N-phenyl-N-piperidin-4-ylpropionamide

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2005-07-20 to 2005-08-02

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference 1

Reference Type:

study report

Title:

Unnamed

Year:

2005

Report date:

2005

Reference 2

Reference Type:

other: Expert statement

Title:

Unnamed

Year:

2014

Report date:

2014

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

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

Deviations:

yes

Remarks:

The relative humidity in the animal room was between 30-87%. However, this deviation did not affect the validity of the study.

GLP compliance:

not specified

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

N-phenyl-N-piperidin-4-ylpropionamide

EC Number:

216-543-3

EC Name:

N-phenyl-N-piperidin-4-ylpropionamide

Cas Number:

1609-66-1

Molecular formula:

C14H20N2O

IUPAC Name:

N-phenyl-N-piperidin-4-ylpropionamide

Test material form:

solid: particulate/powder

Details on test material:

- Name of test material (as cited in study reports): JNJ-1594255-AAA (T000425)
- Physical state: solid (powder)
- Appearance: white

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: 00454795
- Expiration date of the lot/batch: December 31, 2005
- Purity: 100%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature

In vivo test system

Test animals

Species:

mouse

Strain:

CBA/Ca

Remarks:

CBA/CaOlaHsd

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Netherlands, B.V. Postbus 6174, NL-5960 AD Horst, The Netherlands
- Age at study initiation: 5-6 weeks (beginning of acclimatization)
- Weight at study initiation: 18.4- 22.8 grams
- Housing: individually in Makrolon Type I cage with wire mesh top and granulated soft wood bedding
- Diet (e.g. ad libitum): pelleted standard diet, ad libitum
- Water (e.g. ad libitum): tap water, ad libitum
- Acclimation period: no data on period but under test conditions after health examination. Only animals without any visible signs of illness were used for the study.


ENVIRONMENTAL CONDITIONS
- Temperature (deg C): 22 ± 3 deg C
- Humidity (%): 30-87%
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): artificial light 6:00 am - 6:00 pm

Study design: in vivo (LLNA)

Vehicle:

other: ethanol:water (7:3)

Concentration:

pre-test: 2.5, 5, 10 and 25% (w/v)
main test: 5, 10, and 25% (w/v)

No. of animals per dose:

4 females (nulliparous and non-pregnant)

Details on study design:

RANGE FINDING TESTS:
- Compound solubility: 43 g/L in water and 500 g/L in ethanol
- Irritation: A pre-test was performed in two mice with concentrations of 2.5, 5, 10, and 25% (w/v). At these concentrations, the treated animals did not show any signs of toxicity or irritation.
- Lymph node proliferation response: no data

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: Determination of incorporated 3HTdR
- Criteria used to consider a positive response: A test substance is regarded as a sensitizer in the LLNA if the following criteria are fulfilled:
First, that exposure to at least one concentration of the test substance resulted in an incorporation of 3HTdR at least 3-fold or greater than that recorded in the control mice, as indicated by the stimulation index.
Second, that the data are compatible with a conventional dose-response, although allowance must be made (especially at high topical concentrations) for either local toxicity or immunological suppression.
The decision to select a stimulation index (S.I.) of 3 as an arbitrary indication of sensitizing activity was made on the basis of investigations performed with a wide range of chemicals.

TREATMENT PREPARATION AND ADMINISTRATION:
The test item was placed into a volumetric flask glass beaker on a tared balance and the vehicle (Ethanol:Water (7:3)) was quantitatively added. The test item concentrations were prepared serially. Homogeneity of the test item in the vehicle was maintained during treatment with the magnetic stirrer. The preparations were made freshly before each dosing occasion.

Each test group of mice was treated by topical (epidermal) application to the dorsal surface of each ear lobe (left and right) with different test substance concentrations of 5, 10, and 25% (w/v) in Ethanol: water (7:3). The application volume, 25 µL, was spread over the entire dorsal surface ( diameter of approximately 8 mm) of each ear lobe once daily for three consecutive days. A further group of mice was treated with an equivalent volume of the relevant vehicle alone (control animals). A hair dryer was used to dry the ear's surface as quickly as possible to avoid loss of test substance applied.

Positive control substance(s):

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

Statistics:

The mean values and standard deviations were calculated for body weight.
A statistical analysis was conducted for assessment of the dose-response relationship, and the Estimated Concentration for a S.I. of 3 (EC3) value was calculated according to the equation: EC3=(a-c)[(3-d)/(b-d)]+c, where EC3 is the estimated concentration of the test substance required to produce a 3-fold increase in draining lymph node cell proliferative activity; (a,b) and (c,d) are respectively the co-ordinates of the two pairs of data lying immediately above and below the S.I. value of 3 on the local lymph node assay dose response plot.

Results and discussion

Positive control results:

The validation/positive control study was performed with alpha-Hexylcinnamaldehyde in acetone:olive oil, 4:1 (v/v) using CBA/CaOlaHsd mice in March, 2005. S.I. values of 2.29, 3.21, and 8.44 were determined with the positive control substance at concentrations of 5, 10, and 25%, respectively. The EC3 was estimated to be 8.9% (w/v).

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Value:

1.47

Test group / Remarks:

5% w/v group

Results 2

Parameter:

SI

Value:

1.85

Test group / Remarks:

10% w/v group

Results 3

Parameter:

SI

Value:

2.75

Test group / Remarks:

25% w/v group

Cellular proliferation data / Observations:

CELLULAR PROLIFERATION DATA
DPM per lymph node (8 lymph nodes in total):
5% group: 429.0
10% groupo: 542.3
25% group: 804.5

DETAILS ON STIMULATION INDEX CALCULATION: see above

EC3 CALCULATION
The EC3 value could not be calculated, since all SI's are below 3.

VIABILITY/MORTALITY:
No deaths occurred during the study period

CLINICAL OBSERVATIONS:
No symptoms of local toxicity at the ears of the animals and no systemic findings were observed during the study period

BODY WEIGHTS
The body weight of the animals, recorded prior to the first application and prior to treatment with 3HTdR, was within the range commonly recorded for animals for this strain and age.

Any other information on results incl. tables

Calculation and Results of Individual Data
Test item concentration % (w/v)	Group	Measurement DMP	Calculation			Result
			DPM-BG^a	Number of lymph nodes	DPM per lymph nodes	S.I.
---	BG I	2.1	---	---	---	---
---	BG II	12.4	---	---	---	---
---	CG 1	2346.4	2339.2	8	292.4
5	TG 1	3439.5	3432.3	8	429	1.47
10	TG 2	4345.3	4338.1	8	542.3	1.85
25	TG 3	6443.5	6436.3	8	804.5	2.75

BG = Background (1mL 5% trichloroacetic acid) in duplicate

CG = Control Group

TG = Test Group

a = the mean value was taken from the figures BG I and BG II.

Due to the dose-related response and the fact that the SI at the highest tested concentration was just below 3, it was decided to classify this compound as a mild skin sensitizer for occupational health perspectives.

Applicant's summary and conclusion

Interpretation of results:

Category 1B (indication of skin sensitising potential) based on GHS criteria

Conclusions:

The test substance was not a skin sensitiser under the described conditions. However, based on the dose-related response and the fact that the SI at the highest tested concentration was just below 3, it was decided to classify this test item as a mild sensitizer for occupational health perspectives (subcategory 1B).