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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:
July - Aug 1993
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
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
GLP compliance:
yes
Test type:
traditional method
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
4,4'-methylenedicyclohexyl diisocyanate
EC Number:
225-863-2
EC Name:
4,4'-methylenedicyclohexyl diisocyanate
Cas Number:
5124-30-1
Molecular formula:
C15H22N2O2
IUPAC Name:
1,1'-methylenebis(4-isocyanatocyclohexane)
Details on test material:
- Purity: > 99.2 %

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Strain: Hsd/Win: WU (SPF)
- Source: Harlan-Winkelmann, Borchen, Germany
- Age at study initiation: 2-3 months
- Weight at study initiation: mean 170-200 g
- Housing: singly in Makrolon Type II cages (according to Spiegel & Goennert, Zschr. Versuchstierkunde 1, 38, 1961 and Meister, Zschr. Versuchstierkunde 7, 144-153, 1965)
- Diet and Water: ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 2
- Humidity (%): approx. 50 %
- Air changes (per hr): approx. 10
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
inhalation: aerosol
Type of inhalation exposure:
nose/head only
Vehicle:
air
Mass median aerodynamic diameter (MMAD):
>= 1.31 - <= 1.53 µm
Geometric standard deviation (GSD):
>= 1.71 - <= 1.77
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Mode of exposure: Animals were head-nose exposed to the aerosolized test article in restrainers made of Plexiglas.
- Generation of aerosol: The test substance was nebulized neat under dynamic conditions in a cylindric inhalation chamber. In order to increase the efficiency of the generation of respirable particles and to prevent larger particles from entering the chamber a preseparator/ baffle system was used. The inhalation chamber had the following dimensions: inner diameter = 14 cm, outer diameter = 35 cm (two-chamber system), height = 50 cm (internal volume = about 7.6 L). Details of this modular chamber and its validation have been published previously (Pauluhn, Journal of Applied Toxicology, 14, 55-62, 1994).
- Generation of atmosphere: Atmospheres were generated under dynamic conditions using a Braun Infusion pump or a Hamilton Microlab pump and a binary nozzle.
- Conditioning the compressed air: Compressed air was supplied by Boge compressors and was conditioned (i.e. freed from water, dust, and oil) automatically by a VIA compressed air dryer. Adequate control devices were employed to control supply pressure.
- Inhalation chamber equilibrium concentration: The test atmosphere generation conditions provide an adequate number of air exchanges per hour (approx. 237). Under such test conditions used chamber equilibrium is attained in less than one minute of exposure (t95% = 3 x chamber volume/chamber airflow). The ratio between the air supplied and exhausted was chosen so that approximately 90% of the supplied air is removed via the exhaust system.
- Exhaust air treatment: The exhaust air was purified via filter systems.
- Temperature and humidity measurements were performed by the computerized Data Acquisition and Control System using FTF11-sensors (Fa. Elka Electronic, Lüdenscheid).

TEST ATMOSPHERE
- The integrity end stability of the aerosol generation and exposure system was measured by using a RAS-2 real-time aerosol photometer (MIE, Bedford, Massachusetts, USA).
- Samples taken from breathing zone: yes
- Brief description of analytical method used: HPLC and additionally gravimetric analysis of filter samples (filter: Glass-Fibre-Filter, Sartorius, Göttingen, Germany; digital balance).
HPLC-method: Nitro-reagent-treated glass fibres were exposed to the test atmosphere. The content of isocyanate was analytically detected via HPLC.
- Particle size distribution: The particle-size distribution was analyzed using a BERNER critical orifice cascade impactor. 90% of the particle mass had an aerodynamic diameter - MMAD (Mass median aerodynamic diameter): The respirability of the aerosol was adequate, i.e. the mass median aerodynamic diameter (MMAD) was ~ 1.4 µm; geometric standard deviation (GSD) ~ 1.8.
Analytical verification of test atmosphere concentrations:
yes
Duration of exposure:
4 h
Concentrations:
151, 388, 418, 552, 730, 865, and 1352 mg/m³ (gravimetric conc.)

