Salicylonitrile

Eye irritation

Administrative data

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

29th September 2011 - 14th May 2012

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

The study was conducted according to a national guideline method, essentially equivalent to a OECD TG 405 (1987 edition). Although not conducted under GLP (OECD), the study was conducted under a similar national quality assurance system. Adequately documented study, although not all raw data was reported, this did not effect the conclusions, especially considering the detailed description of the observed effects and persistence. Purity data available separately for test batch.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Test guideline 1

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Version / remarks:

1987 edition

Deviations:

yes

Remarks:

Only total scores for cornea and conjunctivae reported.

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: Guidelines for the Testing of Chemicals: Test No. 405: Acute eye irritation/corrosion test (MEP, People's Republic of China, 2004)

Deviations:

not specified

Principles of method if other than guideline:

The methodology followed was equivalent to the 1987 version of OECD TG 405, which deviates from the latest 2002 version, particularly in the number of animals used. The inital test with 4 animals ("unwashed test") was followed by two further tests of 3 animals each, with eye irrigation at 4 seconds after instillation, and irrigation at 30 seconds, respectively. Evaluation was according to Kay&Calandra, although not explicitly referenced.

GLP compliance:

no

Remarks:

The study was conducted in compliance with a national quality assurance system, with similar objectives to GLP (OECD).

Test material

Test material information

Constituent 1

Reference substance name:

Salicylonitrile

EC Number:

210-259-3

EC Name:

Salicylonitrile

Cas Number:

611-20-1

Molecular formula:

C7H5NO

IUPAC Name:

2-hydroxybenzonitrile

Details on test material:

- Name of test material (as cited in study report): 2-cyanophenol
- Substance type: powder
- Physical state: solid
- Purity test date: not reported
- Expiration date of the lot/batch: not reported
- Storage condition of test material: not reported

Test animals / tissue source

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Age at study initiation: not reported
- Weight at study initiation: ca. 2.3kg
- Housing: Suspended, wire bottom, stainless steel, 1 animal per cage
- Diet: ad libitum (Statement that contaminents in feed and water effecting study outcome not expected)
- Water: ad libitum (Municipal water supply, analyzed by City Water Quality Monitoring Center of Zhejiang Province)
- Acclimation period: 5 days
- Sex: male and female, nulliparous & non-pregnant

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20±3°C
- Humidity (%): 30-70%
- Air changes (per hr): 10-12
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: not reported

Test system

Vehicle:

unchanged (no vehicle)

Remarks:

Ground to a fine powder and sieved through 0.154mm mesh

Controls:

not required

Amount / concentration applied:

TEST MATERIAL
- Amount applied: 0.1g

Duration of treatment / exposure:

- Initial Test: 24 hours
- Second Test: 4 seconds
- Third Test: 30 seconds

Observation period (in vivo):

21 days with observations at 1h, 24h, 48h, 72h, 96h, 7 days, 14 days, 21 days.

Number of animals or in vitro replicates:

- Initial Test: 4
- Second Test: 3
- Third Test: 3

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing: gently rinsed for 30s with 0.9% saline water
- Initial Test, time after start of exposure: 24h
- Second Test, time after start of exposure: 4s and 24h
- Third Test, time after start of exposure: 30s and 24h

SCORING SYSTEM:
As described in The Guidelines for the Testing of Chemicals: Test No. 405: Acute Eye Irritation/Corrosion Test (Ministry of Environmental Protection of the People's Republic of China.

TOOL USED TO ASSESS SCORE:
Corneas of all treated eyes were examined with sodium fluorescein ophthalmic solution at 24h. Any corneas exhibiting staining were followed at subsequent observation points with fluorescein solution until staining no longer occurred.

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

cornea opacity score

Basis:

animal: #1, #4

Time point:

24/48/72 h

Score:

40

Reversibility:

not fully reversible within: 21 days

Results 2

Irritation parameter:

cornea opacity score

Basis:

animal #2

Time point:

24 h

Score:

20

Reversibility:

fully reversible within: 21 days

Results 3

Irritation parameter:

cornea opacity score

Basis:

animal #2

Time point:

48 h

Score:

20

Reversibility:

fully reversible within: 21 days

Results 4

Irritation parameter:

cornea opacity score

Basis:

animal #2

Time point:

72 h

Score:

15

Reversibility:

fully reversible within: 21 days

Results 5

Irritation parameter:

cornea opacity score

Basis:

animal #3

Time point:

24 h

Score:

40

Reversibility:

not fully reversible within: 21 days

Results 6

Irritation parameter:

cornea opacity score

Basis:

animal #3

Time point:

48 h

Score:

