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Toxicological information

Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2018
Report Date:
2018

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Version / remarks:
Adopted: 22 July 2010
Deviations:
not specified
GLP compliance:
yes (incl. certificate)
Type of study:
mouse local lymph node assay (LLNA)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Details on test material:
- Name of test material: 4,4'-bis(2-methoxystyryl)-1,1'-biphenyl
- IUPAC name: 1-[(E)-2-(2-methoxyphenyl)ethenyl]-4-{4-[(E)-2-(2-methoxyphenyl)ethenyl]phenyl}benzene
- Molecular formula: C30H26O2
- Molecular weight: 418.53 g/mol
- Smiles notation: COC1=CC=CC=C1/C=C/C2=CC=C(C3=CC=C(/C=C/C4=CC=CC=C4OC)C=C3)C=C2
- InChl: 1S/C30H26O2/c1-31-29-9-5-3-7-27(29)21-15-23-11-17-25(18-12-23)26-19-13-24(14-20-26)16-22-28-8-4-6-10-30(28)32-2/h3-22H,1-2H3/b21-15+,22-16+
- Substance type: Organic
- Physical state: Solid

In vivo test system

Test animals

Species:
mouse
Strain:
CBA/Ca
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: AnLab, Prague, Czech Republic
- Females nulliparous and non-pregnant: yes
- Microbiological status of animals, when known:
- Age at study initiation: 10-11 weeks
- Weight at study initiation: Not available
- Housing: The animals were housed in IVC polycarbonate cages (5 animals per cage) suspended on stainless steel racks
- Diet: ad libitum
- Water: tap water, supply of drinking water was unlimited
- Acclimation period: 6 days
- Indication of any skin lesions: Animals were healthy, without visible signs of disease.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3 °C
- Humidity (%): 50 - 60 %
- Air changes (per hr): Equipped with central air-conditioning
- Photoperiod (hrs dark / hrs light): 12-hour light / 12-hour dark cycle

Study design: in vivo (LLNA)

Vehicle:
acetone/olive oil (4:1 v/v)
Concentration:
the test item at three concentrations (25%, 50% and 100% w/v)
No. of animals per dose:
The doses were selected from the concentration series 100%, 50%, 25%, 10%, 5%, 2.5% etc. according to OECD Guideline No. 429.
The starting concentration was determined according to pre-screen test result.
Number of animals:
5 females – negative control (vehicle)
5 females – positive control
15 females – test item
4 females - pre-screen test, plus spare animals
Details on study design:
PRE-SCREEN TESTS:
- Compound solubility: soluble
- Irritation: all mice (2 mice/group) were observed daily at the application site.
- Systemic toxicity: all mice (2 mice/group) were observed daily at the application site.
- Ear thickness measurements: measured by a calliper on Day 1 (pre-dose), Day 3 and Day 6
- Erythema scores: observed daily at the application site.

MAIN STUDY
- Name of test method: Clinical Observations, Body Weights, Lymph Node Proliferation
ANIMAL ASSIGNMENT AND TREATMENT: Criteria used to consider a positive response: excessive local skin irritation is indicated by an erythema score ≥ 3 and/or an increase in ear thickness of ≥25%.

TREATMENT PREPARATION AND ADMINISTRATION: Preparation of solutions, Dose Preparation and Dose Administration, Preparation of cell suspensions
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Statistics:
For calculation of mean, S.D. and t-test values of body weights MS Excel was used.

Results and discussion

Positive control results:
body weight: 20.39 to 20.81 (from Initial body weight to Terminal body weight);
The daily clinical observation of the animals did not show visible clinical signs of toxicity;
SI values: 6.63* (*Calculated with corresponding control value of 1475 DPM);
The lymph node weight: 0.0642g.

In vivo (LLNA)

Resultsopen allclose all
Key result
Parameter:
SI
Value:
0.81
Test group / Remarks:
100 % (w/v) test substance / test item
Key result
Parameter:
SI
Value:
0.91
Test group / Remarks:
50 % (w/v) test substance / test item
Key result
Parameter:
SI
Value:
0.71
Test group / Remarks:
25 % (w/v) test substance / test item
Cellular proliferation data / Observations:
CELLULAR PROLIFERATION DATA : The DPM values for the three treated groups were 742 (25%), 944 (50%) and 914 (100%), respectively.

DETAILS ON STIMULATION INDEX CALCULATION : The SI values for the three treated groups were 0.71 (25%), 0.91 (50%) and 0.81 (100%), respectively.

EC3 CALCULATION : The EC3 value could not be calculated, since none of the tested concentrations induced a S.I. was not greater than the threshold value of 3.

CLINICAL OBSERVATIONS: The daily clinical observation of the animals did not show visible clinical signs of toxicity.

BODY WEIGHTS: The mean body weight of the negative control group, positive control group and the three test item treated groups are shown in below:
20.42 to 20.37 (negative control: from Initial body weight to Terminal body weight)
20.39 to 20.81 (positive control: from Initial body weight to Terminal body weight)
22.44 to 22.58 (25%: from Initial body weight to Terminal body weight)
20.37 to 20.39 (50%: from Initial body weight to Terminal body weight)
19.56 to 19.63 (100%: from Initial body weight to Terminal body weight)

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
The test item 4,4´-bis(2-methoxystyryl)-1,1´-biphenyl is not considered a skin sensitizer under the test conditions of this study.
Executive summary:

The sensitization potential of4,4´-bis(2-methoxystyryl)-1,1´-biphenylwas evaluated using the Local Lymph Node Assay (LLNA). The LLNA has been developed to determine the allergic contact sensitization potential of chemicals.

Based on the recommendations of the OECD Guideline 429, the test item was suspended in Acetone/Olive Oil, 4:1 (v/v). The positive control (a-Hexylcinnamic aldehyde) (25%) was dissolved in the same vehicle.

The Pre-screen testwas performed using the dose of 100 %. Based on the observations recorded in the Pre-screen tests, the concentration of 100 % was selected as top dose for the main test.

Five female mice (CBA/Ca) per group were topically exposed (dorsum of both ears) to the test item at concentrations of 25%, 50% and 100%, to the positive control and to the vehicle only. Lymphocyte proliferation was measured using incorporation of radioactive125I-iododeoxyuridine and 10-5M fluorodeoxyuridinein the draining lymph nodes. The radioactive incorporation was expressed as disintegrations per minute (DPM)/pooled treatment group and compared with DPM value from the vehicle control group and expressed as the Stimulation Index (SI).

After application of the test itemat three concentrations (25%, 50% and 100% w/v) the animals did not show visible clinical symptomsof either local irritation or systemic toxicity.

In this study the Stimulation Indices (SI) of0.71, 0.91 and 0.81 were determined with the test item at concentrations of 25%, 50%, and 100% inAcetone/Olive Oil 4:1, respectively. The EC3 value could not be calculated, since none of the tested concentrations induced a SI greater than the threshold value of 3.  

The test item4,4´-bis(2-methoxystyryl)-1,1´-biphenylis not considered a skin sensitizer under the test conditions of this study.