9-(2-carboxyphenyl)-3,6-bis(diethylamino)xanthylium acetate

Endpoint summary

Description of key information

Skin Irritation:

The dermal irritation potential of test chemical was assessed in various experimental studies conducted on rabbits. Based on the available data for the key and supporting studies, it can be concluded that the test chemical is unable to cause skin irritation and thus considered as not irritating. Comparing the above annotations with the criteria of CLP regulation, it can be classified under the category “Not Classified”.

Eye irritation:

An ocular irritation potential of target chemical was assessed in various experimental studies conducted on test animals. Based on the available data for the key and supporting studies, it can be concluded that the test chemical is able to cause severe eye damage and thus can be considered as irritating. Comparing the above annotations with the criteria of CLP regulation, it can be classified under the category “Category 2”.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vivo

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Remarks:

experimental data from various test chemicals

Justification for type of information:

Data is summarized based on the available information from various read across test chemicals.

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

other: As mentioned below

Principles of method if other than guideline:

WoE report is based on 2 skin irritation studies as - WoE 2 and WoE 3
Skin irritation test was carried out to study the effects of the test chemicals on rodents.

GLP compliance:

not specified

Species:

other: 2. rabbit 3.rat

Strain:

other: 2. New Zealand White 3.Sprague-Dawley

Details on test animals or test system and environmental conditions:

2.TEST ANIMALS
Source: No data
Age at study initiation: No data
Weight at study initiation: 3 kg
Fasting period before study: No data
Housing: singly housed, in suspended cages
Diet (e.g. ad libitum): Purina Rabbit Chow, ad libitum
Water (e.g. ad libitum): water, ad libitum
Acclimation period: No data
ENVIRONMENTAL CONDITIONS
Temperature (°C): No data
Humidity (%):No data
Air changes (per hr): No data
Photoperiod (hrs dark / hrs light): 12 hour light and 12 hr dark cycle

IN-LIFE DATES: From: No data To: No data

3.Source: National Institute of Biosciences, Pune.
- Females nulliparous and non-pregnant: No data available
- Age at study initiation: Young adult male and female rats aged between 8 – 12 weeks were used.
- Weight at study initiation: The weight range of approximately 216.9 to 251.9 grams at initiation of dosing.
Male
Mean : 246.28 g (= 100 %)
Minimum : 240.9 g (- 2.18 %)
Maximum : 254.3 g (+ 3.26 %)
Total No. of animals : 5
Female
Mean : 219.02 g (= 100 %)
Minimum : 215.0 g (- 1.84 %)
Maximum : 223.4 g (+ 2.00 %)
Total No. of animals : 5

- Identification: Each rat was individually identified by the cage number.
- Fasting period before study: No data available
- Housing: The rats were individually housed in polycarbonate cages with paddy husk as bedding.
- Diet (e.g. ad libitum): Rodent feed supplied by the Nutrivet Life Sciences, Pune, was provided ad libitum
from individual feeders.
- Water (e.g. ad libitum): Water was provided ad libitum from individual bottles attached to the cages. All water was from a local source

ENVIRONMENTAL CONDITIONS

- Temperature (°C): 19.6 to 21.6 degree centigrade
- Humidity (%): 55.0% to 58.4%
- Air changes (per hr): Ten to fifteen air changes per hour.
- Photoperiod (hrs dark / hrs light): An artificial light and dark cycle of 12 hours each was provided to the
room.

