Registration Dossier

Toxicological information

Acute Toxicity: dermal

Currently viewing:

Administrative data

Endpoint:
acute toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Study period:
6 Jan - 9 March 1993
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1993
Report Date:
1993

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
EPA OTS 798.1100 (Acute Dermal Toxicity)
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
Deviations:
no
GLP compliance:
yes
Test type:
standard acute method
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Specific details on test material used for the study:
A sample of isobutanol, Lot No. TS3370114, CAS No. 78-83-1, was used. The test substance was a colorless, transparent, low viscosity liquid.
Gas chromatography-mass spectrometry (GC/MS) and nuclear magnetic resonance spectroscopy (NMR) techniques were independently used to confirm the sample's identity. Sample purity, measured by capillary GC, is ~ 99.9%.

Test animals

Species:
rabbit
Strain:
New Zealand White
Sex:
male/female
Details on test animals and environmental conditions:
Male and female New Zealand White rabbits were received from Hazleton Research Products, Inc. (Denver, PA). The strain and species were selected because of their availability and existing historical data. Rabbits were ordered to be between 2.0 and 2.3 kg (designated by the supplier to be approximately 12 to 14 weeks of age). The females were nulliparous and nonpregnant.

Periodically, a Clinical Veterinarian examined rabbits for any signs of health deficiencies. Within 1 or 2 days of receipt, all animals were assigned a unique number which was marked on the animal cage card. The rabbit number was also marked in indelible ink on 1 ear at the time of dosing.

The rabbits were housed individually in cages with wire floors (approximately 61.0 x 46.0 x 36.0 cm.). DACBQ (Deotized Animal Cage Board; Shepherd Specialty Papers, Inc.) was placed under each cage and changed regularly. An automatic timer was set to provide fluorescent lighting for a 12-hour photoperiod (approximately 0500 to 1700 hours for the light phase). Temperature and relative humidity were recorded (Cole-Parmer Hygrothermograph Seven-Day Continuous Recorder, Model No. 8368-00, Cole-Parmer Instrument Co., Chicago, IL). Temperature was routinely maintained at 61-70°F during the test period; relative humidity was routinely maintained at 40-70%. Any minor exceptions to these specified ranges were noted in the raw data.

Tap water (Municipal Authority of Westmoreland County, Greensburg, PA) was available ad libitum (except during dosing) and was delivered by an automatic watering system with demand control valves mounted on each rack. Water analyses were provided by the supplier, Halliburton NUS Environmental Laboratories, Materials Engineering & Testing Company, and Lancaster Laboratories, Inc. at regular intervals. EPA standards for maximum levels of contaminants were not exceeded. As available, water analysis reports were reviewed by the Study Director. AGWAYe PROLABe Animal Diet High Fiber Rabbit (Agway Inc.) was available ad libitum except during the actual dosing period. No analyses of chemical composition and possible contaminants of the feed were conducted by the supplier.

Animal Acclimation
The animals were acclimated for at least 5 days before dosing. Detailed clinical observations were conducted twice, at the time of receipt and during animal identification and/or dosing. In addition, rabbits were examined and weighed twice prior to dosing. Cage-side observations and mortality checks were conducted at least once daily. Animals considered unacceptable for the study, based on the clinical signs or body weights (rabbits), were rejected for use on this study.

Study Organization
The animals were weighed and inspected for health on the day of the test. Only those exhibiting a healthy state were used. Healthy animals appeared alert, active and well groomed, with no evidence of discharge, diarrhea, breathing difficulties or locomotor abnormalities. A BRRC veterinarian was available for consultation regarding any animal health concerns. Animals were randomly assigned to cages and were designated for dosing according to need and availability.

Administration / exposure

Type of coverage:
occlusive
Vehicle:
unchanged (no vehicle)
Details on dermal exposure:
The fur was removed from the entire trunk of each rabbit using veterinary clippers at least 1 day before application of the test substance. As necessary, the rabbit skin was carefully trimmed (to remove excess regrowth of fur) up to the day before dosing. Only animals with an intact and normal epidermis were used in the study. A double layer of gauze sheeting was wrapped around the trunk and secured with adhesive tape. Polyethylene sheeting was then wrapped around the trunk over the gauze. To secure the polyethylene, plastic ties or rubber bands were added (at the ends of the trunk). The test substance had a tendency to adhere to the inside of the syringe during dosing causing the plunger to stick. Therefore, in order to minimize the potential for exposure by spraying, the undiluted test substance was applied under the plastic wraps for most animals, covering as large a skin area as possible. The area of skin covered/dose level could not be measured except for 1 rabbit at 1.0 g/kg for which the dose was applied directly to the skin prior to wrapping. The amount of test substance applied was recorded for each animal. The sheeting was then protected from removal or tearing by wrapping the rabbit trunk with VETRAP" bandaging tape. The ends of the VETRAP" were secured. After the 24-hour contact period, all coverings were removed.

