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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
Study period:
03-JUN-2004 to 10-AUG-2004
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
Data is from experimental report

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2004
Report Date:
2004

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 405 (Acute Eye Irritation / Corrosion)
Qualifier:
according to
Guideline:
other: Directive 92/69 EEC, B.5. "Acute Toxicity - Eye Irritation", July 31, 1992.
Principles of method if other than guideline:
The purpose of this primary eye irritation study was to assess the possible irritation potential when a single dose of C 010 was placed in the conjunctival sac of rabbit eyes.
The test item was applied at 0.1 g/animal, the dose specified in the test guidelines for a solid test item.
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid
Details on test material:
- Name of test material (as cited in study report): Basic yellow 57
- Molecular formula (if other than submission substance): C19-H22-N5-O.Cl
- Molecular weight (if other than submission substance): 371.87 g/mol
- Smiles notation (if other than submission substance): N1(C([C@@H](\N=N\c2cc([N+](C)(C)C)ccc2)C(=N1)C)=O)c1ccccc1.[ClH-]
- InChl (if other than submission substance): 1S/C19H22N5O.ClH/c1-14-18(19(25)23(22-14)16-10-6-5-7-11-16)21-20-15-9-8-12-17(13-15)24(2,3)4;/h5-13,18H,1-4H3;1H/q+1;/p-1/b21-20+;
- Structural formula attached as image file (if other than submission substance): No data available
- Substance type: Organic
- Physical state: Solid orange powder
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Identification: C 010 (3-[(4,5-dihydro-3-methyl-5-oxo-1-phenyl-1H-pyrazol-4-yl)azo]-N,N,N-trimethylanilinium chloride)
- Lot/batch No.of test material: 15
- Expiration date of the lot/batch: 30-DEC-2023
- Purity: 99.3% HPLC

RADIOLABELLING INFORMATION (Not applicable)
- Radiochemical purity: N/A
- Specific activity: N/A
- Locations of the label: N/A
- Expiration date of radiochemical substance: N/A

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: In the refrigerator (range of 5 ± 3°C), light protected.
- Stability under test conditions: Stable under storage conditions
- Solubility and stability of the test substance in the solvent/vehicle: No data
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium:No data

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: 0.1 g (per animal) of C 010 was weighed and applied undiluted as it was delivered by the Sponsor.
- Preliminary purification step (if any): No data
- Final dilution of a dissolved solid, stock liquid or gel: No data
- Final preparation of a solid: No data

FORM AS APPLIED IN THE TEST (if different from that of starting material)

OTHER SPECIFICS:
Safety precautions : Routine hygienic procedures were used to ensure the health and safety of the personnel.

TEST ITEM PREPARATION
The pH of a 1 % (w/w) solution of the test item was measured for a previous study (RCC Study number 853983, skin irritation with C 010 in rabbits) and was found to be 6.53.

Test animals / tissue source

Species:
rabbit
Strain:
New Zealand White
Remarks:
SPF
Details on test animals or tissues and environmental conditions:
TEST ANIMALS
- Source: Elevage Scientifique des Dombes
F-01400 Chatillon sur Chalaronne / France
- Age at study initiation: 15 - 16 weeks (male),15 - 16 weeks (females)
- Weight at study initiation: Given below
- Identification: By unique cage number and corresponding ear number.
- Allocation: Male No. 82,Female Nos. 83 and 84
- Housing: Individually in stainless steel cages equipped with feed hoppers and drinking water bowls. Wood blocks (RCC Ltd,Füllinsdorf) and haysticks 4642 (batch no.80/03, Provimi Kliba AG) were provided for gnawing.
- Diet (e.g. ad libitum): Pelleted standard Provimi Kliba 3418 rabbit maintenance diet ad Iibitum (batch no.33/04) provided by Provimi Kliba AG, CH-4303 Kaiseraugst.
- Water (e.g. ad libitum):Community tap water from Füllinsdorf, ad libitum.
- Acclimation period: 5 days. Under laboratory conditions after health examination. Only animals without any visual signs of illness were used for the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 17-23°C
- Humidity (%): 30-70 %
- Air changes (per hr): approximately 10-15 air changes per hour.
- Photoperiod (hrs dark / hrs light): The animals were provided with an automatically controlled light cycle of 12 hours light and 12 hours dark. Music was played during
the daytime light period.

