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Administrative data

Description of key information

The results from the acute inhalation and acute dermal toxicity studies indicated that there was no indication of toxicity. The results from the acute oral toxicity study indicated that there was a positive indication of toxicity.

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records
Reference
Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
July 5 2018 - November 03 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Version / remarks:
"Acute Oral Toxicity - Acute Toxic Class Method" (TG 423) published by the Ministry of Environmental Protection of People's Republic of China, 2013.
Deviations:
no
GLP compliance:
yes
Test type:
acute toxic class method
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
female
Details on test animals or test system and environmental conditions:
Justification of Test System: Rats are the preferred species of choice as historically used for the safety evaluations studies and are specified in the appropriate test guidelines.

Test Conditions
Husbandry: Animals were housed in Room Dl20 of the facility's barrier system. Animals were raised in suspended, stainless steel cages (132.0cm xW28.0cmxH20.0cm) on cage racks (Ll67.0cmxW70.0cmxH171.0cm). There were 10 cages per layer, and 4 layers per rack. Animals were housed individually during the test.
Environmental Controls: Temperature and humidity were controlled automatically and daily recorded. The values in the animal room were 20-25°C for temperature, and 40%-70% for humidity. The lighting sequence was 12 hours light, 12 hours dark
Food and Water: Animals .were provided sterilized diet with complete nutrition supplied by Beijing Keaoxieli Feed Cd., Ltd. Analysis reports of diet were supplied by the supplier. All the nutrition components and contaminants were within the permitted limits described in the national standard (GB14924.3-2010 and GB14924.2°2001).
Water was purified by HT-R0l000 purity system. Water analysis was conducted routinely analyzed (annually), and all parameters were within the permitted limits described in the national drinking water standard (GB5749-2006). The diet and water were considered not to contain any contaminants that could reasonably be expected to affect the purpose or integrity of the study.
Water was available to the animals ad libitum during test.
Animal Welfare: Animal use complied with national animal welfare laws and regulations (instructive notions with respect to caring for laboratory animals) (2006, Ministry of Science, P.R.C.). Animals surviving to the end of the study were anesthetized by CO2 and bled by abdominal aorta ·to death. Spare animals were humanely killed by CO2 euthanasia. Their corpse treatments would be entrusted to specialized agencies. The animal care and use activities required for conduct of this study were reviewed and approved by the testing facility Animal .. Care and Use Committee (IACUC).
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
Administration Route
According to the potential route of human contact, oral gavage was selected for administration.

Dose Design
According to the Guidelines for the testing of chemicals "Acute Oral Toxicity-Acute Toxic Class Method" (TG 423) published by the Ministry of Environmental Protection of People's Republic of China in the year of 2013, and that the acute oral LD5o in rat of the test item's analog is 2460 mg/kg, the dose level of 2000 mg/kg b.w. was selected as the starting dose from one of four fixed dose levels (5,. 50, 300, 2000 mg/kg b.w., see Figure 1 in Appendix 2), and 3 animals were used in each step. The first step dosing was 2000 mg/kg b.w. and all animals died, so 300 mg/kg b.w. was selected as the second step dosing, and l animal died. Therefore 300 mg/kg b.w. was selected as the third step dosing and no animals died.

Grouping of Animals
After the acclimatization period, all healthy animals were randomly arranged by Excel 2007, and when administered each time three animals in each group were used sequentially. Each animal, at the commencement of its dosing, its weight fell in an interval within ±20% of the mean body weight of any previously dosed animals. Animals with ID beginning from .2100 to 2102 were used for the first dosing, and 2200 to 2202 were used for the second dosing, and 2300 to 2302 were used for the third dosing.

Animal Identification
All animals were marked by the special animal markers on the hair (the colour was different from the previous ones) and numbers written on cage cards. At the end of the necropsy, animals were also marked on the tails. A cage card was prepared with details.

1.1.2 Formulation Preparation
According to the dose designation and body weights weighed before dosing, the theoretical amounts of the test item were calculated. The test item was weighed in a grinding jar. Put a small amount of vehicle into the grinding jar, and then stirred with a glass rod, and subsequently diluted the test item with vehicle to the scale mark. The prepared formulations were fully homogenized and labeled for use. Computing Formula: Concentration (mg/mL) = Dose (mg/kg)/Dosing Volume (mL/kg b.w.), Theoretical Weighed Test Item (mg)= Prepared Volume (mL) x Concentration (mg/mL).

