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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:
Feb 1992 - Mar 1992
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1995
Report Date:
1995

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
May 1981
Deviations:
no
GLP compliance:
yes
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
The study was performed prior to the implementation of the LLNA method.

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Batch No.of test material: SN 9707
- Expiration date of batch: not available
- Appearance: pale yellow liquid
- Purity: > 95%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature in the dark

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: The test substance was prepared prior to each application on the day of dosing in Alembicol D.

In vivo test system

Test animals

Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: D. Hall, Newchurch, Staffordshire, England
- Females nulliparous and non-pregnant: yes
- Age at study initiation: 6 to 7 weeks
- Weight at study initiation: 294 g to 349 g
- Housing: Group housing (10 animals per group)
- Diet: A vitamin C enriched guinea pig diet FD1, ad libitum
- Water: Drinking water, ad libitum
- Acclimation period: 13 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21
- Humidity (%): 30-70
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12 (light phase from 7 am to 7 pm)

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal
Vehicle:
other: Alembicol D
Concentration / amount:
10% v/v
Adequacy of induction:
highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically
Route:
epicutaneous, occlusive
Vehicle:
other: Alembicol D
Concentration / amount:
as supplied
Day(s)/duration:
Six days after the injection for 48 h
Adequacy of induction:
highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically
Challengeopen allclose all
No.:
#1
Route:
epicutaneous, occlusive
Vehicle:
other: Alembicol D
Concentration / amount:
50% v/v
Day(s)/duration:
24 h
Adequacy of challenge:
highest non-irritant concentration
No.:
#2
Route:
epicutaneous, occlusive
Vehicle:
unchanged (no vehicle)
Concentration / amount:
as supplied
Day(s)/duration:
24 h
Adequacy of challenge:
highest non-irritant concentration
No. of animals per dose:
20
Details on study design:
RANGE FINDING TESTS:
- A preliminary study was performed in order to identify where possible (a) concentrations that will produce irritation suitable for the induction phase of the main study and (b) a maximum non-irritatn concentration by the topical route of adminsitration for the challenge phase

MAIN STUDY
A. INDUCTION EXPOSURE (INTRADERMAL)
- No. of exposures: 1
- Control group: FCA, FCA + V, V
- Site: interscapular area
- Frequency of applications: single application
- Test groups:
1. Freund's complete adjuvant was diluted with an equal volume of water for irrigation (Ph.Eur.).
2. Test substance, 10% v/v in Alembicol D.
3. Test substance, 10% v/v in a 50:50 mixture of Freund's complete adjuvant and Alembicol D.
- Duration: 0-7 d

INDUCTION EXPOSURE (TOPICAL)
- No. of exposures: 1
- Exposure period: 48 h
- Test groups: TS, TS + FCA, FCA
- Control group: FCA, FCA + V, V
- Site: interscapular area
- Frequency of applications: single application
- Day of topical induction: 6 days after the injections
- Concentrations: 10% v/v in Alembicol D and as supplied
- Duration: 0-7 d

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day of challenge: Two weeks after the topical induction
- Exposure period: 24 h
- Test groups: TS
- Control group: TS
- Site: left flank Test substance (TS), and posterior site 50% v/v Alembicol D
- Concentrations: as supplied and 50% v/v in Alembicol D.
- Evaluation (hr after challenge): 24, 48, 72

Challenge controls:
Challenge controls were treated in the same way as test groups.
Positive control substance(s):
yes
Remarks:
A periodically concurrent positive control (formalin) was used

Results and discussion

Positive control results:
The periodic positive control induced hypersensitivity in the test animals.

