2-Propenoic acid, 2-methyl-, 2-(4-benzoyl-3-hydroxyphenoxy)ethyl ester

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was conducted in accordance with the OECD guideline 429 (adopted 24 April 2002) and in compliance with GLP (incl. certificate).

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2008

Report date:

2008

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes (incl. QA statement)

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

2-(4-benzoyl-3-hydroxyphenoxy)ethyl 2-methylprop-2-enoate

Cas Number:

16613-04-0

Molecular formula:

C19 H18 O5

IUPAC Name:

2-(4-benzoyl-3-hydroxyphenoxy)ethyl 2-methylprop-2-enoate

Details on test material:

- Name of test material (as cited in study report): ULS-611
- Test substance No.: 08/0140-1
- Analytical purity: 99.2 % (99.2 g/100 g)
- Impurities (identity and concentrations): not specified
- Storage stability: The stability under storage conditions over the study period was guaranteed by the manufacturer
- Homogeneity: The test substance was homogenous by visual inspection

In vivo test system

Test animals

Species:

mouse

Strain:

CBA

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH, Sandhofer Weg 7, 97633 Sulzfeld
- Age at study initiation: 6-12 weeks
- Weight at study initiation: 18.9-22.3 g
- Housing: single housing
- Diet (ad libitum): Kliba-Labordiät (Maus/Ratte Haltung "GLP"), Provimi Kliba SA, Kaiseraugst, Basel, Switzerland
- Water: tap water, ad libitum
- Acclimation period: 7 days before the first test substance application


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24 °C
- Humidity (%): 30-70 %
- Air changes: fully air-conditioned rooms
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (LLNA)

Vehicle:

propylene glycol

Concentration:

3 %, 10 %, 30 % (w/w); the 30 % preparation was the maximum technically applicable concentration.

No. of animals per dose:

5

Details on study design:

Experimental procedure:
Groups of 5 female CBA/J mice each were treated with several concentrations of the test substance in propylene glycol or with the vehicle alone. The study used 3 test groups and 1 control group. Each test animal was applied with 25 μL per ear of the respective test-substance preparation to the dorsum of both ears for three consecutive days. The control group was treated with 25 μL per ear of the vehicle alone. The animals were checked twice each workday (beginning and end) in terms of mortality and once on Saturdays, Sundays and on public holidays.
Three days after the last application on study day 5, the mice were injected intravenously with 20 μCi of 3H-thymidine in 250 μL of sterile saline into a tail vein.
About 5 hours after the 3H-thymidine injection, the mice were sacrificed. Immediately after the death of each animal, a defined area with a diameter of 0.8 cm was punched out of the apical part of each ear and for each test group the weight of the pooled punches was determined for detecting a potential inflammatory ear swelling.
Immediately after removal of the ear punches, the left and the right auricular lymph nodes were dissected. The weights of each animal’s pooled lymph nodes were determined. Thereafter lymph nodes were pooled group wise and further evaluated by measuring their cellular content and 3H-thymidine incorporation into the lymph node cells (indicators of cell proliferation).
The stimulation indices (fold of change as compared to the vehicle control) for cell count, 3H-thymidine incorporation (measured in a ß-scintillation counter), lymph node weight and ear weight were calculated.

Historical control data:
Vehicle related historical control data, gathered over an appropriate time period, were documented and included in the appendix of the report.

Positive control:
A concurrent positive control (reliability check) with a known sensitizer was not included into the study.
However, studies using the positive control substance Alpha-Hexylcinnamaldehyde are performed twice a year in the conducting testing laboratory in order to show that the test system is able to detect sensitizing compounds under the test conditions chosen.

Positive control substance(s):

hexyl cinnamic aldehyde (CAS No 101-86-0)

Results and discussion

Positive control results:

Positive results using Alpha-Hexylcinnamaldehyde (technical grade, 85 %) were consistently obtained over the years using several variations of the methods and different vehicles.

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Remarks on result:

other: see Remark

Remarks:

Test group 1: vehicle in propylene glycol: 1.00 [test group 1/test group 1] Test group 2: 3 % test substance in propylene glycol: 2.89 [test group 2/test group 1] Test group 3: 10 % test substance in propylene glycol: 1.40 [test group 3/test group 1] Test group 4: 30 % test substance in propylene glycol: 1.23 [test group 4/test group 1]

Results 2

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: see Remark

Remarks:

Test group 1: vehicle in propylene glycol: 424.9 [DPM/Lymph Node Pair] Test group 2: 3 % test substance in propylene glycol: 1226.39 [DPM/Lymph Node Pair] Test group 3: 10 % test substance in propylene glycol: 593.6 [DPM/Lymph Node Pair] Test group 4: 30 % test substance in propylene glycol: 521.1 [DPM/Lymph Node Pair]

Any other information on results incl. tables

- No signs of systemic toxicity were noticed.

- When applied as 3%, 10% and 30% preparation in propylene glycol, the test substance did not induce a concentration dependent and biologically relevant response in the auricular lymph node cell counts.

- The lymph node weights changes followed the lymph node cell count.

- Concomitantly, the increase of 3H-thymidine incorporation into the cells was neither concentration dependent, nor biologically relevant at the concentrations tested.

- None of the test-substance preparations caused an increase in ear weights.

Table 1: Stimulation indices (fold of change as compared to the vehicle control) for cell count, 3H-thymidine incorporation, lymph node weight and ear weight

Test group	Treatment	Stimulation index
		Cell Count	3H-thymidine incorporation	Lymph Node Weight	Ear Weight
1	vehicle propylene glycol	1.00	1.00	1.00	1.00
2	3 % in propylene glycol	1.31	2.89	1.34	1.02
3	10 % in propylene glycol	1.21	1.40	1.25	1.00
4	30 % in propylene glycol	1.05	1.23	1.16	1.03