All formulations were used within two hours of preparation and were assumed to be stable for this period unless specified otherwise by the sponsor. The concentration and homogeneity of the formulations were not determined by analysis. The residual formulated test item after dosing was returned to Test Item Preparation Department and disposed totally.

1.1.3 Administration Method
All test animals were fasted overnight prior to dosing, but water was freely available. Formulations Stirring: Dosing formulations were stirred on the magnetic stirring apparatus for �t least ·10 minutes prior to dosing and were stirred during dosing. Each dosing formulation -·was administrated to animals' stomachs using a standard gavage tube attached to a disposal syringe.
Dosing Frequency: Each animal was dosed once.
Dosing Volume: The actual dosing volume of each animal was calculated according to the fasted body weight weighed before dosing. Dosing volume was 5 mL/kg b.w..
Dosing Time: In the morning
The time interval between treatment groups was determined by the onset, duration, and severity of toxic signs. The dosing interval of next step was determined according to the actual conditions.
Doses:
Dose 1: 2000 mg/kg b.w.
Dose 2: 300 mg/kg b.w.
Does 3: 300 mg/kg b.w.
No. of animals per sex per dose:
3 females per dose
Control animals:
no
Details on study design:
Moribund or Mortality Inspection
Inspections were made twice daily, morning and afternoon, during normal working days (except that it was made once in the dosing and necropsy days), and once daily at weekends and public holidays.

Clinical Observations
Clinical observations were performed once during the first 30 minutes and at 1, 2 and 4 hours after application and then once each day for up to 14 days.
General observations were made once daily for the animals. which have not been administrated with the test item.
Careful observations and records of animal fur changes, eyes and mucosa, digestive, respiratory, circulatory, autonomic and central nervous system, particularly limb activity and behavior changes were made. Attention was directed to observations of tremor, convulsions, salivation, diarrhea, lethargy, sleep and coma.

Body Weights
Individual weights of animals were determined within 24 hours after arrival, at grouping, on Day O (day of dosing), Day 7 and Day 14. At the end of the test surviving animals were weighed. Changes in weights were calculated and recorded when survival exceeding one day.

Necropsy
Animals surviving to the end of the study were anesthetized by CO2 and bled by abdominal aorta to death. A gross necropsy was performed on all animals under test. The necropsy· included carefully· eye examinations of the abdominal, thoracic organs and their contents of all animals.'1
Key result
Sex:
female
Dose descriptor:
approximate LD50
Effect level:
> 300 - < 2 000 mg/kg bw
Based on:
test mat.
Key result
Sex:
female
Dose descriptor:
LD50 cut-off
Effect level:
500 mg/kg bw
Based on:
test mat.
Mortality:
Dose Level-The first dosing (2000 mg/kg b.w.): One animal died on the dosing day and the rest two animals died on the day after dosing day.
Dose Level-The second dosing (300 mg/kg b.w.): One animal died on the second day after dosing day and the rest two animals showed no deaths or moribund status during the test.
Dose Level-The third dosing (300 mg/kg b.w.): All animals showed no deaths or moribund status during the test.
Mortality summary results were given in Appendix 1-Table 1.
Clinical signs:
Dose Level-The first dosing (2000 mg/kg b.w.): One animal showed less movement at 1 hour observation and prostration at 2 hour observation after dosing. The rest 2 animals showed prostration at 2 and 4 hour observations after dosing.
Dose Level-The second dosing (300 mg/kg b.w.): All animals showed no abnormal. symptoms during the course of the test.
Dose Level-The third dosing (300 mg/kg b.w.): All animals showed no abnormal. symptoms during the course of the test.
Body weight:
The results indicated that all surviving animals gained body weights during the test.
Gross pathology:
Not specified
Interpretation of results:
Category 4 based on GHS criteria
Conclusions:
According to the GHS's classification criteria for acute oral toxicity the test item was classified as "Category 4".
Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LD50
Value:
500 mg/kg bw

