(R)-lactic acid

Eye irritation

Administrative data

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1996-01-18 to 1996-03-12

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1996

Report date:

1996

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

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

Version / remarks:

Study performed prior to validation and publication of the test guideline.

Deviations:

yes

Remarks:

No positive controls

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

L-(+)-lactic acid

EC Number:

201-196-2

EC Name:

L-(+)-lactic acid

Cas Number:

79-33-4

Molecular formula:

C3H6O3

IUPAC Name:

(2S)-2-hydroxypropanoic acid

Test animals / tissue source

Species:

other: Chicken Enucleated Eye Test

Test system

Vehicle:

unchanged (no vehicle)

Amount / concentration applied:

Concentration: 100 %
Amount applied: 0.03 mL

Duration of treatment / exposure:

10 seconds

Number of animals or in vitro replicates:

3

Details on study design:

Approximately 7 weeks old, male or female chickens (ROSS, spring chickens), body weight range approximately 2.5-3.0 kg, were used as eye-donors. Heads of these animals were obtained from poultry slaughterhouse v.d. Bor, Amersfoortseweg 118, Nijkerkerveen, the Netherlands. Heads of the animals 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 (3 heads per box) 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. Within 2 hours after kill, eyes were carefully dissected and placed in a superfusion apparatus using the following procedure: First the eye-lids were carefully removed without damaging the cornea and a small drop of Fluorescein sodium BP 2% w/v (Minims, Smith & Nephew Ltd., Romford, England) was applied to the corneal surface for a few seconds and subsequently rinsed off with isotonic saline of ambient temperature. Next, the head with the fluorescein-treated cornea was examined with a slit-lamp microscope (Slit-lamp 900 CN, 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 off 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 (TNO, Zeist, the Netherlands). 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 rate of ca 0.10-0.15 mL/min (peristaltic pump, Desaga STA 131900, Heidelberg, Germany). The chambers of the superfusion apparatus as well as the saline were temperature controlled at 32 ± 1.5 °C (waterpump, Thermomix 1441, B. Braun Melsungen AG, Melsungen, Germany). After placing in the superfusion apparatus, the eyes were examined again with the slit-lamp microscope to ensure that they were not damaged. Corneal thickness was measured using the Depth Measuring Attachment no. II for the Haag-Streit slit-lamp microscope. Thickness of the cornea 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 corneal thickness of the eyes, or 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, if necessary. Per test sample, three eyes were selected for testing, whereas one additional eye was rinsed with isotonic saline only and served as a control of the experimental conditions. After an equilibration period of 45-60 minutes, the corneal thickness of the eyes were measured again to determine the zero reference value for corneal swelling calculations. At time t = 0, i.e. immediately after the zero reference measurement, each of the three test samples was applied to the designated test eyes. The following procedure applied for each test eye:
The clamp holding the eye was placed on paper tissues outside the chamber with the cornea facing upwards. Each of the two liquid samples was applied in an amount of 0.03 mL from a micropipette (Nichiryo Co., Ltd., model 8100, Tokyo, Japan), in such a way that the entire surface of the cornea was bathed with the test material. With the solid sample, the cornea was powdered with 0.03 g of the test material. After a total exposure period of 10 seconds, the corneal surface was rinsed thoroughly with 20 mL of isotonic saline of ambient temperature. After rinsing, each eye in the holder was returned to its chamber.
The control eye and test eyes were examined at 0, 30, 75, 120, 180 and 240 minutes after treatment, using the criteria and scoring system, given in the annex. Fluorescein retention was only scored at 30 minutes after treatment. All examinations were carried out with the slit-lamp microscope.

Results and discussion

In vitro

Results

Results 1

Irritation parameter:

cornea opacity score

Remarks:

after 240 min

Run / experiment:

1

Value:

ca. 4

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

not examined

Remarks on result:

positive indication of irritation

Results 2

Irritation parameter:

cornea opacity score

Remarks:

after 240 min

Run / experiment:

2

Value:

ca. 4

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

not examined

Remarks on result:

positive indication of irritation

Results 3

Irritation parameter:

cornea opacity score

Remarks:

after 240 min

Run / experiment:

3

Value:

ca. 4

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

not examined

Remarks on result:

positive indication of irritation

Results 4

Irritation parameter:

percent corneal swelling

Remarks:

after 240 min

Run / experiment:

1

Value:

ca. 31

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

not examined

Remarks on result:

positive indication of irritation

Results 5

Irritation parameter:

percent corneal swelling

Remarks:

after 240 min

Run / experiment:

2

Value:

ca. 26

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

not examined

Remarks on result:

positive indication of irritation

Results 6

Irritation parameter:

percent corneal swelling

Remarks:

after 240 min

Run / experiment:

3

Value:

ca. 28

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

not examined

Remarks on result:

positive indication of irritation

Results 7

Irritation parameter:

fluorescein retention score

Remarks:

after 30 min

Run / experiment:

1

Value:

ca. 3

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

not examined

Remarks on result:

not determinable

Results 8

Irritation parameter:

fluorescein retention score

Remarks:

after 30 min

Run / experiment:

2

Value:

ca. 3

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

not examined

Remarks on result:

positive indication of irritation

Results 9

Irritation parameter:

fluorescein retention score

Remarks:

after 30 min

Run / experiment:

3

Value:

ca. 3

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

not examined

Remarks on result:

positive indication of irritation

Applicant's summary and conclusion

Interpretation of results:

Category 1 (irreversible effects on the eye) based on GHS criteria

Conclusions:

In conclusion, based on the given conditions the test item is considered severely irritating to eyes.

Executive summary:

Lactic acid 88% (sample code HS88), was examined undiluted for eye irritating/corrosive potential in an ex vivo bioassay, namely the Enucleated Eye Test with chicken eyes (CEET). The eyes were collected as waste material from a slaughterhouse for chickens, which were killed for human consumption. HS88 induced severe corneal effects. On the basis of the results obtained with this in vitro (ex vivo) assay, HS88 can be considered severely irritating to eyes.