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Repeated dose toxicity: inhalation

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Administrative data

Endpoint:
short-term repeated dose toxicity: inhalation
Remarks:
combined repeated dose and reproduction / developmental screening
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
18.03.2008 to 21.11.2008
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study was conducted according to an appropriate OECD test guideline, and in compliance with GLP
Cross-reference
Reason / purpose:
reference to other study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2008
Report Date:
2008

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories
- Age at study initiation: 11-12 weeks.
- Weight at study initiation: Males: 341 g to 418 g; Toxicity phase females:231 g to 282 g ; Reproductive phase females: 218 g to 280 g
- Fasting period before study: None
- Housing: F0 animals were housed individually in clean, stainless steel wire mesh cages suspended above cage board. The cage-board was changed at least 3 times per week. The males and reproductive phase females were paired for mating in the home cage of the male. Following positive evidence of mating, the males were housed in suspended wire mesh cages until the scheduled necropsy, and the females were transferred to plastic maternity cages with nesting material, ground corncob bedding
- Use of restrainers for preventing ingestion (if dermal): yes/no
- Diet (e.g. ad libitum): Ad libitum except during exposure periods, FOB and fasting period prior to blood sampling.
- Water (e.g. ad libitum): Ad libitum except during exposure periods, FOB and fasting period prior to blood sampling.
- Acclimation period: The males and reproductive phase females were housed for an acclimation period of 16 days prior to the first day of treatment. The toxicity phase females were housed for an acclimation period of 23 days prior to the first day of treatment.


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22± 3
- Humidity (%): 50± 20
- Air changes (per hr): At least 10
- Photoperiod (hrs dark / hrs light): 12/12


IN-LIFE DATES: From: 3 April 2008 To: 31 May 2008

Administration / exposure

Route of administration:
inhalation: vapour
Type of inhalation exposure:
whole body
Vehicle:
other: unchanged (no vehicle)
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus:
- Method of holding animals in test chamber:
- Source and rate of air:
- Method of conditioning air:
- System of generating vapour: Trimethylsilanol was generated for exposure as a vapor using a bubbler-type vaporization system located within a heated containment box. Nitrogen flowed into the inlet stem of a gas washing bottle containing an appropriate quantity of test article and bubbled through a fritted disc at the bottom of the bottle in order to create vapors of the test article. The concentrated vapors were piped to the heated chamber inlet where the concentration was reduced by mixing with chamber ventilation air. The control group was exposed to clean, filtered air.

- Temperature, humidity, pressure in air chamber:
- Air flow rate:
- Air change rate:
- Method of particle size determination:
- Treatment of exhaust air:


TEST ATMOSPHERE
- Brief description of analytical method used:
- Samples taken from breathing zone: yes/no


VEHICLE (if applicable)
- Justification for use and choice of vehicle:
- Composition of vehicle:
- Type and concentration of dispersant aid (if powder):
- Concentration of test material in vehicle:
- Lot/batch no. of vehicle (if required):
- Purity of vehicle:
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Actual exposure concentrations within each chamber were measured at approximately 35 minute intervals during each daily exposure period by a validated gas chromatographic method. Exposure atmosphere samples were delivered to the gas chromatograph from the approximate middle of each chamber by a pump and multi-port sampling valve. At least 1 standard was analyzed each day prior to exposure to confirm gas chromatographic calibration.
Homogeneity and temporal stability of the exposure concentrations were evaluated during the method development phase of the study. Four test locations and a reference location were used for these determinations. The test locations were top rear, top front, bottom rear and bottom front. Samples were collected and analyzed on the GC as rapidly as possible by alternating from the reference location to a test location. The measured concentration was calculated as a percent difference for each position from the reference location. Homogeneity was performed in triplicate for each test article exposure chamber.
Duration of treatment / exposure:
Six hours per day, seven days per week.
Frequency of treatment:
Males and reproductive phase females were exposed daily for at least 14 days prior to mating and continuing throughout mating for a minimum of 34 days (males) or through gestation day 20 (reproductive phase females). Toxicity phase females were exposed daily for a minimum of 28 days.
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
0, 61, 303 and 602 ppm (males), 0, 61, 304 and 604 ppm (toxicity phase females) and 0, 61, 304 and 603 ppm (reproductive phase females)
Basis:
analytical conc.
Remarks:
Doses / Concentrations:
0, 60, 300 and 600 ppm
Basis:
nominal conc.
No. of animals per sex per dose:
Ten
Control animals:
other: Filtered air

Examinations

Observations and examinations performed and frequency:
-CAGE SIDE AND DETAILED CLINICAL OBSERVATIONS: Yes
All rats were observed twice daily for moribundity and mortality. Individual clinical observations were recorded daily. Once prior to the initiation of exposure, on the first day of exposure and on a weekly basis throughout the study, all rats were observed outside the home cage in a standard arena and evaluated for changes in gait, posture and response to handling, as well as the presence of clonic or tonic movements, stereotypies or bizarre behavior. Observations were conducted prior to test article exposure during the treatment period and were not performed on the day of Functional Observational Battery (FOB) assessments. Animals visible in the exposure chamber were also observed for signs of toxicity at the mid-point of exposure (3 hours following beginning of exposure) and any positive findings were recorded. All animals were observed for signs of toxicity approximately 1 hour following the exposure period.

