Registration Dossier

Administrative data

Endpoint:
acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
10 July - 24 July, 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
comparable to guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report Date:
2017

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 436 (Acute Inhalation Toxicity: Acute Toxic Class Method)
GLP compliance:
yes
Test type:
acute toxic class method
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Details on test material:
White solid

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
Species: Rat
Strain: Crl: Sprague Dawley (SD)
Source: Beijing Vital River Laboratory Animal Technology Co., Ltd.
Number of Animals: 18 animals (9 males and 9 females) were ordered and 6 of them were used. The females were nulliparous and non-pregnant.
Age: The age was between 69-75 days
Weight: between 336-370 g for male rats and 233-249 g for female rats at the commencement of dosing

Justification for Test System:
According to the Guidelines for the testing of chemicals "Acute Inhalation Toxicity" (TG 436) published by the Ministry of Environmental Protection of People's Republic of China in the year of 2013, rat is the preferred strain for the heredity characters, stability and available background data.

Physical Examination and Acclimation:
A physical examination, weighing and marking on the hair and cage card identifying was made within 24 hours after animals' arrival. After the physical examination a 19-day acclimatization period started. Animals were acclimated to the restraining tubes twice prior to dosing in order to minimize stress
and uncomfortableness about restraining tubes. First pre-adaption was about 1 h. Second pre-adaption was about 2 h. No abnormalities were found during both restraining.

Husbandry
Housing:
Animals were housed in a room in suspended, stainless steel cages. Animals were housed individually during the test.

Environmental Controls:
The temperature and humidity were automatically controlled and recorded. The animal room temperature was 19-25°C, the relative humidity was 40%-70% and light cycle was 12 hour light and 12 hour dark.

Food and Water:
Animals were provided with rodent complete nutrition pellet diet. During the test, diet and water were available to the animals ad libitum except
restraining periods.

Animal Welfare:
The animal use for this study complies with the national animal welfare laws and regulations. The animal care and use activities required to conduct the study were reviewed and approved by the testing facility Animal Care and Use Committee. 10 of the spare animals were used for another test
and 2 of them were euthanized by C02.

Administration / exposure

Route of administration:
inhalation: aerosol
Type of inhalation exposure:
snout only
Vehicle:
other: Fumed silica
Mass median aerodynamic diameter (MMAD):
3.33 µm
Geometric standard deviation (GSD):
1.93
Remark on MMAD/GSD:
Actual mean concentration in the cabinet for four-time measurements was 5064 ± 59 mg/m^3. The average mass median aerodynamic diameter (MMAD) and geometric standard deviation (GSD) for two-time measurements were 3.33 µm and 1.93, respectively. The average aerodynamic particle size less than 4 micron (mass%) for a two-time measurement was 62.37%.
Details on inhalation exposure:
Before exposure, each rat was confined to a transparent polycrylic tube. The exposure tubes were installed in the portholes of the inhalation chamber and
the chamber was sealed up. Filtered and compressed air (0.04 m^3) was mixed with quantitative test item and aerosol was sent to exposure chamber. The moving speed and exposure airflow rate was adjusted. The aerosol was continuously generated from generation system on the top of the chamber with an aerosol producer. The exhausted air was removed from the outlet at the bottom of the chamber to the absorption unit.

Concentration Trial: Before exposure, a test item trial was conducted (without animals) using the inhalation system. After two successive concentrations' error fell within ±20% of target concentration, exposure was conducted.
Analytical verification of test atmosphere concentrations:
not specified
Duration of exposure:
4 h
Remarks on duration:
Rats were exposed to target concentration for 4 hours.
Concentrations:
5,000 mg/m^3
No. of animals per sex per dose:
3/sex/dose
Control animals:
no
Details on study design:
IN-LIFE PROCEDURES, OBSERVATIONS, AND MEASUREMENTS

Exposure Conditions:
The actual concentrations, particle size distribution at the animals' breathing zone, chamber airflow, chamber temperature, relative humidity and oxygen concentration were determined during exposure period.

Actual Concentration:
Actual concentration at the animals' breathing zone was determined using filter gravimetric analysis. Samples were collected by fiber filter fixed in filter holder attached to ESA-3Z auto-sampling device, on which airflow (1 L/min), time (5 min) and volume (5 L) were set. The mass collected filter paper was weighed before (Ml) and after collection (M2). The collected mass (M) = M2-Ml. Actual concentration (C, mg/m^3) = the collected mass (mg)/ the collected volume (L) x 1,000 L/m^3. Frequency of determination was about 1 time per hour

Particle Size Distribution:
Aerosol Instrument for Aerodynamic Particle Sizer was used to assess the particle size distribution of the test atmosphere.
Particle size distribution parameters were collected and calculated by a software. Based on the data, the aerodynamic mass median diameter (MMAD) and and geometric standard deviation (GSD) were determined. Determination frequency was every 2 hours.

Clinical Observations:
Clinical observations were performed once during the exposure, and once after the animals were removed from the confined tubes. Observations of morbidity or moribund states were performed twice from the first day after exposure day to the end of observation period except that they were performed once on necropsy days. Observations and records were conducted including animal fur changes, eyes and mucous membranes, and also respiratory, circulatory, autonomic and central nervous systems, and somatomotor activity and behavior patterns. Attention was directed to tremors, convulsions, salivation, diarrhea, lethargy, sleep and coma. It was determined by the toxic reactions, time of onset and length of recovery.period, which was extended when considered necessary. Animals' poisoning symptoms were observed and the time of their onset, duration and disappearance were recorded.

Body Weights:
he animals were weighed on the day of exposure ( day 0, prior to exposure), and
on day 1, day 3, day 7 and day 14.

Necropsy:
All surviving animals were dissected at the end of the study after anesthetizing with ether or killed by bloodletting. Nose, eyes, pharynx, larynx, trachea and lung were examined. The necropsy included following examinations such as the external features of the carcass, external body orifices, the abdominal, thoracic and their contents of all animals, and the location, size, hardness and the color. All the changes at necropsy were recorded. Due to no abnormalities observed in target organs at necropsy, the histopathological examination was not performed.
Statistics:
Animal number, sex, bodyweight, necropsy and histopathology abnormal findings were determined. The mean and standard deviations of body weight at different times were calculated. The inhalation toxicity LCD0 range was found. According to GHS criteria for the acute inhalation toxicity the test item category was given. Because the unit of LC50 was in mg/m^3, the LC50 was divided by 1000 and was converted to mg/L units when test item classification was conducted.

Results and discussion

Effect levels
Key result
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 5 mg/L air
Exp. duration:
4 h
Mortality:
No deaths of animals were observed after exposure to the end of the test.
Body weight:
The body weights of all test animals continuously increased during the test.
Gross pathology:
No abnormalities were observed for all animals at necropsy.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
Based on the results, the acute inhalation LC50 (4 h) in rats for Difluorosulfonamide HCl is co mg/m^3 and the cut-off value of LC50 (4 h) is co mg/L, According to GHS's classification criteria of acute inhalation toxicity, the test item is classified as "Unclassified".