Registration Dossier

Administrative data

Endpoint:
short-term repeated dose toxicity: inhalation
Remarks:
combined repeated dose and reproduction / developmental screening
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: This study was conducted in accordance with GLP and guidelines (OECD 422, USEPA OPPTS 870.3650) and sufficient data are available for the interpretation of study results.
Cross-reference
Reason / purpose for cross-reference:
reference to same study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2005
Report date:
2005

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: USEPA OPPTS 870.3650
Deviations:
no
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
The purity of three drums of test material used was 99.45, 99.65 and 99.65%, as determined by gas chromatography

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
CD rats (Crl:CD(SD)IGS BR) were selected because of their general acceptance and suitability for toxicity testing, availability of historical background data and the reliability of the commercial supplier (Charles River Laboratories Inc., Portage, Michigan).

Animals were about 8 weeks of age at study start. Each animal was evaluated by a laboratory veterinarian, or a trained animal/toxicology technician under the direct supervision of a lab veterinarian, to determine their general health status and acceptability for study purposes upon arrival at the laboratory. The animals were housed two of same sex per cage in stainless steel cages, in rooms designed to maintain adequate conditions (temperature, humidity, and photocycle), and acclimated to the laboratory for at least one week prior to the start of the study.

After assignment to study, animals were housed one per cage in stainless steel cage in rooms designed to maintain adequate conditions (temperature, humidity, and photocycle). A 12-hour light/dark photocycle was maintained for all animal room(s) with lights on at 6:00 a.m. and off at 6:00 p.m. Room air was exchanged approximately 12-15 times/hour.

Cages had wire-mesh floors and were suspended above catch pans. Cages contained feed containers and pressure activated, nipple-type watering systems. Dams were housed one per cage (with their litter) in plastic cages provided with corn cob nesting material from approximately day 19 of gestation and throughout the lactation phase of the study. Room temperature and relative humidity were recorded daily. The relative humidity was maintained within a range of 40-70%, with the exception of some minor deviations that occurred during the study (data contained in study file). These deviations involved transient excursions in the range of 38-39% humidity. These deviations were minor, had no noticeable impact on the animals, and did not affect the integrity of this study. The room temperature was maintained within the range of 22 ± 1°C (with a maximum permissible excursion range of ± 3°C). These values were within the laboratory recommended range for rats.

For the exposures, animals were transferred each day to stainless steel cages placed inside Rochester-style exposure chambers (see Animal Exposure section) without access to feed or water. All animals were housed one per cage during exposures except during the mating period (see breeding procedures).

Prior to test material administration, animals were stratified by body weight and then randomly assigned to treatment groups using a computer program designed to increase the probability of uniform group mean weights and standard deviations at the start of the study. Animals placed on study were uniquely identified via subcutaneously implanted transponders (BioMedic Data Systems, Seaford, Delaware) which were correlated to unique alphanumeric identification numbers. If a transponder stopped functioning or was lost, it was replaced with a new transponder which was correlated with the unique animal number.

Animals were provided LabDiet Certified Rodent Diet #5002 (PMI Nutrition International, St. Louis, Missouri) in meal form. Feed and municipal water were provided ad libitum except during the 6 hour exposure period when feed and water were withheld. Analyses of the feed were performed by PMI Nutrition International to confirm the diet provided adequate nutrition and to quantify the levels of selected contaminants. Drinking water obtained from the municipal water source was periodically analyzed for chemical parameters and biological contaminants by the municipal water department. In addition, specific analyses for chemical contaminants were conducted at periodic intervals by an independent testing facility. The results of these analyses indicated no contaminants at levels that would interfere with the conduct of this study or interpretation of the results.

Administration / exposure

Route of administration:
inhalation: vapour
Type of inhalation exposure:
whole body
Vehicle:
air
Details on inhalation exposure:
Animals were exposed to the test material daily via inhalation. Each day during the exposure period, rats were transferred to whole-body exposure chambers, exposed to the targeted concentrations of test material for six hours, then returned to their home cages.