Acute toxicity: via inhalation route

Link to relevant study records
Reference
Endpoint:
acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
July 05, 2018 to November 03, 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 436 (Acute Inhalation Toxicity: Acute Toxic Class Method)
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: TG 436
Version / remarks:
"Acute Inhalation Toxicity" (TG 436) published by the Ministry of Enviornmental Protection of the People's Republic of China in the year of 2013.
Deviations:
no
GLP compliance:
yes
Test type:
acute toxic class method
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
Test System
Species: Rat
Strain: Sprague Dawley Grade: SPF
Supplier: Beijing Vital River Laboratory Animal Technology Co., Ltd. Animal Production License: SCXK. (Jing) 2016-0006
Animal Certificate No.: 11400700321610
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 and Body Weight: The age was between 56-62 days, and the ·body weights were between 246-248 g for male rats and 206-216 g for female rats at the commencement of dosing, respectively.
Physical Examination and Acclimatization: A physical examination, weighing and marking on the hair and cage card identifying was made in 24 hours after animals' arrival. After the physical examination a 7-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.

Test conditions:
Husbandry: Animals were housed in Room Al20-1 of the facility. Animals were raised in suspended, stainless steel cages (132.0cm xW28.0cmxH20.0cm) on cage racks (L167.0cmxW70.0cmxH171.0cm). There were 10 cages per layer, and 4 layers per rack. 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 SPF rodent maintenance feed supplied by Beijing Keaoxieli Feed CO., LTD. Analysis report of diet was provided by the supplier. Water was purified using the HT-R0l000 purity system. Drinking water was routinely analyzed. Diet and drinking water were considered not to contain any contaminants that could reasonably be expected to affect the purpose or integrity of the study.. 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 (instructive notions with respect to caring for laboratory animals) (2006, PRC Ministry of Science). The animal care and use activities required for conduct of this study were reviewed and approved by the testing facility Animal Care and Use Committee (IACUC). The spare animals were euthanized by CO2.
Active Ingredient (a.i.): (2-fluoro-3-methoxyphenyl) boronic acid Batch No.: 18051400
Route of administration:
inhalation: aerosol
Type of inhalation exposure:
snout only
Vehicle:
other: fumed silica
Mass median aerodynamic diameter (MMAD):
ca. 1.65 µm
Geometric standard deviation (GSD):
ca. 1.55
Remark on MMAD/GSD:
Aerosol Instrument for Aerodynamic Particle Sizer 3321 was used to assess the particle size distribution of the test atmosphere.
The average aerodynamic particle size less than 4 micron (mass %) for a two-time measurement was 97.66%.
Details on inhalation exposure:
Test Equipment and Administration Method

Equipment: HOPE-MED 8052H dynamic snout only aerosol inhalation instrument was used.

Atmosphere Generation System: The test item was aerosolized using a stainless steel aerosol generation system. There is a pump with a rotating baffle which drives test item to a rotating round plate where the test item is mixed with compressed air. Target concentration was achieved by adjusting air flow rate and pump infusion velocity.

Method: Before exposure, restrain each rat in a confined transparent polycrylic tube. The exposure tubes were installed in the portholes of the inhalation chamber and quantitative test item and aerosol was sent to exposure chamber. The moving speed and exposure airflow rate was adjusted as appropriated. 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 absorption unit.

Concentration Trial: Before exposure, 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:
yes
Duration of exposure:
4 h
Concentrations:
5070 mm/m^3
No. of animals per sex per dose:
First Dose: 3 males
Second Dose: 3 females
Control animals:
no
Details on study design:
Clinical Observations
Clinical observations were performed once during the exposure, and once when and 1 h after the animals' removing from the confined tubes on the exposure day. Observations on death or dying were performed twice (both in the morning and afternoon) from the first day after exposure day to the end of observation period except that they were performed once (in the morning) on necropsy days, weekends and public holidays.
Observations and records were conducted including animal fur changes, eyes and mucousmembranes, 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, and might thus be extended when considered necessary. Animals' poisoning symptoms were observed and the time of their onset, duration and disappearance were recorded.