IN-LIFE DATES:
Experimental Starting Date : 12-06-2017
Experimental Completion Date : 27-06-2017

Type of coverage:

other: 2.Entire inner surface of one ear 3.semiocclusive

Preparation of test site:

other: 2. No data 3.clipped

Vehicle:

other: 2.propylene glycol 3.unchanged (no vehicle)

Controls:

yes

Amount / concentration applied:

2.Dose of 1ml of the test material was applied once daily for five days per week for 2 weeks.
3.2000 mg/kg

Duration of treatment / exposure:

2.once daily, Five days per week for two weeks.
3.24 hours

Observation period:

2.2 weeks
3.14 days

Number of animals:

2.3 rabbits
3.5/sex

Details on study design:

2.TEST SITE
Area of exposure: Ear
% coverage: No data
Type of wrap if used: No data

REMOVAL OF TEST SUBSTANCE
Washing (if done): No data
Time after start of exposure: No data

SCORING SYSTEM: The irritancy produced by repeated application of a chemical or skin care product on the surface epidermis in the rabbit ear is evaluated on a scale of 0 to 5. The grades are summarized as follows:
0 = No irritation
1 = few scales, no Erythema
2 = diffuse scaling, no Erythema
3 = Generalized scaling with Erythema
4 = Scaling, Erythema and Edema
5 = Epidermal necrosis and slough

3.TEST SITE
- Area of exposure: Trunk (dorsal surface and sides from scapular to pelvic area)
- % coverage: Approximately 10% of the body surface area.
- Type of wrap if used: Porous gauze dressing and non-irritating tape.
- Washing (if done): Distilled water was used to remove residual test item.
OBSERVATION TIME POINTS
(indicate if minutes, hours or days) : Dermal reaction was observed daily for study period of 14 days.
SCORING SYSTEM: Draize Method.
OTHER OBSERVATIONS
Type and Frequency of Tests, Analyses and Measurements
Viability: Twice daily.
Clinical Observations and General Appearance:
Animals were observed for clinical signs, mortality, until sacrifice.
Onset, duration and severity of any sign were recorded. The clinical signs and mortality observations w
ere conducted at 10, 30, 60 minutes, 2, 4 and 6 hours on the day of dosing and once daily thereafter for
14 day. Daily observation was done as far as possible at the same time.
The observations were included general clinical signs, observations of eyes, mucous membranes, respira
tory, circulatory system and behavior pattern.
Body weights:
Individual animal body weights were recorded pre-test (prior to administration of the test item), day 7 and
at termination on day 14.
Gross Pathology:
Necropsy was performed on animals surviving at the end of the study. Macroscopicexamination of all the
orifices, cavities and tissues were made and the findings were recorded. All animals surviving the study
period were sacrificed by the carbon dioxide asphyxiation technique (day 15).
Histopathology:
No gross abnormalities were observed in animals sacrificed terminally hence, no histopathology was performed.

Irritation parameter:

overall irritation score

Remarks:

2

Basis:

mean

Time point:

14 d

Score:

0

Max. score:

5

Reversibility:

not specified

Remarks on result:

no indication of irritation

Irritation parameter:

erythema score

Remarks:

3

Basis:

mean

Time point:

14 d

Score:

0

Reversibility:

not specified

Remarks on result:

no indication of irritation

Irritation parameter:

edema score

Remarks:

3

Basis:

mean

Time point:

14 d

Score:

0

Reversibility:

not specified

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

2.No erythema observed
3.Sex : Male
Group I -
Animal treated at the dose level of 2000 mg/kg body weight did not result in any skin reaction during the s
tudy period of 14 days.
Sex : Female
Group I -
Animal treated at the dose level of 2000 mg/kg body weight did not result in any skin reaction during the
study period of 14 days.
Other effects
Other effects:
Clinical Signs of Toxicity and Mortality
Sex : Male
Group I -
Animal treated at the dose level of 2000 mg/kg body weight did not result in any signs of toxicity during the study period of 14 days. All animals survived through the study period of 14 days.
Sex : Female
Group I -
Animal treated at the dose level of 2000 mg/kg body weight did not result in any signs of toxicity during the study period of 14 days. All animals survived through the study period of 14 days.
Body Weight
Sex : Male
Group I (2000 mg/kg) - Percent body weight gain after 7 days and 14 days was found to be 9.30% and
18.14% respectively.
Sex : Female
Group I (2000 mg/kg) - Percent body weight gain after 7 days and 14 days was found to be 4.60% and 9.06% respectively.
Gross Pathological Findings
Gross pathological examination did not reveal any abnormalities in animals from 2000 mg/kg dose group.