BODY WEIGHT: Yes
- Time schedule for examinations: Individual male and toxicity phase female body weights were recorded weekly, beginning 1 week prior to test article exposure, on the first day of exposure and weekly thereafter until the scheduled euthanasia. Body weights were also recorded for the toxicity phase animals assigned to FOB and locomotor activity assessments on the days of these evaluations. Individual reproductive phase female body weights were recorded weekly, beginning 1 week prior to test article exposure, on the first day of exposure and on a weekly basis thereafter until evidence of copulation was observed. Once evidence of mating was observed, reproductive phase female body weights were recorded on gestation days 0, 4, 7, 11, 14, 17 and 20 and on lactation days 1 and 4.

FOOD CONSUMPTION:
Individual food consumption was recorded on the corresponding weekly body weight days until pairing. Once evidence of mating was observed, reproductive phase female food consumption was recorded on gestation days 0, 4, 7, 11, 14, 17 and 20 and on lactation days 1 and 4.

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes, of males and toxicity phase females.
- Time schedule for collection of blood: At scheduled necropsy
- Anaesthetic used for blood collection: Yes (isoflurane)
- Animals fasted: Yes
- How many animals: All
- Parameters checked in table 2 were examined.

CLINICAL CHEMISTRY: Yes, of males and toxicity phase females.
- Time schedule for collection of blood: At scheduled necropsy.
- Animals fasted: Yes
- How many animals: All
- Parameters checked in table [No.2] were examined.

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION:
- Time schedule for examinations: During the week prior to initiation of test article exposure (baseline evaluation) and prior to exposure during study week 4 (the last week of the study), following a minimum of 28 days of exposure (females) or 34 days of exposure (males).
- Dose groups that were examined: All males and toxicity phase females
- Battery of functions tested: sensory activity / grip strength / motor activity / other: See Table 1 for complete list of parameters investigated.
Sacrifice and pathology:
GROSS PATHOLOGY: Yes (see table 3)
HISTOPATHOLOGY: Yes (see table 3)
Statistics:
Analyses were conducted using two-tailed tests (except as noted otherwise) for minimum significance levels of 1% and 5%, comparing each test article-treated group to the control group by sex. Each mean was presented with the standard deviation, standard error and the number of animals used to calculate the mean. Data obtained from nongravid females were excluded from statistical analyses following the mating period. Where applicable, the litter was used as the experimental unit. Mean body weights (weekly, gestation and lactation), body weight changes and food consumption, absolute and relative organ weights, clinical pathology values (except gamma glutamylransferase) and FOB data values were subjected to a parametric one way analysis of variance (ANOVA) to determine intergroup differences. If the ANOVA revealed statistically significant (p<0.05) intergroup variance, Dunnett's test was used to compare the test article-exposed groups to the control group. FOB parameters that yielded scalar or descriptive data and qualitative histopathological findings in the test article exposed groups were compared to the control group using Fisher’s Exact test. Gamma glutamyltransferase values were subjected to the Kruskal-Wallis nonparametric ANOVA to determine intergroup differences. If the ANOVA revealed statistically significant (p<0.05) intergroup variance, Dunn’s test was used to compare the test article-exposed groups to the control group.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
effects observed, treatment-related
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
not examined
Details on results:
CLINICAL SIGNS AND MORTALITY:
F0 males: All F0 males survived to the scheduled necropsy.
Toxicity phase females: One toxicity phase female in the 300 ppm group was found dead on study day 35, the day of scheduled necropsy. The cause of death for this female could not be determined and there were no clinical findings observed for this female during the study. However, because there were no mortalities at 600 ppm, this mortality at 300 ppm was not considered test substance-related.
Reproductive phase females: All reproductive phase females survived to the scheduled necropsy on lactation day 4, post-mating day 25 or post-cohabitation day 25. No test article-related clinical findings were noted.
In all groups no test article-related clinical findings were noted during the daily examinations, at the midpoint of exposure or approximately 1 hour following the end of exposure. Clinical findings were observed in the exposed groups during the daily examinations and approximately 1 hour following the end of exposure, and included hair loss, scabbing and/or red material on various body surfaces. However, these findings were observed in single animals, similarly in the control group and/or in a manner that was not exposure related. Therefore, no relationship to the test article was apparent.