40

Reversibility:

not fully reversible within: 21 days

Results 7

Irritation parameter:

cornea opacity score

Basis:

animal #3

Time point:

72 h

Score:

20

Reversibility:

not fully reversible within: 21 days

Results 8

Irritation parameter:

iris score

Basis:

animal: #1, #3, #4

Time point:

24/48/72 h

Score:

5

Reversibility:

not fully reversible within: 21 days

Results 9

Irritation parameter:

iris score

Basis:

animal #2

Time point:

24/48/72 h

Score:

5

Reversibility:

fully reversible within: 21 days

Results 10

Irritation parameter:

conjunctivae score

Basis:

animal #1

Time point:

24 h

Score:

12

Reversibility:

not fully reversible within: 21 days

Results 11

Irritation parameter:

conjunctivae score

Basis:

animal #1

Time point:

48 h

Score:

12

Reversibility:

not fully reversible within: 21 days

Results 12

Irritation parameter:

conjunctivae score

Basis:

animal #1

Time point:

72 h

Score:

18

Reversibility:

not fully reversible within: 21 days

Results 13

Irritation parameter:

conjunctivae score

Basis:

animal #2

Time point:

24/48/72 h

Score:

16

Reversibility:

not fully reversible within: 21 days

Results 14

Irritation parameter:

conjunctivae score

Basis:

animal #3

Time point:

24/48/72 h

Score:

16

Reversibility:

not reversible

Results 15

Irritation parameter:

conjunctivae score

Basis:

animal #3

Time point:

24/48/72 h

Score:

18

Reversibility:

not fully reversible within: 21 days

Results 16

Irritation parameter:

overall irritation score

Remarks:

Sum of cornea, iris and conjunctivae scores

Basis:

animal #1

Time point:

21 d

Score:

41

Reversibility:

not fully reversible within: 21 days

Remarks on result:

other: No eye irrigation; see tables for full data

Results 17

Irritation parameter:

overall irritation score

Remarks:

Sum of cornea, iris and conjunctivae scores

Basis:

animal #2

Time point:

21 d

Score:

10

Reversibility:

not fully reversible within: 21 days

Remarks on result:

other: No eye irrigation; see tables for full data

Results 18

Irritation parameter:

overall irritation score

Remarks:

Sum of cornea, iris and conjunctivae scores

Basis:

animal #3

Time point:

21 d

Score:

28

Reversibility:

not fully reversible within: 21 days

Remarks on result:

other: No eye irrigation; see tables for full data

Results 19

Irritation parameter:

overall irritation score

Remarks:

Sum of cornea, iris and conjunctivae scores

Basis:

animal #4

Time point:

21 d

Score:

22

Reversibility:

not fully reversible within: 21 days

Remarks on result:

other: No eye irrigation; see tables for full data

Results 20

Irritation parameter:

chemosis score

Basis:

animal: #1, #2, #3, #4

Time point:

24/48/72 h

Remarks on result:

other: not reported

Irritant / corrosive response data:

Observation results of eye reaction (unwashed test) after the application were shown as diffuse beefy red, swelling with lids more than half closed, discharge with moistening of the lids and hairs, and a considerable area around the eye in the conjunctivae. The main symptoms affecting the cornea include easily discernible translucent area, details of iris slightly obscured, the damage area covered greater than three quarters, or up to whole area. There was observed fluorescein staining with scales in corneas and congestion in iris. Area of angiogenesis was greater than one quarter, but less than half in cornea surface (moderately. The corneas displayed slight dulling of normal luster. The highest average of total score until 4d after instillation was 55.50. Duration of irritation reaction was the average of total score on day 7>40.

Observation results of eye reaction (washed test, 4s after instillation) were shown as diffuse beefy red, swelling with lids about half closed, discharge with moistening of the lids and hairs, and a considerable area around the eye in the conjunctivae. The main symptoms affecting the cornea include easily discernible translucent area, details of iris slightly obscured, the damage area covered greater than three quarters. There was observed fluoresdcein staining within the corneal opacity area and congestion in iris. Area of angiogenesis was greater than ten percent, but less than one quarter in cornea surface (mildly). The corneas displayed slight dulling of normal luster. The highest average of total score until 4d after instillation was 40.67. Duration of irritation reaction was the average of total score on day 7>20.

Observation results of eye reaction (washed test, 30s after instillation) were shown as diffuse beefy red, swelling with lids about half closed, discharge with moistening of the lids and hairs, and a considerable area around the eye in the conjunctivae. The main symptoms affecting the cornea include easily discernible translucent area, details of iris slightly obscured, the damage area covered greater than three quarters. There was observed fluorescein staining within the corneal opacity area and congestion in iris. Area of angiogenesis was greater than ten percent, but less than one quarter in cornea surface (mildly) . The corneas were slight dulling of normal luster. The highest average of total score until 4d after instillation was 53.00. Duration of irritation reaction was the average of total score on day 7<40, more than half animals>30, but no animals>60.

