Sodium 6,14-diethyl-1,1,1,2,2,18,18,19,19,19-decafluoro-8,12-dioxo-10-({[1-(2,2,3,3,3-pentafluoropropoxy)butan-2-yl]oxy}carbonyl)-4,7,13,16-tetraoxanonadecane-9-sulfonate

Eye irritation

Administrative data

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

02 FEB - 16 MAR 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying Ocular Corrosives and Severe Irritants)

Version / remarks:

7 SEP 2009

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU method B.47 (Bovine corneal opacity and permeability test method for identifying ocular corrosives and severe irritants)

Version / remarks:

30 MAY 2008

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

sodium 1,5-dioxo-1,5-bis({[1-(2,2,3,3,3-pentafluoropropoxy)butan-2-yl]oxy})-3-({[1-(2,2,3,3,3-pentafluoropropoxy)butan-2-yl]oxy}carbonyl)pentane-2-sulfonate

EC Number:

700-541-9

Cas Number:

1472634-24-4

Molecular formula:

C27H34F15NaO12S

IUPAC Name:

sodium 1,5-dioxo-1,5-bis({[1-(2,2,3,3,3-pentafluoropropoxy)butan-2-yl]oxy})-3-({[1-(2,2,3,3,3-pentafluoropropoxy)butan-2-yl]oxy}carbonyl)pentane-2-sulfonate

Test material form:

solid

Test animals / tissue source

Species:

cattle

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: slaughterhouse (Odenwaldschlachthof Brensbach, D-64395 Brensbach)
Freshly isolated bovine eyes of donor cattle. Excess tissue was removed from the eyes. The eyes were kept and transported in transport medium.

Test system

Vehicle:

other: 0.9% sodium chloride solution

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

TEST MATERIAL
- Concentration (if solution): 20 % in 0.9 % sodium chloride

VEHICLE
- Amount(s) applied (volume or weight with unit): 750 µL
- Concentration (if solution): 0.9 %
- Lot/batch no. (if required): 0191C15

Duration of treatment / exposure:

incubation for 240 min

Duration of post- treatment incubation (in vitro):

permeability determination: 90 min

Number of animals or in vitro replicates:

3

Details on study design:

SELECTION AND PREPARATION OF CORNEAS
The corneas were prepared immediately after delivery of the eyes to the laboratory. All eyes were carefully examined macroscopically for defects. Those presenting defects such as vascularization, pigmentation, opacity or scratches were discarded. The corneas were carefully removed from the eyes using scalpel and rounded scissors. A rim of about 2 mm of tissue (sclera) was left for stability and handling of the isolated cornea. All corneas used in the study were collected in incubation medium (EMEM, pre-warmed at 32 °C).
Each cornea was mounted in a cornea holder (LAB Research, Hungary) with the endothelial side against the sealing ring (0-ring) of the posterior part of the holder. The cornea was gently flattened over the 0-ring without stretching the cornea. Afterwards, the anterior part of the holder was positioned on top of the cornea and fixed in place with screws. Both compartments of the holder were filled with incubation medium (EMEM). The posterior compartment was filled first to return the cornea to its natural convex form.

QUALITY CHECK OF THE ISOLATED CORNEAS
Yes, Opacity Measurement before Treatment:
The opacitometer measured the light transmission passing through the corneas and displayed a lux value. This value was recorded in a table and converted into an opacity value (baseline opacity values). The opacitometer was calibrated as described in the manual and the measured values were within the range of the acceptance criteria. The opacity of each cornea was determined by reading each holder placed in the photoreceptor compartment of the opacitometer. Any corneas that showed macroscopic tissue damage (e.g., scratches, pigmentation, neovascularization) or an opacity > 7 opacity units were discarded. The mean opacity of all equilibrated corneas was calculated. Three corneas with opacity values close to the median value for all corneas as were selected as negative control corneas. The remaining corneas were distributed into treatment and positive control groups.

NUMBER OF REPLICATES
3

NEGATIVE CONTROL USED
yes, see solvent control

SOLVENT CONTROL USED (if applicable)
yes, 0.9 % sodium chloride solution

POSITIVE CONTROL USED
yes, Imidazole

APPLICATION DOSE AND EXPOSURE TIME
Test material: 20 % suspension in 0.9 % sodium chloride, 240 min
Solvent control: 0.9 % sodium chloride, 240 min
Positive control: 20 % Imidazole solution, 240 min

POST-INCUBATION PERIOD: 90 min (for permeability determination)

REMOVAL OF TEST SUBSTANCE
- Number of washing steps after exposure period: at least 3 (or until no visual evidence of test chemical can be observed)
- POST-EXPOSURE INCUBATION: no

METHODS FOR MEASURED ENDPOINTS:
- Corneal opacity: baseline opacity was determined with an opacitometer (BASF-OP2.0)
- Corneal permeability: passage of sodium fluorescein dye measured with the aid of microtiter plate reader (BioTek ELx800) (OD490)

SCORING SYSTEM: In Vitro Irritancy Score (IVIS)

DECISION CRITERIA: A substance that induced an IVIS ≥ 55.1 is defined as a corrosive or a severe irritant.

VALIDITY CRITERIA: The test is acceptable if the IVIS of the positive control and the negative control falls within two standard deviations of the current historical mean (IVIS positive control: 90.2 - 148.5; IVIS negative control: -0.9 - 4.2).

