ethanol water extract from husk of Chenopodium quinoa, Chenopodiaceae

Eye irritation

Administrative data

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

21 September 2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Well conducted and well described study in accordance with GLP and OECD Guideline 437 without any deviation.

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Cross-reference 2

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Report date:

2010

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

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

ethanol water extract from husk of Chenopodium quinoa, Chenopodiaceae

IUPAC Name:

ethanol water extract from husk of Chenopodium quinoa, Chenopodiaceae

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Details on test material:

- Name of test material: MEXORYL SCK
- Physical state: Beige powder
- Analytical purity: saponines content (determined by HPLC assay) 55.8 % w/w
- Lot/batch No.: R0069579A 003 P 001
- Shelf life/Retest date: April 2011
- Storage condition of test material: Stored at room temperature and away from light and moisture

Test animals / tissue source

Species:

other: bovine eye

Strain:

not specified

Details on test animals or tissues and environmental conditions:

TEST SYSTEM
- Eyes selection: from reception of eyes: detection of defects of the cornea, elimination of eyes presenting defects, elimination of the too big eyes, all stages were performed by avoiding touching corneas, in order not to damage them.
- Preservation of the corneas: eyes were placed in buffered Hanks medium enriched with antibiotics and preserved at 5 ± 3 °C for maximum duration of 24 h.

Test system

Vehicle:

other: distilled water

Controls:

other: negative control: 0.9 % w/v sodium chloride solution

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 750 µL ± 8 µL per cornea
- Concentration (if solution): Diluted to 20 % (w/w) in distilled water
- pH of the preparation : 5.5

Duration of treatment / exposure:

- Contact timepoint: 4 h ± 10 minutes at 32 ± 1 °C.

Observation period (in vivo):

- Corneal opacity was measured pre-treatment, post-treatment and post-incubation (after 4 h ± 10 minutes of incubation at 32 ± 1 °C).
- Application of Fluorescein (5 mg/mL) and corneal permeability was measured after 90 ± 5 min of incubation 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:

Details of test procedure: Eyes were placed in buffered Hanks medium enriched with antibiotics and preserved at 5 ± 3 °C for maximum duration of 24 h. Corneas were mounted in corneal holders and preincubated in nutritive medium in water-bath at 32 ± 1 °C for at least one hour. Before the treatment, opacity measurement was performed using an OP KIT opacitometer. The test item (750 µL ± 8 µL) was applied on each cornea and incubated for 4 h ± 10 minutes at 32 ± 1 °C. At the end of the exposure period the test item and control items were removed from the anterior chamber and the cornea was rinsed three times with nutritive medium containing phenol red at 32 ± 1 °C before a final rinse with nutritive medium. The anterior chamber was refilled with nutritive medium. Measurement of the opacity OPT2 (OPT “2 hours”) of each cornea vs an empty holder (containing nutritive medium only). Following the opacity measurement the permeability of the corneas to sodium fluorescein was evaluated. The medium from the anterior chamber was removed and replaced with 1 mL of fluorescein solution (5 mg/mL) and the holders were incubated at 32 ± 1 °C for 90 ± 5 minutes. After incubation the medium in the posterior chamber of each holder was decanted and retained. 360 µL of medium representing each cornea was applied to a designated well on a 96-well plate and the optical density at 490 nm (OD490) was measured.

Results and discussion

In vivo

Results

Irritation parameter:

other: In Vitro Irritancy Score (IVIS)

Basis:

mean

Time point:

other: 4 h

Score:

82.2

Reversibility:

not reversible

Remarks on result:

other: score: 82.2 ± 4.9

Irritant / corrosive response data:

- In Vitro Irritancy Score (IVIS) were 82.2 ± 4.9 and 193.0 ± 2.4 for test item and positive controls, respectively.

Other effects:

None

Any other information on results incl. tables

Table 7.3.2/1: In vitro eye irritation – results

Holder No	Treatment	OPT0	OPT2	OPT2 – OPT0	Corrected opacity	DO	Corrected OD	Score OPc + 150DOc
97	Negative control	0	0	0		0.070
440		3	3	0		0.091
86		0	0	0		0.071
Comments: Nil			Mean		0.0	0.077
			SD		0.0	0.012
29	Imidazole 20 %	1	146	145	145.0	3.256	3.179	192.7
300		0	147	147	147.0	3.312	3.235	195.5
210		0	142	142	142.0	3.332	3.255	190.8
Comments: Detachment of epithelium and edema			Mean		144.7	3.223		193.0
			SD		2.5	0.039		2.4
2	MEXORYL SCK	2	27	25	25.0	4.092	4.015	85.2
12	I.E.C: 101382 0000B455	4	24	20	20.0	3.848	3.771	76.6
58	Diluted to 20 %	0	28	28	28.0	3.866	3.779	84.7
Comments: Detachment of epithelium and edema			Mean		24.3	3.855		82.2
			SD		4.0	0.139		4.9

Applicant's summary and conclusion

Interpretation of results:

corrosive

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Under the test conditions, MEXORYL SCK applied diluted to 20 % (w/w) in distilled water, is classified as corrosive or severe irritant for the isolated bovine cornea, after 4 h of contact.

Executive summary:

In an in vitro eye irritation study performed according to the OECD Guideline 437 and in compliance with GLP, test item, MEXORYL SCK diluted to 20 % (w/w) in distilled water (750 µL ± 8 µL per cornea) was applied to isolated bovine corneas mounted in corneal holders, and incubated for 4 h ± 10 minutes at 32 ± 1 °C. Three corneas were used for each treated series (test item; negative control; positive control: Imidazole 20 %). The two endpoints, corneal opacity and corneal permeability were measured and In Vitro Irritancy Score (IVIS) was calculated. 

In Vitro Irritancy Score (IVIS) were 82.2 ± 4.9 and 193.0 ± 2.4 for test item and positive controls, respectively.

Under the test conditions, MEXORYL SCK applied diluted to 20 % (w/w) in distilled water, is classified as corrosive or severe irritant for the isolated bovine cornea, after 4 h of contact.