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Description of key information

A guinea pig maximisation test was performed to determine the sensitization potential of FAT 20013/C according to the OECD Guideline 406 andEU Method B.6 (Skin Sensitisation). 20 animals per dose were subjected to Intradermal induction and epidermal induction at 5 % and 50 % respectively on Day 1 and Day 8, followed by epidermal challenge at a dose of 25 % on Day 22. Animals were evaluated for skin reaction according to Draize method. 2- Mercaptobenzothaizole was used as positive control. No positive reactions were observed in the animals either when treated with vaselinum album alone or when treated with the test article at 25 % in vaselinum album. FAT 20013/C is therefore, classified as a non-sensitiser in albino guinea pigs according to the grading of Magnusson and Kligman. 100 % sensitization was observed with postive control after 48 hour reading.

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records
Reference
Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Experiment start date: 03 May 1995; Experiment end date: 05 June 1995; Study reported date: 28 July 1995.
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Qualifier:
according to guideline
Guideline:
EU Method B.6 (Skin Sensitisation)
GLP compliance:
yes
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
Data from a reliable in vivo test conducted before the enforcement of Commission Regulation (EU) 640/2012 of 06 July 2012 amending, for the purpose of its adaptation to technical progress, Regulation (EC) No. 440/2008 laying down test methods pursuant to Regulation (EC) No 1907/2006 of the European Parliament and of the Council on the Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH) are available.
Specific details on test material used for the study:
Identification: LANACRON GRAU S-B ROH TROCKEN SFO (FAT 20013/C)
Description: Grey solid
Batch Number: 220
Purity / Formulation: 53.15 %
Stability of Test Article: Stable under specified storage conditions; expiration date: DEC-1999
Storage Conditions: In the original container at room temperature (approx. 20 °C), away from direct sunlight.
Safety Precautions: Gloves, goggles and face mask were obligatory to ensure the health and safety of the personnel.
Species:
guinea pig
Strain:
Himalayan
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: BRL, Biological Research Laboratories Ltd. Wölferstrasse 4, 4414 Füllinsdorf / Switzerland
- Females (if applicable) nulliparous and non-pregnant: 30 females /6 females, nulliparous and non-pregnant
- Age at study initiation: 5 - 7 weeks
- Weight at study initiation: Control and Test Group: 324 - 385 g Pretest: 331 - 399 g
- Housing: Individually in Makrol on type-3 cages with autoclaved standard softwood bedding ("Lignocel", Schill AG, CH-4132 Muttenz).
- Diet (e.g. ad libitum): Pelleted standard Kliba 342, Batch no. 68/95 guinea pig breeding/maintenance diet ("Kliba", Klingentalmühle AG, CH-4303 Kaiseraugst), ad libitum.
- Water (e.g. ad libitum): Community tap water from Füllinsdorf, ad libitum. Once weekly additional supply of ascorbic acid (1 g/l) via the drinking water.
- Acclimation period:One week for the control and test group under test conditions after health examination. No acclimatization for the animals of the pretest.
Only animals without any visible signs of illness were used for the study
- Indication of any skin lesions:

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 - 22 °C
- Humidity (%): 56-68 %
- Air changes (per hr): 10-15 air changes per hour
- Photoperiod (hrs dark / hrs light): 12-hour light, 12-hour dark cycle
Route:
intradermal
Vehicle:
other: ethanol
Concentration / amount:
5 %
Day(s)/duration:
Test day 01
Adequacy of induction:
highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically
Route:
epicutaneous, occlusive
Vehicle:
other: Vaselinum album
Concentration / amount:
50 %
Day(s)/duration:
Test day 08
Adequacy of induction:
non-irritant substance, but skin pre-treated with 10% SDS
No.:
#1
Route:
epicutaneous, occlusive
Vehicle:
other: Vaselinum album
Concentration / amount:
25 %
Day(s)/duration:
Test day 22
Adequacy of challenge:
highest non-irritant concentration
No. of animals per dose:
Control Group: 10
Test Group: 20
Intradermal Pretest: 2
Epidermal Pretest: 4
Details on study design:
RANGE FINDING TESTS:
The objective of this investigation was to identify a maximally tolerated concentration of the test article suitable for the induction phase of the main study. In addition, a suitable non-irritant concentration of the test article, by the topical route of administration, was identified for the challenge application. The procedure employed for these investigations was as follows:

INTRADERMAL INJECTIONS:
Intradermal injections (0.1 ml/site) were made into the clipped flank of two guinea-pigs at concentrations of 5, 3 and 1% of the test article in ethanol. The resulting dermal reactions were assessed 24 hours later. For intradermal induction application a 5 % test article dilution was selected.

