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Registration Dossier
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EC number: 213-668-5 | CAS number: 999-97-3
Endpoint summary
Administrative data
Key value for chemical safety assessment
Genetic toxicity in vitro
Description of key information
Gene mutation (Bacterial reverse mutation assay / Ames test): negative
with and without activation in in S. typhimurium strains TA 98, TA 100,
TA 1535, TA 1537 and TA 102 (OECD Test Guideline 471) (Covance, 1997a).
Cytogenicity in mammalian cells: negative with and without metabolic
activation in cultured human lymphocytes (OECD Test Guideline 473)
(Covance 1997b).
Mutagenicity in mammalian cells: negative with and without metabolic activation in L5178Y mouse lymphoma cells (similar to OECD Test Guideline 476) (Microbiological Associates, 1991).
Link to relevant study records
- Endpoint:
- in vitro gene mutation study in bacteria
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 1997-03-21 to 1997-05-12
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- guideline study with acceptable restrictions
- Remarks:
- The study was conducted according to an appropriate OECD test guideline and was compliant with GLP, however testing with positive controls was not fully undertaken, strains TA98 and 100 only were tested with positive controls.
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 471 (Bacterial Reverse Mutation Assay)
- Deviations:
- yes
- Remarks:
- incomplete testing with positive controls with activation
- GLP compliance:
- yes
- Type of assay:
- bacterial reverse mutation assay
- Species / strain / cell type:
- S. typhimurium, other: TA98, TA100, TA1535, TA1537, TA102
- Additional strain / cell type characteristics:
- other: histidine deficient
- Metabolic activation:
- with and without
- Metabolic activation system:
- Aroclor 1254-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: not given - Untreated negative controls:
- no
- Negative solvent / vehicle controls:
- yes
- Remarks:
- with DMSO
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- 2-nitrofluorene
- Remarks:
- strain TA98 without MA: 5.0 ug/plate
- Untreated negative controls:
- no
- Negative solvent / vehicle controls:
- yes
- Remarks:
- DMSO
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- sodium azide
- Remarks:
- with strains TA100 and TA1535 without MA: 2.0 ug/plate
- Untreated negative controls:
- no
- Negative solvent / vehicle controls:
- yes
- Remarks:
- DMSO
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- 9-aminoacridine
- Remarks:
- with strain TA1537 without MA: 50 ug/plate
- Untreated negative controls:
- no
- Negative solvent / vehicle controls:
- yes
- Remarks:
- DSMO
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- other: glutaraldehyde (25.0 ug/plate)
- Remarks:
- with strain TA102 without MA
- Untreated negative controls:
- no
- Negative solvent / vehicle controls:
- yes
- Remarks:
- DMSO
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- other: 2-aminoanthracene
- Remarks:
- strains TA 100 and TA98
- Untreated negative controls:
- no
- Negative solvent / vehicle controls:
- yes
- Remarks:
- DMSO
- True negative controls:
- no
- Positive controls:
- no
- Remarks:
- strains TA1535, TA 1537 and TA102 were not tested with a positive control in the presence of MA.
- Details on test system and experimental conditions:
- METHOD OF APPLICATION: Experiment 1: in agar (plate incorporation); Experiment 2: preincubation
DURATION
- Preincubation period: 1 hour at 37 degC
- Exposure duration: 3 days
NUMBER OF REPLICATIONS: 3 per exposure, 5 with solvent control.
ACTIVATION: S9 mix included 10% Arocolor-induced rat liver S9, and glucose-6-phosphate, NADP, MgCl2, KCl. 0.5 ml was added with 0.1 ml bacterial suspension and 0.1 ml test or control substance to 2.5 ml of agar to give a final concentration of approximately 1.5% S9.
DETERMINATION OF CYTOTOXICITY
- Method: toxicity to background lawn. - Evaluation criteria:
- The test article was considered to be mutagenic if a dose related and reproducible increase in the number of revertants was induced at one or more test concentration. An increase in revertant colonies was at least two times the mean negative control counts in strains TA98, TA100 and TA102, or three times in strains TA1535 and TA1537.
- Statistics:
- Colonies were counted electronically using a Seescan Colony Counter (Seescan plc) and the background lawn inspected for signs of toxicity.
- Key result
- Species / strain:
- S. typhimurium TA 1535
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- not examined
- Positive controls validity:
- valid
- Key result
- Species / strain:
- S. typhimurium TA 100
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- not examined
- Positive controls validity:
- valid
- Key result
- Species / strain:
- S. typhimurium TA 1537
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- not examined
- Positive controls validity:
- valid
- Key result
- Species / strain:
- S. typhimurium TA 98
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- not examined
- Positive controls validity:
- valid
- Key result
- Species / strain:
- S. typhimurium TA 102
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Remarks:
- at 2500 μg/plate with MA
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- not examined
- Positive controls validity:
- valid
- Additional information on results:
- TEST-SPECIFIC CONFOUNDING FACTORS
- Other confounding effects: in water the test substance hydrolyses into ammonia.
RANGE-FINDING/SCREENING STUDIES: 8, 40, 200, 1000 and 5000 μg/plate with TA100 without S9 resulted in no evidence of toxicity (results with S9 were not scorable due to contamination). The results were used in the mutagenic assessment and are presented in Table 2.
COMPARISON WITH HISTORICAL CONTROL DATA: Following Experiment 1 treatments of strains TA98 and TA102 in the absence and presence of S-9, Experiment 2 treatments of strain TA98 in the absence and presence of S9 and strain TA100 in the absence of S9, mean revertant colony plate counts for the solvent control treatments were just outside the historical range of this laboratory. The mean solvent control plate counts were however well within other published ranges, and so these strains were considered to have been behaving characteristically, and these data were considered valid.
ADDITIONAL INFORMATION ON CYTOTOXICITY: In Experiment 1 a slight thinning of the background lawn was observed at the highest concentrations with strain TA1537 with S9. In Experiment 2 cytotoxicity was observed in the form of complete mortality of the bacteria at the maximum concentration of 2500 μg/plate with strain TA102 with S9. - Remarks on result:
- other: all strains/cell types tested
- Conclusions:
- 1,1,1,3,3,3-Hexamethyldisilazane (CAS 999-97-3, EC 213-668-5) has been tested according to OECD Test Guideline 471 and in compliance with GLP. No test substance induced increase in the number of revertants in S. typhimurium strains TA 98, TA100, TA 1535, TA 1537 and TA 102 was observed. The results obtained with and without metabolic activation in the initial plate incorporation test and in the repeat pre-incubation assay were in agreement. Appropriate solvent and positive controls were included and gave expected results. The test material was therefore considered to be negative for mutagenicity to bacteria under the conditions of this test.
- Endpoint:
- in vitro cytogenicity / chromosome aberration study in mammalian cells
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 1997-06-18 to 1997-11-28
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Remarks:
- The study was conducted according to the appropriate OECD test guideline, and in compliance with GLP.