The animals were exposed to filtered air or test material vapors in 2 cubic meter stainless steel and glass Rochester-type whole-body exposure chambers [1.3 m x 1.2 m wide x 1.2 m deep with a pyramidal top and bottom]. Chamber airflow was maintained at approximately 450 liters per minute. This flow rate was sufficient to provide the normal concentration of oxygen to the animals and 12-15 calculated air changes per hour. The chambers were operated at a slightly negative pressure, relative to the surrounding area. Chamber airflow data were collected using Setra Differential Pressure Transducers (Setra System, Inc., Acton, Massachusetts). The signal from the pressure transducer was sent to the CAMILE TG4 Acquisition and Control System and recorded in liters per minute. The differential pressure transducer was calibrated with a gas meter (Singer Aluminum Diaphragm Meter, Model AL-2300, American Meter Division, Philadelphia, Pennsylvania) prior to the start of the study. Chamber temperature and relative humidity data were collected using a resistance temperature device (RTD) (Omega HX94C, Omega Engineering Inc., Stamford, CT) coupled to the CAMILE TG4 Data Acquisition and Control System. The chamber temperature and relative humidity was controlled by a system designed to maintain values of approximately 22 ± 3°C and 30 to 70%, respectively. Chamber temperature, relative humidity, and airflow data were automatically logged into data files once per hour by CAMILE, and subsequently extracted and printed for inclusion in the study file with the exception of chamber temperature and relative humidity data generated beginning July 17 through the last day of exposure. The Omega RTDs normally used to measure chamber temperature and relative humidity were inadvertently sprayed with water during post-exposure sanitization of the chambers on July 16, causing them to lose their ability to measure data. Therefore, chamber temperature and relative humidity data for each exposure chamber were manually recorded once per hour from calibrated thermometers and hygrometers stationed inside the chambers. This equipment was in place for data collection in the event of a failure of the CAMILE® TG4 Data Acquisition and Control System or its associated equipment.

The various concentrations of test material were generated using a glass J-tube method. Liquid test material was pumped into the glass J-tube assembly and vaporized by compressed nitrogen gas passing through the bead bed of the glass J-tube approximately 40 liters per minute. The nitrogen was heated as needed with a flameless heat torch (FHT-4, Master Appliance Corporation, Racine, Wisconsin) to the minimum extent necessary to vaporize the test material. The generation system was electrically grounded and the J-tubes were changed as needed. The compressed nitrogen and test material vapors were mixed and diluted with supply air to achieve a total flow of 450 liters per minute at the desired test chamber concentration.

Animals in the negative control group were exposed to an atmosphere of approximately 410 L/minute of humidified, HEPA-filtered air and approximately 40 L/minute of compressed nitrogen.

The chamber concentrations of test substance, measured approximately in the center of the breathing zone of the animals, was determined at least once per hour with a Miran 1A infrared (IR) spectrophotometer (Foxboro/Wilks, South Norwalk, Connecticut). The IR spectrophotometer was calibrated and a standard curve was compiled prior to the start of the study, using air standards prepared by vaporizing measured volumes of test substance into Tedlar sample bags (Series 233, SKC, Eighty Four, Pennsylvania) along with the metered volumes of dry, compressed air. The analytical concentration during the exposure was interpolated by the CAMILE TG4 Data Acquisition and Control System using the standard curve. The analytical system was checked prior to each exposure with a standard gasbag containing a known concentration of test substance. The nominal concentration of the test material in each chamber was estimated based on the amount of test material used and the total airflow through the chamber. Prior to the start of the study, each of the chambers was checked to ensure that a uniform distribution of vapor was present throughout the breathing zone of the animals.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The chamber concentrations of test substance, measured approximately in the center of the breathing zone of the animals, was determined at least once per hour with a Miran 1A infrared (IR) spectrophotometer (Foxboro/Wilks, South Norwalk, Connecticut). The IR spectrophotometer was calibrated and a standard curve was compiled prior to the start of the study, using air standards prepared by vaporizing measured volumes of test substance into Tedlar sample bags (Series 233, SKC, Eighty Four, Pennsylvania) along with the metered volumes of dry, compressed air. The analytical concentration during the exposure was interpolated by the CAMILE TG4 Data Acquisition and Control System using the standard curve. The analytical system was checked prior to each exposure with a standard gasbag of known concentration of test substance. The nominal concentration of the test substance in each chamber was estimated based on the amount of test substance used and the total airflow through the chamber. Prior to the start of the study, each of the chambers was checked to ensure that a uniform distribution of vapor was present throughout the breathing zone of the animals.
Duration of treatment / exposure:
Males were exposed 6 hours per day for 14 days prior to mating and continuing throughout mating until necropsy for a total of 33 consecutive days. Females were exposed 6 hours per day for 14 days premating, continuing throughout mating (two weeks) and gestation (up to and including GD 20).
Frequency of treatment:
Daily, 6h/day, 7day/week
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
50, 200, 600 ppm
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
50.1 ± 1.0, 201.0 ± 2.1, 601.8 ± 9.1 ppm
Basis:
analytical conc.
No. of animals per sex per dose:
12
Control animals:
yes, concurrent vehicle
Details on study design:
An inhalation range-finding study was conducted to set the doses for the main study. In this study, five rats/sex were exposed to 0, 100, 450 or 900 ppm test substance for 6 hr/day for 14 days. Clinical observations were conducted daily and body weight, body weight gain, and feed consumption were recorded weekly. An ophthalmic examination was conducted on the last day of exposure using a Welch Allyn halogen light. A gross examination of each animal was performed and liver and kidney weights were recorded.