Duration of exposure:
24 hours
Doses:
Probe study
8000 mg/kg (female)

Definitive study
1000, 2000 and 4000 mg/kg (females)
2000 mg/kg (males)
No. of animals per sex per dose:
Probe study
1 female

Definitive study
5 females/dose
3 males/dose
Control animals:
no
Details on study design:
In the definitive percutaneous toxicity test, 5 female rabbits were included on each of several dose levels in order to determine an LD50. Three male rabbits were included on an intermediate dose level for comparison. One female rabbit was used for preliminary percutaneous toxicity testing. For individual animals, the dose volume was adjusted according to body weight. Treated rabbits wera observed frequently for signs of toxic effect on the first day of the test and twice a day thereafter (except on weekends or holidays when they were examined for death alone). Weights were recorded on the day of dosing and at 7 and 14 days after dosing or at death.

After 14 days, all survivors were sacrificed by ear vein injection using Euthanasia-6 Solution (Veterinarian Laboratories Inc., Lenexa, KS). Necropsies were performed on all animals that died or were sacrificed. The following tissues (unless excessively autolyzed) were collected and retained in 10% neutral buffered formalin: kidneys, urinary bladder, liver, sciatic nerve and spleen. Because of possible lung damage as based on clinical signs, these tissues were also saved from selected animals.
Statistics:
LD50 value, along with evaluations of 95% confidence limits, were calculated by the moving average method (Thompson, 1947). Estimates of the slope will use the method of Weil (1983).

References
Thompson, W. R. (1947). Use of moving averages and interpolation to estimate median effective dose. Bact. Rev. 2, 115-145.
Weil, C. (5. (1983). Economical LD50 and slope determination. Druq Chem. Toxicol 595-603.

Results and discussion

Effect levelsopen allclose all
Sex:
female
Dose descriptor:
LD50
Effect level:
2 460 mg/kg bw
Based on:
test mat.
95% CL:
1 790 - 3 390
Sex:
male
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Remarks on result:
other: 0 of 3 died at 2000 mg/kg
Mortality:
One rabbit was dosed with 8.0 g/kg of isobutanol in preliminary percutaneous toxicity testing (24-hour occluded contact) and died.

Of the 5 female rabbits/group dosed with 1000, 2000 or 4000 mg/kg, there were 0, 1 and 5 animals that died, respectively. One animal from the 2000 and the 4000 mg/kg groups died on the after exposure. All other deaths occurred on the day of exposure. The earliest death occurred 3 hours after test material was applied to the skin.

None of 3 male rabbits died following application of 2000 mg/kg.
Clinical signs:
Dermal reactions included erythema, edema, necrosis, ecchymoses (on 2), fissuring, ulceration (on l), desquamation, scabs and alopecia. Signs of
toxicity observed included sluggishness, lacrimation (in l), transient tremors (in I), prostration, an unsteady gait (in l), abnormal breathing (slow and/or labored), red eyes (conjunctivae, iris and/or nictitating membrane) and wetness of the periurogenital fur (of 1). For 1 to 2 days, 1 rabbit held its head abnormally low with its eyes directed upward; this animal eventually returned to normal.
Body weight:
Several animals exhibited a weight loss by 7 days, but most recovered by 14 days.
Gross pathology:
Gross pathologic evaluation of animals that died revealed red patches or areas on the lungs, dark red lungs (in l), discolored and/or mottled livers (tan or darkened), gas-filled (characterized by bubbles) intestines (in 2), darkened spleens (in 2), dark red foci on 1 spleen, enlarged adrenals (in l), kidneys with a pitted surface (in 1) and a trace amount of blood in the urine of 1 (positive by HEMASTIX@ Reagent Strips). Necropsy of survivors revealed red to dark red patches or areas on the lungs (in 2 ) , gas-filled intestines (in l), 1 mottled dark maroon and light tan spleen, kidneys with a pitted surface (in 1) and tan kidneys (in 2).
Other findings:
No additional information available.

Any other information on results incl. tables

No additional information available.

Applicant's summary and conclusion

Interpretation of results:
Category 5 based on GHS criteria
Conclusions:
The dermal LD50 in male and female rabbits is >2000 and 2460 mg/kg, respectively.
Executive summary:

The acute dermal toxicity was examined. The dermal LD50 in male and female rabbits is >2000 and 2460 mg/kg, respectively.