Tall oil, compd. with ethanolamine

Eye irritation

Administrative data

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

1 April 2011 to 30 May 2011

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study conducted to GLP in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2011

Report date:

2011

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 438 (Isolated Chicken Eye Test Method for Identifying Ocular Corrosives and Severe Irritants)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Fatty acids, tall-oil, compds. with ethanolamine

EC Number:

268-640-5

EC Name:

Fatty acids, tall-oil, compds. with ethanolamine

IUPAC Name:

Fatty acids, tall-oil, compds. with ethanolamine

Test material form:

other: turbid liquid

Details on test material:

- Appearance: turbid, yellow liquid
- Storage Conditions: ambient temperature

Test animals / tissue source

Species:

other: Isolated Chicken Eye

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Approximately 7 week old male or female chickens (ROSS, spring chickens), bodyweight range approximately 1.5 to 2.5 kg, were used as eye-donors. Heads of these animals were obtained from a poultry slaughterhouse. Heads were cut off immediately after sedation of the animals by electric shock and incision of the neck for bleeding, and before they reached the next station on the process line. The heads were placed in small plastic boxes on a bedding of paper tissues moistened with isotonic saline. Next, they were transported to the testing facility. During transportation, the heads were kept at ambient temperature.

- Preparation of eyes: Within 2 hours of the animals being killed, eyes were carefully dissected and placed in a superfusion apparatus using the following procedure:
The eye-lids were carefully removed without damaging the cornea and a small drop of Fluorescein sodium BP 2.0 % w/v was applied to the corneal surface for a few seconds then subsequently rinsed off with isotonic saline at ambient temperature.
The head with the fluorescein-treated cornea was examined with a slit-lamp microscope (Slit-lamp 900 BP, Haag-Streit AG, Liebefeld-Bern, Switzerland) to ensure that the cornea was not damaged. If undamaged, the eye was further dissected from the head without damaging the eye or cornea. Care was taken to remove the eye-ball from the orbit without cutting the optical nerve too short.
The enucleated eye was placed in a stainless steel clamp with the cornea positioned vertically and transferred to a chamber of the superfusion apparatus. The clamp holding the eye was positioned in such a way that the entire cornea was supplied with isotonic saline from a bent, stainless steel tube, at a target rate of 0.10 to 0.15 mL/min (peristaltic pump set at speed 5.00, Watson-Marlow 205CA, Rotterdam, The Netherlands). The chambers of the superfusion apparatus, as well as the saline, were temperature controlled at approximately 32 °C (water pump set at 36.4 °C; Lauda 103, Germany).
After placing in the superfusion apparatus, the eyes were examined once again with the slit-lamp microscope to ensure that they were not damaged. Corneal thickness was measured using the Depth Measuring Attachment No. I for the Haag-Streit slit-lamp microscope. Corneal thickness was expressed in instrument units. An accurate measurement was taken at the corneal apex of each eye.
Eyes with a corneal thickness deviating more than 10 % of the average, eyes showing opacity (score higher than 0.5), eyes that were unacceptably stained with fluorescein (score higher than 0.5, indicating the cornea to be permeable) or eyes that showed any other signs of damage were rejected as test eyes and replaced.

Test system

Vehicle:

unchanged (no vehicle)

Controls:

other: concurrent in vitro positive and negative controls

Amount / concentration applied:

30 µL

Duration of treatment / exposure:

10 seconds of exposure, follwed by a 20 mL saline rinse.

Observation period (in vivo):

The eyes were examined at regular intervals after exposure; 30, 75, 120, 180 and 240 minutes.

Number of animals or in vitro replicates:

3 isolated eyes were exposed to the test material.

Details on study design:

Three test eyes for the test material, one negative control eye (exposed to physiological saline 0.9 %) and two positive control eyes (exposed to 5 % w/w aqueous benzalkonium chloride) were selected for testing. Each eye provided its own baseline values for corneal swelling, corneal opacity and fluorescein retention. For that purpose, after an equilibration period of 45 to 60 minutes, the corneal thickness of the eyes was measured again to determine the zero reference value for corneal swelling calculations.

