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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
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Justification for type of information:
Data is from publication.

Data source

Reference
Reference Type:
publication
Title:
Studies of new short-period method for delayed contact hypersensitivity assay in the guinea pig (I). Development and comparsi on with other methods
Author:
Ryuichi kashma,Yumiko oyake,Joshin okada and yuzo ikeda
Year:
1993
Bibliographic source:
Contact Dermatitis,1993.vol.28 p 235-242

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: As mention below
Principles of method if other than guideline:
The test chemical was assessed for its possible contact allergenic potential by AP2 test in female guinea pigs.
GLP compliance:
not specified
Type of study:
other: AP2 test
Justification for non-LLNA method:
Not specified

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
SMILES:BrC=Cc1ccccc1

In vivo test system

Test animals

Species:
guinea pig
Strain:
Hartley
Sex:
female
Details on test animals and environmental conditions:
Details on test animal
TEST ANIMALS
- Source: Japan SLC
- Age at study initiation:6 weeks old
- Weight at study initiation:250 g
- Housing: Groups of 5 animals were housed in aluminium cages .
- Diet (e.g. ad libitum): Solid diet (Labo G standard. ihon ousan) ad libitum.
- Water (e.g. ad libitum): tap water sterilized by UV available ad ibitum.
- Acclimation period: 2 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23 ± 2 C
- Humidity (%):55 ± 5%.
- Air changes (per hr): Ventilation of15 cycles /hour
- Photoperiod (hrs dark / hrs light): Animal rooms were maintained on a 12-h light-dark cycle with light on at 7:00 and off at 19:00.

Study design: in vivo (non-LLNA)

Induction
Route:
intradermal and epicutaneous
Vehicle:
other: Ethanol
Concentration / amount:
10%
Day(s)/duration:
24 hour
Adequacy of induction:
highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically
Challengeopen allclose all
Route:
epicutaneous, open
Vehicle:
other: Ethanol
Concentration / amount:
0.02 ml of 30%,10% and 3% ( for 1st and 2nd challenge )
Day(s)/duration:
72 hours
Adequacy of challenge:
highest non-irritant concentration
No.:
#3
Route:
epicutaneous, open
Vehicle:
other: ethanol
Remarks:
3rd challenge
Concentration / amount:
0.1 ml at the concentration 0.1%,0.03%,0.01%
Day(s)/duration:
72 hours
Adequacy of challenge:
highest non-irritant concentration
No. of animals per dose:
Test group - 10 animals
Control group-5 animals
Details on study design:
Details on study design
RANGE FINDING TESTS: Before the main study the concentration of test chemical causing slight erythema was determined by preliminary primary skin irritation testing.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures; 1
- Exposure period:No data available
- Test groups: 10
- Control - 5
- Site: No data available .
- Frequency of applications: - No data available .
-Duration: No data available .
- Concentrations: 10%
OTHER-On the first sensitization day,an area of 5 x I 0 cm on the scapular region was clipped with an electric clipper and shaved with an electric shaver 0.1 ml of FCA (undiluted) was injected intraderrnally (i.d.) nto the scapular region at a small inner site on each side of the 2 x 4 cm area. A second FCA (undiluted) i.d. injection was administered around the inside or the first injection site.

B. CHALLENGE EXPOSURE
- No. of exposures:3
- Day(s) of challenge:1st challenge=day 11,2nd challenge=day 32,3rd challenge= day 39
- Exposure period: 3rd challenge =24 hour
- Test groups:10
- Control group: 5
- Site- The flank skin was clipped and shaved. The 1st and second challenge were carried out by non -occlusive topical application 0.02 ml of 30%,10% and 3%. The 3rd challenge was carried out by occlusive topical application of 0.1 ml at the concentration 0.1%,0.03%,0.01% .
- Concentrations: 0.1 ml at the concentration 0.1%,0.03%,0.01% .
- Evaluation (hr after challenge): 24,48,72Hour

Other- Before challenge. The Highest concetraiion of test chemical causing no irritation was determined by preliminary primary skin irritation testing. For this test. animals were treated with 0.1 ml of FCA (undiluted) i.d. injection into 2 sites in the scapular region 7 days before the test. After non-occlusive topical application challenge, animal were placed individually or I to 2 h (long enough for drying) in 5 series of cages allowing no interference.
Challenge controls:
Control group animals were only treated with FCA (undiluted) i.d. injections.
Positive control substance(s):
yes
Remarks:
Benzyl alcohol

