Sodium 2-amino-4,6-dinitrophenoxide

Endpoint summary

Description of key information

Skin Irritation:

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 female Sprague Dawley rats under the experimental conditions tested and classified as “Category- Not Classified” as per CLP Classification.

Eye Irritation:

Mild conjunctival inflammation was observed in each animal. It can be considered that 2.5% aqueous test chemical solution (buffered to pH 7) was not an ocular irritant.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Justification for type of information:

data is from experimental reports

Qualifier:

according to guideline

Guideline:

other: Acute Dermal toxicity (OECD 402 Guidelines)

Principles of method if other than guideline:

To determine the dermal reaction profile of the test chemical in Sprague Dawley rats

GLP compliance:

yes

Species:

rat

Strain:

Sprague-Dawley

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: National Institute of Biosciences, Pune
- Sex: Female. Females were nulliparous and non-pregnant
- Age at study initiation: Young adult (8 to 10 weeks old) female rats
- Weight at study initiation: weight range of approximately 221.5 to 227.4 grams at initiation of dosing.
Body weights at the start :
Female
Mean : 225.23 g (= 100 %)
Minimum : 221.5 g (- 1.66 %)
Maximum : 227.4 g (+ 0.96 %)
Total No. of animals : 3
- 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, ad libitum
- Water (e.g. ad libitum): Water was provided ad libitum from individual bottles attached to the cages. All water was from a local source and passed through the reverse osmosis membrane before use.
- Acclimation period: All animals were acclimatized to laboratory conditions for minimum of 5 days.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): Room temperature was maintained at 19.0 to 21.8 degree centigrade
- Humidity (%): room humidity was maintained at 50.4% to 60.3%.
- Air changes (per hr): The animal room was independently provided with at least ten to fifteen air changes per hour of 100% fresh air that had been passed through the HEPA filters
- Photoperiod (hrs dark / hrs light): An artificial light and dark cycle of 12 hours each was provided to the room

IN-LIFE DATES: From: 29/05/2018 To: 15/06/2018

Type of coverage:

semiocclusive

Preparation of test site:

clipped

Vehicle:

unchanged (no vehicle)

Controls:

not specified

Amount / concentration applied:

Dose range finding study: 2000 mg/kg
Main Study: 2000 mg/kg

Duration of treatment / exposure:

24 hours exposure

Observation period:

Dermal reaction was observed daily for study period of 14 days.

Number of animals:

3 - [1- dose-range finding study;2 - main study]

Details on study design:

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.
REMOVAL OF TEST SUBSTANCE
- 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:
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 we
re 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 included general clinical signs, observations of eyes, mucous membranes, respiratory,
circulatory system and behavior pattern.
Evaluation of Dermal Reaction:
Dermal reaction was observed daily for study period of 14 days.
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. Macroscopic examination 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:

erythema score

Basis:

mean

Time point:

14 d

Score:

0

Reversibility:

not specified

Remarks on result:

no indication of irritation

Irritation parameter:

edema score

Basis:

mean

Time point:

14 d

Score:

0

Reversibility:

not specified

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

Evaluation of Dermal Reaction

Sex : Female
Dose Range Finding Study:
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.
Main Study:
Group II : Animals 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:

Clinical Signs of Toxicity and Mortality

Sex : Female
Dose Range Finding Study:
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. The animal survived through the study period of 14 days.
Main Study:
Group II : Animals 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 : Female
Dose Range Finding Study:
Group I (2000 mg/kg) - Percent body weight gain after 7 days and 14 days was found to be 8.35% and 15.85% respectively.
Main Study:
Group II (2000 mg/kg) - Percent body weight gain after 7 days and 14 days was found to be 5.88% and 12.57% respectively.

Gross Pathological Findings
Gross pathological examination did not reveal any abnormalities in animals from 2000 mg/kg dose group from dose range finding study and main study sacrificed terminally.

