Registration Dossier

Administrative data

skin irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
September 11, 2018 - September 15, 2018
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
OECD 439

Data source

Reference Type:
study report
Report Date:

Materials and methods

Test guideline
according to
OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
GLP compliance:
yes (incl. certificate)

Test material

Test material form:
solid: particulate/powder

In vitro test system

Test system:
human skin model
Source species:
Cell type:
non-transformed keratinocytes
Cell source:
other: reconstructed human epidermis (SkinEthicTM RHE)
Details on animal used as source of test system:
The test system used for the in vitro skin irritation test was reconstructed human epidermis (SkinEthicTM RHE) as recommended by the OECD 439 guideline. The SkinEthicTM RHE model consists of normal human keratinocytes cultured for 17-days on an insert of 0.5 cm2 polycarbonate filter at the air-liquid interface in a chemically defined growth medium. The cells form a multilayered, highly differentiated and stratified epidermis model of the human epidermis that consists of main basal, supra basal, spinous and granular layers and a functional stratum corneum.
Reconstructed human epidermis tissues, SkinEthicTM RHE model, were procured from SkinEthic Laboratories, Episkin 4, Rue Alexander Fleming, 69366 Lyon Cedex 07, France; were used in the study (Lot N°18-RHE-111)
unchanged (no vehicle)
Control samples:
yes, concurrent negative control
yes, concurrent positive control
Amount/concentration applied:
16 ± 2 mg of test item/0.5 cm2
Duration of treatment / exposure:
incubated at room temperature for 42 minute exposure
Duration of post-treatment incubation (if applicable):
at 37 ± 1°C in 5 ± 1% CO2 in a humidified incubator for 42 hours
Number of replicates:
Three tissues for each treatment (test item, Negative control and Positive control)

Results and discussion

In vitro

Irritation / corrosion parameter:
% tissue viability
ca. 90.67
Vehicle controls validity:
Negative controls validity:
Positive controls validity:
Remarks on result:
no indication of irritation

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Based on the results of this study, the classification for the tesi item is as mentioned below:
Globally Harmonized System of Classification and: No Category (Non Skin Irritant)
Executive summary:

This study was performed to evaluate the non-irritant and irritant potential of CZ-HABI using reconstructed human epidermis (RHE) tissue in accordance with the United Nations Globally Harmonized System of Classification and Labelling of Chemicals (UN GHS).

Tissues were exposed tothenegative control (Dulbecco’s Phosphate Buffered Saline (DPBS)), positive control (sodium dodecyl sulfate,5% aqueous (SDS)) and the test ithem in triplicate for 42 minutes at room temperature.

The mean cell viability in tissues treated with the test item was 90.67% after 42 minutes exposure. A significant reduction in percent cell viability was not observed in treated tissues when compared with that of the concurrent negative control.

The Optical density (OD) values for the negative control replicates were between 1.274 and 1.305, against the guideline requirement of ≥ 0.8 and ≤ 3.0 (≥ 1.2 as per SkinEthic SOP). The OD of the blankwasbetween0.042and0.045which met the guideline requirement of OD< 0.1.The positive control showed a 1.52% cell viability, against the acceptance criteria of <40% for the SkinEthic RHE model, when compared with that of the concurrent negative control. Variation between tissue replicates (i.e., CV% value) was 0.18% for the test item group, 5.92% for positive control and 0.77% for negative control against the guideline requirement of ≤ 18%, which demonstrate the efficiency of the test system, SkinEthicTMRHE model. All criteria for a valid study were met as describe in the study plan.

Based on results of this study, the classification for the substance is as follows:

Globally Harmonized System of Classification and  Labeling of Chemicals:         No Category (Non Skin Irritant)