IN-LIFE DATES: From:03-JUN-2004 To:18-JUN-2004

Test system

Vehicle:
unchanged (no vehicle)
Controls:
yes, concurrent no treatment
Amount / concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit):0.1 g
- Concentration (if solution): No data

VEHICLE (No vehicle)
- Amount(s) applied (volume or weight with unit): No data
- Concentration (if solution): No data
- Lot/batch no. (if required): No data
- Purity: No data
Duration of treatment / exposure:
24 hours after treatment.
Observation period (in vivo):
1, 24, 48 and 72 hours, as well as 7 and 10 days after instillation.
Number of animals or in vitro replicates:
3 (Animals of both sexes were used)
Details on study design:
REMOVAL OF TEST SUBSTANCE
- Washing (if done): The treated and untreated eyes were rinsed with lukewarm tap water.
- Time after start of exposure: 24 hours

SCORING SYSTEM: The ocular reaction was assessed according to the numerical scoring system listed in the EEC Commission Directive 92/69/EEC, July 31, 1992 at approximately 1, 24, 48 and 72 hours, as well as 7 and 10 days after instillation.

TOOL USED TO ASSESS SCORE: Eye examinations were made with a Varta Cliptrix diagnostic-Iamp (Roth AG, CH-4153 Reinach/Switzerland) .

Results and discussion

In vivo

Resultsopen allclose all
Irritation parameter:
cornea opacity score
Basis:
animal: 82,83,84
Time point:
24/48/72 h
Score:
0
Max. score:
3
Reversibility:
other: not applicable
Remarks on result:
no indication of irritation
Irritation parameter:
iris score
Basis:
animal: 82,83,84
Time point:
24/48/72 h
Score:
0
Max. score:
3
Reversibility:
other: not applicable
Remarks on result:
no indication of irritation
Irritation parameter:
conjunctivae score
Basis:
animal: 82,83
Time point:
24/48/72 h
Score:
1.67
Max. score:
3
Reversibility:
other: Effects were reversible and were no longer evident 10 days after treatment,the end of the observation period for all animals.
Remarks on result:
other: After 24 hours:moderately reddened; moderate swelling with partial eversion of lids. After 48 hours:moderately reddened; slight swelling. After 72 hours:slightly reddened; slight swelling
Irritation parameter:
conjunctivae score
Basis:
animal: 84
Time point:
24/48/72 h
Score:
1
Max. score:
3
Reversibility:
other: Effects were reversible and were no longer evident 10 days after treatment,the end of the observation period for all animals.
Remarks on result:
other: slightly reddened
Irritation parameter:
chemosis score
Basis:
animal: 82
Time point:
24/48/72 h
Score:
1.33
Max. score:
3
Reversibility:
other: Effects were reversible and were no longer evident 10 days after treatment,the end of the observation period for all animals.
Irritation parameter:
chemosis score
Basis:
animal: 83
Time point:
24/48/72 h
Score:
0.67
Max. score:
3
Reversibility:
other: Effects were reversible and were no longer evident 10 days after treatment,the end of the observation period for all animals.
Irritation parameter:
chemosis score
Basis:
animal: 84
Time point:
24/48/72 h
Score:
0
Max. score:
3
Reversibility:
other: not applicable
Remarks on result:
no indication of irritation
Irritation parameter:
cornea opacity score
Basis:
animal: 82,83,84
Time point:
7 d
Score:
0
Max. score:
3
Reversibility:
other: not applicable
Remarks on result:
no indication of irritation
Irritation parameter:
iris score
Basis:
animal: 82,83,84
Time point:
7 d
Score:
0
Max. score:
3
Reversibility:
other: not applicable
Remarks on result:
no indication of irritation
Irritation parameter:
conjunctivae score
Basis:
animal: 82,83
Time point:
7 d
Score:
1
Max. score:
3
Reversibility:
other: Effects were reversible and were no longer evident 10 days after treatment,the end of the observation period for all animals.
Remarks on result:
other:
Remarks:
slightly reddened
Irritation parameter:
conjunctivae score
Basis:
animal: 84
Time point:
7 d
Score:
0
Max. score:
3
Reversibility:
other: not applicable
Remarks on result:
no indication of irritation
Irritation parameter:
cornea opacity score
Basis:
animal: 82,83,84
Time point:
10 d
Score:
0
Max. score:
3
Reversibility:
other: not applicable
Remarks on result:
no indication of irritation
Irritation parameter:
iris score
Basis:
animal: 82,83,84
Time point:
10 d
Score:
0
Max. score:
3
Reversibility:
other: not applicable
Remarks on result:
no indication of irritation
Irritation parameter:
conjunctivae score
Basis:
animal: 82,83,84
Time point:
10 d
Score:
0
Max. score:
3
Reversibility:
other: not applicable
Remarks on result:
no indication of irritation
Irritation parameter:
chemosis score
Basis:
animal: 82,83,84
Time point:
10 d
Score:
0
Max. score:
3
Reversibility:
other: not applicable
Remarks on result:
no indication of irritation
Irritant / corrosive response data:
IRRITATION
The mean score was calculated across 3 scoring times (24, 48 and 72 hours after instillation) for each animal for corneal opacity, iris, redness and chemosis of the conjunctivae, separately. The individual mean scores for corneal opacity and iris were 0.00 for all three animals. The individual mean scores for the conjunctivae were 1 .67, 1 .67 and 1.00 for reddening and 1.33,0.67 and 0.00 for chemosis, respectively.
No abnormal findings were observed in the cornea or iris of any animal at any of the measurement intervals.
Slight to moderate reddening of the conjunctivae was noted in all animals from the 1-hour upto the 48-hour reading. Slight reddening was still present in all animals 72 hours after treatment and persisted in two animals up to the 7-day examination.
Moderate swelling (chemosis) of the conjunctivae with partial eversion of lids to marked swelling with half-closed lids was observed in all animals at the 1-hour reading. Moderate swelling with partial eversion of the lids was still visible in two animals 24 hours after treatment and slight swelling of the conjunctivae was present in one animal at the 48- and 72-hour examination.
The reddening of the sclerae was not assessable in two animals at the 1-hour reading due to swelling of the conjunctivae while moderate reddening was visible in the third animal. Slight to moderate reddening of the sclerae was present in all animals at the 24-hour examination and moderate reddening was observed in two animals 48 hours after treatment. Slight reddening was still evident in two animals at the 72-hour reading.
Discharge with moistening of the lids and hairs just adjacent to the lids was noted in all animals at the 1-hour examination. Slight discharge was still present in two animals 24 hours after treatment.
No abnormal findings were observed in the treated eye of any animal 10 days after treatment, the end of the observation per iod for all animals.