Summary of Evaluation of Dermal Reaction

 

Laboratory Test Item Code :TAS/122/084

Test System : Sprague Dawley Rat

Sex : Female

Finding Study:

Group No.	Dose mg/kg	Dermal Reaction	Total Number of Animals	Animal No.	Period of signs in days  From – to
I	2000	No dermal reaction observed	1	1	Day 0 - Day 14

 

Main Study:

Group No.	Dose mg/kg	Dermal Reaction	Total Number of Animals	Animal Nos.	Period of signs in days  From – to
II	2000	No dermal reaction observed	2	2, 3	Day 0 - Day 14

Individual Animal - Evaluation of Dermal Reaction

 

Laboratory Test Item Code :TAS/122/084

Test System : Sprague Dawley Rat

Sex : Female  

Finding Study:

Group : I                                                                    Dose  : 2000 mg/kg body weight

Animal	Dermal	D A Y S
No.	Reaction	0	1	2	3	4	5	6	7	8	9	10	11	12	13	14
1	Erythema	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	Oedema	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0

 

Main Study:

Group : II                                                                   Dose  : 2000 mg/kg body weight

Animal	Dermal	D A Y S
No.	Reaction	0	1	2	3	4	5	6	7	8	9	10	11	12	13	14
2	Erythema	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	Oedema	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
3	Erythema	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
	Oedema	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0

Interpretation of results:

other: not irritating

Conclusions:

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 not irritating to the skin of Sprague Dawley rats under the experimental conditions tested and classified as “Category-Not Classified” as per CLP Classification.

Executive summary:

A 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. 5 female young adult, non-pregnant rats were used for the 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.

\ A single dose of 2000 mg/kg body weight of the test item was applied to 1 female animal respectively in the dose range finding study. Since no signs of irritation were observed at the maximum dose of 2000 mg/kg in the test animals, 2 additional animals were tested in the main study with 2000 mg/kg. The animals were applied with a dose of 2000 mg/kg body weight in sequential manner at 48 hours intervals. Dermal reaction was observed daily for study period of 14 days.

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 female Sprague Dawley rats under the experimental conditions tested and classified as “Category- Not Classified” as per CLP Classification.

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:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

data is from peer reviewed journals

Qualifier:

according to guideline

Guideline:

other: as mentioned below

Principles of method if other than guideline:

To assess the eye irritation potential of the test chemical in albino rabbits

GLP compliance:

not specified

Species:

rabbit

Strain:

not specified

Details on test animals or tissues and environmental conditions:

no data available

Vehicle:

other: 2.5% aqueous test chemical (buffered to pH 7)

Controls:

yes, concurrent no treatment

Amount / concentration applied:

0.1ml of 2.5% aqueous test chemical (buffered to pH 7)

Duration of treatment / exposure:

the lids were held together for 1 second

Observation period (in vivo):

24, 48, and 72 hours

Duration of post- treatment incubation (in vitro):

no data available

Number of animals or in vitro replicates:

3

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing (if done): yes
- Time after start of exposure: At 10 seconds post-instillation, the eyes were rinsed with 20 mL of distilled water

Observation period:Reactions were scored at 24, 48, and 72 hours post-instillation.

TOOL USED TO ASSESS SCORE: hand-slit lamp / biomicroscope / fluorescein: no data available

Irritation parameter:

overall irritation score

Basis:

mean

Time point:

24/48/72 h

Reversibility:

fully reversible within: 72 hours

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

Mild conjunctival inflammation was observed in each animal.

Interpretation of results:

other: not irritating

Conclusions:

Mild conjunctival inflammation was observed in each animal. It can be considered that 2.5% aqueous test chemical solution (buffered to pH 7) was not an ocular irritant

Executive summary:

The eye irritation potential of the test chemical was assessed in albino rabbits.

0.1 ml of 2.5% aqueous test chemical (buffered to pH 7) was instilled into the eyes of albino rabbits (N = 3) and the lids were held together for 1 second. At 10 seconds post-instillation, the eyes were rinsed with 20 mL of distilled water. Untreated eyes served as controls. Reactions were scored at 24, 48, and 72 hours post-instillation.

Mild conjunctival inflammation was observed in each animal. It can be considered that 2.5% aqueous test chemical solution (buffered to pH 7) was not an ocular irritant

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin Irritation

Various studies have been summarized to determine the degree of dermal irritation caused by the test chemical in living organisms. These results include in vivo experimental studies on rats as well as rabbits for the test chemicals.

A 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. 5 female young adult, non-pregnant rats were used for the 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.

A single dose of 2000 mg/kg body weight of the test item was applied to 1 female animal respectively in the dose range finding study. Since no signs of irritation were observed at the maximum dose of 2000 mg/kg in the test animals, 2 additional animals were tested in the main study with 2000 mg/kg. The animals were applied with a dose of 2000 mg/kg body weight in sequential manner at 48 hours intervals. Dermal reaction was observed daily for study period of 14 days.

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 female Sprague Dawley rats under the experimental conditions tested and classified as “Category- Not Classified” as per CLP Classification.

