Methyl 2-benzoylbenzoate

Eye irritation

Administrative data

Endpoint:

eye irritation, other

Remarks:

An in vivo eye irritation study was conducted because of negative results in the in vitro eye irritation study.

Type of information:

experimental study

Adequacy of study:

key study

Study period:

September 2014

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2014

Report date:

2014

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Methyl 2-benzoylbenzoate

EC Number:

210-112-3

EC Name:

Methyl 2-benzoylbenzoate

Cas Number:

606-28-0

Molecular formula:

C15H12O3

IUPAC Name:

methyl 2-benzoylbenzoate

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Details on test material:

- Name of test material (as cited in study report): RCX 14-672
- Chemical name: methyl-2-benzoylbenzoate
- Substance type: organic
- Physical state: white to light yellowish powder
- Analytical purity: >99%
- Impurities (identity and concentrations): unknown
- Lot/batch No.: N14003
- Expiration date of the lot/batch: 30 November 2015
- Stability under test conditions: stable
- Storage condition of test material: controlled room temperature (15-25°C, below 70 RH%), protected from light and humidity

Specific details on test material used for the study:

Identification: RCX 14-672
Chemical name: methyl-2-benzoylbenzoate
Batch no.: N14003
CAS no.: 606-28-0
EC no.: 210-112-3
Molecular formula: C15H12O3
Molecular mass: 240.3 g/mol
Description: white to light yellowish powder
Purity: >99% (gas chromatography)
Water solubility: 117.7 mg/l
Test item storage: at room temperature, protected from light
Stability: stable under storage conditions
Expiry date: 30 November 2015

Test animals / tissue source

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

Species and strain: New Zealand White rabbits
Source: S&K-LAP Kft.
2173 Kartal, Császár út 135, Hungary
Number of animals: 3 animals
Age of animals at treatment: ~11 weeks old (adult)
Sex: Male
Body weight range
on the day of treatment: 2711 g and 2836 g
before euthanasia: 2825 g and 2900 g
Date of receipt: 27 August 2014
Acclimatization time: at least 6 days
Animal identification: The individual identification was by engraved ear tag. The cages were marked with individual identity cards with information about study code, sex, dose, cage number and individual animal number.

Light: 12 hours daily, from 6.00 a.m. to 6.00 p.m.
(and during of the analgesic/anaesthetic treatment)
Temperature
during the study: 20 ± 3°C
Relative humidity
during the study: 42 - 80 %
Housing/Enrichment: Rabbits were individually housed in AAALAC approved metal wire rabbit cages. Cages were of an open wire structure and cages were placed together to allow some social interaction with rabbit(s) in adjoining cages
Ventilation: 15-20 air exchanges/hour

The temperature and relative humidity values were measured continuously. The measured range was checked at least daily during the acclimatisation and experimental phases.

Animals received UNI diet for rabbits produced by Cargill Takarmány Zrt. (formerly Agribrands Europe Hungary), H-5300 Karcag, Madarasi út, Hungary, ad libitum.

The animals received municipal tap water, as for human consumption, ad libitum, from an automatic system. The quality control analysis is performed once every three months and microbiological assessment is performed monthly, by Veszprém County Institute of State Public Health and Medical Officer Service (ÁNTSZ, H-8201 Veszprém, József A.u.36., Hungary).

Test system

Vehicle:

unchanged (no vehicle)

Controls:

yes

Amount / concentration applied:

A single dose of 0.1 g of test item RCX 14-672 was administered to the animal. The test substance was placed in the conjunctival sac of the left eye of each animal after gently pulling the lower lid away from the eyeball. The lids were then gently held together for at least one second in order to prevent loss of the material. The untreated contralateral eye was served as the control.

Duration of treatment / exposure:

The treated eye was rinsed with physiological saline solution at the first observation time point in all animals at one hour after the application of test item as the irritation scores were greater than 1 and residual test item was noted on the eye.

Observation period (in vivo):

The eyes were examined at 1, 24, 48 and 72 hours after treatment.

Number of animals or in vitro replicates:

3

Details on study design:

Identification of pH:
The pH of the test item was determined. The pH was found to be 5.5, so the test item is permitted for use in animal studies.

Pre-study examination:
Three male animals in acceptable health condition were selected for the test. Care was taken to select only those animals that had a normal eye condition and any with ocular lesions were rejected.

Chronology of animal use:
Initially only one rabbit was treated with test item. As the effect in the first rabbit was non-irritant (all scores are zero) at 24 hours, then two further rabbits were treated with test item.

Analgesic and anaesthetic treatment:
Sixty minutes (60 ±10 min) prior to test substance application, a systemic opiate analgesic was administered subcutaneous injection (SC) under direct Veterinary supervision. Five minutes (5 ±1.5 min) prior to test substance application, a topical ocular anaesthetic was applied to each eye (including the control eye to ensure direct comparison of any ocular observations). Eight hours (8 to 9 hr) after test substance application, a systemic opiate analgesic and a nonsteroidal anti-inflammatory drug (NSAID) were administered subcutaneous injection (SC) under direct Veterinary supervision. The systemic opiate analgesic was injected ~12 hours, until the ocular lesions were resolved and no clinical signs of pain or distress were present.

