Fatty acids, tall-oil, reaction products with linseed oil, maleic anhydride and methanol

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2015

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

Guideline study under GLP. This in vivo study was conducted prior to passage of Commission Regulation (EU) 863/2016.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2015

Report date:

2015

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Principles of method if other than guideline:

This in vivo study was conducted prior to passage of Commission Regulation (EU) 863/2016.

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:

Addition product of maleic anhydride, tall oil fatty acids, linseed oil and methanol

EC Number:

926-195-0

Cas Number:

1176284-65-3

Molecular formula:

not available for the UVCB substance

IUPAC Name:

Addition product of maleic anhydride, tall oil fatty acids, linseed oil and methanol

Test material form:

liquid

Details on test material:

UVCB, Purity: 100%, Batch Number: 210150084, Expiry Date: April 29 2017 (requalified)

In vivo test system

Test animals

Species:

mouse

Strain:

CBA/Ca

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Laboratories S.r.l.
- Females (if applicable) nulliparous and non-pregnant: yes
- Microbiological status of animals, when known: SPF
- Age at study initiation: 9 weeks
- Weight at study initiation:19.8-21.9 g
- Housing:
- Diet (e.g. ad libitum): ad libitum, ssniff SM
- Water (e.g. ad libitum): ad libitum, municipal water supply
- Acclimation period: 14
- Indication of any skin lesions: no

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18.0-25.8
- Humidity (%): 24-86
- Air changes (per hr): 15-20
- Photoperiod (hrs dark / hrs light): 12/12
- IN-LIFE DATES: From: To:

Study design: in vivo (LLNA)

Vehicle:

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

Concentration:

25, 10 and 5% (w/v) for LLNA

No. of animals per dose:

4

Details on study design:

PRE-SCREEN TESTS:
- Compound solubility: Yes
- Irritation: Yes
- Systemic toxicity:
- Ear thickness measurements: Yes
- Erythema scores: Yes

MAIN STUDY

ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: standard LLNA
- Criteria used to consider a positive response: SI of 3 or greater is an indication of a positive response. SI is the mean DPN of treated group divided by mean DPN of control group.

TREATMENT PREPARATION AND ADMINISTRATION:
Test item was administered to the dorsal side of ears on days 1, 2 and 3. No treatments occurred on days 4, 5 and 6. Mice were then injected with 25 microCi of 3HtdR and left for 5 hours. The mice were euthanised by asphyxiation with CO2 and draining auricular lymph nodes were excised and processed on an individual basis. A single cell suspension of lymph node cells was prepared and collected, washed thrice with PBS and pelleted. After overnight incubation at 2-8 degrees C, pellets were resuspended in 5% TCA solution, mixed with 10 ml of scintillation liquid in vials, counted in a scintillation counter and expressed as DPM.

Positive control substance(s):

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

Statistics:

means of duplicate DPM counts indiviidual animals within the dose groups

Results and discussion

Positive control results:

HCA dissolved in the same vehicle demonstrated an appropriate lymphoproliferative response in the study, with a SI of 5.5.

In vivo (LLNA)

Results

Results 1

Key result

Parameter:

EC3

Value:

12.6

Results 2

Parameter:

SI

Value:

4.4

Test group / Remarks:

25% (high) dose group

Results 3

Parameter:

SI

Value:

2.7

Test group / Remarks:

10% (mid) dose group

Results 4

Parameter:

SI

Value:

1.6

Test group / Remarks:

5% (low) dose group

Any other information on results incl. tables

As the stimulation index of two of the three dose groups was over 3, the substance is considered a dermal sensitiser. The EC3 is calculated at 12.6%. No systemic toxicity was observed in the mice.

Applicant's summary and conclusion

Interpretation of results:

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

Remarks:

Stimulation index (SI) over 3

Conclusions:

The test substance, solubilized in acetone:olive oil (4:1 v/v), was shown to be a sensitiser in the Local Lymph Node Assay in CBA/Ca mice. Irritation and thickening of the skin of the ear was found in a preliminary skin irritation test. The stimulation index values were 4.4, 2.7 and 1.6 at concentrations of 25, 10 and 5% (w/v) respectively. Under the conditions of this study, the calculated EC3 value of the substance is 12.6% (w/v).