Registration Dossier

Administrative data

Description of key information

Repeated dose toxicity:
Oral (subacute):
Repeated dose toxicity: OECD TG 422 in rats: LOAEL = 100 mg/kg bw/day.
At all dose levels (100, 250 and 500 mg/kg bw/day) during the histopathology examination, perivascular lymphoid cell infiltration and transitional cell hyperplasia of the urinary bladder were observed in males and females.
Inhalation (subacute):
Repeated dose toxicity: OECD TG 412 in rats: NOAEC > 620 mg/m³.
No systemic toxicity was observed after repeated exposure to concentrations up to 620 mg/m³.
Dermal toxicity: no measured data are available.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: oral
Remarks:
combined repeated dose and reproduction / developmental toxicity screening
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2009-01-14 to 2009-10-20
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
Remarks:
no significant deviation
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
other: Han Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Labs Ltd., Wölferstrasse 4, 4414 Fűllinsdorf, SWITZERLAND
- Age at study initiation: 11 wk
- Weight at study initiation: 294-330 g (m), 178-213 g (f)
- Housing: 1/Makrolon type 3 cage
- Diet: standard diet ad libitum
- Water: drinking water ad libitum
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 20-70
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 2009-01-14 To: 2009-04-06
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS:
Prepared weekly. Homogeneous suspension maintained with magnetic stirrer during dosing .
VEHICLE
Dried deacidified corn oil
- Justification for use and choice of vehicle (if other than water): none given
- Amount of vehicle (if gavage): dose volume 5 mL/kg bw
- Lot/batch no.: 37899577
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
CG-FID. Stability and homogeneity verified at start of study and during 2nd last week of dosing.
Duration of treatment / exposure:
toxicity males: from 2 weeks prior to mating for at least 4 wk
toxicity/reproductive females: from 2 weeks prior to mating for at about 7 wk
Frequency of treatment:
daily. 7 days/wk
Dose / conc.:
100 mg/kg bw/day (actual dose received)
Dose / conc.:
250 mg/kg bw/day (actual dose received)
Dose / conc.:
500 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
toxicity males: 10
toxicity/reproductive females: 10
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: prior range finding study (Harlan B88762)
- Rationale for animal assignment (if not random): random with consideration for body weight
Positive control:
no
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly

BODY WEIGHT: Yes
- Time schedule for examinations: daily

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined weekly

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: just prior to sacrifice
- Anaesthetic used for blood collection: Yes (isoflurane)
- Animals fasted: Yes (18 h)
- How many animals: 5

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: just prior to sacrifice
- Animals fasted: Yes (18 h)
- How many animals: 5

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION/FOB: Yes
- Time schedule for examinations: just prior to sacrifice
- Dose groups that were examined: 5/sex, all groups
Cage-side observations: unusual body movements (e.g. tremors, convulsions), abnormal behavior (e.g. circling, stereotypy) and posture as well as resistance to removal.
Hand-held observations: palpebral closure, pinna reflex, lacrimation, pupil size, pupil reactivity, salivation, muscle tone, extensor thrust response, righting reflex and reaction to handling.
Open field observations: level of ambulatory activity including rearing (one minute evaluation), responsiveness to sharp noise, paw pinch, gait evaluation, quantity of urine and fecal pellets voided.
Categorical observations (can be made any time during the FOB): hair coat, behavior, respiration, muscle movements, eyes, hearing ability (Preyer’s reflex), urine or feces, soiling, general abnormalities, posture.
Measurements / Counts: hind limb / fore limb grip strength, landing foot splay, rectal temperature.



Sacrifice and pathology:
GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes (range of tissues as specified in OECD 422)
Statistics:
Means and standard deviations of various data were calculated. The 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. Fisher's exact-test was applied to breeding data and the macroscopical findings.
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
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:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
not examined
Details on results:
CLINICAL SIGNS AND MORTALITY
No treatment-related effects.

BODY WEIGHT AND WEIGHT GAIN
In males: at 500 mg/kg bw/day mean body weight gain was statistically significantly reduced during the pre-pairing period. This resulted in a statistically significantly decrease in body weight throughout the whole study. In females: at 500 mg/kg bw/day mean body weight gain was statistically significantly reduced between Days 8 and 14 of the gestation resulting in a decrease of mean body weight between Days 11 and 20 of gestation.

HAEMATOLOGY
No treatment-related effects.

CLINICAL CHEMISTRY
In males, at 250 and 500 mg/kg bw/day, the concentration of urea was statistically significantly increased in dose-dependent manner. The concentration of bile acids was statistically significantly increased but without showing a dose-dependent pattern. At 500 mg/kg bw/day, the concentration of cholesterol was also statistically significantly increased.

NEUROBEHAVIOUR/FOB
No treatment-related effects.

ORGAN WEIGHTS
The report highlights that at 500 mg/kg bw/day, absolute and relative weight of the kidneys was statistically significantly increased in males. In addition, in females at this dose, thymus weight was reduced (absolute, relative to body and relative to brain), although these values were within the range of historical controls. Female liver and kidney weights (relative to body) were also increased at the top dose; again within the range of historical controls. [The report incorrectly notes increased thymus and reduced liver weights for this group.]

GROSS PATHOLOGY
The urinary bladder was thickened in all treated groups (100 mg/kg bw/day 6/10 m, 2/10 f; 250 mg/kg bw/day 7/10 m, 6/10 f; 500 mg/kg bw/day 9/10 m, 7/10 f). This finding in the urinary bladder was said mainly to correlate with transitional cell hyperplasia observed at microscopic level.

