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Toxicological information

Acute Toxicity: inhalation

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Administrative data

acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference Type:
study report
Report date:

Materials and methods

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

Test material

Test animals

Details on test animals or test system and environmental conditions:
- Source: Charles River (UK) Ltd., Margate
- Age at study initiation: 6 to 8 weeks old
- Weight at study initiation: 140 - 200 g
- Fasting period before study: no data
- Housing: groups of 5 animals by sex in grid-floor stainless steel cages
- Diet (e.g. ad libitum): ad libitum; SQC Rat and Mouse Maintenance Diet No. 1 Expanded, Special Diets Services Ltd. Witham
- Water (e.g. ad libitum): ad libitum; mains water
- Acclimation period: 25 days

- Temperature (°C):19 - 25 °C
- Humidity (%): 40 - 70 %
- Air changes (per hr): 15 air changes per hour
- Photoperiod (hrs dark / hrs light): 12 hours light and 12 hours darkness

Administration / exposure

Route of administration:
inhalation: aerosol
Type of inhalation exposure:
head only
clean air
Details on inhalation exposure:
- Exposure apparatus: cylindrical anodised aluminium exposure chamber
- Exposure chamber volume: 30 L
- Method of holding animals in test chamber: head-only
- Source and rate of air: The compressed air supply used was from a clean dry filtered source.
- Method of conditioning air: -
- System of generating particulates/aerosols: DeVilbiss model 646 liquid nebulizer generator, dynamic (continuous flow)
- Method of particle size determination: The particle size was determined using a Delron C55 Cascade Impactor, with 5 separation
stages corresponding to maximum mass median aerodynamic diameters of 0.5, 1.0. 2.0. 4.0 and 8 µm. The samples were obtained
hourly, over periods of up to 8 minutes, -during the exposure period.
- Treatment of exhaust air: The atmospheres were filtered, exhausted to the outside of the building and vented.
- Temperature, humidity, pressure in air chamber: The temperature and relative humidity inside the exposure
chamber were measured continuously and recorded at hourly interva1s throughout th'e 4-hour exposure peri od, using a
digital thermometer with remote probe located in the chamber and a paper hygrometer located in the exhaust duct of the chamber.

- Brief description of analytical method used: The concentration of the test article was determined gravimetrically. The atmosphere
was sampled by drawing a known volume through an open face glass fibre filter positioned at a site representative of that
occupied by the external nares of the experimental animals.
- Samples taken from breathing zone: yes

TEST ATMOSPHERE (if not tabulated)
- Particle size distribution:
- MMAD (Mass median aerodynamic diameter) / GSD (Geometric st. dev.):

Analytical verification of test atmosphere concentrations:
Duration of exposure:
4 h
0.513, 2.118, 4.585 and 6.518 mg/L
No. of animals per sex per dose:
5 aminals per sex per group
Control animals:
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: twice daily for dead or moribund. The body weight of each animal was recorded immediately before and
after exposure, on days 8 and 15 of the study and at necropsy.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight,organ weights, histopathology, other:
Clinical signs were observed at hourly intervals during the exposure period, then for the remainder of the working day, and once daily thereafter for
14 days.
Necropsy: A full internal and external examination was made under the general supervision of a pathologist. The nasal passages were examined and
an assessment made of any irritation of the respiratory tract.
Organ weights: The lungs, bronchi and trachea were dissected free from fat and other contiguous tissue and weighed together.
Histology: Samples of all gross lesions were fixed in 10% neutral buffered formalin and retained without further processing.
The median lethal concentrations (LC50) together·with 95% fiducial limits were calculated separately for males and females, and for
sexes combined, from the recorded mortality rate using a probit analysis method (Finney, D.J. (971)
Probit Analysis, 3rd ed., Cambridge University Press).

Results and discussion

Effect levelsopen allclose all
Dose descriptor:
Effect level:
3.181 mg/L air (analytical)
Based on:
test mat.
95% CL:
0.567 - 50.411
Exp. duration:
4 h
Dose descriptor:
Effect level:
7.655 mg/L air (analytical)
Based on:
test mat.
Exp. duration:
4 h
Key result
Dose descriptor:
Effect level:
5.319 mg/L air (analytical)
Based on:
test mat.
95% CL:
2.981 - 34.296
Exp. duration:
4 h
Qveral-l there was a positive correlation between mortality and chamber concentration. As the minimum level at which death
occurred was 4.585 mg/l for males and 2.118 mg/l for females, the possibility of a sex difference cannot be eliminated.
One male in group 5 died during exposure. Except for one death on day four all deaths occurred on days one or two of the study.
Seven animals were killed for humane reasons on day one. Five of these decedents were in moribund condition.
Clinical signs:
other: Marked signs of toxicity were observed at 4.585 and 6.518 mg/L immediately after exposure. The signs included ataxia, occasionally prostration and irregular respiration. and convulsion and signs of ejaculation at 4.585 mg/l. During and/or following expos
Body weight:
Body weight losses occurred as a result of the r~straint procedures in test and control groups. The losses tended to be greater in
treated groups and persisted in a few individuals until day 8 of the study. However, overall there was no marked effect on body
Gross pathology:
Mean absolute and relative lung weights at termination for treated groups tended to be slightly higher than the corresponding control va1ues. but all values fell withi n the normal range and the numerical differences were too small to conclude there was a treatment-related effect.
Occasional lung weights for decedents were near or ablove the upper limits of the normal range.
This was probably due to agonal congestive changes.
Animals surviving to termination showed only incidental changes. Some decedents were unremarkable but others had discoloured and
inflated lungs suggestive of acute effects on the cardiopulmonary system. Four of the flve group 3 females had sore forefeet
possible due to self-inflicted trauma.

Applicant's summary and conclusion

Interpretation of results:
Category 4 based on GHS criteria
Based on the LC50 female of 3.181 mg/L air the test substance has to be classified according to CLP, EU GHS (Regulation (EC) No. 1272/2008) in Toxicity Category IV.
Executive summary:

In an acute inhalation toxicity study performed according to OECD Guideline 403 5 young Sprague-Dawley rats per sex per group were exposed at single chamber concentrations of 0.513, 2.118, 4.585 and 6.518 mg/L of Guanidine hydrochloride (98.5 % a.i.) by inhalation (head-only) over a period of 4 hours. Corresponding nominal concentrations were in the range 2.124 to 15.280 mg/L. The mass median aerodynamic diameters of the particles in the atmospheres fell in the range 1.88 to 5.62 µm. The aerosol for animal exposures was prepared from an aqueous solution of the test article. A similar group of 5 males and 5 females was exposed to filtered air as a control. Animals then were observed for 14 days.

The acute inhalation median lethal concentrations, calculated by a probit method, were:

LC50 combined: 5.319 mg/L

LC50 Males: 7.655 mg/L

LC50 Females: 3.181 mg/L

Deaths occurred in males at a level of 4.585 mg/l and above, and in females at a level of 2.118 mg/l and above. Overall there was a dose-related relationship between mortality and chamber concentration. Nearly all deaths occurred on days one or two of the study. Marked clinical signs were first observed on the day of exposure. The signs included ataxia and occasionally prostration, irregular respiration, convulsion and signs of ejaculation. A few individuals showed body weight loss that persisted one week after treatment, but overall there was no marked treatment related effect on body weight.

There was no evidence of a treatment-related, effect on lung weight in survivors. Occasional increases in lung weights in decedents were probably due to agonal congestive changes. Animals surviving to termination were unremarkable macroscopically. A few decedents had discolored and inflated lungs.