Registration Dossier

Administrative data

Description of key information

LD50 (oral, rat) > 5000 mg/kg bw 
LC50 (inhalation, rat) > 11 mg/L air
LD50 (dermal, 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
Study period:
2006-09-05 to 2006-11-06
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study according to OECD Guideline 423: Study under UK GLP Regulation SI 1999/3106;
Qualifier:
according to
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Test type:
acute toxic class method
Limit test:
yes
Species:
rat
Strain:
Sprague-Dawley
Sex:
female
Details on test animals and environmental conditions:
The animals 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 bodyweights fell within an interval of ± 20% of the mean initial bodyweight of the first treated group.

The animals were housed in groups of three 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 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.

The temperature and relative humidity were set to achieve limits of 19 to 25°C and 30 to 70% respectively. Any occasional deviations from these targets were considered not to have affected the purpose or integrity of the study. The rate of air exchange was at least fifteen changes per hour and the lighting was controlled by a time switch to give twelve hours continuous light (06:00 to 18:00) and twelve hours darkness.
Route of administration:
oral: gavage
Vehicle:
unchanged (no vehicle)
Details on oral exposure:
MAXIMUM DOSE VOLUME APPLIED: 4.96 ml/kg

CLASS METHOD (if applicable)
- Rationale for the selection of the starting dose: based on available historical data/information
Doses:
5000 mg/kg
No. of animals per sex per dose:
3 female animals
Control animals:
not specified
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations: 30 min, 1h, 2h, 4h, then daily up to 14 days
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight,organ weights
Sex:
female
Dose descriptor:
LD50
Effect level:
> 5 000 mg/kg bw
Based on:
test mat.
Mortality:
There were no deaths.
Clinical signs:
Hunched posture and pilo-erection were noted in one animal two days after dosing and in one animal three days after dosing. Animals appeared normal three or four days after dosing.
Body weight:
Animals showed expected gain in bodyweight over the study period.
Gross pathology:
No abnormalities were noted at time of 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 5000 mg/kg bodyweight. Therefore DBE is not classified according to Annex VI of the Directive 67/548/CEE and according to EU Regulation 1272/2008 (CLP).
Executive summary:

DBE has been tested in an acute oral toxicity study using female Sprague Dawley rats, according to OECD guideline n° 423 and EU guideline n° B.1 tris in compliance with Good Laboratory Practice.

The test item was administered without vehicle once by oral route (gavage) to a group of three fasted female rats at a dose level of 5000 mg/kg.

Clinical signs, mortality and body weight gain were checked for a period of up to 14 days after the single administration of DBE. All animals were subjected to necropsy.

No mortality was recorded during the study. Body weight gain of the treated animals was not affected by treatment. At necropsy, no abnormalities were observed in any animal.

As the LD 50 is higher than 5000 mg/kg, DBE is not classified according to Annex VI of the Directive 67/548/CEE and according to EU Regulation 1272/2008 (CLP).

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
LD50
5 000 mg/kg bw
Quality of whole database:
GLP and OECD 423 guideline compliant (Klimish score = 1)

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:
April 16 to May 18, 1990
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Study similar to OECD Guideline 403;
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
Deviations:
no
GLP compliance:
no
Remarks:
but with quality assurance documentation
Test type:
standard acute method
Limit test:
no
Species:
rat
Strain:
Crj: CD(SD)
Sex:
male/female
Details on test animals and environmental conditions:
Young adult male and female Crl:CD BR rats were received from Charles River Breeding Laboratories. Rats were quarantined for approximately 1 week prior to testing and were weighed and observed 3 times during the quarantine period. During the quarantine period rats were housed singly in 5” x 11" x 7" suspended, stainless steel, wire mesh cages. After exposure, rats were housed (sexes separate) either 1 or 2 per cage in 8” x 14" x 8”cages. Prior to exposure, rats’ tails and cage cards were color-coded so that individual rats could be identified after exposure. For rats housed in pairs, the rat with the lower number was identified by a slash in the right ear.

