Registration Dossier

Diss Factsheets

Administrative data

Endpoint:
acute toxicity: inhalation
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2012
Report date:
2012

Materials and methods

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

Test material

Constituent 1
Reference substance name:
54074-94-1
Cas Number:
54074-94-1
IUPAC Name:
54074-94-1
Constituent 2
Reference substance name:
isopropyl-3-hydroxybutyrate
IUPAC Name:
isopropyl-3-hydroxybutyrate
Test material form:
other: colorless liquid
Details on test material:
The test substance, identified as Isopropyl-3-hydroxybutyrate, was received on July 19, 2012 and was further identified with PSL Reference Number 120719-5D. The test substance was stored at room temperature. The sample was aerosolized as received, shaken before dispensing. Documentation of the methods of synthesis, fabrication, or derivation of the test substance is retained by the Sponsor.
The following information related to the characterization of the test substance was provided by the Sponsor:
Composition: Isopropyl-3-hydroxybutyrate > 99.9% CAS #54074-94-1
Physical Description: Colorless liquid
Solubility: Soluble in water.
Stability: Test substance was expected to be stable for the duration of testing.
Expiration Date: December 31,2012

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
4.A.1 Housing: The animals were singly housed in suspended stainless steel caging with mesh floors, which conform to the size recommendations in the most recent Guide for the Care and Use of Laboratory Animals (Natl. Res. Council of the National Academies, 2011). Litter paper was placed beneath the cage and was changed at least three times per week.

4.A.2 Animal Room Temperature and Relative Humidity Ranges: 19-23°C and 64-76%, respectively. Humidity was above the targeted upper limit for 10 days during the study. A portable dehumidifier was used to lower the humidity levels dw-ing this time.

Animal Room Air Changes/Hour: 12 or 13. Airflow measurements are evaluated regularly and the records are kept on file at Product Safety Labs.

Photoperiod: 12-hour light/dark cycle

Acclimation Period: 9 days

Food: Harlan Teklad Global16% Protein Rodent Diet® 2016. The diet was available ad libitum, except during the exposure.

Water: Filtered tap water was supplied ad libitum by an automatic water dispensing system except during exposure.

Contaminants: There were no known contaminants reasonably expected to be found in the food or water at levels which would have interfered with the results of this study. Analyses of the food and water are conducted regularly and the records are kept on file at Product Safety Labs.

HOUSING
Cage: Each cage was identified with a cage card indicating at least the study number and identification and sex of the animal.

Animal: A number was allocated to each rat on receipt and a stainless steel ear tag bearing this number was attached to the rat. This number, together with a sequential animal number assigned to study 34939, constituted unique identification.

Administration / exposure

Route of administration:
inhalation: mist
Type of inhalation exposure:
nose only
Vehicle:
air
Details on inhalation exposure:
PRE-TEST TRIALS
Prior to initiation of the full inhalation study, pre-test trials were conducted to establish generation procedures to achieve, to the extent possible, the desired chamber concentration (5.0 mg/L) and desired pmiicle size distribution (mass median aerodynamic diameter between 1 and 4 um). In these trials, the following adjustments were made in an attempt to achieve these objectives:
Air Pressure: constant
Compressed Generator Airflow: constant
Compressed Mixing Airflow: varied
Total Airflow: constant
Pump Setting: varied
Pump Type: constant
Tube Size: constant
Atomization System: constant
Fluid Cap: constant
Air Cap: constant

The procedures and aerosolization equipment used in the full test were based on the results of pre-test trial number 6 which provided a gravimetric concentration of 5.15 mg/L and a mass median aerodynamic diameter of 2.50 um.

INHALATION PROCEDURES
The exposure chamber, air supply and equipment used to measure particle size distribution, airflow and chamber concentration were the same as used during the pre-test trials and are described below.

Nose-Only Exposure Chamber: A nose-only inhalation chamber with an intemal volume of approximately 28 liters (Nose-Only Inhalation Chamber, ADG Developments LTD) was used for exposure. Animals were individually housed in polycarbonate holding tubes which seal to the chamber with an "0" ring during exposure. The base unit terminates the chamber with a 0.5-inch diameter tube for discharged air.

Air Supply: Approximately 30.0 liters per minute (Lpm) of filtered air was supplied by an air compressor (Powerex Model: SES05) to the spray atomization nozzle. An additional 6.0 Lpm of compressed mixing air from the air compressor was introduced into the chamber to help uniformly distribute the test atmosphere by creating a vortex at the chamber inlet. Compressed airflow was measured with a Mass Flowmeter (Omega, Model #FMA-5613). Chamber airflow was monitored throughout the exposure period and recorded periodically. Total airflow was 36.0 Lpm. Based on the volume of the inhalation chamber, this airflow provided approximately 77 air changes per hour during the study.

Ambient Conditions: The exposure tube temperature and relative humidity ranges during exposure were 22-24°C and 47-66%, respectively. The room temperature and relative humidity ranges during exposure were 21-22°C and 71-79%, respectively. A dehumidifier was used in the room during exposure. The measurements inside the exposure tube were made with a Temperature-Humidity Recorder (Fisher Scientific, Model #11-661-18) and room conditions were measured with a TemperatureHumidity Monitor (Fisher Scientific, Model #11-661-18). Temperature and relative
humidity values were recorded every 15 minutes for the first hour of exposure and every 30 minutes thereafter.

