Registration Dossier

Diss Factsheets

Administrative data

Description of key information

Not sensitising (OECD 406, GLP, K, rel.1).

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:
Between 05 August 2019 and 06 September 2019
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
GLP study conducted according to OECD Guideline 406 without deviations
Qualifier:
according to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
17 July 1992.
Deviations:
no
Principles of method if other than guideline:
Not applicable
GLP compliance:
yes (incl. QA statement)
Remarks:
Signed on 2019-10-14
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
LLNA can lead to a false positive response (Ku et al., 2008) with fatty acids like nonanoic acid. This substance is one of the constituents of the registered substance. Therefore, the LLNA could not be performed to assess the skin sensitising potential of the registered substance. A GPMT was therefore conducted instead.

Ref: Hyun-Ok Ku et al., Analysis of differential gene expression in auricular lymph nodes draining skin exposed to sensitizers and irritants, Toxicology Letters Volume 177, Issue 1, 28 February 2008, Pages 1-9, DOI: 10.1080/000155598442728
Specific details on test material used for the study:
- Date received: 15 July 2019
Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Male albino guinea pigs of Dunkin-Hartley strain 5-6 weeks old.
- Source: Charles River (F-69210 Saint-Germain-Nuelles)
- Weight at study initiation: 269-312 g
- Housing: Group of 5 max in polycarbonate containers, the flooring of which was covered with dust-free cuttings ans the top fitted with a stainless steel lid with a feeding device and drinking device of 500 mL.
- Diet: ENVIGO, 2040C ad libitum
- Water: Tap water from public distribution system ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature: 19-25 °C
- Humidity: 30-70 %
- Rate of air exchange: at least 10 changes/hour
- Lighting cycle: 12 hours continuous light/12 hours darkness
Route:
intradermal
Vehicle:
corn oil
Concentration / amount:
Three pairs of intradermal injections of 0.1 mL with:
-FCA diluted at 50% in physiological saline
- a test item at 100%
- FCA at 50% + test item at 100% (equal volumes v/v)
Day(s)/duration:
10 days
Adequacy of induction:
highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically
Route:
epicutaneous, occlusive
Vehicle:
corn oil
Concentration / amount:
topical application of 0.5 mL at 100%
Day(s)/duration:
2 days
Adequacy of induction:
non-irritant substance, but skin pre-treated with 10% SDS
No.:
#1
Route:
epicutaneous, occlusive
Vehicle:
corn oil
Concentration / amount:
topical application at 100 and 50%
Day(s)/duration:
24 hours
Adequacy of challenge:
highest non-irritant concentration
No. of animals per dose:
20 males for treated group and 10 males for negative control group
Details on study design:
PRELIMINARY TESTS:
Maximum Non Necrotizing Concentration (M.N.N.C.) determination: 2 male guinea pigs were used. The test item was injected by intradermal route at the following concentrations: 100%, diluted at 50%, 20% and 10% in corn oil. A discrete erythema was note in one animal (1/2) and no cutaneous reaction was noted in the other animal at the tested concentration of 100%. No cutaneous reaction was noted at the tested concentrations of 50%, 20% and 10%.
The first induction of the main study was carried out by intradermal injection at the maximal non necrosing concentration of 100%.

Pre-Maximum Non Irritant Concentration (Pre -M.N.I.C.) determination: 2 male guinea pigs were used. The test item was applied under an occlusive dressing during 24 hours, at the following concentrations 100%, diluted at 50%, 25% and 10% in corn oil. 24 hours after the removal of the occlusive dressings, no cutaneous reaction was recorded. In view of these results, the concentration selected was 100% for the 2nd induction of the main study and the MNIC determination began at this concentration of 100%.

Maximum Non Irritant Concentration (M.N.I.C.) determination: 3 males guinea pigs were used. After induction by intradermal injection with corn oil and by topical application with corn oil and a 10-day rest phase, the challenge phase under occlusive dressing for 24 hours consists in a single topical application of the test item at the following concentrations: 100%, diluted at 50%, 20% and 10% in corn oil. 24 hours and 48 hours after the removal of the occlusive dressings, no cutaneous reaction was recorded. In view of this result, the concentrations selected were 100% (MNIC) and 50% (1/2 MNIC) for the challenge phase.

MAIN STUDY
A. INDUCTION EXPOSURE (INTRADERMAL - 1st induction: Day 0)
- 3 pairs of intradermal injections of 0.1 mL were performed in the negative control group: 2 intradermal injections of Freund’s Complete Adjuvant diluted at 50% in a physiological saline solution; 2 intradermal injections of corn oil and 2 intradermal injections of a mixture with equal volumes - Freund’s Complete Adjuvant at 50% and corn oil.
- 3 pairs of intradermal injections of 0.1 mL were performed in the treated group: 2 intradermal injections of Freund’s Complete Adjuvant diluted at 50% in a physiological saline solution; 2 intradermal injections of the test item at 100% and 2 intradermal injections of a mixture with equal volumes - Freund’s Complete Adjuvant at 50% and the test item at 100%.

