Reaction products of diazotised sodium 2-amino-5-{[2-(sulfonatooxy)ethyl]sulfonyl}benzenesulfonate coupled with sodium 2-(4-aminobenzene-1-sulfonyl)ethyl sulfate, subsequently coupled with diazotised sodium 2-(4-aminobenzene-1-sulfonyl)ethyl sulfate

Endpoint summary

Description of key information

Acute oral toxicity:

1. In acute toxicity study by oral route the LD50 value for the Brilliant Black PN was found to be >2000 mg/kg for mice.

2. The lethal concentration value (LD50) for acute oral toxicity test was considered to be >2000 mg/kg bw,when female Sprague Dawley rats were treated with test chemical orally via gavage according to OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method).

Acute Inhalation Toxicity :

Waiver

Acute Dermal Toxicity : 

1.It was concluded that the acute dermal median lethal dose (LD50) of Disodium 5-acetylamino-4-hydroxy-3-(phenylazo)naphthalene-2,7-disulphonate supplied by Sustainability Support Services (Europe) AB, Sweden, when administered to male and female Sprague Dawley rats was found to be greater than 2000 mg/kg body weight. Thus by considering the CLP criteria for acute toxicity rating for the chemicals, it infers that Disodium 5-acetylamino-4-hydroxy-3-(phenylazo)naphthalene-2,7-disulphonate does not exhibits acute toxicity by the dermal route.

2. It was concluded that the acute dermal median lethal dose (LD50) of Disodium 5-acetylamino-4-hydroxy-3-(phenylazo)naphthalene-2,7-disulphonate supplied by Sustainability Support Services (Europe) AB, Sweden, when administered to male and female Sprague Dawley rats was found to be greater than 2000 mg/kg body weight. Thus by considering the CLP criteria for acute toxicity rating for the chemicals, it infers that Disodium 5-acetylamino-4-hydroxy-3-(phenylazo)naphthalene-2,7-disulphonate does not exhibits acute toxicity by the dermal route.

 

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records

Reference

Reference 1

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Remarks:

Read across data

Adequacy of study:

weight of evidence

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 journal.

Qualifier:

equivalent or similar to guideline

Guideline:

other: as mentioned below

Principles of method if other than guideline:

To determine the acute oral toxicity of the test chemical.

GLP compliance:

not specified

Test type:

other: not specified

Limit test:

yes

Species:

mouse

Strain:

other: DDY

Sex:

male

Details on test animals or test system and environmental conditions:

Details on test animal
TEST ANIMALS
Source: Japan SLC Co., Shizuoka, Japan
Age at study initiation: 7 weeks
Weight at study initiation: no data
Fasting period before study: no data
Housing: no data
Diet (e.g. ad libitum): commercial pellets MF (Oriental Yeast Industries Co., Tokyo, Japan), ad libitum
Water (e.g. ad libitum): Tap Water, ad libitum
Acclimation period: 1 week of acclimatization

ENVIRONMENTAL CONDITIONS
Temperature (°C): The animal room was kept at 20–24°C.
Humidity (%): No data
Air changes (per hr): No data
Photoperiod (hrs dark / hrs light): 12 h Light – dark cycle.

Route of administration:

oral: unspecified

Vehicle:

physiological saline

Details on oral exposure:

MAXIMUM DOSE VOLUME APPLIED:
0.5 × LD50 or the limit dose of 2000 mg/kg.

Doses:

0.5 × LD50 or the limit dose of 2000 mg/kg.

No. of animals per sex per dose:

4 - 5 male mice

Control animals:

not specified

Details on study design:

no data

Statistics:

no data

Preliminary study:

no data

Sex:

male

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Remarks on result:

other: no mortality was observed

Mortality:

no mortality observed at dose 2000 mg/kg bw.

