Propanenitrile, 2,3,3,3-tetrafluoro-2-(trifluoromethyl)-

Repeated dose toxicity: inhalation

Administrative data

Endpoint:

short-term repeated dose toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

22 August 2013 to 16 October 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study was conducted in compliance with OECD and US EPA GLP regulations.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2014

Report date:

2014

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 412 (Subacute Inhalation Toxicity: 28-Day Study)

Deviations:

no

GLP compliance:

yes

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

MTDID 28136

IUPAC Name:

MTDID 28136

Test material form:

other: gas

Details on test material:

- Name of test material (as cited in study report: MTDID 28136
- Substance type: Mono-constituent
- Physical state: Gas
- Analytical purity: 99.98%
- Purity test date: 17 June 2013
- Lot/batch no.: 41-2601-2772-9, PDC Lot 2

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Inc.
- Age at study initiation: 38 days old
- Weight at study initiation:
- Fasting period before study: None
- Housing: Individually
- Diet (e.g. ad libitum): PMI Nutrition International, LLC, Certified Rodent LabDiet 5002 ad libitum
- Water (e.g. ad libitum): Reverse osmosis-treated tap water ad libitum
- Acclimation period: 13 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21-22
- Humidity (%): 40.3-54.7
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12
IN-LIFE DATES: From: 22 August 2013 To: 16 October 2013

Administration / exposure

Route of administration:

inhalation: gas

Type of inhalation exposure:

whole body

Vehicle:

other: unchanged (no vehicle)

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: 1.0 cubic meter stainless steel and glass whole-body inhalation exposure chambers. Exposure atmospheres were generated by releasing the test article in gas form from the original cylinder.
- Exposure chamber volume: 1.0 cubic meter
- Method of holding animals in test chamber: Individually caged in exposure caging
- Source and rate of air: Provided from a HEPA and charcoal-filtered, temperature and humidity-controlled source.
- Method of conditioning air: No data
- Treatment of exhaust air: HEPA and charcoal-filtered
- Temperature, humidity, pressure in air chamber: Between 20-26 C, 30-70% humidity, and 12-15 air changes per hour
TEST ATMOSPHERE
- Brief description of analytical method used: Exposure concentrations were determined at approximately 60-minute intervals using an appropriate gas chromatography method.
- Samples taken from breathing zone: yes

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Exposure concentrations were determined at approximately 60-minute intervals using an appropriate gas chromatography method.

Duration of treatment / exposure:

6 hours per day

Frequency of treatment:

5 days per week for 4 weeks (20 total exposures)

Doses / concentrations

Remarks:

Doses / Concentrations:
0, 252, 516, 1033, or 1512 ppm
Basis:
analytical conc.

No. of animals per sex per dose:

5 (5 additional per sex in control and high dose groups for additional 14-day non-dosing recovery period)

Control animals:

yes, concurrent no treatment

Details on study design:

- Duration of observation period following administration: 5 animals/sex/group were euthanized on the day following the last exposure for the group; the remaining 5 animals/sex in Groups 1 and 5 were euthanized following a 14-day non-dosing recovery period.
- Frequency of observations and weighing: Clinical observations were performed daily, prio to exposure, during exposure, and 0-1 hour following the end of exposure. Body weights were recorded within 4 days of receipt, the week prior to randomization, on the day of randomization, on Study Day 0, and twice weekly therafter.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight,organ weights, histopathology

Positive control:

Not applicable

Examinations

Observations and examinations performed and frequency:

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Clinical observations were performed daily, prior to exposure, during exposure, and 0-1 hour following the end of exposure.
BODY WEIGHT: Yes
- Time schedule for examinations: Body weights were recorded within 4 days of receipt, the week prior to randomization, on the day of randomization, on Study Day 0, and twice weekly thereafter.
FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No
HAEMATOLOGY: Yes
- Time schedule for collection of blood: Study Day 28 or 42 (at necropsy)
- Anaesthetic used for blood collection: Yes, isoflurane
- Animals fasted: Yes, overnight
- How many animals: All animals
- Parameters examined: total leukocyte count, erythrocyte count, hemoglobin, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, platelet count, prothrombin time, activated partial thromboplastin time, reticulocyte count (percent and absolute), mean platelet volume, differential leukocyte count (percent and absolute: neutrophil, lymphocyte, monocyte, eosiniphil, basophil, large unstained cell, red cell distribution width, hemoglobin distribution width, platelet estimate, red cell morphology
CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Study Day 28 or 42 (at necropsy)
- Animals fasted: Yes, overnight
- How many animals: All animals
- Parameters examined: albumin, total protein, globulin, albumin/globulin ratio, total bilirubin, urea nitrogen, creatinine, alkaline phosphatase, alanine aminotransferase, aspartate aminotransferase, gamma glutamyltransferase, glucose, total cholesterol, calcium, chloride, phosphorous, potassium, sodium, sorbitol dehydrogenase, triglycerides
URINALYSIS: Yes
- Time schedule for collection of urine: Study Day 28 or 42 (at necropsy)
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes, overnight
- Parameters examined: specific gravity, pH, urobilinogen, total volume, color, clarity, protein, glucose, ketones, bilirubin, occult blood, leukocytes, nitrites, microscopy of sediment

