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Eye irritation

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Administrative data

Endpoint:
eye irritation: in vitro / ex vivo
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Cross-referenceopen allclose all
Reason / purpose:
read-across source
Reference
Endpoint:
eye irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying i) Chemicals Inducing Serious Eye Damage and ii) Chemicals Not Requiring Classification for Eye Irritation or Serious Eye Damage)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Species:
cattle
Strain:
not specified
Details on test animals or tissues and environmental conditions:
Eyes from adult cattle (typically 12 to 60 months old) were obtained from a local abattoir as a by-product from freshly slaughtered animals. The eyes were excised by an abattoir employee after slaughter, and were placed in Hanks’ Balanced Salt Solution (HBSS) supplemented with antibiotics (penicillin at 100 IU/mL and streptomycin at 100 μg/mL). They were transported to the test facility over ice packs on the same day of slaughter. The corneas were prepared immediately on arrival.
Vehicle:
physiological saline
Controls:
yes, concurrent vehicle
yes, concurrent positive control
Amount / concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.75 mL
- Concentration (if solution): 20% w/v solution in sodium chloride 0.9% w/v.
Duration of treatment / exposure:
240 minutes
Duration of post- treatment incubation (in vitro):
Not applicable
Number of animals or in vitro replicates:
Three corneas
Details on study design:

SELECTION AND PREPARATION OF CORNEAS
The medium from both chambers of each holder was replaced with fresh complete EMEM. A pre-treatment opacity reading was taken for each cornea using a calibrated opacitometer. Three corneas were randomly allocated to the negative control. Three corneas were also allocated to the test item and three corneas to the positive control item.

NUMBER OF REPLICATES
Triplicate

NEGATIVE CONTROL USED
Sodium chloride 0.9% w/v

POSITIVE CONTROL USED
Imidazole

APPLICATION DOSE AND EXPOSURE TIME
The EMEM was removed from the anterior chamber of the BCOP holder and 0.75 mL of the test item preparation or control items were applied to the appropriate corneas. The holders were gently tilted back and forth to ensure a uniform application of the item over the entire cornea. Each holder was incubated, anterior chamber uppermost, at 32 ± 1 ºC for 240 minutes.

POST-INCUBATION PERIOD: no.

REMOVAL OF TEST SUBSTANCE
- Number of washing steps after exposure period: At least three.

METHODS FOR MEASURED ENDPOINTS:
- Corneal opacity: A post-treatment opacity reading was taken and each cornea was visually observed.
- Corneal permeability: passage of sodium fluorescein dye measured with the aid of microtiter plate reader (OD492)

SCORING SYSTEM: In Vitro Irritancy Score (IVIS)

DECISION CRITERIA: Decision criteria as indicated in the TG.
Irritation parameter:
in vitro irritation score
Run / experiment:
Mean
Value:
18
Vehicle controls validity:
valid
Negative controls validity:
not applicable
Positive controls validity:
valid
Other effects / acceptance of results:
OTHER EFFECTS:
- Visible damage on test system: The corneas treated with the test item were clear post treatment, with some test item remaining on the cornea.


ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: The negative control gave opacity of ≤2.4 and permeability ≤0.072. The negative control acceptance criteria were therefore satisfied.
- Acceptance criteria met for positive control: The positive control In Vitro Irritancy Score was within the range of 65.1 to 123.3. The positive control acceptance criterion was therefore satisfied.
Interpretation of results:
GHS criteria not met
Conclusions:
Calcium Titanium Trioxide produced an IVIS of 0.8 during an in vitro eye irritation study. No prediction can be made in regards to its classification as irritant or damaging to the eye.
Executive summary:

An in vitro Bovine Corneal Opacity and Permeability (BCOP) assay was performed in line with OECD Testing Guideline 437 (Bovine Corneal Opacity and Permeability Test Method). The study was GLP-compliant.

A solution of 20% w/v test material in sodium chloride 0.9% w/v was applied to three cattle corneas obtained from an abattoir at a volume of 0.75 mL, after which each cornea was incubated at 32±1°C for 240 minutes. Opacity and permeability of corneas were measured following exposition.

Negative and positive controls were performed and considered as valid.

