Registration Dossier

Administrative data

Description of key information

LD50 (oral, rat): >2000 mg/kg bw

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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP study in compliance with international guidelines
Reference:
Composition 0
Qualifier:
according to
Guideline:
OECD Guideline 420 (Acute Oral Toxicity - Fixed Dose Method)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.1 bis (Acute Oral Toxicity - Fixed Dose Procedure)
Deviations:
no
GLP compliance:
yes
Test type:
fixed dose procedure
Limit test:
no
Test material information:
Composition 1
Species:
rat
Strain:
other: Sprague-Dawley CD strain
Sex:
female
Details on test animals and environmental conditions:
Female Sprague-Dawley CD strain rats were supplied by Charles River (UK) Ltd, Margate, Kent, UK. On receipt the animals were randomly allocated to cages. The females were nulliparous and non-pregnant. After an acclimatisation period of at least five days the animals were selected at random and given a number unique within the study by indelible ink-marking on the tail and a number written on a cage card.
At the start of the study the animals were eight to twelve weeks of age. The bodyweight variation did not exceed ±20% of the mean bodyweight of any previously dosed animal.
The animals were housed in groups of up to four in suspended solid-floor polypropylene cages furnished with woodflakes. With the exception of an overnight fast immediately before dosing and for approximately three to four hours after dosing, free access to mains drinking water and food (Certified Rat and Mouse Diet) was allowed throughout the study. The diet, drinking water and bedding were routinely analysed and were considered not to contain any contaminants that would reasonably be expected to affect the purpose or integrity of the study.
Route of administration:
oral: gavage
Vehicle:
unchanged (no vehicle)
Details on oral exposure:
All animals were dosed once only by gavage using a metal cannula attached to a graduated syringe. The volume administered to each animal was calculated according to its fasted bodyweight at the time of dosing.
Doses:
2000 mg/Kg
No. of animals per sex per dose:
5 female rats
Control animals:
no
Details on study design:
Using available information on the toxicity of the test material, 2000 mg/kg was chosen as the starting dose.
In the absence of toxicity at a dose level of 2000 mg/kg, an additional group of animals was treated.
A total of five animals were therefore treated at a dose level of 2000 mg/kg in the study.
All animals were dosed once only by gavage using a metal cannula attached to a graduated syringe. The volume administered to each animal was calculated according to its fasted bodyweight at the time of dosing.
Clinical observations were made 1/2, 1,2, and 4 hours after dosing and subsequently once daily for fourteen days. Morbidity and mortality checks were made twice daily.
Individual bodyweights were recorded on Day 0 (the day of dosing) and on Days 7 and 14.
At the end of the observation period the animals were killed by cervical dislocation. All animals were subjected to gross necropsy. This consisted of an external examination and opening of the abdominal and thoracic cavities. The appearance of any macroscopic abnormalities was recorded. No tissues were retained.

Statistics:
not applicable
Preliminary study:
Using available information on the toxicity of the test material, 2000 mg/kg was chosen as the starting dose.
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
there were no death
Clinical signs:
No signs of systemic toxicity were noted
Body weight:
Individual bodyweights and bodyweight changes are given in Table 2.
All animals showed expected gains in bodyweight over the study period.
Gross pathology:
No abnormalities were noted at necropsy.
Interpretation of results:
study cannot be used for classification
Remarks:
Migrated information
Conclusions:
The acute oral median lethal dose (LD50) of the test material in the female Sprague-Dawley CD strain rat was estimated to be greater than 2000 mg/kg bodyweight (Globally Ilarmonised Classification System Category — Unclassified).
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
LD50
2 000 mg/kg bw