Other effects:

None reported.

Any other information on results incl. tables

Table 1: Initial and final body weights for initial test (“unwashed”).

	Animal 1	Animal 2	Animal 3	Animal 4
Start weight	2.40	2.42	2.30	2.13
Finish weight	2.46	2.50	2.42	2.21

 

Table 2: Ocular reactions for the initial test (“unwashed”).

Time after treatment	Animal number	Score			Total score	Average of total score
		Cornea	Iris	Conjunctivae
1h	1	-	0	6	6	6.5
	2	-	0	6	6
	3	-	0	6	6
	4	-	0	8	8
24h	1	40	5	12	57	55.50
	2	20	5	16	41
	3	40	5	16	61
	4	40	5	18	63
48h	1	40	5	12	57	55.50
	2	20	5	16	41
	3	40	5	16	61
	4	40	5	18	63
72h	1	40	5	18	63	50.75
	2	15	5	16	36
	3	20	5	16	41
	4	40	5	18	63
96h	1	40	5	18	63	50.75
	2	15	5	16	36
	3	20	5	16	41
	4	40	5	18	63
7 day	1	40	5	18	63	44.75
	2	15	5	16	36
	3	20	5	14	39
	4	20	5	16	41
14 day	1	20	5	18	43	30.5
	2	5	5	12	22
	3	10	5	18	33
	4	5	5	14	24
21 day	1	20	5	16	41	25.25
	2	0	0	10	10
	3	5	5	18	28
	4	5	5	12	22

The highest average total score up to 4 days after instillation: 55.50

Duration of irritation reaction: the average total score on day 7: >40

Evaluation: “very severely irritating” (rating 7)

Table 3: Ocular reactions at 21 days for the post-instillation irrigation satellite groups (second and third tests), with eye irrigation at 4s and 30s after instillation respectively.

 

Time after treatment	Animal number	Score			Total score	Average of total score
		Cornea	Iris	Conjunctivae
Second test (4s), 21 days	1	0	0	0	0	0
	2	0	0	0	0
	3	0	0	0	0
Evaluation	The highest average total score up to 4 days after instillation: 40.67 Duration of irritation reaction: the average total score on day 7: >20 Evaluation: “severely irritating” (rating 6)
Third test (30s), 21 days	1	5	5	10	20
	2	5	5	8	18
	3	0	0	8	8
Evaluation	The highest average total score up to 4 days after instillation: 53.00 Duration of irritation reaction: the average total score on day 7: <40, more than half animals >30, but no animals >60 Evaluation: “severely irritating” (rating 6)

Applicant's summary and conclusion

Interpretation of results:

Category 1 (irreversible effects on the eye) based on GHS criteria

Conclusions:

The current OECD and EU guidelines deviate from the 1987 OECD guideline primarily with respect to numbers of animals and use of satellite groups. Despite only the total scores for cornea and conjunctivae being reported, the descriptive summary allows reconstruction of the maximum scores for corneal opacity (2), the area involved (4), and iris (1). A combination of the descriptive summary of the observations and the effects being observed to 21 days, means the report is adequate for classification and risk assessment purposes.

Executive summary:

In a primary eye irritation study, 0.1g of 2-cyanophenol was instilled into the conjunctival sac of the right eye of male or female New Zealand white rabbits (4) for 24 hours. Treated eyes were gently washed with 0.9% saline solution after 24 hours. Animals then were observed for 21 days.  Irritation was scored by the method of Draize. Ocular reactions after instillation were diffuse beefy red conjunctivae, swelling with lids more than half closed, discharge with moistening of the lids and hairs, and a considerable area around the eye. The cornea showed discernible translucent areas, details of iris slightly obscured, and the damaged area covered greater than three quarters, or up to whole area. There was congestion evident in the iris. Effects were persistent to 21 days. In this study, 2 -cyanophenol was found to be very severely irritating (rating 7) according to Guidelines for the testing of chemicals: Test No. 405: Acute eye irritation/corrosion test (Ministry of Environmental Protection of the People's Republic of China, 2004).

Two satellite groups, each of 3 animals, were instilled with 0.1g of 2-cyanophenol into the conjunctival sac of the right eye for 24 hours. Treated eyes were gently washed with 0.9% saline solution after 4 and 30 seconds respectively. Animals then were observed for 21 days.  Irritation was scored by the method of Draize.