N,N,N-tributylbutan-1-aminium ({[(2S,5R)-2-carbamoyl-7-oxo-1,6-diazabicyclo[3.2.1]oct-6-yl]oxy}sulfonyl)oxidanide

Eye irritation

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

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)

Version / remarks:

2nd October, 2012

Principles of method if other than guideline:

The objective of the study was to assess the irritant and/or corrosive effects of “Tetrabutylammonium bromide (CAS No. - 1643-19-2)” on eye, when exposed by the ocular route in rabbits.

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Tetrabutylammonium bromide

EC Number:

216-699-2

EC Name:

Tetrabutylammonium bromide

Cas Number:

1643-19-2

Molecular formula:

C16H36N.Br

IUPAC Name:

N,N,N-tributylbutan-1-aminium bromide

Details on test material:

- Name of test material (as cited in study report):Tetrabutylammonium bromide
- Molecular formula :C16H36N.Br
- Molecular weight :322.37 g/mol
- Substance type:Organic
- Physical state:White Solid (Crystals)
- Lot/batch No.:Lot 1/02
- AI Content:99.9 %
- Storage condition of test material:The test item was stored in a cool, dry place and kept in a closed.
- Other:
Handling and Disposal
Safety precautions : Aprons, masks, caps, gloves and goggles were used to ensure the health and safety of the personnel.
Disposal : No test item was disposed during the conduct of the study and the same was documented in the raw data.

Test animals / tissue source

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Rabbiroof, Hyderabad, India.
- Age at study initiation: Appr. 4.5 - 5.5 months
- Weight at study initiation: 1.932 kg- 2.438 kg
- Health Status: Healthy young adult, females were nulliparous and non pregnant.
- Housing: The animals were housed individually in stainless steel cages.
- Diet: All animals were provided conventional laboratory rabbit diet (Nutrivet Life Sciences, Pune), ad libitum.
- Water: Aqua guard filtered tap water, ad libitum.
- Acclimation period: At laeast 6 days.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19.5 - 22.7
- Humidity (%): 41.2 - 67.3
- Air changes (per hr): More than 12
- Photoperiod (hrs dark / hrs light): 12/12

Test system

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent no treatment

Amount / concentration applied:

TEST MATERIAL
- Amount applied:100 mg
- Test Item Application Procedure: Test item (pulverized) was placed in the conjunctival sac of eye in three rabbits, after gently pulling the lower lid away from the eyeball. The lids were held together to prevent loss of test item. Other eye which was untreated, served as control. Treated and control eyes of rabbits were examined with the help of ophthalmoscope at one hour after instillation of test item.

Duration of treatment / exposure:

Single instillation

Observation period (in vivo):

1, 24, 48, 72 hours and at day7 after instillation of test item

Number of animals or in vitro replicates:

3 female rabbits

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing: The treated eye of rabbit was washed with normal saline
- Time after start of exposure: 24 hours
Both the eyes of rabbit were further examined for corneal epithelial cell damage with the help of fluorescent strips. The strips were wetted with one drop of normal saline and placed in both the eyes one by one, gently closed for 30 seconds and then washed with normal saline. The area of corneal epithelial cell damage was scored using ophthalmoscope (Ri-Mini2, 5 V-guru; Model No. 3002). Initially, a single rabbit was used for testing. The residual test item was removed after 24 hours and the response was graded. No severe lesions were observed after test item application, hence additional two rabbits (Animal No. 2 and 3) were used to confirm the response.

SCORING SYSTEM: Draize Method

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

cornea opacity score

Basis:

animal #1

Time point:

24/48/72 h

Score:

1

Max. score:

4

Reversibility:

fully reversible within: 7 days

Results 2

Irritation parameter:

cornea opacity score

Basis:

animal #2

Time point:

24/48/72 h

Score:

1

Max. score:

4

Reversibility:

fully reversible within: 7 days

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: 7 days

Results 4

Irritation parameter:

iris score

Basis:

animal: all 3 rabbits

Time point:

other: 24, 48, 72 hours and day 7

Score:

0

Max. score:

