Tetradecyl acrylate

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

2008-01-22 to 2008-05-09

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

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)

Version / remarks:

April 24, 2002

Deviations:

no

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:

Reaction mass of (D)-2-Propylheptyl acrylate and (L)-2-Propylheptyl acrylate

EC Number:

604-669-5

Cas Number:

149021-58-9

Molecular formula:

C13 H24 O2

IUPAC Name:

Reaction mass of (D)-2-Propylheptyl acrylate and (L)-2-Propylheptyl acrylate

Test material form:

other: liquid

Details on test material:

- Name of test material: 2-Propylheptylacrylat rein
-Test substance number: 07/0846-1
- Analytical purity: 98.5 area-%
- Isomers composition: about 87 % 2 Propylheptylacrylate and 10 % 4-methyl-2-propylhexylacrylate
- Analytical report no.: 07L00384
- Lot/batch No.: B4112/13 - 03122007
- Expiration date of the lot/batch: 2008-12-03
- pH-value: Ca. 5.5 (undiluted test substance)

In vivo test system

Test animals

Species:

mouse

Strain:

CBA

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Mouse / CBA/J from 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: 18.3 g – 21.4 g
- Housing:single housing, Makrolon cage, type II
- Diet: Kliba-Labordiaet (Maus / Ratte Haltung “GLP”), Provimi Kliba SA, Kaiseraugst, Basel, Switzerland, ad libitum
- Water: tap water ad libitum
- Acclimation period: 13 days before the first test-substance application

ENVIRONMENTAL CONDITIONS
- Temperature: 20-24°C
- Humidity: 30-70%
- Photoperiod (hrs dark / hrs light): 12 h / 12 h (6.00 a.m. – 6.00 p.m. / 6.00 p.m. – 6.00 a.m.)

Study design: in vivo (LLNA)

Vehicle:

other: AOO 4:1 (mixture of acetone:olive oil Ph.Eur./DAB in a ratio 4:1 parts by volume)

Concentration:

3%, 10%, 30% test substance solution (w/w) in AOO

No. of animals per dose:

4 animals in control group, 5 animals in test groups

Details on study design:

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Criteria used to consider a positive response: The increase SI of cell count by a factor of >1.5 and/or of 3H-thymidine incorporation by a factor of ≥ 3 as compared to the concurrent vehicle control group is generally considered as indicating a sensitizing potential of a test substance.

TREATMENT PREPARATION AND ADMINISTRATION:
-Randomization: Prior to first application, the animals were distributed to the individual groups, received their animal numbers and were allocated to the respective cages.

-Body weight determination: Individual body weights on day 0 prior to the first application and on day 5 prior to the sacrifice of the animals.

-Signs and symptoms: No detailed clinical examination of the individual animals was performed but any obvious signs of systemic toxicity and/or loc
al inflammation at the application sites were noted in the raw data.

-Form of application:
Epicutaneous application is simulating dermal contact with the compound which is possible to occur under practical use conditions.

-Application volume:
25 μL per ear

-Site of application:
Dorsal part of both ears

-Frequency of application:
3 consecutive applications (day 0 – day 2) to the same application site

-Selection of doses:
The selection of vehicle and concentrations takes into account available information on the chemical/physical properties and the composition of the test substance. In addition the results of a pretest with a 50% test-substance preparation in AOO were considered, which showed slightly increased ear weights and increased lymph node weights as indication of ear irritation. Additionally the 50% preparation caused slight scaling on the backside of the ears.

-Mortality check:
Twice each workday (beginning and end) and once on Saturdays, Sundays and on public holidays.

-3H-Thymidine injection:
On study day five (about 66 to 72 hours after the last application of test substance to the ears) the mice were injected intravenously with 20 μCi of 3H
-thymidine in 250 μL of sterile saline into the tail vein. The animals were sacrificed on study day 5 about 5 hours after 3H-thymidine injection by cervical dislocation.

-Determination of ear weight: Immediately after the death of each animal a circular piece of tissue (diameter 0.8 cm) was punched out of the apical part of each ear of all animals. The weight of the pooled punches was determined for each test group. These measurements serve for detecting a potential inflammatory ear swelling.

-Removal and weight determination of the lymph nodes:
Immediately after removal of the ear punches the left and right auricular lymph nodes were dissected. The weight of the pooled lymph nodes from both sides was determined for each animal.

-Determination of cell count was performed by electronic measurement with a Casy(R)-Counter.

-Measurement of 3H-thymidine incorporation of the lymph node cells:
The remaining cell suspensions were washed twice with phosphate buffered saline (PBS) and precipitated with 5% trichloro-acetic acid (TCA). Each precipitate was transferred to scintillation fluid and incorporation of 3H-thymidine into the cells was measured in a ß-scintillation counter.

Positive control substance(s):

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

Statistics:

Calculation of EC (estimated concentration leading to the respective SI values): calculation by linear or semi-logarithmical regression between the data points directly below and above the SI if possible or using the two nearest points below or above the SI.

Results and discussion

Positive control results:

Stimulation index cell counts (1%, 3%, 10%): 1.10, 1.34, 1.77
Stimulation index 3H-thymidine incorporation (1%, 3%, 10%): 1.84, 2.35, 3.26

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Remarks on result:

other: see Remark

Remarks:

Stimulation index cell count (3%, 10%, 30%): 0.95, 1.05, 1.36. Stimulation index 3H-thymidine incorporation (3%, 10%, 30%): 1.27, 1.81, 3.29. The estimated concentration that leads to the SI of 1.5 for cell count (EC 1.5) and the estimated concentration that leads to the SI of 3.0 for 3H-thymidine incorporation (EC 3) were calculated by linear regression from the results of the 30% and 10% concentration to be 38.8% and 26.1%, respectively.

Results 2

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: DPM were 609 in the vehicle control and increased to 2002.6 in the group treated with 30% test substance (for details see Table 1).

Any other information on results incl. tables

Table 1: 3H-Thymidine incorporation: test group mean values and stimulation indices.

Test group	Treatment	^3H-thymidine incorporation
		(DPM/lymph node pair)	Stimulation index
1	Vehicle AOO	609.0	1
2	3% in AOO	770.8	1.27
3	10% in AOO	1105.0	1.81
4	30% in AOO	2002.6	3.29

Applicant's summary and conclusion

Interpretation of results:

sensitising