Mean chamber concentration values for the range-finding study were 0, 105.1 ± 2.2, 450.8 ± 6.4, and 900.0 ± 15.9 ppm for the 0, 100, 450, and 900 ppm chambers, respectively. Actual mean chamber concentration values deviated by < 5% from the targeted values.

All animals survived to the scheduled necropsy. Significant clinical signs of toxicity were observed in the 900 ppm exposure group which included decreased activity, incoordinated gait, cloudy eyes, and periocular, perinasal, perineal or perioral soiling. Incoordinated gait was noted on test day 1 in all 900 ppm male and female rats, and in one male or one female rat exposed to 900 ppm on each of test days 2-4. In addition, decreased activity (1/5), perineal soiling (1/5), and periocular soiling (3/5) were also observed in the 900 ppm females on test day 1. Later in the exposure period (test days 8-14), cloudy eyes were noted clinically in 4/5 males and 1/5 females. In addition, cloudy corneas were noted in four males and three females at the pre-necropsy ophthalmic examination. Feed consumption was decreased in both males and females during the first week of exposure to 900 ppm, while body weights were decreased only in the males from this group. Liver weights were increased in males and females exposed to >= 450 ppm. At necropsy, four males and three females exposed to 900 ppm exhibited unilateral or bilateral cloudy corneas. Cloudy corneas were characterized by a pale linear area at the site of palpebral closure, consistent with drying of the cornea along this site. There were no clinical signs or signs of toxicity in males or females exposed to 100 ppm.
Positive control:
None

Examinations

Observations and examinations performed and frequency:
IN-LIFE OBSERVATIONS:
Twice each day a cage-side examination was conducted and to the extent possible the following parameters were evaluated: skin, fur, mucous membranes, respiration, nervous system function (including tremors and convulsions), animal behavior, moribundity, mortality, and the availability of feed and water.

CLINICAL OBSERVATIONS:
Clinical examinations were conducted on all animals once daily throughout the study. These examinations were conducted following exposures. Clinical observations included a careful, hand-held examination of the animal with an evaluation of abnormalities in the eyes, urine, feces, gastrointestinal tract, extremities, movement, posture, reproductive system, respiration, skin/hair-coat, and mucous membranes, as well as an assessment of general behavior, injuries or palpable mass/swellings.

DETAILED CLINICAL OBSERVATIONS (DCO):
DCO were conducted on all rats pre-exposure, and then weekly throughout the study. Mated females were given DCO examinations on GD 0, 7, 14, and 20, and lactation day (LD) 3. The DCO examinations were conducted at approximately the same time each examination day (before exposure) according to an established format. The examination included cage-side, hand-held and openfield observations that were recorded categorically or using explicitly defined scales.

FUNCTIONAL TESTS:
The functional tests included a sensory evaluation, rectal temperature, grip performance and motor activity. Functional tests were conducted pre-exposure and during the last week of the treatment period. For male rats, this took place on test day 30. For female rats, this took place on LD 4.

Sensory Evaluation
The sensory evaluation included a test for nociception (responsiveness to tail pinch) and for startle response (responsiveness to sharp noise). The evaluation was conducted in a clear plastic box.

Rectal Temperature
Rectal temperature was measured by carefully placing a rectal thermistor (Physitemp RET-2, T-type) approximately 4 cm into the rectum for about 15-20 seconds. Temperature was then recorded. The thermistor was validated at 37°C before and after the study. The instrument was re-calibrated if the validation temperature recordings differ from the reference thermometer by more than ±0.5°C.