Results and discussion

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
other: 1st challenge
Hours after challenge:
24
Group:
test group
Dose level:
0.02 ml of 30%,
No. with + reactions:
10
Total no. in group:
10
Clinical observations:
Senstization reaction was observed
Remarks on result:
positive indication of skin sensitisation
Reading:
other: 1st challenge
Hours after challenge:
48
Group:
test group
Dose level:
0.02 ml of 30%,
No. with + reactions:
10
Total no. in group:
10
Clinical observations:
Skin sensitization reaction observed.
Remarks on result:
positive indication of skin sensitisation
Reading:
other: 1st challenge
Hours after challenge:
72
Group:
test group
Dose level:
0.02 ml of 30%,
No. with + reactions:
10
Total no. in group:
10
Clinical observations:
Skin sensitization reaction observed .
Remarks on result:
positive indication of skin sensitisation
Reading:
other: 1st challenge
Hours after challenge:
24
Group:
test group
Dose level:
0.02 ml of 10%
No. with + reactions:
2
Total no. in group:
10
Clinical observations:
No skin sensitization effect were observed.
Remarks on result:
no indication of skin sensitisation
Reading:
other: 1st challenge
Hours after challenge:
48
Group:
test group
Dose level:
0.02 ml of 10%
No. with + reactions:
6
Total no. in group:
10
Clinical observations:
No skin sensitization effect were observed.
Remarks on result:
no indication of skin sensitisation
Reading:
other: 1st challenge
Hours after challenge:
72
Group:
test group
Dose level:
0.02 ml of 10%
No. with + reactions:
6
Total no. in group:
10
Clinical observations:
No skin sensitiztion effect were observed.
Remarks on result:
no indication of skin sensitisation
Reading:
other: 1st challenge
Hours after challenge:
24
Group:
test group
Dose level:
0.02 ml of 3%
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No indication of skin sensitization .
Remarks on result:
no indication of skin sensitisation
Reading:
other: 1st challenge
Hours after challenge:
48
Group:
test group
Dose level:
0.02 ml of 3%
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No skin sensitization effect were observed.
Remarks on result:
no indication of skin sensitisation
Reading:
other: 1st challenge
Hours after challenge:
72
Group:
test group
Dose level:
0.02 ml of 3%
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No skin sensitization effect were observed.
Remarks on result:
no indication of skin sensitisation
Reading:
other: 2nd challenge
Hours after challenge:
24
Group:
test group
Dose level:
0.02 ml of 30%
No. with + reactions:
10
Total no. in group:
10
Clinical observations:
Skin sensitization effect were observed.
Remarks on result:
positive indication of skin sensitisation
Reading:
other: 2nd challenge
Hours after challenge:
48
Group:
test group
Dose level:
0.02 ml of 30%
No. with + reactions:
10
Total no. in group:
10
Clinical observations:
Skin sensitization effect were observed.
Remarks on result:
no indication of skin sensitisation
Reading:
other: 2nd challenge
Hours after challenge:
72
Group:
test group
Dose level:
0.02 ml of 30%
No. with + reactions:
10
Total no. in group:
10
Clinical observations:
Skin sensitization effect were observed.
Remarks on result:
positive indication of skin sensitisation
Reading:
other: 2nd challenge
Hours after challenge:
24
Group:
test group
Dose level:
0.02 ml of 10%
No. with + reactions:
8
Total no. in group:
10
Clinical observations:
No indication of skin sensitization
Remarks on result:
no indication of skin sensitisation
Reading:
other: 2nd challenge
Hours after challenge:
48
Group:
test group
Dose level:
0.02 ml of 10%
No. with + reactions:
9
Total no. in group:
10
Clinical observations:
skin sensitization effect were observed.
Remarks on result:
positive indication of skin sensitisation
Reading:
other: 2nd challenge
Hours after challenge:
72
Group:
test group
Dose level:
0.02 ml of 10%
No. with + reactions:
6
Total no. in group:
10
Clinical observations:
No skin sensitization effect were observed
Remarks on result:
no indication of skin sensitisation
Reading:
other: 2nd challenge
Hours after challenge:
24
Group:
test group
Dose level:
0.02 ml of 3%
No. with + reactions:
2
Total no. in group:
10
Clinical observations:
No skin sensitization effect were observed.
Remarks on result:
no indication of skin sensitisation
Reading:
other: 2nd challenge
Hours after challenge:
48
Group:
test group
Dose level:
0.02 ml of 3%
No. with + reactions:
2
Total no. in group:
10
Clinical observations:
No indication of skin sensitization was observed.
Remarks on result:
no indication of skin sensitisation
Reading:
other: 2nd challenge
Hours after challenge:
72
Group:
test group
Dose level:
0.02 ml of 3%
No. with + reactions:
2
Total no. in group:
10
Clinical observations:
No indication of skin sensitization was observed.
Remarks on result:
no indication of skin sensitisation
Reading:
other: 3rd challenge
Hours after challenge:
24
Group:
test group
Dose level:
0.1 ml of 0.1(w/w)%,
No. with + reactions:
10
Total no. in group:
10
Clinical observations:
Skin sensitization effect were observed
Remarks on result:
positive indication of skin sensitisation
Reading:
other: 3rd challenge
Hours after challenge:
48
Group:
test group
Dose level:
0.1 ml of 0.1(w/w)%,
No. with + reactions:
10
Total no. in group:
10
Clinical observations:
indication of skin sensitization were observed
Remarks on result:
positive indication of skin sensitisation
Reading:
other: 3rd challenge
Hours after challenge:
72
Group:
test group
Dose level:
0.1 ml of 0.1(w/w)%,
No. with + reactions:
10
Total no. in group:
10
Clinical observations:
Skin sensitization effect were observed.
Remarks on result:
positive indication of skin sensitisation
Reading:
other: 3rd challenge
Hours after challenge:
24
Group:
test group
Dose level:
0.1 ml of 0.03%
No. with + reactions:
9
Total no. in group:
10
Clinical observations:
Skin sensitization effect were observed
Remarks on result:
positive indication of skin sensitisation
Reading:
other: 3r challenge
Hours after challenge:
48
Group:
test group
Dose level:
0.1 ml of 0.03%
No. with + reactions:
10
Total no. in group:
10
Clinical observations:
Skin sensitization effect were observed.
Remarks on result:
positive indication of skin sensitisation
Reading:
other: 3rd challenge
Hours after challenge:
72
Group:
test group
Dose level:
0.1 ml of 0.03%
No. with + reactions:
10
Total no. in group:
10
Clinical observations:
Skin sernsitization effect was observed.
Remarks on result:
positive indication of skin sensitisation
Reading:
other: 3rd challenge
Hours after challenge:
24
Group:
test group
Dose level:
0.1 ml of 0.01%
No. with + reactions:
7
Total no. in group:
10
Clinical observations:
Skin sensitization effect were observed.
Remarks on result:
positive indication of skin sensitisation
Reading:
other: 3rd challenge
Hours after challenge:
48
Group:
test group
Dose level:
0.1 ml of 0.03%
No. with + reactions:
10
Total no. in group:
10
Clinical observations:
Skin sensitization effect were observed
Remarks on result:
positive indication of skin sensitisation
Reading:
other: 3rd challenge
Hours after challenge:
72
Group:
test group
Dose level:
0.1 ml of 0.03%
No. with + reactions:
10
Total no. in group:
10
Clinical observations:
Skin sensitization effect were observed
Remarks on result:
positive indication of skin sensitisation

