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Toxicological information

Acute Toxicity: inhalation

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Administrative data

Endpoint:
acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
December 15, 1994 - February 3, 1995
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1995
Report Date:
1995

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
GLP compliance:
yes
Test type:
standard acute method
Limit test:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Hilltop Lab Animals, Scottdale
- Weight at study initiation: 228-239 g (males), 220-235 g (females)
- Housing: individually (in suspended stainless steel caging with mesh floors)
- Diet: Purina Rodent Chow
- Water: Tap water (ad libitum except during exposure)
- Acclimation period: 23 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20.6 – 23.9
- Photoperiod (hrs dark / hrs light): 12/12


Administration / exposure

Route of administration:
inhalation: aerosol
Type of inhalation exposure:
whole body
Vehicle:
clean air
Mass median aerodynamic diameter (MMAD):
2.2 µm
Geometric standard deviation (GSD):
1.93
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Rectangular whole body perspex chamber with prechamber operated under slight negative pressure
- Exposure chamber volume: 150 L
- Source and rate of air: Approximately 20 liters per minute (Lpm) was supplied from a compressed gas cylinder (Airco #3001) of dry, breathing-grade air to the spray atomization nozzle. Compressed airflow was measured with a Dwyer Rotameter Model #VFB (0-40 Lpm). Approximately 20.6 Lpm of filtered conditioned room air was supplied as diluent air. Room airflow was measured with an Omega Mass Flow Meter, Model #FMA 5613. Chamber airflow was monitored throughout the exposure period and recorded periodically. Total airflow ranged from 40.3 to 40.7 with a mean of 40.6 Lpm,
- System of generating aerosols: The test atmosphere was generated using a 1/4 inch JCO atomizer, FC4 fluid cap and 1502 air cap (Spraying Systems Inc.). Compressed air was supplied at 25 psi. The test substance was metered to the atomization nozzle through Size 14 Master Flex Tygon tubing, using a Master Flex Pump Model 7520-35.
- Method of particle size determination: An eight-stage Andersen cascade impactor was used. Samples were withdrawn from the breathing zone of the animals on two occasions. The filter paper collection stages were weighed before and after sampling to determine the mass collected at each stage. The aerodynamic mass median diameter and geometric standard deviation were determined graphically using two cycle logarithmic probit axes.
- Temperature, humidity, pressure in air chamber: 21.7-23.3°C, 40-45%, 25 psi

TEST ATMOSPHERE
- Brief description of analytical method used: Gravimetric samples were withdrawn on eight occasions from the breathing zone of the animals. Samples were collected using 25 mm glass fiber filters (GF/B Whatman) in filter holders attached by % inch tygon tubing to a General Electric vacuum pump Model 5KH10GGR28. Filter papers were weighed before and after collection to determine the mass collected. This value was divided by the total volume of air sampled to determine the chamber concentration. The collections were carried out for 3 minutes at airflows of 4 Lpm. Sample airflows were measured using an Omega Flow Meter Model #FMA 5610.
- Samples taken from breathing zone: yes


Analytical verification of test atmosphere concentrations:
yes
Duration of exposure:
4 h
Concentrations:
2.12 mg/L (gravimetric chamber concentration)
10.93 mg/L (nominal concentration)
No. of animals per sex per dose:
5 male, 5 female
Control animals:
no
Details on study design:
- Duration of observation period following administration: 50 days
- Frequency of observations and weighing: Individual weights of the animals were recorded just prior to test substance exposure (initial) and again on days 7, 14, 21, 28, 35, 42 and 49 or after death. Cage-Side observations were made at least every 30 minutes during exposure, upon chamber removal and at least once daily thereafter for 50 days.
- Necropsy of survivors performed: yes
- Other examinations performed: histopathology (lung tissue)