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
- Deviations:
- yes
- Remarks:
- but minor, see below
- Principles of method if other than guideline:
- The minor deviations from the guideline were as follows:
(1) Concentration of dosing solutions: a slight variation in concentration between proposed and achieved concentrations (22.34 µg/ml) was apparent following formulation and dilution.
(2) Colchicine treatment: cultures receiving the two highest doses of the test article and all doses levels of NQO in the absence of S9 and scheduled to be sampled at 20 hours were sampled at 21.5 hours. The mistake had no bearing on the study outcome.
(3) Selection of doses: two doses were analysed at the 44+0 hour time point to further investigate the induction of aberrations.
(4) Untreated controls: negative controls were included to be analysed if the frequency of cells with aberrations in solvent control cultures was unusually high (an effect which can sometimes be seen following the use of acetone as a solvent). - GLP compliance:
- yes
- Type of assay:
- in vitro mammalian chromosome aberration test
- Species / strain / cell type:
- lymphocytes: from female human donor
- Details on mammalian cell type (if applicable):
- - Type and identity of media: 0.4 ml heparinised blood into 9.0 ml Hepes-buffered RPMI medium containing 30% (v/v) foetal calf serum and 50 µg/ml gentamycin.
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: no, because fresh blood
- Periodically checked for karyotype stability: no, because fresh blood
- Periodically "cleansed" against high spontaneous background: no, because fresh blood - Additional strain / cell type characteristics:
- not applicable
- Metabolic activation:
- with and without
- Metabolic activation system:
- Aroclor 1254 induced rat liver enzymes (S9 mix)
- Test concentrations with justification for top dose:
- 0-1614 µg/ml
- Vehicle / solvent:
- - Vehicle(s)/solvent(s) used: acetone
- Justification for choice of solvent/vehicle: a preliminary test showed that the test article was miscible with acetone. - Untreated negative controls:
- yes
- Remarks:
- untreated
- Negative solvent / vehicle controls:
- yes
- Remarks:
- acetone
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- 4-nitroquinoline-N-oxide
- Remarks:
- without metabolic activation Migrated to IUCLID6: 2.5 ug/ml
- Untreated negative controls:
- yes
- Remarks:
- untreated
- Negative solvent / vehicle controls:
- yes
- Remarks:
- acetone
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- cyclophosphamide
- Remarks:
- with metabolic activation Migrated to IUCLID6: 25 ug/ml
- Details on test system and experimental conditions:
- ACTIVATION: S9 mix included 2 parts Aroclor induced rat liver S9 to one part each of glucose-6-phosphate (180 mg/l), NADP (25 mg/l) and 150 mM KCl. The final concentration of liver homogenate in the test system was 2%.
METHOD OF APPLICATION: in suspension
DURATION
- Exposure duration: see Table 2.
- Expression time (cells in growth medium): see Table 2.
- Fixation time (start of exposure up to fixation or harvest of cells): same as expression time
SPINDLE INHIBITOR (cytogenetic assays): colchicine (1 µg/ml)
STAIN (for cytogenetic assays): Giemsa stain
NUMBER OF REPLICATIONS: 4 for negative control, and 2 for the rest. Two solvent controls were initially to be analysed for chromosome aberrations; slides from the remaining solvent control cultures were only to be analysed if considered necessary, for example to resolve an equivocal result. The untreated controls were only to be included in the analysis if the frequency of cells with aberrations in the solvent control cultures was unusually high. A single positive control dose level, which gave satisfactory responses in terms of quality and quantity of mitosis and extent of chromosomal damage was to be analysed.
NUMBER OF CELLS EVALUATED: >160 for each treatment
DETERMINATION OF CYTOTOXICITY
- Method: mitotic index
OTHER EXAMINATIONS:
- Determination of polyploidy: yes, scored separately
- Determination of endoreplication: yes, scored separately
- Other: hyperdiploid cells - Evaluation criteria:
- The test substance was considered positive if: a statistical significant increase in the proportion of cells with structural aberrations (excluding gaps) occurred at one or more concentrations and the proportion of cells with structural aberrations at such doses exceeded the normal range. Cells with exchange aberrations or cells with greater than one aberration were to be considered of particular biological significance.
- Statistics:
- Slides were scored manually. The total of cells with structural aberrations excluding gaps were used to determine whether the assay was acceptable or not. The proportions of cells in each replicate were used to establish acceptable heterogeneity between replicates by means of a binomial dispersion test. Fisher's exact test was used to compare negative controls and cells with structural aberrations excluding gaps, probability values of p < or = 0.05 were accepted as significant.
- Key result
- Species / strain:
- other: human lymphocytes
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Additional information on results:
- TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of osmolality: in the range finding test effects of osmolality were excluded from affecting the outcome of the test.
- Water solubility: in water the test article hydrolyses into ammoniac
RANGE-FINDING/SCREENING STUDIES:
COMPARISON WITH HISTORICAL CONTROL DATA: The proportions of cells (cells with structural aberrations including gaps, cells with structural aberrations excluding gaps and polyploid, endoreduplicated or hyperdiploid cells) were examined in relation to historical negative control (normal ranges). Frequencies of cells with numerical aberrations fell within historical negative control ranges under most treatment conditions. The number of aberrant cells fell outside the normal range in a single culture at each of the two concentrations analysed following treatment for 44 hours without MA, but because the increase was small and not seen in replicate culture, the effect was considered to be of no biological significance.
ADDITIONAL INFORMATION ON CYTOTOXICITY: Mitotic inhibition occurred as the following at the highest analysed dose:
20+0 hours -MA: 271.3, 387.5, 553.6 µg/ml = 51% mitotic inhibition
3+17 hours _MA: 790.9, 1130, 1614 µg/ml = 28% mitotic inhibition
44+0 hours -MA: 387.5, 553.6 µg/ml = 52% mitotic inhibition
3+41 hours +Ma: 1614 µg/ml = 9% mitotic inhibition - Remarks on result:
- other: all strains/cell types tested
- Conclusions:
- 1,1,1,3,3,3-Hexamethyldisilazane (CAS 999-97-3, EC 213-668-5) was tested in human lymphocyte cells according to OECD Test Guideline 473 and in compliance with GLP, and did not induce chromosome aberrations. The test material was negative under the conditions of the test.
- Endpoint:
- in vitro gene mutation study in mammalian cells
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 1991
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- guideline study without detailed documentation
- Remarks:
- Guideline study; GLP; limited information re: plating and use of positive/negative controls
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)
- GLP compliance:
- yes
- Type of assay:
- mammalian cell gene mutation assay
- Species / strain / cell type:
- mouse lymphoma L5178Y cells
- Metabolic activation:
- with and without
- Metabolic activation system:
- Aroclor induced rat lived S9
- Test concentrations with justification for top dose:
- Trial 1: 3.0 ul/ml to 10.0 µl/ml (non-activated); 0.5 µl/ml to 5.0 µl/ml (activated). Trial 2: 6.9 µl/ml to 9.0 µl/ml (non-activated); 3.9 µl/ml to 5.1 µl/ml (activated). Trial 3: 0.8 µl/ml to 5.0 µl/ml (non-activated and activated).