Clinical signs:

other: no data

Gross pathology:

no data

Other findings:

no data

Table 1: Acute Oral Toxicity study results

 

Test chemical	Vehicle	Source*	LD50** mg/kg
TEST CHEMICAL	Saline	T	>2000

 

*T = Tokyo Kasei Kogyo Industry Ltd., Tokyo, Japan;

** In order to set appropriate doses for the assay, we determined approximate LD50 by simple acute toxicity experiments on four–five animals. When no death was observed at 2000 mg/kg, the LD50 was defined as >2000 mg/kg.

Interpretation of results:

other: not classified

Conclusions:

A simple acute oral toxicity experiment was conducted prior to Comet assay in male DDY mice.When no death was observed at 2000 mg/kg, the LD50 was defined as >2000 mg/kg.The acute oral LD50 in male DDY mice was determined to be greater than 2000 mg/kg after dosing with test chemical.

Executive summary:

A simple acute oral toxicity experiment was conducted prior to Comet assay in male DDY mice. The test was performed to access the toxicity potential of the read across substance Amaranth dye (CAS no.: 915-67-3). Male ddY mice were obtained from Japan SLC Co., Shizuoka, Japan, at 7 weeks of age and used after 1 week of acclimatization. They were fed commercial pellets MF (Oriental Yeast Industries Co., Tokyo, Japan) and tap water ad libitum throughout the acclimatization period and the experiment. The animal room was kept at 20–24 °C with a 12 h light – dark cycle. In order to set appropriate doses for the assay, we determined approximate LD50 by simple acute toxicity experiments on four–five animals. When no death was observed at 2000 mg/kg, the LD50 was defined as >2000 mg/kg. The acute oral LD50 in male DDY mice was determined to be greater than 2000 mg/kg after dosing with test chemical. Hence, the test substance was considered to be not classified as per the CLP criteria.

Reference 2

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Remarks:

Read across data

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Justification for type of information:

Data is from publication

Qualifier:

according to guideline

Guideline:

other: as per mentioned below

Principles of method if other than guideline:

Oral dosages of compound were administered by oral intubation to the mouse. Animals were observed usually for 14 days during which time the development of toxic signs was followed and time of death recorded.

GLP compliance:

not specified

Test type:

standard acute method

Limit test:

yes

Specific details on test material used for the study:

- Name of test material (as cited in study report): Brilliant Black PN
- Molecular formula (if other than submission substance): C28H17N5Na4O14S4
- Molecular weight (if other than submission substance):867.66716 g\mol
- Substance type: Organic
- Physical state: Solid
-Purity:83.6%

Species:

mouse

Strain:

other: ICI Alderley Park Strain 1 SPF

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: No data available
- Age at study initiation: No data available
- Weight at study initiation: No data available
- Fasting period before study: 18 hr prior to dosing
- Housing: No data available
- Diet (e.g. ad libitum): No data available
- Water (e.g. ad libitum): No data available
- Acclimation period: No data available

ENVIRONMENTAL CONDITIONS
- Temperature (°C): No data available
- Humidity (%):No data available
- Air changes (per hr): No data available
- Photoperiod (hrs dark / hrs light): No data available

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

Route of administration:

oral: gavage

Vehicle:

water

Details on oral exposure:

No data available

Doses:

no data

No. of animals per sex per dose:

10 mice/sex

Control animals:

yes

Details on study design:

no data available

Statistics:

LD50 values with 95 % confidence limits were calculated according to Litchfield & Wllcoxon (1949).

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Remarks on result:

other: with 95 %confidence limits

Mortality:

No mortality observed

Clinical signs:

other: no toxic signs observed

Gross pathology:

No data available

Other findings:

Orally, a substantial amount of coloured material was excreted in the faeces.

Interpretation of results:

other: Not classified

Conclusions:

In acute toxicity study by oral route the LD50 value for the Brilliant Black PN was found to be >2000 mg/kg for mice

Executive summary:

An acute oral toxicity study was performed to access the toxicity potential of the read across substance Brilliant Black PN (CAS no. 2519-30-4, E.C. no.: 219-746-5. The test substance was administered in 10 male and female mice of ICI Alderley Park Strain 1 SPF strain via oral route. The test substance was administered at a dose of 2000mg/kg. The treated animals were further observed for clinical signs of toxicity and mortality. The LD50 value with 95% confidence limits were calculated according to LItchfield & Wilcoxon. No clinical signs of toxicity or mortality was observed in any of the animal at the dose of 2000mg/kg. Therefore, the LD 50 value of Brilliant Black PN was found to be >2000 mg/kg for mice. Hence, the test substance was considered to be "not classified", as per the CLP criteria.

