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Administrative data

Endpoint:
eye irritation: in vitro / ex vivo
Remarks:
In vitro
Type of information:
experimental study
Adequacy of study:
key study
Study period:
25 April 2017 to 28 April 2017
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions

Data source

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

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 492 (Reconstructed Human Cornea-like Epithelium (RhCE) Test Method for Identifying Chemicals Not Requiring Classification and Labelling for Eye Irritation or Serious Eye Damage)
Deviations:
no
GLP compliance:
no
Remarks:
See'Justification for type of information'

Test material

Constituent 1
Reference substance name:
Active enzyme protein of sedolisin (EC no. 848318-58-1, CAS no. 814-114-0, EC name Sedolisin, Enzyme class no 3.4.21.100 )
Molecular formula:
Not available
IUPAC Name:
Active enzyme protein of sedolisin (EC no. 848318-58-1, CAS no. 814-114-0, EC name Sedolisin, Enzyme class no 3.4.21.100 )
Constituent 2
Reference substance name:
Protein as a constituent of enzyme deriving from the fermentation or extraction process
Molecular formula:
Not available
IUPAC Name:
Protein as a constituent of enzyme deriving from the fermentation or extraction process
Constituent 3
Reference substance name:
Inorganic salts as a constituent of enzyme deriving from the fermentation or extraction process
Molecular formula:
Not available. See remarks.
IUPAC Name:
Inorganic salts as a constituent of enzyme deriving from the fermentation or extraction process
Constituent 4
Reference substance name:
Carbohydrates constituent of enzyme deriving from the fermentation or extraction process
Molecular formula:
Not available. See remarks.
IUPAC Name:
Carbohydrates constituent of enzyme deriving from the fermentation or extraction process
Constituent 5
Reference substance name:
Lipids as a constituent of enzyme deriving from the fermentation or extraction process
Molecular formula:
Not available. See remarks.
IUPAC Name:
Lipids as a constituent of enzyme deriving from the fermentation or extraction process
Test material form:
liquid
Details on test material:
- Lot/batch No.: PPF38268
- Expiration date of the lot/batch: 22-april-2025
- Storage condition of test material: Frozen

Test animals / tissue source

Species:
human
Strain:
other: SkinEthic Human Corneal Epithelium (HCE)

Test system

Vehicle:
unchanged (no vehicle)
Controls:
yes, concurrent positive control
yes, concurrent negative control
Amount / concentration applied:
30 µL of undiluted test substance.
Duration of treatment / exposure:
30 min
Duration of post- treatment incubation (in vitro):
30 minutes ± 2 minutes at 37°C, 5% CO2, ≥ 95% humidity.
Number of animals or in vitro replicates:
3
Details on study design:
Performed according to In vitro Prediction Assay for Acure Ocular Irritation of Liquid Chemicals SkinEthic Human Corneal Epithelial Model (SkinEthicTM HCE).
Treatment:
Apply 30 μL of test liquid to apical side of insert.
Treat tissue with adapted intervals according to the rinsing-off intervals (3 min).
Treat negative control with PBS
Treat positive control with Methyl Acetate
Incubate for 30 minutes ± 2 minutes at 37°C, 5% CO2, ≥ 95% humidity.

Test liquid were rinsed away after exposure:
Adjust distribution to 10 mL and rinse every tissue 2 times (20 mL).
Transfer the rinsed tissue to a new well containing 750 µL fresh maintenance medium (2nd line on the plate).
Apply 750 µL fresh maintenance medium apically to the tissue.
Make sure no air bubbles are trapped underneath the inserts.
Incubate for 30 minutes ± 2 minutes at 37°C, 5% CO2, ≥ 95% humidity.

Results and discussion

In vitro

Results
Irritation parameter:
in vitro irritation score
Value:
0
Vehicle controls validity:
not applicable
Negative controls validity:
valid
Positive controls validity:
valid

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
Sedolisin, batch PPF38268 tested in human corneal epithelial cells revealed a viability of 88.1%. Therefore, it is not irritating to the eye (No Category), when tested in the present model under the test conditions.
Executive summary:

The Human Corneal Epithelium Assay (HCE) from SkinEthic (France) was used to assess the eye irritation potential of the test substance. The MTT conversion assay, which measures the NAD(P)H-dependent microsomal enzyme reduction of MTT to a blue formazan precipitate, was used to assess cellular metabolism after exposure to a test substance for the defined exposure time.

The test system and procedure described in In Vitro Prediction Assay for Acute Ocular Irritation of Liquid Chemicals SkinEthic Human Corneal Epithelial Model (SkinEthicTM HCE) was used.

Sedolisin, batch PPF38268 tested in human corneal epithelial cells revealed a viability of 88.1%. Therefore, it was not irritating to the eye (No Category), when tested in the present model under the test conditions.

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