4-Decenal, 9-hydroxy-5,9-dimethyl-

Eye irritation

Administrative data

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

May 24, 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP study conducted according to OECD test Guideline No. 437 without any deviation.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Test guideline 1

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:

No. 1152/2010

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

September 2009

Deviations:

no

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes (incl. QA statement)

Remarks:

German GLP Certificate (Signed on May 12, 2014)

Test material

Test material information

Constituent 1

Reference substance name:

9-hydroxy-5,9-dimethyldec-4-enal

EC Number:

940-437-2

Cas Number:

926-50-1

Molecular formula:

C12H22O2

IUPAC Name:

9-hydroxy-5,9-dimethyldec-4-enal

Test material form:

liquid

Details on test material:

Appearance: colourless to pale yellow liquid
Expiry date: 18 March 2014, extended to 19 April 2014, and then 22 May 2015

Test animals / tissue source

Species:

other: Freshly isolated bovine cornea

Details on test animals or tissues and environmental conditions:

TEST SYSTEM
- Eyes from at least 9 month old donor cattle were obtained from a local abattoir. Excess tissue was removed from the excised eye. The isolated eyes were transported to the laboratory Hanks' Balanced Salt Solution (HBSS) supplemented with streptomycin/penicillin at ambient temperature. The corneae were isolated on the same day after delivery of the eyes.

Test system

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.75 mL was applied on each cornea
- Concentration: Undiluted

Duration of treatment / exposure:

The undiluted test item was applied for 10 minutes at 32 ± 1 °C.

Observation period (in vivo):

- The opacitometer determines changes in the light transmission passing through the corneae, and displays a numerical opacity value. This value was recorded. The opacitometer OP_KiT opacitometer (Electro Design, 63-Riom France) was calibrated and the opacity of each of the corneae was determined by reading each holder placed in the photoreceptor compartment for treated cornea. Corneal opacity was measured after exposure, after rinsing and after incubation of the corneae for two hours (t130).
- Permeability endpoint was measured after the final opacity measurement when the complete medium was removed from the anterior compartment and replaced by 1 mL of a 0.5% w/v Sodium Fluorescein solution in HBSS. Corneawere incubated again in a horizontal postion for 90 minutes in a water-bath at 32 +/- 1°C. Complete medium from the posterior compartment was removed, well mixed and the optical density at 490 nm (OD490) was determined with a spectrophotometer.

Duration of post- treatment incubation (in vitro):

After the incubation phase the test item, positive and the negative controls were rinsed from the cornea and incubated for another 120 minutes at 32 ± 1 °C

Number of animals or in vitro replicates:

Total: 9 corneas - 3 corneas/group for test item, negative and positive controls

Details on study design:

PREPARATION OF CORNEAE
All eyes were carefully examined macroscopically for defects. Those presenting defects such as vascularization, pigmentation, opacity and scratches were discarded. The cornea was carefully removed from the eye using scalpel and rounded scissors. A rim of about 2 mm of tissue (sclera) was left for stability and handling of the isolated cornea. The corneae were directly used in the BCOP test on the same day.
Each isolated cornea was mounted in a specially designed cornea holder according to the description given in guideline that consists of anterior and posterior compartments, which interface with the epithelial and endothelial sides of the cornea, respectively. The endothelial side of the cornea was positioned againstthe sealing ring (O-ring) of the posterior part of the holder. The cornea was gently flattened over the O-ring but stretching was avoided. 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. The posterior compartment was filled first to return the cornea to its natural convex position. Care was taken to assure no air bubbles were present within the compartments. For equilibration, the corneae in the holder were incubated in a vertical position for about one hour at 32 +/- 1 °C in a water-bath.
At the end of the incubation period, the basal opacity was determined (t0) and recorded. Each corneae with a value of the basal opacity > 7 was discarded.
Sets of three corneae were used for treatment with the test item and the negative and positive controls.

DETAILS ON TEST PROCEDURE
The anterior compartment received the test item or negative or positive control at a volume of 0.75 mL on the surface of the corneae and was incubated at 32 +/- 1°C in the water-bath, while the corneae were in a horizontal position. The incubation time lasted ten minutes.

After the test item and control items were rinsed off from the application side with saline. The corneae was then incubated at 32 ± 1 °C for further two hours in a vertical position, followed by a second opacity reading (t130). In the second step of the assay, permeability of the cornea was determined.

Results and discussion

In vitro

Results

Irritation parameter:

in vitro irritation score

Value:

7.03

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Any other information on results incl. tables

See the attached document for information on tables of results

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

According to the current study and under the experimental conditions reported, the test item is not corrosive/not severely irritating to the eye according to the Regulation (EC) No. 1272/2008 (CLP) and to the Globally Harmonized System of Classification and Labelling of Chemicals (GHS).

Executive summary:

In an in vitro eye irritation study performed according to the OECD Guideline 437 and in compliance with GLP, 0.75 mL of undiluted test item was applied to isolated bovine corneas for 10 minutes followed by an incubation period of two hours at 32 °C. Three corneas were used for undiluted test item, negative control (saline solution) and positive control (2-Ethoxyethanol). 

After a first opacity measurement of the fresh bovine corneae (t0), the neat test item; the positive and the negative controls were applied to corneae and incubated for 10 minutes at 32 +/- 1°C. After the incubation phase the test item, positive and negative controls were rinsed from the corneae. Further, the corneae were incubated for another 120 minutes at 32 +/- 1°C in incubation medium, and opacity was measured a second time (t130). After the opacity measurements, permeability of the corneae was determined by measuring spectrophotometrically the transfer of sodium fluorescein after incubation in a horizontal position for 90 minutes at 32 +/- 1°C.

With the negative control neither an increase of opacity nor permeability of the corneae could be observed (mean IVIS = 1.44).

The positive control (2-Ethoxyethanol) showed clear opacity and distinctive permeability of the corneae (mean IVIS = 65.13) corresponding to a classification as corrosive/severe irritant to the eye (CLP/GHS Cat.1).

Relative to the negative control, the test item caused a slight increase of the corneal opacity and permeability. The calculated mean IVIS was 7.03 (threshold for corrosivity/severe irritancy: IVIS ≥ 55.1). According to OECD 437, the test item is classified as not corrosive/not severe irritant to the eye.

 

According to the current study and under the experimental conditions reported, the test item is not corrosive/not severely irritating to the eye according to the Regulation (EC) No. 1272/2008 (CLP) and to the Globally Harmonized System of Classification and Labelling of Chemicals (GHS).