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Toxicological information

Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation, other
Remarks:
an in vivo skin sensitization (non-LLNA) study was performed based on ambigious results in the in vitro test battery
Type of information:
experimental study
Adequacy of study:
key study
Study period:
23.03.2016 - 07.07.2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2016
Report Date:
2016

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
The aim of the test was to assess the contact sensitizing potential of the test item in follow-up of 3 in vitro skin sensitisation studies that could not give a clear result. Structural similar substances gave stimulation indices (SI) > 3 which were considered not relevant for humans. For that reason, GPMT was performed in order to avoid false positive results.

Test material

Reference
Name:
Unnamed
Type:
Constituent

In vivo test system

Test animals

Species:
guinea pig
Strain:
other: CrlOri:Ha
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Weight at study initiation: male: 255 - 282 g, female: 337 - 395 g
- Age at study initiation: male: 4 weeks, female: 5 weeks
- Housing: 1 animal in stainless wire mesh cages, 210W×350D×180H (mm)
- Diet: ad libitum, Pellet feed for experimental animals (Purina experimental diet for guinea pig 38065)
- Water: ad libitum, public tap water
- Acclimation period: 12 days
- Microbiological status of animals, when known: SPF

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20.0–23.8
- Humidity (%): 33.9–52.1
- Air changes (per hr): 10–15 times/h
- Photoperiod (hrs dark / hrs light): 12 hour light/dark cycle (7 AM to 7 PM via automated timer), 150-300 Lux

Study design: in vivo (non-LLNA)

Induction
Route:
intradermal and epicutaneous
Vehicle:
olive oil
Concentration / amount:
first induction 25% and second induction 100%
Day(s)/duration:
48 hours
Adequacy of induction:
highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically
Challenge
No.:
#1
Route:
epicutaneous, occlusive
Vehicle:
unchanged (no vehicle)
Concentration / amount:
100%
Day(s)/duration:
24 hours
No. of animals per dose:
10
Details on study design:
PRELIMINARY TEST:
Intradermal Injection and topical application:
- Solvent: olive oil
- Concentration: 100, 50, 25, 12.5, 6.25, and 3.13%
- Amount animals: 2 x 2 (2 intradermal dosing and 2 topical application)
- Recordings: 48 h after treatment.
- Application method:
-- Sites: shoulder was clipped and shaved, 6 sites: 3 on the left and 3 on the right side
-- Concentration: 100, 50, 25, 12.5, 6.25 and 3.13%
-- Procedure: closed patch by means of an occlusive bandage
-- Recordings: 24 and 48h post application
-- Patches: 2×2 cm patch, animals were wrapped with adhesive tape and secured with surgical tape

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 2
First induction: Intradermal
Second induction and challenge: Dermal (topical application): on Day 7 after first induction
- Exposure period: 48 h
- Site: 2 x 4 cm clipped and shaved, application sites were pretreated with 0.5 mL of 10% SDS
- Concentration: first induction: 25%; second induction: 100%
- Test group:
1. WFI-FCA emulsion
2. 25% Test substance
3. 25% Test substance-FCA emulsion
- Control group:
1. WFI-FCA emulsion
2. Olive oil
3. Olive oil-FCA emulsion

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge:
- Site: 2 x 2 cm clipped and shaved
- Concentrations:100%
- Evaluation (hr after challenge): 24 hours
- Test and control group: 100% test substance was applied to the left challenge sites of the test substance and control groups, while at the right challenge sites of the test substance and control groups olive oil was applied

EVALUATION
- Clinical signs: daily
- Body weight: on day 0, once weekly and on day 24
- Skin sensitisation: see scoring system

SCORING SYSTEM: Magnusson and Kligman
No reaction: 0
Scattered mild redness: 1
Moderate & diffuse redness: 2
Intense redness & swelling: 3

STATISTICAL ANALYSIS: Statistical analysis was not performed. Mean scores and values are presented.
Challenge controls:
Olive oil
Positive control substance(s):
yes
Remarks:
1-Chloro-2,4-dinitrobenzene (CDNB): A positive control group was not set in this study, a positive control study was conducted periodically

Results and discussion

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
100%
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
no indication of skin sensitisation
Reading:
1st reading
Hours after challenge:
48
Group:
test group
Dose level:
100%
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
no indication of skin sensitisation

Any other information on results incl. tables

CLINICAL OBSERVATIONS: No abnormal signs or symptoms were observed in any animal of any group throughout the course of the study.

BODY WEIGHTS: All animals exhibited normal body weight gain.

POSITIVE CONTROL: The sensitization rate of CDNB was 100% and the sensitization grade was classified to be ‘V (Extreme)’. Consequently, the study was considered to be valid.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
No animal showed an allergic response, the test article is not sensitising.
Executive summary:

The aim of the current study was to assess the skin sensitizing potential of the test item, by means of a maximization test using guinea pigs according to OECD 406 and GLP.

 

In a preliminary test topical application did not reveal any evidence of adverse skin reactions, therefore, the induction sites in the main test for each group was pretreated with 10% sodium dodecyl sulfate prior to the second induction.

There were 2 groups, one for the test substance, this group consisted of 10 animals, while the second group was the control group made of 5 animals. In the test group, a concentration of 25% of the test substance was injected intradermally for the first induction. The second induction was conducted with 100% of the test substance. The sites were occluded for 48 hours, whereafter the challenge was conducted with 100% of the test substance. Hereafter, the sites were occluded for 24 hours. In the control group, the first and second inductions was done with olive oil and the challenges with the 100% of the test substance and olive oil.

Both in the control and the test group no skin reactions such as redness or swelling were observed in any animal at 24 and 48 hours after the challenge patch was removed. During the observation period, no abnormalities in clinical signs or body weight gain were observed in any animal of any group. Based on the result of this study, the test substance did not produce skin sensitization under the conditions of this study.

Therefore, the test article does not have to be classified as sensitising.