Registration Dossier

Administrative data

Endpoint:
acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
28 May 2010 - 14 June 2010
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study under GLP, according to international guidelines and with full documentation

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010
Report Date:
2010

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.2 (Acute Toxicity (Inhalation))
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.1300 (Acute inhalation toxicity)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- The analysis was performed for the reference sample "Sample 2010" as defined in Section 1.4
- Lot/batch No.: 10013593
- Expiration date of the lot/batch: March 2011
- Storage condition of test material: In refrigerator at 2-8 ºC, under inert gas, away from light

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
Species / Strain: Rat, CRL (WI) BR of Wistar origin
Source: Toxi-Coop Zrt. 1103 Budapest, Cserkesz u. 90.
Hygienic level: SPF at arrival and kept in good conventional environment during the study.
Age of animals at arrival: Young adult rats, 7-9 weeks old
Body weight range at treatment: Male: 278-306 g; Female: 196-214 g
Number of animals: 5 male and 5 female (nulliparous and non pregnant animals) rats
Number of animals/group: 5 animals/ sex
Acclimatisation time: 15 days
Acclimatisation to the test apparatus: 11 days
- Fasting period before study: no
- Housing: Group caging (2 or 3 animals, by sex, per cage)
- Diet (e.g. ad libitum): The animals received ssniff® SM R/M-Z+H complete diet produced by ssniff Spezialdiäten GmbH, D-59494 Soest Germany, ad libitum.
- Water (e.g. ad libitum): Animal received tap water from watering bottles.

Environmental conditions:
Illumination: Artificial light, from 6 a.m. to 6 p.m.
Temperature: 22 ± 3 °C
Relative humidity: 30 - 70 %
Ventilation: 8-12 air exchanges/hour by central air-condition system.
Environmental conditions were maintained by an air-condition system. Temperature and relative humidity were verified and recorded daily during the study.


Administration / exposure

Route of administration:
inhalation: vapour
Type of inhalation exposure:
nose only
Vehicle:
air
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: anodised aluminium Flow Past Exposure Chamber (CR Equipment SA, Switzerland)
- Exposure chamber volume: 6 L
- Method of holding animals in test chamber: polycarbonate restraining tubes
- Source and rate of air: rate: Source: no data; 1.0 L/min
- Method of conditioning air: The air was supplied by an oil-free air compressor and was filtered in a two-stage filter set (Festo: 1μm and 0.01 μm). The temperature of the air was regulated by a heat exchanger. The air was not humidified in order to avoid modification of particle size distribution.
- System of generating particulates/aerosols: The test atmosphere was generated by a compressed air nebulizer (TSE Systems GmbH, Bad Homburg, Germany.) In the nebulizer the air flow rate was 15 L/min.
- Method of particle size determination: 7-stage cascade impactor type 02-150 (IN-TOX Products, N.M., USA)
- Treatment of exhaust air: no data
- Temperature, humidity, pressure in air chamber: 23.8-26.3 °C; 26.5-9.1 % relative humidity; pressure: no data

TEST ATMOSPHERE
- Brief description of analytical method used: The actual test atmosphere concentration was measured five times during exposure by pulling a volume of 10 L of test atmosphere (flow rate: 2.0 L/min) through sampling cylinders (length: 100mm, dia. 30mm) tightly filled with about 20g of granulated activated carbon (Reanal Private Ltd., Prod. No. 29903-1-99-33, pellet diameter 4mm). The amount of adsorbed test item was determined gravimetrically by the balance type Mettler PM100 (d = 1mg).
- Samples taken from breathing zone: yes

VEHICLE
no vehicle used

TEST ATMOSPHERE (if not tabulated)
- Particle size distribution: see any other information on materials and methods
- MMAD (Mass median aerodynamic diameter) / GSD (Geometric st. dev.):
MMAD: Mass Median Aerodynamic Diameter (mean): 0.59 μm
GSD: Geometric Standard Deviation (mean): 1.35
Inhalable Fraction: Aerodyn. Diameter < 4μm: 100 %

CLASS METHOD (if applicable)
- Rationale for the selection of the starting concentration: upper limit of the classification into acute toxicity category 4 according to the CLP-regulation.
Analytical verification of test atmosphere concentrations:
yes
Remarks:
method see above
Duration of exposure:
4 h
Remarks on duration:
nose-only exposure
Concentrations:
21.0 mg/L (Mean Achieved Concentration)
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:
Animals were checked twice daily during the observation period for morbidity and/or mortality.
All animals were observed for clinical signs at 1st, 2nd and 3rd hours during exposure, as soon as practicable following removal from restraint, one hour after exposure and subsequently once daily for fourteen days.
Individual bodyweights were recorded on the day of exposure Day 0 (prior to exposure) and Days 1, 3, 7 and 14.

- Necropsy of survivors performed: yes; At the end of the fourteen day observation period, the animals were euthanized by exsanguination under anaesthesia (intra-peritoneal injection of Euthasol® 40%) and gross necropsies were performed. These included a detailed examination of the abdominal and thoracic cavities with special attention given to the respiratory tract for macroscopic signs of irritancy or local toxicity.