Interpretation of results:

other: Not irritating

Conclusions:

The test chemical was considered to be not irritating to the skin of rabbits and rats.

Executive summary:

Various studies were performed on rabbits and rats to assess the dermal irritation potential of test chemical which have been summarized as follows:

The test uses a single rabbit ear to indicate the Comedogenicity and irritancy of the test chemical. The test chemical was mixed in propylene glycol at a 9 to 1 dilution for testing unless otherwise indicated (10% concentration). A colony of New Zealand albino rabbits that have genetically good ears and free from mites were used. Three rabbits, weighing two to three kilograms, were used for each assay. Animals were housed singly in suspended cages and fed Purina Rabbit Chow and water ad libitum. Animals were maintained on a 12-hour light and 12-hour dark cycle. A dose of 1 ml of the test material was applied and spread once daily to the entire inner surface of once for five days per week for two weeks. The opposite untreated ear of each animal served as an untreated control.

The test chemical falls under Grade 0 (no irritation observed). Hence it can be concluded that the test chemical was not irritating to rabbit ears.

Another study was designed and conducted to determine the dermal reaction profile of the test chemical in Sprague Dawley rats. The study was performed as per OECD Guidelines 402 and complying to the GLP procedures. Ten rats (5 male and 5 female) were used for conducting dermal irritation /corrosion study.

The animals were kept in their cages for at least 5 days prior to administration for acclimatization to the laboratory condition and after acclimatization period, animals were randomly selected. Approximately 24 hours before application, the hair of each rat was closely clipped from the trunk (dorsal surface and sides from scapular to pelvic area) with an electric clipper, so as to expose at least 10% of the body surface area. The test item was applied directly onto the exposed skin of the animal, taking care to spread the test item evenly over the entire area of approximately 10% of the total body surface area or as much of the area as can reasonably be covered. The test item was held in contact with the skin using a porous gauze dressing and non irritating tape around the animal to cover the exposure site for first 24 hours exposure period. Elizabethan collar was placed on each animal for first 24 hours after application of the test item. These collars prevent ingestion of test item. Following 24 hours of exposure, the wrapping was removed and the test site wiped free of excess test item. Distilled water was used to remove residual test item.The test item was applied to shorn skin of 5 male and 5 female animals at 2000 mg/kg body weight. Administration of the test item at 2000 mg/kg did not result in any skin reaction at the site of application during the study period of 14 days. Administration of the test item did not result in any signs of toxicity and mortality during the study period of 14 days. Animals exhibited normal body weight gain through the study period of 14 days. Gross pathological examination did not reveal any abnormalities attributable to the treatment. The overall irritation score of the substance was determined to be 0 and no erythema and edema (skin irritation) were found at the end of 14 days observation period after patch removal. Hence, it was concluded that the test chemical was Non-Irritating to the skin of Sprague Dawley rats under the experimental conditions tested and classified as “Category- Not Classified” as per CLP Classification.

All these studies lead to a conclusion that Test chemical is indeed not irritating to skin. Hence, comparing the above annotations with the criteria of CLP regulation, Test chemical can be classified under the category “Not Classified”.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vivo

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Remarks:

experimental data from various test chemicals

Justification for type of information:

Data is summarized based on the available information from various read across test chemicals.

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

other: As mentioned below

Principles of method if other than guideline:

WoE report is based on 3 eye irritation studies as - WoE 3, WoE 4 and WoE 5.
Eye irritation test was carried out to study the effects of the test chemicals on rodents.

