Ethyl 4-oxopiperidine-1-carboxylate

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

from 2006-07-18 to 2006-08-29

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Report date:

2006

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

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:

Ethyl 4-oxopiperidine-1-carboxylate

EC Number:

249-984-5

EC Name:

Ethyl 4-oxopiperidine-1-carboxylate

Cas Number:

29976-53-2

Molecular formula:

C8H13NO3

IUPAC Name:

ethyl 4-oxopiperidine-1-carboxylate

Test material form:

liquid

Details on test material:

- Name of test material (as cited in study reports): JNJ-126971-AAA (T000509)
- Physical state: liquid
- Appearance: clear colourless to light yellow liquid

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: 00451178HT000509G3I381
- Expiration date of the lot/batch: 21 September 2008
- Purity:100%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: At room temperature
- Solubility and stability of the test substance in the solvent/vehicle:
solubility in vehicle: methanol, dichloromethane, acetone, ethanol, 2-propanol, N,N-dimethylformamide: > 500 g/L
solubility in water: > 500 g/L
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: no data

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing:
The test item was placed into a volumetric flask glass beaker on a tared balance and the vehicle (acetone:olive oil (4+1)) 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.

In vivo test system

Test animals

Species:

mouse

Strain:

CBA

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: female mice (nulliparous and non-pregnant), CBA/CaOlaHsd; Harlan Netherlands, B.V., The Netherlands
- Age at beginning of acclimatization: 7 to 9 weeks
- Initial weight: 18.6 - 21.4 grams (mean = 20.2 grams)
- Housing: single caging in Makrolon Type I, with wire mesh top, 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: under test conditions after health examination. Only animals without any visible signs of illness will be used for the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3°C
- Humidity (%): 30 - 87%
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12/12 (artificial light 6.00 a.m. - 6.00 p.m.)

Study design: in vivo (LLNA)

Vehicle:

acetone/olive oil (4:1 v/v)

Concentration:

0, 1, 50 and 100%

No. of animals per dose:

4 females/group; 3 groups and 1 control group

Details on study design:

PRE-SCREEN TESTS:
To determine the highest non-irritant test concentration or the highest technically applicable concentration, a non-GLP pretest was performed in two mice (pretest excluded from the Statement of Compliance). The data showed tha tthe highest test item concentration, which can be used is 100%. At this concentration the treated mouse did not show any signs of irritation.
- Compound solubility: see specific details on test material
- Irritation: no signs of irritation observed at the maximum concentration of 100%
- Systemic toxicity: no data
- Ear thickness measurements: no data
- Erythema scores: no data

ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: Local Lymph Node Assay
- Criteria used to consider a positive response:
A test item is regarded as a sensitiser in the LLNA if the following criteria are fulfilled:
- First, that exposure to at least one concentration of test item resulted in an incorporation of ³HTdR at least 3-fold or greater than that recorded in 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.

TREATMENT PREPARATION AND ADMINISTRATION:
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 item concentrations of 1, 50 and 100% in acetone:olive oil (4+1). The application volume, 25 µL, was spread over the entire dorsal surface (diameter ~ 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).

Positive control substance(s):

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

Statistics:

The mean values and standard deviations were calculated for the body weights.

Results and discussion

Positive control results:

- background I: 16.02 dpm (background = 1 mL 5% trichloroacetic acid)
- background II: 17.55 dpm (background = 1 mL 5% trichloroacetic acid)
- control group: 5083.65 dpm; 5066.87 dpm-bg*; 633.4 dpm per lymph node**
- test group 1 (5%): 11655.70 dpm; 11638.92 dpm-bg*; 1457.0 dpm per lymph node**; S.I. = 2.3
- test groupd 2 (10%): 21860.70 dpm; 21843.92 dpm-bg*; 2732.6 dpm per lymph node**; S.I. = 4.31
- test group 3 (25%): 56809.20 dpm; 56792.42 dpm-bg*; 7101.2 dpm per lymph node**; S.I. = 11.21
- EC3 = 6.7%
* The mean value was taken from the figures of backgroud I and background II
** Since the lymph nodes of the animals of a dose group were pooled, DPM/node was determined by dividing the measured value by the number of lymph nodes pooled.
Number of lymph nodes per test item concentration was 8

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Value:

0.82

Test group / Remarks:

1% group

Results 2

Parameter:

SI

Value:

0.66

Test group / Remarks:

50% group

Results 3

Parameter:

SI

Value:

0.72

Test group / Remarks:

100% group

Cellular proliferation data / Observations:

CELLULAR PROLIFERATION DATA
- background I: 16.28 dpm (background = 1 mL 5% trichloroacetic acid)
- background II: 18.44 dpm (background = 1 mL 5% trichloroacetic acid)
- control group: 3278.86 dpm; 3261.5 dpm-bg*; 407.7 dpm per lymph node**
- test group 1 (1%): 2704.57 dpm; 2687.2 dpm-bg*; 335.9 dpm per lymph node**; S.I. = 0.82
- test groupd 2 (50%): 2173.81 dpm; 2156.5 dpm-bg*; 269.6 dpm per lymph node**; S.I. = 0.66
- test group 3 (100%): 2371.81 dpm; 2354.5 dpm-bg*; 294.3 dpm per lymph node**; S.I. = 0.72
* The mean value was taken from the figures of backgroud I and background II
** Since the lymph nodes of the animals of a dose group were pooled, DPM/node was determined by dividing the measured value by the number of lymph nodes pooled.
Number of lymph nodes per test item concentration was 8

DETAILS ON STIMULATION INDEX CALCULATION
See results

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

CLINICAL OBSERVATIONS:
- Viability/mortality: No deaths occurred during the study period.
- Clinical signs: 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 of this strain and age.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

The test item was not observed to be a skin sensitiser under the described conditions. Based on the available data and the criteria of the CLP Regulation, the substance has not to be classified for skin sensitisation.