Registration Dossier

Administrative data

Endpoint:
eye irritation
Remarks:
other: In vitro preliminary test and in vivo main test
Type of information:
experimental study
Adequacy of study:
key study
Study period:
18 September 2012 to 25 March 2013
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Fully GLP compliant and in accordance with current test guidelines

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2013
Report Date:
2013

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying Ocular Corrosives and Severe Irritants)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.5 (Acute Toxicity: Eye Irritation / Corrosion)
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 405 (Acute Eye Irritation / Corrosion)
Deviations:
no
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
other: Viscous semi-solid
Details on test material:
Name: TOFA_TETA_PAA_BADGE_CGE_Adduct
CAS number: 186321-96-0
Batch number: WA520
Purity: 100%
Expiry date: 28 Feb 2014
Date of receipt: 03 Sep 2012
Storage: stored in a sealed container, at 15 to 25ºC, in the dark.

Test animals / tissue source

Species:
rabbit
Strain:
New Zealand White
Details on test animals or tissues and environmental conditions:
Bovine corneas supplied by a local abattoir. The eyes were removed after slaughter, completely immersed in Hanks’ Balanced Salt Solution (containing penicillin at 100 IU/mL and streptomycin at 100 µg/mL) in a suitably sized container and transported on the same day to the testing facility. All corneas were preserved in 10% Neutral Buffered Formalin.

Main test details
TEST ANIMALS
- Source: Harlan UK Ltd., Bicester
- Age at study initiation: 17 to 18 weeks
- Weight at study initiation: Not reported
- Housing: the rabbit was housed in a cage that conformed to the 'Code of Practice for the Housing and Care of Animals Used in Scientific Procedures' (Home Office, London, 1989).
- Diet (e.g. ad libitum): Global Diet 2930C (Harlan Teklad, Bicester, UK), ad libitum
- Water (e.g. ad libitum): Mains water, ad libitum
- Acclimation period: 7 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 15 to 22°C
- Humidity (%): 45%
- Air changes (per hr): The animal room was designed to permit 15 to 20 air changes per hour.
- Photoperiod (hrs dark / hrs light): The rooms were illuminated by fluorescent strip-lights for twelve hours daily.

IN-LIFE DATES: From: 10 December 2012 To: 14 December 2012

Test system

Vehicle:
unchanged (no vehicle)
Amount / concentration applied:
Preliminary test: A volume of 750 µL of the test article was applied to each of three corneas
Main test: One dose consisting of 0.1 mL of undiluted test article was instilled into the left conjunctival sac of a single New Zealand White rabbit (the sentinel).
Details on study design:
Preliminary test: A volume of 750 µL of the test article was applied to each of three corneas followed by a 10 minute incubation at 32°C ± 1°C. After this incubation, each cornea was washed with media containing phenol red (as a pH indicator) until this indicator showed no pH effect occurring (and demonstrating that the test article had been removed successfully). The corneas were then washed once in media without phenol red and incubated at 32°C ± 1°C for 120 ± 10 minutes, the opacities measured and the the anterior chamber emptied. For the permeability endpoint, 1 mL of sodium fluorescein (4 mg/mL solution) was added into the anterior chamber and the corneas were incubated in the vertical position for 1.5 hours ± 5 minutes. Following this period, the media in the posterior chamber was removed and held in a labelled tube. Three 350 μL aliqots of this media (per cornea) were analysed for optical density at 490 nanometers (OD490).
A volume of 750 µL of the negative or positive control was similarly applied to further groups of three corneas. These groups were subject to the procedures detailed above.
Main test: Before the first animal could be dosed, the pH of the test article was checked. Since this was within the acceptable range of pH 2.0 to 11.5, the study continued.
One dose consisting of 0.1 mL of undiluted test article was instilled into the left conjunctival sac of a single New Zealand White rabbit (the sentinel). The lower eyelid was gently prised away from the eyeball to create a receptacle for the dose. After instillation the eyelids were held closed for a few seconds to prevent loss of the dose. The right eye remained untreated and served as a control to the treated eye. The day of dosing was designated as Day 1.
The condition of the treated eye of the sentinel was assessed for a period of three days. As severe ocular responses were observed, the animal was humanely killed and no further animals were treated.