- Vehicle / solvent:
- - Vehicle(s)/solvent(s) used: Trial 1 and 2: DMSO; Trial 3: ethanol.
- Justification for choice of solvent/vehicle: None given in study report. It was reported that the test substance was more miscible in ethanol than in DMSO. - Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- 3-methylcholanthrene
- Remarks:
- with activation
- Untreated negative controls:
- yes
- Remarks:
- untreated
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- ethylmethanesulphonate
- Remarks:
- without activation
- Details on test system and experimental conditions:
- METHOD OF APPLICATION: in medium; in agar (plate incorporation); preincubation; in suspension; as impregnation on paper disk
DURATION
- Preincubation period: none
- Exposure duration: 48 hours
- Expression time (cells in growth medium): 48 hours
- Fixation time (start of exposure up to fixation or harvest of cells): 48 hours
SELECTION AGENT (mutation assays): not indicated in study report
NUMBER OF REPLICATIONS: Two
DETERMINATION OF CYTOTOXICITY
- Method: relative total growth
OTHER EXAMINATIONS:
- Other: Colony size distribution was recorded and reported as number of mutant colonies per million surviving cells per colony size.
OTHER:
Trial 1: an unacceptably high solvent mutation frequency in the presence of metabolic activation was reported, so a second trial was carried out. A second toxicity test was run before the third trial. Trial 3 was carried out to use a new sample and change the solvent and the order of addition for dosing to achieve a better consistency of data. - Evaluation criteria:
- No evaluation criteria described in study report.
- Statistics:
- The number of revertant colonies was counted and the size distribution of colonies was recorded.
- Key result
- Species / strain:
- mouse lymphoma L5178Y cells
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- other: Trial 1 with test substance in DMSO -complete toxicity at 9 and 10 µl/ml (non-activated) and at 5 µl/ml (activated).|Trial 3 with test substance in ethanol- complete toxicity at 5.0 µl/ml (non-activated and activated).
- Additional information on results:
- Other confounding effects: the test article's reactivity with water prevented the testing of higher concentrations
RANGE-FINDING/SCREENING STUDIES: See table below
COMPARISON WITH HISTORICAL CONTROL DATA: Not presented in study report
- Conclusions:
- 1,1,1,3,3,3-Hexamethyldisilazane (CAS 999-97-3, EC 213-668-5) has been tested in a mammalian mutagenicity study conducted according to OECD Test Guideline 476 and in compliance with GLP. No test substance related increase in mutant frequency relative to solvent control was observed in any of the three trials conducted in this study using L5178Y TK+/- cells in both the presence and absence of metabolic activation. In addition, there was no evidence of an increase in small colonies when treated cultures were compared to solvent cultures. It is concluded that the test substance is neither mutagenic nor clastogenic under the conditions of the test.
Referenceopen allclose all
Table 2a. Experiment 1 -Mutagenicity Assay. Number of revertants per plate (average).
Positive control was 2-nitrofluorene without metabolic activation and 2-aminoanthracene with metabolic activation.
Strain |
TA 98 |
||
Conc. (μg/plate) |
- MA |
+ MA |
Cytotoxic (yes/no) |
0* |
34.0 |
50.2 |
no |
8 |
49.7 |
43.3 |
no |
40 |
43.0 |
51.7 |
no |
200 |
46.7 |
47.7 |
no |
1000 |
50.0 |
49.0 |
no |
5000 |
44.0 |
47.3 |
no |
Positive control |
766.3 |
1774.3 |
no |
* = solvent control with DMSO
Table 2b. Experiment 1 -Mutagenicity Assay. Number of revertants per plate (average).
Positive control was sodium azide for strains without metabolic activation, the positive control with metabolic activation was 2-aminoanthracene with strain TA100 and none for strain TA1535
Strain |
TA 100 |
TA 1535 |
||||
Conc.(μg/plate) |
-MA |
+MA |
Cytotoxic (yes/no) |
-MA |
+MA |
Cytotoxic (yes/no) |
0* |
115.0 |
145.4 |
no |
17.6 |
23.8 |
no |
80 |
126.0 |
162.3 |
no |
28.0 |
26.3 |
no |
40 |
106.7 |
161.3 |
no |
26.0 |
26.7 |
no |
200 |
116.7 |
162.3 |
no |
21.7 |
26.7 |
no |
1000 |
109.7 |
183.7 |
no |
19.7 |
25.7 |
no |
5000 |
109.0 |
165.7 |
no |
22.3 |
24.3 |
no |
Positive control |
593.0 |
1448.0 |
no |
387.0 |
- |
no |
* = solvent control with DMSO
Table 2c. Experiment 1 - Mutagenicity Assay. Number of revertants per plate (average).
Positive control was 9-aminoacridine without metabolic activation, and no positive control with metabolic activation.
Strain |
TA 1537 |
||
Conc. (μg/plate) |
- MA |
+ MA |
Cytotoxic (yes/no) |
0* |
9.4 |
12.6 |
no |
80 |
16.7 |
14.7 |
no |
40 |
15.0 |
6.3 |
no |
200 |
11.3 |
16.3 |
no |
1000 |
12.3 |
15.0 |
no |
5000 |
15.7 |
14.3 |
no |
Positive control |
419.0 |
- |
no |
* = solvent control with DMSO
Table 2d. Experiment 1 - Mutagenicity Assay. Number of revertantsper plate (average).
Positive contrrol was glutaraldehyde without metabolic activation, and there was no positive control with metabolic activation.
Strain |
TA 102 |
||
Conc. (μg/plate) |
- MA |
+ MA |
Cytotoxic (yes/no) |
0* |
404.8 |
454.8 |
no |
80 |
429.3 |
506.0 |
no |
40 |
452.0 |
444.3 |
no |
200 |
430.0 |
475.3 |
no |
1000 |
410.3 |
472.0 |
no |
5000 |
326.0 |
538.7 |
no |
Positive control |
575.7 |
- |
no |
* = solvent control with DMSO
Table 3a. Experiment 2 -Mutagenicity Assay. Number of revertants per plate (average).
Positive control was 2-nitrofluorene without metabolic activation and 2-aminoanthracene with metabolic activation.
Strain |
TA 98 |
||
Concentration |
- MA |
+ MA |
Cytotoxi (yes/no) |
0* |
29.8 |
39.2 |
no |
1 |
36.3 |
39.3 |
no |
2 |
29.7 |
36.7 |
no |
3 |
38.3 |
31.7 |
no |
4 |
32.0 |
35.0 |
no |
5 |
40.3 |
28.7 |
yes |
Positive control |
776.0 |
1412.7 |
no |
* = solvent control with DMSO
Concentrations: 0, 1, 2, 3, 4 & 5 without metabolic activation were respectively = 0, 312.5, 625, 1250, 2500 and 5000 μg/plate; and with metabolic activation respectively = 0, 156.25, 312.5, 625, 1250, 2500 μg/plate.
