Fatty acids, C16-18 and C18-hydroxy, oligomeric reaction products with adipic acid, decanoic acid, isooctadecanoic acid, octanoic acid, pentaerythritol and stearic acid

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

03 Apr - 04 May 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

adopted in 2010

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:

921-836-0

EC Number:

921-836-0

IUPAC Name:

921-836-0

Details on test material:

- Name of test material (as cited in study report): [trade name]
- Physical state/appearance: dark yellow liquid
- Analytical purity: no data
- Batch No.: 103991

In vivo test system

Test animals

Species:

mouse

Strain:

other: CBA/CaOlaHsd

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Winkelmann, Borchen, Germany
- Age at study initiation: 8-9 weeks
- Housing: 5 per cage in IVC cages, type II L
- Diet: Altromin 1324 maintenance diet for rats and mice; ad libitum
- Water: tap water (sulphur acidified to a pH value of approx. 2.8); ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 55 ± 10
- Air changes (per hr): at least 10
- Photoperiod (hrs dark / hrs light): 12/ 12

Study design: in vivo (LLNA)

Vehicle:

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

Concentration:

25, 50 and 100%

No. of animals per dose:

- Main test: 5
- Pre-test: 3

Details on study design:

RANGE FINDING TESTS:
2 animals were treated by topical application with the undiluted test substance on three consecutive days. 1 animal was treated with AOO (negative control).
- Compound solubility: The maximum technically applicable concentration of the test item was found to be 100% in AOO.
- Irritation: Both ears of each animal were observed daily for erythema and edema as well as for ear thickness before the first application and approx. 48 hours after the first application and before sacrifice (day 6).
- Lymph node proliferation response: not examined
Neither signs of systemic toxicity nor signs of irritation at the application site could be detected in any animal.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Criteria used to consider a positive response: A substance is considered a sensitiser in the LLNA if at least one concentration of the test substance results in a 3-fold or greater increase in ³H-methyl thymidine incorporation into lymph node cells of the test group animals, relative to that recorded for the lymph nodes of control group animals (SI >= 3).

TREATMENT PREPARATION AND ADMINISTRATION:
- Topical application: Each mouse was treated by topical application of 25 µL of the solution to the entire dorsal surface of each ear. Topical applications were performed once daily over three consecutive days.
- Administration of ³H-Methyl Thymidine: Five days after the first topical application all mice were dosed with 20 µCi ³H-methyl thymidine by intravenous injections of 250 µL of ³H-methyl thymidine, diluted to a working concentration of 80 µCi/mL.
- Preparation of cell suspension:
Approx. 5 hours after the injection of ³H-methyl thymidine all mice were sacrificed by cervical dislocation. Draining auricular lymph nodes were excised, individually pooled for each animal (2 lymph nodes per animal) and collected in PBS. A single cell suspension of pooled lymph node cells was prepared by gentle mechanical disaggregation through polyamide gauze. After washing the gauze with PBS the cell suspension was pelleted in a centrifuge. The supernatant was discarded and the pellets were resuspended with PBS. The washing procedure was repeated.
After the final wash each pellet was resuspended in 5% TCA at 4 °C for approx. 18 hours. Each precipitate was washed again, resuspended in 5% TCA and scintillation fluid was added. The solution was transferred into scintillation vials and stored at room temperature overnight.
- Determination of incorporated ³H-methyl thymidine: The incorporation was measured in a beta counter and expressed as the number of disintegrations per minute (DPM). Determination of radioactivity was performed individually for each animal.

Positive control substance(s):

other: P-Phenylenediamine (in a separate study)

Results and discussion

Positive control results:

Stimulation index (mean value out of 5 animals treated with phenylene diamine): 10.9 ± 3.3

In vivo (LLNA)

Results

Results 1

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: Mean DPM values (out of 5 animals, respectively) ± SD: - Negative control: 1897.6 ± 302.2 - 25%: 2948.2 ± 801.5 - 50%: 3243.6 ± 1639.1 - 100%: 4198.2 ± 1213.2

Results 2

Key result

Parameter:

SI

Remarks:

mean value out of 5 animals

Value:

1.6

Test group / Remarks:

25%

Results 3

Key result

Parameter:

SI

Remarks:

mean value out of 5 animals

Value:

1.7

Test group / Remarks:

50%

Results 4

Key result

Parameter:

SI

Remarks:

mean value out of 5 animals

Value:

2.2

Test group / Remarks:

100%

Any other information on results incl. tables

All animals survived throughout the test period without showing any clinical signs. All animals showed the expected weight development.

Applicant's summary and conclusion

Interpretation of results:

other: CLP/EU GHS criteria not met, no classification required according to Regulation(EC) No. 1272/2008

Conclusions:

CLP: not classified