Eugenol

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2006

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study design reported to follow OECD Guideline 429 (2002) with deviations (no positive control was included in the experiment, group housing instead of individual housing, and acclimatisation period was not reported).

Data source

Reference

Reference Type:

publication

Title:

Investigation of the Dermal sensitization potential of various essential oils in the local lymph node assay.

Author:

Lalko J., and Api A.M.

Year:

2006

Bibliographic source:

Food and Chemical Toxicology. 2006; Volume 44: 736-746

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)

Deviations:

yes

Remarks:

No positive control was included in the experiment, group housing instead of individual housing, and acclimatisation period was not reported.

GLP compliance:

not specified

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

Eugenol

EC Number:

202-589-1

EC Name:

Eugenol

Cas Number:

97-53-0

Molecular formula:

C10H12O2

IUPAC Name:

2-methoxy-4-(prop-2-en-1-yl)phenol

Details on test material:

- Name of test material (as cited in study report): Eugenol
- Analytical purity: 99.9%
Further details not reported.

In vivo test system

Test animals

Species:

mouse

Strain:

other: CBA/Ca

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Interfauna UK, Shaw's Farm, Blackthorne, Bicester, Oxon, UK
- Age at study initiation: 8 to 12 weeks old
- Weight at study initiation: 17 to 21 g
- Housing: Housed in groups of 4 per cage
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 to 25°C
- Humidity (%): 30 to 70%
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (LLNA)

Vehicle:

other: 1:3 ethanol:diethyl phthalate

Concentration:

0, 2.5, 5, 10, 25, 50%

No. of animals per dose:

4 mice/dose group

Details on study design:

RANGE FINDING TESTS: none

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: LLNA
- Criteria used to consider a positive response: A test material was considered a sensitizer if at least one concentration was observed to have a stimulation index value of 3 or more.

TREATMENT PREPARATION AND ADMINISTRATION:
Groups of female mice (n=4) were dosed topically on the dorsum of both ears with 25 µL eugenol. Each group received one of five test concentrations (2.5, 5, 10, 25, or 50%). A vehicle control group was similarly treated with 1:3 EtOH:DEP. Dosing occurred daily for 3 consecutive days. The animals “rested” for 2 days and on the 6th day after the first application, all mice were injected intravenously by the tail vein with phosphate buffered saline containing [3H]methyl thymidine. Five hours later, the mice were euthanized and the draining auricular lymph nodes were excised and pooled for each experimental group.
For each concentration, a stimulation index relative to the concurrent vehicle-control was calculated. The stimulation index value was calculated by dividing the mean dpm at a given dose level by the mean dpm of the vehicle control group.

Positive control substance(s):

other: Study was reported to be performed according to OECD guideline 429, which requires that a positive experiment be conducted. Therefore, it was assumed that a positive control experiment was performed but was not reported.

Results and discussion

In vivo (LLNA)

Results

Results 1

Parameter:

EC3

Value:

5.4

Test group / Remarks:

eugenol

Results 2

Parameter:

SI

Value:

1.2

Test group / Remarks:

2.5% eugenol

Results 3

Parameter:

SI

Value:

2.7

Test group / Remarks:

5.0% eugenol

Results 4

Parameter:

SI

Value:

6

Test group / Remarks:

10.0% eugenol

Results 5

Parameter:

SI

Value:

14.3

Test group / Remarks:

25.0% eugenol

Results 6

Parameter:

SI

Value:

19.4

Test group / Remarks:

50.0% eugenol

Any other information on results incl. tables

Mean values of DPM per dose group are provided below.

Vehicle group = 386 dpm/lymph node

2.5% eugenol = 464 dpm/lymph node

5.0% eugenol = 1,051 dpm/lymph node

10.0% eugenol = 2,322 dpm/lymph node

25.0% eugenol = 5,534 dpm/lymph node

50.0% eugenol = 7,476 dpm/lymph node

A stimulation index value greater than or equal to 3 are considered to result in a positive response.

Mean values of SI per dose group are provided below.

Vehicle group = not applicable

2.5% eugenol = 1.2

5.0% eugenol = 2.7

10.0% eugenol = 6.0

25.0% eugenol = 14.3

50.0% eugenol = 19.4

The estimated concentration (EC3) giving rise to a stimulation index of 3 was calculated by linear interpretation of the dose response data for each assay. The lower the EC3 value, the greater the skin sensitization potential.

The EC3 for eugenol was reported to be 5.4%.

Applicant's summary and conclusion

Interpretation of results:

sensitising

Remarks:

Migrated information Category 1B Criteria used for interpretation of results: EU

Executive summary:

The potential for skin sensitisation following exposure to eugenol was assessed using the local lymph node assay in groups of 4 female mice at test concentrations of 0, 2.5, 5, 10, 25, or 50%. The estimated concentration (EC3) giving rise to a stimulation index of 3 was calculated by linear interpretation of the dose response data for each assay. The lower the EC3 value, the greater the skin sensitization potential. The authors concluded that eugenol was regarded as a potential skin sensitizer at concentrations at and above 10.0%

(specifically, based on the data for the EC3, at a concentration >5.4%).