GLP compliance:

not specified

Species:

other: 3.human 4.rabbit 5.rabbit

Strain:

other: 3.not specified 4. Russian breed 5.not specified

Details on test animals or tissues and environmental conditions:

3.Age at study initiation: 16 males of mean age 39 years (25-50) and one female 40 year old.
4.TEST ANIMALS
Sex: Male and females
- Source: obtained from the breeder
- Age at study initiation: no data available
- Weight at study initiation: 1.5 – 1.8 kg
- Fasting period before study: no data available
- Housing: suspended metal cages (47*32*34 cm)
- Diet (e.g. ad libitum): standard diet of Nafag, ad libitum
- Water (e.g. ad libitum): drinking water, ad libitum
- Acclimation period: the rabbits were acclimatized to the lab environment (duration not specified)


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18±1 deg C
- Humidity (%): 55±5%
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 14 hours light/ day

5.not specified

Vehicle:

other: 3.not specified 4.unchanged (no vehicle) 5.unchanged (no vehicle)

Controls:

yes, concurrent no treatment

other: 3. No 4.Right eye served as control

Amount / concentration applied:

3. (10 LBS) 4535.92 mg
4. 0.1 g
5. undiluted

Duration of treatment / exposure:

3.26 min
4.1 minute for rinsed eyes , not specified incased of unrinsed eyes
5.single exposure

Observation period (in vivo):

3.24hrs
4.1,6 hours and 1,2 ,3, 6 ,8 days
5.24,48, 72 hours till 8 days

Duration of post- treatment incubation (in vitro):

no data available

Number of animals or in vitro replicates:

3.17 individuals
4.6 (3 males and 3 females)
5. 6

Details on study design:

3.REMOVAL OF TEST SUBSTANCE
- Washing :
Eyes were rinsed with physiological saline.
- Time after start of exposure:
1 minute after application.

SCORING SYSTEM:
Not specified

TOOL USED TO ASSESS SCORE: hand-slit lamp
4.TEST SITE
- Area of exposure: conjunctival sac

REMOVAL OF TEST SUBSTANCE
- Washing (if done): yes, the eyes of 3 animals were washed
- Time after start of exposure: 1 minute after exposure

TOOL USED TO ASSESS SCORE: hand-slit lamp / biomicroscope / fluorescein: hand-slit lamp
SCORING SYSTEM: Grading Scale for Eye Irritation:
Cornea Grade
Opacity: degree of density (readings should be taken from most dense area)*
No ulceration or opacity = 0
Scattered or diffuse areas of opacity (other than slight dulling of normal lustre); details of iris slightly obscured = 1
Easily discernible translucent area; details of iris slightly obscured = 2
Opalescent area; no details of iris visible; size of pupil barely discernible = 3
Opaque cornea; iris not discernible through the opacity = 4
Iris
Normal = 0
Markedly deepened rugae, congestion, swelling, moderate circumcorneal hyperaemia; or injection; iris reactive to light (a sluggish reaction is considered to be an effect = 1
Hemorrhage, gross destruction, or no reaction to light = 2
Maximum possible: 2
Conjunctivae
Redness (refers to palpebral and bulbar conjunctivae; excluding cornea and iris)
Vessels Normal = 0
Vessels definitely injected above normal = 1
Diffuse, crimson colour; individual vessels not easily discernible = 2
Diffuse beefy red = 3
Chemosis
Swelling (refers to lids and/or nictating membranes)
Normal = 0
Any swelling above normal (includes nictating membrane)= 1
Obvious swelling, with partial eversion of lids = 2
Swelling, with lids about half closed = 3
Swelling, with lids more than half closed= 4

Overall Maximum Possible Score = 110

The overall irritation index
0 – 10 – minimal
10 – 25 – slight
26 – 55 – moderate
56 -65 – marked
86 – 110 - extreme

5. SCORING SYSTEM: Draize method

Irritation parameter:

overall irritation score

Remarks:

3

Basis:

mean

Time point:

24 h

Reversibility:

fully reversible

Remarks on result:

positive indication of irritation

Irritation parameter:

overall irritation score

Remarks:

4

Basis:

mean

Time point:

72 h

Score:

74.2

Max. score:

110

Reversibility:

not specified

Remarks on result:

positive indication of irritation

Irritation parameter:

overall irritation score

Remarks:

4

Basis:

mean

Time point:

72 h

Score:

72

Max. score:

110

Reversibility:

not specified

Remarks on result:

positive indication of irritation

Irritation parameter:

overall irritation score

Remarks:

