Registration Dossier

Administrative data

Endpoint:
acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1997-10-09 to 1998-05-05
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline-conform study conducted under GLP.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1998
Report Date:
1998

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
Deviations:
no
GLP compliance:
yes
Test type:
fixed concentration procedure
Limit test:
yes

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: Dr. K. Thomae GmbH, Biberach, Germany
- Age at study initiation: approx. 8 - 9 weeks
- Weight at study initiation: mean body weight males 269 g, mean body weight females 202 g
- Fasting period before study:
- Housing: Animals were housed singly in cages type DK III - Diet: KLIBA rat/mouse/hamster laboratory diet 10 mm pellets, Klingentalmühle AG, Kaiseraugst, Switzerland, ad libitum
- Water: tap water ad libitum
- Acclimation period: at least 1 week

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24°C
- Humidity (%): 30-70%
- Air changes (per hr): no data, however the cages were housed in fully air-conditioned rooms
- Photoperiod (hrs dark / hrs light): 12 h/12 h

IN-LIFE DATES: From: 1997-10-09 To: 1997-10-23

Administration / exposure

Route of administration:
inhalation: vapour
Type of inhalation exposure:
whole body
Vehicle:
air
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus:
Whole-body inhalation system IKA 02 (glass-steel construction), BASF Aktiengesellschaft.
Technical equipment: continuous infusion pump Perfusor VII (B. Braun), vaporizer with thermostat (glass, BASF), mixing vessel (glass, BASF)
- Exposure chamber volume: 200 L
- Method of holding animals in test chamber: the animals were kept singly in compartmentalized wire cages, and were exposed inside the chamber
- Source and rate of air: A Supply air flow (compressed air) of 3000 l/h was used for the test group. The exhaust air flow was 3200 L/h. An air changes of about 15 times per hour can be calculated by dividing the supply air flow by the volume of the inhalation system. The about 7% higher amount of exhaust air, which was adjusted by means of a separate exhaust air system, achieved a slightly negative pressure inside the exposure system. This ensured that no contamination of the laboratory occurred as result of possible leakages from the inhalation chamber. Additionally, the exposure system was located inside an exhaust cabin in the air-conditioned laboratory. The inhalation atmosphere was offered to the animals for inhalation for 4 hours after equilibration of the inhalation System (t99 about 20 min).
- Method of conditioning air: A vapour-air mixture was generated. For the test group the test substance was supplied to the heated vapouriser (about 49.5°C) by means of the continuous infusion pump. The vapours that developed were taken up by the supply air passed into the exposure System. Flow rate of the test substance to the atomizer was 25.0 mL/h.
- Treatment of exhaust air: no data
- Temperature, humidity, pressure in air chamber: 24.2 °C, 12%, slightly negative pressure inside the exposure system

TEST ATMOSPHERE
- Brief description of analytical method used:
For the quantitative determination of the vapour concentration a gas chromatographical method was used.
Gas chromatographical analysis

Equipment: GC HP 5840 A (Hewlett Packard)
The following gaschromatographical conditions were used:
column: glass, 2 m x 3 mm
separation phase: 10% Ucon LB 550X
solid phase: Chromosorb W AW /HP 100/120 mesh
carrier gas: helium
carrier gas flow rate: 25.1 mL/min
hydrogen: 30 mL/min
air: 240 mL/min
temperature . 130°C/min
detector temp. (FID) : 2000C
injector temp.: 200°C
internal standard (i.st.): C12 KW
injection volume: 1µL
retention time (i.st.): 4.18 min
retention time sample: 2.90 min

Calibration:
Method set up:
During the set up of the analytical procedure a calibration curve was prepared in the solvent with the test substance to be investigated to show linearity in the suitable concentration range of the samples. The calibration curve of the substance in the solvent used had a linearity range from 20.2 to 60.6 mg/50 mL (method dated Oct. 06, 1997; for details see raw data).

Routine analysis during test period:
For routine analysis a one-point calibration of the analytical procedure was prepared for the analytical campaign based on the method mentioned above.

Sample preparation:
The samples obtained were taken up in 2-Propanole in a 50-mL calibrated flask. The flask was filled up to the calibration mark after an internal standard was added using a pipette.

Calculation of the concentration:
The concentrations were calculated in mg/L from the analytically determined mass values of the test substance and the sample volumes of the inhalation atmosphere.

- Samples taken from breathing zone: yes, sampling position immediately adjacent to the animals' noses
Analytical verification of test atmosphere concentrations:
yes
Duration of exposure:
4 h
Concentrations:
Nominal: 7.48 mg/L
Analytical: 6.3 mg/L (maximum attainable vapour concentration)
No. of animals per sex per dose:
5
Control animals:
yes
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: The body weight of the animals was determined just prior to exposure, after 7 days and at the end of the observation period. A check for overt clinical signs of toxicity or mortality as well as a check for the presence of feed and drinking water was made twice a day on workdays and once daily on weekends and public holidays.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, irregular respiration, dragging respiration, attempts to escape, squatting posture, piloerection, body weight

Results and discussion

Effect levels
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 6.3 mg/L air (analytical)
Based on:
test mat.
Exp. duration:
4 h
Mortality:
None
Clinical signs:
Attempts to escape, irregular and dragging respiration as well as squatting posture and piloerection. No clinical signs could be detected from post exposure day 5 onward.
Body weight:
Body weight development of the animals was not influenced.
Gross pathology:
No macroscopic pathologic findings were noted in animals examined at the end of the study.

Applicant's summary and conclusion