Body Weights
The 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 and 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 arid their contents of all animals, and the location, size, hardness and the color. All the changes at necropsy were recorded. Due to no abnormalities were observed in target organs at necropsy, the histopathological examination was not performed.

Evaluation of Data
Animal number, sex,. bodyweight, necropsy and histopathology abnormal findings were summed up. The mean and standard deviations of body weight at different times were calculated.
The inhalation toxicity LCso range was found. According to GHS criteria for the acute inhalation toxicity (as shown in Table 3 below) the test item category was given. Because unit of LCso was mg/m3, the LCso divided by 1000 was converted to mg/L units when test item classification was conducted.
Key result
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 5 070 mg/m³ air
Based on:
test mat.
Mortality:
No deaths of animals under test were observed after exposure till the end of the test.
Clinical signs:
other: no signs of abnormality
Body weight:
The body weights of the animals under test showed an increasing trend.
Interpretation of results:
GHS criteria not met
Conclusions:
Based on above results, the acute inhalation LC50 (4 h) in rats for FMPBA is more than 5070 mg/m3 and the cut-off value of LC50 (4h) is- infinity mg/L. According to GHS's classification criteria of acute inhalation toxicity, the test item is classified as "Unclassified".
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
LC50
Value:
5 070 mg/m³

Acute toxicity: via dermal route

Link to relevant study records
Reference
Endpoint:
acute toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Study period:
July 05 2018 - November 03 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
other: "Acute Dermal Toxicity" (TG 402) published by the Ministry of Environmental Protection of People's Republic of China (2013)
Deviations:
no
Qualifier:
according to guideline
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity: Fixed Dose Procedure)
Deviations:
no
GLP compliance:
yes
Test type:
fixed dose procedure
Limit test:
yes
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
Justification of Test System: Rats are the preferred species of choice as historically used for the safety evauluations studies and are specified in the appropriate test guidelines.

11.2 Test Conditions
Husbandry
Animals were housed in Room A115 of the facility. Animals were individually raised in stainless steel cages (W70 cmxL80cmxH75 cm during the study.
Environmental Controls
Temperature and humidity were controlled automatically and daily recorded. The values in the aninJal room were 17-23°C for temperature, and 40%-70% for humidity. The lighting sequence was 12 hours light, 12 hours dark.
Food and Water
Animals were provided with rabbit maintenance feed supplied by Beijing Keaoxieli Feed Co., Ltd. Analysis reports of diet were supplied. by the supplier. All the nutrition components and contaminants were within the permitted limits described in the national standard.
Drinking water was purified by the HT-ROlOOO purity system. Water analysis was conducted routinely analyzed (annually), and all parameters were within the permitted limits described in the national standard.
Diet and drinking water were considered not to contain any contaminants that could reasonably be expected to affect the result, purpose and integrity of the study.
Diet and drinking water were available to the animals ad libitum during the test. Animal Welfare
Animal use comply with national animal welfare laws and regulations (instructive notions with respect to caring for laboratory aninJals) (2006, Ministry of Science, P.R.C.).The aninJal care and use activities required for conduct of this study were reviewed and approved by the testing facility Animal Care and Use Committee (IACUC).
The surviving animals were humanely killed by 10% KCl injection application (1.5 mL/kg) after anesthetized by Zoletil (5 mg/kg) at the end of the study. Their corpse treatments were entrusted to specialized agencies.
Type of coverage:
semiocclusive
Vehicle:
water
Details on dermal exposure:
Justification for the Route of Administration
The dermal route was chosen to assess the safety by a possible dermal exposure.

Testing Strategy
To investigate if the test material has dermal-irritation potential, an in vivo test in rabbits was conducted in a sequential way.
Initial Test: The test was performed initially using one animal with one control patch and one test patch for an exposure period of 4 hours.
Confirmatory Test: No irritant effects were observed in the initial test within 24hours after patch removal of the test item, a confirmatory test was conducted by exposing two additional animals simultaneously. Each animal received application of one control patch and one test patch for an exposure period of 4 hours.

Dose Design
Based on the Guidelines for the testing of chemicals "Acute Dermal Irritation/Corrosion Test" (TG 404) published by the Ministry of Environmental Protection of People's Republic of China in the year of 2013, 0.5 g oftest item was applied to the test site of skin.

