Cocoa, powd., alkalized

Eye irritation

Administrative data

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study performed according to GLP.

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Cross-reference 2

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying Ocular Corrosives and Severe Irritants)

Principles of method if other than guideline:

The BCOP test method is an organotypic model that provides short-term maintenance
of normal physiological and biochemical function of the bovine cornea in vitro.
Damage by the test article is assessed by quantitative measurements of changes in
corneal opacity and permeability. Both measurements are used to calculate an In Vitro

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Cocoa Ext.

IUPAC Name:

Cocoa Ext.

Constituent 2

Reference substance name:

Cocoa, ext.

EC Number:

283-480-6

EC Name:

Cocoa, ext.

Cas Number:

84649-99-0

IUPAC Name:

283-480-6

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Details on test material:

- Name of test material (as cited in study report): High Fat Cocoa powder
- Substance type: UVCB
- Physical state: Brown powder
- Composition of test material, percentage of components:
- Purity test date:
- Lot/batch No.: ESTS 165/12
- Expiration date of the lot/batch: 4th Sept. 2015
- Stability under test conditions:
- Storage condition of test material:

Test system

Vehicle:

physiological saline

Remarks:

0.9% sodium chloride solution

Amount / concentration applied:

The test article was prepared as a 20% w/v solution in 0.9% sodium chloride solution.
The formulation was prepared on the day of use.

Duration of treatment / exposure:

4hours

Details on study design:

Upon arrival at the test facility, the corneas were excised from the eyes and loaded onto the specifically designed holders. Both chambers of each holder were filled with pre-warmed Eagle’s Minimal Essential Medium (EMEM) (posterior chamber first), ensuring that no bubbles were formed. The holders were incubated at 32°C ± 1°C for at least 1 hour. After the incubation, the media was removed from both the anterior and posterior chambers. Fresh media was added to the posterior chamber first and then the anterior chamber (this media replacement order ensured the cornea retained
its natural curvature as much as possible). The opacity of each cornea was measured using an opacitometer. Any corneas found to have scratches or increased neovascularization or an opacity of >7 opacity units when examined prior to treatment were discarded.

Treatment of corneas:
A volume of 750 μL of the test article formulation was applied to each of three corneas followed by a four-hour incubation at 32°C. After this incubation, each cornea was washed with media containing phenol red (as a pH indicator) until this indicator showed no pH effect occurring (and demonstrating that the test article had been removed successfully). The corneas were then washed once in media without phenol red, the opacities measured and the anterior chamber emptied. For the permeability endpoint, 1 mL of sodium fluorescein (5 mg/mL solution) was added into the anterior chamber and the corneas were incubated in the vertical position for 1.5 hours ± 5 minutes. Following this period, the media in the posterior chamber was
removed and held in a labelled tube. Three x 350 μL aliquots of this media (per cornea) were analysed for optical density at 490 nanometers (OD490).
A volume of 750 μL of the negative or positive control was similarly applied to further groups of three corneas. These groups were subject to the procedures detailed above.

Results and discussion

In vivo

Results

Irritation parameter:

cornea opacity score

Basis:

animal: Bovine corneal

Score:

ca. 5.54

Remarks on result:

other: not considered to be corrosive or severely irritating to the eye

Applicant's summary and conclusion

Interpretation of results:

other: Not corrosive or severely irritating to the eye

Remarks:

Criteria used for interpretation of results: EU

Conclusions:

The test article, High fat cocoa powder, produced an IVIS score of 5.54 and was considered not to be corrosive or severely irritating to the eye.

Executive summary:

This study was conducted to determine whether the test article causes corrosion or severe irritation to excised bovine corneas using the bovine corneal opacity and permeability (BCOP) assay.The test article was prepared as a 20% w/v solution in 0.9% sodium chloride solution. A volume of 750 µL of the test article formulation was applied to each of three corneas followed by a four-hour incubation at 32°C. After this incubation, each cornea was washed with media containing phenol red followed by media without phenol red, the opacities measured and then the anterior chamber emptied. For the permeability endpoint, sodium fluorescein solution was added into the anterior chamber and the corneas were incubated in the vertical position for 1.5 hours ± 5 minutes.

The test article, High Fat Cocoa Powder, produced an IVIS score of 5.54 and was considered not to be corrosive or severely irritating to the eye.