Atmosphere Generation: The test atmosphere was generated using a 1;4 inch JCO atomizer (Spraying Systems Co.), FC3 fluid cap (Robert Miller Associates) and 70SS air cap (Spraying Systems Co.). Compressed air was supplied at 30 psi. The test substance was metered to the atomization nozzle through size 14 BPT tubing, using a peristaltic pump (Master Flex, Model #7520-35).

Chamber Concentration Measurements: Gravimetric samples were withdrawn at 6 intervals from the breathing zone of the animals. Samples were collected using 3 7 mm glass fiber filters (GF/B Whatman) in a filter holder attached by 1/4 inch tygon tubing to a vacuum pump (GAST, Model #1531-107B-G557X). Filter papers were weighed before and after collection to detetmine the mass collected. This value was divided by the total volume of air sampled to detennine the chamber concentration. The collections were carried out for 1 minute at airflows of 4.0 Lpm. Sample airflows were measured using a Mass Flowmeter (Aalborg, Model #GFC-17).

Particle Size Distribution: An eight-stage ACFM Andersen Ambient Particle Size Sampler was used to assess the particle size distribution of the test atmosphere. Samples were withdrawn from the breathing zone of the animals at two intervals. The filter paper collection stages were weighed before and after sampling to determine the mass collected upon each stage. The aerodynamic mass median diameter and geometric standard deviation were calculated using two-cycle logarithmic pro bit axes.

Exposure Period: The animals were exposed to the test atmosphere for 4 hours and 4 minutes. The exposure period was extended beyond 4 hours to allow the chamber to reach equilibrium (T99). The times for 90 and 99% equilibration of the chamber atmosphere were 1.8 and 3.6 minutes, respectively. At the end of the exposure period, the generation was tenninated and the chamber was operated for a further 15 minutes with clean air. At the end of this period the animals were removed from the exposure tube. Prior to being returned to their cages excess test substance was
removed from the fur of each animal.
Analytical verification of test atmosphere concentrations:
no
Duration of exposure:
ca. 4 h
Concentrations:
>5.11 mg/L
No. of animals per sex per dose:
5
Control animals:
no
Details on study design:
Selection of Animals
On the day of and prior to exposure, the rats were examined for health and weighed. Ten healthy naive rats (five males and five females; not previously tested) were selected for test.

Body Weights
Individual body weights of the animals were recorded prior to test substance exposure (initial) and again on Days 1, 3, 7, and 14 (termination).

Cage-Side Observations
All animals were observed for mortality during the exposure period. The animals were examined for signs of gross toxicity, and behavioral changes upon removal from the exposure tube and at least once daily thereafter for 14 days. Observations included gross evaluation of skin and fur, eyes and mucous membranes, respiratory, circulatory, autonomic and central nervous systems, somatomotor activity and behavior pattern. Patiicular attention was directed to observation of tremors, convulsions, salivation, diarrhea, and coma.

Necropsy
All rats were euthanized via C02 inhalation on Day 14. Gross necropsies were performed on all animals. Tissues and organs ofthe thoracic and abdominal cavities were examined.

Results and discussion

Effect levelsopen allclose all
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 5.1 mg/L air (analytical)
Based on:
test mat.
Sex:
male/female
Dose descriptor:
LC0
Effect level:
> 5.1 mg/L air (analytical)
Based on:
test mat.
Mortality:
none
Clinical signs:
other: All animals survived exposure to the test atmosphere.
Body weight:
Following exposure, all rats exhibited irregular respiration. However, they recovered by Day 4 and appeared active and healthy for the remainder of the observation period. Although all animals lost body weight on Day 1, they showed continued weight gain thereafter and gained weight through Day 14 observation period.
Gross pathology:
no abnormalities observed

Applicant's summary and conclusion

Interpretation of results:
practically nontoxic
Remarks:
Migrated information Criteria used for interpretation of results: expert judgment
Conclusions:
Under the conditions of this study, the single exposure acute inhalation LC50 of Isopropyl-3- hydroxybutyrate as a mist is greater than 5.11 mg/L in male and female rats.
Executive summary:

An acute inhalation toxicity test was conducted with rats to determine the potential for Isopropyl- 3-hydroxybutyrate to produce toxicity from a single exposure via the inhalation (nose-only exposure) route. Under the conditions of this study, the single exposure acute inhalation LC50 of the test substance as a mist is greater than 5.11 mg/L in male and female rats. After establishing the desired generation procedures during pre-test trials, ten healthy rats (5/sex) were exposed to the test atmosphere for 4 hours. Chamber concentration and particle size distributions of the test substance were determined periodically during the exposure period. The animals were observed for mortality, signs of gross toxicity, and behavioral changes at least once daily for 14 days following exposure. Body weights were recorded prior to exposure and again on Days 1, 3, 7, and 14 (termination). Necropsies were performed on all animals at terminal sacrifice. The gravimetric chamber concentration was 5.11 mg/L. The mass median aerodynamic diameter was calculated to be 2.26 11m based on graphic analysis of the particle size distribution as measured with an ACFM Andersen Ambient Particle Sizing Sampler.

All animals survived exposure to the test atmosphere. Following exposure, all rats exhibited irregular respiration. However, they recovered by Day 4 and appeared active and healthy for the remainder of the observation period. Although all animals lost body weight on Day 1, they showed continued weights gain thereafter and gained weight through Day 14. No gross abnormalities were noted for any of the animals when necropsied at the conclusion of the 14-day observation period.