HISTOPATHOLOGY: NON-NEOPLASTIC
Microscopically, the test item-related lesions recorded were:
Kidneys: At 500 mg/kg bw/day, multifocal tubular degeneration/regeneration was noted in all males and two females, and in one male this was associated with minimal hyaline casts. Tubular simple dilation was noted in four males and all females. Increased incidence of focal tubular degeneration/regeneration was observed in females. Transitional cell hyperplasia was noted in all males and females. At 250 mg/kg bw/day, multifocal tubular degeneration/regeneration was observed in three males. Transitional cell hyperplasia was noted in all males and females. The hyperplastic lesions were accompanied by an increased incidence of renal pelvic dilation in six/six males and five/five females and correlated the hyperplastic findings in urinary bladders.
Urinary Bladder: Perivascular lymphoid cell infiltration was noted in ten/ten males and six/seven females at 100 mg/kg bw/day, in ten/ten males and seven/eight females at 250 mg/kg bw/day and in seven/ten males and ten/ten females at 500 mg/kg bw/day. At all dose levels, minimal to moderate transitional cell hyperplasia was observed in all males and females. These hyperplastic lesions were accompanied by minimal to slight dilation in eacheight/ten males at 100 and 250 mg/kg bw/day, and ten/ten males at 250 mg/kg bw/day. In females, dilation in three/seven at 100 mg/kg bw/day, four/eight at 250 mg/kg bw/day, and ten/ten at 500 mg/kg bw/day. Minimal to moderate bladder congestion in all males at all dose levels. In females, six/seven at 100 mg/kg bw/day, seven/eight at 250 mg/kg bw/day, and ten/ten at 500 mg/kg bw/day, in each one male and one female at 250 mg/kg bw/day associated with slight hemorrhage.
Jejunum: At 500 mg/kg bw/day, multifocal lymphangiectasis of villi was noted in all males and females and in two males and one female at 250 mg/kg bw/day.
Liver and Thyroid: At 500 mg/kg bw/day, the liver cell hypertrophy noted in two/five females and consequent increase of follicular cell hypertrophy in the thyroid gland was considered to be an adaptive effect and therefore, not adverse.

HISTORICAL CONTROL DATA (if applicable)
See comment on organ weights above.

OTHER
Measured doses were in the range 94-103% of nominal.
Dose descriptor:
NOAEL
Sex:
male/female
Basis for effect level:
other: effects in urinary bladder at all doses (>=100 mg/kg bw/day)
Remarks on result:
not determinable
Remarks:
no NOAEL identified
Dose descriptor:
LOAEL
Effect level:
100 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: effects on urinary bladder
Critical effects observed:
yes
Lowest effective dose / conc.:
100 mg/kg bw/day (actual dose received)
System:
urinary
Organ:
bladder
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
not specified

Increased urea and cholesterol can both be indicative for kidney effects.

At all dose levels (100, 250 and 500 mg/kg bw/day) during the histopathology examination, perivascular lymphoid cell infiltration and transitional cell hyperplasia of the urinary bladder were observed in males and females.Therefore, based on the findings in urinary bladder noted in all test item-treated groups, a general NOAEL could not be established.

Conclusions:
A well reported oral combined repeated dose/reproductive and developmental screening study, conducted according to OECD 422 and in accordance with GLP, did not identify a NOAEL for the registered substance; effects to the urinary bladder were reported at the lowest tested dose of 100 mg/kg bw/day.
Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LOAEL
100 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
The study was conducted according to the appropriate OECD test guideline, and in compliance with GLP.
System:
urinary
Organ:
bladder

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1979-06-11 - 1979-07-12
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
It was not compliant with GLP. Due to an accident, the mid dose group had to be terminated and the respective exposure was repeated in a following experiment.
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 412 (Subacute Inhalation Toxicity: 28-Day Study)
Deviations:
yes
Remarks:
(longer exposure duration (6.5 h) and no detailed information on test material purity)
GLP compliance:
no
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Bio-Breeding Laboratories Ltd., Ottawa, Ontario, Canada (received on 1979-05-24)
- Age at study initiation: 10-14 weeks
- Weight at study initiation: 200-250 g
- Housing: individually in in wire mesh-bottomed metal cages
- Diet: standard commercial laboratory diet (Purina) ad libitum (no food was available for the animals during exposure)
- Water: tab water available through a water sipper system ad libitum (no water was available for the animals during exposure)
- Acclimation period: 2 weeks
- Subsequent to arrival the animals received a qulified vetinary aid to ensure "normal" health status.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): animal room: 20.0-23.9, inhalation room: 21.4-23.6
Route of administration:
inhalation: vapour
Type of inhalation exposure:
whole body
Vehicle:
other: clean, dry air
Remarks on MMAD:
MMAD / GSD: not applicable
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: stainless steel whole body exposure chambers (400 l volume)
- Method of holding animals in test chamber: in a stainless steel wire mesh compartmentalised cage (each compartment measuring 7 x 4 x 3 inches)
- System of generating aerosols: Delivery of the test article into the chambers was accomplished by forcing air through gas generators fittet with sintered glass frits and containing the test item, and metering vapours into the chamber.
- Temperature, humidity, pressure in air chamber: Airflow through each chamber was measured in the exhaust line of the chamber by reading the pressure drop across an orifice plate on a magnehelic gauge. These gauges were calibrated against conventional ball-type flow meters. The magnitude of negative pressure within each chamber with respect to the outside atmosphere was always less than 0.5 inches of water. Temperature and relative humidity within both the inhalation chamber and the inhalation room were measured at hourly intervals during each exposure period. Chamber temperature and relative humidity were measured by means of remote sensors placed within each inhalation chamber. A mean daily chamber temperature of 22-26 °C was maintained throughout the study with the exception of study days 0.1 and 2.1 when higher temperatures were recorded in Chambers 1 to 2 and 3, respectively. Inhalation chamber relative humidity during exposure was generally 35-65% A higher relative humidity was observed in Chamber 1 (air controls) compared with the inhalation room, and a decrease in chamber relative humidity tended to occur as chamber test substance concentration increased.
- Air flow rate: One gas generator was used for Chamber 2 with an airflow of 6 l/min and two generators in parallel with an airflow of 30 l/min for Chamber 3. In the case of Chamber 4 a total airflow of 40 l/min was passed through two parallel generators partially immersed in a water bath at approximately 65 °C. The two generators were each serially connected to a pair of generators maintained at room temperature. The outputs from the vapour generation systems were each diluted with room air to a total chamber airflow of 45 l/min.

