Dimethyl sebacate

Eye irritation

Administrative data

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

23 October 2012 to 7 May 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Fully GLP compliant and in accordance with current test guidelines

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Dimethyl sebacate

EC Number:

203-431-4

EC Name:

Dimethyl sebacate

Cas Number:

106-79-6

Molecular formula:

C12H22O4

IUPAC Name:

1,10-dimethyl decanedioate

Test material form:

other: Solid to liquid (melting point 23°C)

Details on test material:

Name: Dimethyl sebacate
CAS number: 106.79.6
Batch number: 120801
Purity: 99.08%
Expiry date: 31 August 2013
Date of receipt: 11 October 2012
Storage details: stored in a sealed container, at room temperature in the dark,

Test animals / tissue source

Species:

other: Bovine

Strain:

not specified

Details on test animals or tissues and environmental conditions:

Upon arrival at the test facility, the corneas were excised from the eyes and loaded onto the specifically designed holders. Both chambers of each holder were filled with pre-warmed Eagle's Minimal Essential Medium (EMEM) (posterior chamber first), ensuring that no bubbles were formed. The holders were incubated at 32±1°C for at least 1 hour. After the incubation, the media was removed from both the anterior and posterior chmabers. Fresh media was added to the the posterior chamber first and then the anterior chamber (this media replacement order ensured the cornea retained its natural curvature as much as possible). The opacity of each cornea was measured using an opacitometer. Any corneas found to have scratches or increased neovascularization or an opacity of >7 opacity units when examined prior to treatment were discarded.

Test system

Vehicle:

unchanged (no vehicle)

Controls:

not required

Amount / concentration applied:

A volume of 750 µL of undiluted test article was applied to each of the three corneas.

Duration of treatment / exposure:

After the initial application the corneas were incubated for 10 minutes.

Observation period (in vivo):

After the initial 10 minute exposure period the corneas were washed with media containing phenol red (as a pH indicator) until the indicator showed no effect. The corneas were then washed with fresh media and incubated (horixontally) for 2 hours ± 10 minutes. For the permeability endpoint, 1 mL of sodium flourescein (4 mg/mL solution) was added into the anterior chamber and the corneas were incubated in the vertical position for 1.5 hours ± 5 minutes.

Number of animals or in vitro replicates:

Three corneas were used for both dosed and control groups

Details on study design:

A volume of 750 µL was applied to each of the three corneas followed by a ten minute incubation at 32°C. After this incubation the corneas were waswashed with media containing phenol red (as a pH indicator) until the indicator showed no pH effect occuring (demonstrating that the test article had been removed successfully). The corneas were then washed once in media without phenol red and the opacities measured. The corneas were incubated (horizontally) for 2 hours ± 10 minutes after which, the opacities were measured and then the anterior chamber emptied. For the permeability endpoint, 1 mL of sodium flourescein (4 mg/mL solution) was added into the anterior chamber and the corneas were incubated in the vertical position for 1.5 hours ± 5 minutes. Following this period, the media in the posterior chamber was removed and held in a labelled tube. Three 350 µL aliquots of this media (per cornea) were analysed for optical density at 490 nanometers (OD490) using a spectrophotometer.
A volume of 750 µL of the negative or positive control was similarly applied to further groups of three corneas. These groups were subject to the procedures detailed above.
All corneas were preserved in 10% Neutral Buffered Formalin.

Results and discussion

In vitro

Results

Results 1

Irritation parameter:

cornea opacity score

Value:

-0.7

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Results 2

Irritation parameter:

other: mean group corrected optical density

Value:

-0.158

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Results 3

Irritation parameter:

in vitro irritation score

Value:

-3.04

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

In vivo

Irritant / corrosive response data:

The mean opacity reading for the test article was -0.7 and the mean group corrected optical density was -0.158.
The mean opacity reading and the mean group corrected optical density for the negative control were 0.0.
The mean opacity reading for the positive control was 54.7 and the mean group corrected optical density was 1.017.

Applicant's summary and conclusion

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: OECD GHS

Conclusions:

The test article, Dimethyl Sebacate, produced an IVIS score of -3.04 and was considered not to be corrosive or severely irritating to the eye.

Executive summary:

A bovine corneal opacity and permeability (BCOP) assay was conducted. Three corneas were treated by application of 750 µL of the undiluted test article onto the anterior surface of the cornea followed by a 10 minute incubation at 32°C ± 1°C. A volume of 750 µL of the negative or positive control was similarly applied to further groups of three corneas.

The corneas were then washed and the anterior chamber filled with pre-warmed EMEM (without phenol red). The corneas were then incubated for an additional 2 hours at 32°C ± 1°C. At the end of the post-incubation period the corneas were assessed for opacity. Permeability was determined by the amount of sodium fluorescein dye that penetrated all corneal layers. The media in the anterior chamber was replaced with 4 mg/mL sodium fluorescein, while the posterior chamber was filled with fresh EMEM. The corneas were then incubated for 90 minutes ± 5 minutes at 32 ± 1°C after which the media in the posterior chamber was assessed for presence of sodium fluorescein by measuring the optical density at 490 nanometers using a spectrophotometer.

The mean opacity reading for the test article was -0.7 and the mean group corrected optical density for the test article was -0.158. The test article produced an IVIS score of -3.04 and was considered not to be corrosive or severely irritating to the eye. The in vivo test was therefore conducted.