Table 3b. Experiment 2 -Mutagenicity Assay. Number of revertantsper plate (average).
Positive control was sodium azide for both strains without metabolic activation, the positive control with metabolic activation was 2-aminoanthracene with strain TA100 and none for strain TA1535
Strain |
TA 100 |
TA 1535 |
||||
Concentration |
-MA |
+MA |
Cytotoxic (yes/no) |
-MA |
+MA |
Cytotoxic (yes/no) |
0* |
126.6 |
133.0 |
no |
22.0 |
18.0 |
no |
1 |
126.5 |
136.0 |
no |
20.3 |
30.0 |
no |
2 |
121.7 |
138.3 |
no |
19.3 |
26.0 |
no |
3 |
125.0 |
129.0 |
no |
22.7 |
21.3 |
no |
4 |
139.0 |
145.3 |
no |
30.5 |
26.0 |
no |
5 |
121.0 |
134.7 |
no |
20.7 |
25.0 |
no |
Positive control |
717.0 |
1061.0 |
no |
549.3 |
- |
no |
* = solvent control with DMSO
Concentrations: 0, 1, 2, 3, 4 & 5 without metabolic activation were respectively = 0, 312.5, 625, 1250, 2500 and 5000 μg/plate; and with metabolic activation respectively = 0, 156.25, 312.5, 625, 1250, 2500 μg/plate.
Table 3c. Experiment 2 - Mutagenicity Assay. Number of revertants per plate (average).
Positive control was 9-aminoacridine without metabolic activation, and no positive control with metabolic activation.
Strain |
TA 1537 |
||
Concentration |
- MA |
+ MA |
Cytotoxic (yes/no) |
0* |
12.6 |
12.4 |
no |
1 |
10.3 |
11.0 |
no |
2 |
11.7 |
11.7 |
no |
3 |
10.7 |
12.0 |
no |
4 |
10.0 |
16.0 |
no |
5 |
11.7 |
9.0 |
no |
Positive control |
853.0 |
- |
no |
* = solvent control with DMSO
Concentrations: 0, 1, 2, 3, 4 & 5 without metabolic activation were respectively = 0, 312.5, 625, 1250, 2500 and 5000 μg/plate; and with metabolic activation respectively = 0, 156.25, 312.5, 625, 1250, 2500 μg/plate.
Table 3d. Experiment 2 -Mutagenicity Assay. Number of revertantsper plate (average).
Positive control was glutaraldehyde without metabolic activation, and there was no positive control with metabolic activation.
Strain |
TA 102 |
||
Concentration |
- MA |
+ MA |
Cytotoxic (yes/no) |
0* |
336.8 |
370.8 |
no |
1 |
315.7 |
354.7 |
no |
2 |
325.0 |
362.7 |
no |
3 |
298.0 |
344.7 |
no |
4 |
330.3 |
351.7 |
no |
5 |
287.3 |
- |
yes |
Positive control |
619.0 |
- |
no |
* = solvent control with DMSO
Concentrations: 0, 1, 2, 3, 4 & 5 without metabolic activation were respectively = 0, 312.5, 625, 1250, 2500 and 5000 μg/plate; and with metabolic activation respectively = 0, 156.25, 312.5, 625, 1250, 2500 μg/plate.
Table 4a. Experiment 1 - Results of chromosome analysis, 20+0 hours without metabolic activation.
Mitotic index is the average of 200 cells.
|
Control* |
271.3μg/ml |
387.5μg/ml |
553.6μg/ml |
Positive control |
|
Cytotoxicity |
no |
yes |
yes |
yes |
yes |
|
|
Total of 200 cells |
|||||
Chromatid aberrations*** |
gaps** |
2 |
3 |
3 |
4 |
6 |
deletions |
2 |
3 |
4 |
6 |
14 |
|
interchanges |
0 |
0 |
0 |
0 |
19 |
|
Chromosome aberrations*** |
gaps |
- |
- |
- |
- |
- |
deletions |
0 |
0 |
0 |
2 |
2 |
|
interchanges |
0 |
0 |
0 |
0 |
0 |
|
Mitotic index |
2.1 |
1.4 |
1.4 |
1.3 |
- |
|
Polyploidy and Endoreplication |
NR |
NR |
NR |
NR |
NR |
|
Total number of cells with aberrations, excluding gaps |
2 |
3 |
4 |
7 |
23 |
|
* solvent control with acetone
** It is not clear in the study report if the gaps recorded are chromatid or chromosome gaps
*** per culture
NR not reported
Table 4b. Experiment 1 - Results of chromosome analysis, 3+17 hours with metabolic activation.
Mitotic index is the average of 200 cells.
|
Control* |
790.9 μg/ml |
1130 μg/ml |
1614 μg/ml |
Positive control |
|
Cytotoxicity |
yes/no |
yes/no |
yes/no |
yes/no |
yes/no |
|
|
Total of 200 cells on average |
|||||
Chromatid aberrations |
gaps** |
0 |
4 |
3 |
1 |
28 |
deletions |
0 |
2 |
0 |
0 |
26 |
|
interchanges |
0 |
0 |
0 |
0 |
8 |
|
Chromosome aberrations |
gaps |
- |
- |
- |
- |
- |
deletions |
1 |
0 |
0 |
0 |
9 |
|
interchanges |
0 |
1 |
0 |
0 |
0 |
|
Mitotic index |
5.3 |
3.1 |
2.5 |
3.8 |
- |
|
Polyploidy and Endoreplication |
NR |
NR |
NR |
NR |
NR |
|
Total number of cells with aberrations, excluding gaps |
1 |
3 |
0 |
0 |
28 |
|
* solvent control with acetone
** It is not clear in the study report if the gaps recorded are chromatid or chromosome gaps
*** per culture
Table 4c. Experiment 2 - Results of chromosome analysis, 44+0 hours without metabolic activation.
Mitotic index is the average of 200 cells.
|
Control* |
387.5 μg/ml |
553.6 μg/ml |
|
Cytotoxicity |
yes/no |
yes/no |
yes/no |
|
|
Total of 200 cells on average |
|||
Mitotic index |
2.9 |
2.0 |
1.4 |
|
Total number of cells with aberrations, excluding gaps |
1 |
7 |
12 |
|
* = solvent control with acetone
Table 4d. Experiment 2 - Results of chromosome analysis, 3+41 hours with metabolic activation.
Mitotic index is the average of 200 cells.
|
Control* |
1614 μg/ml |
|
Cytotoxicity |
yes/no |
yes/no |
|
|
Total of 200 cells on average |
||
Mitotic index |
3.9 |
3.5 |
|
Total number of cells with aberrations, excluding gaps |
1 |
1 |
|
* = solvent control with acetone
¥ = numbers exceeding historical control range
Trial 1: the results of Trial 1 are not presented here as an unacceptably high solvent mutation frequency in the presence of metabolic activation was reported.