TEST ATMOSPHERE
- Brief description of analytical method used: Nominal chamber concentrations were calculated from the total airflow and the weight loss from the generator(s) during the exposure. Actual chamber test material concentrations were measured hourly using a MIRAN 1A Infrared gas analyser at an analytical wave length of 9.2 µm and a path length of 0.75 m. The Infrared gas analyser had been previously calibrated by injection of known microliter quantities of the test article into a closed loop system and allowing the liquid to vaporise.
- Samples taken from breathing zone: yes
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Target chamber concentrations for Groups 2 and 3 were generally achieved by the end of the 30 min equilibration time. In case of Group 4 approximately two hours of equilibration were required before the chamber concentration reached the targeted value. This was concluded by the study authors to be possibly related to the low volatility of the test material.
Mean daily actual chamber concentrations were close to the target concentrations on all exposure occasions except for Chamber 3 on study day 1.4 when the mean test article concentration reached a recorded maximum of 134 ppm. This overexposure was found to result from a technical error. The measured concentration in Chamber 1 (control chamber) was approximately 2 ppm throughout the study although no test article was introduced into this chamber. The possible reason for this spurious value was concluded to be caused by infrared absorbance at the analytical wavelength by vapours emanating from rat faeces and urine in the chambers. Consequently, the chamber test article concentrations should be considered as 2 ppm higher than the true concentrations existing in the chambers.
Duration of treatment / exposure:
4 weeks
Frequency of treatment:
6.5 h/day, 5 days/week
Each daily exposure period was 6.5 hours in duration, starting after the 30 min equilibration time when 95% of the target concentration was established within the chamber. After 6.5 hours of exposure at the desired concentration, delivery of the test article into the chamber was stopped and the chamber was allowed to equilibrate with room air for 30 min prior to the removal of the animals. Thus, the animals remained in the chamber for 7.5 hours/day.
Dose / conc.:
10 ppm (nominal)
Dose / conc.:
50 ppm (nominal)
Dose / conc.:
80 ppm (nominal)
Dose / conc.:
11.5 ppm (analytical)
Remarks:
mean analytical conc. over the whole study period
Dose / conc.:
54.4 ppm (analytical)
Remarks:
mean analytical conc. over the whole study period
Dose / conc.:
78.5 ppm (analytical)
Remarks:
mean analytical conc. over the whole study period
Dose / conc.:
9.5 ppm (analytical)
Remarks:
corrected mean analytical cnc. over the whole study period due to additional absorbance of vapours emanating from rat faeces and urine in the chambers
Dose / conc.:
52.4 ppm (analytical)
Remarks:
corrected mean analytical cnc. over the whole study period due to additional absorbance of vapours emanating from rat faeces and urine in the chambers
Dose / conc.:
76.5 ppm (analytical)
Remarks:
corrected mean analytical cnc. over the whole study period due to additional absorbance of vapours emanating from rat faeces and urine in the chambers
Dose / conc.:
77 mg/m³ air (analytical)
Remarks:
corrected mean analytical conc. over the whole study period due to additional absorbance of vapours emanating from rat faeces and urine in the chambers; values calculated with (molecular weight [g/mol] x mean corrected exposure conc. [ppm])/24.45
Dose / conc.:
425 mg/m³ air (analytical)
Remarks:
corrected mean analytical conc. over the whole study period due to additional absorbance of vapours emanating from rat faeces and urine in the chambers; values calculated with (molecular weight [g/mol] x mean corrected exposure conc. [ppm])/24.45
Dose / conc.:
620 mg/m³ air (analytical)
Remarks:
corrected mean analytical conc. over the whole study period due to additional absorbance of vapours emanating from rat faeces and urine in the chambers; values calculated with (molecular weight [g/mol] x mean corrected exposure conc. [ppm])/24.45
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
Group 1: vehicle control, Group 2: 10 ppm exposure, Group 3: 50 ppm exposure, Group 4: 80 ppm exposure

During exposure an incident occured in Group 3. The line supplying suppressed air to the Chamber 3 gas generators was improperly connected resulting in test article being sprayed over ten animals in the chamber. Four rats were thoroughly soaked with test article, and the chamber concentration of the test substance increased to 134 ppm. Two rats were semi-conscious and ataxic within 45 min, whereas one female rat died after approximately 8 hours. Ten rats were observed to have wet fur on their ventral surface post-treatment and two of them were semi-conscious and lethargic. Three rats were found dead the next morning, and the deaths, as well as the lethargy and the ataxia, were concluded by the study authors to be attributed to an acute toxic effect of the dermal absorption of the test article.
As a consequence of this accident a second group of twenty rats were treated with the test article at 50 ppm and the data generated was presented as an addendum to this report (Bio-Research Laboratories No. 1981-I-0065-1573-6 (Addendum), 1981).
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily; druing the treatment period after each exosure, and additionally during exposure for signs of overt toxicity

DETAILED CLINICAL OBSERVATIONS: No

BODY WEIGHT: Yes
- Time schedule for examinations: Individaul body weights were measured twice weekly during both acclimatisation and treatment periods.

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at study termination
- Anaesthetic used for blood collection: No
- Animals fasted: Yes
- How many animals: all animals
- Parameters checked: red blood cell count, haemoglobin concentration, packed cell volume, and total and differential leucocyte count

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at the termination of the treatment period
- Animals fasted: Yes
- How many animals: all
- Parameters checked: fasting serum glucose, blood urea nitrogen (BUN), serum alkaline phospahtase (SAP), serum glutamic oxalacetic transaminase (SGOT), and serum glutamic pyruvic transaminase (SGPT)