This is supported by the results of a similar study performed as per Consumer Product Safety Commission of the USA (Code of Federal Regulations, Title 16, Sec. 1500.41) to evaluate the irritation potential of the test chemical.2.5% (w/v) Suspension of the test chemical in 0.5% aqueous gum tragacanth, containing 0.05% Na2SO3was applied to the intact and abraded skin of 3 New Zealand White albino rabbits. The rabbits were observed for signs of irritation till 72 hours and the reactions were scored based on the definition of skin irritation given in the  (Code of Federal Regulations, Title 16, Sec.1500.41).

None of the rabbits treated with the test chemical showed any signs of irritation during the 72 hours observation period. The Primary Irritation Index was 0.0.

Based on the scores and observations, the test chemical was considered to be not irritating to albino New Zealand White rabbit skin.

The above results are further supported by a similar study performed as per Consumer Product Safety Commission of the USA (Code of Federal Regulations, Title 16, Sec. 1500.41) to determine the irritation potential of the test chemical.2.5% (w/v) Suspension of the test chemical in 0.5% aqueous gum tragacanth, containing 0.05% Na2SO3was applied to the intact and abraded skin of 3 New Zealand White albino rabbits. The rabbits were observed for signs of irritation till 72 hours and the reactions were scored based on the definition of skin irritation given in the Code of Federal Regulations, Title 16, Sec.1500.41.

None of the rabbits treated with the test chemical showed any signs of irritation during the 72 hours observation period. The Primary Irritation Index was 0.0.

Based on the scores and observations, the test chemical was considered to be not irritating to albino New Zealand White rabbit skin.

Considering the results of the in vivo studies, the test chemical can be considered to be not irritating to skin. Comparing the above annotations with the criteria of CLP Regulation, the test chemical can be classified under the category “Not Classified”.

Eye Irritation

Various studies have been summarized to determine the extent of ocular damage caused by the test chemical in living organisms. These results include in vivo experimental studies on rabbits for the test chemicals.

The eye irritation potential of the test chemical was assessed in albino rabbits.

0.1 ml of 2.5% aqueous test chemical (buffered to pH 7) was instilled into the eyes of albino rabbits (N = 3) and the lids were held together for 1 second. At 10 seconds post-instillation, the eyes were rinsed with 20 mL of distilled water. Untreated eyes served as controls. Reactions were scored at 24, 48, and 72 hours post-instillation.

Mild conjunctival inflammation was observed in each animal. It can be considered that 2.5% aqueous test chemical solution (buffered to pH 7) was not an ocular irritant.

This is supported by a study performed according to the method prescribed in the Code of Federal Regulations (Title 16, Sec. 1500.42) to assess the ocular irritation of the test chemical. The test chemical was prepared as a 2.5% (w/v) solution or suspension. (Suspension in 0.5% aqueous gum tragacanth, containing 0.05% Na2SO3). Eleven groups of three rabbits were used to test solutions, each of which was instilled into one eye of each of three rabbits. The eyes of all the rabbits were irrigated with 50 ml of lukewarm (37°C) water, 10 sec after instillation of the test material.

Treatment with the test chemical gave rise to a yellow coloration of the cornea 1 hr after instillation, but the cornea was clear by the 24-hr reading, and there was no observable irritant response in any of the animals in this group.

According to the Code of Federal Regulations, Test for Eye Irritants, the test chemical was scored as ± .

Based on the scores and observations, the test chemical can be considered to be not irritating to eyes.

In another study conducted according the procedure based on that prescribed in the Code of Federal Regulations (Title 16, Sec. 1500.42), the irritation potential of the test chemical was assessed in rabbits. The test chemical was prepared as a 2.5% (w/v) solution or suspension. (Suspension in 0.5% aqueous gum tragacanth, containing 0.05% Na2SO3). Eleven groups of three rabbits were used to test solutions, each of which was instilled into one eye of each of three rabbits. The eyes of all the rabbits were irrigated with 50 ml of lukewarm (37°C) water, 10 sec after instillation of the test material.

None of the three animals treated with the test material showed any response to treatment, and although occasional transient mild conjunctival inflammation was observed. The reactions did not persist for more than 24 hr.

Thus, the substance was considered to be non irritating to eyes.

Considering the results of the in vivo studies, the test chemical can be considered to be not irritating to eyes. Comparing the above annotations with the criteria of CLP Regulation, the test chemical can be classified under the category “Not Classified”.

Justification for classification or non-classification

Available studies for the test chemical indicate that the test chemical lacks the potential to cause any irritation to eyes and skin. Hence, the test chemical can be considered to be not irritating to eyes and skin. It can further be classified under the category "Not Classified” as per CLP regulation.