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Administrative data

Description of key information

Ethylvanillin is not a skin sensitizer in a LLNA study performed in 2003 (CIT, 2003 - reliability 1) selected as a key study.

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
Study period:
From 29 apr 2003 to 06 oct 2003
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
according to guideline
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
GLP compliance:
Type of study:
mouse local lymph node assay (LLNA)
Details on test animals and environmental conditions:
- Source: Charles River Laboratories France, L'Arbresle, France.
- Age at study initiation: approximately 9 weeks old
- Weight at study initiation: 21.7 +/- 1.4 g
- Housing: individually in disposable crystal polystyrene cages (22*8.5*8 cm)
- Diet (e.g. ad libitum): free access to A04 C pelleted diet (SAFE, Villemoisson, Epinay-sur-Orge, France).
- Water (e.g. ad libitum): free access to tap water (filtered using a 0.22 micron filter) contained in bottles.
- Acclimation period: at least 5 days before the start of the study.

- Temperature (°C): 22 +/- 2°C
- Humidity (%): 30 to 70% relative humidity
- Air changes (per hr): 12 cycles per hour of filtered, non-recycled air.
- Photoperiod (hrs dark / hrs light): 12h / 12h

IN-LIFE DATES: From 05 jun 2003 to 10 jun 2003
acetone/olive oil (4:1 v/v)
0, 2.5, 5, 10, 25 and 50 %
No. of animals per dose:
Details on study design:
- Compound solubility: the test item was freely soluble in the vehicle, consequently, the concentrations selected for the preliminary test were 5, 10, 25 and 50%.
- Irritation: no cutaneous reactions and no noteworthy increase in ear thickness were observed in the animals of the treated groups.
- Lymph node proliferation response: no. No positive lymphoproliferative response was noted and no dose-related increase in the stimulation index was observed at the tested concentrations.

- Name of test method: animals will be assigned to the treatment groups by hand procedure. Identification is individually by a number on the tail using an indelible marker.
- Criteria used to consider a positive response: The test item should be regarded as a skin sensitizer when the SI for a dose group is >=3. Other relevant criteria such as cellularity, radioactivity levels and ear thickness will be also taken into account to evaluate the data.
When appropriate, the EC3 value (theoretical concentration resulting in a SI value of 3) will be determined by linear interpolation of points on the dose-response curve, immediately above and below the 3-fold threshold. The equation used for calculation of EC3 will be:
EC3 = c + [(3 - d)/(b - d)] x (a - c)
Where a = the lowest concentration giving stimulation > 3; b = the actual stimulation index caused by a; c = the highest concentration failing to produce a stimulation index of 3; and d = the actual stimulation index caused by c.

TREATMENT PREPARATION AND ADMINISTRATION: 28 female mice were allocated to 7 groups: five treated groups (2.5, 5, 10, 25 and 50%), one negative control group (vehicle), one positive control group (HCA).
On days 1, 2 and 3, a dose-volume of 25 µL of the control or dosage form preparations was applied to the dorsal surface of both ears, using an adjustable pipette fitted with a plastic tip.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Positive control results:
SI = 13.5 with 25% of hexyl cinnamic aldehyde
other: disintegrations per minute (DPM)
Remarks on result:
other: Per node: from 135 to 116.
Test group / Remarks:
2.5% / 4 animals
Test group / Remarks:
5% / 4 animals
Test group / Remarks:
10% / 4 animals
Test group / Remarks:
25% / 4 animals
Test group / Remarks:
50% / 4 animals

Groups 1 2 3 4 5 6 7
Concentrations (%) 0
(Acetone/olive oil)
2,5 5 10 25 50 HCA at 25%
Cell count viable 66 100 142 78 118 102 192
Cell count dead 10 9 14 9 Il 10 27
Viability (%) 86.84 91.74 91.03 89.66 91.47 91.07 87.67
Amount of cells (*106) cells) 3.3 5.00 7.10 3.90 5.90 5.1 19.2
Cellularity index   1.52 2.15 1.18 1.79 1.55 5.82
Number of nodes per group 8 8 8 8 8 8 8
Disintegrations per min per gp (dpm) 579.34 1083.16 989.82 833.98 822.97 930.8 7820.28
Disintegrations per min per node (dpm) 72.42 135.4 123.73 104.25 102.87 116.35 977.54
stimulation index (SI) 1.87 1.71 1.44 1.42 1.61 13.5
Increase in ear thickness (% between day 1 and day 6) 0.00 1.98 6.86 7.84 4.85 -2.91  
EC3 value   NA  

- Positive control results:at the concentration of 25%, a moderate increase in cellularity and a stimulation index exceeding the threshold value of 3 (SI=13.50) was noted. The study was therefore considered valid.

Local irritation:

No cutaneous reactions and no noteworthy increase in ear thickness were observed in the animals at any of the tested concentrations.

Proliferation assay:

In the treated groups, no positive lymphoproliferative response was noted and no dose-related increase in the stimulation index was observed at the tested concentrations.

Interpretation of results:
GHS criteria not met
Not sensitizing
Executive summary:

In a dermal sensitization study (Klein, 2003) with Ethylvanillin in acetone/olive oil (4/1), female CBA/J mice (4 animals per dose) were tested in LLNA, according to the guideline OECD 429 at doses: 0, 2.5, 5, 10, 25 and 50 %.

During the study, no cutaneous reactions (ears irritation) were observed, nor a positive lymphoproliferative response.

In absence of effect no EC3 value could be calculated.

Based on these results, Ethylvanillin is not sensitizing according to CLP 1272/2008/EC criteria.

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

Four studies were available for this endpoint. Two studies are reliable, one with reliability 1 and the other one with reliability 2 according to Klimisch scale. The two other studies are not reliable, one with reliability 3 and the other one with reliability 4. Among the two reliable studies, one was chosen as key study (CIT, 2003 - reliability 1), and the other one was selected as "supporting study". These two LLNA studies showed that Ethylvanillin is not sensitizing to skin.

Conclusion: Based on the two LLNA reliable studies ethylvanillin is not sensitizing to skin.

Justification for classification or non-classification

Based on the results obtained in two LLNA studies, ethylvanillin is not classified for skin sensitization, according to CLP 1272/2008/EC criteria.