5

Basis:

mean

Time point:

72 h

Score:

6.56

Max. score:

110

Reversibility:

not specified

Remarks on result:

positive indication of irritation

Irritant / corrosive response data:

3.Burning of the eyes (82%), excessive tearing (47%), nasal burning (41%), nasal itching (35%), chest pain/tightness (35%), rhinorhea (29%), cough (29%), dyspnea (29%), burning of the throat (24%). burning/ pruritic skin (24%), chest burning (12%), headache (6%), and nausea (6%).
4.The mean irritation index for unrinsed eyes was 74.2/110 ; and 72/110 for rinsed eyes.
Rinsing the eyes following instillation was of no effect.

5.Primary Eye Irritation Score = 6.56*
*-no reading was possible due to substance-induced staining of the cornea or iris.

Interpretation of results:

Category 2 (irritating to eyes) based on GHS criteria

Conclusions:

The Test chemical was considered to be irritating to the eyes of treated test animals.

Executive summary:

Various studies were performed on test animals to assess the ocular irritation potential of test chemical which have been summarized as follows:

The first study reports the evaluation of 17 patients who were exposed to aerosolized test chemical. Approximately 10 lbs =(4.53kg) 4535.92 mg of test chemical powder was spilled onto the floor of a vehicle maintenance shop. A cleanup crew began to sweep up the powder from the floor . This resulted in an aerosol of test chemical dust that dispersed throughout the building contaminating the hair , skin, eyes, and clothing of most of the shop employees. Sixteen of the patients (94%) complained of symptoms such as redness, burning sensation..etc. One patient (6%) denied any symptoms despite being exposed to the dust for 15 minutes without a surgical mask. The most frequently reported symptom was burning o f the eyes which occurred in 82% of the patients . Four of the patients (24%) were placed on humidified oxygen because of respiratory symptoms. Five of the patients (29%) underwent irrigation of their eyes secondary to eye burning. After being discharged from the hospital, follow up was maintained. Ten of the patients (59%) reported that their symptoms had resolved within 4 hours of their exposure to the dust. One patient (6%) reported that his symptoms lasted 12 hours and 2 patients (12%) reported their symptoms lasted 24 hours. Three of the patients (18%) could not remember the duration of their symptoms. Burning of eyes was observed in 82% of the patients. Based on these effects test chemical may be considered as irritating to eyes.

In another study, the irritation potential of test chemical was evaluated. The test chemical was tested on 6 rabbits (3 males, 3 females) of the Russian breed (obtained from the breeder, weighing 1.5-1.8 kg). Rabbits were housed separately in metal cages and fed with standard diet of Nafag and drinking water ad libitum. Only the rabbits which showed normal ophthtalmic findings were used for the study. 0.1 g of test chemical was instilled into the conjunctival sac of the left eye of the rabbits. After application, the eyelids were held open for few seconds. The right eye served as control. The eyes of 3 animals were rinsed 1 minute after exposure with physiological saline. The reactions were assessed by hand slit lamp after 1,6 hours and 1,2,3,6,8 days according to the scoring pattern. The maximum possible score was 110. Under the conditions of the study, test chemical caused marked irritation when applied to the rabbit eyes. The mean irritation index for unrinsed eyes was 74.2/110 ; and 72/110 for rinsed eyes. Rinsing the eyes following instillation was of no effect. Hence, test chemical can be considered as an eye irritant.

Another study was performed to assess the eye irritation potential of the test chemical in rabbits. The study was performed according to the procedures outlined in Federal Register 38 No 187 § 1500 .42 (Eye), 5. 27019 of 27.09.73. Undiluted test chemical was instilled in the right eye of 6 rabbits, the untreated eye served as control. The treated eyes were observed and scored for ocular lesions at 24,48 and 72 hours. The eyes were scored according to the mehtod of Draize and the maximum attainable score was 110. The Primary Eye Irritation Score was calculated to be 6.56.The slight signs of irritation to the mucous membrane predominantly regressed within 8 days, but not the coloring of the eyeball. Because of these findings, the substance must be described as slightly irritating to eyes.

