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Toxicological information

Eye irritation

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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:
2015-06-10 to 2015-06-10
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

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:
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
Qualifier:
according to
Guideline:
EU method B.47 (Bovine corneal opacity and permeability test method for identifying ocular corrosives and severe irritants)
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent

Test animals / tissue source

Species:
cattle
Strain:
not specified
Details on test animals or tissues and environmental conditions:
SOURCE OF COLLECTED EYES
- Source: Slaughterhouse, Germany
- Characteristics of donor animals: at least 9 month old
- Storage, temperature and transport conditions of ocular tissue: transported to the laboratory at ambient temperature
- Time interval prior to initiating testing: corneae were isolated on the same day after delivery of the eyes, corneae were directly used in the BCOP test on the same day
- indication of any existing defects or lesions in ocular tissue samples: eyes presenting defects were discarded

Test system

Vehicle:
unchanged (no vehicle)
Controls:
yes, concurrent positive control
yes, concurrent negative control
Amount / concentration applied:
TEST MATERIAL
- Amount applied: 0.75 mL
- Concentration: undiluted
Duration of treatment / exposure:
10 min
Duration of post- treatment incubation (in vitro):
2 h
Number of animals or in vitro replicates:
3
Details on study design:
SELECTION AND PREPARATION OF CORNEAS
Eyes presenting defects such as vascularization, pigmentation, opacity and scratches were discarded.

QUALITY CHECK OF THE ISOLATED CORNEAS
The basal opacity of all corneae was recorded. Each corneae with a value of the basal opacity > 7 was discarded.

NUMBER OF REPLICATES
Sets of three corneae were used for treatment with the test item and the negative and positive controls.

NEGATIVE CONTROL USED
Saline (0.9% NaCl in deionised water) was used as negative control.

POSITIVE CONTROL USED
2-Ethoxyethanol (purity: 99%) was used as positive control.

APPLICATION DOSE AND EXPOSURE TIME
The anterior compartment received a volume of 0.75 mL on the surface of the corneae.
The incubation time lasted ten minutes.

TREATMENT METHOD:
Each isolated cornea was mounted in a specially designed cornea holder according to the description given in OECD guideline 437 with a closed chamber.

REMOVAL OF TEST SUBSTANCE
- POST-EXPOSURE INCUBATION: After the test item or control items, respectively, were rinsed off from the application side with
saline, the corneae were incubated for further two hours in a vertical position.

METHODS FOR MEASURED ENDPOINTS:
- Corneal opacity: changes in light transmission with an opacitometer
- Corneal permeability: passage of sodium fluorescein dye measured with the aid of UV/VIS spectrophotometry

SCORING SYSTEM: In Vitro Irritancy Score (IVIS)

DECISION CRITERIA: according to OECD guideline 437: =3 no category, >3; =55 no prediction can be made; >55 Serious eye damage according to CLP/EPA/GHS (Cat 1)

Results and discussion

In vitro

Results
Irritation parameter:
in vitro irritation score
Value:
3.1
Negative controls valid:
yes
Positive controls valid:
yes
Remarks on result:
other: No indication of serious eye damage
Other effects / acceptance of results:
ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: The negative control responses result in opacity and permeability values that are less than the established upper limits for background opacity and permeability values for bovine corneae treated with the respective negative control.
- Acceptance criteria met for positive control: The positive control gives an IVIS that falls within two standard deviations of the current historical mean (updated every three months).

Any other information on results incl. tables

Test Group

Opacity value = Difference (t130-t0) of Opacity

Permeability at 490 nm (OD490)

IVIS

Mean IVIS

 

Proposed in vitro Irritancy Score

 

 

Mean

 

Mean

 

 

 

Negative Control

-1

-0.33

0.074

0.067

0.11

0.67

Not categorized

0

0.061

0.92

0

0.066

0.99

Positive Control

55.33*

1.042*

70.96

76.02

Category 1

55.33*

1.312*

73.01

67.33*

1.116*

84.07

ETHYLPROPIONATE

0.33*

0.048*

1.05

3.10

No prediction can be made

3.33*

0.005*

3.41

4.33*

0.033*

4.83

*corrected values

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
With the negative control (saline) neither an increase of opacity nor permeability of the corneae could be observed (mean IVIS = 0.67).
The positive control (2-Ethoxyethanol) was tested undiluted and showed clear opacity and distinctive permeability of the corneae (mean IVIS = 76.02) corresponding to a classification as serious eye damaging (CLP/EPA/GHS (Cat 1)).

Relative to the negative control, the test item caused a slight increase of the corneal opacity. The calculated mean IVIS was 3.10 (threshold for serious eye damage: IVIS >= 55 or <=03). According to OECD 437 no prediction for the damage hazard of the test item to the eye can be made.
Executive summary:

The corneal damage potential of the test item by means of the BCOP assay using fresh bovine corneae was assessed. With the negative control (0.9% (w/v) NaCl solution in deionised water) neither an increase of opacity nor permeability of the corneae could be observed. The positive control (2-Ethoxyethanol) showed clear opacity and distinctive permeability of the corneae corresponding to a classification as serious eye damaging (CLP/EPA/GHS (Cat 1)). The calculated mean in vitro irritancy score was 3.10. According to OECD 437 (see table in chapter 3.10.3) the test item is not classified as causing serious eye damage but relative to the negative control, the test item caused a slight increase of the corneal opacity. Permeability effects did not occur.