At time t = 0 (i.e. immediately after the zero reference measurement), the following procedure was applied for each test eye:
The clamp holding the test eye was placed on paper tissues outside the chamber with the cornea facing upwards.
The eyes (corneas) were treated with 30 µL of the appropriate material for 10 seconds, followed by rinsing with 20 mL of saline. After rinsing, each eye in the holder was returned to its chamber.
The eyes were examined at approximately 0, 30, 75, 120, 180 and 240 minutes after treatment. Fluorescein retention was only scored at approximately 30 minutes after treatment. All examinations were carried out with the slit-lamp microscope.

After the final examination, the eyes were preserved in a neutral aqueous phosphate-buffered 4 % solution of formaldehyde. The corneas were embedded in paraffin wax, sectioned at 5 µm and stained with PAS (Periodic Acid-Schiff). The microscopic slides were filed in the archives of the testing facility and kept available for histopathological examination if considered relevant.

EVALUATION OF THE RESULTS
The eyes were examined to determine ocular effects using the parameters of corneal thickness (swelling), corneal opacity and fluorescein retention. Defined scoring scales were used for each parameter to define the severity of effects into four categories (I-IV) as specified in the guideline.

Results and discussion

In vivo

Results

Irritation parameter:

other: Irritation Index

Basis:

mean

Score:

38

Remarks on result:

other: Irritation index = maximum mean corneal swelling + maximum mean opacity (x 20) + mean fluorescein score (x 20)

Irritant / corrosive response data:

The test material caused very slight corneal swelling, slight corneal opacity and very slight fluorescein retention.
The negative control eyes did not show any corneal effect and demonstrated that the general conditions during the test were adequate.
The positive control eyes showed severe corneal effects (calculated Irritation Indices ranged from 152 to 155) and demonstrated the suitability and sensitivity of the ICE to detect severe eye irritants.

Any other information on results incl. tables

Table 1 Mean Values and Irritancy Categories

Time Interval (minutes)	Swelling %	Opacity	Fluorescein Retention
30 75 120 180 240	1 1 2 3 4	0.5 1.0 1.0 1.0 1.0	0.7
Irritancy Category	I	II	II

 

Table 2 Individual Values

Time Interval (minutes)	Eye Number	Swelling %	Opacity	Fluorescein Retention
30	22 25 28	2 2 0	0.5 0.5 0.5	1.0 0.5 0.5
75	22 25 28	2 2 0	1.0 1.0 1.0
120	22 25 28	2 3 0	1.0 1.0 1.0
180	22 25 28	2 3 3	1.0 1.0 1.0
240	22 25 28	3 5 5	1.0 1.0 1.0

Applicant's summary and conclusion

Interpretation of results:

not classified

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Under the conditions of this study, the test material did not cause any corrosive or severe irritancy effects and as such requires no classification in accordance with EU criteria.

Executive summary:

The potential of the undiluted test material to cause corrosive or severe irritancy effects to the eye was investigated in vitro in an Isolated Chicken Eye (ICE) study conducted in accordance with the standardised guideline OECD 438 under GLP conditions.

The isolated chicken eyes were exposed to the test material for 10 seconds followed by a 20 mL saline rinse. The eyes were evaluated after 30, 75, 120, 180 and 240 minutes for the adverse eye effects of corneal thickness (expressed as corneal swelling), corneal opacity and fluorescein retention of damaged epithelial cells. Physiological saline (0.9 %) was used as negative control whilst benzalkonium chloride (as a 5 % w/w aqueous solution) served as positive control.

The test material caused very slight corneal swelling, slight corneal opacity and very slight fluorescein retention. The negative control eyes did not show any corneal effect and demonstrated that the general conditions during the test were adequate. The positive control eyes showed severe corneal effects (calculated Irritation Indices ranged from 152 to 155) and demonstrated the suitability and sensitivity of the ICE to detect severe eye irritants.

Under the conditions of this study, the test material did not cause any corrosive or severe irritancy effects and as such requires no classification in accordance with EU criteria.