Registration Dossier

Toxicological information

Skin sensitisation

Currently viewing:

Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2004
Report Date:
2004

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
GLP compliance:
yes
Type of study:
Buehler test

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
- Physical state: liquid
- Analytical purity: ca. 99 %
- Lot/batch No.: 030013p040
- Stability under storage and test conditions: The stability under storage conditions was confirmed by reanalysis. The stability of the test substance in
the vehicle was determined indirectly by the homogeneity analysis.
- Storage condition of test material: room temperature, protected from light, storage under air

In vivo test system

Test animals

Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Winkelmann GmbH, Borchen, Germany
- Age at study initiation: about 6 weeks
- Weight at study initiation: 355-433 g
- Housing: 5 animals/cage in stainless steel wire cages with plastic-coated grating, minimum floor area: 2000 cm2
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 15 days before the first test substance application


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24 °C
- Humidity (%): 30-70 %
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
epicutaneous, occlusive
Vehicle:
water
Concentration / amount:
undiluted test substance was used for all three inductions, test substance 75% in doubly distilled water was used for the 1st challenge, test substance
50% in doubly distilled water was used for the 2nd challenge.
Challengeopen allclose all
Route:
epicutaneous, occlusive
Vehicle:
water
Concentration / amount:
undiluted test substance was used for all three inductions, test substance 75% in doubly distilled water was used for the 1st challenge, test substance
50% in doubly distilled water was used for the 2nd challenge.
No. of animals per dose:
20 animals in the test group and 10 animals in the control group
Details on study design:
RANGE FINDING TESTS:
In accordance with the guideline a slightly irritating concentration should be used in the main test for induction whereas the maximum non-irritant
concentration should be applied for challenge.
Amount applied :
2 x 2 cm gauze patches (6 layers surgical gauze Ph . Eur. from Lohmann GmbH & Co. KG) containing 0.5 ml of the test substance / test substance
formulation were applied to the skin of the flanks under an occlusive dressing. The dressing consisted of rubberized linen patches (4 x 4 cm from
Russka), patches of Idealbinde (5 x 5 cm from Pfälzische Verbandstoff-Fabrik) and Fixomull Stretch (adhesive fleece) from Beiersdorf AG.
Duration of exposure:
- 6 hours
Site of application:
- right and left flank
Application frequency:
- one application
Number of test animals:
- 3 per test concentration
Readings:
- 1, 24 and 48 h after removal of the patches

Results:
24 hours after removal of the patch: discrete or patchy erythema was noted in two animals treated with the undiluted test substance. In one of these
animals erythema persisted until 48 hours. The animals treated with a 75% test substance preparation did not show any skin reactions 24 and 48 hours after removal of the patch. Therefore undiluted test substance was chosen for the inductions.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: three inductions were conducted
- Exposure period: 6 hours
- Test groups: treated with 0.5 ml of the undiluted test material
- Control group: The control group was not treated, since the test substance was applied undiluted and thus no vehicle was used
- Site: intact flank, always on the same application area
- Frequency of applications: one application per week; days 0, 7 and 14
- Duration: the animals were exposed to the test substance for 6 hours
- Concentrations: undiluted


B. CHALLENGE EXPOSURE
- No. of exposures: two
- Day(s) of challenge: the 1st challenge was carried out 14 days after the third induction. A 2nd challenge was performed one week after the 1st
challenge
- Exposure period: 6 hours
- Test groups: treated with 0.5 ml of the test substance solutions (75% 1st challenge, 50% 2nd challenge)
- Control group: treated with 0.5 ml of the test substance solutions (75% 1st challenge, 50% 2nd challenge)
- Site: intact flank
- Concentrations: 1st challenge with 0.5 ml of a 75% solution of the test material, 2nd challenge with 0.5 ml of a 50% solution of the test material
- Evaluation (hr after challenge): 24 and 48 hours after removal of the patch
Challenge controls:
A positive control (reliability check) with a known sensitizer is not included in this study. However, a separate study is performed twice a year in the
laboratory . The positive controls with Alpha-Hexylcinnamaldehyde, techn. 85% showed that the test system was able to detect sensitizing compounds under the laboratory conditions chosen.
Positive control substance(s):
yes
Remarks:
Alpha-Hexylcinnamaldehyde, techn. 85%

