[1,1'-Biphenyl]-4-pentanoic acid, .gamma.-[(3-carboxy-1-oxopropyl)amino]-.alpha.-methyl-, 4-ethyl ester, calcium salt (2:1), (.alpha.R,.gamma.S)-

Eye irritation

Administrative data

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Nov 2007 - March 2008

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

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:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2400 (Acute Eye Irritation)

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

calcium bis(3-{[(2S,4R)-1-{[1,1'-biphenyl]-4-yl}-5-ethoxy-4-methyl-5-oxopentan-2-yl]carbamoyl}propanoate)

EC Number:

935-847-3

Cas Number:

1369773-39-6

Molecular formula:

C48H56CaN2O10

IUPAC Name:

calcium bis(3-{[(2S,4R)-1-{[1,1'-biphenyl]-4-yl}-5-ethoxy-4-methyl-5-oxopentan-2-yl]carbamoyl}propanoate)

Test material form:

solid: particulate/powder

Test animals / tissue source

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

Albino rabbit, New Zealand White (SPF-quality)
Location:
Specific Pathogenic Free area: animal room no. 7.
Room temperature:
20.0 to 21.4° C.
Room relative humidity:
29 – 60%.
Lighting cycle:
Fluorescent light for a 12-hour light/12-hour dark cycle.
Animal caging:
Individually housed in labelled cages with perforated floors (Scanbur, Denmark, dimensions 56x44x37.5 cm).
Acclimatization:
Acclimatization period was at least 5 days before start of treatment under laboratory conditions.
Food:
Free access to pelleted diet for rabbits (K-H from SSNIFF® Spezialdiäten GmbH, Soest, Germany). In addition, hay (TecniLab-BMI BV, Someren, The Netherlands) was provided three times a week.
Analysis of food:
Results of analysis for each batch of diet (nutrients and contaminants) and hay were assessed and did not reveal any findings that were considered to have affected the study integrity.
Water:
Free access to tap-water.
Analysis of water:
Certificates of analysis were examined and archived. Analysis of water did not reveal any findings that were considered to have affected study integrity.

Test system

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent no treatment

Amount / concentration applied:

Animals were treated by instillation of the test substance at a volume of approximately 0.1 ml (on average, 30.5 mg (range 30.1 – 30.9 mg) in the conjunctival sac of one of the eyes after gently pulling the lower lid away from the eyeball.

Duration of treatment / exposure:

Single samples of AHU377 at a volume of approximately 0.1 ml were instilled into one eye of each of three rabbits.

Observation period (in vivo):

Observations were made 1, 24, 48 and 72 hours and 7 and/or 14 Days after instillation.

Number of animals or in vitro replicates:

three rabbits

Details on study design:

Animals were treated by instillation of the test substance at a volume of approximately 0.1 ml (on average, 30.5 mg (range 30.1 – 30.9 mg) 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. The other eye remained untreated and served as the reference control.
Immediately after the 24-hour observation, a solution of 2% fluorescein (Merck, Darmstadt, Germany) in water (adjusted to pH 7.0) was instilled into both eyes of each animal to quantitatively determine corneal epithelial damage. This procedure was repeated to assess recovery. Any bright green stained area, indicating epithelial damage, was estimated as a percentage of the total corneal area.
Immediately after fluorescein examination on Day 2, the treated eye of animal 681 was rinsed with approx 50 mL tepid tap water, using a velocity of flow, which did not affect the eye, to remove residual test substance. For reference control the other eye was also rinsed.
After the final observation, the animals were sacrificed by intra-venous injection of pentobarbital Euthesate®

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

cornea opacity score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0.7

Max. score:

4

Reversibility:

fully reversible within: 7 d

Remarks on result:

probability of weak irritation

Results 2

Irritation parameter:

cornea opacity score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0.7

Max. score:

4

Reversibility:

fully reversible within: 14d

Remarks on result:

probability of weak irritation

Results 3

Irritation parameter:

cornea opacity score

Basis:

animal #3

Time point:

24/48/72 h

Score:

1

Max. score:

4

Reversibility:

fully reversible within: 14d

Remarks on result:

probability of weak irritation

Results 4

Irritation parameter:

iris score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0.3

Max. score:

4

Reversibility:

fully reversible within: 48 h

Remarks on result:

probability of weak irritation

Results 5

Irritation parameter:

iris score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0.3

Max. score:

4

Reversibility:

fully reversible within: 48 h

Remarks on result:

probability of weak irritation

Results 6

Irritation parameter:

iris score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0.7

Max. score:

4

Reversibility:

fully reversible within: 72

Remarks on result:

probability of weak irritation

Results 7

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

24/48/72 h

Score:

2.3

Max. score:

4

Reversibility:

fully reversible within: 7d

Remarks on result:

probability of mild irritation

Results 8

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

24/48/72 h

Score:

1.3

Max. score:

4

Reversibility:

fully reversible within: 14d

Remarks on result:

probability of weak irritation

Results 9

Irritation parameter:

chemosis score

Basis:

animal #3

Time point:

24/48/72 h

Score:

1.3

Max. score:

4

Reversibility:

fully reversible within: 14d

Remarks on result:

probability of weak irritation

Results 10

Irritation parameter:

conjunctivae score

Basis:

animal #1

Time point:

24/48/72 h

Score:

3

Max. score:

4

Reversibility:

fully reversible

Remarks on result:

probability of moderate irritation

Results 11

Irritation parameter:

conjunctivae score

Basis:

animal #2

Time point:

24/48/72 h

Score:

3

Max. score:

4

Reversibility:

fully reversible

Remarks on result:

probability of mild irritation

Results 12

Irritation parameter:

conjunctivae score

Basis:

animal #3

Time point:

24/48/72 h

Score:

2.7

Max. score:

4

Reversibility:

fully reversible

Remarks on result:

probability of mild irritation

Irritant / corrosive response data:

There was no evidence of ocular corrosion.
No symptoms of systemic toxicity were observed in the animals during the test period and no mortality occurred.

Other effects:

Remnants of the test substance were present in the eye on Days 1 and/or 2.

Applicant's summary and conclusion

Interpretation of results:

Category 2 (irritating to eyes) based on GHS criteria

Conclusions:

Based on these results:
- according to the Globally Harmonized System of Classification and Labelling of Chemicals (GHS) of the United Nations (2004), AHU377 should be classified as : irritating to eyes (Category 2).