Registration Dossier

Administrative data

Endpoint:
eye irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
13 Jun 2017 - 13 Jun 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017

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:
2013
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid
Details on test material:
Identification: Soybean oil, epoxidized, Me ester, reaction products with propylene glycol
Test item 207811/B
Appearance: Yellow liquid (determined by Charles River Den Bosch)
Purity/Composition: UVCB
Test item storage: At room temperature
Stable under storage conditions until: 17 May 2018 (expiry date)
Test item handling: No specific handling conditions required
Chemical name (IUPAC), synonym or trade name: Not indicated

Test animals / tissue source

Species:
cattle
Strain:
not specified
Details on test animals or tissues and environmental conditions:
TISSUE SOURCE
- Source: Vitelco, 's Hertogenbosch, The Netherlands
- Age at study initiation: young cattle

OTHER: the eyes were excised by a slaughterhouse employee as soon as possible after slaughter. Eyes were collected and transported in physiological saline in a suitable container under cooled conditions.

Test system

Vehicle:
unchanged (no vehicle)
Controls:
yes, concurrent no treatment
yes, concurrent positive control
Amount / concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 750µL
- Concentration (if solution): Undiluted
Duration of treatment / exposure:
10 +/- 1 minutes
Duration of post- treatment incubation (in vitro):
120 +/- 10 minutes
Number of animals or in vitro replicates:
triplicate
Details on study design:
SELECTION AND PREPARATION OF CORNEAS
The eyes were checked for unacceptable defects, such as opacity, scratches, pigmentation and neovascularization by removing them from the physiological saline and holding them in the light. Those exhibiting defects were discarded.
The isolated corneas were stored in a petri dish with cMEM (Earle’s Minimum Essential Medium) containing 1% (v/v) L-glutamine and 1% (v/v) Foetal Bovine Serum. The isolated corneas were mounted in a corneal holder (one cornea per holder) of BASF with the endothelial side against the O-ring of the posterior half of the holder. The anterior half of the holder was positioned on top of the cornea and tightened with screws. The compartments of the corneal holder were filled with cMEM of 32C. The corneas were incubated for the minimum of 1 hour at 32 C.

QUALITY CHECK OF THE ISOLATED CORNEAS
After the incubation period, the medium was removed from both compartments and replaced with fresh cMEM. Opacity determinations were performed on each of the corneas using an opacitometer. The opacity of each cornea was read against a cMEM filled chamber, and the initial opacity reading thus determined was recorded. Corneas that had an initial opacity reading higher than 7 were not used.

NUMBER OF REPLICATES
Three corneas were selected at random for each treatment group.

NEGATIVE CONTROL USED
Unexposed

POSITIVE CONTROL USED
Ethanol

APPLICATION DOSE AND EXPOSURE TIME
Undiluted 10 min exposure time

TREATMENT METHOD: closed chamber

REMOVAL OF TEST SUBSTANCE
- Number of washing steps after exposure period: at least 2

- POST-EXPOSURE INCUBATION: yes 120 +/- 10 minutes

METHODS FOR MEASURED ENDPOINTS:
- Corneal opacity: measured by the diminution of light passing through the cornea using an opacitometer
- Corneal permeability: microtiter plate reader (OD490) TECAN Infinite® M200 Pro Plate Reader
- Others (e.g, pertinent visual observations, histopathology): (please specify)

SCORING SYSTEM: In Vitro Irritancy Score (IVIS)

DECISION CRITERIA: according to guideline

Results and discussion

In vitro

Results
Irritation parameter:
in vitro irritation score
Run / experiment:
Main experiment (mean of triplicate)
Value:
-0.3
Vehicle controls validity:
not applicable
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
no indication of irritation
Other effects / acceptance of results:
OTHER EFFECTS:
- Visible damage on test system: not reported

DEMONSTRATION OF TECHNICAL PROFICIENCY:
The negative control responses for opacity and permeability were less than the upper limits of the laboratory historical range indicating that the negative control did not induce irritancy on the corneas. The mean in vitro irritancy score of the positive control (Ethanol) was 53 and was within two standard deviations of the current historical positive control mean. It was therefore concluded that the test conditions were adequate and that the test system functioned properly.

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: Yes
- Acceptance criteria met for positive control: Yes
- Range of historical values:

Negative control
Opacity: -2.9 - 3.0, Mean=0.10, SD=1.04, n=78
Permeability: -0,016 - 0,042, Mean=0,01, SD=0,01, n=71
In vitro Irritancy Score: -2.8 - 3.0, Mean=0.20, SD=1.14, n=53

Positive control
In vitro Irritancy Score: 34.7-78.2, Mean=56.47, SD=12.48, n=53


Applicant's summary and conclusion

Interpretation of results:
other: Not classified
Remarks:
Based on CLP (1272/2008/EC).
Conclusions:
Soybean oil, epoxidized, Me ester, reaction products with propylene glycol does not need to be classified for Eye irritation in accordance with the criteria outlined in Annex I of the CLP Regulation (1272/2008/EC).
Executive summary:

The eye irritation hazard potential of Soybean oil, epoxidized, Me ester, reaction products with propylene glycol was as measured Acoording to OECDTG437 (BCOP test). The eye damage of

"Soybean oil, epoxidized, methyl ester, reaction products with propylene glycol" was tested through topical application for 10 minutes.  The test item was applied as it is (750 µl) directly on top of the corneas. The negative and positive were considered valid. It was therefore concluded that the test conditions were adequate and that the test system functioned properly.  The test item did not induce ocular irritation through both endpoints (no corneal opacity or permeability), resulting in a mean in vitro irritancy score of -0.3 after 10 minutes of treatment.  In conclusion, since the test item induced an IVIS ≤ 3, no classification is required for eye irritation or serious eye damage, in accordance with the criteria outlined in Annex I of the CLP Regulation (1272/2008/EC).