2

Results 5

Irritation parameter:

conjunctivae score

Basis:

animal #1

Time point:

24/48/72 h

Score:

1.67

Max. score:

3

Reversibility:

fully reversible within: 7 days

Results 6

Irritation parameter:

conjunctivae score

Basis:

animal #2

Time point:

24/48/72 h

Score:

2

Max. score:

3

Reversibility:

fully reversible within: 7 days

Results 7

Irritation parameter:

conjunctivae score

Basis:

animal #3

Time point:

24/48/72 h

Score:

2

Max. score:

3

Reversibility:

fully reversible within: 7 days

Results 8

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

24/48/72 h

Score:

1.33

Max. score:

4

Reversibility:

fully reversible within: 7 days

Results 9

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

24/48/72 h

Score:

2

Max. score:

4

Reversibility:

fully reversible within: 7 days

Results 10

Irritation parameter:

chemosis score

Basis:

animal #3

Time point:

24/48/72 h

Score:

1.67

Max. score:

4

Reversibility:

fully reversible within: 7 days

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 was seen in animal no. 1 and 3 whereas scattered or diffuse areas of opacity (other than slight dulling of normal lustre) details of iris clearly visible was seen in animal no. 2; Area of Opacity- Zero in animal no. 1 and 3 whereas one quarter (or less) but not zero was seen in animal no.2; Iris: Normal in all the animals. Conjunctivae - Diffuse, crimson color; individual vessels not easily discernible in animal no. 2 and some blood vessels definitely hyperaemic (injected) animal no. 1 and 3; Chemosis: Some swelling above normal (includes nictitating membranes) in animal no. 1 and obvious swelling with partial eversion of lids was observed in animal no. 2 and 3.

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- Greater than one quarter, but less than half was seen in animal no. 1 and 2 whereas one quarter (or less) but not zero was seen in animal no.3; Iris: Normal in all the animals. Conjunctivae - Diffuse, crimson color; individual vessels not easily discernible was seen in all 3 animals; Chemosis: Some swelling above normal (includes nictitating membranes) was seen in animal no.1 whereas obvious swelling with partial eversion of lids was seen in animal no.2 and 3.

At 24 hours observation the rabbits were examined for corneal epithelium cell damage using sodium fluorescein strips and noticed 50%, 60% and 45% damage in animal no. 1, 2 and 3 respectively.

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 was seen in animal no.1 and 3 whereas greater than one quarter, but less than half was seen in animal no. 2; Iris: Normal in all the animals. Conjunctivae - Some blood vessels definitely hyperaemic (injected) was seen in animal no. 1 whereas diffuse, crimson color; individual vessels not easily discernible was seen in animal no. 2 and 3; Chemosis: Obvious swelling with partial eversion of lids was seen in all 3 animals.

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 was seen in all 3 animals; Iris: Normal in all the animals. Conjunctivae - Diffuse, crimson color; individual vessels not easily discernible was seen in all 3 animals; Chemosis: Some swelling above normal (includes nictitating membranes) was seen in animal no. 1 and 3 whereas obvious swelling with partial eversion of lids was seen in animal no. 2.

Observation on day 7 after instillation of test item revealed: Cornea- No ulceration or opacity in all 3 animals; Area of Opacity- Zero percent was seen in all 3 animals; Iris: Normal in all the animals. Conjunctivae - Blood vessels normal was seen in all 3 animals; Chemosis: No swelling (Normal) was seen in all the 3 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 1.33, 0.00, 1.67, 1.33; 1.67, 0.00, 2.00, 2.00; 1.00, and 1.00 0.00, 2.00, 1.67, respectively.

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

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- Greater than one quarter, but less than half was seen in animal no. 1 and 2 whereas one quarter (or less) but not zero was seen in animal no.3; Iris: Normal in all the animals. Conjunctivae - Diffuse, crimson color; individual vessels not easily discernible was seen in all 3 animals; Chemosis: Some swelling above normal (includes nictitating membranes) was seen in animal no.1 whereas obvious swelling with partial eversion of lids was seen in animal no.2 and 3.