URINALYSIS: Yes
- Time schedule for collection of urine: during the night prior to terminal necropsy
- Metabolism cages used for collection of urine: Yes
- Animals fasted: No data
- Parameters checked: volume, colour, odour, transparency, specific gravity, occult blood, ketones, glucose, protein, pH, bilirubin, and microscopic examination of urinary sediment
Sacrifice and pathology:
GROSS PATHOLOGY: Yes (complete gross pathology of all animals)
HISTOPATHOLOGY: Yes (brain (cerebrum, cerebellum, medulla, and pons), spinal cord, olfactory nerve and/or olfactory bulb, both eyes and optic nerves, pituitary gland, thyroide, thymus, adrenal glands, heart, spleen, thoracic aorta, oesophagus, submandibular salivary gland, stomach, duodenum, jejunum, ileum, cecum, colon, rectum, liver, pancreas, gonads, lymph nodes (mesenteric and mediastinal), uterus, prostate, both kidneys, urinary bladder, sciatic nerve, skeletal muscle, bone marrow section (sternum), skin, tongue and buccal mucosa (bilateral), nasal and paranasal sinuses with vestibulum and adjacent bone, nasal septum, larynx, trachea, and major bronchi)
ORGAN WEIGHTS: Yes (left and right adrenal glands, brain, left and right gonads, heart, left and right kidneys, liver, pituitary gland, spleen, lungs and thyroid)
Other examinations:
MYELOGRAMS: Femoral bone marrow smears and sections from all control and high dose animals were examined by a qualified haematologist.
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 examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
no effects observed
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Details on results:
CLINICAL SIGNS AND MORTALITY
No abnormal clinical observations were noted for the animals of Groups 1-4 during exposure except for the animals of Group 3 as a consequence of the accident as described above. The majority of abnormal clinical observations recorded during the daily physical examinations consisted of signs of pulmonary congestion. Both treated and control groups exhibited a low incidence of congestion throughout the study, suggesting that this finding was unrelated to the treatment with the test item. Incidental observations included reddish brown fur discolouration of the perinasal and periorbital regions. Additionally, Groups 2 and 3 showed wet fur. The low incidence of these observations coupled with the absence of a wet fur in Group 4 animals post treatment and the presence of reddish brown fur discolouration perinasally and periorbitally in three control rats pretreatment suggests that exposure to the test item at concentrations up to 80 ppm had no adverse effect.

BODY WEIGHT AND WEIGHT GAIN
Statistical analysis (Dunnett’s t test), comparing the mean body weights of treated and control animals, did not reveal any significant differences between groups which could be attributed to treatment with the test article. Body weight of treated and control rats increased in a similar fashion throughout the study with the exception of several animals, which lost weight for a short period of time: one control male, one female of the 10 ppm dose group, and one male and female each of the high dose group, whereas the two latter animals showed congestion during these periods and the effect might be related to the observed weight loss. Due to a lack of time- and dose-dependence of the observed weight loss and the additional occurrence in the control group, it is suggested that the effects are not treatment related.

FOOD CONSUMPTION
No significant differences were observed between the absolute food consumption of treated and control groups. With respect to relative food consumption, significantly greater food intake was observed in Group 4 females during week 3 compared with the control group. A review of the individual data reveals that this is mainly contributed to one female animal of Group 4. This period of high food intake followed several weeks of reduced food consumption during which the body weight had decreased.
Infrequent periods of reduced food intake were noted in several rats of both the control and the high dose group and occurred at the same time as the body weight loss previously mentioned. The authors conclude that the effects seen are not related to the test material exposure.

HAEMATOLOGY
No significant differences in blood parameters existed between treated and control groups. Lymphocyte count was slightly elevated in some Group 4 animals, while segmented neutrophil count was slightly lower in Group 4 males and females as compared to controls. These small differences were not considered to be of any biological importance and the test material was concluded not to cause any adverse haematological effect in rats.

CLINICAL CHEMISTRY
The mean blood serum glucose level of Group 2 (10 ppm) females was significantly higher than that of the control group. Since no such effects were observed at higher doses, this effect was considered to be not treatment related. Although some individual values fell outside the range of normal values (e. g. increased BUN in one male of the control group, increased SAP in one male of the low dose group, increased SGOT in one female of the control group and one male of the high dose group (the latter in combination with an increased SGPT level)) none of these findings was considered to be exposure related.

URINALYSIS
Large intersubject variations existed in each group with respect to urine volume and specific gravity. However, the range for these parameters was similar in both treated and control groups. The high urine volume and associated low specific gravity in a proportion of the animal was considered possibly to be due to leakage of water from the water sipper in each cage. Hence, the test item was concluded not to cause any effect on urinary parameters measured in this study.

ORGAN WEIGHTS
The left adrenal gland (but not the right) of Group 4 (80 ppm) males and the spleens of Group 2 (10 ppm) males were significant lighter than those of control rats. There were no differences compared with the controls for the spleens of the high dose males. Hence, the slight effects observed were considered not to be treatment related. Individual data revealed that one male rat of the high dose, which had lost weight in the last week of the study, had high relative lung and kidney weights. One male each of the control group ant the low dose group had small left thyroide lobes and on male rat of the low dose group had small testes in terms of both absolute and relative weight. One female of the high dose group had small ovaries, one female of the low dose group had a small spleen, and two males of the low dose group had large lungs in terms of relative weight. Since these observations were isolated, they were not considered treatment related.

GROSS PATHOLOGY
The most commonly observed gross pathological findings were pulmonary congestion, enlargement and/or congestion of the lymph nodes and enlargement and/or fluid in the uterus and surrounding the ovaries of females. Each of these findings appeared as frequently in the treated groups as in the control group with the exception of fluid surrounding the ovary. The frequency of the latter finding was not dose related, and hence, gross pathological findings were concluded not to be treatment related. Further incidental findings not associated with the treatment were small testes or spleen, focal to general discolouration of the lung, liver, and kidney, and congestion of the thymus and salivary glands.

HISTOPATHOLOGY: NON-NEOPLASTIC
Pulmonary lesions were observed in both treated and control animals and consisted of lymphatic infiltration about air passages and blood vessels, some bronchiectasis, interstitial fibrosis, neutrophilic exudation, and alveolar macrophages. Similar lymphatic inflammatory was observed in the trachea, the larynx, and air passages. In most cases, lymphocytic aggregates were focal or multifocal in distribution and substantial amount of pulmonary parenchyma was intact and uninvolved. These changes were possibly the result of a mild to moderate murine respiratory mycoplasmosis. In some animals of both control and treated groups there was evidence of pulmonary oedema with accumulation of an eosinophilic proteinaceous material in alveolar spaces. This change was considered to be of agonal and post mortem occurrence. Hence, the test material was concluded not to produce any pathological changes.