B. INDUCTION EXPOSURE (TOPICAL - 2nd induction: Day 7 to Day 10)
- Topical application, on the same zone, with the test item t 100%, 24 hours after brushing with 0.5 mL of a solution of Sodium lauryl sulfate at 10%.
- Duration: 3 days
- Concentrations: 100%

C. REST PHASE: 10 days

D. CHALLENGE EXPOSURE (Day 20 to day 23)
- Day(s) of challenge: Day 20
- Exposure period: 24 h
- Challenge phase (Groups 1 & 2): topical application under occlusive dressing at the following concentrations: 100% & 50%.
- Evaluation: 24 and 48 h after challenge
Challenge controls:
No. of animals: 10 males
Challenge: 100% and 50%
Positive control substance(s):
no
Remarks:
Results of the last 11 positive groups with a-Hexylcinnamaldehyde were available
Positive control results:
Historical data: α-Hexylcinnamaldehyde at 30 or 75% in corn oil induced skin sensitisation in more than 30% of the treated animals indicating the validity of the study.
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
100 and 50%
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
None
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
100 and 50%
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
None
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
100 and 50%
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
None
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
100 and 50%
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
None
Remarks on result:
no indication of skin sensitisation
Group:
positive control
Remarks on result:
other: Historical data: last 11 positive control data

Sensitising potential assessment: No macroscopic cutaneous reactions attributable to allergy was recorded during the examination following the removal of the occlusive dressing (challenge phase) from the animals of the treated group with the test item. No cutaneous intolerance reaction was recorded in animals from the negative control group.

Weight evolution: No abnormality was recorded in the weight growth of both groups.

Mortality: No mortality occurred during this study.

Interpretation of results:
GHS criteria not met
Conclusions:
Under the test conditions, test substance is not classified as sensitiser according to the criteria of the Regulation EC No. 1272/2008 (CLP) and of the GHS.
Executive summary:

In a Magnusson & Kligman maximisation study (GPMT) performed according to OECD Guideline 406 and in compliance with GLP, Guinea pigs were treated as follows:

After induction (intradermic injection at 100% and topical application at 100%) of 20 Guinea Pigs of treated group with the test substance and a 10 -day rest phase, the challenge phase, under occlusive dressing for 24 hours, consisted to a single topical application of the test item at 100% and diluted at 50% in corn oil. The test concentrations for the main study were determined from a preliminary study.

 

No macroscopic cutaneous reactions attributable to allergy was recorded during the examination following the removal of the occlusive dressing (challenge phase) from the animals of the treated group with the test item. No cutaneous intolerance reaction was recorded in animals from the negative control group.

 

The sensitivity of the guinea-pig was checked with known sensitiser i.e., α-Hexylcinnamaldehyde indicating the validity of the study. 

Under the test conditions, test substance is not classified as sensitiser according to the criteria of the Regulation EC No. 1272 /2008 (CLP) and of the GHS.

This study is considered as acceptable and satisfies the requirement for skin sensitisation endpoint.

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

A key study was identified (Phycher, 2019). In this dermal sensitisation study performed according to the OECD guideline No. 406 and in compliance with GLP, the test material was tested in twenty Guinea Pigs (20 males) using the Magnusson and Kligman method (20 treated animals + 10 controls). The preliminary study determined the concentration to be used for the induction and challenge phases of the main study. The test material diluted in corn oil at 100% was administered by injection for intradermal induction. On Day 7, topical induction was performed with the test material as supplied, 24h after brushing with 0.5 mL of sodium lauryl sulfate at 10%. For the challenge, on Day 20, the test material was tested at 50 diluted in corn oil and at 100%.

The historical positive control α-Hexylcinnamaldehyde produced skin sensitisation. The test system was therefore considered to be valid.

In both groups, no clinical signs and no deaths were noted during the study. The body weight gain of the treated animals was similar to that of the control animals. After the challenge application, no cutaneous reaction was noted. The overall sensitisation rate was 0%.

Under the test conditions, the test material is not a skin sensitizer.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

Harmonised classification:

The substance has no harmonised classification according to the Regulation (EC) No. 1272/2008 (CLP).

Self classification:

Based on the available data no additional self-classification is proposed according to the CLP and to the GHS.

No data was available for respiratory sensitisation. However, this substance is not a skin sensitizer, therefore according to Figure R.7.3 -2 of the Chapter R.7 (V 4.1 - October 2015) the chemical is not considered as a respiratory sensitiser.

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