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Description of key information

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Well documented and reported study fully adequate for assessment. The study was conducted according to internationally accepted technical guidelines and in compliance with GLP in a recognized contract research organization.
according to
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Version / remarks:
of 2010
according to
EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)
Version / remarks:
of 2008 as amended 2012
GLP compliance:
yes (incl. certificate)
Type of study:
mouse local lymph node assay (LLNA)
Details on test animals and environmental conditions:
- Mouse (healthy females only), strain: CBA/Ca with appropriate range of bodyweight at study start.
- Source: Charles River (UK) Ltd.
- Age at treatment start (1st induction): nine to eleven weeks.
- Weight at treatment start (1st induction): Minimum 16.9 g, maximum 21.0 g.
- Housing: Two animals per polycarbonate cage inside a barriered rodent facility.
- Bedding material: Woodflake bedding.
- Cage enrichment: Nestlets and plastic shelter
- Diet (ad libitum): Standard rodent diet (Rat and Mouse No. 1 Maintenance Diet) containing no added antibiotic,
chemotherapeutic or prophylactic agent.
- Water (ad libitum): Tap water
- Acclimation period: At least 7 days before treatment start under laboratory conditions.

Analysis of the batch of diet used and water did not provide evidence of contamination that might have prejudiced the study.


Air conditioned room kept at positve pressure without re-circulation of the filtered fresh air supplied to the room.
Controlled environmental conditions were set at:
- Air changes per hour in the animal room: controlled periodically
- Temperature (°C): 21 ± 2°C
- Relative Humidity (%): 40 to 70%
- Photoperiod (artificial lighting): 12 hrs day / 12 hrs night
There was no mentioning of any deviations from these ranges, which compromised the integrity or validity of the study.

acetone/olive oil (4:1 v/v)
Main Study:
0, 5, 10, and 25%

WS400505 was unsuitable for dosing at 50% in any of the vehicles stated in the protocol; 25% was the maximum achievable concentration.
No. of animals per dose:
Main Study:
4 female animals per dose
Details on study design:
Compound Solubility:

WS400505 was unsuitable for dosing at 50% in any of the vehicles stated in the protocol; 25% was the maximum achievable concentration forming a bitty white suspension.

Treatment Preparation and Administration:

On three consecutive days, groups of 4 female mice were treated by topical application to the entire dorsal surface of both ears with 25 μL/ear/day at the following concentrations (w/v) of test substance in the vehicle: 0, 5, 10, and 25%, respectively.

The test substance preparations were prepared freshly on each day of administration.

Observations, Measurements and Endpoints (Pooled treatment group approach):

All animals were checked daily for signs of ill health or toxicity. The ears were also examined daily for signs of irritation. In addition, bodyweights were recorded on Days 1 (prior to treatment) and 6 (prior to termination). On Day 6, all animals were injected into the tail vein 3H-methyl thymidine diluted in phosphate buffered saline at a nominal dose of 20 µCi per mouse, in order to measure lymphocyte proliferation by radioactive labelling. Five hours afterwards the draining (auricular) lymph nodes were excised and pooled for each experimental group. Single cell suspensions were prepared for determination of incorporated radioactivity.

Criteria Used to Consider a Positive Response:

The test substance is regarded as a sensitizer if at least one concentration of the test substance produces a stimulation index (SI) ≥ 3.

Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Data were not statistically analysed.
Positive control results:
A stimulation index (SI) of 9.1 was attained in a contemporaneous positive control assay with the same strain of mice (CBA/Ca) in response to 25% v/v hexyl cinnamic aldehyde in AOO, thus demonstrating the reliability and sensitivity of this test system and assay to detect skin sensitization potential in this laboratory.
Key result
Test group / Remarks:
5% w/v test substance dilution
Key result
Test group / Remarks:
10% w/v test substance
Key result
Test group / Remarks:
25% w/v test substance
other: disintegrations per minute (DPM)
Remarks on result:
other: Overall DPM/lymph node values for the experimental groups treated with 5, 10, and 25% w/v test substance dilutions were 411, 423, and 248 DPM/node, respectively. For the vehicle control group, 349 DPM/node were recorded.

Mortality / clinical signs:

There were no deaths, no signs of ill health or toxicity and no signs of local irritation over the treated area. Greasy fur on the head was noted following each dosing occasion.

Body weight: There was no indication of adverse effects on bodyweight attributable to treatment with the test substance.

Interpretation of results:
not sensitising
Migrated information Criteria used for interpretation of results: EU
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 low stimulation indices observed in the local lymph node assay, WS400505 is considered not to be a skin sensitizer and does not warrant any classification regarding skin sensitisation according to European classification rules [Regulation (EC) 1272/2008].