The results obtained from these studies lead to a conclusion that Test chemical is indeed irritating to eye. Hence, comparing the above annotations with the criteria of CLP regulation, Test chemical can be classified under the category “ Category 2 (irritating to eyes) based on GHS criteria”.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin Irritation:

Various studies were performed on rabbits and rats to assess the dermal irritation potential of test chemical which have been summarized as follows:

The test uses a single rabbit ear to indicate the Comedogenicity and irritancy of the test chemical. The test chemical was mixed in propylene glycol at a 9 to 1 dilution for testing unless otherwise indicated (10% concentration). A colony of New Zealand albino rabbits that have genetically good ears and free from mites were used. Three rabbits, weighing two to three kilograms, were used for each assay. Animals were housed singly in suspended cages and fed Purina Rabbit Chow and water ad libitum. Animals were maintained on a 12-hour light and 12-hour dark cycle. A dose of 1 ml of the test material was applied and spread once daily to the entire inner surface of once for five days per week for two weeks. The opposite untreated ear of each animal served as an untreated control.

The test chemical falls under Grade 0 (no irritation observed). Hence it can be concluded that the test chemical was not irritating to rabbit ears.

Another study was designed and conducted to determine the dermal reaction profile of the test chemical in Sprague Dawley rats. The study was performed as per OECD Guidelines 402 and complying to the GLP procedures. Ten rats (5 male and 5 female) were used for conducting dermal irritation /corrosion study. The animals were kept in their cages for at least 5 days prior to administration for acclimatization to the laboratory condition and after acclimatization period, animals were randomly selected. Approximately 24 hours before application, the hair of each rat was closely clipped from the trunk (dorsal surface and sides from scapular to pelvic area) with an electric clipper, so as to expose at least 10% of the body surface area. The test item was applied directly onto the exposed skin of the animal, taking care to spread the test item evenly over the entire area of approximately 10% of the total body surface area or as much of the area as can reasonably be covered. The test item was held in contact with the skin using a porous gauze dressing and non irritating tape around the animal to cover the exposure site for first 24 hours exposure period. Elizabethan collar was placed on each animal for first 24 hours after application of the test item. These collars prevent ingestion of test item. Following 24 hours of exposure, the wrapping was removed and the test site wiped free of excess test item. Distilled water was used to remove residual test item.The test item was applied to shorn skin of 5 male and 5 female animals at 2000 mg/kg body weight. Administration of the test item at 2000 mg/kg did not result in any skin reaction at the site of application during the study period of 14 days. Administration of the test item did not result in any signs of toxicity and mortality during the study period of 14 days. Animals exhibited normal body weight gain through the study period of 14 days. Gross pathological examination did not reveal any abnormalities attributable to the treatment. The overall irritation score of the substance was determined to be 0 and no erythema and edema (skin irritation) were found at the end of 14 days observation period after patch removal. Hence, it was concluded that the test chemical was Non-Irritating to the skin of Sprague Dawley rats under the experimental conditions tested and classified as “Category- Not Classified” as per CLP Classification.

All these studies lead to a conclusion that Test chemical is indeed not irritating to skin. Hence, comparing the above annotations with the criteria of CLP regulation, Test chemical can be classified under the category “Not Classified”.

Eye Irritation (In-vitro):

The ocular irritation potential of test article was determined according to the OECD 492 test guideline for this study. The MatTek EpiOcular™ model was used to assess the potential ocular irritation of the test articles by determining the viability of the tissues following exposure to the test article via MTT. Tissues were exposed to solid test articles and control for approx.6 hours, followed by a 25 minute post-soak and approximately 18 hours recovery after the post-soak. The viability of each tissue was determined by MTT assay.

The MTT data show the assay quality controls were met, passing the acceptance criteria.