An In Vitro Irritation Score (IVIS) of 18.0 was calculated for Calcium Titanium Trioxide from corneal opacity and permeability measurements. The substance is not irritant to the eye according to the guideline.

Since the result is between 3 and 55 no prediction can be made regarding the classification of Calcium Titanium Trioxide as irritant or damaging to the eye in accordance with with Regulation (EC) No.1272/2008.

Reason / purpose:
read-across source
Reference
Endpoint:
eye irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
From 19 July 2018 to 2 August 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
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
Remarks:
No deviation impacting on the quality of the study
GLP compliance:
yes (incl. certificate)
Species:
hamster
Strain:
other: Reconstructed human Cornea-like Epithelium
Details on test animals or tissues and environmental conditions:
The EpiOcularTM tissue model is composed of stratified human keratinocytes in a three-dimensional structure, reflecting the morphology and function of the human corneal epithelium found in vivo
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): ca. 50 mg
Duration of treatment / exposure:
Application for 6 hours ± 15 minutes
Duration of post- treatment incubation (in vitro):
25 minutes ± 2 minutes post-treatment immersion, and 18 hours ± 15 minutes post-treatment incubation
Number of animals or in vitro replicates:
Three replicates
Details on study design:
Preliminary test

The test item is first checked for its potential for MTT interference and solvent interference (water and isopropanol).

Main test overview

Day 0: On the day of receipt, EpiOcular tissues were pre-incubated overnight at 37°C, 5%
CO2, ≥95% RH.

Day 1: Exposure to, and removal of 50 mg of test item using a dosing spoon or 50µl of reference items for 6 hours ± 15 minutes, followed by a 25 minutes ± 2 minutes post-treatment immersion, and 18 hours ± 15 minutes post-treatment incubation.

Day 2: End of MTT viability test, optical density readings at 570 nm without reference filter
Irritation parameter:
other: viability
Remarks:
in %
Run / experiment:
Mean of three replicates
Value:
86.657
Vehicle controls validity:
not applicable
Negative controls validity:
valid
Positive controls validity:
valid

Name

Code

Mean of OD

SD of OD

Mean of viability
(%)

SD of viability (%)

CV %

Corrected Mean of viability (%)

Classification

Sterile water

NC

1.839

0.039

100.000

2.11

2.115

N/A

No-Category

Methyl Acetate

PC

0.668

0.262

36.312

14.243

39.225

N/A

No Prediction

Calcium Titanium Trioxide

TA1

1.594

0.138

86.661

7.497

8.650

86.657%

No-Category

Interpretation of results:
GHS criteria not met
Conclusions:
The percentage viability of Calcium Titanium Trioxide was 86.657%. It was concluded that the substance did not require a classification as irritant or damaging to eye in accordance with Regulation (EC) No.1272/2008.
Executive summary:

A Reconstructed human Cornea-like Epithelium (RhCE) test for identifying chemicals not requiring classification and labelling for eye irritation or serious eye damage was performed on Calcium Titanium Trioxide in accordance with the OECD Testing Guideline 492. The study was GLP-compliant.

After 6 hours ± 15 minutes exposure on the surface of EpiOcular reconstructed ocular epithelium and 18 hours ± 15 mins post-incubation time, the viability of the tissues was assessed and compared to a negative control. The percentage viability of Calcium Titanium Trioxide was determined to be 86.657%. All acceptance criteria were met during the study.

According to the OECD Testing Guideline 492 A test item is considered as "No Category" (not requiring classification under Regulation (EC) No.1272/2008), if the eye model viability after exposure and post-treatment incubation is >60%. It can therefore be concluded that Calcium Titanium Trioxide does not meet the criteria for classification as irritant or damaging to the eye.

Data source

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying i) Chemicals Inducing Serious Eye Damage and ii) Chemicals Not Requiring Classification for Eye Irritation or Serious Eye Damage)
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent

Test animals / tissue source

Species:
cattle

Results and discussion

In vitro

Results
Irritation parameter:
in vitro irritation score
Remarks on result:
no indication of irritation

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
The percentage viability of Magnesium Titanium Trioxide was 86.657%. It was concluded that the substance did not require a classification as irritant or damaging to eye in accordance with Regulation (EC) No.1272/2008. No prediction can be made in regards to its classification as irritant or damaging to the eye.