Registration Dossier

Administrative data

Endpoint:
skin irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From 2017-04-18 to 2017-05-05
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)
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Test material form:
solid: particulate/powder
Specific details on test material used for the study:
STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: room temperature


In vitro test system

Test system:
human skin model
Source species:
human
Vehicle:
unchanged (no vehicle)
Details on test system:
RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE

- Model used: Episkin SA, RHE/S/17
- Tissue batch number(s): 17-RHE-050
- Delivery date: 2017-03-03
- Date of initiation of testing: 2017-03-03
- Killed tissues were frozen the 2017-03-14 and defrost on the treatment day.

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: room temperature
- Temperature of post-treatment incubation : 37°C

REMOVAL OF TEST MATERIAL AND CONTROLS
- Number of washing steps: 1 (25 x 1 mL of DPBS (VWR, Batch No. 6MB235). The rinsed tissues were checked for any coloration and noted to be blue and to present residue of test item).
- Observable damage in the tissue due to washing: none

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 300 μL of a MTT solution at 1.0 mg/mL
- Incubation time: 3 hours
- Spectrophotometer: ELx800 absorbance microplate reader
- Wavelength: 570 nm
Control samples:
yes, concurrent negative control
yes, concurrent positive control
Amount/concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 16 mg

NEGATIVE CONTROL
(DPBS – VWR - Batch No. 6MB235)

POSITIVE CONTROL
- Amount(s) applied (volume or weight):
- Concentration (if solution): solution of SDS at 5%. The 5% SDS solution was prepared by weighing 0.5 g of SDS (SIGMA Batch No. STBF1623V) in a 10 mL volumetric flask qsp 10 mL of distilled water.
Duration of treatment / exposure:
42 min
Duration of post-treatment incubation (if applicable):
42 hr
Number of replicates:
3 living and 2 killed Reconstructed Human epidermis (SkinEthic RHE® model) previously moistened with 10 μL of distilled water.

Results and discussion

In vitro

Results
Irritation / corrosion parameter:
% tissue viability
Value:
ca. 80.2
Negative controls validity:
valid
Positive controls validity:
valid
Other effects / acceptance of results:
OTHER EFFECTS:
- Visible damage on test system: no
- Direct-MTT reduction: no
- Colour interference with MTT: no

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
In accordance with the Regulation EC No. 1272/2008, the test item ADDITIVE 8020 has to be considered as Non-irritant to skin. I
Executive summary:

The aim was to evaluate the possible irritating effects of the test item after topical application on in vitro human reconstructed epidermis (SkinEthic RHE® model).

The test item was applied at the dose of 16 mg during 42 minutes to 3 living and 2 killed Reconstructed Human epidermis (SkinEthic RHE® model) previously moistened with 10 μL of distilled water. The application was followed by a rinse with 25 mL of DPBS and a 41 hours and 55 minutes post-incubation period at 37°C, 5% CO2. Cell viability was then measured by enzymatic conversion of the vital dye MTT into a blue formazan salt that was quantitatively measured after extraction from tissues.

The experimental protocol was established in accordance with O.E.C.D. Test Guideline No. 439 adopted 28 July 2015 and the Test method B.46 of Council regulation No. 761/2009 dated 23 July 2009 (EU Journal L220) - ATP Council regulation No. 440/2008 of 30 May 2008 (E.U. Journal L142).

The mean corrected percent viability of the treated tissues was 80.2%, versus 1.7% in the positive control (5% Sodium Dodecyl Sulfate).

In accordance with the Regulation EC No. 1272/2008, the test item has to be considered as Non-irritant to skin. It corresponds to UN GHS No Category.

No hazard statement or signal word is required.