Grip Performance
Hind-limb grip performance was tested according to the procedure described by Mattsson et al. (1986). Briefly, the observer places the animal’s forelegs on a plastic bench and the hind- feet are set on a horizontal screen attached to an electronic strain gauge (Chatillon, Greensboro, North Carolina). The observer then smoothly but firmly pulls backward on the tail until the animal’s grip on the screen was broken. An electronic strain gauge reading was used to record the animal’s resistance to the pull in grams. The average of three trials was used for statistical analysis. Forelimb grip performance was similarly tested. In this application, a bench was not used, and the animal was placed so that the forefeet are on the screen and the hind-feet were suspended approximately 10 cm above the smooth horizontal plastic surface.
A standard 500 g weight attached to a fine-gauge wire was suspended from the load cell (weight was measured from the load cell in grams). A 1% tolerance (i.e. 500 ± 5 grams) was acceptable and was checked just before and just after testing.

Motor Activity:
An automated system was used for motor activity (MA) data collection. No entry into the MA test room was allowed during the testing period. Each test session consisted of ten 8- minute epochs, totaling 80 minutes of testing per animal per test session. This duration was chosen based on the results of a validation study indicating that performance of control animals approached asymptote in 30-40 minutes in Fischer 344 rats (Marable and Andrus, 2001). Activity counts for each epoch were recorded.
Motor Activity Cage Calibration: Cages to be used for testing were calibrated prior to testing each day. Calibration was performed with a rod attached to a rotary motor that breaks the infrared beam at a constant speed. The duration of each beam break was calculated to ensure equivalence across chambers.
Motor Activity Cage Allocation: The rats were allocated to the motor activity cages in such a way the counterbalancing of treatment groups and sexes across cages and test times were maximized.

OPHTHALMOLOGY
The eyes of all animals were examined by a veterinarian pre-exposure and prior to the scheduled necropsy using indirect ophthalmoscopy. One drop of 0.5% tropicamide ophthalmic solution was instilled in each eye to produce mydriasis prior to the indirect ophthalmic examinations. At necropsy, the eyes were also examined using a moistened glass slide pressed to the cornea.

BODY WEIGHTS
All rats were weighed at least once during the pre-exposure period and on the first day of exposure. Body weights for males were recorded weekly throughout the course of the study. Females were weighed weekly during the premating and mating periods. During gestation, females were weighed on GD 0, 7, 14, and 20. Females that delivered litters were weighed on LD 1 and 4. Females that failed to mate or deliver a litter were not weighed during the gestation or lactation phases. Body weight gains were determined for the following intervals: GD 0-7, 7-14, 14-20, 0-20, and LD 1-4.

FEED CONSUMPTION:
Feed consumption was determined weekly during the two week pre-breeding period by weighing feed containers at the start and end of a measurement cycle. During breeding, feed consumption was not measured in males or females due to co-housing. Following breeding, feed consumption was not measured for males. For mated females, feed consumption was measured on GD 0, 7, 14, and 20. After parturition, feed consumption was measured on LD 1 and 4. Feed consumption was not recorded for females that failed to mate or deliver a litter.

LITTER OBSERVATIONS:
Females were observed for signs of parturition beginning on or about GD 20. In so far as possible, parturition was observed for signs of difficulty or unusual duration. The day of parturition was recorded as the first day the presence of the litter was noted and was designated as LD 0. Litters were examined as soon as possible after delivery. The following information was recorded on each litter: the date of parturition, the number of live and dead pups on LD 0, 1, and 4, and the sex and body weight of each pup on LD 1 and 4. Any visible physical abnormalities or demeanor changes in the neonates were recorded as they were observed during the lactation period. Any pups found dead or sacrificed in moribund condition were sexed and examined grossly, if possible, for external and visceral defects and discarded.
Sacrifice and pathology:
On the day prior to the scheduled necropsy, all males and females in each dose group were fasted overnight. At necropsy, the animals were anesthetized with CO2, and the blood samples were collected from the orbital sinus.

A complete necropsy of all adults was performed. Males were necropsied on test day 34, while females that delivered litters were necropsied on LD 5. Fasted adult rats submitted alive for necropsy were anesthetized by the inhalation of carbon dioxide, weighed, blood samples collected, their tracheas exposed and clamped, and the animals were then euthanized by decapitation.

