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Toxicological information

Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1995-10-05 to 1996-01-23
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
test procedure in accordance with national standard methods with acceptable restrictions

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1996
Report Date:
1996

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Principles of method if other than guideline:
NA
GLP compliance:
yes
Type of study:
Buehler test
Justification for non-LLNA method:
The study was conducted in 1996 when the LLNA was not yet an established testing approach.

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid
Details on test material:
Purity batch
> 99.0 % 7.32% active oxygenBatch 247408973
99.27 % 9407226339; 247408999
99.6 % 0410105225413
> 97 % 4289070300; 0419101226056
99.1 % 11600 0010-108
99.3 % 247413988
99.4 % 7.35 % active oxygen Batch 247409800
98.9 % 0707518256
> 99.1 % 247412384
98.5 % 0707603153
99.0 % 1703465433
99.5 % 150726; 247413309
99.3 % 000055055

In vivo test system

Test animals

Species:
guinea pig
Strain:
Hartley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Sprague Dawley, Inc., Haslett, MI.
- Age at study initiation: Young adult
- Housing: individually in suspended stainless steel cages.
- Diet: ad libitum, Purina Certified Pig Chow
- Water: ad libitum, municipal tap water
- Acclimation period: for a minimum of five years


ENVIRONMENTAL CONDITIONS
- Temperature: 17.8 - 25.6 °C
- Humidity: 28 - 73 %
- Air changes (per hr): 10 - 15 air changes/hour
- Photoperiod (hrs dark / hrs light): 12-hour light/ 12-hour dark

Study design: in vivo (non-LLNA)

Induction
Route:
epicutaneous, occlusive
Vehicle:
other: mineral oil
Concentration / amount:
25 % w/v t-Butyl Peroctoate
Challenge
Route:
epicutaneous, occlusive
Vehicle:
other: mineral oil
Concentration / amount:
25 % w/v t-Butyl Peroctoate
No. of animals per dose:
10 male, 10 female
Details on study design:
RANGE FINDING TESTS:
Dosing: On the day prior to dose administration, four topical range-finding guinea pigs were weighed and the hair removed from the right and left side of the animals with a small animal clipper. Care was taken to avoid abrading the skin during clipping procedures. On the following day, four concentrations of the test article were prepared and each concentration was applied to the clipped area of each topical range-finding animal.
Following patch application, the trunk of each animal was wrapped with elastic wrap which was secured with adhesive tape to prevent removal of the patches and the animal was returned to its cage. Approximately six hours after patch application, the elastic wrap, tape and patches were removed. The test sites were then wiped with gauze moistened in distilled water to remove test article residue and the animals returned to their cages.


MAIN STUDY
A. INDUCTION EXPOSURE
On the day prior to the first induction dose administration, the hair was removed from the left side of the 20 test animals with a small animal clipper. Care was taken to avoid abrading the skin during clipping procedures. On the following day (day 0), the appropriate concentration of the test article was prepared and applied to the clipped area of the animals.
Following patch application, the trunk of each animal was wrapped with elastic wrap which was secured with adhesive tape to prevent removal of the patch and the animal was returned to its cage. Approximately 6 hours after patch application, the elastic wrap, tape and patches were removed. The test sites were then wiped with gauze moistened in distilled water to remove test article residue and the animals returned to their cages.

Dermal Observations: The test sites were graded for dermal irritation at approximately 24 and 48 hours following patch application using the Dermal Grading System.

The induction procedure was repeated on study day 7 and on study day 14 so that a total of three consecutive induction exposures were made to the 20 test animals.


B. CHALLENGE EXPOSURE
On the day prior to challenge dose administration, the hair was removed from the right side of the 19 test and 10 control animals with a small animal clipper. Care was taken to avoid abrading the skin during clipping procedures. On the following day (day 28), the appropriate concentration of the test article was prepared and applied to a naive site within the clipped area of the animals.
Following patch application, the trunk of each animal was wrapped with elastic wrap which was secured with adhesive tape to prevent removal of the patch and the animal was returned to its cage. Approximately 6 hours after patch application, the elastic wrap, tape and patches were removed. The test sites were then wiped with gauze moistened in distilled water to remove test article residue and the animals returned to their cages.

Dermal Observations: The test sites were graded for dermal irritation at approximately 24 and 48 hours following patch application using the Dermal Grading System.


C. RECHALLENGE
A rechallenge was conducted in order to clarify the results of the challenge phase.
On the day prior to rechallenge dose administration, the hair was removed from the left side of the 19 test and 10 control animals with a small animal clipper. Care was taken to avoid abrading the skin during clipping procedures. On the following day (day 35), the appropriate concentration of the test article was prepared and applied to a naive site within the clipped area of the animals.
Following patch application, the trunk of each animal was wrapped with elastic wrap which was secured with adhesive tape to prevent removal of the patch and the animal was returned to its cage. Approximately 6 hours after patch application, the elastic wrap, tape and patches were removed. The test sites were then wiped with gauze moistened in distilled water to remove test article residue and the animals returned to their cages.

Dermal Observations: The test sites were graded for dermal irritation at approximately 24 and 48 hours following patch application using the Dermal Grading System.
Challenge controls:
see above at "Details on study design"
Positive control substance(s):
yes
Remarks:
Hexylcinnamaldehyde

Results and discussion

Positive control results:
Using Hexylcinnamaldehyde as a mild to moderate positive control, Springborn Laboratories, Inc., Spencerville, Ohio, has compiled historical control data for contact sensitization to this agent utilizing the test system described herein (Modified Buehler Design). As indicated, 95% of animals induced with Hexylcinnamaldehyde elicited a contact sensitization response following challenge with Hexylcinnamaldehyde, thereby demonstrating the susceptibility of the test system to this sensitizing agent.