Table 2 Toxicity evaluation 1 Growth % of control
Strain |
L5178Y |
||
Conc. (μg/ml) |
- MA |
+ MA |
Cytotoxic (yes/no) |
5 |
83 |
0 |
yes |
1 |
101 |
88 |
no |
0.5 |
93 |
95 |
no |
0.1 |
96 |
96 |
no |
0.05 |
93 |
101 |
no |
0.01 |
97 |
96 |
no |
0.005 |
99 |
90 |
no |
| 0.001 | 93 | 93 | no |
Table 2a.Experiment 2 -Mutagenicity Assay. Mutant frequency (average of 3 plates).
Strain |
L5178Y |
|||
Conc. (μg/ml) |
- MA |
Conc. (μg/ml) |
+MA |
cytotoxic (yes/no) |
9 |
0.29 |
5.1 |
0.59 |
no |
9 |
0.30 |
5.1 |
0.51 |
yes |
8.7 |
0.30 |
4.9 |
0.46 |
no |
8.7 |
0.24 |
4.9 |
0.51 |
no |
8.4 |
0.31 |
4.7 |
0.49 |
no |
8.4 |
0.27 |
4.7 |
0.41 |
no |
8 |
0.32 |
4.6 |
0.50 |
no |
8 |
0.30 |
4.4 |
0.63 |
no |
6.9 |
0.36 |
4.2 |
0.53 |
yes |
6.9 |
0.32 |
4.2 |
0.55 |
no |
0* |
0.26 |
0* |
0.56 |
- |
0* |
0.35 |
0* |
0.50 |
- |
0** |
0.24 |
0** |
0.60 |
-- |
0** |
0.28 |
0** |
0.48 |
- |
0** |
0.32 |
0** |
0.56 |
- |
0** |
0.27 |
0** |
0.51 |
- |
|
|
|
|
|
* solvent control with DMSO
** untreated control
Table 4 Toxicity evaluation 2 Growth % of control
Strain |
L5178Y |
||
Conc. (μg/ml) |
- MA |
+ MA |
Cytotoxic (yes/no) |
5 |
0 |
0 |
yes |
1 |
94 |
94 |
no |
0.5 |
88 |
93 |
no |
0.1 |
98 |
101 |
no |
0.05 |
100 |
99 |
no |
0.01 |
98 |
99 |
no |
0.005 |
99 |
107 |
no |
Table 5 Experiment 3 -Mutagenicity Assay. Mutant frequency (average of 3 plates).
Strain |
L5178Y |
|
||
Conc. (μg/ml) |
- MA |
Conc. (μg/ml) |
+MA |
Cytotoxic (yes/no) |
5 |
0.62 |
4.4 |
0.75 |
yes |
5 |
0.78 |
4.4 |
0.93 |
yes |
4.4 |
0.76 |
3.8 |
0.84 |
no |
4.4 |
0.66 |
3.8 |
0.88 |
no |
3.8 |
0.73 |
3.2 |
0.84 |
no |
3.8 |
0.58 |
3.2 |
0.70 |
no |
3.2 |
0.56 |
2.6 |
0.66 |
no |
3.2 |
0.52 |
2.6 |
0.76 |
no |
2.6 |
0.46 |
2 |
0.86 |
no |
2.6 |
0.54 |
2 |
0.64 |
no |
0* |
0.55 |
0* |
- |
- |
0* |
0.55 |
0* |
- |
- |
0** |
0.68 |
0** |
0 .70 |
- |
0** |
0.63 |
0** |
0.96 |
- |
0** |
0.48 |
0** |
0.88 |
- |
0** |
0.52 |
0** |
0.92 |
|
* solvent control with ethanol
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed (negative)
Genetic toxicity in vivo
Description of key information
Mammalian Bone Marrow Chromosome Aberration Test in rat (ip study): the
hydrolysis product trimethylsilanol, CAS 1066-40-6: negative (similar to
OECD Test Guideline 475) (Bioassay Systems Corporation (1982)).
Rat dominant lethal Assay (oral study): the hydrolysis product trimethylsilanol, CAS 1066-40-6: negative (similar to OECD Test Guideline 478) (Dow Corning Corporation, 1983).
Link to relevant study records
- Endpoint:
- in vivo mammalian somatic cell study: cytogenicity / bone marrow chromosome aberration
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 1980-12-22 to 1981-12-6
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- comparable to guideline study with acceptable restrictions
- Remarks:
- The study was conducted according to an appropriate national standard method which is similar to OECD 475, with acceptable restrictions. The restrictions are that the number of cells used for the determination of mitotic index was not stated, but appeared to be about 100 cells per animal, and the report is less detailed than required by the current guideline. Read-across to the registered substance is considered scientifically justified.
- Qualifier:
- according to guideline
- Guideline:
- other: Methodologies in Mutagen Testing (Toxicology & Applied Pharm 22: 269-275, 1972).
- Deviations:
- no
- Qualifier:
- equivalent or similar to guideline
- Guideline:
- OECD Guideline 475 (Mammalian Bone Marrow Chromosome Aberration Test)
- Deviations:
- yes
- Remarks:
- The number of cells evaluated to determine mitotic index was not stated, but appeared to be less than required by the current guideline. Some details were missing from the report.
- GLP compliance:
- not specified
- Type of assay:
- chromosome aberration assay
- Species:
- rat
- Strain:
- Sprague-Dawley
- Sex:
- male/female
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Gofmoor Farms
- Age at study initiation: 10 - 16 weeks
- Weight at study initiation: 200 - 280 g
- Assigned to test groups randomly: no information
- Housing: Six per cage
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
ENVIRONMENTAL CONDITIONS
- Temperature (°F): 68 ± 3 °F
- Humidity (%): approx 50 % - Route of administration:
- intraperitoneal
- Vehicle:
- - Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: solubility properties
- Lot/batch no. (if required): D-128
- Purity: reagent grade - Details on exposure:
- Route of exposure: intraperitoneal
- Duration of treatment / exposure:
- 6 - 48 hours
- Frequency of treatment:
- A single intraperitoneal administration equivalent to doses of 420, 200 and 100 mg/kg.
- Post exposure period:
- Animals were sacrificed at 6, 24 or 48 hours after injection.
- Remarks:
- Doses / Concentrations:
100 mg/kg bw
Basis:
nominal conc. - Remarks:
- Doses / Concentrations:
200 mg/kg bw
Basis:
nominal conc. - Remarks:
- Doses / Concentrations:
420 mg/kg bw
Basis:
nominal conc. - No. of animals per sex per dose:
- 6
- Control animals:
- yes, concurrent vehicle
- Positive control(s):
- - cyclophosphamide
- Justification for choice of positive control(s): standard guideline positive control
- Route of administration: intraperitoneal - Tissues and cell types examined:
- Chromosomes from bone marrow examined for breaks.