BODY WEIGHT AND WEIGHT GAIN:
F0 males: Mean body weights and body weight gains for all groups were similar to the control group throughout the premating (study days 0-13) and entire exposure (study days 0-34) periods. No statistically significant differences were observed. During study days 27-34, mean body weight losses were noted for all groups, including the control group. This was likely due to the fasting period (study day 33-34) prior to blood collection for clinical pathology evaluations.
Toxicity phase females: Differences from the control group were slight and not statistically significant. Like males there were fasting period related mean body weight losses in all groups, including the controls.
Reproductive phase females: No differences between control and treated animals.

FOOD CONSUMPTION:
F0 males, toxicity phase and reproductive phase females: No statistically significant differences were noted.
HAEMATOLOGY:
F0 males: Mean absolute eosinophil and lymphocyte counts for the 600 ppm group males were 42.9% and 26.4% lower, respectively, than the control group. The difference for mean absolute eosinophils was significant (p<0.05). Because these changes were within historical limits for approximately age-matched animals (0.05-0.25 and 5.20 13.78 thousand/µL for eosinophils and lymphocytes, respectively) and did not correlate with any other clinical or pathological consequences, the decreased absolute eosinophil and lymphocyte counts were considered test article-related but nonadverse.
Toxicity phase females: A significantly (p<0.05) higher mean corpuscular volume was noted for the 600 ppm group (55.8 fL) when compared with the control group (53.9 fL). This finding was not considered to be test substance related because there were no correlating clinical pathological findings. Furthermore, the change was considered toxicologically insignificant because the small magnitude of change and because the increase in MCV fell within the WIL historical control data range.

CLINICAL CHEMISTRY:
F0 males: The mean alanine aminotransferase value for the 600 ppm group males was 20.5% higher than the control group. The difference was significant (p<0.05). While this change was considered test article-related, it was not considered adverse because the magnitude of change from the control group was small, the value was within historical limits and it did not correlate with any other clinical or pathological abnormalities.
Toxicity phase females: The mean alanine aminotransferase value for the 600 ppm group females was 11.1% higher than the control group, but did not achieve statistical significance. While this change was considered test substance-related, it was not considered adverse because the magnitude of change from the control group was small, the value was within historical limits and it did not correlate with any other clinical or pathological abnormalities. In addition, a significantly (p<0.05) lower creatinine level was noted for the 300 ppm toxicity phase females when compared with the control group. However, because this decrease was not observed in an exposure-related manner, no relationship to the test substance was apparent.

NEUROBEHAVIOUR:
F0 males: No test article-related findings were observed for F0 males at any exposure level during the functional observational battery. One male in the 600 ppm group was observed with Straub tail (increased tail elevation). However, this finding was only noted in 1 of the 20 animals observed. Therefore, no relationship to the test article was apparent.
Locomotor activity patterns (total activity as well as ambulatory activity counts) in F0 males were unaffected by test article exposure during the study week 4 evaluation. While no statistically significant differences in locomotor activity were observed during the exposure period, mean ambulatory counts for the 60 ppm group males were significantly (p=0.003) higher than the control group during the first 10-minute subinterval of the pretest evaluation. Because this change occurred prior to the initiation of the exposure period, there was no relationship to the test article. In addition, mean total motor activity counts for the overall 60-minute session were 24.9% higher for the 600 ppm males when compared to the control group during study week 4. However, this apparent increase was primarily due to 1 male that also had Straub tail. Therefore, no relationship to the test article was apparent.
Toxicity phase females: there were no test substance-related effects.

ORGAN WEIGHTS:
F0 males: No test substance-related changes.
Toxicity phase females: Higher mean adrenal gland, kidney and liver weights relative to final body weight were noted for the 600 ppm group when compared with the control group. Although the differences in mean adrenal gland weight achieved statistical significance (p<0.01), the value for the 600 ppm group (0.031 g) was within the range of the WIL historical control data (0.02 g-0.05 g) and was not considered toxicologically important. The increases in mean liver and kidney weights relative to final body weight were not toxicologically significant and fell within the WIL historical control data range. In addition, the elevation in mean liver and kidney weights in the high dose females may in part be due to one animal with histopathologic findings consistent with Caroli’s disease, which is a combination of polycystic kidney disease and bile duct dilatation. These findings in the liver and kidney of this particular animal were associated with relatively high liver and kidney weights, which would have contributed to the statistically significant higher mean kidney and liver weights (relative to final body weight) in this group.

GROSS PATHOLOGY:
F0 males: No test substance-related internal findings.
Toxicity phase females: One toxicity phase female in the 300 ppm group was found dead on study day 35 (day of scheduled necropsy). This female had no macroscopic findings that indicated a cause of death. There were no test substance-related changes.
Reproductive toxicity phase females: No test substance-related changes.

