1,1,1,3,3,3-hexamethyldisilazane

Endpoint summary

Description of key information

In a 90-day repeated dose toxicity study conducted according to OECD Test Guidelines 413 and in compliance with GLP, nose-only inhalation exposure of 1,1,1,3,3,3-hexamethyldisilazane (CAS 999-97-3, EC 213-668-5) at concentrations of 25, 75 or 400 ppm to Sprague-Dawley rats for 6 hours per day, 5 days per week for 13 weeks resulted in minor and reversible changes at the highest concentration. Renal effects in male rats were observed at all concentrations, but these findings were consistent with alpha-2u-nephropathy and therefore of no toxicological significance to humans. Based on the results in this study, the NOAEC was considered to be higher than or equal to 400 ppm (equivalent to 2640 mg/m³), the highest concentration tested (Harlan 2014).

1,1,1,3,3,3-Hexamethyldisilazane is rapidly hydrolysed in contact with moisture releasing trimethylsilanol and ammonia.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records

Reference

Endpoint:

sub-chronic toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

17 April to 27 August 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Study was conducted in accordance with an appropriate guideline and in compliance with GLP.

Qualifier:

according to guideline

Guideline:

OECD Guideline 413 (Subchronic Inhalation Toxicity: 90-Day Study)

Deviations:

yes

Remarks:

(relative humidity of chamber atmosphere)

Qualifier:

according to guideline

Guideline:

EU Method B.29 (Sub-Chronic Inhalation Toxicity:90-Day Study)

Deviations:

yes

Remarks:

(relative humidity of chamber atmosphere)

Principles of method if other than guideline:

For technical reasons, the relative humidity value of the chamber atmosphere was below the range given by the guidelines. The reason is the dried, pressurized air used for vapor generation, in combination with heating the test item in the nebulizer to 68 - 76 °C. This deviation from the guidelines has no influence on the results of the study, based on the absence of any findings in the respiratory tract and the circumstance that dry air generally worsens any findings in the respiratory tract due to loss of the protective mucus layer on the lining epithelium.

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Species:

rat

Strain:

other: Crl:CD(SD)IGS

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River, Germany
- Age at study initiation: 8 weeks
- Weight at study initiation: 268-297g (males); 163-200g (females)
- Fasting period before study: no
- Housing: Makrolon type-4 cages with wire mesh tops and sterilized standard softwood bedding (Lignocel), including paper enrichment (Enviro-dri)
- Diet (ad libitum): Pelleted standard Harlan Teklad 2914C rodent maintenance diet (Provimi Kliba AG, Switzerland),
- Water (ad libitum): community tap water
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22+/- 3
- Humidity (%): 30-70
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: To: 17 April to 27 August 2013

Route of administration:

inhalation: vapour

Type of inhalation exposure:

nose only

Vehicle:

air

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: nose-only, flow-past exposure chamber
- Method of holding animals in test chamber: restraint tubes
- Source and rate of air:
- Method of conditioning air:
- System of generating vapour: 1,1,1,3,3,3-Hexamethyldisilazane was pumped into a glass flask. Compressed, filtered, and dried air was supplied into the glass flask through a metal nebulization tube. The glass flask was kept between a temperature of 68 and 76 °C with a thermal regulating device set to facilitate the process of vaporization.
- Temperature, humidity, pressure in air chamber: 19.9 to 22.6 degrees C: 0.0 to 4.0% ; slight positive pressure to outside
- Air flow rate: 0.75-1.0L/min
- Air change rate:
- Method of particle size determination:
- Treatment of exhaust air:

TEST ATMOSPHERE
- Brief description of analytical method used: nominal - wieighing of test susbtance; chemical - on-line GC
- Samples taken from breathing zone: yes

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The concentration of 1,1,1,3,3,3-Hexamethyldisilazane was measured at least once per hour of exposure per on-line GC for groups 1 to 4.