On the day of exposure, rats were approximately 7-8 weeks old; male rats weighed 238-281 g and female rats weighed 174-222 g. Except during exposure, Purina Certified Rodent Chow and water were available ad libidum. Animal rooms were maintained on a timer-controlled, 12 hour/12 hour light/dark cycle.

Environmental conditions of the animal rooms were targeted for a temperature of 23°+ 2°C and relative humidity of 50 % + 10 %. Excursions outside these ranges were judged to have been of insufficient magnitude and/or duration to have adversely affected the validity of the study.
Route of administration:
inhalation: aerosol
Type of inhalation exposure:
nose only
Vehicle:
clean air
Details on inhalation exposure:
For the LC50 determination, groups of 5 male and 5 female rats were individually restrained in stainless steel restrainers with conical nose pieces and exposed nose-only to DBE. Each restrainer was inserted into a 38-L cylindrical exposure chamber such that only the nose of each protruded into the chamber. To determine whether the ocular response was diminished by nose-only exposure, an additional group of 4 male rats was placed in a stainless steel and aluminum wire mesh cage within the exposure chamber and exposed whole-body.

Each group of rats was exposed for a single. 4-hour period to an aerosol / vapor mixture of DBE in air. Rats were observed for clinical signs of toxicity (e.g., general physical and clinical disposition. including unusual movements, respiration. etc.) before and during exposure. upon removal from the animal restrainers approximately 30 minutes after the cessation of exposure, and daily thereafter (weekends and holidays excluded unless warranted by the rats' condition). During the exposure. the use of restrainers limited the observations mainly to the presence of nasal or oral discharges. In addition. a sharp rap was periodically delivered to the exposure chamber and rats were observed for a general startle response (rapid movement) during the exposure. This procedure was used primarily as a gross indicator of unconsciousness or death. Rats were weighed prior to exposure, and were weighed daily. weekends and holidays excluded except when warranted by the rats' condition, during the 14-day recovery period.

Atmosphere Generation

DBE was generated by atomization. Test atmospheres of DBE were generated by atomizing the liquid test material using an Airlife, Solo-Sphere Nebulizer. Conditioned, filtered houseline air (approximately 8-16 L/min) was used to generate the aerosol and to carry the aerosol directly into the 38 L cylindrical, glass exposure chamber. Located immediately inside the chamber was a plastic deflection plate (approximately 12.5 cm diameter) which acted as a baffle to promote turbulent flow and facilitate uniform distribution of the aerosol within the chamber. Chamber atmospheres were exhausted through tandem, dry-ice cold traps and an MSA activated charcoal/HEPA filter cartridge prior to discharge into a fume hood
Analytical verification of test atmosphere concentrations:
yes
Duration of exposure:
4 h
Concentrations:
3.5, 5.6, 11.0 mg/L DBE;
No. of animals per sex per dose:
5 males;
5 females;
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing:starting 30 min after exposure, then daily for 14 days
- Necropsy of survivors performed: no data
- Other examinations performed: opthalmology
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 11 mg/L air
Based on:
test mat.
Exp. duration:
4 h
Mortality:
No rats died following exposure to DBE at atmospheric concentrations as great as 11 mg/L, the highest concentration tested.
Clinical signs:
Clinical signs of toxicity observed in some male and female rats during the recovery period included red nasal, ocular or oral discharges, wet, yellow-stained perineum, and hunched posture. All clinical signs were transient and had resolved by study day 4.
Body weight:
Rats from all exposure groups exhibited slight to severe body weight losses 1 day after exposure (group average weight losses ranged from approximately 5-11% and 2-9% of initial body weight in male and female rats, respectively). In addition, some transient, slight to moderate weight losses occurred in some male and female rats during the first or second weeks of the recovery period.
Other findings:
Prior to exposure, the eyes of rats from the 11 mg/L group were evaluated and considered ophthalmoscopically normal. After exposure, the eyes of rats from the 3.5 or 11 mg/L groups were considered to be ophthalmoscopically normal; the corneal and pupillary reflexes were normal. At 5.6 mg/L, however, bilateral mild chemosis (edema/swelling) of the bulbar conjunctivae was found in all rats. Although a subepithelial corneal opacity was also noted in 1 rat exposed to 5.6 mg/L, this finding was believed to be due to a preexisting injury and not compound-related.
Interpretation of results:
practically nontoxic
Remarks:
Migrated information Criteria used for interpretation of results: OECD GHS
Conclusions:
Under the conditions of this study, the 4-hour LC50 for DBE exceeds 11 mg/L, the highest concentration tested.
Executive summary:

DBE has been tested in an acute inhalation toxicity study using Crj: CD(SD) rats, according to OECD guideline n° 403 and EU guideline n° B.2 with quality assurance documentation.

5 males and 5 females per dose were exposed to 3.5, 5.6 or 11.0 mg/L DBE for 4 h via inhalation (nose only). Clinical signs, mortality and body weight gain were checked for a period of up to 14 days after the single exposure to DBE. All animals were subjected to necropsy.

No rats died following exposure to DBE at atmospheric concentrations as great as 11 mg/L, the highest concentration tested. Rats from all exposure groups exhibited slight to severe body weight losses 1 day after exposure (group average weight losses ranged from approximately 5-11% and 2-9% of initial body weight in male and female rats, respectively). In addition, some transient, slight to moderate weight losses occurred in some male and female rats during the first or second weeks of the recovery period.

Clinical signs of toxicity observed in some male and female rats during the recovery period included red nasal, ocular or oral discharges, wet, yellow-stained perineum, and hunched posture. All clinical signs were transient and had resolved by study day 4.

Ophthalmoscopic findings in the 5.6 mg/L group were believed to be due to a preexisting injury and not compound related as no comparable findings were seen in the 3.5 or 11 mg/L groups.

As the LC 50 is higher than 11 mg/L, DBE is not classified according to Annex VI of the Directive 67/548/CEE and according to EU Regulation 1272/2008 (CLP).

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
LC50
11 000 mg/m³
Quality of whole database:
study equivalent to OECD guideline 403 study (Klimish score = 2)

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:
January 09 to March 26, 1987
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study equivalent to OECD Guideline 402; Study under GLP Regulation;
Qualifier:
according to
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Test type:
standard acute method
Limit test:
yes
Species:
rat
Strain:
Crj: CD(SD)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River, F-76410 Saint Aubin les Elbeuf, France;
- Weight at study initiation: males: 266 ± 2 g, females: 247 ±10 g;
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 6 days;


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 60 ± 20
- Photoperiod (hrs dark / hrs light): 12/12;
Type of coverage:
semiocclusive
Vehicle:
unchanged (no vehicle)
Details on dermal exposure:
TEST SITE
- Area of exposure: 5 x 7 cm²; shaved backs and flanks
- % coverage: < 10%;
- Type of wrap if used: gauze


REMOVAL OF TEST SUBSTANCE
- Washing (if done): not specified;
- Time after start of exposure: 24 hours


TEST MATERIAL
- Amount(s) applied (volume or weight with unit): not specified;
- Concentration (if solution): 2000 mg/kg bw;
Duration of exposure:
24 hours
Doses:
2000 mg/kg bw;
No. of animals per sex per dose:
5 males;
5 females;
Control animals:
not required
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: observations: at least once daily; weighting: at days 1 (pre-administration), 5, 8, and 15
- Necropsy of survivors performed: yes
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
No deaths occured.
Clinical signs:
No clinical signs of toxicity were detected.
Body weight:
Body weight was not affected by exposure to the test article.
Gross pathology:
No signs of gross pathology have been detected
Interpretation of results:
practically nontoxic
Remarks:
Migrated information Criteria used for interpretation of results: OECD GHS
Conclusions:
Under the conditions of the current study, the test article DBE showed no signs of toxicity in female and male rats over a period of 14 days when applied dermally once at 2000 mg/kg bw. The LD50 can therefore be concluded to be greater than 2000 mg/kg bw. Based on this Result DBE is not classified according to Annex VI of the Directive 67/548/CEE and according to EU Regulation 1272/2008 (CLP).
Executive summary:

