Registration Dossier

Diss Factsheets

Administrative data

Endpoint:
acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
19-03-2012 to 16-04- 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study performed under GLP. All relevant validity criteria were met; with some generally acceptable deviations in accordance with the specifications of the guidelines.
Justification for type of information:
Information as to the availability of the in vivo study is provided in 'attached justification'.

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 436 (Acute Inhalation Toxicity: Acute Toxic Class Method)
Deviations:
yes
Remarks:
It was not possible to generate a MMAD < 4 µm atmosphere and maintaining the concentration at 2 mg/L despite attempts to generate such an atmosphere. Therefore expert judgement was used to apply study to classification and lablleing
GLP compliance:
yes (incl. QA statement)
Remarks:
inspected: July 2011; signature: August 2011
Test type:
acute toxic class method
Limit test:
yes

Test material

Constituent 1
Chemical structure
Reference substance name:
2-methoxy-4-methylphenyl methyl carbonate
EC Number:
700-673-7
Cas Number:
132638-45-0
Molecular formula:
C10H12O4
IUPAC Name:
2-methoxy-4-methylphenyl methyl carbonate
Test material form:
solid
Details on test material:
- Physical state: Extremely pale yellow crystalline solid
- Storage condition of test material: Approximately 4°C in the dark, under nitrogen

Test animals

Species:
rat
Strain:
Wistar
Remarks:
RccHan: WIST
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Recognised supplier
- Age at study initiation: 8 to 12 weeks
- Weight at study initiation: 200 to 250g
- Fasting period before study: yes
- Housing: groups of up to 5 by sex in solid-floor polypropylene cages with stainless steel lids, furnished with softwood flakes and cage enrichment (wooden chew blocks and cardboard fun tunnels).
- Diet (ad libitum): 2014C Harlan Rodent Diet from recognised supplier, provided ad libitum (except for exposure period period)
- Water (ad libitum): mains drinking water, ad libitum (except for exposure period period)
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 to 25
- Humidity (%): 30 to 70
- Air changes (per hr): at least 15
- Photoperiod (hrs dark / hrs light): 12 h light / 12 h dark

IN-LIFE DATES: From: 19-03-2012 To: 16-04-2012

Administration / exposure

Route of administration:
inhalation: dust
Type of inhalation exposure:
nose only
Vehicle:
clean air
Mass median aerodynamic diameter (MMAD):
5.56 µm
Geometric standard deviation (GSD):
3.07
Remark on MMAD/GSD:
MMAD/GSD relates to: 47.6 mg/L (nominal), 3.52 mg/L (maximum mean attainable concentration) dose
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Dust atmosphere was produced from the test item using a SAG 410 Solid Aerosol Generator located adjacent to the exposure chamber. The generator was connected to a metered compressed air supply
- Exposure chamber volume: approximately 30 litres (dimensions: 28 cm diameter x 50 cm high)
- Method of holding animals in test chamber: Prior to the day of exposure each rat was acclimatized (for approximately 2 hours) to a tapered polycarbonate restraining tube. During the exposure period, each rat was individually held in a tapered, polycarbonate restraining tube fitted onto a single tier of the exposure chamber and sealed by means of a rubber ‘O’ ring. Only the nose of each animal was exposed to the test atmosphere.
- Source and rate of air: filtered air; chamber flow rate was maintained at 60 L/min providing 120 air changes per hour.
- Method of conditioning air: Compressed air was supplied by means of an oil free compressor and passed through a water trap and respiratory quality filters before it was introduced to the Solid Aerosol Generator.
- System of generating particulates/aerosols: The concentration within the chamber was controlled by adjusting the air flow settings and test item feed rate from the SAG 410. The chamber flow rate was maintained at 60 L/min providing 120 air changes per hour.
- Method of particle size determination: Particle size was determined using a cascade impactor. The device consisted of six impactors stages. The mean amount for each stage was used to determine the cumulative amount below each cut-off point size to calculate the proportional (%) aerosol of defined size ranges. The resulting values were converted to probits and plotted against Log10 cut-point size. From this plot, the Mass Median Aerodynamic Diameter (MMAD) was determined (as the 50% point) and the geometric standard deviation was calculated. In addition the proportion (%) of aerosol less than 4 μm (considered to be the inhalable fraction) was determined.
- Treatment of exhaust air: The extract from the exposure chamber passed through a ‘scrubber’ trap and was connected with a high efficiency filter to a metered exhaust system. The chamber was maintained under negative pressure. A schematic of the dynamic (continuous flow) system is presented in the full study report.
- Temperature, humidity, in air chamber: The temperature and relative humidity inside the exposure chamber were measured by an electronic thermometer/humidity meter: 19-20°C, 50-51% humidity.

