1-Piperazinecarboxylic acid, 4-(6-amino-3-pyridinyl)-, 1,1-dimethylethyl ester

Eye irritation

Administrative data

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 28 May to 13 June 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study run to a method comparable with current guidelines and to GLP

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

tert-butyl 4-(6-aminopyridin-3-yl)piperazine-1-carboxylate

EC Number:

695-268-4

Cas Number:

571188-59-5

Molecular formula:

C14H22N4O2

IUPAC Name:

tert-butyl 4-(6-aminopyridin-3-yl)piperazine-1-carboxylate

Details on test material:

Batch No.: E010013662
Purity: 98.6%

Test animals / tissue source

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Harlan Laboratories Ltd.
- Age at study initiation: twelve to twenty weeks old
- Weight at study initiation: 2.41 to 2.90 kg
- Housing: individually housed in suspended cages
- Diet (e.g. ad libitum): Free access to mains food (2930C Teklad Global Rabbit diet supplied by Harlan Laboratories UK Ltd., Oxon, UK) was allowed throughout the study.
- Water (e.g. ad libitum): Free access to mains drinking water
- Acclimation period: at least five days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 17 to 23 °C
- Humidity (%): 30 to 70%
- Air changes (per hr): at least fifteen changes per hour
- Photoperiod (hrs dark / hrs light): twelve hours continuous light (06:00 to 18:00) and twelve hours darkness.

IN-LIFE DATES: From To:

Test system

Vehicle:

unchanged (no vehicle)

Controls:

not required

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): A volume of 0.1 mL of the test item, which was found to weigh approximately 70 mg
- Concentration (if solution):

VEHICLE
- Amount(s) applied (volume or weight with unit):
- Concentration (if solution):
- Lot/batch no. (if required):
- Purity:

Duration of treatment / exposure:

8 hours

Observation period (in vivo):

72 hours

Number of animals or in vitro replicates:

3 males

Details on study design:

Immediately before the start of the test, both eyes of the provisionally selected test rabbits were examined for evidence of ocular irritation or defect with the aid of a light source from a standard ophthalmoscope. Only animals free of ocular damage were used.
Initially, a single rabbit was treated. A subcutaneous injection of buprenorphine 0.01 mg/kg was administered to each eye 60 minutes prior to test item application to provide a therapeutic level of systemic analgesia. Five minutes prior to test item application, a pre-dose anaesthesia of ocular anesthetic (two drops of 0.5% tetracaine hydrochloride) was applied to each eye.
A volume of 0.1 mL of the test item, which was found to weigh approximately 70 mg (as measured by gently compacting the required volume into an adapted syringe) was placed into the conjunctival sac of the right eye, formed by gently pulling the lower lid away from the eyeball. The upper and lower eyelids were held together for about one second immediately after treatment, to prevent loss of the test item, and then released. The left eye remained untreated and was used for control purposes. Immediately after administration of the test item, an assessment of the initial pain reaction was made according to the six point scale.
Eight hours after test item application, a subcutaneous injection of post-dose analgesia, buprenorphine 0.01 mg/kg and meloxicam 0.5 mg/kg, was administered to provide a continued therapeutic level of systemic analgesia. The treated animal was checked for signs of pain and suffering approximately 12 hours later. No further analgesia was required.
After consideration of the ocular responses produced in the first treated animal, two additional animals were similarly treated.
Assessment of ocular damage/irritation was made approximately 1 hour and 24, 48 and 72 hours following treatment, according to the numerical evaluation (Draize, J.H, 1977).
Any other ocular effects were also noted. Examination of the eye was facilitated by the use of the light source from a standard ophthalmoscope.
Any clinical signs of toxicity, if present, were also recorded.
Individual body weights were recorded on Day 0 (the day of dosing) and at the end of the observation period.

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

cornea opacity score

Basis:

mean

Time point:

other: mean of 24, 48 and 72 hours

Score:

0

Max. score:

0

Results 2

Irritation parameter:

iris score

Basis:

mean

Time point:

other: mean of 24, 48 and 72 hours

Score:

ca. 0.11

Max. score:

1

Reversibility:

fully reversible within: 48 hours

Remarks on result:

other: fully reversible within 48 hours in one animal and within 24 hours in other two animals

Results 3

Irritation parameter:

conjunctivae score

Basis:

mean

Time point:

other: mean of 24, 48 and 72 hours

Score:

ca. 0.89

Max. score:

2

Reversibility:

fully reversible within: 72 hours

Remarks on result:

other: fully reversible within 72 hours in all three animals

Results 4

Irritation parameter:

chemosis score

Basis:

mean

Time point:

other: mean of 24, 48 and 72 hours

Score:

ca. 0.67

Max. score:

2

Reversibility:

fully reversible within: 72 hours

Remarks on result:

other: fully reversible within 72 hours in all three animals

Irritant / corrosive response data:

No corneal effects were noted during the study.
Iridial inflammation was noted in all treated eyes one hour after treatment and in one treated eye at the 24-Hour observation.
Moderate conjunctival irritation was noted in all treated eyes one hour after treatment with minimal conjunctival irritation noted at the 24 and 48-Hour observations.
All treated eyes appeared normal at the 72-Hour observation.

Other effects:

All animals showed expected gain in body weight during the study.

Any other information on results incl. tables

Individual Scores and Individual Total Scores for Ocular Irritation

Rabbit Number and Sex	Time after application	Cornea opacity	Iris	Conjunctivae
				Redness	Chemosis
73206 Male	1 hours 24hours 48hours 72hours	0 0 0 0	1 1 0 0	2 1 1 0	2 1 1 0
73247 Male	1 hour 24 hours 48 hours 72 hours	0 0 0 0	1 0 0 0	2 2 1 0	2 1 1 0
73248 Male	1 hour 24 hours 48 hours 72 hours	0 0 0 0	1 0 0 0	2 2 1 0	2 1 1 0

Applicant's summary and conclusion

Interpretation of results:

slightly irritating

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The test item produced a maximum group mean score of 15.0 and was classified as a mild irritant (Class 4 on a 1 to 8 scale) to the rabbit eye according to a modified Kay and Calandra classification system.
The test item was not classified as an irritant according to GHS and EC CLP 1272/2008.