Tetraol

Eye irritation

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

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. QA statement)

Test material

Test material information

Constituent 1

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