Analytical concentrations were determined by GC. The method of analysis was performed according to the conditions listed below.
- Column: HP-5MS (30 m, 0.25 mm, 0.25 μm)
- Injector: 250 °C
- Oven: 40 °C for 1 min, then 40 °C/min to 100 °C for 0 min, then 50 °C/min to 230 °C for 1 min
- Detector: 280 °C

- Calibration:
A calibration curve consisting of at least 6 points and ranging between concentrations of approximately 10 ppm to approximately 500 ppm were constructed from the test item in gas bags as part of the technical trials. The calibration gas bags were prepared at each concentration and at least two independent gas bags were used.

- Acceptance Criteria:
The coefficients of variation were below 3.1% (acceptance criteria of >10%) for all calibration gas bag samples at each concentration. The correlation coefficient of the used regression was >0.9989 (acceptance criteria of >0.985). Accordingly, the acceptance criteria were met.

Daily standards constructed from the test item in gas bags were sampled at one of the chamber-lines and used to check the integrity of the sampling line and check the GC calibration. Plots of the peak height used for the calibration were used to assess trends regarding system stability. The acceptance criterion for standard samples was an accuracy of 90 - 110% of the theoretical value. The acceptance criteria for standard samples were met for each exposure session.

The test item was used as the analytical standard.

Duration of treatment / exposure:

6 hours

Frequency of treatment:

5 days/week for 13 weeks, followed by 4 week recovery period for sub-group of Control and high dose

Dose / conc.:

0 ppm (nominal)

Remarks:

Group 1 - Control Group

Dose / conc.:

25 ppm (nominal)

Remarks:

Group 2 (165 mg/m3)

Dose / conc.:

75 ppm (nominal)

Remarks:

Group 3 (495 mg/m3)

Dose / conc.:

400 ppm (nominal)

Remarks:

Group 4 (2640 mg/m3)

No. of animals per sex per dose:

20 Control and high dose, 10 low and intermediate dose

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: dose concentrations based on previous OECD 422 inhalation study
- Post-exposure recovery period in satellite groups: 4 weeks

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily during dosing period, once daily during recovery period

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: twice daily during dosing period, once weekly during recovery period

BODY WEIGHT: Yes
- Time schedule for examinations: twice weekly during dosing period, once weekly during recovery period

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/animal/day: Yes
- Time schedule for examinations: weekly

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: before dosing started and Week 13
- Dose groups that were examined: all (not recovery animals in Week 13)

HAEMATOLOGY: Yes
- Time schedule for collection of blood: prior to necropsy at end of dosing or recovery period
- Anaesthetic used for blood collection: Yes (isoflurane)
- Animals fasted: Yes
- How many animals: 10 sex/group
- Parameters checked in table No. 1 were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: prior to necropsy at end of dosing or recovery period
- Animals fasted: Yes
- How many animals: 10 sex/group
- Parameters checked in table No. 2 were examined.

URINALYSIS: Yes
- Time schedule for collection of urine: 18 hour period prior to necropsy at end of dosing or recovery period
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes
- Parameters checked in table No. 3 were examined.

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes (see table)
HISTOPATHOLOGY: Yes (see table)

Statistics:

The following statistical methods were used to analyze the food consumption, body weight, ophthalmoscopic examinations, macroscopic findings, organ weights and ratios, as well as clini¬cal laboratory data:

• The two-sided Dunnett-test (many to one t-test) based on a pooled variance estimate was applied if the variables could be assumed to follow a normal distribution for the comparison of the treated groups and the control groups for each sex.

• The Steel-test (many-one rank test) was applied instead of the Dunnett-test when the data could not be assumed to follow a normal distribution.

• The two-sided Fisher's exact-test was applied to the ophthalmo-scopic and macroscopic findings.

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Ataxia and decreased activity were noted in all animals in the high dose from treatment start onwards. Ataxia was recorded until weeks 8 and 11 in males and females, respectively, and decreased activity was recorded until week 3 for both sexes. Based on adaptation the animals showed during the treatment and on the complete reversibility of the findings during recovery, these findings are considered to be non-adverse.