- Details of tissue and slide preparation:
- CRITERIA FOR DOSE SELECTION: rang-finding experiment to determine the maximum tolerated dose
DETAILS OF SLIDE PREPARATION: a small amount of cells suspended in fixative (3:1, methanol: acetic acid) were dropped onto slides which were subsequently air dried. Slides were stained with 5 % Giemsa.
METHOD OF ANALYSIS: Slides were scanned and only those of acceptable quality were retained. Suitable cells were photographed using a 100x objective. A minimum of 100 metaphase cells were analyzed per animal, and 5 animals were analysed for each dose. - Evaluation criteria:
- The test substance is cytogenic if it causes a statistically significant, dose related increase in the frequency of chromosomal breaks or aberrations.
- Statistics:
- The significance of differences in break frequency were examined using "goodness to fit" to a Poisson distribution, compared to historical negative control data, the Wilcoxon test and Chi square.
- Sex:
- male/female
- Genotoxicity:
- negative
- Toxicity:
- no effects
- Remarks:
- on bone marrow
- Vehicle controls validity:
- valid
- Negative controls validity:
- not applicable
- Positive controls validity:
- valid
- Additional information on results:
- RESULTS OF RANGE-FINDING STUDY
- Dose range: experiment 1: 175, 500, 1750, 5000 mg/kg bw; experiment 2: 75, 50, 175, 500, 1750 mg/kg bw
- Clinical signs of toxicity in test animals: none reported. Deaths occurred at the top dose in experiment 1 and in the top two doses in experiment 2. - Conclusions:
- Trimethylsilanol (CAS 1066-40-6) has been tested in chromosome aberration study conducted according to a protocol that is similar to OECD Test Guideline 475. The test substance did not cause a statistically significant, dose related increase in chromosome breaks or aberrations when administered by intraperitoneal injection to Sprague Dawley rats up to the maximum tolerated dose. It is concluded that the test substance is not clastogenic in rat bone marrow cells under the conditions of the test.
- Endpoint:
- in vivo mammalian germ cell study: cytogenicity / chromosome aberration
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- comparable to guideline study
- Remarks:
- The study was conducted according to an appropriate OECD test guideline. It was not compliant with GLP.
- Qualifier:
- according to guideline
- Guideline:
- other: OECD Method No. 426 (August 21, 1981; FR 42472)
- Version / remarks:
- 1981
- Qualifier:
- equivalent or similar to guideline
- Guideline:
- OECD Guideline 478 (Genetic Toxicology: Rodent Dominant Lethal Test)
- Version / remarks:
- 1984
- Principles of method if other than guideline:
- Extended Dominant Lethal Assay as recommended by the U.S. Food and Drug Administration (Green, et al 1977)
- GLP compliance:
- not specified
- Type of assay:
- rodent dominant lethal assay
- Species:
- rat
- Strain:
- Sprague-Dawley
- Sex:
- male
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Harlan Research, Haslett, Michigan
- Age at study initiation: 10 - 12 weeks
- Weight at study initiation: mean 355 - 375 g
- Assigned to test groups randomly: [yes, under following basis: randomly selected using a computer program]
- Housing: wire-bottomed stainless steel cages
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 7 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 24 °C
- Humidity (%): 45 - 55 % relative humidity
- Photoperiod (hrs dark / hrs light): 12 hours light / 12 hours dark
- Route of administration:
- oral: gavage
- Vehicle:
- - Vehicle(s)/solvent(s) used: none
- Justification for choice of solvent/vehicle: N/A
- Concentration of test material in vehicle: test material was given neat
- Amount of vehicle (if gavage or dermal): none
- Type and concentration of dispersant aid (if powder): N/A
- Lot/batch no. (if required): N/A
- Purity: N/A - Details on exposure:
- PREPARATION OF DOSING SOLUTIONS:
DIET PREPARATION
- Rate of preparation of diet (frequency): daily, 5 days per week for 8 weeks
- The test substance was administered undiluted by oral gavage.
- Storage temperature of food: no data - Duration of treatment / exposure:
- The exposure period lasted 8 weeks
- Frequency of treatment:
- Daily, for 5 days per week
- Post exposure period:
- 2 weeks
- Remarks:
- Doses / Concentrations:
0, 20, 100, 200 mg/kg
Basis:
analytical conc. - No. of animals per sex per dose:
- 15 males per dose group. 4 females per male (2 matings per week over 2 weeks)
- Control animals:
- yes, concurrent no treatment
- Positive control(s):
- triethylenemelamine
- Justification for choice of positive control(s): no data
- Route of administration: Oral gavage
- Doses / concentrations: 0.05 mg/kg bw/day - Tissues and cell types examined:
- Ovaries and uteri examined at necropsy. The number of corpora lutea and living and dead implantations were counted and recorded for each pregnant female.
- Details of tissue and slide preparation:
- CRITERIA FOR DOSE SELECTION: A two week subchronic dosing assay was conducted on sexually mature male Sprague-Dawley rats to determine a maximum tolerated dose level (MTD). The test material was administered undiluted by oral gavage. A level was selected which gave some evidence of biological effect (transient sedation) without producing sufficient toxicity to interfere with the test system. Based on this determination, male rats were randomly assigned to one of five experimental dosing groups (15 males/group).
DETAILS OF SLIDE PREPARATION: microscopic examination (no other information)
METHOD OF ANALYSIS: Number of corpora lutea, preimplantation loss, total number of implants, live and dead implants (restorations) counted for each mated female.
- Evaluation criteria:
- Examine for evidence that the test substance reaches mammalian germinal tissue and induces chromosomal aberrations. The analysis considers both effects per litter and total number of effects / total number of pups.
- Statistics:
- Data analysed for statistical significance (p <0.05 for all cases) by a computer program specifically designed for analysis of dominant lethal studies.
The program first tests the data for goodness to fit. Parametrically distributed data is analysed for statistical significance by Fishers Exact Probability Test while non-parametric data is automatically shifted to an analysis by the Wilcoxon Test as modified by Haseman and Hoel (1974). - Sex:
- male
- Genotoxicity:
- negative
- Toxicity:
- no effects
- Vehicle controls validity:
- valid
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Additional information on results:
- One male from the high dose group died acutely due to lung damage caused by a dosing accident. One animal from the intermediate dose group failed to gain weight and was eating and drinking poorly. The animal was sacrificed and evidence of erosion of the gastric mucosa noted. The death was not attributed to TMS.
- Conclusions:
- Trimethylsilanol (CAS 1066-40-6) has been tested in a rodent dominant lethal assay according to OECD Method No 426 (August 21, 1981; FR42472) and equivalent to OECD Test Guideline 478. The test substance was administered by oral gavage to male rats at three dose levels, five days per week over eight weeks, and gave no evidence of inducing chromosomal damage in germinal tissue; there was no substance related effect on fertility index, pre-implantation loss or resorption rate. A clear dominant lethal syndrome was evident in the positive control group. Although the test material was administered at a high enough level to induce transient sedation without toxicity, no reduction in male fertility or increase in dominant lethality was obtained. It is concluded that the test substance is negative for genetic toxicity to germ cells under the conditions of the test.
