Registration Dossier

Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
from 1994-09-06 to 1994-10-06
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: This study followed the procedures indicated by internationally accepted guidelines and recommendations as Commission Regulation (EC) No 440/2008 (B.6: Skin Sensitisation).
Cross-reference
Reason / purpose:
reference to other study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1994
Report Date:
1994

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
EU Method B.6 (Skin Sensitisation)
Version / remarks:
directive 84/449/EEC
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
no
Principles of method if other than guideline:
NA
GLP compliance:
yes
Type of study:
guinea pig maximisation test

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid: viscous

In vivo test system

Test animals

Species:
guinea pig
Strain:
other: Himalayan albino
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: BRL Ltd., Basel, Switzerland
- Age at study initiation: Approx. 6 weeks
- Weight at study initiation: 311 - 437 grams
- Housing: group housing of 2 animals per labelled metal cage with wire-mesh floors and equipped with an automatic drinking system (ITL, Bergen, The Netherlands)
- Diet: free access to standard guinea pig diet, including ascorbic acid (1600 mg/kg); LC 23-B, pellet diameter 4mm (Hope farms, Woerden, The Netherlands).
- Water: free access to tap water, diluted with decalcified water
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature: 21 °C
- Humidity: 50 % rel. humidity
- Air changes: approximately 15 air changes per hour
- Photoperiod: 12 hours artificial fluorescent light and 12 hours per day

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal and epicutaneous
Vehicle:
other: distilled water and physiological saline
Concentration / amount:
Concentrations used at each stage of induction:
Intradermal injection: 5 % (w/w) in physiological saline
epidermal application: 100 %

Concentrations used at each stage of challenge: 100, 50 and 25 % (w/w) in distilled water
Challengeopen allclose all
Route:
epicutaneous, semiocclusive
Vehicle:
other: distilled water and physiological saline
Concentration / amount:
Concentrations used at each stage of induction:
Intradermal injection: 5 % (w/w) in physiological saline
epidermal application: 100 %

Concentrations used at each stage of challenge: 100, 50 and 25 % (w/w) in distilled water
No. of animals per dose:
Preliminary study: 5 females
Experimental group; 10 females
Control group: 5 females
Details on study design:
Main study:
Intradermal injections:
Day 1: The experimental animals were intradermally injected with three pairs of injections (0.1 mL/site) in the clipped scapular region as follows:
A) Freunds` Complete Adjuvant 50:50 with water for injection.
B) The test substance at a 5 % (w/w) concentration in physiological saline.
C) The test substance, at twice the concentration used in (B), emulsified in a 50:50 mixture of Freunds` Complete Adjuvant.

Epidermal applications:
Day 7: The clipped scapular area was rubbed with 10 % sodium-dodecyl-sulfate in petrolatum using a spatula. This concentration of SDS enhances sensitisation by provoking a mild inflammatory reaction.
Day 8: The clipped area between the injection sites was treated with 0.5 mL of the undiluted test substance using a Scotchpak-non-woven patch (2 x 4 cm) mounted on Micropore tape and secured with Coban elastic bandage. After 48 hours, the dressing was removed and residual test substance removed using a tissue moistened with tap water. The skin reaction was assessed immediately after bandage removal according to the grading system described above.
Challenge controls:
Day 22: All animals were treated epidermally with 0.05 mL of each of the following test substance concentrations 100 %, 50 % and 25 %, w/w in distilled water and with the vehicle on the clipped and shaved flank, using Square chambers attached to Micropore tape and secured with Coban elastic bandage. After 24 hours, the dressing was removed and residual test substance removed using a tissue moistened with tap water. The treated sites were assessed for redness and swelling 24 and 48 hours after removal of dressings, using the numerical grading system described below. After the first reading, the test sites were shaved with an electric razor.

At the end of the study period all animals were killed by oxygen/carbon dioxide asphyxiation.
Positive control substance(s):
yes
Remarks:
alpha-hexylcinnamicaldehyde

Study design: in vivo (LLNA)

Concentration:
NA
No. of animals per dose:
NA
Details on study design:
NA
Statistics:
NA

Results and discussion

Positive control results:
A positive control experiment is performed once every six months as a sensitivity check of the system.
Taking into account the intensity of the skin reactions noted in the control animals, it was considered that positive reactions, indicative of sensitisation, were observed in all experimental animals in response to one or more of the test substance concentrations. These results leads to the maximum sensitisation rate of 100 %, which indicates that alpha-hexylcinnamicaldehyde has extreme sensitizing properties in this test applying the rating of allergenicity described by Kligman A.M. (1966).
From these results, it can be concluded that the Himalayan strain of guinea pig is an appropriate animal model for the performance of studies designed to evaluate the sensitising potential of a substance.

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
100 %
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
none
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 100 %. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: none.
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
50 %
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
none
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 50 %. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: none.
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
25 %
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
none
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 25 %. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: none.
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
100 %
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
none
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 100 %. No with. + reactions: 0.0. Total no. in groups: 5.0. Clinical observations: none.
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
50 %
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
none
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 50 %. No with. + reactions: 0.0. Total no. in groups: 5.0. Clinical observations: none.
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
25 %
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
none
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 25 %. No with. + reactions: 0.0. Total no. in groups: 5.0. Clinical observations: none.
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
100 %
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
none
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 100 %. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: none.
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
50 %
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
none
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 50 %. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: none.
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
25 %
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
none
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 25 %. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: none.
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
100 %
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
none
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 100 %. No with. + reactions: 0.0. Total no. in groups: 5.0. Clinical observations: none.
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
50 %
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
none
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 50 %. No with. + reactions: 0.0. Total no. in groups: 5.0. Clinical observations: none.
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
25 %
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
none
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 25 %. No with. + reactions: 0.0. Total no. in groups: 5.0. Clinical observations: none.

Applicant's summary and conclusion

Interpretation of results:
not sensitising
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
The test item was considered to have no skin sensitizating properties in the Maximization test with guinea pigs.
Executive summary:

The skin sensitizating properties of the test item were evaluated in a Maximization test with guinea pigs according to OECD guideline 406/EU method B.6. A test group of 10 albino guinea pigs and a control group of 5 animals were investigated for signs of skin hypersensitivity after intradermal and epidermal exposure.

After a preliminary study for assessment of the slightly irritating and the non-irritating test substance concentrations, a main study was performed with the selected test substance concentrations. Ten experimental animals were intradermally injected with a 5% concentration and epidermally exposed to the undiluted test substance, while five control animals were similarly treated without test substance and epidermally with a dry patch. Immediately after the epidermal exposure, the skin irritation was scored. Two weeks after the epidermal application all animals were challenged with test substance concentrations of 100%, 50% and 25% and the vehicle (distilled water). The challenge reactions were assessed 24 and 48 hours after bandage removal.

The epidermal exposure of the test item in the induction phase resulted in slight skin irritation in the majority of experimental animals. The epidermal exposure of the test item in the challenge phase resulted in no positive reactions in response to the test substance concentration tested.

Under the conditions used in this study, exposure of the test item induced no sensitisation.