MYELOGRAMS: There were no differences in cell/fat and myeloid/erythroid ratios between control and treated animals. Megakaryocytes were plentiful in all rats and there was no evidence of tumour or granuloma infiltration. Bony spicules were normal in treated and untreated animals alike. The eosinophilia observed in one high dose female could be associated with extramedullary haematopoiesis in the liver of this animal, however, the eosinophilia, neutrophilia, and lymphocytosis observed in other control or treated animals cannot be correlated with any histological changes. Hence, the test material was concluded not to cause effects on bone marrow haematopoiesis.
Dose descriptor:
NOAEC
Effect level:
>= 80 ppm (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: highest dose tested
Dose descriptor:
NOAEC
Effect level:
>= 620 mg/m³ air (analytical)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: highest dose tested
Critical effects observed:
no
Conclusions:
The test item was tested for repeated inhalation toxicity similar or equivalent to the OECD TG 412, but not in compliance with GLP. The test article did not exhibit any systemic toxicity after repeated exposure to concentrations up to 620 mg/m³. Hence, the NOAEC was set at >= 620 mg/m³, which was the highest dose tested.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEC
620 mg/m³
Study duration:
subacute
Species:
rat
Quality of whole database:
The study was conducted to a protocol that is comparable to the OEDCD TG 412. It was not compliant with GLP.

Repeated dose toxicity: inhalation - local effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1979-06-11 - 1979-07-12
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
It was not compliant with GLP. Due to an accident, the mid dose group had to be terminated and the respective exposure was repeated in a following experiment.
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 412 (Subacute Inhalation Toxicity: 28-Day Study)
Deviations:
yes
Remarks:
(longer exposure duration (6.5 h) and no detailed information on test material purity)
GLP compliance:
no
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Bio-Breeding Laboratories Ltd., Ottawa, Ontario, Canada (received on 1979-05-24)
- Age at study initiation: 10-14 weeks
- Weight at study initiation: 200-250 g
- Housing: individually in in wire mesh-bottomed metal cages
- Diet: standard commercial laboratory diet (Purina) ad libitum (no food was available for the animals during exposure)
- Water: tab water available through a water sipper system ad libitum (no water was available for the animals during exposure)
- Acclimation period: 2 weeks
- Subsequent to arrival the animals received a qulified vetinary aid to ensure "normal" health status.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): animal room: 20.0-23.9, inhalation room: 21.4-23.6
Route of administration:
inhalation: vapour
Type of inhalation exposure:
whole body
Vehicle:
other: clean, dry air
Remarks on MMAD:
MMAD / GSD: not applicable
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: stainless steel whole body exposure chambers (400 l volume)
- Method of holding animals in test chamber: in a stainless steel wire mesh compartmentalised cage (each compartment measuring 7 x 4 x 3 inches)
- System of generating aerosols: Delivery of the test article into the chambers was accomplished by forcing air through gas generators fittet with sintered glass frits and containing the test item, and metering vapours into the chamber.
- Temperature, humidity, pressure in air chamber: Airflow through each chamber was measured in the exhaust line of the chamber by reading the pressure drop across an orifice plate on a magnehelic gauge. These gauges were calibrated against conventional ball-type flow meters. The magnitude of negative pressure within each chamber with respect to the outside atmosphere was always less than 0.5 inches of water. Temperature and relative humidity within both the inhalation chamber and the inhalation room were measured at hourly intervals during each exposure period. Chamber temperature and relative humidity were measured by means of remote sensors placed within each inhalation chamber. A mean daily chamber temperature of 22-26 °C was maintained throughout the study with the exception of study days 0.1 and 2.1 when higher temperatures were recorded in Chambers 1 to 2 and 3, respectively. Inhalation chamber relative humidity during exposure was generally 35-65% A higher relative humidity was observed in Chamber 1 (air controls) compared with the inhalation room, and a decrease in chamber relative humidity tended to occur as chamber test substance concentration increased.
- Air flow rate: One gas generator was used for Chamber 2 with an airflow of 6 l/min and two generators in parallel with an airflow of 30 l/min for Chamber 3. In the case of Chamber 4 a total airflow of 40 l/min was passed through two parallel generators partially immersed in a water bath at approximately 65 °C. The two generators were each serially connected to a pair of generators maintained at room temperature. The outputs from the vapour generation systems were each diluted with room air to a total chamber airflow of 45 l/min.

TEST ATMOSPHERE
- Brief description of analytical method used: Nominal chamber concentrations were calculated from the total airflow and the weight loss from the generator(s) during the exposure. Actual chamber test material concentrations were measured hourly using a MIRAN 1A Infrared gas analyser at an analytical wave length of 9.2 µm and a path length of 0.75 m. The Infrared gas analyser had been previously calibrated by injection of known microliter quantities of the test article into a closed loop system and allowing the liquid to vaporise.
- Samples taken from breathing zone: yes
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Target chamber concentrations for Groups 2 and 3 were generally achieved by the end of the 30 min equilibration time. In case of Group 4 approximately two hours of equilibration were required before the chamber concentration reached the targeted value. This was concluded by the study authors to be possibly related to the low volatility of the test material.
Mean daily actual chamber concentrations were close to the target concentrations on all exposure occasions except for Chamber 3 on study day 1.4 when the mean test article concentration reached a recorded maximum of 134 ppm. This overexposure was found to result from a technical error. The measured concentration in Chamber 1 (control chamber) was approximately 2 ppm throughout the study although no test article was introduced into this chamber. The possible reason for this spurious value was concluded to be caused by infrared absorbance at the analytical wavelength by vapours emanating from rat faeces and urine in the chambers. Consequently, the chamber test article concentrations should be considered as 2 ppm higher than the true concentrations existing in the chambers.
Duration of treatment / exposure:
4 weeks
Frequency of treatment:
6.5 h/day, 5 days/week
Each daily exposure period was 6.5 hours in duration, starting after the 30 min equilibration time when 95% of the target concentration was established within the chamber. After 6.5 hours of exposure at the desired concentration, delivery of the test article into the chamber was stopped and the chamber was allowed to equilibrate with room air for 30 min prior to the removal of the animals. Thus, the animals remained in the chamber for 7.5 hours/day.
Dose / conc.:
10 ppm (nominal)
Dose / conc.:
50 ppm (nominal)
Dose / conc.:
80 ppm (nominal)
Dose / conc.:
11.5 ppm (analytical)
Remarks:
mean analytical conc. over the whole study period
Dose / conc.:
54.4 ppm (analytical)
Remarks:
mean analytical conc. over the whole study period
Dose / conc.:
78.5 ppm (analytical)
Remarks:
mean analytical conc. over the whole study period
Dose / conc.:
9.5 ppm (analytical)
Remarks:
corrected mean analytical cnc. over the whole study period due to additional absorbance of vapours emanating from rat faeces and urine in the chambers
Dose / conc.:
52.4 ppm (analytical)
Remarks:
corrected mean analytical cnc. over the whole study period due to additional absorbance of vapours emanating from rat faeces and urine in the chambers
Dose / conc.:
76.5 ppm (analytical)
Remarks:
corrected mean analytical cnc. over the whole study period due to additional absorbance of vapours emanating from rat faeces and urine in the chambers
Dose / conc.:
77 mg/m³ air (analytical)
Remarks:
corrected mean analytical conc. over the whole study period due to additional absorbance of vapours emanating from rat faeces and urine in the chambers; values calculated with (molecular weight [g/mol] x mean corrected exposure conc. [ppm])/24.45
Dose / conc.:
425 mg/m³ air (analytical)
Remarks:
corrected mean analytical conc. over the whole study period due to additional absorbance of vapours emanating from rat faeces and urine in the chambers; values calculated with (molecular weight [g/mol] x mean corrected exposure conc. [ppm])/24.45
Dose / conc.:
620 mg/m³ air (analytical)
Remarks:
corrected mean analytical conc. over the whole study period due to additional absorbance of vapours emanating from rat faeces and urine in the chambers; values calculated with (molecular weight [g/mol] x mean corrected exposure conc. [ppm])/24.45
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
Group 1: vehicle control, Group 2: 10 ppm exposure, Group 3: 50 ppm exposure, Group 4: 80 ppm exposure

