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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:
weight of evidence
Study period:
2015-07-07 to 2015-07-28
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2016
Report Date:
2016

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 material form:
liquid
Details on test material:
Purity of Test material:
60.9 % Disodium (sulphonatothio)acetate
20.3 % NaCl
4.3 % Na2S2O3
0.6 % NasSO4
5.0 % H2O
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No. of test material: Sample 14/078 from Mollescal HW, charge 10924344R0
- Expiration date of the lot/batch: April 28, 2019

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature

Test animals / tissue source

Species:
cattle
Strain:
not specified
Details on test animals or tissues and environmental conditions:
SOURCE OF COLLECTED EYES
- Source: Schlachthof Mannheim, Schlachthofstr. 21, 68165 Mannheim, Germany
- Characteristics of donor animals (age): 12 to 60 months
- indication of any existing defects or lesions in ocular tissue samples: No

Test system

Vehicle:
water
Controls:
yes, concurrent vehicle
yes, concurrent positive control
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.

QUALITY CHECK OF THE ISOLATED CORNEAS
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 < 542 opacity units were discarded.

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: closed chamber

POST-INCUBATION PERIOD: no

REMOVAL OF TEST SUBSTANCE
- Number of washing steps after exposure period: at least 3 times with Eagle’s MEM (containing phenol red) and once with Eagle’s MEM (without phenol red)

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
Run / experiment:
test substance
Value:
0.5
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 = 11.1 ± 6.9 (-2.7 - 24.9)
Permeability = 0.003 ± 0.001 (0.001 - 0.006)

Historic range PC
Opacity = 76.3 ± 15.1 (46.2 - 106.4)
Permeability = 2.803 ± 0.616 (1.571 - 4.034)
IVIS = 118.3 ± 18.7 (81 - 155.6)

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

13

5.1

11.8

6.6

0

0.5

0.8

14

6

9

3

0

15

4.1

12.7

8.6

1.4

NC

1

5.4

11.7

6.3

NA

7.3

3.3

2

5.1

9.8

4.6

NA

3

4.1

15.1

10.9

NA

PC

4

3.5

107

103.4

96.1

70.2

25.4

5

3.4

79.9

76.5

69.2

6

3

55.6

52.6

45.4

Permeability score of test substance, NC and PC

Cornea No.

Mean OD490

Dilution factor

Mean corrected OD490*

Mean

SD

test substance

13

0.008

1

0.004

0.004

0.001

14

0.009

1

0.005

15

0.008

1

0.003

NC

1

0.004

1

NA

0.005

0.001

2

0.004

1

NA

3

0.006

1

NA

PC

4

0.699

5

3.492

2.992

0.533

5

0.611

5

3.052

6

0.487

5

2.432

 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

13

0

0.004

0.1

0.5

0.8

14

0

0.005

0.1

15

1.4

0.003

1.4

NC

1

6.3

0.004

6.3

7.3

3.3

2

4.6

0.004

4.7

3

10.9

0.006

11

PC

4

96.1

3.492

148.5

115.1

33.3

5

69.2

3.052

115

6

45.4

2.432

81.2

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met