Registration Dossier

Toxicological information

Eye irritation

Currently viewing:

Administrative data

Endpoint:
eye irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
2014-11-17 to 2015-05-20
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 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)
Version / remarks:
26 July 2013
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Lot/batch No.of test material: 003
- Expiration date of the lot/batch: 01 Jan 2020

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature, exclusion of direct sunlight, test substance is hygroscopic
- Stability under test conditions: stable

Test animals / tissue source

Species:
cattle
Strain:
not specified
Details on test animals or tissues and environmental conditions:
SOURCE OF COLLECTED EYES
- Source: Schlachthof Bensheim, Am Schlachthof 7-9, 64625 Bensheim, Germany
- Characteristics of donor animals: 12 to 60 months of age
- indication of any existing defects or lesions in ocular tissue samples: Not observed.

Test system

Vehicle:
water
Controls:
yes
Amount / concentration applied:
TEST MATERIAL
- Amount applied: 750 µL
- Concentration (if solution): 20 % (w/v)

VEHICLE
- Amount applied: 750 µL
Duration of treatment / exposure:
4 hours
Number of animals or in vitro replicates:
3
Details on study design:
SELECTION AND PREPARATION OF CORNEAS
Corneas free of defects (opacity, scratches, pigmentation etc.) were dissected with a 2 to 3 mm rim of sclera. Isolated corneas were mounted in cornea holders that consists of anterior and posterior chambers. Both chambers were filled to excess with pre-warmed Eagles’s MEM (without phenol red) and then equilibrated in a vertical position at about 32 °C for at least 1 hour. After the equilibration period the medium in both chambers was replaced with fresh prewarmed medium and initial corneal opacity readings were taken for each cornea with an opacitometer. Any corneas that showed macroscopic tissue damage or an opacity value < 537 opacity units 1 were discarded. The remaining corneas were then distributed into negative control, positive control and treatment groups. Each corneal holder was uniquely identified with a number on the chambers.

NUMBER OF REPLICATES
3 corneas were used per treatment group.

SOLVENT CONTROL USED
De-ionized water

POSITIVE CONTROL USED
Imidazole (CAS No. 288-32-4) 20 % (w/v) solution in deionized water for non-surfactant solid test substances

TREATMENT METHOD: open chamber

POST-INCUBATION PERIOD: no

REMOVAL OF TEST SUBSTANCE
- Number of washing steps after exposure period: The test substance could not be removed with a syringe. Therefore the corneas were rinsed with the open chamber method.

METHODS FOR MEASURED ENDPOINTS:
- Corneal opacity: was observed visually. Final corneal opacity readings were taken for each cornea with an opacitometer.
- Corneal permeability: passage of sodium fluorescein dye measured with the aid of microtiter plate reader (OD490)

SCORING SYSTEM: In Vitro Irritancy Score (IVIS)

DECISION CRITERIA: Evaluation of results were based on the decision criteria used in OECD guideline 437.

Results and discussion

In vitro

Results
Irritation parameter:
in vitro irritation score
Value:
0
Vehicle controls validity:
valid
Negative controls validity:
not examined
Positive controls validity:
valid
Remarks on result:
no indication of irritation
Other effects / acceptance of results:
OTHER EFFECTS:
- Visible damage on test system: Not observed

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: Yes
- Acceptance criteria met for positive control: Yes
- Range of historical values: Mean ± SD (Mean-2SD - Mean+2SD)

Historic range NC
Opacity = 6.6 ± 2.0 (2.6 - 10.5)
Permeability = 0.001 ± 0.006 (-0.011 - 0.013)

Historic range PC
Opacity = 70.1 ± 13.7 (42.7 - 97.4)
Permeability = 3.039 ± 0.659 (1.721 - 4.358)
IVIS = 115.8 ± 12.0 (91.9 - 139.7)

Any other information on results incl. tables

Opacity score of test substance, NC and PC

Cornea No.

Initial opacity

final opacity

opacity change

corrected opacity change

mean

SD

test substance

7

6.1

9.8

3.7

0.0

0.0

0.0

8

2.2

4.6

2.4

0.0

9

3.5

11.1

7.6

0.0

NC

1

2.6

18.3

15.7

NA

8.4

6.4

2

3.4

8.8

5.4

NA

3

2.1

6.1

4

NA

PC

4

4.9

97.6

92.7

84.4

86.2

12.3

5

1.9

85.3

83.4

75

6

2.2

109.9

107.7

99.3

Permeability score of test substance, NC and PC

Cornea No.

Mean OD490

Dilution factor

Mean corrected OD490*

mean

SD

test substance

7

0.005

1

0.003

0.003

0.003

8

0.008

1

0.006

9

-0.002

1

0

NC

1

-0.001

1

NA

0.002

0.002

2

0.002

1

NA

3

0.003

1

NA

PC

4

0.559

5

2.792

2.625

0.275

5

0.462

5

2.308

6

0.556

5

2.777

  In Vitro Irritancy score (IVIS) of the test substance, NC and PC

Cornea No.

Opacity per cornea

Permeability per cornea

IVIS

per cornea

mean

SD

test substance

7

0.0

0.003

0.0

0.0

0.0

8

0.0

0.006

0.1

9

0.0

0.0

0.0

NC

1

15.7

0.0

15.7

8.4

6.4

2

5.4

0.002

5.4

3

4

0.003

4.1

PC

4

84.4

2.792

126.2

125.6

15.7

5

75

2.308

109.7

6

99.3

2.777

141

 

* Negative values are set to zero for further calculation

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met