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Toxicological information

Eye irritation

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Administrative data

Endpoint:
eye irritation: in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
07 August 2018 - 18 September 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
Based on two previous performed in vitro eye irritation tests with Potassium Ferricyanide, no conclusion could be drawn on the eye irritating potential of the substance. In order to clarify classification it was decided to perform an in vivo study.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2018

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 405 (Acute Eye Irritation / Corrosion)
Version / remarks:
2017
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: EC No 440/2008, part B: "Acute Toxicity: Eye Irritation/Corrosion"
Version / remarks:
May 2008
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.2400 (Acute Eye Irritation)
Version / remarks:
August 1998
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: Appendix to Director General Notification, No. 12-Nousan-8147. Agricultural Production Bureau, Ministry of Agriculture, Forestry and Fisheries of Japan (JMAFF)
Version / remarks:
November 2000, including the most recent revisions.
Deviations:
no
GLP compliance:
yes

Test material

Constituent 1
Chemical structure
Reference substance name:
Tripotassium hexacyanoferrate
EC Number:
237-323-3
EC Name:
Tripotassium hexacyanoferrate
Cas Number:
13746-66-2
Molecular formula:
C6FeN6.3K
IUPAC Name:
tripotassium hexacyanoferrate
Test material form:
solid: particulate/powder
Details on test material:
Appearance: Dark-red crystalline powder
Storage: At room temperature protected from light

Test animals / tissue source

Species:
rabbit
Strain:
New Zealand White
Details on test animals or tissues and environmental conditions:
TEST ANIMALS
- Source: Charles River France, L’Arbresle, France
- Age at study initiation: approximately 13-14 weeks old
- Weight at study initiation: 2976 to 3221g
- Housing: individually, in cages with perforated floors. For psychological/environmental enrichment, animals were provided with shelters and wooden sticks.
- Diet: Pelleted diet for rabbits (Global Diet 2030 Teklad®, Mucedola, Milanese, Italy), once daily throughout the study period. Hay was fed in addition.
- Water: municipal tap-water, ad libitum
- Acclimation period: at least 5 days

It is considered that there were no known contaminants in the feed and water that would interfere with the objectives of the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 to 21
- Humidity (%): 48 to 90
- Air changes (per hr): 10 or more
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 09 August 2018 to 18 September 2018

Test system

Vehicle:
unchanged (no vehicle)
Controls:
yes, concurrent no treatment
Amount / concentration applied:
- Amount applied: 99.9mg - 110.2mg (0.1 mL) to the conjunctival sac of the eye.
Duration of treatment / exposure:
single treatment
Observation period (in vivo):
14 days and/or 21 days
Number of animals or in vitro replicates:
3
Details on study design:
REMOVAL OF TEST SUBSTANCE
- Washing (if done): no

SCORING SYSTEM: according to Draize scale

TOOL USED TO ASSESS SCORE: 2% fluorescein in water (pH adjusted to 7.0).

OBSERVATIONS:
- Mortality/moribundity checks: Twice daily, in the morning and at the end of the working day.
- Clinical observations: Once daily
- Body weights: Animals were weighed individually on day 1 (pre instillation) and on the day of the final observation.
- Irritation: The eyes of each animal were examined approximately 1, 24, 48 and 72 hours and 7, 14 and/or 21 days after instillation of the test item. The irritation scores and a description of all other (local) effects were recorded.

PAIN MANAGEMENT:
- One hour prior to instillation of the test item, buprenorphine 0.01 mg/kg was administered by subcutaneous injection in order to provide a therapeutic level of systemic analgesia.
- Five minutes prior to instillation of the test item, two drops of the topical anaesthetic 0.5% proparacaine hydrochloric ophthalmic solution were applied to both eyes.
- Immediately after fluorescein examination on Day 2, in order to provide a continued level of systemic analgesia, buprenorphine 0.01 mg/kg and meloxicam 0.5 mg/kg were administered by subcutaneous injection. Additional injections of buprenorphine 0.01 mg/kg and meloxicam 0.5 mg/kg were administered immediately after the 48-hour observation to reduce pain and distress.