Mortality:

no mortality observed

Description (incidence):

There were no unscheduled deaths during the course of the study.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Lower body weight gain for high dose animals in both sexes during treatment period. An increased body weight gain was recorded in males during the recovery period.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

Lower for high dose males during treatment period and for high dose females during first two weeks of treatment period. An increased food intake was noted during recovery.

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not specified

Ophthalmological findings:

no effects observed

Description (incidence and severity):

No ophthalmoscopic findings were observed which were considered to be attributable to exposure with the test item

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

Total red blood cells and hematocrit were slightly but statistically significantly higher at the end of the treatment period in females at 400 ppm. These findings had completely reversed at the end of the recovery period.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

Sodium, potassium and chloride levels were higher at the end of the treatment period in females at 400 ppm. In addition, total protein was lower for those animals. These findings had completely reversed at the end of the recovery period.

Urinalysis findings:

effects observed, treatment-related

Description (incidence and severity):

The pH value was lower at the end of the treatment period in males at 400 ppm. This finding had completely reversed at the end of the recovery period.

Behaviour (functional findings):

not specified

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

Absolute and relative liver weights were higher in females at 400 ppm. In the absence of any histopathological findings in the livers and based on the reversibility
that was observed during the recovery period, this finding is considered to be adaptive.

Gross pathological findings:

no effects observed

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

In comparison to controls the incidence and severity of increased intra-epithelial hyaline droplets and focal or multifocal basophilic tubules was greater in all the male treated groups. In addition in 2/10 males given 400 ppm granular casts were present at the cortico-medullary junction of the kidneys. These findings were all considered to be consistent with a diagnosis of alpha-2µ-nephropathy and therefore of no toxicological significance to humans.

Histopathological findings: neoplastic:

no effects observed

Other effects:

not specified

Key result

Dose descriptor:

NOAEC

Effect level:

>= 400 ppm (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other:

Remarks on result:

not determinable due to absence of adverse toxic effects

Key result

Critical effects observed:

yes

Lowest effective dose / conc.:

400 ppm (nominal)

System:

urinary

Organ:

kidney

Treatment related:

yes

Dose response relationship:

not specified

Relevant for humans:

no

Table 1 - Exposure Conditions

Group	Temperature [°C]	Relative Humidity [%]	Oxygen Concentration [%]
1	22.4 ± 0.2 (n=66)	0.0 ± 0.0 (n=66)	20.2 ± 0.0 (n=66)
2	22.2 ± 0.1 (n=66)	3.0 ± 0.1 (n=66)	20.2 ± 0.0 (n=66)
3	22.1 ± 0.2 (n=66)	3.0 ± 0.5 (n=66)	20.0 ± 0.0 (n=66)
4	22.2 ± 0.1 (n=66)	3.9 ± 0.1 (n=66)	20.2 ± 0.1 (n=66)

For technical reasons, the relative humidity value of the chamber atmosphere was below the range given by the guidelines. The reason is the dried, pressurized air used for vapor generation, in combination with heating the test item in the nebulizer to 68 – 76 °C. This deviation from the guidelines has no influence on the results of the study, based on the absence of any findings in the respiratory tract and the circumstance that dry air generally worsens any findings in the respiratory tract due to loss of the protective mucus layer on the lining epithelium.

Table 2 - Test Atmosphere Concentrations

Group	Chemically Determined AtmosphereConcentration [ppm]	TargetAtmosphereConcentration [ppm]	Atmosphere ConcentrationsRelative to Target
2	25.0 ± 0.5 (n=66, CV=1.8%)	25	100.0%
3	75.0 ± 1.6 (n=66, CV=2.2%)	75	100.0%
4	400 ± 4 (n=66, CV=1.1%)	400	100.0%

 

Conclusions:

In a repeated dose toxicity study conducted according to OECD Test Guidelines 413 and in compliance with GLP, nose-only inhalation exposure of 1,1,1,3,3,3-hexamethyldisilazane (CAS 999-97-3, EC 213-668-5) at concentrations of 25, 75 or 400 ppm to Sprague-Dawley rats for 6 hours per day, 5 days per week for 13 weeks resulted in minor and reversible changes at the highest concentration. Renal effects in male rats were observed at all concentrations, but these findings were consistent with alpha-2u-nephropathy and therefore of no toxicological significance to humans. Based on the results in this study, the NOAEC was considered to be higher than or equal to 400 ppm, the highest concentration tested.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEC

2 640 mg/m³

Study duration:

subchronic

Species:

rat

Repeated dose toxicity: inhalation - local effects

Link to relevant study records

Reference

Endpoint:

sub-chronic toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

17 April to 27 August 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Study was conducted in accordance with an appropriate guideline and in compliance with GLP.

Qualifier:

according to guideline

Guideline:

OECD Guideline 413 (Subchronic Inhalation Toxicity: 90-Day Study)

Deviations:

yes

Remarks:

(relative humidity of chamber atmosphere)

Qualifier:

according to guideline

Guideline:

EU Method B.29 (Sub-Chronic Inhalation Toxicity:90-Day Study)

Deviations:

yes

Remarks:

(relative humidity of chamber atmosphere)

Principles of method if other than guideline:

For technical reasons, the relative humidity value of the chamber atmosphere was below the range given by the guidelines. The reason is the dried, pressurized air used for vapor generation, in combination with heating the test item in the nebulizer to 68 - 76 °C. This deviation from the guidelines has no influence on the results of the study, based on the absence of any findings in the respiratory tract and the circumstance that dry air generally worsens any findings in the respiratory tract due to loss of the protective mucus layer on the lining epithelium.

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Species:

rat

Strain:

other: Crl:CD(SD)IGS

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River, Germany
- Age at study initiation: 8 weeks
- Weight at study initiation: 268-297g (males); 163-200g (females)
- Fasting period before study: no
- Housing: Makrolon type-4 cages with wire mesh tops and sterilized standard softwood bedding (Lignocel), including paper enrichment (Enviro-dri)
- Diet (ad libitum): Pelleted standard Harlan Teklad 2914C rodent maintenance diet (Provimi Kliba AG, Switzerland),
- Water (ad libitum): community tap water
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22+/- 3
- Humidity (%): 30-70
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: To: 17 April to 27 August 2013

Route of administration:

inhalation: vapour

Type of inhalation exposure:

nose only

Vehicle:

air

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: nose-only, flow-past exposure chamber
- Method of holding animals in test chamber: restraint tubes
- Source and rate of air:
- Method of conditioning air:
- System of generating vapour: 1,1,1,3,3,3-Hexamethyldisilazane was pumped into a glass flask. Compressed, filtered, and dried air was supplied into the glass flask through a metal nebulization tube. The glass flask was kept between a temperature of 68 and 76 °C with a thermal regulating device set to facilitate the process of vaporization.
- Temperature, humidity, pressure in air chamber: 19.9 to 22.6 degrees C: 0.0 to 4.0% ; slight positive pressure to outside
- Air flow rate: 0.75-1.0L/min
- Air change rate:
- Method of particle size determination:
- Treatment of exhaust air:

TEST ATMOSPHERE
- Brief description of analytical method used: nominal - wieighing of test susbtance; chemical - on-line GC
- Samples taken from breathing zone: yes

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The concentration of 1,1,1,3,3,3-Hexamethyldisilazane was measured at least once per hour of exposure per on-line GC for groups 1 to 4.

Analytical concentrations were determined by GC. The method of analysis was performed according to the conditions listed below.
- Column: HP-5MS (30 m, 0.25 mm, 0.25 μm)
- Injector: 250 °C
- Oven: 40 °C for 1 min, then 40 °C/min to 100 °C for 0 min, then 50 °C/min to 230 °C for 1 min
- Detector: 280 °C

- Calibration:
A calibration curve consisting of at least 6 points and ranging between concentrations of approximately 10 ppm to approximately 500 ppm were constructed from the test item in gas bags as part of the technical trials. The calibration gas bags were prepared at each concentration and at least two independent gas bags were used.