Results and discussion

Preliminary study:
Prior to initiation of the full inhalation study, pre-test trials were conducted to establish generation procedures for achieving as closely as possible the desired (2.0 mg/L) or highest possible chamber concentration and desired particle size distribution (mass median aerodynamic diameter less than or equal to 4/ µm). The exposure procedures and atomization equipment used were based on results of pre-test trial number 2 which provided a gravimetric concentration of 2.0 mg/L and a mass median aerodynamic diameter of 2.1 µm.
Effect levels
Key result
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 2.12 mg/L air
Based on:
test mat.
Exp. duration:
4 h
Mortality:
One male animal died on test day 11.
Clinical signs:
In chamber animal observation included ocular and nasal discharge, facial staining, irregular respiration, dyspnea, gasping, hunched posture and hypoactivity. Upon chamber removal, similar clinical signs persisted in all animals. Within several days of exposure animals also developed piloerection, rales, emaciation, ano-genital staining and an unthrifty appearance. Prior to death, the decedent exhibited reduced food consumption, reduced fecal volume and abdominal distention. Transient signs of abnormal respiration (rales and irregular respiration! were noted in several rats for an extended period of time.

Body weight:
Transient body weight loss was noted during the study.
Gross pathology:
Gross necropsy of the decedent revealed discoloration of the lungs, gastro intestinal tract and liver, gaseous distention of the gastrointestinal tract, edema of the lungs and retraction of the testes into the abdominal cavity. Gross necropsy findings at terminal sacrifice were generally unremarkable. Apart from red lung discoloration which is consistent with euthanasia by CO2 inhalation, all tissues and organs appeared normal.
Other findings:
- Histopathology: Histological evaluation of the lung tissue of the survivors revealed pulmonary congestion: the presence of alveolar macrophages; the accumulation of mucous in the bronchi and bronchioles; and chronic interstitial inflammation. Although the congestion may have been due to the euthanasia procedure, it along with interstitial inflammation may have resulted from exposure to a very mild irritant.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
Based on the results of testing, the single exposure Acute Inhalation Toxicity LC50 of the test substance is greater than 2.12 mg/L.
Executive summary:

An Acute Inhalation Toxicity test according OECD Guideline 403 was conducted with rats to determine the potential of the test item to produce toxicity via the inhalation route.

After establishing the desired generation procedures during pre-test trials, ten healthy rats were exposed to the test atmosphere for 4 hours. Chamber concentration and particle size distribution of the test substance were determined periodically during the exposure period. The animals were observed for signs of gross toxicity and mortality at least once daily for 50 days. Bodyweights were recorded just prior to exposure and again or days 71 4, 21, 28, 35, 42 and 49 or after death. Gross necropsies were performed on all animals. The lungs of all survivors were removed at terminal sacrifice, preserved in formalin and evaluated histopatholggically.

One male died on day 11 of the study. All other animals survived exposure to the test atmosphere. The gravimetric chamber concentration was 2.12 mg/L. Based on graphic analysis of the particle size distribution as measured with an Anderson Cascade Impactor, the mass median aerodynamic diameter was estimated to be 2.2 µm.

In chamber animal observations included ocular and nasal discharge, facial staining, irregular respiration, dyspnea, gasping, hunched posture and hypoactivity. Upon chamber removal, similar clinical signs persisted in all animals. Within several days of exposure animals also developed piloerection, rales, emaciation, ano-genital staining and an unthrifty appearance. Prior to death, the decedent exhibited reduced food consumption, reduced fecal volume and abdominal distention. Transient signs of abnormal respiration were noted in several rats for an extended period of time. In order to assess the reversal of symptoms, the observation period was extended to 50 days. Although clinical signs persisted and transient bodyweight loss was noted during the study, all surviving rats gained weight over the entire 50 day observation period. Gross necropsy of the decedent revealed discoloration of the lungs, gastro-intestinal tract and liver, gaseous distention of the gastro-intestinal tract, edema of the lungs and retraction of the testes into the abdominal cavity. Gross necropsy findings at terminal sacrifice were generally unremarkable. Apart from red lung discoloration which is consistent with euthanasia by CO2 inhalation, all tissues and organs appeared normal. In order to further investigate the abnormal respiratory findings noted during the observation period, the lungs of all surviving animals were removed at gross necropsy and preserved in neutral buffeted formalin. Histological evaluation of the lung tissue revealed pulmonary congestion: the presence of alveolar macrophages; the accumulation of mucous in the bronchi and bronchioles; and chronic interstitial inflammation. Although the congestion may have been due to the euthanasia procedure, it along with interstitial inflammation may have resulted from exposure to a very mild irritant.

Based on the results of testing, the single exposure Acute Inhalation Toxicity LC50 of the test substance is greater than 2.12 mg/L.