Registration Dossier

Administrative data

Endpoint:
skin irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report Date:
2017

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Product no. B4429, Lot no. SLBK5235V
- Expiration date of the lot/batch: 30 April 2018
- Purity test date: na

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

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: no

In vitro test system

Test system:
human skin model
Source species:
human
Vehicle:
unchanged (no vehicle)
Details on test system:
TISSUE
- Model used: EPISKIN(TM) Small Model (SM)
- Tissue batch number(s): 16-EKIN-026
- Production date: not specified
- Shipping date: not specified
- Delivery date: not specified
- Date of initiation of testing: 29.06.2016

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: room temperature
- Temperature of post-treatment incubation (if applicable): 37°C for 42 hours

REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: 25 mL PBS solution used to thoroughly rinse the tissue once
- Observable damage in the tissue due to washing: none
- Modifications to validated SOP: none

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 2 mL of 0.3 mg/mL MTT per well
- Incubation time: 3 hours +/- 5 min
- Spectrophotometer: not specified
- Wavelength: 570 nm
- Filter: not specified
- Filter bandwidth: not specified
- Linear OD range of spectrophotometer: not specified

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA: secured to batch release quality control

NUMBER OF REPLICATE TISSUES: 3

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE: no direct MTT interference

NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: 1

PREDICTION MODEL / DECISION CRITERIA (choose relevant statement)
- The test substance is considered to be non-irritating to skin if the viability is greater than or equal to 50%
Control samples:
yes, concurrent negative control
yes, concurrent positive control
Amount/concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 10 mg (to moistened skin)

NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 10 µL

POSITIVE CONTROL
- Amount(s) applied (volume or weight): 10 µL
- Concentration (if solution): 5% aq.
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:
1
Value:
96
Vehicle controls validity:
not applicable
Negative controls validity:
valid
Positive controls validity:
valid
Other effects / acceptance of results:
- OTHER EFFECTS:
- Visible damage on test system: none
- Direct-MTT reduction: no
- Colour interference with MTT: no

DEMONSTRATION OF TECHNICAL PROFICIENCY:

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
- Range of historical values if different from the ones specified in the test guideline: -

Any other information on results incl. tables

In an experimental study according to OECD test guideline 439 under GLP conditions, the test substance did not reduce cell viability (96% with an SD of 14.52).

Therefore, it is considered not irritating to skin.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Executive summary:

The skin irritation potential of the test substance was investigated in an experimental study according to OECD test guideline 439 under GLP conditions using the EPISKINTMSM. The EPISKINTMSM is provided in kits, which are manufactures according to defined quality assurance procedures.

The testing laboratory, which had demonstrated its technical proficiency in conducting the test, received the kit used for testing in good order.

The test substance was applied (10 mg) for 15 minutes to three tissues. After thorough rinsing, the treated tissue were incubated for 42h at 37°C. Then the MTT test was performed, followed by formazan extraction and cell viability measurement with a spectrometer at 570 nm.

Concurrent negative and positive control fulfilled all acceptance criteria, as did the test substance, which did not directly reduce MTT.

With a mean cell viability of 96% (as compared to the negative control), the test substance did only very marginally reduce tissue viability.

Using the standard prediction (with a threshold of 50%) the test substance is considered to be not skin irritating and does not require to be classified (UN GHS 'No Category').