EPIDERMAL APPLICATIONS:
Both flanks of each of 4 guinea pigs were clipped and shaved just prior to the application. Thereafter 4 patches* of filter paper ( 2 x 2 cm) were saturated with the test article at A = 50% (this concentration used was found to be the most qualified to assure an optimum technical application procedure), B = 25%, C = 15% and D = 10% of the test article in vaselinum album and applied to the clipped and shaved flanks. The patches were covered by a strip of aluminum foil and firmly secured by elastic plaster wrapped around the trunk and covered with impervious adhesive tape. This procedure ensured the intensive contact of the test article. The dressings were removed after an exposure period of 24 hours. 21 hours after removal of the dressing the application site was depilated with an approved depilatory cream (VEET Cream, Reckitt & Colman AG, CH-4123 Allschwil) to clean the application site from staining produced by the test article, so that possible erythema reactions were clearly visible at that time. The depilatory was placed on the patch sites and surrounding areas, and left on for 3-5 minutes. It was then thoroughly washed off with a stream of warm, running water. The animals were then dried with a disposable towel, and returned to their cages. The reaction sites were assessed 24 and 48 hours after removal of the bandage for erythema and oedema on a numerical basis according to Draize described above.
For the epidermal induction the test article at 50% and for challenge procedure the test article at 25% in vaselinum album was selected.

MAIN STUDY
A. INDUCTION EXPOSURE (intradermal)
- No. of exposure: 01
- Exposure period: not applicable
- Test groups: 3 injections administered
1) 1:1 (v/v) mixture of Freund's Complete Adjuvant and physiological saline.
2) The test article, diluted to 5% with ethanol.
3) The test article diluted to 5 % by emulsion in a 1:1 (v/v) mixture of Freund's Complete Adjuvant and physiological saline.
- Control group: 3 injections administered
1) 1:1 (v/v) mixture of Freund's Complete Adjuvant and physiological saline.
2) Ethanol
3) 1:1 (w/w) mixture of vaselinum album in a 1:1 (v/v) mixture of Freund's Complete Adjuvant and physiological saline.
- Site: scapular region (approximately 6 x 8 cm)
- Frequency of applications: Once

B. INDUCTION EXPOSURE (epidermal)
- No. of exposure: 01
- Exposure period: 4 hours
- Site: : scapular area (approximately 6 x 8 cm)
- Frequency of applications: Once
- Concentration: 50 % in vaselinum album

C. CHALLENGE EXPOSURE
- No. of exposures: 01
- Day(s) of challenge: Test day 22
- Exposure period: 04 hours
- Site: left and right flank of each guinea-pig
- Concentrations: 25 %
- Evaluation (hr after challenge): 24 and 48 hours after the removal of the dressing the application sites were assessed for erythema and oedema using the numerical scoring system according to Draize.

Erythema and eschar formation:
No erythema.................................................................................................................... 0
Very slight erythema (barely perceptible)............................................................................ 1
Well-defined erythema...................................... .................................................................2
Moderate to severe erythema............................................................................................. 3
Severe erythema (beet redness) to slight eschar formation (injuries in depth)........................... 4

Oedema formation:
No oedema........................................................................................................................0
Very slight oedema (barely perceptible) ...............................................................................1
Slight oedema (edges of area well-defined by definite raising) .................................................2
Moderate oedema (raised approximately 1 millimeter) ...........................................................3
Severe oedema (raised more than 1 millimeter and extending beyond the area of exposure) .....4
Positive control substance(s):
yes
Remarks:
2- mercaptobenzothiazol
Positive control results:
In this study 95 % and 90 % of the animals of the test group were observed with positive skin reactions at the 24-hour and 48-hour reading respectively after treatment with a non-irritant test substance concentration of 5 % in peanut oil. No skin reactions were observed in the control group. The results obtained with the test article at 10 % in peanut oil were not taken into consideration since the both control group were observed with erythematous reactions.
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
25 %
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
No clinical observation recorded
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
25 %
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
No clinical observation recorded
Remarks on result:
no indication of skin sensitisation
Reading:
1st reading
Hours after challenge:
24
Group:
positive control
Dose level:
5 % Intradermal induction
No. with + reactions:
19
Total no. in group:
20
Remarks on result:
positive indication of skin sensitisation
Reading:
2nd reading
Hours after challenge:
48
Group:
positive control
Dose level:
5 % Intradermal induction
No. with + reactions:
18
Total no. in group:
20
Remarks on result:
positive indication of skin sensitisation
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
no indication of skin sensitisation
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
no indication of skin sensitisation
Interpretation of results:
GHS criteria not met
Conclusions:
FAT 20013/C is considered as not a sensitiser.
Executive summary:

A guinea pig maximisation test was performed to determine the sensitization potential of FAT 20013/C according to the OECD Guideline 406 and EU Method B.6 (Skin Sensitisation). 20 animals per dose were subjected to Intradermal induction and epidermal induction at 5 % and 50 % respectively on Day 1 and Day 8, followed by epidermal challenge at a dose of 25 % on Day 22. Animals were evaluated for skin reaction according to Draize method. 2- Mercaptobenzothaizole was used as positive control. No positive reactions were observed in the animals either when treated with vaselinum album alone or when treated with the test article at 25 % in vaselinum album. FAT 20013/C is therefore, classified as a non-sensitiser in albino guinea pigs according to the grading of Magnusson and Kligman. 100 % sensitization was observed with postive control after 48-hour reading.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

Based on the available data of the GPMT selected as key study, FAT 20013 does not warrant classification for skin sensitisation under the CLP (1272/2008) Regulation.