Registration Dossier

Administrative data

Endpoint:
eye irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
December 14, 2018 - December 15, 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
OECD 437

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2018
Report Date:
2019

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying i) Chemicals Inducing Serious Eye Damage and ii) Chemicals Not Requiring Classification for Eye Irritation or Serious Eye Damage)
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Type:
Constituent
Test material form:
solid: particulate/powder

Test animals / tissue source

Species:
cattle
Strain:
not specified
Details on test animals or tissues and environmental conditions:
Source: Deonar Abattoir slaughter house, Mumbai, Maharashtra

Bovine Age: Between 1 to 5 years (the age of the animals was determined based on the teeth count and horn ring count in addition to the Horizontal Diameter of corneas and central corneal thickness)

Transportation Condition: Transported (in a sealed plastic container) under cold condition in Hanks’ Balanced Salt Solution containing antibiotics [e.g., penicillin at 100 IU/mL and streptomycin at 100 μg/mL]

Test system

Amount / concentration applied:
750 μL of the prepared test item [20% (w/v) concentration in corn oil]
Duration of treatment / exposure:
Corneas were exposed for 4 h at 32 ± 1 °C
Number of animals or in vitro replicates:
3 replicates for each conditions (Normal saline, Imidazole 20%, Corn Oil, the test item at 20% in corn Oil)
Details on study design:
This study was performed to evaluate the ocular irritancy of test item as measured by its ability to induce opacity and increase permeability in an isolated bovine cornea. Effects were measured by: 1) decreased light transmission through the cornea (opacity) and 2) increased passage of sodium fluorescein dye through the cornea (permeability). The opacity and permeability assessments of the cornea following exposure to a test item are considered individually and also combined to derive an In Vitro Irritancy Score, which is used to classify the irritancy level of the test item. This study was conducted in compliance with the OECD Principles of GLP (1998).

In this test, damage by the test item is assessed by quantitative measurements of changes in corneal opacity and permeability using an opacitometer and visible light (ELISA reader), respectively. These measurements are used to calculate an in vitro irritancy score (IVIS) to assign an in vitro irritancy hazard classification category of a test item.
Isolated corneas from eyes of freshly slaughtered cattle are used in this test. Corneal opacity is measured quantitatively, as the amount of light transmission through the cornea. Permeability is measured quantitatively as the amount of sodium fluorescein dye that passes across the full thickness of the cornea, as detected in the medium in the posterior chamber. The test item is applied to the epithelial surface of the cornea by addition to the anterior chamber of the corneal holder.

Results and discussion

In vitro

Resultsopen allclose all
Irritation parameter:
cornea opacity score
Value:
ca. 1.21
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Irritation parameter:
fluorescein leakage
Value:
0.028
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Irritation parameter:
in vitro irritation score
Value:
1.62
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 IVIS score, for the corneas treated with 750 μL test item (suspension) at 20% (w/v) concentration in corn oil, were found to be 1.62.
Based on results of this study, the classification for the test item is as follows:
Classification (OECD 437 UN GHS): No Category
Executive summary:

This study was conducted to evaluate theocular irritancy of test item,in compliance with the OECD437 for theBCOP test.

Four sets, each consisting of three corneas, were tested. The first set served as the control and was treated with750 μLnormal saline. The second set served as the positive control and was treated with750 μL20% (w/v) imidazole innormal saline. The third setwas treated with corn oil and served as vehicle control, and the fourth set was treated with750 μLtest item (suspension) at 20% (w/v) concentration in corn oil. Post application, corneas were incubated for approximately 4 h at32 ºC.At the end of incubation, opacity readings were taken.

The post opacity permeability reading was measured by applying1 mL of fluorescein sodium solution
(5 mg/mL)
on to the anterior surface of the cornea followed by incubation for approximately90 min at
32 ºC. At the end of incubation, the Optical Density (OD) was measured at 490 nm from fluid collected from the posterior chamber.

The mean in-vitro Irritancy Score (IVIS) of normal saline, corn oil and750 μL20% (w/v) imidazole innormal saline(positive control) treated corne as were found to be 1.54, 1.20 and 107.04,respectively. The IVIS score for the corneas treated with750 μLtest item (suspension) at 20% (w/v) concentration in corn oil was found to be 1.62.

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

Classification (OECD 437 UN GHS):          No Category