Referenceopen allclose all
Table 1: Results of chromosome analysis in Rat Bone Marrow Cells
|
Solvent Control (DMSO) |
Positive Control Cyclophosphamide |
Low dose 100 mg/kg |
Mid dose 200 mg/kg |
High dose 420 mg/kg |
|||||||||
Sampling time (h) |
6 |
24 |
48 |
24 |
6 |
24 |
48 |
6 |
24 |
48 |
6* |
24 |
48 |
|
Number of cells analysed |
551 |
592 |
523 |
286 |
552 |
568 |
502 |
546 |
593 |
509 |
499 |
574 |
534 |
|
Toxicity |
No |
No |
No |
No |
No |
No |
No |
No |
No |
No |
No |
No |
No |
|
Total number of Chromosome aberrations |
Gaps |
ND |
ND |
ND |
0 |
2 |
7 |
12 |
3 |
5 |
11 |
6 |
5 |
8 |
Breaks |
0 |
2 |
4 |
>99 |
2 |
7 |
5 |
6 |
3 |
8 |
3 |
5 |
12 |
|
Interchanges |
ND |
ND |
ND |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
|
Polyploidy |
ND |
ND |
ND |
ND |
ND |
ND |
ND |
ND |
ND |
ND |
ND |
ND |
ND |
|
Endo reduplication |
ND |
ND |
ND |
ND |
ND |
ND |
ND |
ND |
ND |
ND |
ND |
ND |
ND |
|
ND Not determined
Table 2: Number of males / pregnant females / non-pregnant females
Dose Group (mg / kg) |
0 |
20 |
100 |
200 |
Positive Control |
Total Number of Males |
15 |
15 |
14 |
14 |
15 |
Number of fertile Males |
13 |
14 |
12 |
12 |
13 |
Number of Pregnant females |
42 |
40 |
46 |
36 |
52 |
Number of non-pregnant females |
18 |
20 |
10 |
20 |
8 |
Table 3: Effect on Fertility index
Dose (mg/kg) |
Fertile/Total Males |
Fertility Index* |
||
Mating # 1 |
Mating # 2 |
Mating # 1 |
Mating # 2 |
|
0 |
8/15 |
13/15 |
53.3 |
86.7 |
20 |
6/15 |
14/15 |
40 |
93.3 |
100 |
11/14 |
12/14 |
78.6 |
85.7 |
200 |
6/14 |
12/14 |
42.9 |
85.7 |
Positive control |
13/15 |
13/15 |
86.7 |
86.7 |
*Number of fertile males / Total number of males x 100
Table 4: Effect on pre-implantation loss
Week of Mating |
Dose Level (mg/kg) |
Preimplantation Loss Rate (a) |
% Loss (b) |
Average % Response ± S.D (c) |
1 |
0 |
5/8 |
62.5 |
10.85 ± 13.93 |
20 |
1/6 |
16.7 |
1.23 ± 3.14 |
|
100 |
7/11 |
63.6 |
7.02 ± 8.84 |
|
200 |
4/6 |
66.7 |
6.67 ±5.95 |
|
Positive Control |
5/8 |
62.5 |
7.97 ± 17.81 |
|
2 |
0 |
7/13 |
53.8 |
9.05 ± 14.42 |
20 |
9/14 |
64.3 |
7.11 ± 8.82 |
|
100 |
7/13 |
53.8 |
2.75 ± 4.8 |
|
200 |
7/12 |
58.3 |
12.69 ± 24.55 |
|
Positive Control |
8/13 |
61.5 |
10.68 ± 13.8 |
|
3 |
0 |
10/13 |
76.9 |
9.44 ± 10.02 |
20 |
9/14 |
64.3 |
5.33 ± 5.6 |
|
100 |
7/12 |
58.3 |
4.11 ± 4.29 |
|
200 |
8/12 |
66.7 |
7.83 ± 10.39 |
|
Positive Control |
9/13 |
69.2 |
10.42 ± 10.83 |
(a) Total number of males producing preimplantation / Total number of fertile males
(b) Preimplantation loss rate x 100
(c) Total preimplantation loss / Total number Corpora Lutea x 100 = Mean (x) percent loss
Table 5 : Effect on Resorption rate
Week of Mating |
Dose Level (mg/kg) |
Resorption Rate (a) |
Resorption% (b) |
Average % Response ± S.D (c) |
1 |
0 |
1/8 |
12.5 |
1.14 ± 3.22 |
20 |
3/6 |
50 |
3.35 ± 3.68 |
|
100 |
3/11 |
27.3 |
3.05 ± 7.02 |
|
200 |
1/6 |
16.7** |
0.63 ± 1.55 |
|
Positive Control |
10/13 |
76.9 |
18.95** ± 13.34 |
|
2 |
0 |
5/13 |
38.5 |
2.19 ± 3.12 |
20 |
4/14 |
28.6 |
2.61 ± 5.6 |
|
100 |
5/12 |
41.7 |
2.3 ± 2.99 |
|
200 |
3/12 |
25 |
1.69 ± 3.48 |
|
Positive Control |
9/13 |
69 |
13.8** ± 21.17 |
|
3 |
0 |
6/13 |
46.2 |
1.99 ± 2.29 |
20 |
6/14 |
42.9 |
2.32 ± 3.24 |
|
100 |
6/12 |
50 |
2.59 ± 4.17 |
|
200 |
4/12 |
33.3 |
1.65 ± 3.26 |
|
Positive Control |
12/13 |
92.3* |
16.08** ± 11.54 |
(a) Total number of males producing preimplantation / Total number of fertile males
(b)Resorption rate x 100
(c) Total average percent resorption rate (Total resorptions/ Total implants per fertile male / Total number fertile males
* Significant at p <0.05
** Significant at p <0.01
No dose-related deaths, clinical symptoms or behavioural changes were observed.
Male rats receiving mid to high dose showed signs of sedation immediately after dosing.
There was no significant difference between the mean body weight gain for any of the dosing groups compared to negative control group.
There was no statistically significant difference in fertility index / pre-implantation loss or resorption rate in any of the groups dosed with TMS compared to the negative control group.
A clear dominant lethal syndrome was evident in the positive control group.
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed (negative)
Additional information
Information is available from reliable studies for all the required in vitro endpoints. The most recent of the most reliable studies were selected as key. They were conducted according to OECD Test Guidelines and in compliance with GLP.
Additional information is available for the hydrolysis product, trimethylsilanol (CAS 1066 -40 -6), from an in vitro cytogenicity study, and in vivo assays for clastogenicity to somatic and to germ cells.