At 24 hours observation the rabbits were examined for corneal epithelium cell damage using sodium fluorescein strips and noticed 50%, 60% and 45% damage in animal no. 1, 2 and 3 respectively.

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 was seen in animal no.1 and 3 whereas greater than one quarter, but less than half was seen in animal no. 2; Iris: Normal in all the animals. Conjunctivae - Some blood vessels definitely hyperaemic (injected) was seen in animal no. 1 whereas diffuse, crimson color; individual vessels not easily discernible was seen in animal no. 2 and 3; Chemosis: Obvious swelling with partial eversion of lids was seen in all 3 animals.

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 was seen in all 3 animals; Iris: Normal in all the animals. Conjunctivae - Diffuse, crimson color; individual vessels not easily discernible was seen in all 3 animals; Chemosis: Some swelling above normal (includes nictitating membranes) was seen in animal no. 1 and 3 whereas obvious swelling with partial eversion of lids was seen in animal no. 2.
Observation on day 7 after instillation of test item revealed: Cornea- No ulceration or opacity in all 3 animals; Area of Opacity- Zero percent was seen in all 3 animals; Iris: Normal in all the animals. Conjunctivae - Blood vessels normal was seen in all 3 animals; Chemosis: No swelling (Normal) was seen in all the 3 animals.
No effects were seen in the untreated eyes.

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
All rabbits gained weight during experimental period.

Any other information on results incl. tables

Table 1 : Individual Animal Eye Irritation Scores

 

In Treated area Dose:100 mg of test item                                                            Sex:Female

Animal Numbers		1							2								3
Application Side		Left							Right								Right
Eye Reactions		At hour						Day		At hour						Day			At hour						Day
		*		1		24		48		72		7		*		1		24		48		72			7		*		1		24		48		72			7
Corneal Opacity		0		0		1		1		1		0		0		1		1		1		1			0		0		0		1		1		1			0
Area of Opacity		0		0		2		1		1		0		0		1		2		2		1			0		0		0		1		1		1			0
Iris		0		0		0		0		0		0		0		0		0		0		0			0		0		0		0		0		0			0
Conjunctiva		0		2		2		1		2		0		0		1		2		2		2			0		0		1		2		2		2			0
Chemosis		0		1		1		2		1		0		0		2		2		2		2			0		0		2		2		2		1			0
Corneal Damage%		50%							60%								45%

 

Dose:Untreated (Control Eye)                                                                       Sex:Female

Animal Numbers	1							2							3
Application Side	Right							Left							Left
Eye Reactions	At hour					Day		At hour					Day		At hour					Day
	*	1	24	48	72		7	*	1	24	48	72		7	*	1	24	48	72		7
Corneal Opacity	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
Iris	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
Chemosis	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	1.33	1.67	1.00
Iris	0.00	0.00	0.00
Conjunctiva	1.67	2.00	2.00
Chemosis	1.33	2.00	1.67

 

 

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:Female

Animal No.	Days (Post application observation)
	0	1	2	3	4	5	6	7
1	1	1	1	1	1	1	1	1
2	1	1	1	1	1	1	1	1
3	1	1	1	1	1	1	1	1

Key:1 = Normal

 

Table 3: Individual Animal Body Weight

Sex :Female

Animal No.	Animal Body Weight (kg)
	Prior to application	At termination
1	2.208	2.387
2	1.932	2.214
3	2.438	2.646

Key:kg = Kilogram

Applicant's summary and conclusion

Interpretation of results:

Category 2 (irritating to eyes) based on GHS criteria

Conclusions:

Based on the results of an acute Eye Irritation/Corrosion Study was performed according to OECD guideline with tetrabutylammonium bromide, it is concluded that tetrabutylammonium bromide is “Mildly Irritating to Eyes” and is being classified as an eye irritant in 'category 2' as per the CLP regulation. This result is read across to SAM-3.

Executive summary:

Based on the results of an acute Eye Irritation/Corrosion Study was performed according to OECD guideline with tetrabutylammonium bromide, it is concluded that tetrabutylammonium bromide is “Mildly Irritating to Eyes” and is being classified as an eye irritant in 'category 2' as per the CLP regulation. This result is read across to SAM-3.