- Acceptance Criteria:
The coefficients of variation were below 3.1% (acceptance criteria of >10%) for all calibration gas bag samples at each concentration. The correlation coefficient of the used regression was >0.9989 (acceptance criteria of >0.985). Accordingly, the acceptance criteria were met.

Daily standards constructed from the test item in gas bags were sampled at one of the chamber-lines and used to check the integrity of the sampling line and check the GC calibration. Plots of the peak height used for the calibration were used to assess trends regarding system stability. The acceptance criterion for standard samples was an accuracy of 90 - 110% of the theoretical value. The acceptance criteria for standard samples were met for each exposure session.

The test item was used as the analytical standard.

Duration of treatment / exposure:

6 hours

Frequency of treatment:

5 days/week for 13 weeks, followed by 4 week recovery period for sub-group of Control and high dose

Dose / conc.:

0 ppm (nominal)

Remarks:

Group 1 - Control Group

Dose / conc.:

25 ppm (nominal)

Remarks:

Group 2 (165 mg/m3)

Dose / conc.:

75 ppm (nominal)

Remarks:

Group 3 (495 mg/m3)

Dose / conc.:

400 ppm (nominal)

Remarks:

Group 4 (2640 mg/m3)

No. of animals per sex per dose:

20 Control and high dose, 10 low and intermediate dose

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: dose concentrations based on previous OECD 422 inhalation study
- Post-exposure recovery period in satellite groups: 4 weeks

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily during dosing period, once daily during recovery period

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: twice daily during dosing period, once weekly during recovery period

BODY WEIGHT: Yes
- Time schedule for examinations: twice weekly during dosing period, once weekly during recovery period

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/animal/day: Yes
- Time schedule for examinations: weekly

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: before dosing started and Week 13
- Dose groups that were examined: all (not recovery animals in Week 13)

HAEMATOLOGY: Yes
- Time schedule for collection of blood: prior to necropsy at end of dosing or recovery period
- Anaesthetic used for blood collection: Yes (isoflurane)
- Animals fasted: Yes
- How many animals: 10 sex/group
- Parameters checked in table No. 1 were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: prior to necropsy at end of dosing or recovery period
- Animals fasted: Yes
- How many animals: 10 sex/group
- Parameters checked in table No. 2 were examined.

URINALYSIS: Yes
- Time schedule for collection of urine: 18 hour period prior to necropsy at end of dosing or recovery period
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes
- Parameters checked in table No. 3 were examined.

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes (see table)
HISTOPATHOLOGY: Yes (see table)

Statistics:

The following statistical methods were used to analyze the food consumption, body weight, ophthalmoscopic examinations, macroscopic findings, organ weights and ratios, as well as clini¬cal laboratory data:

• The two-sided Dunnett-test (many to one t-test) based on a pooled variance estimate was applied if the variables could be assumed to follow a normal distribution for the comparison of the treated groups and the control groups for each sex.

• The Steel-test (many-one rank test) was applied instead of the Dunnett-test when the data could not be assumed to follow a normal distribution.

• The two-sided Fisher's exact-test was applied to the ophthalmo-scopic and macroscopic findings.

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Ataxia and decreased activity were noted in all animals in the high dose from treatment start onwards. Ataxia was recorded until weeks 8 and 11 in males and females, respectively, and decreased activity was recorded until week 3 for both sexes. Based on adaptation the animals showed during the treatment and on the complete reversibility of the findings during recovery, these findings are considered to be non-adverse.

Mortality:

no mortality observed

Description (incidence):

There were no unscheduled deaths during the course of the study.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Lower body weight gain for high dose animals in both sexes during treatment period. An increased body weight gain was recorded in males during the recovery period.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

Lower for high dose males during treatment period and for high dose females during first two weeks of treatment period. An increased food intake was noted during recovery.