In vivo (non-LLNA)

Resultsopen allclose all
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
5 % t-Butyl Peroctoate
No. with + reactions:
9
Total no. in group:
19
Clinical observations:
Discrete or patchy erythema
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
5 % t-Butyl Peroctoate
No. with + reactions:
3
Total no. in group:
19
Clinical observations:
Discrete or patchy erythema
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
other: challenge control
Dose level:
5 % t-Butyl Peroctoate
No. with + reactions:
3
Total no. in group:
10
Clinical observations:
Discrete or patchy erythema
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
other: challenge control
Dose level:
5 % t-Butyl Peroctoate
No. with + reactions:
2
Total no. in group:
10
Clinical observations:
Discrete or patchy erythema
Key result
Reading:
rechallenge
Hours after challenge:
24
Group:
test group
Dose level:
2 % t-Butyl Peroctoate
No. with + reactions:
5
Total no. in group:
19
Clinical observations:
Discrete or patchy erythema
Key result
Reading:
rechallenge
Hours after challenge:
48
Group:
test group
Dose level:
2 % t-Butyl Peroctoate
No. with + reactions:
0
Total no. in group:
19
Clinical observations:
none
Key result
Reading:
rechallenge
Hours after challenge:
24
Group:
other: challenge control
Dose level:
2 % t-Butyl Peroctoate
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
none
Key result
Reading:
rechallenge
Hours after challenge:
48
Group:
other: challenge control
Dose level:
2 % t-Butyl Peroctoate
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
none
Group:
positive control
Remarks on result:
not measured/tested
Key result
Group:
negative control
Remarks on result:
not measured/tested

Applicant's summary and conclusion

Interpretation of results:
Category 1 (skin sensitising) based on GHS criteria
Conclusions:
Due to the irritation noted in both the test and challenge control animals, challenge with tert.-Butylperoxy- 2-ethylhexanoat was considered to be ambiguous. Following rechallenge with 2% tert.-Butylperoxy- 2-ethylhexanoat, slight evidence of sensitization was observed in 5/19 test animals at the 24 hour scoring interval; however, the dermal responses did not persist to the 48 hour scoring interval. Therefore, based on the results of this study, tert.-Butylperoxy- 2-ethylhexanoat is considered to be a possible weak sensitizer in guinea pigs. The results of the Hexylcinnamaldehyde Historical Control demonstrates that the test design utilized by Springborn Laboratories, Inc., could detect potential mild to moderate contact sensitizers.
Executive summary:

The dermal sensitization potential of tert.-Butylperoxy- 2-ethylhexanoat was evaluated in Hartley-derived albino guinea pigs. Ten male and ten female guinea pigs were topically treated with 25 % w/v tert.-Butylperoxy- 2-ethylhexanoat in mineral oil, once per week, for 3 consecutive weeks. Following a two week rest period, a challenge was performed whereby the 19 test and 10 previously untreated (naive) challenge control guinea pigs were topically treated with 5 % w/v tert.-Butylperoxy- 2-ethylhexanoat in mineral oil. Challenge responses in the test animals were compared to those of the challenge control animals. Following a one week rest period, a rechallenge was performed whereby the 19 test and the 10 challenge control guinea pigs were topically treated with 2% w/v tert.-Butylperoxy- 2-ethylhexanoat in mineral oil. Rechallenge responses in the test animals were compared to those of the challenge control animals.

Following induction 1 with 25 % w/v tert.-Butylperoxy- 2-ethylhexanoat in mineral oil, mild irritation was noted in the test animals. The dermal irritation increased slightly at induction 2 and 3. Following challenge with 5 % w/v tert-butyl 2-ethylperoxyhexanoate in mineral oil, dermal scores of 1 were noted in 9/19 test animals and 3/10 challenge control animals at the 48 hour scoring interval and in 3/19 test animals and 2/10 challenge control animals at the 48 hour scoring interval. Dermal scores of 0 were noted in the remaining test and challenge control animals. Group mean dermal scores were noted to be similar in the test animals as compared to the challenge control animals. Following rechallenge with 2 % w/v tert.-Butylperoxy- 2-ethylhexanoat in mineral oil, dermal scores of 1 were noted in 5/19 test animals at the 24 hour scoring interval. Dermal scores of 0 were noted in the remaining test and all challenge control animals. Group mean dermal scores were noted to be slightly higher in the test animals as compared to the challenge control animals.

One test animal (1431/F) was found dead on study day 27. Gross necropsy observations included dark red mandibular and axillary lymph nodes, an adhesion in the thoracic cavity, mottled lungs, mottled liver, enlarged spleen and congested meningeal vessels in the brain. The majority of sensitization study animals gained weight during the test period and generally appeared in good health.

Due to the irritation noted in both the test and challenge control animals, challenge with tert.-Butylperoxy- 2-ethylhexanoat was considered to be ambiguous. Following rechallenge with 2% tert-butyl 2-ethylperoxyhexanoate, slight evidence of sensitization was observed in 5/19 test animals at the 24 hour scoring interval; however, the dermal responses did not persist to the 48 hour scoring interval. Therefore, based on the results of this study, tert.-Butylperoxy- 2-ethylhexanoat is considered to be a possible weak sensitizer in guinea pigs.