In vivo (non-LLNA)

Resultsopen allclose all
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
as supplied
No. with + reactions:
12
Total no. in group:
20
Clinical observations:
Erythema (12/20) and oedema (1/20)
Remarks on result:
positive indication of skin sensitisation
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
50% v/v in Alembicol D
No. with + reactions:
3
Total no. in group:
20
Clinical observations:
erythema (3/20)
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
as supplied
No. with + reactions:
18
Total no. in group:
20
Clinical observations:
erythema (18/20) and oedema (7/20)
Remarks on result:
positive indication of skin sensitisation
Reading:
other: 3rd reading
Hours after challenge:
72
Group:
test group
Dose level:
as supplied
No. with + reactions:
18
Total no. in group:
20
Clinical observations:
erythema (18/20) and oedema (7/20)
Remarks on result:
positive indication of skin sensitisation
Reading:
other: 3rd reading
Hours after challenge:
72
Group:
test group
Dose level:
50% v/v in Alembicol D
No. with + reactions:
14
Total no. in group:
20
Clinical observations:
erythema (14/20) and oedema (2/20)
Remarks on result:
positive indication of skin sensitisation

Any other information on results incl. tables

CLINICAL SIGNS:

No signs of ill health or toxicity were recorded.

BODY WEIGHT:

Body weight increases were recorded for all guinea pigs over the period of the study.

INDUCTION:

Injections:

Necrosis was recorded at sites receiving Freund's Complete Adjuvant in test and control animals. Irritation was seen in test animals at sites receiving the test substance, 10% v/v Alembicol and very slight irritation was observed in control animals receiving Alembicol D.

Topical application:

Slight erythema was observed in test animals following topical application with the test substance, as supplied. Very slight erythema was seen in the control guinea pigs.

CHALLENGE:

The dermal reactions seen in the 17 test animals were more marked than the controls. In one further test animal the dermal reactions were slightly more marked but not clearly different from the maximum reaction seen in the controls. The dermal response seen in the two remaining test animals was similar to the controls.

Applicant's summary and conclusion

Interpretation of results:
Category 1B (indication of skin sensitising potential) based on GHS criteria
Conclusions:
In this test, performed in twenty albino guinea pigs the test substance produced evidence of skin sensitization (delayed contact hypersensitivity) in seventeen animals. An inconclusive result was recorded in one further animal.
Executive summary:

A study was performed to assess the skin sensitization potential of the test substance using the guinea pig. The method followed was that described in:

- EPA Health Effects Testing Guidelines, Subpart E - Specific Organ/Tissue Toxicity §798.4100 Dermal sensitization, September 27, 198S (described in Federal Register Vol. 50, No. 188) and subsequent revisions. Subpart E provides detailed information relating to data requirements of 40 CFR Part 798 and supports the Toxic Substances Control Act (TSCA).

- EEC Methods for the determination of toxicity, Directive 84/449/EEC (OJ No. L251, 19 .9 .84), Part B, Method B.6. Acute toxicity (skin sensitization).

- OECD Guideline for Testing of Chemicals No. 406 "Skin Sensitisation". Adopted: 12 May 1981.

- MAGNUSSON, B. and KLIGMAN, A.M. (1970) Allergic Contact Dermatitis in the Guinea-pig: Identification of contact allergens, Thomas, C.C., Springfield, Illinois, U.S.A.

Based on the results of a preliminary study and in compliance with the guidelines, the following dose levels were selected:

Intradermal injection: 10% v/v in Alembicol D.

Topical application: As supplied.

Challenge application: As supplied and 50% v/v in Alembicol D.

Twenty test and twenty control guinea pigs were used in this study. In this study performed using twenty guinea pigs the test substance produced evidence of skin sensitization (delayed contact hypersensitivity) in seventeen test animals. An inconclusive response was observed in one further animal. Based upon the percentage of animals sensitized the test substance is considered to be a Grade III ("Moderate" sensitizer; MAGNUSSON B. and KLIGMAN AM. (1969) The Identification of Contact Allergens by Animal Assay. The Guinea pig Maximisation Test. J. Invest. Derm., 52.3, 268).

The test substance does require labelling with the risk phrase R43 "May cause sensitization by skin contact" in accordance with Council Directive 79/831/EEC Annex VI, Part II(D) as described in The Commission Directive 91/325/EEC.