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
Study period:
April 2003
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2003
Report Date:
2003

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)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.2600 (Skin Sensitisation)
Deviations:
no
GLP compliance:
yes
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
Assessment of the skin sensitisation potential was performed in a guinea pig maximisation test, prior to publication of the LLNA method as described in OECD guideline 429 as the preferred method.

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid
Details on test material:
No further details
Specific details on test material used for the study:
- Name of test material (as cited in study report): XTJ 568
- Physical state: Clear colourless liquid
- Analytical purity: 97.8%
- Impurities (identity and concentrations): no data
- Purity test date: no data
- Lot/batch No.: 8191-34
- Expiration date of the lot/batch: 2004-01-01
- Stability under test conditions: no data
- Storage condition of test material: at room temperature in the dark, stable under storage conditions
- Other: specific gravity: > 0.94 g/cm3

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: Source animal no. 605: 'Charles River France, L'arbresie Cedex, France. Source of other animals: Charles River Deutschland, Kisslegg, Germany
- Age at study initiation: young adult animals (approx. 4 weeks old) were selected
- Weight at study initiation: control females 362 +/- 8 g; exposed females 351 +/- 25 g
- Housing: Group housing of maximally 5 animals per labelled cage (74 cm x 54 cm x 25 cm height) containing purified sawdust as bedding material (SAWI, Jelu Werk, Rosenberg, Germany). Certificates of analysis were examined and retained in the NOTOX archives.
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: at least 5 days before the start of treatment under laboratory conditions

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19.0 - 24.4 °C
- Humidity (%): 56 - 89 %; cleaning procedures in the room might have caused the temporary fluctuations above the optimal level of 70% for relative humidity. Based on laboratory historical data these conditions were considered not to have affected the study integrity.
- Air changes (per hr): 15 air changes per hour
- Photoperiod (hrs dark / hrs light): 12 / 12

Study design: in vivo (non-LLNA)

Induction
Route:
intradermal and epicutaneous
Vehicle:
corn oil
Concentration / amount:
Concentration of test material and vehicle used at induction:
Intradermal: 0.1% in corn oil
Epidermal: 2% in corn oil
Vehicle: corn oil.
Concentration of test material and vehicle used for each challenge: 1% in corn oil.
Challenge
Route:
epicutaneous, semiocclusive
Vehicle:
corn oil
Concentration / amount:
Concentration of test material and vehicle used at induction:
Intradermal: 0.1% in corn oil
Epidermal: 2% in corn oil
Vehicle: corn oil.
Concentration of test material and vehicle used for each challenge: 1% in corn oil.
No. of animals per dose:
Number of animals in test group: 10
females
Number of animals in negative control group: 5 females
Details on study design:
RANGE FINDING TESTS: the selection of concentrations was based on the following criteria:
- The concentrations are well-tolerated systematically by the animals
- For the induction exposures: the highest possible concentration that produced mild to moderate irritation (grade 2-3)
- For challenge exposure: the maximum non-irritant concentration
Series of test substance concentrations were tested. Practical feasibility of administration determined the highest starting-concentration for each route. The starting-and subsequent concentrations were taken from the series: 100% (undiluted), 50%, 20%, 10%, 5%, 2%, 1% and if needed, further lower concentrations using the same steps. The test system and procedures were identical to those used during the main study. The six animals selected were between 4 and 9 weeks old. No body weights were determined.

Intradermal injections:
Initially, a series of four test substance concentrations was used: the highest concentration was chosen based on the corrosive properties of the test substance. Each of two animals received two different concentrations in duplicate (0.1 mL/site) in the clipped scapular region. The resulting dermal reactions were assessed 24 and 48 hours after treatment. Based on the results in the initially treated animals, two additional animals were treated in a similar manner with four lower concentrations at a later stage.