Results and discussion

In vivo

Resultsopen allclose all
Irritation parameter:
cornea opacity score
Basis:
mean
Time point:
other: 72 Hours
Score:
2
Max. score:
2
Reversibility:
not reversible
Irritation parameter:
iris score
Basis:
mean
Time point:
other: 72 hours
Score:
1
Max. score:
1
Reversibility:
not reversible
Irritation parameter:
conjunctivae score
Remarks:
Redness
Basis:
mean
Time point:
other: 72 hours
Score:
3
Max. score:
3
Reversibility:
not reversible
Irritation parameter:
conjunctivae score
Remarks:
Chemosis
Basis:
mean
Time point:
other: 72 hours
Score:
4
Max. score:
4
Reversibility:
not reversible
Irritation parameter:
conjunctivae score
Remarks:
Discharge
Basis:
mean
Time point:
other: 72 hours
Score:
2
Max. score:
2
Reversibility:
not reversible
Irritant / corrosive response data:
Scattered or diffuse areas of cornea opacity were noted 4 hours after instillation, with easily discernible translucent areas of corneal opacity noted from 24 to 72 hours after treatment.
Iridial inflammation was noted at all time points from 30 minutes to 72 hours after instillation.
Moderate conjunctival irritation was noted 30 minutes, 1, 4 and 24 hours after instillation with severe conjunctival irritation noted 28, 48 and 72 hours after instillation.
Maximal conjunctival inflammation (swelling grade 4) was noted 24 hours after instillation and was associated with redness grade 3 at 48 hours after instillation. As there was no evidence of recovery by the 72-hour observation, the animal was humanely killed after the 72-hour observation in accordance with the UK Home Office Guidelines on Eye Irritation Tests (published March 2006).

Applicant's summary and conclusion

Interpretation of results:
Category 1 (irreversible effects on the eye) based on GHS criteria
Remarks:
Migrated information
Conclusions:
The test article was classified as causing irreversible effects on the eye (Category 1) according to the Globally Harmonised System of Classification and Labelling of Chemicals (GHS).
Executive summary:

This study was conducted to determine whether the test article has a potential to cause corrosion or irritation to the eye.

Initially the test article was evaluated using an in vitro test system, the bovine corneal opacity and permeability assay (BCOP). A volume of 750 µL of the test article formulation was applied to each of three corneas followed by a four hour incubation at 32°C ± 1°C. After this incubation, each cornea was washed with media containing phenol red followed by media without phenol red and then incubated at 32°C ± 1°C for 120 ± 10 minutes. The opacities were then measured and then the anterior chamber emptied. For the permeability endpoint, sodium fluorescein solution was added into the anterior chamber and the corneas were incubated in the vertical position for 1.5 hours ± 5 minutes. Following this period, the media in the posterior chamber was removed and three 350 μL aliquots of this media (per cornea) were analysed for optical density at 490 nanometers (OD490).

A volume of 750 µL of the negative or positive control was similarly applied to further groups of three corneas. These corneas were subject to the procedures detailed above.

Corneas treated with the test article were noted to be slightly opaque. Corneas treated with the positive control article were cloudy and blistered. The mean opacity reading for the test article was 8.3, for the negative control was 0.0 and for the positive control was 75.7. The mean group corrected optical density for the test article was 0.226 for the negative control was 0.0 and for the positive control was 0.595. The test article produced an IVIS score of 11.73 and was not considered to be corrosive or severely irritating to the eye according to the BCOP assay.

An in vivo eye irritation test was therefore conducted. The undiluted test article (0.1 mL) was instilled into one conjunctival sac of a New Zealand White rabbit on Day 1. Ocular reactions were assessed for three days after treatment.

Scattered or diffuse areas of cornea opacity were noted 4 hours after instillation, with easily discernible translucent areas of corneal opacity noted from 24 to 72 hours after treatment. Iridial inflammation was noted at all time points from 30 minutes to 72 hours after instillation. Moderate conjunctival irritation was noted 30 minutes, 1, 4 and 24 hours after instillation with severe conjunctival irritation noted 24, 48 and 72 hours after instillation.

Maximal conjunctival inflammation (swelling grade 4) was noted 24 hours after instillation and was associated with redness grade 3 at 48 hours after instillation. As there was no evidence of recovery by the 72-hour observation, the animal was humanely killed after the 72-hour observation in accordance with the UK Home Office Guidelines on Eye Irritation Tests (published March 2006).