Registration Dossier

Administrative data

Endpoint:
acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2013-2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP and OECD guideline compliant study

Data source

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

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Test type:
acute toxic class method
Limit test:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Substance type: orange solid
- Physical state: solid powder
- Analytical purity: 95.6 % w/w
- Expiration date of the lot/batch:
- Stability under test conditions: stable
- Storage condition of test material: room temperature

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Laboratories B.V., 5961 NM Horst, The Netherlands
- Age at study initiation: male animals approx. 9 weeks, female animals approx. 11 weeks
- Weight at study initiation: Animals of comparable weight (± 20% of the mean weight)
- Fasting period before study:
- Housing: Single housing
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 – 24°C
- Humidity (%): 30 – 70%
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 2013-12-02 To: 2013-12-18

Administration / exposure

Route of administration:
inhalation: dust
Type of inhalation exposure:
nose only
Vehicle:
air
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Head nose inhalation system INA 10 (glass steel construction)
- Exposure chamber volume: 34L
- Method of holding animals in test chamber: restraining tubes
- Source and rate of air: Air passed through activated charcoal and a HEPA filter. Flow rates: 0.8 m³/h (during the first 2 hours), 1.0 m³/h (during hour 3), 1.3 m³/h (during hour 4)
The flow was adjusted and continuously measured with a flowmeter (Yokogawa).
During technical trial, an atmospheric concentration of slightly higher than 5 mg/L had been achieved using a supply air flow of 0.8 m³/h and an exhaust air flow of 0.6 m³/h. During the study however, the measured concentrations were below 5 mg/L, probably due to the presence of animals. In order to achieve a higher atmospheric concentration, the supply air flow was increased to 1.0 mg/m³ in hour 3, and subsequently increased to 1.3 m³/h.

- Method of conditioning air: Dosing-wheel dust generator
- System of generating particulates/aerosols: The test substance was des-agglomerated in a mixer (mixing for 10 seconds) under addition 1% AEROSIL® 200 and 1% AEROSIL® R 972 before introduction into the dust generator.) The dust was produced inside the dust pre-chamber using a
dosing wheel dust generator and compressed air. The dust was passed via the cyclonic separator into the inhalation system.

- Method of particle size determination: Stack Sampler Mark III (Andersen).
Before sampling, to reduce rebounding effect, the impactor stages were coated with parafine oil. Thereafter, the impactor were assembled with preweighed glass fiber collecting discs, and equipped with a backup particle filter. The impactor was connected to the vacuum pump and two samples were taken from the breathing zone of the animals sampling occurred 30 minutes (or later) after the beginning of the exposure.
The sample volume for each sample was 6 L.

- Treatment of exhaust air: The exhaust air was filtered and conducted into the exhaust air of the building.
- Temperature, humidity, pressure in air chamber: 21.4 °C, 25.2% humidity. Lower amount of exhaust air, which was adjusted by means of a separate exhaust air system, achieved a positive pressure inside the exposure system. This ensured that the mixture of test substance and air was not diluted with laboratory air in the breathing zones of the animals

TEST ATMOSPHERE
- Brief description of analytical method used: Preweighed filters were placed into the filtration equipment (MN 85/90 BF (d = 47 mm)). By means of the vacuum pump metered volumes of the dust were drawn through the filter. For each sample the dust concentration in mg/L was calculated from the difference between the preweight of the filter and the weight of the filter after sampling, with reference to the sample volume of the inhalation atmospheres. Mean and standard deviation were calculated for the concentration from the results of the individual measurements. The concentration was corrected for the amount of additive used. Samples were taken every hour from a total sample volume of 6L air.

- Samples taken from breathing zone: yes

VEHICLE
- Composition of vehicle (if applicable): To improve the flowability, 1 % AEROSIL® 200 was added to the test substance. In addition 1% AEROSIL® R 972 was added as an antihydroscopic agent.


TEST ATMOSPHERE
- Particle size distribution: See figure

Sample MMAD (µm) Geometrical standard deviation
1 4.4 3.3
2 2.8 2.3


CLASS METHOD (if applicable)
- Rationale for the selection of the starting concentration: The limit dose was chosen because the substance was found to be non-toxic in all available toxicity studies.
Analytical verification of test atmosphere concentrations:
yes
Remarks:
Gravimetric measurement
Duration of exposure:
4 h
Concentrations:
4.651 mg/L +/- 0.581 (measured; >5 mg/L nominal)
No. of animals per sex per dose:
5
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Individual body weights once during the acclimatization period, shortly before exposure (day 0) and at least on days 1, 3 and 7, and before the sacrifice of the animals at the end of the observation period. Detailed clinical observations were recorded for each animal separately several times during exposure and at least once daily on the pre-exposure day and during the observation period.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight
Statistics:
not required

Results and discussion

Effect levelsopen allclose all
Sex:
male/female
Dose descriptor:
LC0
Effect level:
>= 4.651 mg/L air (analytical)
Based on:
test mat.
Exp. duration:
4 h
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 4.651 mg/L air (analytical)
Based on:
test mat.
Exp. duration:
4 h
Mortality:
none
Clinical signs:
other: See table 1
Body weight:
The mean body weights of the animals decreased during the first post exposure observation day and increased from study day 3 onward.
Gross pathology:
No adverse findings were observed.

Any other information on results incl. tables

Table 1: Clinical signs

Test group (4.651 mg/L)

Male animals

Female animals

Fur, yellow discolored

d6 - d14

d5 – d14

Fur, substance contaminated

d0 – d11

d0 – d4

Respiration, intermittent

d0 – d1

d0 – d4

Respiration, labored

h2 – h4

h2 – h4

Piloerection

d0 – d7

d0 – d5

Table 2: Concentration in the test chamber

time point of sampling measured concentration
1h 4.06
2h 4.246
3h 5.101
4h 5.196
Mean 4.6508
Mean (rounded) 4.651
Standard deviation of the mean 0.581

Applicant's summary and conclusion

Interpretation of results:
practically nontoxic
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
No mortality was observed in rats exposed to a dust at a measured concentration of 4.651 mg/L for 4hours.