ADULT NECROPSY
The necropsy was conducted by a veterinary pathologist assisted by a team of trained individuals. The necropsy included an examination of the external tissues, and all orifices. The head was removed, the cranial cavity opened and the brain, pituitary and adjacent cervical tissues were examined. The eyes were examined in situ by application of a moistened microscope slide to each cornea. The nasal cavity was flushed via the nasopharyngeal duct and the lungs were distended to an approximately normal inspiratory volume with neutral, phosphate-buffered 10% formalin using a hand-held syringe and blunt needle. The skin was reflected from the carcass, the thoracic and abdominal cavities were opened and the viscera examined. All visceral tissues were dissected from the carcass, re-examined and selected tissues were incised. The uteri of all females were stained with a 10% solution of sodium sulfide for approximately two minutes and the numbers of implantation sites were recorded. After evaluation, uteri were gently rinsed with saline and preserved in neutral phosphate-buffered 10% formalin. The following tissues were trimmed and weighed: testes, epididymides, liver, kidneys, adrenals, thymus, spleen, brain, thyroid/parathyroid (after fixation) and heart. The organ to body weight ratios were calculated.

Representative samples of the following tissues were collected and preserved in neutral, phosphate-buffered 10% formalin, except that the testes and epididymides were preserved by immersion in Bouin’s fixative. Transponders were removed and placed in jars with the tissues.
Adrenals; aorta; auditory sebaceous glands; bone (including joint); bone marrow; brain (cerebrum, brainstem, cerebellum); cecum; cervix; coagulating glands; colon; cranial nerve – optic; duodenum; epididymides; eyes; gross lesions; heart; ileum; jejunum; kidneys; lacrimal/harderian glands; larynx; liver; lungs; mammary gland – females only; mediastinal lymph node; mediastinal tissues; mesenteric lymph node; mesenteric tissues; nasal tissues/pharynx; oesophagus; oral tissues; ovaries; oviducts; pancreas; parathyroid glands; peripheral nerve –tibial; pituitary; prostate; rect um; salivary glands; seminal vesicles; skeletal muscle; skin and subcutis; spinal cord (cervical, thoracic, lumbar); spleen; stomach; testes; thymus; thyroid gland; tongue; trachea; urinary bladder; uterus; vagina.

HISTOPATHOLOGY
Histopathologic examination of the tissues listed above, and tissues with relevant gross lesions was conducted on all adult rats from the control and high-dose groups. The histopathological examination of the testes included a qualitative assessment of stages of spermatogenesis. A cross section through the approximate center of both testes of control and high-dose males was embedded in paraffin, sectioned at 5 mm and stained with modified periodic acid-Schiff’s-hematoxylin. The presence and integrity of the 14 stages of spermatogenesis were qualitatively evaluated following the criteria and guidance of Russell et al. (1990). Microscopic evaluation included a qualitative assessment of the relationships between spermatogonia, spermatocytes, spermatids, and spermatozoa seen in cross sections of the seminiferous tubules. The progression of these cellular associations defined the cycle of spermatogenesis. In addition, sections of both testes were examined for the presence of degenerative changes, e.g., vacuolation of the germinal epithelium, multinucleated giant cells, a decrease in the thickness of the germinal epithelium, a preponderance of Sertoli cells, sperm stasis, inflammatory change s, mineralization, and fibrosis. Examination of tissues from the remaining groups was limited to liver and relevant gross lesions. Other paraffin embedded tissues were sectioned approximately 6 μm thick, stained with hematoxylin and eosin and examined by a veterinary pathologist using a light microscope.
Other examinations:
HAEMATOLOGY
Samples were mixed with ethylenediamine-tetraacetic acid (EDTA) and blood smears were prepared, stained with Wright-Giemsa stain and archived for potential future evaluation if warranted. Hematologic parameters were assayed using a Technicon H1E Hematology Analyzer (Bayer Corporation, Tarrytown, New York): haematocrit; haemoglobin concentration; red blood cell count; total white blood cell count; platelet count; differential WBC count; reticulocyte count; mean corpuscular haemoglobin, volume, and haemoglobin concentration.

