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Administrative data

Endpoint:
eye irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
26 February 2015 to 10 March 2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
Compliant to GLP and testing guidelines; coherence between data, results and conclusion.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2015
Report date:
2015

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying Ocular Corrosives and Severe Irritants)
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: OECD series on testing and assesment no. 160
Qualifier:
according to guideline
Guideline:
other: Use of an alternate testing framework for classification of eye irritation potential of EPA pesticide products - Office of Pesticide Programs, U.S. Environmental Protection Agency (31 May 2013).
GLP compliance:
yes

Test material

Constituent 1
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder
Details on test material:
Test item: Acid Red EAY 9656

Test animals / tissue source

Species:
cattle
Strain:
not specified
Remarks:
bovine cornea
Details on test animals or tissues and environmental conditions:
Test system: isolated corneas from the eyes of freshly slaughtered bovine.
Age of animals: 6-12 months

Transport solution: Hanks balanced salt solution (HBSS) Modified supplemented with Penicillin sulphate and Streptomycin sulphate.
Transport conditions: in transport solution at approximately 4°C.

Test system

Vehicle:
physiological saline
Controls:
yes, concurrent vehicle
Amount / concentration applied:
The test item was spread on each epithelial surface of three idoneous bovine corneas as supplied (150 mg/cornea); 0.75 mL of physiological saline was then added on each cornea in order to better distribute the substance on the cornea surface (final concentration on cornea = 200 mg/mL, approximately 20% w/v)
Number of replicates: 3
Duration of treatment / exposure:
Corneas were exposed in horizontal position for 4 hours ± 10 minutes, incubated in a liquid bath at 32 ± 1°C.
There was no post-exposure period.
Details on study design:
Positive control item: corneas treated 20% (w/v) Imidazole (Sigma, batch no. 0001422794) in physiological saline (0.9% NaCl) (Baxter, batch no. 13D0406).
Negative control item: corneas treated with physiological saline (0.9% NaCl) (Baxter, batch no. 13D0406).
Volume of treatment: 0.75 mL
Number of replicates: 3

Results and discussion

In vitro

Resultsopen allclose all
Irritation parameter:
cornea opacity score
Remarks:
mean score of three corneae
Value:
22.7
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Irritation parameter:
fluorescein retention score
Remarks:
mean permeability OD490 score
Value:
0.186
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Irritation parameter:
in vitro irritation score
Remarks:
IVIS = mean opacity score + (15 x mean permeability OD490 score)
Value:
25.5
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Other effects / acceptance of results:
The mean opacity detected with an opacitometer at the end of the test item exposure period was 22.7. This was confirmed at the macroscopic observation in which slight opacity of the treated corneas was noted. Plentiful residual test item, adhering on the surface of all treated corneas, was observed. Slight increases in cornea permeability were observed after treatment with the test item, with a mean permeability OD490 value of 0.1863.

The following results were obtained in the negative control:
– mean opacity value of the negative control = 0.00
– mean permeability value of the negative control = 0.0077

The following results were obtained in the positive control:
– IVISpositive control = 72.4

Based on the evaluation criteria the experiment was accepted as valid.

Any other information on results incl. tables

The following results were obtained:

– mean opacity value of the negative control = 0.00

– mean permeability value of the negative control = 0.0077

– IVISpositive control = 72.4

Based on the stated criteria (§ 4.6.2) the experiment was accepted as valid.

The calculated in vitro irritancy score for the test item was:

– IVIStest item = 25.5

indicating that the test item cannot be clearly classified.

Applicant's summary and conclusion

Interpretation of results:
study cannot be used for classification
Remarks:
Migrated information
Conclusions:
The test item Acid Red EAY 9656 cannot be clearly classified.
Executive summary:

The potential of the test item, Acid Red EAY 9656, to cause corrosion/severe irritation by using the Bovine Corneal Opacity and Permeability (BCOP) assay, was examined in agreement with OECD Guideline no. 437 (adopted on 26 July 2013), the Guidance Document OECD series on testing and assessment no. 160 and the method described in the document issued by U.S. EPA/OPP (31 May 2013). The test item was spread on each epithelial surface of three idoneous bovine corneas as supplied (150 mg/cornea); 0.75 mL of physiological saline was then added on each cornea in order to better distribute the substance on the cornea surface (final concentration on cornea = 200 mg/mL, approximately 20% w/v); an exposure period of 4 hours ± 10 minutes was used. The mean opacity detected with an opacitometer at the end of the test item exposure period was 22.7. This was confirmed at the macroscopic observation in which slight opacity of the treated corneas was noted. Plentiful residual test item, adhering on the surface of all treated corneas, was observed. After the determination of opacity, the epithelial surface was treated with a 0.5% solution of sodium fluorescein in DPBS for 90 minutes, to investigate alteration in cornea permeability. The calculated mean permeability OD490 value of the corneas treated with the test item was 0.1863. The calculated in vitro irritancy score (IVIS) for the test item was 25.5. Positive and negative controls [a 20 % (w/v) Imidazole solution in physiological saline and physiological saline alone, respectively] were concurrently tested in similar conditions and gave the expected results. According to the OECD Guideline no. 437, the test item can not be clearly classified.

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