Acute toxicity: via inhalation route

Link to relevant study records
Reference
Endpoint:
acute toxicity: inhalation
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
May 8 to July 8, 2003
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: guideline test for read across, well documented
Reference:
Composition 0
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
Deviations:
no
Principles of method if other than guideline:
Two groups of 6 male rats each were exposed to aerosol atmospheres of test aritcle in air. Rats were exposed nose-only for a single, 4-hour period. Following exposure, 3 rats from the 0.10 mg/L exposure were retained for a 1-day recovery period; the remaining 3 rats were retained for a 14-day recovery period. Rats from the 1.8 mg/L exposure were retained for a 14-day recovery period.
GLP compliance:
not specified
Test type:
standard acute method
Limit test:
no
Test material information:
Composition 1
Species:
rat
Strain:
Crj: CD(SD)
Sex:
male
Details on test animals and environmental conditions:
B. Test Species
Young adult, male Crl:CD®(SD)IGS BR rats were received from Charles River Laboratories, Inc., Raleigh, North Carolina. The rats were approximately 7 or 8 weeks old on the day of arrival.
Rats have historically been used in safety evaluation studies for inhalation toxicity testing. The Crl:CD®(SD)IGS BR rat was selected based on consistently acceptable health status and on extensive experience with the strain at Haskell Laboratory.
C. Animal Husbandry
1. Quarantine
Rats were quarantined after arrival for 6 days prior to testing. During the quarantine period, rats were weighed and observed for clinical signs of disease 3 times.
2. Animal Selection
Prior to each exposure, 6 male rats were selected for use on the study from the rats that were released from quarantine. The selected animals had been gaining weight at a normal rate and demonstrated no overt signs of disease, and were the appropriate age and body weight. No attempt was made to randomly group animals.
3. Identification
Each rat was assigned an animal number which was recorded on a card affixed to the cage. Prior to exposure, the tail of each animal and cage card were marked with water-insoluble pens so that each animal could be identified after exposure and during the recovery period.
4. Housing
Except during exposure, rats were housed singly in stainless steel, wire-mesh cages suspended above cage boards.
5. Animal Room Environment
Rats were housed in proximity to the inhalation chambers. Animal rooms were maintained at a temperature of 18-26°C (targeted to 22-24ºC) and a relative humidity of 30-70% (targeted to 40-60%). Animal rooms were artificially illuminated (fluorescent light) on an approximate 12 hour light/dark cycle. Excursions outside of these ranges were of insufficient magnitude and/or duration to have adversely affected the validity of the study.
6. Feed and Water
Except during exposure, PMI® Nutrition International, LLC Certified Rodent LabDiet® 5002 and tap water were available ad libitum.
7. Animal Health and Environmental Monitoring Program
As specified in the Haskell Laboratory animal health and environmental monitoring program, the following procedures are performed periodically to ensure that contaminant levels are below those that would be expected to impact the scientific integrity of the study:
• Water samples are analyzed for total bacterial counts, and the presence of coliforms, lead, and other contaminants.
• Feed samples are analyzed for total bacterial, spore, and fungal counts.
• Samples from freshly washed cages and cage racks are analyzed to ensure adequate
sanitation by the cagewashers.
Certified animal feed is used, guaranteed by the manufacturer to meet specified nutritional requirements and not to exceed stated maximum concentrations of key contaminants, including specified heavy metals, aflatoxin, chlorinated hydrocarbons, and organophosphates. The presence of these contaminants below the maximum concentration stated by the manufacturer would not be expected to impact the integrity of the study.
The animal health and environmental monitoring program is administered by the attending laboratory animal veterinarian. Evaluation of these data did not indicate any conditions that affected the validity of the study.
Route of administration:
inhalation: vapour
Type of inhalation exposure:
nose only
Vehicle:
air
Details on inhalation exposure:
Inhalation Exposure System
1. Atmosphere Generation
Chamber atmospheres were generated by aerosolization of the solid test substance in air with a glass, round-bottom flask placed in a heating mantle. The flask was heated to approximately 100ºC to melt the test substance. Filtered, high-pressure air, metered into the flask by a Brooks model 5851E mass flow controller, carried the evolved vapor through heated glass tubing. For the 0.10 mg/L exposure, 2 chambers were used. The vapor passed from the heated tubing into the first chamber where a condensation aerosol was formed. The aerosol then passed into the second chamber for animal exposure. For the 1.8 mg/L exposure, the vapor passed from the heated tubing in the exposure chamber where an aerosol was formed.
Test atmospheres were exhausted through an MSA charcoal/HEPA filter cartridge prior to discharge into the fume hood.
2. Chamber Construction and Design
The exposure chambers were constructed of glass (cylindrical) with a nominal internal volume of 34 L. A baffle inside each chamber promoted uniform distribution of the test atmosphere.
3. Exposure Mode
During exposure, animals were individually restrained in perforated stainless steel cylinders with conical nose pieces. The restrainers were inserted into a polymethylmethacrylate faceplate which was attached to the exposure chamber so that the nose of each animal extended into the exposure chamber.
Characterization of Chamber Atmosphere
1. Test Substance Sampling and Analysis
The atmospheric concentration of H-25777 was determined by gravimetric analysis at approximately 30-minute intervals during each exposure. Known volumes of chamber
atmosphere were drawn from the sampling port through a 25 mm filter cassette that contained a pre-weighed Gelman glass fiber (Type A/E) filter. The filters were weighed on a Cahn model C-33 Microbalance®. The atmospheric concentration of H-25777 was calculated from the difference in the pre- and post-sampling filter weights divided by the volume of chamber atmosphere sampled.
2. Particle Size Determination
A sample to determine particle size distribution (mass median aerodynamic diameter and percent particles less than 1, 3, or 10 µm diameter) was taken during each exposure with a Sierra® Series 210 cyclone preseparator/Cascade impactor and Sierra® series 110 constant flow air sampler.
3. Environmental Monitoring
Chamber airflow was set at the beginning of each exposure to achieve at least 10 air changes per hour. The airflow was monitored continually with a calibrated Brooks model 5851E mass flow controller and recorded initially and whenever changes were made during each exposure.
Chamber temperature was targeted at 22 ± 2°C. The temperature was monitored continually with a digital traceable thermometer and recorded 3 times during each exposure. Chamber relative humidity was targeted at 50 ± 10%. The relative humidity was measured with an Omega model RH5100C digital psychrometer and recorded 3 times during each exposure. Chamber oxygen concentration was targeted to be at least 19%. The oxygen concentration was measured with a Biosystems model 3100R oxygen analyzer and recorded 3 times during each exposure.





