[No public or meaningful name is available]

Eye irritation

Administrative data

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

September 20, 2004 - October 19, 2004

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2004

Report date:

2004

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Version / remarks:

(2002)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.5 (Acute Toxicity: Eye Irritation / Corrosion)

Version / remarks:

(1992)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2400 (Acute Eye Irritation)

Version / remarks:

(1998)

Deviations:

no

Test guideline 4

Qualifier:

according to guideline

Guideline:

other: Japanese Ministry of Agriculture, Forestry and Fisheries (JMAFF), 12 Nousan, Notification No 8147, November 2000, including the most recent partial revisions.

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

453-140-3

EC Name:

-

Cas Number:

667465-46-5

Molecular formula:

C20H38N6O13 (not dissociated) C20H44N6O13 (dissociated in water)

IUPAC Name:

3,6,9-tris(2-{[(2-hydroxyethyl)amino]oxy}-2-oxoethyl)-3,6,9-triazaundecan-1,11-dioic acid

Constituent 2

Reference substance name:

Water

EC Number:

231-791-2

EC Name:

Water

Cas Number:

7732-18-5

Molecular formula:

H2O

IUPAC Name:

Water

Test material form:

liquid

Details on test material:

- Name of test material: DHX2
- CAS no.: 667465-46-5
- Storage condition of test material: at room temperature in the dark

Test animals / tissue source

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

- Source: Charles River Deutschland, Kisslegg, Germany
- Age at study initiation: Animals used withihn the study were at least 6 weeks old.
- Weight at study initiation: Body weights were at least 1.0 kg.
- Housing: Individually housed in cages with perforated floors.
- Diet: Free access to pelleted diet for rabbits (Charles River Breeding and Maintenance Diet for Rabbits, Altromin, Lage, Germany). Hay was provided at least three times a week.
- Water: Free access to tap water.
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19.7 – 22.4
- Humidity (%): 33 - 71
- Air changes (per hr): approx. 15
- Photoperiod (hrs dark / hrs light): 12/12

Test system

Vehicle:

unchanged (no vehicle)

Controls:

other: One eye of each animal remained untreated and served as the reference control.

Amount / concentration applied:

TEST MATERIAL- Amount(s) applied: 0.1 mL

Duration of treatment / exposure:

Single instillation on Day 1.

Observation period (in vivo):

7 days.

Number of animals or in vitro replicates:

3 males

Details on study design:

STUDY DESIGN
Since no severe harm for the animals was to be expected, this in-vivo eye irritation study was performed and was started by treatment of a single rabbit (sentinel). The two other animals were treated in a similar manner 3 weeks later, after considering the degree of eye irritation observed in the first animal.

TREATMENT
Each animal was treated by instillation of the test substance, in the conjunctival sac of one of the eyes after gently pulling the lower lid away from the eyeball. The lids were then gently held together for about one second to prevent loss of the test substance.
Immediately after the 24-hour observation, a solution of 2% fluorescein in water was instilled into both eyes of each animal to quantitatively determine corneal epithelial damage. Any bright green stained area, indicating epithelial damage, was estimated as a percentage of the total corneal area.

REMOVAL OF TEST SUBSTANCE
-Washing (if done): No

OBSERVATIONS
- Mortality/Viability: Twice daily.
- Toxicity: At least once daily.
- Body Weight: Day of treatment (prior to instillation) and at termination.
- Necropsy: No data available.
- Irritation:
The eyes of each animal were examined approximately 1, 24, 48 and 72 hours and 7 days after instillation of the test substance.
The irritation scores and a description of all other (local) effects were recorded. The irritation was assessed according to OECD 405.

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

cornea opacity score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

4

Remarks on result:

no indication of irritation

Results 2

Irritation parameter:

cornea opacity score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

4

Remarks on result:

no indication of irritation

Results 3

Irritation parameter:

cornea opacity score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

4

Remarks on result:

no indication of irritation

Results 4

Irritation parameter:

iris score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

2

Remarks on result:

no indication of irritation

Results 5

Irritation parameter:

iris score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

2

Remarks on result:

no indication of irritation

Results 6

Irritation parameter:

iris score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

2

Remarks on result:

no indication of irritation

Results 7

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0.3

Max. score:

4

Reversibility:

fully reversible within: 48 hours

Results 8

Irritation parameter:

conjunctivae score

Remarks:

(redness)

Basis:

animal #1

Time point:

24/48/72 h

Score:

2.3

Max. score:

3

Reversibility:

fully reversible within: 7 days

Results 9

Irritation parameter:

conjunctivae score

Remarks:

(redness)

Basis:

animal #2

Time point:

24/48/72 h

Score:

1.3

Max. score:

3

Reversibility:

fully reversible within: 7 days

Results 10

Irritation parameter:

conjunctivae score

Remarks:

(redness)

Basis:

animal #3

Time point:

24/48/72 h

Score:

1.7

Max. score:

3

Reversibility:

fully reversible within: 7 days

Results 11

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0.3

Max. score:

4

Reversibility:

fully reversible within: 48 hours

Results 12

Irritation parameter:

chemosis score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0.7

Max. score:

4

Reversibility:

fully reversible within: 48 hours

Irritant / corrosive response data:

Instillation of the substance into one eye of each of three animals resulted in irritation of the conjunctivae, which consisted of redness, chemosis and discharge. The irritation had completely resolved within 7 days in all animals.
No iridial irritation or corneal opacity was observed, and treatment of the eyes with 2% fluorescein, 24 hours after test substance instillation revealed no corneal epithelial damage.
There was no evidence of ocular corrosion.

Other effects:

No staining of (peri) ocular tissues by the test substance was observed and no test substance remnants were seen.
No symptoms 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: not irritating according to EC regulation 1272/2008 and its amendments.

Conclusions:

In an eye irritation study with rabbits, performed according to OECD 405 test guidelines, limited irritation was observed. The irritation had completely resolved within 7 days in all animals.

Executive summary:

Three male rabbits were exposed to DHX2 for 24 hours, according to OECD 405 test guideline and GLP principles.

Instillation of the substance into one eye of each of three animals resulted in irritation of the conjunctivae, which consisted of redness, chemosis and discharge. The irritation had completely resolved within 7 days in all animals. No iridial irritation or coneal opacity was observed. No symptoms of systemic toxicity were observed in the animals during the test period and no mortality occurred.

It can be concluded that DHX2 is slightly irritating to eyes but no classification is needed according to EC regulation 1272/2008.