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not specified

Ophthalmological findings:

no effects observed

Description (incidence and severity):

No ophthalmoscopic findings were observed which were considered to be attributable to exposure with the test item

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

Total red blood cells and hematocrit were slightly but statistically significantly higher at the end of the treatment period in females at 400 ppm. These findings had completely reversed at the end of the recovery period.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

Sodium, potassium and chloride levels were higher at the end of the treatment period in females at 400 ppm. In addition, total protein was lower for those animals. These findings had completely reversed at the end of the recovery period.

Urinalysis findings:

effects observed, treatment-related

Description (incidence and severity):

The pH value was lower at the end of the treatment period in males at 400 ppm. This finding had completely reversed at the end of the recovery period.

Behaviour (functional findings):

not specified

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

Absolute and relative liver weights were higher in females at 400 ppm. In the absence of any histopathological findings in the livers and based on the reversibility
that was observed during the recovery period, this finding is considered to be adaptive.

Gross pathological findings:

no effects observed

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

In comparison to controls the incidence and severity of increased intra-epithelial hyaline droplets and focal or multifocal basophilic tubules was greater in all the male treated groups. In addition in 2/10 males given 400 ppm granular casts were present at the cortico-medullary junction of the kidneys. These findings were all considered to be consistent with a diagnosis of alpha-2µ-nephropathy and therefore of no toxicological significance to humans.

Histopathological findings: neoplastic:

no effects observed

Other effects:

not specified

Key result

Dose descriptor:

NOAEC

Effect level:

>= 400 ppm (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other:

Remarks on result:

not determinable due to absence of adverse toxic effects

Key result

Critical effects observed:

yes

Lowest effective dose / conc.:

400 ppm (nominal)

System:

urinary

Organ:

kidney

Treatment related:

yes

Dose response relationship:

not specified

Relevant for humans:

no

Table 1 - Exposure Conditions

Group	Temperature [°C]	Relative Humidity [%]	Oxygen Concentration [%]
1	22.4 ± 0.2 (n=66)	0.0 ± 0.0 (n=66)	20.2 ± 0.0 (n=66)
2	22.2 ± 0.1 (n=66)	3.0 ± 0.1 (n=66)	20.2 ± 0.0 (n=66)
3	22.1 ± 0.2 (n=66)	3.0 ± 0.5 (n=66)	20.0 ± 0.0 (n=66)
4	22.2 ± 0.1 (n=66)	3.9 ± 0.1 (n=66)	20.2 ± 0.1 (n=66)

For technical reasons, the relative humidity value of the chamber atmosphere was below the range given by the guidelines. The reason is the dried, pressurized air used for vapor generation, in combination with heating the test item in the nebulizer to 68 – 76 °C. This deviation from the guidelines has no influence on the results of the study, based on the absence of any findings in the respiratory tract and the circumstance that dry air generally worsens any findings in the respiratory tract due to loss of the protective mucus layer on the lining epithelium.

Table 2 - Test Atmosphere Concentrations

Group	Chemically Determined AtmosphereConcentration [ppm]	TargetAtmosphereConcentration [ppm]	Atmosphere ConcentrationsRelative to Target
2	25.0 ± 0.5 (n=66, CV=1.8%)	25	100.0%
3	75.0 ± 1.6 (n=66, CV=2.2%)	75	100.0%
4	400 ± 4 (n=66, CV=1.1%)	400	100.0%

 

Conclusions:

In a repeated dose toxicity study conducted according to OECD Test Guidelines 413 and in compliance with GLP, nose-only inhalation exposure of 1,1,1,3,3,3-hexamethyldisilazane (CAS 999-97-3, EC 213-668-5) at concentrations of 25, 75 or 400 ppm to Sprague-Dawley rats for 6 hours per day, 5 days per week for 13 weeks resulted in minor and reversible changes at the highest concentration. Renal effects in male rats were observed at all concentrations, but these findings were consistent with alpha-2u-nephropathy and therefore of no toxicological significance to humans. Based on the results in this study, the NOAEC was considered to be higher than or equal to 400 ppm, the highest concentration tested.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEC

2 640 mg/m³

Study duration:

subchronic

Species:

rat

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

In the key repeated dose toxicity study conducted according to OECD Test Guideline 413 and in compliance with GLP, Sprague-Dawley rats were exposed, nose-only, to 1,1,1,3,3,3-hexamethyldisilazane vapour (CAS 999-97-3, EC 213-668-5) for 6 hours a day, 5 days a week for 13 weeks, followed by a 4 -week recovery period, at 0, 25, 75 or 400 ppm.

Clinical signs of decreased activity and ataxia was recorded after dose administration but not the following day and was therefore considered not to be adverse.

Minor effects on body weight and food consumption recorded at 400 ppm at the end of the exposure period but not after the 4 -week recovery period were considered not to be adverse. The changes in some haematology, clinical chemistry and urinalysis parameters were slight and/or only noted in one sex and/or without pathological correlate and had completely reversed by the end of the recovery period so were considered not to be adverse. The increase in liver weight was in the absence of any histopathological changes in the liver and was reversible during the recovery period and so was considered to be adaptive and therefore not adverse. The renal changes, including increased incidence and severity of intra-epithelial hyaline droplets or multifocal basophilic tubules in males at all doses was considered to be consistent with alpha-2u-nephropathy and therefore of no toxicological relevance to humans (Swenberg, 1993). The NOAEC was therefore considered to be 400 ppm (equivalent to 2640 mg/m³), the highest concentration tested (Harlan, 2014).

In a dose range-finding study based on a protocol equivalent to OECD Test Guideline 412 with some deviations and no compliance with GLP, undiluted 1,1,1,3,3,3-hexamethyldisilazane was administered to Sprague-Dawley rats (5/sex/dose) as a vapour at target concentrations of 0, 30, 100 and 400 ppm for six hours/day on seven consecutive days. Body and feeder weights were recorded prior to the first exposure and on Day 8. Daily observations for clinical signs of toxicity, as well as mortality and morbidity were conducted. Each animal underwent a gross necropsy on Day 8. Post-exposure observations in the highest dose group (actual concentration 584 ppm) consisted of decreased to absent activity and severe uncoordinated gait, or the inability to walk. Decreased activity and slight uncoordinated gait were noted occasionally in a few of the mid dose group (actual concentration 307 ppm). In all cases the effects were transient and animals recovered before the next exposure. The gross necropsy revealed no abnormal findings. Mean body weights, on Day 8, in the 307 and 584 (statistically significant) ppm groups were reduced in a dose-dependent manner. Body weight gains in the 307 and 584 ppm groups were statistically significantly reduced in a dose-dependent manner, and in the highest dose group animals lost weight over the eight-day period. There was a statistically significant reduction in food consumption in 307 and 584 ppm females, and 584 ppm males. The NOAEC for 1,1,1,3,3,3-hexamethyldisilazane was concluded to be 107 ppm based on clinical observations and statistically significant and/or exposure-related differences observed for body weights, body weight gains and food consumption when compared to controls. The study has been used to provide a rational basis to select the exposure levels for the subsequent repeated dose inhalation toxicity with the reproduction/developmental toxicity study, which is described below (Dow Corning Corporation 2007).