Analytical verification of test atmosphere concentrations:
yes
Duration of exposure:
4 h
Concentrations:
0.1 and 1.8 mg/l
No. of animals per sex per dose:
3
Control animals:
no
Details on study design:
Two groups of 6 male rats each were exposed to aerosol atmospheres of H-25777 in air. Rats were exposed nose-only for a single, 4-hour period. Following exposure, 3 rats from the 0.10 mg/L exposure were retained for a 1-day recovery period; the remaining 3 rats were retained for a 14-day recovery period. Rats from the 1.8 mg/L exposure were retained for a 14-day recovery period.
Rats were approximately 8 weeks old and weighed between 258 and 279 grams at the time of exposure.
Animals were observed for mortality 4 times during each exposure and observed for mortality and clinical signs of toxicity immediately after exposure. During the recovery period, all rats were observed each day for mortality. Rats were weighed and observed for clinical signs of toxicity on the day following exposure and at least twice more during the recovery period. At the end of both recovery periods, all rats from the 0.10 mg/L group were sacrificed for anatomic pathology evaluation. All rats from the 1.8 mg/L exposure were sacrificed by carbon dioxide asphyxiation and discarded.
Anatomic Pathology Evaluation
All animals from the 0.10 mg/L exposure underwent a gross pathology examination at sacrifice. In addition, the lungs, trachea, nose (4 cross sections), larynx, and pharynx were processed for histopathology and microscopically examined. Lung weights were obtained for the 3 animals sacrificed 1-day after exposure to 0.10 mg/L.
Statistics:
no applicable
Sex:
male
Dose descriptor:
LC50
Effect level:
> 1.8 mg/L air (analytical)
Based on:
test mat.
Exp. duration:
4 h
Remarks on result:
other: Other than slight body weight losses measured in 3 of 6 rats one day after exposure to 1.8 mg/L of H-25777, there were no significant clinical signs observed in the exposed rats at either test concentration.
Mortality:
No deaths occurred during the study.
Clinical signs:
No clinical signs of toxicity were observed during the remainder of the recovery period.
Body weight:
No body weight losses were observed in rats exposed to 0.10 mg/L immediately after exposure or after the 14-day recovery period.
Three of 6 rats exposed to 1.8 mg/L showed slight weight loss (<10 grams) on the day following exposure, followed by a normal weight gain rate (approximately 5-10 g/day) for the remainder of the recovery period.
Gross pathology:
There were no appreciable differences between the 2 groups. The alveolar hemorrhages were very minimal, and were likely agonal changes occurring at the time of sacrifice.
Lung weights in 3 rats sacrificed 1 day after exposure to H-25777 at 0.10 mg/L did not appear to be different from control rats of similar weights in a recent Haskell Laboratory inhalation study.
Although no concurrent controls were provided, the changes present in the tissues examined were considered to be of the type and severity expected in this strain of rats, and none of them appeared to be the result of treatment.

Other findings:
Exposure Conditions
Animals were exposed to H-25777 at mean concentrations of 0.10 or 1.8 mg/L. The atmospheres generated in this study were considered to be marginally respirable in rats, as the mass median aerodynamic diameters (MMAD) were 5.8 and 6.8 µm. Several attempts were made to produce a smaller, more respirable aerosol size; however, it appeared that this could not be accomplished at these high concentrations due to the condensation aerosol forming large particle agglomerates.
Chamber temperature ranged from 23 to 24°C, chamber relative humidity ranged from 46 to 68%, chamber airflow ranged from 15 to 25 L/min, and the oxygen concentration was 21%.
Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
Under the conditions of this study, it can be concluded that LC50 of test article is considered to be greater than 1.8 mg/L after nose-only for a single, 4-hour exposure.
However, the mass median aerodynamic diameter (MMAD) of the tested aerosol at this concentration was 6.8 µm. Targeted MMADs in inhalation studies are from 1 - 4 µm to allow maximum lung exposure. Several attempts were made to produce smaller, more respirable aerosol size; however, it appeared that this could not be accomplished at these high concentrations due to the condensation aerosol forming large particle agglomerates. Thus the inhalation hazard cannot be determined since the aerosols formed were not respirable to the rats, particularly the pulmonary region.
Executive summary:

The study was performed to determine a 4-hour inhalation approximate lethal concentration (ALC) of test article in male rats. Two groups of 6 male rats each were exposed to aerosol atmospheres of test article in air. Rats were exposed nose-only for a single, 4-hour period. Following exposure, 3 rats from the 0.10 mg/L exposure were retained for a 1-day recovery period. Rats from the 1.8 mg/L exposure were retained for a 14- day recovery period. At the end of both recovery periods, all rats from the 0.10 mg/L group were sacrificed for anatomic pathology evaluation. All rats from the 1.8 mg/L exposure were sacrificed by carbon dioxide asphyxiation and discarded. Other than slight body weight losses measured in 3 of 6 rats one day after exposure to 1.8 mg/L of H-25777, there were no significant clinical signs observed in the exposed rats at either test concentration. Histological examination of rats exposed to 0.10 mg/L showed no treatment- related effects.

Therefore, LC50 of test article is considered to be greater than 1.8 mg/L based on the results of this study.

However, the mass median aerodynamic diameter (MMAD) of the tested aerosol at this concentration was 6.8 µm. Targeted MMADs in inhalation studies are from 1 - 4 µm to allow maximum lung exposure. Several attempts were made to produce smaller, more respirable aerosol size; however, it appeared that this could not be accomplished at these high concentrations due to the condensation aerosol forming large particle agglomerates. Thus the inhalation hazard cannot be determined since the aerosols formed were not respirable to the rats, particularly the pulmonary region.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
The mass median aerodynamic diameter (MMAD) of the tested aerosol at this concentration was 6.8 µm. Targeted MMADs in inhalation studies are from 1 - 4 µm to allow maximum lung exposure. Several attempts were made to produce smaller, more respirable aerosol size; however, it appeared that this could not be accomplished at these high concentrations due to the condensation aerosol forming large particle agglomerates. Thus the inhalation hazard cannot be determined since the aerosols formed were not respirable to the rats, particularly the pulmonary region.

Acute toxicity: via dermal route

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

The acute oral study was conducted following OECD guideline under the GLP conditions. A total of five animals were treated at a dose level of 2000 mg/kg in the study. Individual bodyweights were recorded on Day 0 (the day of dosing) and on Days 7 and 14.At the end of the observation period the animals were killed by cervical dislocation. All animals were subjected to gross necropsy. No abnormality was observed at gross necropsy. No signs of systemic toxicity and were noted. There was no death during the test period. Therefore, LD50 of test article for acute oral toxicity is considered to be greater than 2000 mg/kg/day.

There is no direct information of inhalation route for target substance. Considering the structural similarity,another study conducted with the Acrylate Monomer - C8 analogue was chosen for read across. In this study, 6 male rats were exposed nose-only for a single, 4-hour period to test item in airat mean concentrations of 0.10 or 1.8 mg/L. Other than slight body weight losses measured in 3 of 6 rats one day after exposure to 1.8 mg/L of test article, there were no significant clinical signs observed in the exposed rats at either test concentration. Histological examination of rats exposed to 0.10 mg/L showed no treatment-related effects. Thus, 1.8 mg/L was considered to be the LC50 of test article for acute inhalation. The LC50 of target test is therefore expected to be >1.8 mg/L. However, the mass median aerodynamic diameter (MMAD) of the tested aerosol at this concentration was 6.8 µm. Targeted MMADs in inhalation studies are from 1 - 4 µm to allow maximum lung exposure. Several attempts were made to produce smaller, more respirable aerosol size; however, it appeared that this could not be accomplished at these high concentrations due to the condensation aerosol forming large particle agglomerates. Thus the inhalation hazard cannot be determined since the aerosols formed were not respirable to the rats, particularly the pulmonary region.

According to REACH 1972/2006,Annex VIII,column 2,8.5, in addition to oral route, for substance other than gases, the information mentioned under 8.5.2 to 8.5.3 shall be provided for at least on other route. Thus, the acute dermal toxicity study can be waived since the data of acute toxicity by oral and inhalation route is available and sufficient for assessment of acute toxicity of test article.


Justification for selection of acute toxicity – dermal endpoint
According to REACH 1972/2006,Annex VIII,column 2,8.5, in addition to oral route, for substance other than gases, the information mentioned under 8.5.2 to 8.5.3 shall be provided for at least on other route. Thus, the acute dermal toxicity study can be waived since the data of acute toxicity by oral and inhalation route is available and sufficient for assessment of acute toxicity of test article.

Justification for classification or non-classification

Based on the available data, target substance should be classified neither for acute oral toxicity nor for acute inhalation toxicity under the CLP (Regulation EC No. 1272/2008).