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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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
data is from experimental reports

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2014
Report Date:
2014

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 405 (Acute Eye Irritation / Corrosion)
Principles of method if other than guideline:
Eye irritation study was performed for test chemical.
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid
Details on test material:
- Name of test material: Reaction mass of 4-(2,6,6-trimethylcyclohex-2-ene-1-yl)-but-3-ene-2-one and 4-(2,6,6-trimethylcyclohex-1-ene-1-yl)-but-3-ene-2-one
- IUPAC name: Reaction mass of 4-(2,6,6-trimethylcyclohex-2-ene-1-yl)-but-3-ene-2-one and 4-(2,6,6-trimethylcyclohex-1-ene-1-yl)-but-3-ene-2-one
- Molecular formula: C13H20O
- Molecular weight: 192.3 g/mole
- Smiles : C1([C@@H](C(=CCC1)C)\C=C\C(C)=O)(C)C
- Inchl: 1S/C13H20O/c1-10-6-5-9-13(3,4)12(10)8-7-11(2)14/h6-8,12H,5,9H2,1-4H3/b8-7+
- Substance type: Organic
- Physical state: Solid (yellow)

Test animals / tissue source

Species:
rabbit
Strain:
New Zealand White
Details on test animals or tissues and environmental conditions:
TEST ANIMALS
- Sex: Male
- Source: Procured from RABBI ROOF Animal Facility, A. P., India. (CPCSEA Reg. No. 1655/PO/b/12/CPCSEA).
- Age at study initiation:4.5 to 5.5 Months (Approximately)
- Health Status: Healthy young adult
- Weight at study initiation: Minimum: 1.696 kg and Maximum: 1.942 kg (Prior to Treatment)
- Identification: During acclimatization marking was done with non toxic marker pen in the inside of left ear of rabbits and after acclimatization, animals were marked with permanent number in the inner side of right ear of rabbits. Permanent marker and cage card was used for identification.
- Housing: The animals were housed individually in stainless steel cages.
- Diet (e.g. ad libitum): All animals were provided conventional laboratory rabbit diet (Nutrivet Life Sciences, Pune) ad libitum. Batch No.: 200012
- Water (e.g. ad libitum): Aqua guard filtered tap water was provided ad libitum
- Acclimation period: Rabbits were acclimatised to the test conditions for a period of 6 days (Animal No.-1) and 10 days (Animal No. 2 and 3) prior to the application of the test item
- Room -Sanitation: The experimental room floor and work tops were swept and mopped with disinfectant solution every day.
- Cages and water bottle: All the cages and water bottles were changed minimum twice a week.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): Minimum: 19.50 °C and Maximum: 22.20 °C
- Humidity (%): Minimum: 41.20 % and Maximum: 59.20 %
- Air changes (per hr): More than 12 changes per hour
- Photoperiod (hrs dark / hrs light): 12:12

Test system

Vehicle:
unchanged (no vehicle)
Controls:
yes, concurrent no treatment
Amount / concentration applied:
0.1 ml
Duration of treatment / exposure:
24 hours
Observation period (in vivo):
Animals were observed at 24 hours intervals for 72 hours.
Duration of post- treatment incubation (in vitro):
no data available
Number of animals or in vitro replicates:
3 males
Details on study design:
REMOVAL OF TEST SUBSTANCE
- Washing (if done): yes
- Time after start of exposure: After 24 hours observation period, the treated eye of rabbit was washed with normal saline

SCORING SYSTEM: Draize method

TOOL USED TO ASSESS SCORE: hand-slit lamp / biomicroscope / fluorescein:The area of corneal epithelial cell damage was scored using ophthalmoscope (Ri-Mini2, 5 V-guru; Model No. 3002).

Results and discussion

In vitro

Other effects / acceptance of results:
no data available

In vivo

Resultsopen allclose all
Irritation parameter:
cornea opacity score
Basis:
animal: 1,2,3
Time point:
24/48/72 h
Score:
0
Reversibility:
not specified
Remarks on result:
no indication of irritation
Irritation parameter:
iris score
Basis:
animal: 1,2,3
Time point:
24/48/72 h
Score:
0
Reversibility:
not specified
Remarks on result:
no indication of irritation
Irritation parameter:
conjunctivae score
Basis:
animal: 1,2,3
Time point:
24/48/72 h
Score:
1
Reversibility:
not specified
Remarks on result:
no indication of irritation
Irritation parameter:
chemosis score
Basis:
animal: 1,2,3
Time point:
24/48/72 h
Score:
0.33
Reversibility:
not specified
Remarks on result:
no indication of irritation
Irritant / corrosive response data:
The following were observed in treated rabbits.
Observation at 1 hour after instillation of test item revealed: Cornea- No ulceration or opacity in all 3 animals; Area of Opacity- Zero in all the animals; Iris: Normal in all the animals. Conjunctivae - Some blood vessels definitely hyperaemic (injected) in animal no. 1 and 2 whereas animal no.3 showed no ocular lesion; Chemosis: No swelling (Normal) were observed in all the animals.
Observation at 24 hours after instillation of test item revealed: Cornea- No ulceration or opacity in all 3 animals; Area of Opacity- Zero in all the animals; Iris: Normal in all the animals. Conjunctivae - Diffuse, crimson color; individual vessels not easily discernible was observed in all the treated animals; Chemosis: Some swelling above normal (includes nictating membranes) was observed in animal number 1 and no swelling was observed in animal number 2 and 3.
At 24 hours observation the rabbits were examined for corneal epithelium cell damage using sodium fluorescein strips and noticed 40 % and 15 % damage in animal no 1 and 2 whereas no damage was seen in animal no. 3.
Observation at 48 and 72 hours after instillation of test item revealed: Cornea- No ulceration or opacity in all the animals; Area of Opacity- Zero in all the animals; Iris: Normal in all the animals. Conjunctivae - blood vessels were normal in all the animals; Chemosis: No swelling was observed in all the animals.
The individual mean score for Animal Nos. 1, 2 and 3 at 24, 48, 72 hours for Corneal opacity, iris, conjunctiva, chemosis were found 0.00, 0.00, 1.00; 0.33; 0.00, 0.00; 1.00, 0.00 and 0.00, 0.00, 1.00, 0.00, respectively.
Other effects:
Clinical Observation
No systemic toxicity was observed at treated rabbits during the experimental period.
Mortality
No mortality was observed during the observation period.
Body weight
All rabbits were weighed on test day 0 (prior to application) and at termination. The body weights were marginally increased in animal number 1, 2 and 3 when compared with pre-treatment body weights.