TEST ATMOSPHERE
- Brief description of analytical method used: The test atmosphere was sampled seventeen times during the exposure period. The actual chamber concentration was measured at regular intervals during the exposure period. The Gravimetric Method used glass fibre filters placed in a filter holder. The holder was temporarily sealed in a vacant port in the exposure chamber in the animals’ breathing zone and a suitable, known volume of exposure chamber air was drawn through the filter using a vacuum pump. Each filter was weighed before and after sampling in order to calculate the weight of collected test item. The difference in the two weights, divided by the volume of atmosphere sampled, gave the actual chamber concentration. The nominal chamber concentration was calculated by dividing the mass of test item used by the total volume of air passed through the chamber. The nominal concentration is 1353% of the actual mean achieved atmosphere concentration and shows that keeping the aerosol airborne was extremely difficult.
Full details of the analytical method are provided in the full study report.
- Samples taken from breathing zone: yes

TEST ATMOSPHERE (if not tabulated)
- Particle size distribution: The particle size of the generated atmosphere inside the exposure chamber was determined three times during each exposure period using a cascade impactor. The particle size distribution for each group is reported in table 1.
- MMAD (Mass median aerodynamic diameter) / GSD (Geometric st. dev.): The Mass Median Aerodynamic Diameter (MMAD) was determined and is reported for each group in table 1.

CLASS METHOD (if applicable)
- Rationale for the selection of the starting concentration: maximum achievable
Analytical verification of test atmosphere concentrations:
no
Duration of exposure:
4 h
Concentrations:
3.52 mg/L
No. of animals per sex per dose:
3 per sex per dose. Full details are provided in table 2.
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Clinical signs at hourly intervals during exposure, immediately on removal from the restraining tubes at the end of exposure, one hour after termination of exposure and subsequently once daily for up to fourteen days. Any evidence of mortality or overt toxicity was recorded at each observation. Individual body weights were recorded on arrival, prior to treatment on the day of exposure and on Days 1, 3, 7 and 14 or after mortality.
- Necropsy of survivors performed: yes (and in the event of any mortalities)
- Other examinations performed: clinical signs, body weight, organ weights, and any other relevant toxicological effects were reported.

Results and discussion

Effect levels
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 3.52 mg/L air
Based on:
test mat.
Exp. duration:
4 h
Mortality:
No mortality.
Clinical signs:
other: Hunched posture, piloerection and red/brown staining around eyes or snout and wet fur noted when animals removed from restraint tubes and considered associated with restraint procedure. Increased respiratory rate noted during and 1 hour after exposure. Al
Body weight:
Slight loss of body weight noted for 3 males and 1 female on day after exposure. Slight body weight loss also noted for 1 female between Days 1 and 3. All males and females gained weight during the study.
Gross pathology:
Dark patches on lungs noted for 1/3 males and 1/3 females.

Any other information on results incl. tables

Additional comments regarding test atmosphere generation:

It was noted that four samples were outside 20% of the mean achieved atmosphere concentration (two low and two high). The test item was generated at the maximum speed that the generation system could run at in order to achieve the maximum attainable atmosphere concentration during the exposure. The nominal concentration (47.6 mg/L) also shows that keeping the aerosol airborne was extremely difficult. These deviations were therefore considered to be unavoidable.