No. of animals per sex per dose:
5
Control animals:
yes
Remarks:
air control
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations: Several times on day of exposure, twice on postexposure days, also on weekends.
- Frequency of weighing: Before exposure, on day of exposure, and on days 3, 7 and 14 post-exposure
- Necropsy of survivors performed: yes
- Other examinations performed: rectal temperature, reflexes
Statistics:
Calculation of LC50 was according to A.P. Rosiello et al., J. Tox. and Environ. Health 3, 797-809 (1977) and modified by J. Pauluhn; Bayer Report No. 11845 (1983)

Results and discussion

Effect levelsopen allclose all
Key result
Sex:
male/female
Dose descriptor:
LC50
Effect level:
434 mg/m³ air
95% CL:
>= 355 - <= 533
Exp. duration:
4 h
Sex:
male
Dose descriptor:
LC50
Effect level:
456 mg/m³ air
95% CL:
>= 313 - <= 667
Exp. duration:
4 h
Sex:
female
Dose descriptor:
LC50
Effect level:
431 mg/m³ air
95% CL:
>= 363 - <= 513
Exp. duration:
4 h
Mortality:
Mortalities occurred from 388 mg/m³ onwards from exposure day through post exposure day 4.
Number of deaths at each dose (time of death):
male rats: 0/5 at 151 mg/m³, 2/5 at 388 mg/m³ (1d-2d), 2/5 at 418 mg/m³ (1d-2d), 5/5 at 552 mg/m³ (1d-4d), 4/5 at 730 mg/m³ (1d-3d), 2/5 at 865 mg/m³ (1d-2d), 5/5 at 1.352 mg/m³ (0d-1d)
female rats: 0/5 at 151 mg/m³, 2/5 at 388 mg/m³ (1d), 2/5 at 418 mg/m³ (1d), 4/5 at 552 mg/m³ (1d-2d), 5/5 at 730 mg/m³ (1d), 5/5 at 865 mg/m³ (1d-3d), 5/5 at 1.352 mg/m³ (0d-1d)
Clinical signs:
other: see section "Other findings"
Body weight:
A significant and concentration dependent effect on body weight was observed in the post exposure period at and above 151 mg/m³.
Gross pathology:
Rats that died during the observation period: Hydrothorax, lung oedematous, trachea filled with foamy content, lung does not collapse after opening of thorax, lung reddish up to hepatoid appearance, liver with lobular pattern and pale, duodenumcontent mucous-reddish, spleen and kidneys pale.
Rats that were sacrificed at termination of the study no indications for specific concentration dependent effects on organs were found.
Other findings:
- Other observations:
Rectal temperature: a significant and concentration dependent decrease was seen.
Reflex examination: no indications for specific neurological changes were found.

Clinical signs were observed for all test substance treated group. Signs occurred during exposure and up to 12 days thereafter for males and 9 days thereafter for females and consisted of bradypnea, dyspnea, laboured breathing, rales, irregular breathing pattern, ungroomed coat, piloerection, motility reduced, tremor, hunched posture, highlegged gait, staggered gait, sluggish, cyanosis, vocalization, emaciation, serous discharge from nose, reddish rhinarium, wheezing, nose with red encrustations, nose-muzzle/red encrustations, and prostration (lying on belly).
At the end of the observation period all surviving animals were without symptoms.

