Registration Dossier

Administrative data

Endpoint:
skin irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
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 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
Version / remarks:
2015
Deviations:
no
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Lot # 20003, unit 0022
- Expiration date of the lot/batch: 30 March, 2018
- Purity test date: 30 March, 2016

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: At room temperature

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: None
- Preliminary purification step (if any): None

FORM AS APPLIED IN THE TEST: The test article was applied to the tissue undiluted.

In vitro test system

Test system:
human skin model
Source species:
human
Cell type:
non-transformed keratinocytes
Cell source:
other: Source specified as "Adult donors, Batch 16-EKIN-025"
Details on animal used as source of test system:
SOURCE ANIMAL
- Source: Adult human donors
Justification for test system used:
The test system has been validated an included in the OECD protocol.
Vehicle:
unchanged (no vehicle)
Details on test system:
SKIN DISC PREPARATION
- Procedure used: Tissues were purchased from EpiSkin
TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 37 C
- Temperature of post-treatment incubation (if applicable): 37 C
REMOVAL OF TEST MATERIAL AND CONTROLS
- Number of washing steps: 1
DYE BINDING METHOD
- Dye used in the dye-binding assay: MTT
- Spectrophotometer: TECAN Infinite M200 Pro Plate Reader
- Wavelength: 570 nm
NUMBER OF INDEPENDENT TESTING RUNS / EXPERIMENTS TO DERIVE FINAL PREDICTION: 3
PREDICTION MODEL / DECISION CRITERIA
A test article is considered an irritant in the skin irritation test if:
- The relative mean issue viability of three individual tissues after 156 minutes of exposure to the test article and 42 hours post incubation is less than or equal to 50% of the mean viability of the negative controls.

A test article is considered a non-irritant in the in vitro skin irritation test if:
-The mean relative tissue viability of three individual tissues after 15 minutes of exposure to the test article and 42 hours post incubation is greater than 50% of the mean viability of the negative controls.
Control samples:
yes, concurrent negative control
yes, concurrent positive control
Amount/concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 25 uL

NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 25 uL

POSITIVE CONTROL
- Amount(s) applied (volume or weight): 25 uL
- Concentration (if solution): 5%
Duration of treatment / exposure:
15 minutes
Duration of post-treatment incubation (if applicable):
42 hours
Number of replicates:
3

Results and discussion

In vitro

Results
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
Mean
Value:
ca. 108
Negative controls validity:
valid
Positive controls validity:
valid
Other effects / acceptance of results:
- OTHER EFFECTS:
- Visible damage on test system: None
- Colour interference with MTT: None

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: Yes
- Acceptance criteria met for positive control: Yes
- Acceptance criteria met for variability between replicate measurements: Yes, the standard deviation value of the percentage viability of three tissues treated identically was less than 11%, indicating that the test system functioned properly.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
Based on the results of the study, the test article is not a dermal irritant.
Executive summary:

The dermal irritation potential of the test article was evaluated using a three dimensional human skin model (EPISKIN Small Model). The study was conducted according to OECD 439 (2015) in compliance with OECD GLP (1997). The test article was applied undiluted (25 uL) directly on top of the skin tissue for a 15 minute exposure period at 37 C. Negative (PBS) and posivite (5% SDS) controls were tested concurrently. All exposures were run in triplicate. After washing, the tissue it was incubated at 37 C for 42 hours. Following incubation, tissue viability was measured by formazan production from MTT and compared to the negative control tissue viability. The relative mean tissue viability obtained after 15 minutes of treatment with the test article compared to the negative control tissues was 108%. Based on the results of the study, the test article is not a dermal irritant.