Registration Dossier

Administrative data

Endpoint:
skin corrosion: 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:
2018

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
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
Test material form:
solid

In vitro test system

Test system:
human skin model
Vehicle:
other: Dulbecco’s Phosphate Buffered Saline (DPBS buffer)
Details on test system:
The test system is a commercially available EpiDermTM-Kit, procured by MatTek.
The EpiDermTM tissue consists of human-derived epidermal keratinocytes which have been cultured to form a multi-layered, highly differentiated model of the human epidermis. It consists of organized basal, spinous and granular layers, and a multi-layered stratum corneum containing intercellular lamellar lipid layers representing main
lipid classes analogous to those found in vivo. The EpiDermTM tissues are cultured on spe-cially prepared cell culture inserts.
Control samples:
yes, concurrent vehicle
yes, concurrent positive control

Test system

Amount / concentration applied:
Tissue Amount
1 26.2 mg
2 27.2 mg
3 26.9 mg
Details on study design:
One plate (3 tissues) was used as negative control; each tissue was treated with 30 µL DPBS buffer, a nylon mesh was added in order to ensure sufficient contact with the tissue surface.
One plate was used as positive control; each tissue was treated with 30 µL 5% SDS-solution, a nylon mesh was added in order to ensure sufficient contact with the tissue sur-face.
One plate was used for treatment with the test item:
The tissues were wetted with 25 µL DPBS buffer before applying the test item and spread-ing it to match the tissue size.

Tissues were dosed in 1-minute-intervals. After dosing the last tissue, all plates were transferred into the incubator for 35 minutes at 37 ± 1°C and 5.0 ± 0.5% CO2.
1 hour after the first application, the inserts were removed from the plates using sterile forceps and rinsed immediately in 1-minute-intervals.
After rinsing, each tissue was blotted with sterile cellulose tissue and then transferred into a new 6-well-plate with fresh assay medium (0.9 mL).
Then, the tissues were set in the incubator for 23 hours and 20 minutes at 37 ± 1°C and 5.0 ± 0.5% CO2.

Results and discussion

In vitro

Resultsopen allclose all
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
tissue 1
Value:
70.4
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
tissue 2
Value:
80.5
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
tissue 3
Value:
69.8
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
mean value
Value:
73.5
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
The test item L-Tyrosine, 3,5-dinitro- (wet product) is considered as non-irritant to skin.
After the treatment, the mean value of relative tissue viability was reduced to 73.5%. This value is above the threshold for skin irritation (50%).
The optical density of the negative control was well within the required acceptability criteri-on of 0.8 ≤ mean OD ≤ 2.8.
The positive control has met the acceptance criterion too, for thus ensuring the validity of the test system.
Variation within replicates was within the accepted range for negative control, positive con-trol and test item (required: ≤ 18%).
For these reasons, the result of the test is considered valid.
Executive summary:

Three tissues of the human skin model EpiDermTMwere treated with L-Tyrosine, 3,5-dinitro- (wet product) for 60 minutes.

The test item was applied directly to each tissue and spread to match the tissue size (0.63  cm2; as indicated by the supplier).

DPBS-buffer was used as negative control and 5% SDS solution was used as positive control.

After treatment with the negative control, the mean absorbance value was within the required acceptability criterion of 0.8 ≤ mean OD ≤ 2.8, OD was 1.5. The positive control showed clear irritating effects. The mean value of relative tissue viability was reduced to 2.8 % (required: ≤ 20%).

The variation within the tissue replicates of negative control, positive control and test item was acceptable (required: ≤ 18%).

 

After the treatment with the test item, the mean value of relative tissue viability was reduced to 73.5 %. This value is above the threshold for skin irritation potential (50%). Test items that induce values above the threshold of 50% are considered non- irritant to skin