Reference 3

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Remarks:

Read across data

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

Data is from experimental study report of read across substance

Qualifier:

according to guideline

Guideline:

OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)

Principles of method if other than guideline:

The purpose of this study was to assess the Toxicological profile of test item to a single administration via oral route to Sprague Dawley rats. This study was designed to determine the acute toxicity at fixed dose levels by oral route of the test item.

GLP compliance:

yes

Test type:

acute toxic class method

Limit test:

yes

Species:

rat

Strain:

Sprague-Dawley

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: National Institute of Biosciences, Pune.
- Females nulliparous and non-pregnant: yes
- Age at study initiation: Female rats of the age of approximately 8 to 12 weeks old were used.
- Weight at study initiation: Body weight range was 199.1 to 219.9 grams.
Body weights at the start : Female Mean: 206.81 g (= 100 %); Minimum : 199.1 g (- 3.73 %); Maximum : 219.9 g (+ 6.33 %)
Total No. of animals : 12
- Fasting period before study: Approximately 16 hours or more.
- Housing: The rats were housed in polycarbonate cages.
- 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 and passed through the reverse osmosis membrane before use.
- Acclimation period: 5 days.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20.6 to 23.2 degree centigrade.
- Humidity (%): 55.1% to 58.6%.
- 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: 26-09-2016 to 15-10-2016

Route of administration:

oral: gavage

Vehicle:

water

Remarks:

(Distilled water)

Details on oral exposure:

VEHICLE
- Concentration in vehicle: 300 mg/kg, 300 mg/kg, 2000 mg/kg and 2000 mg/kg
MAXIMUM DOSE VOLUME APPLIED: 10 ml per kg of body weight.

Doses:

Dose Group I : 300 mg/kg
Dose Group I : 300 mg/kg
Dose Group II : 2000 mg/kg
Dose Group II : 2000 mg/kg

No. of animals per sex per dose:

Three females were used at each step.

Control animals:

not specified

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Twice daily
- Necropsy of survivors performed: yes
- Other examinations performed: Clinical Observations and General Appearance: Animals were observed for clinical signs, mortality and morbidity, until sacrifice.
Onset, duration and severity of any sign were recorded. The clinical signs and mortality observations were conducted at immediately (0 to 5 minutes), 5, 10, 30, 60 minutes, 2, 4 and 6 hours on the day of dosing and once daily therea fter for 14 day. Daily observation was done as far as possible at the same time.
Body weights: Individual animal body weights were recorded, before fasting, prior to administration of the test item (fasting body weights), weekly thereafter and at termination on day 14. Weight changes were calculated and recorded.
Gross Pathology: Necropsy was performed on all animals at the end of the study period on day 15. 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.
Histopathology: No gross abnormalities were observed in animals sacrificed terminally hence, no histopathology was performed.

Statistics:

No data available

Preliminary study:

no data available

Sex:

female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Remarks on result:

other: No mortality was noted

Mortality:

All animals treated at the dose level of 300 mg/kg body weight and 2000mg/kg body weight survived through the study period of 14 days.

Clinical signs:

other: Group I Step I : Animals treated at the dose level of 300 mg/kg body weight did not result in any signs of toxicity during the study period of 14 days. Group I Step II : Animals treated at the dose level of 300 mg/kg body weight did not result in any sign

Gross pathology:

Gross pathological examination did not reveal any abnormalities in animals from 300 mg/kg and 2000 mg/kg dose groups.

