Registration Dossier

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 study report

Data source

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

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 405 (Acute Eye Irritation / Corrosion)
Principles of method if other than guideline:
The objective of the study was to assess the irritant and/or corrosive effects of test chemical on eye, when exposed by the ocular route in rabbits.
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report):Solvent violet 8 - 4, 4’-bis (dimethyl amino)-4”-(methyl amino) trityl alcohol
- Molecular formula : C24H29N3O
- Molecular weight :375.51 g/mol
- Substance type:Organic
- Physical state:Indigo solid
- Lot/batch No.:Lot 1/10
- AI Content:99.65 %
- Storage condition of test material:Kept container tightly closed in a cool, dry and well- ventilated place.
- Other:
Handling and Disposal
Safety precautions : Avoided inhalation and spilling. Smoking, eating and drinking should be prohibited in appropriate area. Aprons, masks, caps, gloves and goggles were used to ensure the health and safety of the personnel.
Disposal : The remaining unused test item was disposed as per internal SOPs of sa-FORD.

Test animals / tissue source

Species:
rabbit
Strain:
New Zealand White
Details on test animals or tissues and environmental conditions:
TEST ANIMALS
- Source:Procured from RABBI ROOF Animal Facility, A. P., India.
- Age at study initiation:3.5 to 4.5 Months (Approximately)
- Weight at study initiation:Minimum: 1.576 kg and Maximum: 2.746 kg (Prior to Treatment)
- Health Status :Healthy young adult.
- Housing:The animals were housed individually in stainless steel cages.
- 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.
- Diet (e.g. ad libitum):All animals were provided conventional laboratory rabbit diet (Nutrivet Life Sciences, Pune) ad libitum.
- 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 5 days (Animal No.-1) and 9 days (Animal No. 2 and 3) prior to the application of the test item.

ENVIRONMENTAL CONDITIONS
- Temperature (°C):Minimum: 19.20 °C and Maximum: 22.70 °C
- Humidity (%):Minimum: 42.00 % and Maximum: 74.70 %.
- 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
Amount / concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit):0.1 gm
- Concentration (if solution):N/A

VEHICLE
- Amount(s) applied (volume or weight with unit):N/A
- Concentration (if solution):N/A
- Lot/batch no. (if required):N/A
- Purity:N/A
Duration of treatment / exposure:
24 hrs
Observation period (in vivo):
All the animals were observed at 1, 24, 48, 72 hours, day 7, 14 and 21 after instillation of test item.
Number of animals or in vitro replicates:
3 male rabbits
Details on study design:
REMOVAL OF TEST SUBSTANCE
- Washing (if done):The treated eye of rabbit was washed with normal saline.
- Time after start of exposure:24 hours

SCORING SYSTEM:Draize Method

TOOL USED TO ASSESS SCORE: Ophthalmoscope and fluorescein strips.