During exposure an incident occured in Group 3. The line supplying suppressed air to the Chamber 3 gas generators was improperly connected resulting in test article being sprayed over ten animals in the chamber. Four rats were thoroughly soaked with test article, and the chamber concentration of the test substance increased to 134 ppm. Two rats were semi-conscious and ataxic within 45 min, whereas one female rat died after approximately 8 hours. Ten rats were observed to have wet fur on their ventral surface post-treatment and two of them were semi-conscious and lethargic. Three rats were found dead the next morning, and the deaths, as well as the lethargy and the ataxia, were concluded by the study authors to be attributed to an acute toxic effect of the dermal absorption of the test article.
As a consequence of this accident a second group of twenty rats were treated with the test article at 50 ppm and the data generated was presented as an addendum to this report (Bio-Research Laboratories No. 1981-I-0065-1573-6 (Addendum), 1981).
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily; druing the treatment period after each exosure, and additionally during exposure for signs of overt toxicity

DETAILED CLINICAL OBSERVATIONS: No

BODY WEIGHT: Yes
- Time schedule for examinations: Individaul body weights were measured twice weekly during both acclimatisation and treatment periods.

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at study termination
- Anaesthetic used for blood collection: No
- Animals fasted: Yes
- How many animals: all animals
- Parameters checked: red blood cell count, haemoglobin concentration, packed cell volume, and total and differential leucocyte count

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at the termination of the treatment period
- Animals fasted: Yes
- How many animals: all
- Parameters checked: fasting serum glucose, blood urea nitrogen (BUN), serum alkaline phospahtase (SAP), serum glutamic oxalacetic transaminase (SGOT), and serum glutamic pyruvic transaminase (SGPT)

URINALYSIS: Yes
- Time schedule for collection of urine: during the night prior to terminal necropsy
- Metabolism cages used for collection of urine: Yes
- Animals fasted: No data
- Parameters checked: volume, colour, odour, transparency, specific gravity, occult blood, ketones, glucose, protein, pH, bilirubin, and microscopic examination of urinary sediment
Sacrifice and pathology:
GROSS PATHOLOGY: Yes (complete gross pathology of all animals)
HISTOPATHOLOGY: Yes (brain (cerebrum, cerebellum, medulla, and pons), spinal cord, olfactory nerve and/or olfactory bulb, both eyes and optic nerves, pituitary gland, thyroide, thymus, adrenal glands, heart, spleen, thoracic aorta, oesophagus, submandibular salivary gland, stomach, duodenum, jejunum, ileum, cecum, colon, rectum, liver, pancreas, gonads, lymph nodes (mesenteric and mediastinal), uterus, prostate, both kidneys, urinary bladder, sciatic nerve, skeletal muscle, bone marrow section (sternum), skin, tongue and buccal mucosa (bilateral), nasal and paranasal sinuses with vestibulum and adjacent bone, nasal septum, larynx, trachea, and major bronchi)
ORGAN WEIGHTS: Yes (left and right adrenal glands, brain, left and right gonads, heart, left and right kidneys, liver, pituitary gland, spleen, lungs and thyroid)
Other examinations:
MYELOGRAMS: Femoral bone marrow smears and sections from all control and high dose animals were examined by a qualified haematologist.
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 examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
no effects observed
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Details on results:
CLINICAL SIGNS AND MORTALITY
No abnormal clinical observations were noted for the animals of Groups 1-4 during exposure except for the animals of Group 3 as a consequence of the accident as described above. The majority of abnormal clinical observations recorded during the daily physical examinations consisted of signs of pulmonary congestion. Both treated and control groups exhibited a low incidence of congestion throughout the study, suggesting that this finding was unrelated to the treatment with the test item. Incidental observations included reddish brown fur discolouration of the perinasal and periorbital regions. Additionally, Groups 2 and 3 showed wet fur. The low incidence of these observations coupled with the absence of a wet fur in Group 4 animals post treatment and the presence of reddish brown fur discolouration perinasally and periorbitally in three control rats pretreatment suggests that exposure to the test item at concentrations up to 80 ppm had no adverse effect.