Test Item Preparation and Dosage
0.5 g oftest item was weighed ahd moistened with vehicle, and then was applied to the gauze patch for the test site.

Preparation of Animals
Approximately 24 hours before the test, the back and flanks of each animal were clipped free· of fur. Care was taken to avoid abrading the skin, and only animals with healthy, intact skin by gross observation were used for the study.
Depilated area was approximately 6 cm x 6 cm for each side of the mid-line of each animal. The site (about 2.5cmx2.5cm) of depilated area below the suprascapular on each side was selected. Site A was for the control patch served as a control site, and Site B was for the test patch, both for an exposure period of 4 hours.

Administration Method
Dosing: A dose 0.5 g of the test item was applied to the gauze patch (2.5cm x 2.5cm), which was on no-irritating adhesive tape covered by medical film. 0.8 mL vehicle was dripped into the test item and modulated into a paste for uniform distribution of the substance, and which was served as test patch. The untreated gauze patch (2.5cm x 2.5cm) on no-irritating adhesive tape covered by medical film was served as control patch. The test and control patches were applied to the appointed and depilated skin, all animals were wrapped using elastic bandage and medical paper tape to form a semi-occlusive dressing, by which there was a good contact with skin and prevented animals from ingestion or inhalation of the test item. One control patch and one test patch were applied to each animal for an exposure period of 4 hours. Site A was covered with the control patch and Site B was covered with the test patch, removed after 4 hours.
Removal of Residual Test Item: Patches were removed at the end of the exposure period. The residual test substance was gently wiped off the application site by cotton moistened with tepid water. Care was taken not to alter the existing response or the integrity of the epidermis.
Dosing Frequency: Each animal was dosed once.
Dosing Time: Morning
The time interval between initial and confirmatory tests was 1 day.
Duration of exposure:
24 hours
Doses:
2000 mg/kg b.w.
No. of animals per sex per dose:
Dose 1 - 5 males
Dose 2 - 5 females
Control animals:
no
Details on study design:
Clinical Observations
All animals were observed for clinical signs once daily throughout the study.

Skin Reactions Examination
The skin sites after patch removal were examined and recorded for dermal irritation ( erythema/eschar, oedema and all local toxic effects of skin) and corrosion (ulcers, bleeding, bloody scabs, etc., by the end of observation).
The skin sites of each animal were examined and recorded for signs of skin reaction immediately, at approximately 1, 24, 48 and 72 hours after patch removal.

Grading of Skin Reactions
Dermal reactions for erythema/eschar or oedema of test sites of each animal were scored and recorded at approximately 1, 24, 48 and 72 hours after patch removal according to the grades in Table2 below.

Body Weights
Individual animal body weight was recorded within 24 hours after arrival, on the day of dosing and on the completion day of the final observations of dermal irritant symptoms.

Histopathological Examination
Histopathological examination was not conducted because there were no equivocal responses.

Evaluation of Data
Swnmary data in tables covered animal ID, gender, dosage, clinical observations, skin reactions examination, grading of skin reactions and body weights.
Mean scores for erythema/eschar and oedema at 24, 48 and 72 hours after patch removal of the test item for each animal were calculated to assess the degree of dermal irritation/corrosion of the test item in conjunction with the nature and severity of lesions, and their reversibility or lack of reversibility.
The test item was classified according to GHS criteria for the Acute Dermal Irritation/Corrosion.
Key result
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Remarks on result:
no indication of skin irritation up to the relevant limit dose level
Key result
Sex:
male
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Remarks on result:
no indication of skin irritation up to the relevant limit dose level
Mortality:
No deaths or moribund status were observed during the test.
Clinical signs:
There were no abnormal findings in all animals during the test.
Body weight:
The results indicated that most of the body weight gains of animals showed growth.
Gross pathology:
Not specified
Interpretation of results:
GHS criteria not met
Conclusions:
According to the GHS's classification criteria of acute dermal toxicity, the test item was classified as "Category 5" or "Unclassified". However, since category 5 is not implemented in the EU, GHS critieria was considered to be not met.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
LD50
Value:
2 000 mg/kg bw

Additional information

Justification for classification or non-classification