DBE has been tested in an acute dermal toxicity study using Crj: CD(SD) rats, according to OECD guideline n° 402 and EU guideline n° B.3 in compliance with Good Laboratory Practice.

The test item was applied once under semiocclusive conditions to the shaved backs and flanks of the animals at a dose level of 2000 mg/kg for 24 h (5 males + 5 females per dose, 5 x 7 cm²).

Clinical signs, mortality and body weight gain were checked for a period of up to 14 days after the single administration of DBE. All animals were subjected to necropsy.

No mortality was recorded in both groups during the study. Body weight gain of the treated animals was not affected by treatment. At necropsy, no abnormalities were observed in any animal.

As the LD 50 is higher than 2000 mg/kg, DBE is not classified according to Annex VI of the Directive 67/548/CEE and according to EU Regulation 1272/2008 (CLP).

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
LD50
2 000 mg/kg bw
Quality of whole database:
GLP and OECD 402 guideline compliant (Klimish score = 1)

Additional information

Oral route:

Two studies, both of reliability 1 according to Klimisch cotation criteria, are available for oral route (Pooles, 2006 and Clouzeau, 1987). The 2006 study was selected as the key study since the tested dose (5000 mg/kg) was higher than that tested in the other study (2000 mg/kg), although 2000 mg/kg was administered to a higher number of test animals of both genders (5 male and 5 female rats compared to 3 female rats). No death occurred during the 2-week observation period in either study. The LD50 for oral route in rats was higher than 5000 mg/kg. No classification for acute oral toxicity is therefore warranted on the basis of these in vivo data.

 

Inhalation route:

One study, of reliability 2 according to Klimisch cotation criteria, is available for inhalation route (Valentine, 1990). Following a 4-hour nose-only exposure of 5 male and 5 female rats to concentrations up to 11 mg/L, no death occurred during the 2-week observation period. The 4-hour LD50 for inhalation in rats was higher than 11 mg/L. No classification for acute inhalation toxicity is therefore warranted on the basis of these in vivo data.

 

Dermal route:

Two studies are available for dermal route. One study in rats (Clouzeau, 1987) was of reliability 1 according to Klimisch cotation criteria and was therefore selected as the key study. The other study, conducted in rabbits (Ford, 1981) was considered of low reliability (3) due to major uncertainties. In the rat study, no deaths occurred following dermal application of 2000 mg/kg for 24 hours to 5 males and 5 females during a 2-week observation period. Similarly in the rabbit study, no deaths occurred following dermal application of doses up to 2250 mg/kg for 24 hours to 6 males during a 2-week observation period. Based on the key study, the LD50 for dermal route in rats was higher than 2000 mg/kg. No classification for acute dermal toxicity is therefore warranted on the basis of these in vivo data.


Justification for selection of acute toxicity – oral endpoint
This study was selected as the key study since the tested dose (5000 mg/kg) was higher than that tested in the other study (2000 mg/kg)

Justification for selection of acute toxicity – inhalation endpoint
only one study available

Justification for selection of acute toxicity – dermal endpoint
the most reliable study (Klimish score 1 vs. 3 for the other one)

Justification for classification or non-classification

Due to the absence of mortality following exposure of rats to limit test doses or concentrations and a 2-week observation period whatever the route of administration, no classification for acute toxicity is warranted according to the criteria of Annex VI Directive 67/548/EEC or EU Regulation 1272/2008 (CLP).