

+

1.3

86.0

9.4

236.2

151.6

+

+

Positive control 1

82.5

44.2

817.8

733.3

+

+

 

RCE = Relative cloning efficiency    GEF = Global evaluation factor

RTG = Relative total growth

MF = Mutant frequency                     IMF = Induced mutant frequency

Conclusions:
Triethoxy(3-thiocyanatopropyl)silane has been tested for mutagenicity to mammalian cells in a reliable study conducted according to OECD TG 476 and in compliance with GLP using L5178Y cells. A dose-related increase in mutant frequency above the global evaluation factor was observed at concentrations of 1mM and above in the main experiment with metabolic activation. Appropriate solvent and positive controls were included and gave expected results. It is therefore concluded that the test substance is mutagenic in the in vitro mammalian cell gene mutation assay (thymidine kinase locus) in mouse lymphoma L5178Y cells. No clastogenic effects (increase in number of small colonies) were observed in this study.
Executive summary:

The test item triethoxy(3-thiocyanatopropyl)silane was assessed for its potential to induce mutations at the mouse lymphoma thymidine kinase locus using the cell line L5178Y.

The selection of the concentrations used in the main experiment was based on data from the pre-experiment. In the main experiment 1.3 mM (with metabolic activation) and 1 mM (without metabolic activation) were selected as the highest concentrations. The experiment with and without metabolic activation was performed as a 4 h short-term exposure assay. Based on the solubility test DMSO was used as solvent (final concentration 1% DMSO v/v).

The test item was investigated at the following concentrations:

with metabolic activation:

0.05, 0.1, 0.2, 0.5, 0.7, 0.9, 1.1 and 1.3 mM

and without metabolic activation:

0.005, 0.01, 0.02, 0.05, 0.1, 0.2, 0.5 and 1 mM

Precipitation of the test item was noted in the pre-experiment at a concentration of 2.5 mM. Growth inhibition was observed in the main experiment with and without metabolic activation.

In the main experiment with metabolic activation the relative total growth (RTG) was 9.4% for the highest concentration (1.3 mM) evaluated. The highest concentration evaluated without metabolic activation was 1 mM with an RTG of 13.1%. In the main experiment a biologically relevant increase of mutants was found after treatment with the test item with metabolic activation, without metabolic activation no biologically relevant increase of mutants was found after treatment with the test item. The Global Evaluation Factor (GEF; defined as the mean of the negative/vehicle mutant frequency plus one standard deviation; data gathered from ten laboratories) was exceeded by the induced mutant frequency at a concentration of 1.1 mM and higher in the main experiment with metabolic activation.

A dose-response relationship was observed.

However, in the main experiment colony sizing showed no clastogenic effects induced by the test item under the experimental conditions (with and without metabolic activation).

EMS, MMS and B[a]P were used as positive controls and showed distinct and biologically relevant effects in mutation frequency. Additionally, MMS and B[a]P significantly increased the number of small colonies, thus proving the efficiency of the test system to indicate potential clastogenic effects.

This study is classified as acceptable. This study satisfies the requirements for Test Guidelines OPPTS 870.5300, OECD 476 for in vitro mutagenicity (mammalian forward gene mutation) data.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (positive)

Genetic toxicity in vivo

Description of key information

Based on the positive response in the in vitro studies, an in vivo comet assay is being carried out. When available, the sections herein will be updated accordingly.

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Triethoxy(3-thiocyanatopropyl)silane has been tested in a reliable study conducted in accordance with OECD 471 (1997) and in compliance with GLP using Salmonella typhimurium strains TA 98, 100, 1535, 1537 and 1538, and E.coli WP2uvrA (Evonik Degussa, 2000). No evidence of test substance induced increase in the number of revertants was observed up to limit concentrations in any of the bacteria tested with and without metabolic activation in either the initial plate incorporation assay or the repeat experiment using the preincubation method. Appropriate untreated, solvent and positive controls were included and gave expected results. It is concluded that the test substance is non-mutagenic under the conditions of the test.

Information on the potential for triethoxy(3-thiocyanatopropyl)silane to cause chromosome aberrations in mammalian cells is available from a reliable study conducted according to OECD 473 and in compliance with GLP (BSL Bioservice, 2012). A dose-related increase in the number of aberrations was observed at all concentrations tested with and without metabolic activation. Appropriate solvent and positive controls were included and gave expected results. It is concluded that the test substance is clastogenic (causes structural aberrations in chromosomes) under the conditions of the test. No evidence of polyploidy was observed.

Triethoxy(3-thiocyanatopropyl)silane has been tested for mutagenicity to mammalian cells in a reliable study conducted according to OECD 476 and in compliance with GLP using L5178Y cells (BSL Bioservice, 2011). A dose-related increase in mutant frequency above the global evaluation factor was observed at concentrations of 1mM and above in the main experiment with metabolic activation. Appropriate solvent and positive controls were included and gave expected results. It is therefore concluded that the test substance is mutagenic in the in vitro mammalian cell gene mutation assay (thymidine kinase locus) in mouse lymphoma L5178Y cells. No clastogenic effects (increase in number of small colonies) were observed in this study.

Evidence of clastogenicity and mutagenicity to mammalian cells was obtained from the in vitro cytogenicity and mutagenicity studies using mammalian cells. Therefore, an in vivo comet assay is being conducted to evaluate the potential in vivo.


Justification for classification or non-classification