HISTOPATHOLOGY:
F0 males: No test substance-related changes.
Toxicity phase females: There were no test article-related histopathologic findings in toxicity phase females at any exposure level. One 600 ppm group female was noted with lesions of the kidney and liver compatible with congenital polycystic kidney disease and cystic dilatation of intrahepatic bile ducts. This combination of changes is a congenital autosomal recessive disease recognized in Crl:CD rats and therefore, these occurrences in 1 animal were considered unrelated to test article exposure.

Effect levels

Dose descriptor:
NOAEL
Effect level:
>= 600 ppm (nominal)
Sex:
male/female
Basis for effect level:
other: No toxicologically significant test substance-related adverse effects were observed at any dose tested.

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
In this OECD test guideline 422 screening study, which was conducted to GLP, test substance-related effects were limited to changes in hematology (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 600 ppm was considered to be the no-observed-adverse-effect level (NOAEL). It is considered appropriate to use this result in support of the repeated dose endpoint for general systemic toxicity of chlorotrimethylsilane as this substance is hydrolysed very rapidly in the presence of moisture to trimethylsilanol and hydrogen chloride. Since chlorotrimethylsilane is hydrolysed to trimethylsilane and hydrogen chloride, additional local irritation can be expected due to the acidic nature of the HCl.
Executive summary:

Three groups of Crl:CD(SD) rats, each group consisting of 10 males, 10 toxicity phase females and 10 reproductive phase females, were exposed via whole-body inhalation to vapor atmospheres of the test article, trimethylsilanol, 6 hours/day, 7 days/week. Target exposure concentrations were 60, 300 and 600 parts per million (ppm). A concurrent control group was exposed to filtered air on a comparable regimen. F0 males and reproductive phase females were exposed daily for 14 days prior to mating, throughout the mating period and continuing through the day prior to euthanasia (males) or gestation day 20 (reproductive phase females). F0 reproductive phase females with no evidence of mating or that failed to deliver were exposed through post-mating or post-cohabitation day 24 (the day prior to euthanasia) for a total of 52 consecutive days. F0 toxicity phase females were exposed for 28 consecutive days.

All animals were observed twice daily for mortality and moribundity. Clinical observations, body weights and food consumption were recorded at appropriate intervals.  Functional observational battery (FOB) and locomotor activity data were recorded for all males and toxicity phase females prior to initiation of exposure for each sex (baseline evaluation) and near the end of exposure. F0 males were euthanized following completion of the mating period and F0 toxicity phase females were euthanized following 28 days of exposure. Clinical pathology evaluations (hematology and serum chemistry) were performed on all F0 males and toxicity phase females at necropsy. Complete necropsies were conducted on all F0 males and toxicity phase females, and selected organs were weighed; selected tissues in the control and high‑exposure groups were examined microscopically.

All F0 reproductive phase females were allowed to deliver and rear their pups until lactation day 4. F1clinical observations and body weights were recorded on postnatal days (PND) 1 and 4. Reproductive phase dams and pups were euthanized and examined on lactation day/PND 4.

One F0 toxicity phase female in the 300 ppm group was found dead on study day 35, the day of scheduled necropsy.  While the cause of death for this female was not determined, there were no mortalities in the 600 ppm group.  Therefore, this mortality at 300 ppm was not attributed to test article exposure.  All other toxicity phase females and all F0males and reproductive phase females survived to the scheduled necropsies.  There were no test article-related macroscopic findings or effects on organ weights.There were no remarkable clinical observations noted at any exposure level.  Mean body weights, body weight gains and food consumption for males and females at all exposure levels were similar to the control group for all phases.  There were no test article-related effects on FOB parameters, including home cage, handling, open field, sensory, neuromuscular or physiological observations and no effects on locomotor activity patterns for males and toxicity phase females.  No remarkable shifts in the pattern of habituation occurred during locomotor activity assessments in any of the test article‑exposed groups when the F0 males and toxicity phase females were evaluated prior to the initiation of exposure or during the study week 4 evaluation.

Decreased mean eosinophil and lymphocyte counts were noted for the 600 ppm group F0 males.  In addition, increased mean alanine aminotransferase values were noted for F0 males and toxicity phase females in the 600 ppm group when compared with the control group.  Although these changes were considered test article‑related, they were not considered adverse in the absence of correlating histopathology.

Test article-related effects were limited to changes in hematology (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 600 ppm was considered to be the no-observed-adverse-effect level (NOAEL) for reproductive, systemic and neonatal toxicity of trimethylsilanol when administered via whole-body inhalation exposure to Crl:CD(SD) rats.