In a supporting repeated dose inhalation toxicity with the reproduction/developmental toxicity screening test conducted according to OECD Test Guideline 422 and in compliance with GLP, Sprague-Dawley rats were exposed, whole body, to 1,1,1,3,3,3-hexamethyldisilazane vapour for 6 hours/day, 7 days/week for 4 weeks at 0, 25, 100 or 400 ppm. Each exposure group consisted of 10 male and 20 female rats. Females were further divided into a toxicity and reproductive group consisting of 10 animals each (the results of the reproductive and developmental toxicity elements of the study are presented in Section 7.8). Males were used to evaluate toxicity and reproductive parameters. Males and toxicity group females were treated for 29 and 28 days, respectively. Animals were observed throughout the exposure period for mortality and signs of toxicity, FOB and motor activity evaluations were conducted on males and toxicity phase females. Body weights and food consumption were monitored. Haematology and clinical chemistry investigations were conducted on blood samples taken at termination. Complete necropsies were performed on males and toxicity phase females, and selected organs were weighed. Microscopic examinations were performed on selected tissues from the control and high dose toxicity subgroup animals. There were no deaths or adverse findings in the FOB and motor activity tests. Clinical signs were limited to uncoordinated gait and decreased activity immediately after exposure in the highest exposure group. There was a statistically significant decrease in absolute body weights for females (-15%) with a corresponding decrease  in total weight gain compared to controls for both sexes (62 and 25% for females and males, respectively) in the highest dose group. Absolute body weights for the mid dose group females were identified as statistically decreased (-7%) from controls, however, total and percent weight gain were not different. Total food consumption was statistically decreased for the highest dose group males  and females. Increased red blood cell counts, hemoglobin, and haematocrit were noted in the highest dose group females. Decreased glucose along with increased cholesterol levels, were noted in the highest dose group females with cholesterol and sodium levels statistically increased in the highest dose group males. Organ weight changes included statistically significant decreases in absolute epididymides weight in the high dose males and lung weights in the high dose group females, with increases in kidney-to-body weight ratios for mid and high dose group females and high dose group males. Increased liver-to-body  weight ratios were also noted in the high dose group females. There were no gross findings. Histopathological examination of tissues and organs for control and high exposure animals demonstrated an increased incidence of centrilobular hypertrophy in the liver of high dose group females.  Evaluation of intermediate dose levels confirmed this finding was limited to high dose group females.  There were no other test article-related histopathological findings. The NOAEC for repeated dose toxicity endpoints for 1,1,1,3,3,3-hexamethyldisilazane, based on the clinical observation, body weight changes and histological findings was 100 ppm (660 mg/m³). There were no test article-related effects observed on reproductive and developmental parameters (Dow Corning Corporation, 2008).

There is another inhalation study conducted according to OECD Test Guideline 422 and in compliance with GLP on the hydrolysis product, trimethylsilanol (CAS 1066-40-6) in which three groups of Sprague-Dawley rats were exposed, via whole-body inhalation to vapor atmospheres of the test article for 6 hours/day, 7 days/week for 4 weeks, at 0, 60, 300 and 600 ppm.

Test substance-related effects were limited to changes in haematology (lower eosinophil and lymphocyte counts for males) and serum chemistry (higher alanine aminotransferase for males and toxicity phase females) at 600 ppm. These changes occurred in the absence of correlating histologic changes and were not considered adverse. Therefore, under the conditions of this screening study, an exposure level of at least 600 ppm (2214 mg/m3) was considered to be the NOAEC (WIL Research Laboratories, 2008).

Repeat dose information (OECD SIDS, 2007) for the non-silanol hydrolysis product, ammonia, indicates that in studies conducted by the inhaled route, the main effects are irritation and inflammation of the respiratory tract. Growth retardation and effects on the CNS were also noted in inhaled studies in rats conducted with ammonium sulfate. Although effects on the respiratory tract were not noted in the repeat dose inhalation study with 1,1,1,3,3,3-hexamethydisilazane, the clinical signs and body weight effects may be attributable to its rapid hydrolysis and production of ammonia and/or trimethylsilanol.

In a brief summary of a non-standard inhalation toxicity study (reliability score 4) the lowest Toxic Concentration for 1,1,1,3,3,3-hexamethyldisilazane was 98 mg/m³ in rats following 17 weeks of intermittent exposure.

There were no repeat dose data for 1,1,1,3,3,3-hexamethyldisilazane via the oral and dermal routes.

Justification for classification or non-classification

Based on the available data, 1,1,1,3,3,3-hexamethyldisilazane does not require classification for specific target organ toxicity according to Regulation (EC) 1272/2008.