Results and discussion

In vitro

Results

Results 1

Irritation parameter:

in vitro irritation score

Run / experiment:

1

Value:

5.879

Vehicle controls validity:

valid

Negative controls validity:

valid

Remarks:

see vehicle control

Positive controls validity:

valid

Results 2

Irritation parameter:

in vitro irritation score

Run / experiment:

2

Value:

3.991

Vehicle controls validity:

valid

Negative controls validity:

valid

Remarks:

see vehicle control

Positive controls validity:

valid

Results 3

Irritation parameter:

in vitro irritation score

Run / experiment:

3

Value:

4.233

Vehicle controls validity:

valid

Negative controls validity:

valid

Remarks:

see vehicle control

Positive controls validity:

valid

Results 4

Irritation parameter:

in vitro irritation score

Run / experiment:

mean

Value:

4.7

Vehicle controls validity:

valid

Negative controls validity:

valid

Remarks:

see vehicle control

Positive controls validity:

valid

Results 5

Irritation parameter:

cornea opacity score

Run / experiment:

1

Value:

4.851

Vehicle controls validity:

valid

Negative controls validity:

valid

Remarks:

see vehicle control

Positive controls validity:

valid

Results 6

Irritation parameter:

cornea opacity score

Run / experiment:

2

Value:

3.913

Vehicle controls validity:

valid

Negative controls validity:

valid

Remarks:

see vehicle control

Positive controls validity:

valid

Results 7

Irritation parameter:

cornea opacity score

Run / experiment:

3

Value:

1.109

Vehicle controls validity:

valid

Negative controls validity:

valid

Remarks:

see vehicle control

Positive controls validity:

valid

Results 8

Irritation parameter:

fluorescein retention score

Run / experiment:

1

Value:

0.069

Vehicle controls validity:

valid

Negative controls validity:

valid

Remarks:

see vehicle control

Positive controls validity:

valid

Results 9

Irritation parameter:

fluorescein retention score

Run / experiment:

2

Value:

0.005

Vehicle controls validity:

valid

Negative controls validity:

valid

Remarks:

see vehicle control

Positive controls validity:

valid

Results 10

Irritation parameter:

fluorescein retention score

Run / experiment:

3

Value:

0.208

Vehicle controls validity:

valid

Negative controls validity:

valid

Remarks:

see vehicle control

Positive controls validity:

valid

Any other information on results incl. tables

Validity of the Test

After treatment with the negative control (0.9% sodium chloride solution) the calculated IVIS was 1.1 and, thus, within two standard deviations of the current historical mean (IVIS: -0.9 - 4.2). After treatment with the positive control (20% Imidazole) the calculated IVIS was 94. 7 and, thus, also within two standard deviations of the current historical mean (IVIS 90.2 - 148.5). Therefore, the study fulfilled the validity criteria.

Purpose

Thisin vitroGLP study was performed to assess the ocular severe irritant or corrosive potential of the test item by means of the BCOP (Bovine Corneal Opacity and Permeability Assay) according to OECD Guideline 437 and the Commission Regulation (EU) No. 1152/2010.

For this purpose fresh bovine corneas were exposed to a 20% suspension of the test material and toxic effects to the cornea were measured by decreased light transmission (opacity) and increased passage of sodium fluorescein dye (permeability). The opacity and permeability assessments of the corneas were combined to determine anIn VitroIrritancy Score (IVIS), which was used to classify the irritancy potential of the test material.

 

Study Design

In this study the induced opacity and increased permeability was investigated in isolated bovine corneas after exposure to the test material as a 20% suspension in a 0.9% sodium chloride solution. As negative control 0.9% sodium chloride solution and as positive control 20% imidazole was used. 3 corneas were used per group (test material, negative and positive control group). After a first opacity measurement of the fresh bovine corneas, 750 µL of the suspended test material, positive or negative control were applied on the corneas and incubated for 240 minutes at 32 ± 1 °C. After the incubation phase the test material, the positive, and the negative control were rinsed from the corneas and the opacity was measured again. After the opacity measurements, the permeability of the corneas was determined by application of 1 mL of a fluorescein solution for 90 minutes at 32 ± 1 °C. The amount of fluorescein solution that crossed the cornea was measured spectrophotometrically. The opacity and permeability assessments were combined to determine an In Vitro Irritancy Score (IVIS). 

Results

The treatment of the corneas with the negative control (0.9% sodium chloride solution) showed neither an increase of opacity nor an increase of permeability. After treatment with the positive control (20% imidazole) the calculated IVIS was 94.7 and, thus, within two standard deviations of the current historical mean (IVIS: 90.2 - 148.5). Therefore, the study fulfilled the validity criteria. The IVIS obtained after treatment with the test material was 4.7 and, thus, lower than 55.1. Therefore, the test material is not considered to possess an ocular severe irritant or corrosive potential. 

Conclusions

Under the given experimental conditions, the test material did not show an ocular severe irritant or corrosive potential.

Applicant's summary and conclusion

Interpretation of results:

other: not Category 1 (irreversible effects on the eye) based on EU GHS criteria

Conclusions:

The IVIS obtained after treatment with the test material was 4.7 and, thus, lower than 55.1. Therefore, the test material is not considered to possess an ocular severe irritant or corrosive potential.