Any other information on results incl. tables

Table 1 : Individual Animal Eye Irritation Score

Eye Irritation Scores - Mean Values after 24, 48, 72 Hours (Treated eye)

            Animal No.

 Eye Reaction

1

2

3

Corneal Opacity

0.00

0.00

0.00

Iris

0.00

0.00

0.00

Conjunctiva

1.00

1.00

1.00

Chemosis

0.33

0.00

0.00

 

 

Formula :

 

Mean Eye Irritation Score =

                          Sum of the Individual Animal Score for eye reactionat24, 48 and 72 hours

Number of the Observations (3)

Table 2 : Individual AnimalClinicalSigns

 

Sex:Male

Animal No.

Days (Post application observation)

0

1

2

3

1

1

1

1

1

2

1

1

1

1

3

1

1

1

1

Key:1 = Normal

 

Table 3: Individual Animal Body Weight

Sex :Male

Animal No.

Animal Body Weight (kg)

Prior to application

At termination

1

1.696

1.812

2

1.942

1.982

3

1.800

1.826

Key:kg = Kilogram

Applicant's summary and conclusion

Interpretation of results:
other: not irritating
Conclusions:
The individual mean score for Animal Nos. 1, 2 and 3 at 24, 48, 72 hours for Corneal opacity, iris, conjunctiva, chemosis were found  0.00, 0.00, 1.00; 0.33; 0.00, 0.00; 1.00, 0.00 and 0.00, 0.00, 1.00, 0.00,  respectively.
Hence, under the experimental conditions, the test chemical is “Non Irritant” to New Zealand White male rabbit eyes
Executive summary:

Acute Eye Irritation/Corrosion Study of Test chemical in Rabbits, was performed in an OECD GLP laboratory (Sustainability Support Services (Europe) has letter of access).The study was performed according to OECD 405 Guidelines. Rabbits free from injury of eye were selected for the study. The eyes of all the rabbits were examined 24 hours prior to treatment. One eye of each rabbit served as control and other as treated. Control eye was left untreated whereas;0.1 ml of test item (as such) was instilled in the other (treated) eye of rabbits. The eye was observed at 1, 24, 48, 72 hours after test item instillation. Ophthalmoscope was used for scoring of eye lesions. In the initial test,0.1 ml of test item was applied into the conjunctival sac of the right eye of Animal No.1. The left eye of the rabbit served as the control. Animal No. 1 presented ocular lesions at 1 hour observation period. Hence the confirmatory test was conducted on additional two rabbits (Animal No. 2 and 3);0.1 ml of test item was instilled into the conjunctival sac of right eye and left eye served as the control. Ocular lesions were observed at 1, 24 and 48 hour in Animal Number 2 whereas in Animal Number 3 ocular lesions presented only at 1 hour observation period. Untreated eye of the treated rabbits was normal throughout the experimental period of 72 hours. The following grading scores were observed in treated eye of tested rabbits. Observation at 1 hour after instillation of test item revealed: Cornea-No ulceration or opacity in all 3 animals; Area of Opacity-Zero in all the animals;Iris:Normal in all the animals. Conjunctivae -Some blood vessels definitely hyperaemic (injected) in animal no. 1 and 2 whereas animal no.3 showed no ocular lesion;Chemosis:No swelling (Normal) were observed in all the animals. Observation at 24 hours after instillation of test item revealed: Cornea-No ulceration or opacity in all 3 animals; Area of Opacity-Zero in all the animals;Iris:Normal in all the animals.Conjunctivae -Diffuse, crimson color; individual vessels not easily discernible was observed in all the treated animals;Chemosis:Some swelling above normal (includes nictating membranes) was observed in animal number 1 and no swelling was observed in animal number 2 and 3. At 24 hours observation the rabbits were examined for corneal epithelium cell damage using sodium fluorescein strips and noticed 40 % and 15 % damage in animal no 1 and 2 whereas no damage was seen in animal no. 3. Observation at 48 and 72 hours after instillation of test item revealed: Cornea-No ulceration or opacity in all the animals; Area of Opacity-Zero in all the animals;Iris:Normal in all the animals. Conjunctivae- blood vessels were normal in all animals; Chemosis: No swelling was observed in all the animals. Under the experimental conditions tested, eye irritation and reversibility of effects on eyes of rabbits was observed at 72 hours. The individual mean score for Animal Nos. 1, 2 and 3 at 24, 48, 72 hours for Corneal opacity, iris, conjunctiva, chemosis were found  0.00, 0.00, 1.00; 0.33; 0.00, 0.00; 1.00, 0.00 and 0.00, 0.00, 1.00, 0.00,  respectively. Hence, under the experimental conditions, Test chemical is “Non Irritant” to New Zealand White male rabbit eyes.