1,1,1,3,3,3 -Hexamethyldisilazane (CAS 999-97-3, EC 213-668-5) hydrolyses very rapidly to trimethylsilanol (hydrolysis half-life at pH 7 and 1.5°C: ≤ 0.5 min (measured)); the other hydrolysis products, ammonium ions, are not expected to contribute to genetic toxicity, based on information in the public domain (OECD, 2003, 2004, 2005). It is considered, therefore, that read-across to the registered substance is scientifically justified.
No test substance related increase in the number of revertants relative to the solvent control was observed when 1,1,1,3,3,3-hexamethyldisilazane (CAS 999-97-3, EC 213-668-5) was tested in S. typhimurium strains TA 98, TA 100, TA 1535, TA 1537 and TA 102 in accordance with OECD Test Guideline 471 and in compliance with GLP (Covance, 1997a). The results obtained with and without metabolic activation in the initial plate incorporation test and in the repeat pre-incubation assay were in agreement. Appropriate solvent and positive controls were included and gave expected results. The test substance was therefore considered to be negative for mutagenicity to bacteria under the conditions of this test. A number of supporting bacterial mutagenicity studies are available and all produced a negative result.
1,1,1,3,3,3 -Hexamethyldisilazane (CAS 999-97-3, EC 213-668-5) has been tested for mammalian cell cytogenicity in human lymphocyte cells according to OECD Test Guideline 473 and in compliance with GLP (Covance, 1997b). The proportions of cells with aberrations (cells with structural aberrations including gaps, cells with structural aberrations excluding gaps and polyploid, endoreduplicated or hyperdiploid cells) were examined in relation to historical negative control (normal ranges). Frequencies of cells with numerical aberrations fell within historical negative control ranges under most treatment conditions. The number of aberrant cells fell outside the normal range in a single culture at the highest concentration following treatment for 20 hours without metabolic activation, but the increase was small and was not seen in the replicate culture, and was associated with lower cell viability, so it is not considered to be of biological significance. The number of aberrant cells fell outside the normal range in a single culture at each of the two concentrations analysed following treatment for 44 hours without metabolic activation, but because the increase was small and not seen in replicate cultures, and was also associated with lower cell viability, the effect was considered to be of no biological significance. In addition, sampling at this time (approximately 3.3 times the normal cell cycle length) is outside the requirements of method B.10. Mutagenicity - in vitro mammalian chromosome aberration test in Regulation (EC) 440/2008, which is an additional reason for considering the slight increase of no biological significance. It is therefore concluded that the test substance is negative for cytogenicity in human lymphocytes under the conditions of the test.
The conclusion from the key cytogenicity study, (Covance, 1997b), is supported by findings in a second study, where a weak positive response was observed in Chinese hamster ovary cells in the first trial without metabolic activation using a non-standard protocol (NTP 1995). This response was not replicated in the second trial, and the study result is reported to be negative.
Results are available from an in vitro study in which 1,1,1,3,3,3-hexamethyldisilazane (CAS 999-97-3, EC 213-668-5) was tested for mutagenicity to L5178Y TK+/- cells in the presence and absence of metabolic activation according to OECD Test Guideline 476 and in compliance with GLP (Microbiological Associates, 1991). No test substance related increase in mutant frequency relative to solvent control was observed in any of the three trials conducted in this study. In addition, there was no evidence of an increase in small colonies when treated cultures were compared to solvent cultures. It is concluded that the test substance is neither mutagenic nor clastogenic under the conditions of the test.
Additional information on the potential for cytogenicity of the registered substance is available from an in vitro cytogenicity study on the silanol hydrolysis product, trimethylsilanol, CAS 1066-40-6, which was tested according to a protocol that is similar to OECD Test Guideline 473 for cytogenicity to mammalian cells using mouse lymphoma L5178Y cells and testing up to cytotoxic concentrations (Litton Bionetics, 1978). No increase in the number of cells with two or more aberrations was observed in the presence of metabolic activation. A dose-related increase in the frequency of aberrations per cell was reported in the absence of metabolic activation, and it was concluded by the authors of the report that this represented evidence for clastogenicity, though the authors observed that these effects could be the result of cytotoxic stress on the cells. The number of cells with two or more aberrations per cell was not increased. The number of cells with aberrations was not reported. It is concluded by the reviewer that trimethylsilanol does not induce biologically significant aberrations in mouse lymphoma L5178Y cells with or without metabolic activation under the conditions of the test.
The negative conclusion reached after careful consideration of the results obtained in the key in vitro cytogenicity study on 1,1,1,3,3,3-hexamethyldisilazane (CAS 999-97-3, EC 213-668-5) is supported by the lack of evidence of increase in small colonies in the key in vitro mammalian mutagenicity study on the registered substance. Induction of small colonies in such studies is associated with chemicals that induce gross chromosome aberrations (Regulation EC 440/2008 citing Yandell et al, 1990, Mutation Res., 229, p. 89-102). Additional supporting data come from testing on the hydrolysis product trimethylsilanol, CAS 1066-40-6, which include, in addition to a negative interpretation of an in vitro study, in vivo cytogenicity studies on somatic and germ cells.
Trimethylsilanol (CAS 1066 -40 -6) has been tested in an in vivo chromosome aberration study conducted according to a protocol that is similar to OECD Test Guideline 475 (Bioassay Systems Corporation, 1982). The test substance did not cause a statistically significant, dose related increase in chromosome breaks or aberrations when administered by intraperitoneal injection to Sprague Dawley rats up to the maximum tolerated dose. It is concluded that the test substance is not clastogenic in rat bone marrow cells under the conditions of the test. Trimethylsilanol has been tested in an in vivo rodent dominant lethal assay according to OECD Test Guideline 421 (1981) and equivalent to OECD Test Guideline 478 (Dow Corning Corporation, 1983). The test substance when administered to male rats at three dose levels, five days per week over eight weeks, gave no evidence of inducing chromosomal damage in germinal tissue; there was no substance related effect on fertility index, pre-implantation loss or resorption rate. A clear dominant lethal syndrome was evident in the positive control group. Although the test material was administered at a high enough level to induce transient sedation without toxicity, no reduction in male fertility or increase in dominant lethality was obtained. It is concluded that the test substance is negative for genetic toxicity to germ cells under the conditions of the test.
OECD (2003): SIDS Initial Assessment Report for SIAM 17, Arona, Italy, 11 -14 November 2003, Ammonium Chloride, CAS 12125 -02 -9.
OECD (2004): SIDS Initial Assessment Report for SIAM 19, Berlin, Germany, 18-20 October 2004, Ammonium sulfate, CAS 7783 -20 -2.
OECD (2005): SIDS Initial Assessment Report for SIAM 20, Paris, France, 19 -21 April 2005, Persulfates, CAS 7727 -54 -0.
Justification for classification or non-classification
Based on the available in vitro and in vivo genotoxicity data, 1,1,1,3,3,3-hexamethyldisilazane (CAS 999-97-3, EC 213-668-5) does not require classification for mutagenicity according to Regulation (EC) No 1272/2008.
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