BODY WEIGHT AND WEIGHT GAIN
Statistical analysis (Dunnett’s t test), comparing the mean body weights of treated and control animals, did not reveal any significant differences between groups which could be attributed to treatment with the test article. Body weight of treated and control rats increased in a similar fashion throughout the study with the exception of several animals, which lost weight for a short period of time: one control male, one female of the 10 ppm dose group, and one male and female each of the high dose group, whereas the two latter animals showed congestion during these periods and the effect might be related to the observed weight loss. Due to a lack of time- and dose-dependence of the observed weight loss and the additional occurrence in the control group, it is suggested that the effects are not treatment related.

FOOD CONSUMPTION
No significant differences were observed between the absolute food consumption of treated and control groups. With respect to relative food consumption, significantly greater food intake was observed in Group 4 females during week 3 compared with the control group. A review of the individual data reveals that this is mainly contributed to one female animal of Group 4. This period of high food intake followed several weeks of reduced food consumption during which the body weight had decreased.
Infrequent periods of reduced food intake were noted in several rats of both the control and the high dose group and occurred at the same time as the body weight loss previously mentioned. The authors conclude that the effects seen are not related to the test material exposure.

HAEMATOLOGY
No significant differences in blood parameters existed between treated and control groups. Lymphocyte count was slightly elevated in some Group 4 animals, while segmented neutrophil count was slightly lower in Group 4 males and females as compared to controls. These small differences were not considered to be of any biological importance and the test material was concluded not to cause any adverse haematological effect in rats.

CLINICAL CHEMISTRY
The mean blood serum glucose level of Group 2 (10 ppm) females was significantly higher than that of the control group. Since no such effects were observed at higher doses, this effect was considered to be not treatment related. Although some individual values fell outside the range of normal values (e. g. increased BUN in one male of the control group, increased SAP in one male of the low dose group, increased SGOT in one female of the control group and one male of the high dose group (the latter in combination with an increased SGPT level)) none of these findings was considered to be exposure related.

URINALYSIS
Large intersubject variations existed in each group with respect to urine volume and specific gravity. However, the range for these parameters was similar in both treated and control groups. The high urine volume and associated low specific gravity in a proportion of the animal was considered possibly to be due to leakage of water from the water sipper in each cage. Hence, the test item was concluded not to cause any effect on urinary parameters measured in this study.

ORGAN WEIGHTS
The left adrenal gland (but not the right) of Group 4 (80 ppm) males and the spleens of Group 2 (10 ppm) males were significant lighter than those of control rats. There were no differences compared with the controls for the spleens of the high dose males. Hence, the slight effects observed were considered not to be treatment related. Individual data revealed that one male rat of the high dose, which had lost weight in the last week of the study, had high relative lung and kidney weights. One male each of the control group ant the low dose group had small left thyroide lobes and on male rat of the low dose group had small testes in terms of both absolute and relative weight. One female of the high dose group had small ovaries, one female of the low dose group had a small spleen, and two males of the low dose group had large lungs in terms of relative weight. Since these observations were isolated, they were not considered treatment related.

GROSS PATHOLOGY
The most commonly observed gross pathological findings were pulmonary congestion, enlargement and/or congestion of the lymph nodes and enlargement and/or fluid in the uterus and surrounding the ovaries of females. Each of these findings appeared as frequently in the treated groups as in the control group with the exception of fluid surrounding the ovary. The frequency of the latter finding was not dose related, and hence, gross pathological findings were concluded not to be treatment related. Further incidental findings not associated with the treatment were small testes or spleen, focal to general discolouration of the lung, liver, and kidney, and congestion of the thymus and salivary glands.

HISTOPATHOLOGY: NON-NEOPLASTIC
Pulmonary lesions were observed in both treated and control animals and consisted of lymphatic infiltration about air passages and blood vessels, some bronchiectasis, interstitial fibrosis, neutrophilic exudation, and alveolar macrophages. Similar lymphatic inflammatory was observed in the trachea, the larynx, and air passages. In most cases, lymphocytic aggregates were focal or multifocal in distribution and substantial amount of pulmonary parenchyma was intact and uninvolved. These changes were possibly the result of a mild to moderate murine respiratory mycoplasmosis. In some animals of both control and treated groups there was evidence of pulmonary oedema with accumulation of an eosinophilic proteinaceous material in alveolar spaces. This change was considered to be of agonal and post mortem occurrence. Hence, the test material was concluded not to produce any pathological changes.

MYELOGRAMS: There were no differences in cell/fat and myeloid/erythroid ratios between control and treated animals. Megakaryocytes were plentiful in all rats and there was no evidence of tumour or granuloma infiltration. Bony spicules were normal in treated and untreated animals alike. The eosinophilia observed in one high dose female could be associated with extramedullary haematopoiesis in the liver of this animal, however, the eosinophilia, neutrophilia, and lymphocytosis observed in other control or treated animals cannot be correlated with any histological changes. Hence, the test material was concluded not to cause effects on bone marrow haematopoiesis.
Dose descriptor:
NOAEC
Effect level:
>= 80 ppm (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: highest dose tested
Dose descriptor:
NOAEC
Effect level:
>= 620 mg/m³ air (analytical)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: highest dose tested
Critical effects observed:
no
Conclusions:
The test item was tested for repeated inhalation toxicity similar or equivalent to the OECD TG 412, but not in compliance with GLP. The test article did not exhibit any systemic toxicity after repeated exposure to concentrations up to 620 mg/m³. Hence, the NOAEC was set at >= 620 mg/m³, which was the highest dose tested.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEC
620 mg/m³
Study duration:
subacute
Species:
rat
Quality of whole database:
The study was conducted to a protocol that is comparable to the OEDCD TG 412. It was not compliant with GLP.

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

Studies were chosen as key when the available study was of relevance and of sufficient quality for classification, labelling and for risk assessment.

Oral

For the oral route, a reliable key OECD 422 combined repeated dose/reproductive and developmental screening study is available for trimethoxyphenylsilane. This study is a valid investigation of the toxicological effects resulting from repeated oral-gavage administration of the test item to male and female rats for at least 28 days. The test item was administered in dried and deacidified corn oil as vehicle at dosages of 100, 250, and 500 mg/kg body weight/day, and controls received the vehicle only. At 250 mg/kg bw/day, treatment with the test item caused an increase in the concentration of urea and bile acids. Multifocular tubular degeneration/regeneration and transitional hyperplasia of kidney were noted in males and females. During the macroscopic examination, the urinary bladder was observed to be thickened in males and females at all dose levels. This correlated with the findings noted at the microscopic level. At all dose levels (100, 250 and 500 mg/kg bw/day) during the histopathology examination, perivascular lymphoid cell infiltration and transitional cell hyperplasia of the urinary bladder were observed in males and females. Therefore, based on the findings in urinary bladder noted in all test item-treated groups, a general NOAEL could not be established. The LOAEL was 100 mg/kg bw/day (SEHSC, 2009).

