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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:
09 March to 02 April 1993
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1993
Report Date:
1993

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.6 (Skin Sensitisation)
Version / remarks:
Guideline 84/449
Deviations:
no
GLP compliance:
yes
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
Method was not available

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder

In vivo test system

Test animals

Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Winkelmann GmbH
- Females nulliparous and non-pregnant: [yes]
- Age at study initiation: 4 to 7 Weeks
- Weight at study initiation: Mean 317g (280 to 360g)
- Housing: 5 animals per cage
- Diet (e.g. ad libitum): Altromin 3020, ad libitum
- Water (e.g. ad libitum): Tap water, ad libitum
- Acclimation period: at least 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 +/- 1.5
- Humidity (%): 40 - 70
- Air changes (per hr): 12 - 15
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal
Vehicle:
physiological saline
Remarks:
Cremophor EL (2% v/v) in sterile NaCl
Concentration / amount:
5%
Day(s)/duration:
Day 1
Adequacy of induction:
highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically
Route:
epicutaneous, occlusive
Vehicle:
physiological saline
Remarks:
Cremophor EL (2% v/v) in sterile NaCl
Concentration / amount:
10%
Day(s)/duration:
Day 8 / 48 h
Adequacy of induction:
highest technically applicable concentration used
Challengeopen allclose all
No.:
#1
Route:
epicutaneous, occlusive
Vehicle:
physiological saline
Remarks:
Cremophor EL (2% v/v) in sterile NaCl
Concentration / amount:
10%
Day(s)/duration:
Day 22 / 24 h
Adequacy of challenge:
highest non-irritant concentration
No.:
#2
Route:
epicutaneous, occlusive
Vehicle:
physiological saline
Remarks:
Cremophor EL (2% v/v) in sterile NaCl
Concentration / amount:
6%
Day(s)/duration:
Day 22 / 24 h
No. of animals per dose:
One test substance group consisting of 20 experimental animals (No. 21 to 40) and two control groups consisting of 10 animals each (No. 1 to 10 and 11 to 20)
Details on study design:
Intradermal Induction
One day prior to application the skin on the dorsum and the flanks was shorn. Starting behind the nucha,three intradermal injections were administered on each side of and parallel to the spinal column.
The application volume per injection site was 0.1 ml, The animals of the three groups were treated as follows:

a) Test substance group

1. Injection site pair (cranial)
Freund's complete adjuvant (Difco Lab.) 1:1 diluted with physiological saline solution (sterile).
2. Injection site pair (medial)
5% test substance, formulated as a suspension in Cremophor EL (2% v/v) in sterile physiological NaCl
3. Injection site pair (caudal)
5% test substance, formulated as a suspension in Cremophor EL (2% v/v) in sterile physiological NaCl and Freund's complete adjuvant in equal proportions.

b) Control groups
The animals of the control groups were treated like the animals of the test substance group, the formulation for the injection site pairs 2 and 3, however, contained no test compound but a corresponding amount of Cremophor EL (2% v/v) in sterile physiological NaCl .

Topical Induction
One week after intradermal induction topical induction took place. On the eve, the areas to be treated were shorn, and a 10% preparation of sodium lauryl sulphate in parafin oil was spread on these areas. Hypoallergenic patches {2x4 cm) were applied between or on the injection sites; they were covered with aluminum foil and kept in place on the skin with Fermoflex adhesive tape (Transatlantic GmbH, Schwarzenbach).
The hypoallergenic patches were treated as follows:
a) Test substance group: 0.5 ml test substance 10%
b) Control group: 0.5 ml Cremophor EL (2% v/v) in sterile physiological NaCl
At the end of the 48-hour exposure period the substance residues were removed with physiological saline.

Topical Challenge of Sensitization
The topical challenge took place 3 weeks after intradermal induction. Meanwhile, the test compound concentration for challenge had been determined in a range-finding test on 5 guinea pigs treated like control animals during inductions.

One day prior to the treatment the animals were shorn on the dorsum and the flanks. For challenge a hypoallergenic patch saturated with 10% and 6% test compound formulation was placed on the left flank of the animals of the test substance group and the ist control group. The patch was fixed for 24 hours on the skin with Fermoflex adhesive tape. The patches were applied in both concentrations cranially or caudally - alternately from animal to animal. On the right flank two control patches were fixed, saturated with Cremophor EL (2% v/v) in sterile physiological NaCl.
The application volume was 0.5 ml in each case.

At the end of the exposure period, the skin of the animals was depilated in the region of the treated areas. Pilca cream (depilation cream for humans, containing salts of the thioglycolic acid; manufactured by Schwarzkopf GmbH, Hamburg) was used for that purpose.

Challenge controls:
Control groups
The animals of the control groups were treated like the animals of the test substance group, the formulation for the injection site pairs 2 and 3, however, contained no test compound but a corresponding amount of sterile physiological saline solutiothe vehicle Cremophor EL (2% v/v) in sterile physiological NaCl.
Positive control substance(s):
yes

Results and discussion

Positive control results:
positive controls are valid

In vivo (non-LLNA)

Resultsopen allclose all
Key result
Reading:
1st reading
Hours after challenge:
48
Group:
test group
Dose level:
10%
No. with + reactions:
2
Total no. in group:
20
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
72
Group:
test group
Dose level:
10%
No. with + reactions:
1
Total no. in group:
20
Remarks on result:
no indication of skin sensitisation
Reading:
1st reading
Hours after challenge:
48
Group:
test group
Dose level:
6%
No. with + reactions:
1
Total no. in group:
20
Remarks on result:
no indication of skin sensitisation
Reading:
2nd reading
Hours after challenge:
72
Group:
test group
Dose level:
6%
No. with + reactions:
1
Total no. in group:
20
Remarks on result:
no indication of skin sensitisation
Reading:
1st reading
Hours after challenge:
48
Group:
negative control
Dose level:
10%
No. with + reactions:
0
Total no. in group:
10
Reading:
2nd reading
Hours after challenge:
72
Group:
negative control
Dose level:
10%
No. with + reactions:
0
Total no. in group:
10
Reading:
1st reading
Hours after challenge:
48
Group:
negative control
Dose level:
6%
No. with + reactions:
0
Total no. in group:
10
Reading:
2nd reading
Hours after challenge:
72
Group:
negative control
Dose level:
6%
No. with + reactions:
0
Total no. in group:
10

Any other information on results incl. tables

The incidences of skin reddenings after challenge are summarized in the following table

 

 

Test substance Group (20 animals)

1stControl group (10 animals)

Test substance patch

Control patch

Test substance patch

Control patch

Hours

48

72

Total

48

72

48

72

Total

48

72

Challenge 10%

2

1

2

0

0

0

0

0

0

0

Challenge 6%

1

1

1

0

0

0

0

0

0

0

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
The substance was found to be non-sensitising in the maximisation test.
Executive summary:

The test substance was tested for skin sensitizing properties on female guinea pigs in a maximisation test as described by MAGNUSSON and KLIGMAN. Intradermal induction was carried out with a 5% test compound formulation, topical induction with 10%, which was the highest doese which could be formulated. As this concentration had produced no primary skin irritant effects in the range-finding test, the areas to be treated were pre-irritated with sodium lauryl sulphate prior to topical induction.

 

After the challenge exposure one animal (5%) and two animals (10%) of the test substance group reacted with skin reddening to the 6% and 10% test compound formulation, respectively. In the control group no skin reactions were observed.

 

Thus, under the conditions of the maximisation test, the test compound has no skin sensitizing potential.