The mean % tissue viability of test substance was determined to be 121.7%. Hence, under the experimental test conditions it was concluded that test substance was considered to be not irritating to the human eyes and can thus be classified as "Not Classified" as per CLP Regulation.

Eye Irritation (In-vivo):

Various studies were performed on test animals to assess the ocular irritation potential of test chemical which have been summarized as follows:

The first study reports the evaluation of 17 patients who were exposed to aerosolized test chemical. Approximately 10 lbs =(4.53kg) 4530000mg of test chemical powder was spilled onto the floor of a vehicle maintenance shop. A cleanup crew began to sweep up the powder from the floor . This resulted in an aerosol of test chemical dust that dispersed throughout the building contaminating the hair , skin, eyes, and clothing of most of the shop employees. Sixteen of the patients (94%) complained of symptoms such as redness, burning sensation..etc. One patient (6%) denied any symptoms despite being exposed to the dust for 15 minutes without a surgical mask. The most frequently reported symptom was burning o f the eyes which occurred in 82% of the patients . Four of the patients (24%) were placed on humidified oxygen because of respiratory symptoms. Five of the patients (29%) underwent irrigation of their eyes secondary to eye burning. After being discharged from the hospital, follow up was maintained. Ten of the patients (59%) reported that their symptoms had resolved within 4 hours of their exposure to the dust. One patient (6%) reported that his symptoms lasted 12 hours and 2 patients (12%) reported their symptoms lasted 24 hours. Three of the patients (18%) could not remember the duration of their symptoms. Burning of eyes was observed in 82% of the patients. Based on these effects test chemical may be considered as irritating to eyes.

In another study, the irritation potential of test chemical was evaluated. The test chemical was tested on 6 rabbits (3 males, 3 females) of the Russian breed (obtained from the breeder, weighing 1.5-1.8 kg). Rabbits were housed separately in metal cages and fed with standard diet of Nafag and drinking water ad libitum. Only the rabbits which showed normal ophthtalmic findings were used for the study. 0.1 g of test chemical was instilled into the conjunctival sac of the left eye of the rabbits. After application, the eyelids were held open for few seconds. The right eye served as control. The eyes of 3 animals were rinsed 1 minute after exposure with physiological saline. The reactions were assessed by hand slit lamp after 1,6 hours and 1,2,3,6,8 days according to the scoring pattern. The maximum possible score was 110. Under the conditions of the study, test chemical caused marked irritation when applied to the rabbit eyes. The mean irritation index for unrinsed eyes was 74.2/110 ; and 72/110 for rinsed eyes. Rinsing the eyes following instillation was of no effect. Hence, test chemical can be considered as an eye irritant.

Another study was performed to assess the eye irritation potential of the test chemical in rabbits. The study was performed according to the procedures outlined in Federal Register 38 No 187 § 1500 .42 (Eye), 5. 27019 of 27.09.73. Undiluted test chemical was instilled in the right eye of 6 rabbits, the untreated eye served as control. The treated eyes were observed and scored for ocular lesions at 24,48 and 72 hours. The eyes were scored according to the mehtod of Draize and the maximum attainable score was 110. The Primary Eye Irritation Score was calculated to be 6.56.The slight signs of irritation to the mucous membrane predominantly regressed within 8 days, but not the coloring of the eyeball. Because of these findings, the substance must be described as slightly irritating to eyes.

Though the in vitro results for the target indicates that test chemical can be not irritating to eye, but the estimated in-vivo results imply that test chemical is irritating to eye. These results are aptly supported by the results of rabbit study for the rtest chemical.

The results obtained from these studies lead to a conclusion that Test chemical is indeed irritating to eye. Hence, comparing the above annotations with the criteria of CLP regulation, Test chemical can be classified under the category “ Category 2 (irritating to eyes) based on GHS criteria”.

Justification for classification or non-classification

The skin and eye irritation potential of test chemical were observed in various studies. The results obtained from these studies indicates that the chemical is unlikely to cause skin irritation but can cause severe eye damage. Hence the test chemical can be classified under the category “Not Classified” for skin and “Category 2” for eye as per CLP.