For the structural analogue triethoxy(phenyl)silane (CAS 780 -69 -8), a reliable combined repeated dose/reproductive and developmental screening study according to OECD 422 by oral route is available (Eurofins, 2019). The test substance was administered in corn oil as vehicle at dosages of 40, 120, and 360 mg/kg body weight/day, and controls received the vehicle only. Two recovery groups were included which were treated with vehicle only or 360 mg test substance/kg bw/day. 

One rat from the high dose group with 360 mg/kg bw/day was sacrificed moribund on day 15. As to histopathological evaluation, it is assumed that the moribund condition of the animal was a consequence of backflow nephrosis and considered to be treatment-related. There were no treatment-related clinical signs throughout the treatment period up to 360 mg/kg bw/day. There were no treatment-related functional observation changes at the end of the treatment and recovery periods. Treatment-related adverse effects of the test item were found on male and female body weight and food consumption mainly at 360 mg/kg bw/day. Haematology and coagulation, clinical biochemistry, and urinalysis parameters were not affected by the treatment in both genders. At scheduled necropsy, treatment-related gross macroscopic findings were noted in kidneys and urinary bladder. Changes in respective organ weights are considered to be treatment related at 120 and 360 mg/kg bw/day. Test item-related gross lesions consisted in the kidneys of renal pelvis dilatation (in one decedent recovery male it was described as enlarged kidney and abnormal consistency). Histologically, the incidence of pelvic dilatation increased in males at 40 up to 360 mg/kg bw/day, and tubular dilatation was noted in almost all males at 360 mg/kg bw/day. The findings at 120 and 360 mg/kg bw/day, degenerative and inflammatory lesions consisted of increased incidences and/or severities of tubular basophilia, pyelitis, and, at 360 mg/kg bw/day, of interstitial inflammation, interstitial fibrosis and papillary necrosis. Furthermore, there was urothelial hyperplasia in two males at 120 mg/kg bw/day, and in all males and four females at 360 mg/kg bw/day. Males were more affected than females. The findings were still present after the recovery period.

Pelvic dilatation is common in rats, as are the single incidences of tubular basophilia at minor severity degrees. Therefore, findings at 40 mg/kg bw/day are not considered to be test item-related. There were also ureter dilatation and thickening of the urinary bladder wall noted at necropsy. Histologically, these alterations correlated with the ureters of one male at 120 mg/kg bw/day, and with all males and three females at 360 mg/kg bw/day to dilatation, associated in some cases at 360 mg/kg bw/day with mucosal and/or muscularis hyperplasia, and/or inflammation. The highest degree of mucosal hyperplasia was noted at the most distal portions of the ureters.

In the urinary bladder, there was diffuse urothelial hyperplasia in both sexes at 120 and 360 mg/kg bw/day. This finding was associated with an increased incidence and severity of mononuclear cell foci. In one male, the urethra was found in its full length and diameter within the prostate tissue. The urothelium showed moderate hyperplasia accompanied by a minimal subacute inflammation. At some locations, in mucosal folds, precipitation of an unknown material was seen. The precipitation was noted by chance only due to the unusual circumstances of observing the urethra within the prostate gland. This finding explains the higher grading of urothelial hyperplasia at more distal parts of the urinary system. Precipitation is deemed to cause irritative effects to the urothelium that causes backflow nephrosis with dilatation of the renal pelvis (hydronephrosis) and tubular dilatation (nephrohydrosis).

Stress-related lesions were noted in the adrenals by cortical hypertrophy and in the thymus by increased thymic atrophy.

Based on the findings in the urinary tract, the NOAEL for triethoxy(phenyl)silane in this study for general toxicity is considered to be 40 mg/kg bw/day.

 

Comparing the findings in the screening study with both substances it can be seen that their target organ is the same and their toxicity is in the same range. Thus, read across between both substances can be justified.

 

Inhalation

In a key study, an acceptable repeated inhalation toxicity study similar or equivalent to the OECD TG 412, but not in compliance with GLP, the test item was administered to Sprague-Dawley rats (10 per sex and dose) by whole body exposure at concentrations of 9.5, 52.4, and 76.5 ppm, corresponding to 0.077, 0.425, and 0.620 mg/l for 6.5 hours per day, 5 days/week for a total of 28 days.

There were no compound related effects in mortality, clinical signs, body weight, food consumption, haematology, clinical chemistry, organ weights, or gross and histologic pathology.

Based on no systemic toxicity after repeated exposure to concentrations up to 0.62 mg/l, the NOAEC was set at > 620 mg/m³, which was the highest dose tested (Bio-Research Laboratories, 1980).

Justification for classification or non-classification

Following oral exposure there were signs of adverse effects in the urinary bladder of rats at all doses and therefore it is proposed that trimethoxyphenylsilane has to be classified for STOT-RE Cat 2, with the hazard statement 'H373:May cause damage to organs according to Regulation (EC) 1272/2008.

The available data on repeated dose toxicity of the registered substance by the inhalation route do not meet the criteria for classification according to Regulation (EC) 1272/2008 or Directive 67/548/EEC, and is therefore conclusive but not sufficient for classification of the registered substance.

No data are available for the dermal route. However, dermal absorption of the submission substance is expected to be low 0.004 mg/cm²/h (DERMWIN V2.00.2009). Generally, absorption of xenobiotica via the inhalation route is higher than via the dermal route, as the skin is known to have a barrier function and the lung is an exchange organ. Since no specific target organ toxicity was observed after inhalation exposure, a hazard via the dermal route can be excluded.

In conclusion, the hazard statement "H373: May cause damage to urinary bladder after prolonged or repeated oral exposure is" will be applied.