Results and discussion

In vivo

Resultsopen allclose all
Irritation parameter:
cornea opacity score
Basis:
animal #1
Time point:
24/48/72 h
Score:
1
Max. score:
4
Reversibility:
fully reversible within: 7 days
Irritation parameter:
cornea opacity score
Basis:
animal #2
Time point:
24/48/72 h
Score:
0.7
Max. score:
4
Reversibility:
fully reversible within: 72 hours
Irritation parameter:
cornea opacity score
Basis:
animal #3
Time point:
24/48/72 h
Score:
1
Max. score:
4
Reversibility:
fully reversible within: 7 days
Irritation parameter:
iris score
Basis:
animal #1
Time point:
24/48/72 h
Score:
1
Max. score:
2
Reversibility:
fully reversible within: 7 days
Irritation parameter:
iris score
Basis:
animal #2
Time point:
24/48/72 h
Score:
0
Max. score:
2
Reversibility:
fully reversible within: 24 hours
Irritation parameter:
iris score
Basis:
animal #3
Time point:
24/48/72 h
Score:
0.3
Max. score:
2
Reversibility:
fully reversible within: 48 hours
Irritation parameter:
conjunctivae score
Basis:
animal #1
Time point:
24/48/72 h
Score:
2.7
Max. score:
3
Reversibility:
fully reversible within: 14 days
Irritation parameter:
conjunctivae score
Basis:
animal #2
Time point:
24/48/72 h
Score:
2.7
Max. score:
3
Reversibility:
fully reversible within: 14 days
Irritation parameter:
conjunctivae score
Basis:
animal #3
Time point:
24/48/72 h
Score:
2.7
Max. score:
3
Reversibility:
fully reversible within: 14 days
Irritation parameter:
chemosis score
Basis:
animal #1
Time point:
24/48/72 h
Score:
1.7
Max. score:
4
Reversibility:
fully reversible within: 7 days
Irritation parameter:
chemosis score
Basis:
animal #2
Time point:
24/48/72 h
Score:
1
Max. score:
4
Reversibility:
fully reversible within: 72 hours
Irritation parameter:
chemosis score
Basis:
animal #3
Time point:
24/48/72 h
Score:
2
Max. score:
4
Reversibility:
fully reversible within: 7 days
Irritant / corrosive response data:
Instillation of the test item caused corneal injury, iridial irritation and irritation to the conjunctivae.

The corneal injury consisted of opacity and epithelial damage. As a result of the corneal injury, pannus (neovascularization of the cornea) was apparent 7 and/or 14 days after instillation for two animals. The corneal injury resolved within 7 days in one animal and within 14 days in the other two animals.
Iridial irritation was observed and resolved within 24 hours, 48 hours or 7 days.
The irritation of the conjunctivae consisted of redness, chemosis and discharge and was completely resolved within 14 days in all animals.
Other effects:
- Coloration/remnants: Yellow staining of the fur on the head and paws caused by the test item was observed. 72 hours after instillation the staining had dissappeared in all animals.
- Toxicity/mortality: No signs of systemic toxicity were observed in the animals during the test period and no mortality occurred.

Applicant's summary and conclusion

Interpretation of results:
other: Irritating to eyes (Category 2)
Remarks:
according to EC No 1272/2008
Conclusions:
In an in vivo eye irritation test performed according to OECD test guideline 405 and GLP principles, Potassium Ferricyanide caused effects on the cornea, iris and conjunctivae and is classified as irritating to eyes (Category 2).
Executive summary:

An in vivo eye irritation study was performed in new zealand white rabbits with Potassium Ferricyanide according to OECD guideline 405 and GLP principles. A single application of 99.9mg - 110.2mg (approximately 0.1 mL) Potassium Ferricyanide was applied to the conjunctival sac of the eye under systemic and local anesthetic. Observations period were at 1, 24, 48 and 72 hours and 7, 14 and/or 21 days after instillation of the test item. Corneal epithelial damage was assessed with 2% fluorescein in water. Observations indicated corneal injury that consisted of opacity and epithelial damage. As a results of the corneal injury, pannus (neovascularization of the cornea) was apparent 7 and/or 14 days after instillation for two animals.  The corneal injury resolved within 7 days in one animal and within 14 days in the other two animals. Iridial irritation was observed and resolved within 24 hours, 48 hours or 7 days. Irritation of the conjunctivae was observed after instillation and consisted of redness, chemosis and discharge and was completely resolved within 14 days in all animals. No signs of systemic toxicity were observed and no mortality occurred. Based on these results, Potassium Ferricyanide is classified as: mildly irritating to eyes (Category 2B) according to the Globally Harmonized System of Classification and Labelling of Chemicals (GHS) of the United Nations (2017) (including all amendments) and classified as irritating to eyes (Category 2), labeled as H319: Causes serious eye irritation, according to the Regulation (EC) No 1272/2008 on classification, labelling and packaging of items and mixtures (including all amendments).