 

It is also noted that the particle size distribution and the GSD that were achieved during this study were outside of the ranges specified in the test guidelines (1 - 4μm and 1.5-3.0). It was, preferable to expose the animals to a concentration of test item as close to 5 mg/L as possible, even though this may have increased the MMAD as this was considered to result in the animals being exposed to the highest possible concentration of particles < 4μm. During the formal exposure the percentage of particles < 4μm was found to be 38.4%, which means that approximately 1.35 mg/L of test atmosphere could be considered to be in the inhalable range of the test animals (< 4μm). In order to achieve this concentration of particles < 4 μm at 2 mg/L the particle size achieved would have needed to be approximately to 2.5 μm. Even with substantial grinding this would have been impossible to achieve with this particular test item. Even when reducing the achieved concentration to below 2 mg/L during the characterisation phase of the study and by adding particle selection devices into the generation system a particle size <4 μm could not be achieved.

 

Further testing was not considered appropriate given the results and the nature of the test item, within the study.

 

Table 1. Characteristics of the achieved atmosphere:

Mean Maximum Attainable Concentration (mg/L)

Mean Mass Median Aerodynamic Diameter (μm)

Inhalable Fraction

(% <4 μm)

Geometric Standard Deviation

3.52

5.56

38.4

3.07

 

 

 

 

 

Table 2. Achieved atmosphere concentrations:

Atmosphere Concentration

Mean Maximum Attainable (mg/L)

Standard Deviation

Nominal (mg/L)

3.52

0.54

47.6

 

 

 

 

Table 3. Mortality data summary:

Mean Maximum Attainable Concentration (mg/L)

Male Mortalities

Female Mortalities

Total Mortalities

3.52

0/3

0/3

0/6

 

 

 

 

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Remarks:
Criteria used for interpretation of results: EU
Conclusions:
Under the conditions of this study, the mean maximum attainable atmosphere concentration over 4 hours was 3.52 mg/L. The inhalation 4h-LC50 (male/female) was considered to be > 3.52 mg/L within the Wistar (RCCHan: WIST) strain rat.
Executive summary:

The study was performed according to OECD TG 436 guideline in accordance with GLP to assess the acute inhalation toxicity of the test item. A single group of six Wistar (RccHan : WIST) strain rats (three males and three females) were exposed to an dust atmosphere of the test item. The groups were exposed for four hours using a nose only exposure system, followed by a fourteen day observation period. The mean maximum attainable atmosphere concentration was follows: 3.52 mg/L based on a nominal concentration of 74.6 mg/L. The characteristics of the achieved atmosphere where Mean Mass Median Diameter (particle size) and Inhalable Fraction < 4 μm were: 5.56 μm and 38.4% with geometric Standard Deviation 3.07. There was no male and female mortalities in the 3.52 mg/L maximum attainable atmosphere concentration. Common abnormalities noted during the study included increased respiratory rate, hunched posture, pilo-erection, red/brown staining around the eyes or snout and wet fur. All males and females recovered to appear normal from Days 3 to 5 post-exposure. All males and one female exhibited slight bodyweight losses on the first day post-exposure. Reasonable bodyweight development was noted for all males and females during the remainder of the recovery period with the exception of one female animal which exhibited a slight bodyweight loss from Days 1 to 3 post-exposure. All males and females gained weight during the study. Dark patches on the lungs were detected amongst one female and one male at necropsy. All other findings were normal. No mortality occurred and there was no indications of significant toxicity in a group of six rats exposed to a mean maximum attainable atmosphere concentration of 3.52 mg/L for four hours. Under the conditions of this study, the acute inhalation median lethal concentration 4 hr-LC50 was > 3.52 mg/L in the male/female RccHanTM : WIST strain rat.

Categories Display