Registration Dossier

Administrative data

Endpoint:
acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2014-03-24 to 2014-05-27
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline and GLP study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2014
Report Date:
2014

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
Version / remarks:
2009
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.2 (Acute Toxicity (Inhalation))
Version / remarks:
2008
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.1300 (Acute inhalation toxicity)
Version / remarks:
1998
Deviations:
no
GLP compliance:
yes (incl. certificate)
Test type:
acute toxic class method
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Laboratories, B.V., Kreuzelweg 53, 5961 NM Horst/Netherlands
- Age at study initiation: 10 weeks (group 1), 9 weeks (group 2)
- Weight at study initiation: Males (256.4 to 327 g), Females (174.8 to 206.8 g)
- Housing: Makrolon cages type-4
- Diet: Pelleted standard Harlan Teklad 2914 C rodent maintenance diet, ad libitum
- Water: Community tap-water from Itingen, ad libitum
- Acclimation period: Performed for 15 days (group 1) or 7 days (group 2)

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/-3 °C
- Humidity (%): 30-70 %
- Air changes (per hr): 10-15 air changes
- Photoperiod (hrs dark / hrs light): 12 hour fluorescent light/12 hour dark cycle with at least eight hours music during the light period.

Administration / exposure

Route of administration:
inhalation: vapour
Type of inhalation exposure:
nose only
Vehicle:
air
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Nose only, flow past exposure chamber
- Exposure chamber volume: Flow of air at each tube was 1.0 L/min.
- Method of holding animals in test chamber: The animals were confined separately in restraint tubes which were positioned radially around the exposure chamber.
- Source and rate of air: Flow of air at each tube was 1.0 L/min.
- Method of conditioning air: The test item was pumped into a glass flask. Compressed, filtered, and dried air was supplied into the glass flask through a metal nebulization tube. The glass flask was kept at a temperature of 28 to 29 °C with a thermal regulating device set to facilitate the process of vaporization.


TEST ATMOSPHERE
- Brief description of analytical method used: Test atmosphere samples were collected at room temperature in a solvent trap constructed with two fritted wash bottles with approximately 100 mL acetonitrile (100%, lot nos. 1325508002 and 142308006, JT Baker) each, connected one after the other to prevent carry-over and ensure all test item was trapped (samples B1 and B2). The wash bottles were placed in a bucket and cooled with an ice/water mixture. All test item was trapped in the first wash bottle (samples B1). The duration of sampling was 10 min. The samples were filled up to 100 g of Acetonitrile and transferred into appropriate labeled vials and forwarded immediately at ambient temperature to the scientist responsible for formulation analysis and stored at -20 ± 5 °C until analysis. The samples were analyzed using a GC method.
- Samples taken from breathing zone: yes
- Particle size distribution: The concentration of the test item in the inhalation chamber was controlled by regulating the flow of the test item into the atmosphere generation glass flasks and by dilution with compressed, filtered, and dried air to achieve the concentrations required for this study.- Rationale for the selection of the starting concentration: The target concentration of 20 mg/L air for 4 hours is the recommended concentration for a limit test (OECD 403, “Acute Inhalation Toxicity”). The target concentration for group 2 was based on regulatory cutoff values.
Analytical verification of test atmosphere concentrations:
yes
Duration of exposure:
4 h
Concentrations:
Target vapour concentration: 10 and 20 mg/L air.
Chemically determined mean concentration: 10.5 mg/L air and 22.0 mg/L air.
No. of animals per sex per dose:
10 animals per dose
5 males and 5 females
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations: Viability/Mortality: Once daily during the acclimatization period, once before exposure on the day of exposure (test day 1), three times during exposure, immediately and 1 h after exposure on test day 1 and twice daily during the observation period.
Clinical Signs: Once daily during the acclimatization period, once before exposure on the day of exposure (test day 1), three times during exposure at hourly intervals (only grossly abnormal signs were detectable during exposure as animals were restrained in the exposure tubes), immediately and 1 h after exposure on test day 1 and once daily during the observation period.
Body Weights: Once during acclimatization, on test days 1 (before exposure), 2, 4, 8 and 15 (before necropsy).
- Necropsy of survivors performed: yes
Statistics:
No statistical analysis was performed as only two groups were allocated to the study.

Results and discussion

Effect levels
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 10.5 - < 22 mg/L air
Based on:
test mat.
Exp. duration:
4 h
Mortality:
All animals of group 1 exposed to a concentration of 22.0 mg/L air died within 3 hours after exposure start. Two males and one female of group 2 exposed to a concentration of 10.5 mg/L air died within 1 to 7 days after exposure. The remaining seven animals survived the scheduled observation period.
Clinical signs:
other: No clinical signs were visible in animals of group 1 exposed to a concentration of 22.0 mg/L air one hour after exposure start. Two hours after exposure start, four males and two females were unconscious. All animals died within 3 hours after exposure sta
Body weight:
All surviving animals of group 2 exposed to 10.5 mg/L air lost body weight during the first week after treatment but had started to gain body weight on test day 8, with the exception of one female, which already started to gain weight on test day 4. Due to its premature death, no body weight was recorded after exposure of male no. 11 which was found dead on test day 2. The male and the female which were found dead on test day 8 and test day 3, respectively (animal nos. 14 and 17, respectively), had continuously lost body weight until their death. In the second week after exposure, all surviving animals showed normal body weight gain.
Gross pathology:
All animals of group 1 which died during exposure to a concentration of 22.0 mg/L air showed dark red discoloration of the lungs at necropsy. No test item-related findings were noted in all females and the surviving males of group 2 exposed to a concentration of 10.5 mg/L air. The male which was found dead on test day 2 showed reddish discoloration of the lungs, the abdominal cavity of the male which was found dead on test day 8 contained watery fluid.

Applicant's summary and conclusion