Registration Dossier

Toxicological information

Skin sensitisation

Currently viewing:

Administrative data

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
09 May 2011 - 17 June 2011
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Compliant to GLP and testing guideline; adequate coherence between data, comments and conclusions.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2011
Report Date:
2011

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of study:
mouse local lymph node assay (LLNA)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid

In vivo test system

Test animals

Species:
mouse
Strain:
CBA
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Breeder: Janvier, Le Genest-Saint-Isle, France
- Age at study initiation: on the first day of the treatment period, the animals of the preliminary and main tests were approximately 8 weeks old
- Weight at study initiation: a mean body weight +- standard deviation of 21.0 g +/- 0.9 g
- Housing: the animals were group housed by two (preliminary test) or four (main test) in polycarbonate cages with stainless steel lids
- Diet (e.g. ad libitum): free access to SSNIFF R/M-H pelleted maintenance diet
- Water (e.g. ad libitum): tap water (filtered using a 0.22 micron filter)
- Acclimation period: the animals were acclimated to the study conditions from 10 to 14 days before the beginning of the treatment period.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2°C
- Humidity (%): 50 ± 20%
- Air changes (per hr): approximately 12 cycles/hour of filtered, non-recycled air
- Photoperiod (hrs dark / hrs light): 12 h/12 h (7:00 - 19:00).

IN-LIFE DATES: From: 19 May 2011 To: 14 June 2011.

Study design: in vivo (LLNA)

Vehicle:
acetone/olive oil (4:1 v/v)
Concentration:
5, 10, 25, 50 or 100% in a mixture acetone/olive oil (4/1; v/v).
No. of animals per dose:
7 groups of 4 animals.
Details on study design:
RANGE FINDING TESTS:
- Compound solubility: the first choice vehicle was a mixture of Acetone/Olive Oil (4/1; v/v) (AOO). Satisfactory solubility of the test item in AOO
(i.e. solution to the naked eye at the concentration of 50% was obtained).
- Irritation: the test item was not excessively irritant (increase in ear thickness not above 25%) whatever the concentration
- Lymph node proliferation response: incorporation of tritiated methyl thymidine.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- 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 together with
consideration of a dose-response relationship. Other relevant criteria such as radioactivity levels and ear thickness were also taken into account to evaluate the data.

TREATMENT PREPARATION AND ADMINISTRATION:
On days 1, 2 and 3, a dose-volume of 25 µL of the appropriate dosage form preparation was applied to the dorsal surface of both ears (one concentration per ear), using an adjustable pipette fitted with a plastic tip.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)

Results and discussion

Positive control results:
The study was considered valid as the SI for the positive control was higher than 3 (3.15).

In vivo (LLNA)

Results
Key result
Parameter:
EC3
Remarks:
%
Value:
31
Cellular proliferation data / Observations:
CELLULAR PROLIFERATION DATA
See attached table

DETAILS ON STIMULATION INDEX CALCULATION
See attached table

EC3 CALCULATION
EC3 = 31%

CLINICAL OBSERVATIONS:
- Clinical signs and mortality
Neither mortality nor clinical signs were observed during the observation period.
- Local irritation
Dryness of the skin of the ear was noted on day 6 in all animals given 50 and 100%. In addition, erythema and scabs on ear were noted in 1/4 females given 100%.
No notable increase in ear thickness was observed at any of the tested concentrations.

BODY WEIGHTS
The body weight change of test item treated animals was similar to that of vehicle treated animals.

Applicant's summary and conclusion

Interpretation of results:
Category 1B (indication of skin sensitising potential) based on GHS criteria
Conclusions:
TERTIODODECYLMERCAPTAN induced contact hypersensitivity in the murine Local Lymph Node Assay.
According to the EC3 value of 31%, it was a weak sensitizer.
Executive summary:

The potential of the test item, TERTIODODECYLMERCAPTAN, to induce contact hypersensitivity was evaluated using the murine Local Lymph Node Assay (LLNA). This study was conducted in compliance with the principles of Good Laboratory Practice.

 

A preliminary test was first performed in order to define the concentrations of test item to be used in the main test.

In the main test, 28 female CBA/J mice were allocated to 7 groups of 4 animals:

.            five treated groups receiving the test item at the concentration of 5, 10, 25, 50 or 100% in a mixture acetone/olive oil (4/1; v/v) (vehicle),

.            one negative control group receiving the vehicle,

.            one positive control group receiving the positive control, a-hexylcinnamaldehyde (HCA), a moderate sensitizer, at the concentration of 25% in a mixture acetone/olive oil (4/1; v/v).

 

During the induction phase, the test item, vehicle or positive control item was applied over the ears (25 µL per ear) for 3 consecutive days (days 1, 2 and 3). After 2 days of resting, the proliferation of lymphocytes in the lymph node draining the application site was measured by incorporation of tritiated methyl thymidine (day 6). The obtained values were used to calculate Stimulation Indices (SI). The irritant potential of the test item was assessed in parallel by measurement of ear thickness on days 1, 2, 3 and 6. Following the solubility assay, a mixture acetone/olive oil (4/1; v/v) was chosen as vehicle. The concentrations selected for the preliminary test were 10, 25, 50 and 100%. Since the test item was non-irritant in the preliminary test, the highest concentration retained for the main test was the maximal practicable concentration (100%).

 

Neither mortality nor clinical signs were observed during the observation period. Dryness of the skin of the ear was noted on day 6 in all animals given 50 and 100%. In addition, erythema and scabs on ear were noted in 1/4 females given 100%. No notable increase in ear thickness was observed at any of the tested concentrations. As acceptance criteria were met, this experiment was therefore considered valid.

 

The results are presented in the following table:

 

Treatment

Concentration

(%)

Irritation level

Stimulation Index

(SI)

Test item

5

non-irritant

0.18

Test item

10

non-irritant

0.64

Test item

25

non-irritant

2.11

Test item

50

non-irritant

5.94

Test item

100

slightly irritant

4.71

HCA

25

-

3.15

A significant lymphoproliferation (SI > 3) was noted at concentrations=50% with a clear evidence of a dose-response relationship between all concentrations. In the absence of excessive local irritation, the significant lymphoproliferative responses were attributed to delayed contact hypersensibility. The EC3value is equal to 31%.

The test item, TERTIODODECYLMERCAPTAN, induced contact hypersensitivity in the murine Local Lymph Node Assay. According to the EC3value obtained, the test item should be considered as a weak sensitizer.