Any other information on results incl. tables

Data from IUCLID4

RS-Freetext:
VALUE:
LC50 (male): 456 mg/m3 (95% confidence interval of 313 to 667 mg/m3)
LC50 (female): 431 mg/m3 (95% confidence interval of 363 to 513 mg/m3)
LC50 (male/female): 434 mg/m3 (95% confidence interval of 355 to 533  mg/m3)
MORTALITY:
Number of deaths at each dose (time of death), male rats: 151 mg/m3: 0/5;  388 mg/m3: 2/5 (1d-2d); 418 mg/m3: 2/5 (1d-2d); 552 mg/m3: 5/5 (1d-4d);  730 mg/m3: 4/5 (1d-3d); 865 mg/m3: 2/5 (1d-2d); 1,352 mg/m3: 5/5 (0d-1d)
Number of deaths at each dose (time of death), female rats: 151 mg/m3:  0/5; 388 mg/m3: 2/5 (1d); 418 mg/m3: 2/5 (1d); 552 mg/m3: 4/5 (1d-2d);  730 mg/m3: 5/5 (1d); 865 mg/m3: 5/5 (1d-3d); 1,352 mg/m3: 5/5 (0d-1d)
CLINICAL SIGNS:
Number of animals with clinical signs, male rats: >= 151 mg/m3: 5/5 (due  to early death at 1,352 mg/m3 clinical signs were only for 4/5 animals  recorded) showed during the exposure and up to 12d after the exposure  bradypnea, dyspnea, laboured breathing, rales, irregular breathing  pattern, ungroomed coat, piloerection, motility reduced, tremor, hunched  posture, highlegged gait, staggered gait, sluggish, cyanosis,  vocalization, emaciation, serous discharge from nose, reddish rhinarium,  wheezing, nose with red encrustations, nose-Muzzle/red encrustations,  prostration (lying on belly); at the end of the post observation period  all surviving animals were without symptoms
Number of animals with clinical signs, female rats: >= 151 mg/m3: 5/5  (due to early death at 1,352 mg/m3 clinical signs were only for 4/5  animals recorded) showed during the exposure and up to 9d after the  exposure bradypnea, dyspnea, laboured breathing, rales, irregular  breathing pattern, ungroomed coat, piloerection, motiltiy reduced,  tremor, hunched posture, highlegged gait, staggered gait, sluggish,  cyanosis, vocalization, emaciation, serous discharge from nose, reddish  rhinarium, wheezing, nose with red encrustations, nose-Muzzle/red  encrustations, prostration (lying on belly); at the end of the post  observation period all surviving animals were without symptoms
BODY WEIGHT:
>= 151 mg/m3: body weight retardation
RECTAL TEMPERATURE:
concentration dependent decrease
REFLEX EXAMINATION:
no substance induced effects
NECROPSY:
rats died during the post observation period:
pale liver, spleen and kidneys, lobular pattern of the liver, duodenum  was filled with a red mucous mass, trachea was filled with a mucous mass,  less collapsed lungs with red foci, pulmonary edema and hydrothorax;  lungs were red up to hepatoid changes, red secretions in the nose 
rats sacrificed at the end of the post observation period:
>= 151 mg/m3 no specific concentration dependent organ changes

Applicant's summary and conclusion

Executive summary:

4,4´-Methylenedicyclohexyl diisocyanate has been assessed in the OECD HPV programme, 2005.

Partly cited from SIAR of SIAM 20 (Paris, April 19 -22, 2005):

An acute inhalation toxicity study of 4,4´-methylenedicyclohexyl diisocyanate (purity > 99.2 %) was performed "by exposing Wistar rats in seven groups, each containing 5 males and 5 females. The method used was essentially that of OECD TG 403. Each group was nose only exposed to concentrations of the aerosol of the test substance. After exposure (4 hours) the animals were observed for two weeks. The actual mean concentrations of 4,4´-methylenedicyclohexyl diisocyanate were 151, 388, 418, 552, 730, 865 and 1352 mg/m³. The test substance aerosol exhibited a particle-size indicating that this aerosol was of adequate respirability (90 % of the particle mass was ≤ 3 μm; mass median aerodynamic diameter (MMAD) ≈ 1.4 μm; geometric standard deviation (GSD) ≈ 1.8 μm). Rats exposed to ≥ 151 mg/m³ experienced signs of respiratory tract distress (i.e. salivation, bradypnea, stridor). Exposure to concentrations of ≥ 388 mg/m³ caused mortality, which occurred on the exposure day through post exposure day 4. Gross necropsy findings (less collapsed lungs including a hemorrhagic lung edema, red secretions in the nose, pale liver with lobular pattern, pale spleen and kidneys) were observed only in the rats which died during the observation period. The LC50 (4 h) stated in this study is 434 mg/m³ with confidence limits (95 %) of 355 - 533 mg/m³ both sexes combined."