Results and discussion

In vivo

Resultsopen allclose all
Irritation parameter:
cornea opacity score
Basis:
animal: #1,#2 and #3
Time point:
other: 24,48,72 hrs,7days,14 days and 21 days
Score:
0
Max. score:
4
Reversibility:
not specified
Remarks on result:
other: Untreated (Control Eye)
Irritation parameter:
iris score
Basis:
animal: #1,#2 and #3
Time point:
other: 24,48,72 hrs,7days,14 days and 21 days
Score:
0
Max. score:
2
Reversibility:
not specified
Remarks on result:
other: Untreated (Control Eye)
Irritation parameter:
conjunctivae score
Basis:
animal: #1,#2 and #3
Time point:
other: 24,48,72 hrs,7days,14 days and 21 days
Score:
0
Max. score:
3
Reversibility:
not specified
Remarks on result:
other: Untreated (Control Eye)
Irritation parameter:
chemosis score
Basis:
animal: #1,#2 and #3
Time point:
other: 24,48,72 hrs,7days,14 days and 21 days
Score:
0
Max. score:
4
Reversibility:
not specified
Remarks on result:
other: Untreated (Control Eye)
Irritation parameter:
cornea opacity score
Basis:
animal: #1,#2 and #3
Time point:
other: After 24, 48, 72 Hours,Day 7 and Day 14
Score:
1
Max. score:
4
Reversibility:
not specified
Remarks on result:
other: For Treated eye
Irritation parameter:
cornea opacity score
Basis:
animal: #1,#2 and #3
Time point:
other: Day 21
Score:
0
Max. score:
4
Reversibility:
not specified
Remarks on result:
other: For Treated eye
Irritation parameter:
iris score
Basis:
animal: #1,#2 and #3
Time point:
other: After 24, 48, 72 Hours,day 7 ,day 14 and day 21
Score:
0
Max. score:
2
Reversibility:
not specified
Remarks on result:
other: For Treated eye
Irritation parameter:
conjunctivae score
Basis:
animal #1
Time point:
other: After 24 Hours
Score:
1
Max. score:
3
Reversibility:
not specified
Remarks on result:
other: For Treated eye
Irritation parameter:
conjunctivae score
Basis:
animal #2
Time point:
other: After 48 Hours
Score:
1.67
Max. score:
3
Reversibility:
not specified
Remarks on result:
other: For Treated eye
Irritation parameter:
conjunctivae score
Basis:
animal #3
Time point:
other: After 72 Hours
Score:
1.33
Max. score:
3
Reversibility:
not specified
Remarks on result:
other: For Treated eye
Irritation parameter:
conjunctivae score
Basis:
animal: #1,#2
Time point:
other: Day 7 and Day 14
Score:
2
Max. score:
3
Reversibility:
not specified
Remarks on result:
other: For Treated eye
Irritation parameter:
conjunctivae score
Basis:
animal #3
Time point:
other: Day 7 and Day 14
Score:
1
Max. score:
3
Reversibility:
not specified
Remarks on result:
other: For Treated eye
Irritation parameter:
conjunctivae score
Basis:
animal: #1,#2 and #3
Time point:
other: Day 21
Score:
0
Max. score:
3
Reversibility:
not specified
Remarks on result:
other: For Treated eye
Irritation parameter:
chemosis score
Basis:
animal #1
Time point:
other: After 24 Hours
Score:
1
Max. score:
4
Reversibility:
not specified
Remarks on result:
other: For Treated eye
Irritation parameter:
chemosis score
Basis:
animal: #2 and #3
Time point:
other: After 48 Hours and 72 Hours
Score:
1.33
Max. score:
4
Reversibility:
not specified
Remarks on result:
other: For Treated eye
Irritation parameter:
chemosis score
Basis:
animal #1
Time point:
other: Day 7
Score:
2
Max. score:
4
Reversibility:
not specified
Remarks on result:
other: For Treated eye
Irritation parameter:
chemosis score
Basis:
animal: #2,#3
Time point:
other: Day 7
Score:
1
Max. score:
4
Reversibility:
not specified
Remarks on result:
other: For Treated eye
Irritation parameter:
chemosis score
Basis:
animal: #1 and #2
Time point:
other: Day 14
Score:
1
Max. score:
4
Reversibility:
not specified
Remarks on result:
other: For Treated eye
Irritation parameter:
chemosis score
Basis:
animal #3
Time point:
other: Day 14
Score:
0
Max. score:
4
Reversibility:
not specified
Remarks on result:
other: For Treated eye
Irritation parameter:
chemosis score
Basis:
animal: #1,#2 and #3
Time point:
other: Day 21
Score:
0
Max. score:
4
Reversibility:
not specified
Remarks on result:
other: For treated eye
Irritant / corrosive response data:
The following were observed in treated rabbits.
In the initial test, 0.1 gm of test item (Pulverized Form) was applied into the conjunctival sac of the right eye of animal no.1 whereas the left eye of the rabbit served as the control. As animal no. 1 showed ocular lesions hence a confirmatory test was conducted on additional two rabbits (animal no. 2 and 3); 0.1 gm of test item was instilled into the conjunctival sac of right eye of both the rabbits and left eye served as the control. Ocular lesions were seen in animal no. 1, 2 and 3 at till day 14observation which recovered at day 21.