- Other examinations performed: clinical signs, body weight

Results and discussion

Effect levels
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 21 mg/L air (analytical)
Based on:
test mat.
Exp. duration:
4 h
Mortality:
No animal died during the study.
Clinical signs:
In all male and female animals were observed decreased activity and general reaction and incoordination after removal from restraint on completion of the exposure. In four female and in three male animals were found tremor at this time. One hour after the exposure all animals slept.
These clinical signs were considered as test item related effect.
All male and female animals were symptom-free from first day of observation period.
Body weight:
In both genders of some animals body weight loss was observable on the day of inhalation exposure. In both sexes a compensation of body weight loss was found from third days of observation period.
On basis of body weight and body weight gain data, there was no notable test item effect observable in the exposed animals.
Gross pathology:
A single four hours nose-only exposure of reaction mass of 2,2'-oxybisbutane (DSBE), DIPE, SBA and 2-methylpropan-2-ol (TBA) to CRL: (WI) BR rat followed by a 14-day observation period at a dose level 21.0 mg/L was not associated with any test item-related macroscopic findings.

Any other information on results incl. tables

Test atmosphere concentrations

The test atmosphere concentration was sampled five times during the exposure. The actual concentration of the test item was calculated as the average of the concentrations obtained from gravimetry of the samples. The test item appeared as vapour, however, a small amount of aerosol was also detected in the cascade impactor. The aerosol particles presented approximately 0.04 % of the total mass of the test item in the test atmosphere.

Table 1

Mean Achieved Concentration

Standard Deviation

Aerosol

Concentration

Nominal concentration

21.0 mg/L

2.21 mg/L

9.2 µg/L

26.1 mg/L

Individual Body weight data:

 

Sex

 

Animal Number

Body weight (g)

Weekly Increase (g)

Day 0

Day 1

Day 3

Day 7

Day 14

Day 16

Week 1

Week 2

 

Male

5877

285

280

290

321

370

387

36

49

5883

278

275

289

312

356

366

34

44

5888

306

305

321

346

380

396

40

34

5889

292

292

313

335

370

380

43

35

5890

280

274

289

308

341

349

28

33

Mean:

 

288.2

285.2

300.4

324.4

363.4

375.6

36.2

39.0

*Difference in %

 

 

-1.0

5.3

8.0

12.0

3.4

12.6

12.0

Female

5905

206

204

217

231

248

255

25

17

5912

200

200

210

222

245

250

22

25

5914

213

223

233

248

276

272

35

28

5917

214

219

221

229

255

258

15

26

5918

196

191

201

211

227

224

15

16

Mean:

 

205.8

207.4

216.4

228.2

250.2

251.8

22.4

22.4

*Difference in %

 

 

0.7

4.3

5.5

9.6

0.6

11.0

9.6

Applicant's summary and conclusion

Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
Under the experimental conditions of this study, no deaths occurred in a group of ten rats exposed for four hours to the atmosphere containing the test item at the mean achieved concentration of 21.0 mg/L. The acute inhalation median lethal concentration (4hr LC50) of test item reaction mass of 2,2'-oxybisbutane (DSBE), DIPE, SBA and 2-methylpropan-2-ol (TBA), in Wistar Crl:(WI) BR rats, was therefore considered to be greater than 21.0 mg/L.
The clinical signs represented the decreased activity and general reaction, tremor, incoordination and sleeping.
Consequently, reaction mass of 2,2'-oxybisbutane (DSBE), DIPE, SBA and 2-methylpropan-2-ol (TBA) must not be classified for acute toxicity-inhalation.
Executive summary:

Acute inhalation toxicity of test item reaction mass of 2,2'-oxybisbutane (DSBE), DIPE, SBA and 2-methylpropan-2-ol (TBA) was tested under GLP and according to OECD 403 and EU B.2 guidelines. A group of ten Wistar Crl:(WI) BR rats (five males and five females) was exposed to an atmosphere that contained the vapour and in small fraction (0.04 %) the aerosol of the test item. The animals were exposed for four hours using a nose-only exposure system, followed by a fourteen day observation period. The mean achieved atmosphere concentration was 21.0 +/- 2.21 mg/L.

In all male and female animals test item related clinical signs were found after removal from restraint on completion of the exposure. One hour after the exposure all animals slept. 

All male and female animals were symptom-free from first day of observation period. The clinical signs represented the decreased activity and general reaction, tremor, incoordination and sleeping. On basis of body weight and body weight gain data, there was no notable test item effect observable in the exposed animals.

A single four hours nose-only exposure oftest item reaction mass of 2,2'-oxybisbutane (DSBE), DIPE, SBA and 2-methylpropan-2-ol (TBA) to CRL: (WI) BR rat followed by a 14-day observation period at a dose level 21.0 mg/L was not associated with any test item-related macroscopic findings on the male and female animals.