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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:
17 Oct 2017 - 17 Oct 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report Date:
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:
July 26, 2013
Deviations:
no
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid

Test animals / tissue source

Species:
cattle
Strain:
not specified
Details on test animals or tissues and environmental conditions:
SOURCE OF COLLECTED EYES
- Source: Vitelco, 's Hertogenbosch, The Netherlands
- Characteristics of donor animals: 6-60 months
- Storage, temperature and transport conditions of ocular tissue: Eyes were collected and transported in physiological saline without antibioticsin a suitable container under cooled conditions.
- indication of any existing defects or lesions in ocular tissue samples: eyes exhibiting defects were discarded
- Indication of any antibiotics used: none used

Test system

Vehicle:
unchanged (no vehicle)
Controls:
yes
yes, concurrent positive control
Amount / concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 750 mL
Duration of treatment / exposure:
10 +/- 1 minutes
Duration of post- treatment incubation (in vitro):
120 +/- 10 minutes
Number of animals or in vitro replicates:
single sample, measured in 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
Value:
11
Vehicle controls validity:
not applicable
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
not determinable
Remarks:
IVIS > 3 ≤ 55, no prediction on the classification can be made according to UN GHS
Other effects / acceptance of results:

ACCEPTANCE OF RESULTS:
The positive control gives an in vitro irritancy score that falls within two standard deviations of the current historical mean.
The negative control responses should result in opacity and permeability values that are less than the upper limits of the laboratory historical range.

Range of historical values if different from the ones specified in the test guideline: see table

Any other information on results incl. tables

Historical Control Data for the BCOP Studies

 

Negative control

Positive control

 

Opacity

Permeability

In vitroIrritancy Score

In vitroIrritancy Score

Range

-2.9 – 3.0

-0.016 – 0.042

-2.8 – 3.0

34.7 – 78.2

Mean

0.08

0.01

0.17

56.01

SD

1.04

0.01

1.14

12.51

n

84

77

78

55

SD = Standard deviation

n = Number of observations

The above mentioned historical control data range of the controls were obtained by collecting all data over the period of Aug 2014 to Aug 2017.

Applicant's summary and conclusion

Interpretation of results:
other: Not determinable
Remarks:
in accordance with Annex I of 1272/2008/EC (CLP)
Conclusions:
Based on the results of the present study, 3-Methyl Sulfolane induced an IVIS > 3 ≤ 55. Therefore no prediction on the classification can be made according to UN GHS and Annex I of the CLP Regulation (1272/2008/EC).
Executive summary:

The eye irritation hazard potential of 3-Methyl Sulfolane was as measured Acoording to OECDTG437 (BCOP test). The eye damage of the test substance 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.  3-Methyl Sulfolane induced ocular irritation through one endpoint (opacity), resulting in a mean in vitro irritancy score of 11 after 10 minutes of treatment.

In conclusion, since 3-Methyl Sulfolane induced an IVIS > 3 ≤ 55, no prediction on the classification can be made.