Untreated eye of all the three rabbits was normal throughout the experimental period of 21 days.

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 3 animals; Iris: Normal in all 3 animals. Conjunctivae - Some blood vessels definitely hyperaemic (injected) was seen in all 3 animals; Chemosis: some swelling above normal (includes nictitating membranes) was seen in animal numbers 2 and 3 whereas animal number 1 was normal.

Observation at 24 hours after instillation of test item revealed: Cornea- Scattered or diffuse areas of opacity (other than slight dulling of normal lustre); details of iris clearly visible in all 3 animals; Area of Opacity- One quarter (or less) but not zero; Iris: Normal in all 3 animals; Conjunctivae - Some blood vessels definitely hyperaemic (injected) in all the animals; Chemosis: Some swelling above normal (includes nictitating membranes) was seen in all the animals.

At 24 hours observation the rabbits were examined for corneal epithelium cell damage using sodium fluorescein strips and noticed 30 %, 30 % and 20 % damage in animal no. 1, 2 and 3, resectively.

Observation at 48 hours after instillation of test item revealed: Cornea- Scattered or diffuse areas of opacity (other than slight dulling of normal lustre); details of iris clearly visible in all 3 animals; Area of Opacity- One quarter (or less) but not zero; Iris: Normal in all 3 animals; Conjunctivae - Some blood vessels definitely hyperaemic (injected) was seen in animal number 1 whereas diffuse, crimson color; individual vessels not easily discernible in animal number 2 and 3; Chemosis: Obvious swelling with partial eversion of lids was seen in animal numbers. 2 and 3 whereas some swelling above normal (includes nictitating membranes) was seen in animal no.1.

Observation at 72 hours after instillation of test item revealed: Cornea- Scattered or diffuse areas of opacity (other than slight dulling of normal lustre); details of iris clearly visible in all 3 animals; Area of Opacity- One quarter (or less) but not zero; Iris: Normal in all 3 animals; Conjunctivae - Some blood vessels definitely hyperaemic (injected) was seen in animal numbers 1 and 3 whereas diffuse, crimson color; individual vessels not easily discernible in animal number 2; Chemosis: some swelling above normal (includes nictitating membranes) was noticed in all the animals.

Observation at day 7 after instillation of test item revealed: Cornea- Scattered or diffuse areas of opacity (other than slight dulling of normal lustre); details of iris clearly visible in all 3 animals; Area of Opacity- One quarter (or less) but not zero; Iris: Normal in all 3 animals; Conjunctivae - Some blood vessels definitely hyperaemic (injected) was seen in animal number 3 whereas diffuse, crimson color; individual vessels not easily discernible in animal numbers 1 and 2; Chemosis: some swelling above normal (includes nictitating membranes) was noticed in animal numbers 2 and 3 whereas obvious swelling with partial eversion of lids was seen in animal number 1.

Observation at day 14 after instillation of test item revealed: Cornea- Scattered or diffuse areas of opacity (other than slight dulling of normal lustre); details of iris clearly visible in animal numbers 1 and 2 whereas animal number 3 recovered to normal; Area of Opacity- One quarter (or less) but not zero in animal numbers 1 and 2 whereas animal number 3 recovered to normal; Iris: Normal in all 3 animals; Conjunctivae - Some blood vessels definitely hyperaemic (injected) was seen in animal number 3 whereas diffuse, crimson color; individual vessels not easily discernible in animal numbers 1 and 2; Chemosis: some swelling above normal (includes nictitating membranes) was noticed in animal numbers 1 and 2 whereas animal number 3 recovered to normal.

Observation at day 21 after instillation of test item revealed: Cornea- No ulceration or opacity in all 3 animals; Area of Opacity- No opacity was seen in all 3 animals; Iris: Normal in all 3 animals. Conjunctivae: recovered to 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 and chemosis were found 1.00, 0.00, 1.00, 1.00; 1.00, 0.00, 1.67, 1.33 and 1.00, 0.00, 1.33,1.33, respectively.
Other effects:
Clinical Observation
No systemic toxicity was observed in treated rabbits during the experimental period.