Epidermal application:
A series of four test substance concentrations was used; the highest concentration was chosen based on the corrosive properties of the test substance. Two different concentrations were applied (0.5 mL each) per animal to the clipped flank, using Metalline patches (2x3 cm) mounted on Medical tape, which were held in place with Micropore tape and subsequently Coban elastic bandage. The initially used animals receiving intradermal injections were treated with the lowest concentrations and two further animals receiving intradermal injections were treated with the lowest concentrations and two further animals with the highest concentrations. After 24 hours, the dressing was removed and the skin cleaned of residual test substance using water. The resulting dermal reactions were assessed for irritation 24 and 48 hours after exposure. Based on the results in the initially treated animals, two additional animals were treated in a similar manner with two lower concentrations at a later stage.

MAIN STUDY
A. INDUCTION EXPOSURE
Experimental animals
Day 1: The scapular region was clipped and three pairs of intradermal injections (0.1 mL/site) were made in this area as follows:
A) A 1:1 w/w mixture of Freunds' Complete Adjuvant (Difco, Detroit, U.S.A.) with water for injection (Fresenius AG, Bad Homburg, Germany)
B) The test substance at a 0.1% concentration
C) A 1:1 w/w mixture of the test substance, at twice the concentration used in (B) and Freunds' Complete Adjuvant
One of each pair was on each side of the midline and from cranial A) to caudal C).
Day 3: The dermal reactions caused by the intradermal injections were assessed for irritation
Day 8: The scapular area between the injection sites was clipped and subsequently treated with 0.5 mL of a 2% test substance concentration using a Metalline patch (2 x 3 cm) mounted on Medical tape, which was held in place with Micropore tape and subsequently Coban elastic bandage.

The dressing was removed after 48 hours exposure, the skin cleaned of residual test substance using water and the dermal reactions caused by the epidermal exposure were assessed for irritation.

Control animals
The control animals were treated as described for the experimental animals except that, instead of the test substance, vehicle alone was administered

B. CHALLENGE EXPOSURE
All animals:
Day 22: One flank of all animals was clipped and treated by epidermal application of a 1% test substance concentration and the vehicle (0.1 mL each), using Patch test Plasters (Curatest, Lohmann, Almere, The Netherlands). The patches were held in place with Micropore tape and subsequently Coban elastic bandage.

The dressing was removed after 24 hours exposure and the skin cleaned of residual test substance and vehicle using water. The treated sites were assessed for challenge reactions 24 and 48 hours after removal of the dressing.
Positive control substance(s):
yes
Remarks:
alpha-hexylcinnamicaldehyde, tech. 85% (CAS no. 101-86-0)

Results and discussion

Positive control results:
The skin reactions observed in six experimental animals in response to the 20% test substance concentration in the challenge phase were considered indicative of sensitisation, based on the absence of any response in the control animals. These results lead to a sensitisation rate of 60 per cent to the 20% concentration. From these results, it was concluded that the female guinea pig of the Dunkin Hartley strain is an appropriate animal model for the performance of studies designed to evaluate the sensitising potential of a substance in a Maximisation type of test.

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
0 %
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 0 %. No with. + reactions: 0.0. Total no. in groups: 10.0.
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
1 %
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 1 %. No with. + reactions: 0.0. Total no. in groups: 10.0.
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
0 %
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 0 %. No with. + reactions: 0.0. Total no. in groups: 10.0.
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
1 %
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 1 %. No with. + reactions: 0.0. Total no. in groups: 10.0.
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
0 %
No. with + reactions:
0
Total no. in group:
5
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 0 %. No with. + reactions: 0.0. Total no. in groups: 5.0.
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
1 %
No. with + reactions:
0
Total no. in group:
5
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 1 %. No with. + reactions: 0.0. Total no. in groups: 5.0.
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
0 %
No. with + reactions:
0
Total no. in group:
5
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 0 %. No with. + reactions: 0.0. Total no. in groups: 5.0.
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
1 %
No. with + reactions:
0
Total no. in group:
5
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 1 %. No with. + reactions: 0.0. Total no. in groups: 5.0.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
There was no evidence that the substance had caused skin hypersensitivity in the guinea pig, since no responses were observed in the experimental animals in the challenge phase. This result indicates a sensitisation rate of 0%. Based on these results and according to the EC criteria for classification and labelling requirements for dangerous substance and preparations (Council Directive 67/548/EEC), the substance does not have to be classified and has no obligatory labelling requirements for sensitisation by skin contact.