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes
Tissues examined: Adernals, aorta, bone with marrow (femur with joint, sternebrae), brain, cervix, epididymides, eyes with optic nerves, gastrointestinal tract, harderian glands, heart, kidneys, lacrimal gland, larynx, liver, lungs, lymph nodes mammary gland, nasal cavity, ovaries, pancreas, peripheral nerve, Peyer's patches, pituitary, pharynx, prostate, salivary glands, seminal vesicles, skeletal muscle, skin, spinal cord, spleen, testes, thymus, thyroid, trachea, urinary bladder, uterus, gross lesions.

Results and discussion

Results of examinations

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:

effects observed, treatment-related

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

no effects observed

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

no effects observed

Details on results:

CLINICAL SIGNS AND MORTALITY: No mortality or test substance-related clinical observations were noted during the study.
BODY WEIGHT AND WEIGHT GAIN: Test substance-related lower body weight gains and/or losses resulting in lower body weights were noted in the 500, 1000, and 1500 ppm group males and in all test substance-treated females during the exposure period. These effects were statistically significant in the 1000 and 1500 ppm group males and females. There were no treatment-related effects noted during the recovery period.
FOOD CONSUMPTION: Test substance-related lower food consumption was noted in the 1000 and 1500 ppm group males and females during the exposure period. No treatment-related effects were noted during the recovery period.
HAEMATOLOGY: On Study Day 28, mean erythrocytic parameters (RBC count, hemoglobin, and hematocrit values) were slightly higher than controls with a generally dose-related response in the 500, 1000, and 1500 ppm group males and females. Mean absolute reticulocyte counts were lower than controls in the 1000 and 1500 ppm group males (28% and 39%, respectively). Mean hemoglobin distribution width was slightly lower (6%) than controls in the 1500 ppm group males. Following the recovery interval, there were no meaningful consistent alterations in hematology parameters.
CLINICAL CHEMISTRY: No clinical chemistry parameters were altered with a dose-response relationship.
URINALYSIS: There were no test substance-related alterations in urinalysis parameters.
ORGAN WEIGHTS: Test substance-related lower thymus weights were noted in the 1500 ppm group males at the Study Day 28 primary necropsy. Lower mean absolute thymus weights were also noted in the 500 and 1000 ppm group males and lower mean thymus to body weight and thymus to brain weight ratios in the 1000 and 1500 ppm group males were also noted but the differences were not statistically significant. Mean thymus weights were higher than the control group at the Study Day 42 recovery necropsy. There were no correlating histopatholic finding within the thymus.
GROSS PATHOLOGY: No test substance-related gross lesions were observed upon necropsy.
HISTOPATHOLOGY: NON-NEOPLASTIC: Test-substance related microscopic findings were noted in the nasal cavity in the 250, 500, 1000, and 1500 ppm group males and females from the Study Day 28 primary necropsy, and in the 1500 ppm group males and females from the Study Day 42 recovery necropsy. Findings were also noted in the pharynx in the 500, 1000, and 1500 ppm group males and females and in the 250 ppm group males from the Day 28 primary necropsy and in the 1500 ppm group females from the study day 42 recovery necropsy. Hyperplasia of the transitional epithelium, degeneration and regeneration of the respiratory epithelium, and/or degeneration and regeneration of the olfactory epithelium were observed in the nasal cavity in males and females from all test substance-exposed groups at the Day 28 primary necropsy aside from the 250 ppm group males. Degeneration of the respiratory epithelium was characterized bgy loss of cilia, presence of cytoplasmic vacuolation and/or bleb formation, and sparse inflammatory cell cell infiltration. In addition, decreased mucous cells and hyperplasia of the submucosal glands were observed in the rostral levels. The olfactory epithelium was most severely affected in all groups except in the 250 ppm group. In the 250 ppm group animals, the transitional and respiratory epithelia of nasal cavity level II were often most severely affected. Degeneration and regeneration were almost always present concurrently within the same sections, indicating repetitive injury to the epithelial cells. No toxicologically relevant microscopic findings were observed in the larynx, trachea or lungs.
HISTOPATHOLOGY: NEOPLASTIC: No test article-related effects were noted.