Mortality
No mortality was observed during the observation period.

Body weight
Animal Number 1 revealed marginal decrease in body weight where as Animal Numbers 2 and 3 were observed with marginally body weight gain on day 14 as compared to day 0.

Any other information on results incl. tables

Table 1 : Individual Animal Eye Irritation Scores

 

In Treated area Dose:0.1 gm of test item                                                             Sex:Male

Animal Numbers

1

2

3

Application Side

Right

Right

Right

Eye Reactions

At hour

At hour

At hour

*

1

24

48

72

Day 7

Day 14

Day 21

*

1

24

48

72

Day 7

Day 14

Day 21

*

1

24

48

72

Day 7

Day 14

Day 21

Corneal Opacity

0

0

1

1

1

1

1

0

0

0

1

1

1

1

1

0

0

0

1

1

1

1

1

0

Area of Opacity

0

0

1

1

1

1

1

0

0

0

1

1

1

1

1

0

0

0

1

1

1

1

1

0

Iris

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

Conjunctiva

0

1

1

1

1

2

2

0

0

1

1

2

2

2

2

0

0

1

1

2

1

1

1

0

Chemosis

0

0

1

1

1

2

1

0

0

1

1

2

1

1

1

0

0

1

1

2

1

1

0

0

Corneal Damage%

30

30

20

 

 

Dose:Untreated (Control Eye)                                                                         Sex:Male

 

Animal Numbers

1

2

3

Application Side

Left

Left

Left

Eye Reactions

At hour

At hour

At hour

*

1

24

48

72

Day 7

Day 14

Day 21

*

1

24

48

72

Day 7

Day 14

Day 21

*

1

24

48

72

Day 7

Day 14

Day 21

Corneal Opacity

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

Area of Opacity

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

Iris

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

Conjunctiva

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

Chemosis

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

Corneal Damage%

0

0

0

Key:*= Pre-exposure eye examination.

 

 

Table 1 (Continued):Eye Irritation Scores - Mean Values after 24, 48, 72 Hours (Treated eye)

            Animal No.

 Eye Reaction

1

2

3

Corneal Opacity

1.00

1.00

1.00

Iris

0.00

0.00

0.00

Conjunctiva

1.00

1.67

1.33

Chemosis

1.00

1.33

1.33

 

 

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

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

21

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

2

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

3

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

1

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

1.576

1.506

2

1.746

1.891

3

1.686

1.799

Key:kg = Kilogram

 


Applicant's summary and conclusion

Interpretation of results:
Category 2 (irritating to eyes) based on GHS criteria
Conclusions:
Under the experimental conditions tested, eye irritation and reversibility of effects on eyes of all 3 animals was observed at day 21.Hence, test chemical is “Irritant” to New Zealand White Female rabbit eyes
and is thus classified as a eye irritant.



Executive summary:

Acute Eye Irritation/Corrosion Study of test chemical in Rabbits. This study was performed as per OECD guideline no. 405. 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 gm of test item was instilled in the other (treated) eye of each rabbit.The eye was observed at 1, 24, 48, 72 hours, day 7, 14 and 21 after test item instillation.Ophthalmoscope was used for scoring of eye lesions. In the initial test,0.1 gm of test item (Pulverized Form)was applied into the conjunctival sac of the right eye of animal no.1 whereas the left eye of the rabbit served as the control. As animal no. 1 showed ocular lesions, hence confirmatory test was conducted on additional two rabbits (animal no. 2 and 3); 0.1 gmof test itemwas instilled into the conjunctival sac of right eye of both the rabbits and left eye served as the control. Ocular lesions were seen in animal no. 1, 2 and 3 till day 14 observations which recovered at day 21. Untreated eye of all the three rabbits was normal throughout the experimental period of 14 days. 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 3 animals;Iris:Normal in all 3 animals.Conjunctivae -Some blood vessels definitely hyperaemic (injected) was seen in all 3 animals;Chemosis:some swelling above normal (includes nictitating membranes) was seen in animal numbers 2 and 3 whereas animal number 1 was normal. Observation at 24 hours after instillation of test item revealed: Cornea-Scattered or diffuse areas of opacity (other than slight dulling of normal lustre); details of iris clearly visiblein all 3 animals; Area of Opacity-One quarter (or less) but not zero; Iris: Normal in all 3 animals;Conjunctivae-Some blood vessels definitely hyperaemic (injected) in all the animals;Chemosis:Some swelling above normal (includes nictitating membranes) was seen in all the animals. At 24 hours observation the rabbits were examined for corneal epithelium cell damage using sodium fluorescein strips and noticed 30 %, 30 % and 20 % damage in animal no. 1, 2 and 3, resectively.Observation at 48 hours after instillation of test item revealed: Cornea-Scattered or diffuse areas of opacity (other than slight dulling of normal lustre); details of iris clearly visiblein all 3 animals; Area of Opacity-One quarter (or less) but not zero; Iris: Normal in all 3 animals; Conjunctivae -Some blood vessels definitely hyperaemic (injected) was seen in animal number 1 whereas diffuse, crimson color; individual vessels not easily discernible in animal number 2 and 3;Chemosis:Obvious swelling with partial eversion of lids was seen in animal numbers. 2 and 3 whereas some swelling above normal (includes nictitating membranes) was seen in animal no.1. Observation at 72 hours after instillation of test item revealed: Cornea-Scattered or diffuse areas of opacity (other than slight dulling of normal lustre); details of iris clearly visiblein all 3 animals; Area of Opacity-One quarter (or less) but not zero; Iris: Normal in all 3 animals; Conjunctivae -Some blood vessels definitely hyperaemic (injected) was seen in animal numbers 1 and 3 whereas diffuse, crimson color; individual vessels not easily discernible in animal number 2;Chemosis:some swelling above normal (includes nictitating membranes) was noticed in all the animals. Observation at day 7 after instillation of test item revealed: Cornea-Scattered or diffuse areas of opacity (other than slight dulling of normal lustre); details of iris clearly visiblein all 3 animals; Area of Opacity-One quarter (or less) but not zero; Iris: Normal in all 3 animals; Conjunctivae -Some blood vessels definitely hyperaemic (injected) was seen in animal number 3 whereas diffuse, crimson color; individual vessels not easily discernible in animal numbers 1 and 2;Chemosis:some swelling above normal (includes nictitating membranes) was noticed in animal numbers 2 and 3 whereas obvious swelling with partial eversion of lids was seen in animal number 1. Observation at day 14 after instillation of test item revealed: Cornea-Scattered or diffuse areas of opacity (other than slight dulling of normal lustre); details of iris clearly visiblein animal numbers 1 and 2 whereas animal number 3 recovered to normal; Area of Opacity-One quarter (or less) but not zeroin animal numbers 1 and 2 whereas animal number 3 recovered to normal; Iris: Normal in all 3 animals; Conjunctivae -Some blood vessels definitely hyperaemic (injected) was seen in animal number 3 whereas diffuse, crimson color; individual vessels not easily discernible in animal numbers 1 and 2;Chemosis:some swelling above normal (includes nictitating membranes) was noticed in animal numbers 1 and 2 whereas animal number 3 recovered to normal. Observation at day 21 after instillation of test item revealed: Cornea-No ulceration or opacity in all 3 animals; Area of Opacity-No opacity was seen in all 3 animals;Iris:Normal in all 3 animals.Conjunctivae:recovered to normal in all the animals;Chemosis:No swelling was observed in all the animals. The individual mean score for animal nos. 1, 2 and 3at 24, 48, 72 hoursfor corneal opacity, iris, conjunctiva and chemosis were found 1.00, 0.00, 1.00, 1.00; 1.00, 0.00, 1.67, 1.33 and 1.00, 0.00, 1.33,1.33, respectively. Under the experimental conditions tested, eye irritation and reversibility of effects on eyes of all 3 animals was observed at day 21. Hence, test chemical is “Irritant” to New Zealand White Female rabbit eyes and is thus classified as a eye irritant.