2-((E)-(4-((4-(1,3-dioxo-1,3-dihydro-2H-hetereocarbapolycyclo-2-yl)alkyl)(ethyl)amino)-2-methyl)diazenyl)-5-nitro-1,3-dicarbonitrile, polyphenyl

Eye irritation

Administrative data

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2008-08-28

Reliability:

1 (reliable without restriction)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2008

Report date:

2008

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

other: The Bovine Corneal Opacity and Permeability Assay (BCOP) will be performed according to the INVITTOX (UK) protocol no. 98 "The Bovine Corneal Opacity and Permeability Assayn, dated February 1994.

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: Bovine Corneal Opacity and Permeability (BCOP) Assay, SOP of Microbiological Associates Ltd., UK, Procedure Details, April 1997.

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Reference

Name:

Unnamed

Type:

Constituent

Details on test material:

- Name of the test material (as cited in the study report: FAT 40842/A TE
- Substance type: coloring dye
- Physical state: solid, dark bluish green powder
- Analytical purity: 96%
- Lot/batch No.: Blau DRI 2098 Op 1/07
- Expiration date of the lot/batch: June 30, 2014
- Storage condition of test material: at room temperature at about 20 ºC

Test animals / tissue source

Species:

other: in vitro study

Strain:

other: in vitro study

Details on test animals or tissues and environmental conditions:

not applicable

Test system

Vehicle:

unchanged (no vehicle)

Controls:

not required

Amount / concentration applied:

not applicable

Duration of treatment / exposure:

240 ± 5 min.

Observation period (in vivo):

not applicable

Number of animals or in vitro replicates:

not applicable

Results and discussion

In vivo

Results

Irritation parameter:

other: In Vitro Test. See Point "Remarks on Results"

Basis:

other: 3 corneas

Time point:

other: after 240 min treament

Score:

> -2.5 - < -1.6

Max. score:

-1.6

Reversibility:

other: not applicable

Remarks on result:

other: In Vitro Score

Any other information on results incl. tables

Results after 240 Minutes Incubation Time

Test Group	Opacity value = Difference (t240-t0) of Opacity		Permeability at 490 nm (OD490)		In vitroScore	Meanin vitro Score	Proposedin vitro Irritation Scale
		Mean		Mean
Negative Control	2	1.67	0.056	0.058	2.84	2.54	non
Negative Control	2		0.070		3.05
Negative Control	1		0.048		1.72
Positive Control	119.33*		0.820*		131.63	129.35	very severe
Positive Control	120.33*		1.238*		138.90
Positive Control	102.33*		1.011*		117.50
FAT 40842/A TE	-2.67*		0.015*		-2.44	-2.2 = 0	non
FAT 40842/A TE	-2.67*		0.065*		-1.69
FAT 40842/A TE	-2.67*		0.013*		-2.47

*corrected values

SCORING

In vitroScore	Proposedin vitroirritation Scale
0 - 3	Non eye irritant
3.1 – 25	Mild eye irritant
25.1 – 55	Moderate eye irritant
55.1 – 80	Severe eye irritant
> 80.1	Very severe eye irritant

Validity

Criteria: The test was acceptable if the positive control caused an at least moderate effect.

The positive control (10% (v/v) Benzalconium chloride) showed clear opacity and distinctive permeability of the corneas and therefore, is classified as very severe eye irritant. The in vitro score was calculated as 129.35.

Applicant's summary and conclusion

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: other: Invitox (UK) protocol no. 98 aand calculated "in vitro" score = 0

Conclusions:

In conclusion, it can be stated that in this study and under the experimental conditions reported, the test item FAT 40842/A TE is not considered to be an eye irritant.

Executive summary:

This in vitro study was performed to assess the corneal irritation and damage potential of FAT 40842/A TE by means of the BCOP assay using fresh bovine corneas.

After a first opacity measurement of the fresh bovine corneas (t0), the neat test item FAT 40842/A TE, the positive, and the negative controls were applied to corneas and incubated for 240 minutes at 32 ± 2 °C in cMEM medium, supplemented with 10% FCS. After the incubation phase the test item, the positive, and the negative controls were each rinsed from the corneas and opacity was measured again (t240).

After the opacity measurements permeability of the corneas was determined while application of 1 mL of a fluorescein solution for about 90 minutes at 32 ± 2 °C in a horizontal position. The coming out liquid was measured spectrophotometrically.

With the negative control (0.9% NaCl solution) neither an increase of opacity nor permeability of the corneas could be observed.

The positive control (10% (v/v) Benzalconium chloride) showed clear opacity and distinctive permeability of the corneas and therefore, is classified as very severe eye irritant.

The test item FAT 40842/A TE did not cause any opacity or permeability of the corneas compared with the results of the negative control. The calculated in vitro score was 0 and therefore, the test item was classified as non eye irritant.

In conclusion, it can be stated that in this study and under the experimental conditions reported, the test item FAT 40842/A TE is not considered to be an eye irritant.