Any other information on results incl. tables

The reaction was evaluated at 24 ,4 8 and 72 h after challenge. According  to the following  criteria and scores in parentheses:   . 

 -no reaction (0).

 ± slight erytherna  (1)

 +apparent  erytherna  (2).           

+ apparent erythema with edema (3):

 + crust or necrosis  (4). 

Greater than a + reaction  was considered to  be positive.

 

Score after 1st and 2nd challenge

 Challenge conc.%(w/w) 2ndchallenge 24-h 48-h 72-h

Challenge conc.%(w/w)

1stchallenge

24-h

48-h

72-h

30%

10/10(2.6)

10/10(2.9)

10/10(3.1)

10%

2/10(0.9)

6/10(1.6)

6/10(1.6)

3%

0/10(0.2)

0/10(0.5)

0/10(0.6)

 

Challenge conc.%(w/w)

2ndchallenge

24-h

48-h

72-h

30%

10/10(2.7)

10/10(3.0)

10/10(3.0)

10%

8/10(1.7)

9/10(2.1)

6/10(1.6)

3%

2/10(0.7)

2/10(2.1)

0/10(1.6)

 

Challenge conc.%(w/w)

3rd challenege

24-h

48-h

72-h

0.1%

10/10(2.3)

10/10(3.0)

10/10(3.2)

0.03%

9/10(2.1)

10/10(3.0)

10/10(3.0)

0.01%

7/10(1.7)

10/10(2.9)

10/10(2.8)

 

Positive number/Total numberand mean response (score)in parenthesis.

Applicant's summary and conclusion

Interpretation of results:
other: sensitizing
Conclusions:
The test chemical was assessed for its possible contact allergenic potential by AP2 in female guinea pigs. According toAP2, the test chemical was considered to be sensitizing in guinea pigs.
Executive summary:

The test chemical was assessed for its possible contact allergenic potential by AP2 in female guinea pigs.

For this purpose induction exposure was performed as 24-h occlusive patch at the 2 x 4 cm site on the scapular region with the occlusive patch unit containing 0.2 ml of test solution at concentration of 10%. Before challenge exposure test animals were treated with 0.1 ml  of FCA (undiluted)  i.d. injection into 2 sites in the scapular region 7 days before the  test. After non-occlusive  topical  application challenge, animal  were placed  individually  or 1 to  2 h0ur  (long enough   for drying)  in 5 series of cages allowing  no interference. After a rest period of 11 and 32 and 39 days respectively 1st, 2ndand 3rdchallenge were performed .The flank skin was clipped and shaved. The 1st and 2nd challenges were carried out by non-occlusive topical application. 0.02 ml of test solution were applied to 2.0 cm diameter areas of the flank skin with a silicone stick (0 = 5.0 mm) using a test Concentrations of 30%, 10%,3%. The 3rdchallenge was performed by occlusive topical application of 0.1 ml at the concentration 0.1%, 0.03%, 0.01%.The reaction was evaluated at 24, 48 and 72 h after challenge

After third challenge, the test chemical showed positive result for the test concentration 0.1% as the evaluated score was in the range of 3.2. While for the other two concentration 0.03% and 0.01% the evaluated score for sensitization was 3.0 and 2.8 respectively. 

Thus according to AP2 test, the test chemical was observed to be sensitizing in guinea pigs at the tested concentrations.