3-[3-(1,1,1,3,5,5,5-heptamethyltrisiloxan-3-yl)propoxy]- 2-hydroxypropyl methacrylate and 1-[3-(1,1,1,3,5,5,5-heptamethyltrisiloxan-3-yl)propoxy]-3-hydroxypropan-2-yl methacrylate

Eye irritation

Administrative data

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

The study was performed between 03 June 2008 and 05 June 2008.

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

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

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: SkinEthic HCE model

Deviations:

no

Test guideline 2

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 492 (Reconstructed Human Cornea-like Epithelium (RhCE) Test Method for Identifying Chemicals Not Requiring Classification and Labelling for Eye Irritation or Serious Eye Damage)

Deviations:

no

Principles of method if other than guideline:

The test is based on the hypothesis that irritant chemicals are able to penetrate the stratum corneum of the SkinEthic HCE model and are sufficiently cytotoxic to cause cell death in the underlying cell layers.

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

10-hydroxy-2,2,4-trimethyl-4-[(trimethylsilyl)oxy]-3,8-dioxa-2,4-disilaundecan-11-yl 2-methylprop-2-enoate; 11-hydroxy-2,2,4-trimethyl-4-[(trimethylsilyl)oxy]-3,8-dioxa-2,4-disilaundecan-10-yl 2-methylprop-2-enoate

EC Number:

700-334-3

Molecular formula:

C17H38O6Si3

IUPAC Name:

10-hydroxy-2,2,4-trimethyl-4-[(trimethylsilyl)oxy]-3,8-dioxa-2,4-disilaundecan-11-yl 2-methylprop-2-enoate; 11-hydroxy-2,2,4-trimethyl-4-[(trimethylsilyl)oxy]-3,8-dioxa-2,4-disilaundecan-10-yl 2-methylprop-2-enoate

Test material form:

other: liquid

Details on test material:

Sponsor's identification: SiMAA2
Description: clear colourless liquid
Lot number: 003068
Date received: 25 March 2008
Storage conditions: approximately 4°C in the dark

Test animals / tissue source

Species:

other: SkinEthic Reconstituted Human Corneal model

Strain:

other: Reconstituted HCE model

Details on test animals or tissues and environmental conditions:

Tissue: Consists of the transformed human kerotinocytes of the cell line HCE that formed a corneal epithelial tissue (mucosa), devoid of stratum corneum, resembling, histologically, the mucosa of the human eye.
- Source: HCE, SkinEthic Laboratories, Nice, France



ENVIRONMENTAL CONDITIONS
- Temperature (°C): Room temperature
- Storage: wells containing 1ml of maintenance medium
-Incubation: Incubated over night at 37 deg C, 5% CO2 in air

Test system

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 30 µl
- Concentration (if solution): NA

Duration of treatment / exposure:

10 minutes

Observation period (in vivo):

None, at the end of the expsoure period, each skinEthic tissue was rinsed. The rinsed tissues (two per group) were taken for MTT loading. The remaining tissues were retained for possible histopathology. Following MTT loading the reduced MTT was extracted from the tissues.

Number of animals or in vitro replicates:

Not applicable as tissues were used. Triplicate SkinEthic tissues were used.

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing (if done): At the end of the relevant exposure period, each tissue insert was removed from the well using forceps and rinsed using a wash bottle containing DPBS.
- Time after start of exposure: 10 minutes

SCORING SYSTEM: After exposure of tissues to the test material, the tissues are removed and rinsed. The rinsed tissues are taken for MTT loading, were the reduced MTT is extracted from the tissues. After extraction, the absorbency of triplicate aliquots of the extracted MTT solution for each SkinEthic tissue was measured (OD50). Data is presnted in the form of % viability (MTT conversion relative to negative controls).


TOOL USED TO ASSESS SCORE: The test material was classified according to the following criteria:
* If the % relative viability was greater than or equal to 60% the test material was considered to not likely to be a severe ocular irritant.
* If the % relative viability was less than 60% the test material was considered to be irritant.

Results and discussion

In vitro

Results

Irritation parameter:

other: Relative mean viability (%)

Run / experiment:

10 minute exposure

Value:

104.8

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: Not likely to be a severe ocular irritant

Any other information on results incl. tables

Assessment of Direct test Material Reduction of MTT

The test material was not able to directly reduce MTT.

Assessment of Eye Irritation Potential:

The mean OD₅₄₀ values and mean viabilities for each treatment group are given in Table 1.

The relative mean viability of the test material treated tissues after a 10 minute exposure was 104.8%.

It was considered unnecessary to proceed with tissue histopathology.

Table1:

Material	Mean OD540	% Viability
Negative Control	0.888	100*
Positive Control 0.5% w/v	0.566	63.7
Positive Control 1% w/v	0.376	42.3
Test Material	0.931	104.8

* =          The mean viability of the negative tissues is set at 100%

Quantitative Evaluation of Tissue Viability (MTT Uptake Visaul Assessment)

The quantitative evaluation of tissue viability is presented in table 2.

The negative control and test material treated tissues appeared blue. This was considered to be indicative of viable tissue.  The positive control material at each concentration appeared blue/white and was considered to be indicative of semi‑viable tissue.

Table 2:

Material	Score
	Tissue 1	Tissue 2
Negative Control	-	-
Positive Control 0.5% w/v	+	+
Positive Control 1% w/v	+	+
Test Material	-	-

Applicant's summary and conclusion

Interpretation of results:

other: not irritating

Conclusions:

According to the protocol followed the test material was considered not likely to be a severe ocular irritant (NI).

Executive summary:

Introduction.  The purpose of this study was to determine the eye irritation potential of the test materials using the SkinEthic Reconstituted Human Corneal model (HCE, SkinEthic Laboratories, , ) after a treatment period of 10 minutes.  The test is based on the hypothesis that irritant chemicals are able to penetrate the corneal epithelial tissue and are sufficiently cytotoxic to cause cell death.

Methods.  The experimental design of the study consists of a test for Direct Reduction of MTT by the test materials, followed by the main test.

For the main test, triplicate SkinEthic tissues were treated with 30 ml of the test material for 10 minutes.  Triplicate tissues treated with 30 µl of Solution A served as the negative control and triplicate tissues treated with 30 µl of Sodium Dodecyl Sulphate at concentrations of 1% w/v and 0.5% w/v served as the positive control.

At the end of the exposure period each SkinEthic tissue was rinsed.  The rinsed tissues (two per group) were taken for MTT loading.  The remaining tissues were retained for possible histopathology.  Following MTT loading the reduced MTT was extracted from the tissues.

After extraction the absorbency of triplicate aliquots of the extracted MTT solution for each SkinEthic tissue was measured (OD₅₄₀).  Data is presented in the form of % viability (MTT conversion relative to negative controls).

The test material was classified according to the following criteria:

i)                    If the % relative viability was ≥ 60% the test material was considered to not likely to be a severe ocular irritant.

ii)                   If the % relative viability was 60% the test material was considered to be irritant.

Results.  The relative mean viability of the test material treated tissues after a 10 minute exposure was 104.8%.

It was considered unnecessary to proceed with tissue histopathology.

Quality criteria.  The quality criteria required for acceptance of results in the test were satisfied.

Conclusion. According to the protocol followed the test material was considered not likely to be a severe ocular irritant (NI).