Effect levels

Dose descriptor:

NOEC

Effect level:

ca. 500 ppm

Sex:

male/female

Basis for effect level:

other: overall effects clinical signs; mortality; body weight; haematology; clinical chemistry; urinalysis; gross pathology; organ weights; histopathology

Target system / organ toxicity

Critical effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

Based on the results of the study, the No Observed Adverse Effect Concentration (NOAEC) is 516 ppm for the test article.

Executive summary:

The repeat-dose inhalation toxicity potential of the test article was evaluated in male and female Sprague-Dawley rats via whole-body inhalation for 6 hours/day, 5 days/week, for 4 weeks (20 total exposures). The study was performed in compliance with EPA GLP 40 CFR 792 (1989) and OECD GLP (1997). The study method was based on OECD Guideline 412 (2009). Main phase rats (5/sex/group) were exposed to filtered air (control) or filtered air containing 0, 250, 500, 1000, or 1500 ppm (nominal) of the test article, for 6 hour exposures, 5 days per week for 4 weeks (20 total exposures). Recovery rats (5/sex/group) were exposed to filtered air or 1500 ppm test article in the same manner followed by a 2 week recovery period after the exposure period. Mean analyzed exposure concentrations were 252, 516, 1033, and 1512 ppm test article. Clinical observations were performed daily, prior to exposure, during exposure, and 0-1 hour following the end of exposure and detailed physical examinations were performed weekly. Body weights were recorded within 4 days of receipt, the week prior to randomization, on the day of randomization, on Study Day 0, and twice weekly thereafter. Blood and urine were collected at necropsy for all animals (Day 28 for main group animals, Day 42 for recovery animals) and hematology and urinary parameters were evaluated. Macroscopic examinations were performed on all animals upon necropsy. Selected tissues were examined microscopically from all animals in the control and high concentration groups at the primary necropsy (Day 28). Target tissues were examined from all animals in the 250, 500 and 1000 ppm groups at the primary necropsy. No mortality or test substance related clinical observations were noted during the study. Test substance related lower body weight gains and/or losses resulting in lower body weights were noted in the 500, 1000, and 1500 ppm group males and in all test substance treated females during the exposure period. These effects were statistically significant in the 1000 and 1500 ppm group males and females. There were no treatment related effects noted during the recovery period. Test substance related lower food consumption was noted in the 1000 and 1500 ppm group males and females during the exposure period. No treatment related effects were noted during the recovery period. On Study Day 28, mean erythrocytic parameters (RBC count, hemoglobin, and hematocrit values) were slightly higher than controls with a generally dose related response in the 500, 1000, and 1500 ppm group males and females. Mean absolute reticulocyte counts were lower than controls in the 1000 and 1500 ppm group males (28% and 39%, respectively). Mean hemoglobin distribution width was slightly lower (6%) than controls in the 1500 ppm group males. Following the recovery interval, there were no meaningful consistent alterations in hematology parameters. No clinical chemistry or urinalysis parameters were altered with a dose response relationship. Test substance related lower thymus weights were noted in the 1500 ppm group males at the Study Day 28 primary necropsy. Lower mean absolute thymus weights were also noted in the 500 and 1000 ppm group males and lower mean thymus to body weight and thymus to brain weight ratios in the 1000 and 1500 ppm group males were also noted but the differences were not statistically significant. Mean thymus weights were higher than the control group at the Study Day 42 recovery necropsy. There were no correlating histopathologic finding within the thymus. No test substance related gross lesions were observed upon necropsy. Test-substance related microscopic findings were noted in the nasal cavity in the 250, 500, 1000, and 1500 ppm group males and females from the Study Day 28 primary necropsy, and in the 1500 ppm group males and females from the Study Day 42 recovery necropsy. Findings were also noted in the pharynx in the 500, 1000, and 1500 ppm group males and females and in the 250 ppm group males from the Day 28 primary necropsy and in the 1500 ppm group females from the study day 42 recovery necropsy. Hyperplasia of the transitional epithelium, degeneration and regeneration of the respiratory epithelium, and/or degeneration and regeneration of the olfactory epithelium were observed in the nasal cavity in males and females from all test substance exposed groups at the Day 28 primary necropsy aside from the 250 ppm group males. Degeneration of the respiratory epithelium was characterized by loss of cilia, presence of cytoplasmic vacuolation and/or bleb formation, and sparse inflammatory cell cell infiltration. In addition, decreased mucous cells and hyperplasia of the submucosal glands were observed in the rostral levels. The olfactory epithelium was most severely affected in all groups except in the 250 ppm group. In the 250 ppm group animals, the transitional and respiratory epithelia of nasal cavity level II were often most severely affected. Degeneration and regeneration were almost always present concurrently within the same sections, indicating repetitive injury to the epithelial cells. No toxicologically relevant microscopic findings were observed in the larynx, trachea or lungs. Based on the results of the study, the No Observed Adverse Effect Concentration (NOAEC) is 516 ppm for the test article.