Systemic opiate analgesic: Buprenorphine 0.01 mg/kg.
Topical ocular anaesthetic: Humacain (oxybuprocaine) one-two drops per eye.
Nonsteroidal anti-inflammatory drug: Meloxicam 0.5 mg/kg.

Clinical Observations and Evaluation of Ocular Irritation:
The eyes were examined at 1, 24, 48 and 72 hours after treatment. The duration of the observation period was sufficient to identify reversibility or irreversibility of changes. Any clinical signs of toxicity or signs of ill-health during the study were recorded. At the end of the observation period, the animal was sacrificed by intramuscular injections of CP-Ketamin 10% and CP-Xylazin 2% followed by i.v. Pentobarbital sodium (see details in 3.2.). Death was verified by checking pupil and corneal reflex and the absence of respiration. All rabbits were examined for distress at least twice daily, with observations at least 6 hours apart. Clinical observations or signs of ill-health were recorded.

Scoring and Assessment of Local Reaction:
The eye irritation scores were evaluated according to the scoring system by Draize (1977) and OECD 405 (02 October 2012)

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

conjunctivae score

Basis:

mean

Time point:

other: 1 hour

Score:

2

Max. score:

2

Reversibility:

fully reversible within: 24 hours

Remarks on result:

probability of mild irritation

Results 2

Irritation parameter:

chemosis score

Basis:

mean

Time point:

other: 1 hour

Score:

0.33

Max. score:

1

Reversibility:

fully reversible within: 24 hours

Remarks on result:

probability of weak irritation

Results 3

Irritation parameter:

cornea opacity score

Basis:

mean

Time point:

other: 1 hour

Score:

0

Max. score:

0

Remarks on result:

no indication of irritation

Results 4

Irritation parameter:

iris score

Basis:

mean

Time point:

other: 1 hour

Score:

0

Max. score:

0

Remarks on result:

no indication of irritation

Results 5

Irritation parameter:

conjunctivae score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

0

Reversibility:

fully reversible within: 24 hours

Remarks on result:

no indication of irritation

Results 6

Irritation parameter:

chemosis score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

0

Reversibility:

fully reversible within: 24 hours

Remarks on result:

no indication of irritation

Results 7

Irritation parameter:

cornea opacity score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

0

Remarks on result:

no indication of irritation

Results 8

Irritation parameter:

iris score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

0

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

Examination of eye-irritancy
The eyes were examined at 1, 24, 48 and 72 hours after the application. No Initial Pain Reaction (IPR/PR) was observed after application of the test item.
First animal (No: 01669) clinical observation:
At one hour after the application: Conjunctival redness (score 2), chemosis (score 1) and discharge (score 2) were found. Residual test item was noted on the eye. At 24, 48 and 72 hours after the application: There were no clinical signs, and no conjunctival or corneal effects observed. The animal was euthanized after the 72 hours observation.

Second animal (No: 01672) clinical observation:
At one hour after the application: Conjunctival redness (score 2) and discharge (score 1) were found. Residual test item was noted on the eye. At 24, 48 and 72 hours after the application: There were no clinical signs, and no conjunctival or corneal effects observed. The animal was euthanized after the 72 hours observation.

Third animal (No: 01671) clinical observation:
At one hour after the application: Conjunctival redness (score 2) and discharge (score 1) were found. Residual test item was noted on the eye. At 24, 48 and 72 hours after the application: There were no clinical signs, and no conjunctival or corneal effects observed. The animal was euthanized after the 72 hours observation. There were no clinical sign of systemic toxicity observed in the animals during the study and no mortality occurred.

During the study, the control eye of each animal was symptom-free. The body weights of the animal were considered to be within the normal range of variability.

Applicant's summary and conclusion

Interpretation of results:

not irritating

Conclusions:

The test item RCX 14-672, applied to rabbit eye mucosa, caused conjunctival effects in all animals at one hour after application. The effects were fully reversible within 24 hours.

Executive summary:

An acute eye irritation study with RCX 14-672 was performed in New Zealand White rabbits in accordance with OECD Guideline No. 405. The irritation effects of the test item were evaluated according to the Draize method. Three male New Zealand White Rabbits were used in this study. The test item was placed into the conjunctival sac of the left eye of each animal. The untreated right eye served as control. An amount of 0.1 g of the test item was administered as a single dose. The eyes were examined at 1, 24, 48 and 72 hours after the application. No Initial Pain Reaction (IPR/PR) was observed after application of the test item. At one hour after the application: conjunctival redness (score 2), chemosis (score 1) and discharge (score 2) was observed in animal nr. 1. In the other 2 animals conjunctival redness (score 2) and discharge (score 1) were observed at one hour after the application. Residual test item was noted on the eye in all animals. At 24, 48 and 72 hours after the application, no clinical signs, and no conjunctival or corneal effects were observed in any of the animals. All animals were euthanized after the 72 hours observation. There were no clinical sign of systemic toxicity observed in the animals during the study and no mortality occurred. During the study, the control eye of each animal was symptom-free. The body weights of the animal were considered to be within the normal range of

variability.

According to Regulation (EC) No 1272/2008, RCX 14-672 does not require classification as an eye irritant. According to the UN Globally Harmonised System of Classification and Labelling of Chemicals (Rev. 5) (2013), RCX 14-672 does not require classification as an eye irritant.