2-nitro-4-(trifluoromethyl)benzonitrile

Eye irritation

Administrative data

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

07 - 26 Oct 2019

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2019

Report date:

2019

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Version / remarks:

adopted 14 Jun 2021

Deviations:

yes

Remarks:

No topical anesthetics and systemic analgesics were used.

GLP compliance:

yes

Remarks:

The test complied with the Principles of Good Laboratory Practices (GLP) of the Certification and Accreditation Administration of the People’s Republic of China (2013 revised edition).

Test material

Test material information

Constituent 1

Reference substance name:

2-nitro-4-(trifluoromethyl)benzonitrile

EC Number:

212-298-1

EC Name:

2-nitro-4-(trifluoromethyl)benzonitrile

Cas Number:

778-94-9

Molecular formula:

C8H3F3N2O2

IUPAC Name:

2-nitro-4-(trifluoromethyl)benzonitrile

Test animals / tissue source

Species:

rabbit

Strain:

other: Japanese long-eared white rabbit, general grade

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Qingdao Kangda Biological Technology Co., Ltd., Shandong, China
- Age at study initiation: not reported
- Weight at study initiation: 1.70 - 1.85 kg (males)
- Fasting period before study: not reported
- Housing: individually in in hanging stainless steel cages (L38 cm x W50 cm x H35 cm) affixed on rearing shelves (L200 cm x W65 cm x H170 cm)
- Diet: rabbit maintenance feed (Liaoning Changsheng Biotechnology Co., Ltd, Liaoning, China); ad libitum
- Water: grade-one reverse osmosis water (Liaoning Changsheng Biotechnology Co., Ltd, Liaoning, China); ad libitum
- Acclimation period: 25 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20.10 - 24.00
- Humidity (%): 40.20 - 70.00
- Air changes (per hr): not reported
- Photoperiod (hrs dark / hrs light): 12 / 12

IN-LIFE DATES: 07 - 26 Oct 2019

Test system

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent no treatment

Amount / concentration applied:

0.1 g ground test sample

Duration of treatment / exposure:

single application without washing

Observation period (in vivo):

16 days
Reading time points: 1 h, 24 h, 48 h, 72 h, and 4 - 16 d

Duration of post- treatment incubation (in vitro):

not applicable

Number of animals or in vitro replicates:

3 males

Details on study design:

PRE- / POST-TREATMENT OF THE ANIMALS:
No topical anaesthetics and stystemic analgesics were administered prior to nor after test substance application.

REMOVAL OF TEST SUBSTANCE
- Washing: not performed

SCORING SYSTEM: Draize scoring system.

TOOL USED TO ASSESS SCORE: fluorescein
Approximately 24 h before the test started, a headlight magnifier and 2% sodium fluorescein (vehicle: normal saline) were used to examine the exterior, conjunctiva, cornea, and iris of both eyes of the animals. Approximately 1 hour, 24 hours, 48 hours, 72 hours, and 4 - 16 days after the test sample was applied, a headlight magnifier was used to examine both eyes of the test animals. In addition, at observation points approximately 24 hours, 48 hours, 72 hours, 4 - 7 days, and 14 days after the test sample was applied, a fluorescein examination was carried out on both eyes of the test animals.
Fluorescein staining examination method: One drop of 2% sodium fluorescein normal saline solution was dripped into the conjunctival sac of the eye. After 15 sec, normal saline was used to gently rinse the eye, and the headlight magnifier was used to examine whether the cornea had retained the fluorescein.
At the pre-test eye examination, no eye irritation, ocular defects, or corneal damage were visible in the eyes of the test animals.

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

cornea opacity score

Basis:

animal #1

Time point:

24/48/72 h

Score:

2.7

Max. score:

4

Reversibility:

fully reversible within: 14 d

Remarks on result:

positive indication of irritation

Results 2

Irritation parameter:

cornea opacity score

Basis:

animal #2

Time point:

24/48/72 h

Score:

3

Max. score:

4

Reversibility:

fully reversible within: 16 d

Remarks on result:

positive indication of irritation

Results 3

Irritation parameter:

cornea opacity score

Basis:

animal #3

Time point:

24/48/72 h

Score:

3

Max. score:

4

Reversibility:

fully reversible within: 15 d

Remarks on result:

positive indication of irritation

Results 4

Irritation parameter:

iris score

Basis:

animal #1

Time point:

24/48/72 h

Score:

2

Max. score:

2

Reversibility:

fully reversible within: 6 d

Remarks on result:

positive indication of irritation

Results 5

Irritation parameter:

iris score

Basis:

animal #2

Time point:

24/48/72 h

Score:

2

Max. score:

2

Reversibility:

fully reversible within: 7 d

Remarks on result:

positive indication of irritation

Results 6

Irritation parameter:

iris score

Basis:

animal #3

Time point:

24/48/72 h

Score:

2

Max. score:

2

Reversibility:

fully reversible within: 6 d

Remarks on result:

positive indication of irritation

Results 7

Irritation parameter:

conjunctivae score

Basis:

animal #1

Time point:

24/48/72 h

Score:

3

Max. score:

3

Reversibility:

fully reversible within: 15 d

Remarks on result:

positive indication of irritation

Results 8

Irritation parameter:

conjunctivae score

Basis:

animal #2

Time point:

24/48/72 h

Score:

3

Max. score:

3

Reversibility:

fully reversible within: 12 d

Remarks on result:

positive indication of irritation

Results 9

Irritation parameter:

conjunctivae score

Basis:

animal #3

Time point:

24/48/72 h

Score:

3

Max. score:

3

Reversibility:

fully reversible within: 12 d

Remarks on result:

positive indication of irritation

Results 10

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

24/48/72 h

Score:

3

Max. score:

4

Reversibility:

fully reversible within: 16 d

Remarks on result:

positive indication of irritation

Results 11

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

24/48/72 h

Score:

3

Max. score:

4

Reversibility:

fully reversible within: 13 d

Remarks on result:

positive indication of irritation

Results 12

Irritation parameter:

chemosis score

Basis:

animal #3

Time point:

24/48/72 h

Score:

3

Max. score:

4

Reversibility:

fully reversible within: 12 d

Remarks on result:

positive indication of irritation

Irritant / corrosive response data:

After test substance application severe effects on the eyes (iritis, chemosis, conjunctivitis and corneal opacity) occurred starting partly 1 h after test substance application. All observed effects were reversible - latest on day 16 after test material application. For individual and mean ocular scores, see tables 2 and 3 under section "any other information on results incl. tables".

Other effects:

Clinical symptoms: All three animals showed clinical symptoms after test item administration. The initial response was closed eyes for all animals. Chemosis and secretions were noted from 1 h after administration, and corneal opacity occurred from 24 h after administration. Symptoms reversed until 16 days after treatment at the latest. For details on clinical symptoms, see table 1 under section "any other information on results incl. tables".
Body weight: All animals gained body weight (0.40 - 0.45 kg) during the test period and had similar weights at the end of the test (2.50 - 2.55 kg).
Fluorescein retention: All three animals exhibited retention of sodium fluorescein at 24 h, 48 h, 72 h and 4 - 7 days after test chemical administration. On the 14th day after administration, the retained sodium fluorescein had disappeared.

Any other information on results incl. tables

Table 1: Table of Clinical Observations

	Animal 1	Animal 2	Animal 3
Initial response	B	B	B
1 h	BSF	B1SF	B1SF
7 h	BSF	B1SF	B1SF
24 h	B1SFH	B1SFH	B1SFH
30 h	B1SFH	B1SFH	B1SFH
48 h	B1SFH	B1SFH	B1SFH
54 h	B1SFH	B1SFH	B1SFH
72 h	B1SFH	B1SFH	B1SFH
78 h	B1SFH	B1SFH	B1SFH
4 d	B1SFH	B1SFH	B1SFH
5 d	B1SFH	B1SFH	B1SFH
6 d	SFH	B1SFH	SFH
7 d	SFH	SFH	SFH
8 d	SFH	SFH	SFH
9 d	SH	SFH	SH
10 d	SH	SH	SH
11 d	SH	SH	SH
12 d	SH	SH	H
13 d	SH	H	H
14 d	S	H	H
15 d	S	H	N
16 d	N	N	-

N: no abnormalities; B: closed eyes; S: chemosis; F: secretions; H: corneal opacity; B1: half-closed eyes

 

 

Table 2: Table of Individual Eye Response Total Scores

Animal no.	Ocular response	Ocular response total score – observation timepoints
		1 h	24 h	48 h	72 h	4 d	5 d	6 d	7 d	8 d	9 d	10 d	11 d	12 d	13 d	14 d	15 d	16 d
1	Cornea	0	3	3	2	2	2	1	1	1	1	1	1	1	1	0	0	0
	Iris	0	2	2	2	2	2	0	0	0	0	0	0	0	0	0	0	0
	Conjunctival hyperaemia	1	3	3	3	3	2	2	2	2	2	2	1	1	1	1	0	0
	Chemosis	2	3	3	3	3	3	2	2	2	2	1	1	1	1	1	1	0
2	Cornea	0	3	3	3	3	3	3	2	2	2	1	1	1	1	1	1	0
	Iris	0	2	2	2	2	2	2	0	0	0	0	0	0	0	0	0	0
	Conjunctival hyperaemia	1	3	3	3	2	2	2	2	2	2	1	1	0	0	0	0	0
	Chemosis	2	3	3	3	3	3	3	2	2	2	1	1	1	0	0	0	0
3	Cornea	0	3	3	3	3	3	2	2	2	1	1	1	1	1	1	0	-
	Iris	0	2	2	2	2	2	0	0	0	0	0	0	0	0	0	0	-
	Conjunctival hyperaemia	1	3	3	3	2	2	2	2	2	2	1	1	0	0	0	0	-
	Chemosis	2	3	3	3	3	3	2	2	2	1	1	1	0	0	0	0	-

 

Table 3: Individual Eye Response Mean Scores

Animal no.	Ocular response	Individual ocular response – mean score
1	Cornea	2.7
	Iris	2
	Conjunctival hyperaemia	3
	Chemosis	3
2	Cornea	3
	Iris	2
	Conjunctival hyperaemia	3
	Chemosis	3
3	Cornea	3
	Iris	2
	Conjunctival hyperaemia	3
	Chemosis	3

 

Applicant's summary and conclusion

Interpretation of results:

other: Category 1 "Eye Dam. 1" (H318) according to Regulation (EC) No 1272/2008

Conclusions:

Under the present test conditions, the test substance 2-nitro-4-(trifluoromethyl)benzonitrile was corrosive to the eyes of three male rabbits. The mean scores for cornea/iris/conjunctiva hyperaemia/chemosis in the right eye at 24 h, 48 h, and 72 h after test chemical administration were 2.7/2/3/3, 3/2/3/3, and 3/2/3/3 for the three animals, respectively. According to the Regulation (EC) No 1272/2008, the test sample thus meets the criteria for Eye Damage 1 (H318).