CORROSION
No corrosion of the cornea was observed at any of the reading times.
Other effects:
NECROPSY : No necropsy was performed on the animals sacrificed at termination of observation.
All rabbits were sacrificed by an intravenous injection of Vetanarcol into the ear vein at a dose of at least 1 mL/kg body weight (equivalent to 162 mg sodium pentobarbitone/kg body weight) and discarded.

VIABILlTY/MORTALITY AND CLINICAL SIGNS
No clinical signs of systemic toxicity were observed in the animals during the study and no mortality occurred.

COLORATION
No staining of the treated eyes produced by the test item was observed.

BODY WEIGHTS
The body weights of all rabbits were considered to be within the normal range of variability.
Body weight in grams
Animal No. Sex First Day of Acclimatization Day of Treatment Last Day of Observation
82 male 2591 2783 2950
83 female 3036 3277 3543
84 female 2720 2832 2999

Applicant's summary and conclusion

Interpretation of results:
other: Not irritating
Remarks:
based on CLP criteria
Conclusions:
Based on these results and according to the CLP classification criteria: the test item C 010 (3-[(4,5-dihydro-3-methyl-5-oxo-1-phenyl-1H-pyrazol-4-yl)azo]-N,N,N-trimethylanilinium chloride) is considered to be "not irritating" to the rabbit eye.

Executive summary:

The primary eye irritation potential of C 010 (3-[(4,5-dihydro-3-methyl-5-oxo-1-phenyl-1H-pyrazol-4-yl)azo]-N,N,N-trimethylanilinium chloride) was investigated according to OECD test guideline no. 405. The test item was applied by instillation of 0.1 g into the left eye of each of three young adult New Zealand White rabbits. The treated and untreated eyes were rinsed with lukewarm tap water 24 hours after instillation. Scoring of irritation effects was performed approximately 1, 24, 48 and 72 hours, as well as 7 and 10 days after test item instillation.

The mean score was calculated across 3 scoring times (24, 48 and 72 hours after instillation) for each animal for corneal opacity, iris, redness and chemosis of the conjunctivae, separately. The individual mean scores for corneal opacity and iris were 0.00 for all three animals. The individual mean scores for the conjunctivae were 1.67, 1.67 and 1.00 for reddening and 1.33, 0.67 and 0.00 for chemosis, respectively.

The instillation of C 010 (3-[(4,5-dihydro-3-methyl-5-oxo-1-phenyl-1H-pyrazol-4-yl)azo]-N,N,N-trimethyl anilinium chloride) into the eye resulted in mild to moderate, early-onset and transient ocular changes, such as reddening of the conjunctivae and sclerae, discharge and chemosis. These effects were reversible and were no longer evident 10 days after treatment, the end of the observation period for all animals. No abnormal findings were observed in the cornea or iris of any animal at any of the examinations. No corrosion was observed at any of the measuring intervals. No staining of the treated eyes by the test item was observed and no clinical signs were observed.

Thus, the test item did not induce significant or irreversible damage to the rabbit eye.

Based on these results and according to the CLP classification criteria: the test item C 010 (3-[(4,5-dihydro-3-methyl-5-oxo-1-phenyl-1H- pyrazol-4-yl)azo] -N,N,N-trimethylanilinium chloride) is considered to be "not irritating" to the rabbit eye.