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Toxicological information

Eye irritation

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Administrative data

Endpoint:
eye irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
2014
Report Date:
2014
Reference Type:
study report
Title:
Unnamed
Year:
2014
Report Date:
2014
Reference Type:
study report
Title:
Unnamed
Year:
2015
Report Date:
2015

Materials and methods

Test guideline
Guideline:
other: EpiOcular
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): C17 Methacrylate
- Physical state: liquid
- Analytical purity: >99 %
- Lot/batch No.: S732030105

Test animals / tissue source

Species:
other: in vitro
Strain:
other: in vitro

Test system

Vehicle:
unchanged (no vehicle)
Details on study design:
METHOD
TEST SYSTEM

The EpiOcular™ model (OCL-200) is a three-dimensional non-keratinized tissue construct composed of normal human derived epidermal keratinozytes used to model the human corneal epithelium. The EpiOcular™ tissues (surface 0.6 cm2)are cultured on specially prepared cell culture inserts (MILLICELLs®, 10 mm Ø ) and are commercially available as kits (EpiOcular™ 200), containing 24 tissues on shipping agarose.

EXPERIMENT AL PROCEDURE
Two tissues are treated with each test substance, the PC and NC, respectively. There are two separate protocols for liquids and solids, differing in exposure time and post-incubation period. Due to the physical condition of the test substance the protocol for solids is applied.

The tissues will be transferred to sterile 6-well plates with 1 ml pre-warmed assay medium and preconditioned in the incubator at 37°C for 16 - 24 hours. After the pre-incubation the tissues are pre-treated with 20 μl of PSS in order to wet the tissue surface. The tissues are incubated at standard culture conditions for 30 minutes.
After the incubation I postincubation period, the assay medium is replaced by 0.3 ml MTT solution and the tissues are incubated in the incubator for 3 hours. After incubation, the tissues are washed with PBS to stop the MTT-incubation.
The formazan that is metabolically produced by the tissues will be extracted by incubation of the tissues in 2 ml isopropanol at room temperature overnight or for at least 2 hours on a plate shaker (ea. 120 rpm). After shaking the isopropanol extract and piercing the tissues, 2 aliquots of each extract per tissue will be transferred to a 96-well microtiter plate. The optical density (OD570) will be determined spectrophotometrically using a filter with a wavelength of 570 nm.

Results and discussion

Any other information on results incl. tables

Individual and mean OD570 values, individual and mean viability values and inter-tissue variability

Test substance

 

tissue 1

tissue 2

mean

inter-tissue variability [%]

NC

mean OD570

1.809

1.818

1.813

 

viability

[% of NC]

99.7

100.3

100

0.5

14/0100-1

mean OD570

1.905

1.767

1.836

 

viability

[% of NC]

105.0

97.4

101

7.6

PC

mean OD570

0.455

0.562

0.508

 

viability

[% of NC]

25.1

31.0

28

5.9

The test substance is not able to reduce MTT directly.

The mean viability of the test-substance treated tissues was 101%.

Mean tissue viability

(%of negative control)

Prediction

 

50

Irritant

>5060

no prediction*

>60

non-irritant

*At presentnoprediction isperformedif the mean relative tissue viability with a test materialis> 50 ≤ 60%as the cut off valueiscurrently being evaluated toliein this range.

Applicant's summary and conclusion