COAGULATION
Blood samples were collected in sodium citrate tubes, centrifuged and plasma collected and assayed using an ACL9000 (Instrumentation Laboratory, Lexington, Massachusetts) for prothrombin time

CLINICAL CHEMISTRY
Serum was separated from cells as soon as possible following blood collection. Serum parameters were measured using a Hitachi 914 Clinical Chemistry Analyzer (Boehringer-Mannheim, Indianapolis, Indiana): alkaline phosphatase; alanine aminotransferase; aspartate aminotransferase; albumin; cholesterol; creatinine; electrolytes (Na+, K+, PO4---, Cl- and Ca++); glucose; total bilirubin; total protein; urea nitrogen.

URINALYSIS
A timed urine volume was obtained from all male rats in each dose group (nonfasted) during the week prior to necropsy (test day 29). This was accomplished by placing rats in metabolism cages and collecting urine for approximately 16 hours. Urine was assessed for colour, appearance and specific gravity. Semiquantitative analysis (Multistix Reagent Strips, Bayer Corporation, Elkhardt, Indiana on the Clinitek 200+) of: pH; bilirubin; glucose; proteins; ketones; blood; urobilinogen.
Urine was collected by manual compression of the bladder. The urine was pooled for each group, and the microsediment was characterized microscopically.
Statistics:
Descriptive statistics only (means and standard deviations) were reported for chamber concentration, temperature, humidity, airflow, RBC indices, and WBC differential counts.

Parental body weights, gestation and lactation body weight gains, litter mean body weights, feed consumption, organ weights (absolute and relative), clinical chemistry data, urinalysis, and appropriate hematologic data were first evaluated by Bartlett's test for equality of variances (α=0.01). Based upon the outcome of Bartlett's test, either a parametric or nonparametric analysis of variance (ANOVA) was performed. If the ANOVA was significant at
α=0.05, a Dunnett's test or the Wilcoxon Rank-Sum test with Bonferroni's correction was performed (experiment wise α=0.05). Statistical outliers were identified by a sequential test (Grubb's) at α=0.02, and were excluded only for documented scientifically sound reasons (e.g., spilled or scratched feed).

DCO and sensory evaluation incidence scores were statistically analyzed by a z-test of proportions comparing each treated group to the control group at α=0.05. The data were analyzed separately for each time point. Because numerous measurements were statistically compared in the same group of animals, the overall false positive rate (Type I errors) was greater than the normal alpha levels. Therefore, the final interpretation of the data considered statistical analyses along with other factors, such as dose-response relationships and whether the results were consistent with other biological and pathological findings and historical control values.

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Perioral soiling (clear) was found in 6/12 males and 7/12 females at 600ppm. Perioral soiling (red) was found in 1/12 males and 2/12 females at 600ppm. Perinasal soiling (red) was found in 2/12 females at 600ppm.
Mortality:
mortality observed, treatment-related
Description (incidence):
Perioral soiling (clear) was found in 6/12 males and 7/12 females at 600ppm. Perioral soiling (red) was found in 1/12 males and 2/12 females at 600ppm. Perinasal soiling (red) was found in 2/12 females at 600ppm.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Slight (<6%) reduction in body weights in males and females at 600ppm compared to controls. Not statistically significant, but persistent throughout exposure. Final bodyweights of males and females in the 600ppm group were reduced by 5.6% and 5.4% respect
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Slightly decreased in male and females at 600ppm, but not statistically significant except in females during the first week of gestation.
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
no effects observed
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
no effects observed
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Increased liver weight (absolute and relative to bodyweight) in both males and females at 600ppm. Statistical significance only reached for absolute weight in males and relative weights in both sexes.
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Males exposed to 600 ppm had a treatment-related increase in the incidence of very slight centrilobular hypertrophy of hepatocytes. The incidence of the centrilobular hypertrophy was 1, 1, 1, and 9 for males exposed to 0, 50, 200, or 600 ppm respectively.
Histopathological findings: neoplastic:
not specified

Effect levels

open allclose all
Dose descriptor:
NOAEC
Effect level:
200 ppm
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No adverse effects observed
Dose descriptor:
LOAEC
Effect level:
600 ppm
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Perioral and/or perinasal soiling; slight decreases in feed consumption, body weights and/or body weight gains; increased mean liver weight and very slight centrilobular hypertrophy (males only)

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
Based on these results, the no-observed-effect concentration (NOEC) for general toxicity was 200 ppm. The NOEC for reproductive and neurological effects was 600 ppm, the highest concentration tested.