Results and discussion

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
75% test substance
No. with + reactions:
8
Total no. in group:
20
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 75% test substance. No with. + reactions: 8.0. Total no. in groups: 20.0.
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
75% test substance
No. with + reactions:
2
Total no. in group:
10
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 75% test substance. No with. + reactions: 2.0. Total no. in groups: 10.0.
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
75% test substance
No. with + reactions:
6
Total no. in group:
20
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 75% test substance. No with. + reactions: 6.0. Total no. in groups: 20.0.
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
75% test substance
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 75% test substance. No with. + reactions: 0.0. Total no. in groups: 10.0.
Reading:
rechallenge
Hours after challenge:
24
Group:
test group
Dose level:
50% test substance
No. with + reactions:
9
Total no. in group:
20
Remarks on result:
other: Reading: rechallenge. . Hours after challenge: 24.0. Group: test group. Dose level: 50% test substance. No with. + reactions: 9.0. Total no. in groups: 20.0.
Reading:
rechallenge
Hours after challenge:
24
Group:
negative control
Dose level:
50% test substance
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
other: Reading: rechallenge. . Hours after challenge: 24.0. Group: negative control. Dose level: 50% test substance. No with. + reactions: 0.0. Total no. in groups: 10.0.
Reading:
rechallenge
Hours after challenge:
48
Group:
test group
Dose level:
50% test substance
No. with + reactions:
3
Total no. in group:
20
Remarks on result:
other: Reading: rechallenge. . Hours after challenge: 48.0. Group: test group. Dose level: 50% test substance. No with. + reactions: 3.0. Total no. in groups: 20.0.
Reading:
rechallenge
Hours after challenge:
48
Group:
negative control
Dose level:
50% test substance
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
other: Reading: rechallenge. . Hours after challenge: 48.0. Group: negative control. Dose level: 50% test substance. No with. + reactions: 0.0. Total no. in groups: 10.0.

Any other information on results incl. tables

The first until third induction caused discrete or patchy to intense erythema 
and swelling in the test group animals.

After the 1st challenge discrete or patchy to moderate and confluent erythema and swelling could be observed in the test group animals. Due to discrete or patchy erythema observed in two control group animals the test substance concentration was reduced for the 2nd challenge.
The 2nd challenge caused discrete or patchy to moderate and confluent erythema in test group animals only.


The number of animals with skin findings after the 1st challenge and 2nd challenge is summarized in the following table:

1st challenge 2nd challenge
Test substance Test substance 75% in doubly 50% in doubly distilled water distilled water
24 h 48 h Total 24 h 48 h Total
Control group 2/10 0/10 2/10 0/10 0/10 0/10
Test group 8/20 6/20 9/20 9/20 3/20 9/20

x/y: number of positive reactions/number of animals tested (reading at 24 h and/or 48 h after the removal of the patch)

Applicant's summary and conclusion

Interpretation of results:
sensitising
Remarks:
Migrated information
Conclusions:
Based on the results of this study and applying the evaluation criteria (at least 15% of the test animals exhibit skin reactions) it was concluded that
the test substance has a sensitizing effect on the skin of the guinea pig in the BUEHLER Test under the test conditions chosen.
Executive summary:

"confidentials substance name" was tested for its sensitizing effect on the skin of the guinea pig in the BUEHLER Test based on the method of BUEHLER, E .V. (1965).

The test substance concentrations for the main test were selected based on the results of the pretest. All inductions were performed with the undiluted test substance, for the 1 st challenge a 75% and for the 2 nd challenge a 50% test substance preparation in doubly

distilled water was chosen. The study was performed using 1 control group and 1 test group. The inductions were performed on days 0, 7 and 14 . Two challenges were carried out 14 and 21 days after the last induction. The first until third induction caused discrete or patchy to intense erythema and swelling in the test group animals . After the 1 st challenge discrete or patchy to moderate and confluent erythema and swelling could be obsenred in the test group animals . Due to discrete or patchy erythema observed in two control group animals the test substance concentration was reduced for the 2 nd challenge. The 2nd challenge caused discrete or patchy to moderate and confluent erythema in test group animals only. The number of animals with skin findings after the 1 st challenge and 2nd challenge is summarized in the following table :

1 st challenge 2nd challenge

Control group: 2/10 0/10

Test group: 9/20 9/20

Based on the results of this study it was concluded that "confidential substance name" has a sensitizing effect on the

skin of the guinea pig in the BUEHLER Test under the test conditions chosen.