Other findings:

No data available

Table No. I

 

Summary of Clinical Signs of Toxicity and Mortality

Test System : Sprague Dawley Rat

Sex : Female

Group I :   

Step No.	Dose mg/kg	Observed Signs	Total Number of Animals	Animal Nos.	Period of signs in days From - to	Mortality
I	300	No clinical signs observed	3	1 - 3	0 to 14	0/3

 

 

Group I :

Step No.	Dose mg/kg	Observed Signs	Total Number of Animals	Animal Nos.	Period of signs in days From - to	Mortality
II	300	No clinical signs observed	3	4 - 6	0 to 14	0/3

 

 

Group II :

Step No.	Dose mg/kg	Observed Signs	Total Number of Animals	Animal Nos.	Period of signs in days From - to	Mortality
I	2000	Diarrhoea (Black colour stools)	3	7,9 8	4 hrs. - 6 hrs. 2 hrs. - 6 hrs.	0/3

 

Group II :

Step No.	Dose mg/kg	Observed Signs	Total Number of Animals	Animal Nos.	Period of signs in days From - to	Mortality
II	2000	Diarrhoea (Black colour stools)	3	10,11,12	4 hrs. - 6 hrs.	0/3

 

Staining of the stool is attributed to the black colour of the test item.

 

 

Table No.II

 

Mean Body Weight and Percent Body Weight Gain (g)

Test System : Sprague Dawley Rat

Sex : Female

Group I :

Step No.	Dose (mg/kg body weight)		Before Fasting Body weight	Body weight Day 7	% body weight gain day 0-7	Body weight Day 14	% body weight gain day 7- 14	% body weight gain day 0- 14
I	300	Mean	201.47	210.43	4.45	227.17	7.95	12.76
		± SD	2.63	2.15	0.39	2.19	0.25	0.68

 

 

Group I :

Step No.	Dose (mg/kg body weight)		Before Fasting Body weight	Body weight Day 7	% body weight gain day 0-7	Body weight Day 14	% body weight gain day 7- 14	% body weight gain day 0- 14
II	300	Mean	203.23	210.47	3.56	228.60	8.62	12.49
		± SD	1.35	1.86	0.76	1.71	0.87	1.33

 

 

Group II :

Step No.	Dose (mg/kg body weight)		Before Fasting Body weight	Body weight Day 7	% body weight gain day 0-7	Body weight Day 14	% body weight gain day 7- 14	% body weight gain day 0- 14
I	2000	Mean	207.00	213.33	3.06	229.73	7.69	10.98
		± SD	2.52	2.51	0.67	3.33	0.29	0.81

 

 

Group II :

Step No.	Dose (mg/kg body weight)		Before Fasting Body weight	Body weight Day 7	% body weight gain day 0-7	Body weight Day 14	% body weight gain day 7- 14	% body weight gain day 0- 14
II	2000	Mean	215.53	220.20	2.17	236.77	7.52	9.86
		± SD	4.45	3.22	0.67	3.50	0.29	0.91

 

 

Table No.III

 

Summary of Gross Pathological Findings

Test System : Sprague Dawley Rat

Sex : Female

Group I :

Step No.	Dose mg/kg	Animal Numbers	Animal Fate	Gross Pathological Findings
I	300	1 - 3	TS	No abnormality detected

 

Group I :

Step No.	Dose mg/kg	Animal Numbers	Animal Fate	Gross Pathological Findings
II	300	4 - 6	TS	No abnormality detected

 

Group II :

 

Step No.	Dose mg/kg	Animal Numbers	Animal Fate	Gross Pathological Findings
I	2000	7 - 9	TS	No abnormality detected

 

    Group II :

Step No.	Dose mg/kg	Animal Numbers	Animal Fate	Gross Pathological Findings
II	2000	10 - 12	TS	No abnormality detected

  TS = Terminal Sacrifice

Interpretation of results:

other: not classified

Conclusions:

The lethal concentration value (LD50) for acute oral toxicity test was considered to be >2000 mg/kg bw,when female Sprague Dawley rats were treated with test chemical orally via gavage according to OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method).

Executive summary:

The reported study was designed and conducted to determine the acute oral toxicity profile of the given test chemical in Sprague Dawley rats. Initially, three female animals were treated at the dose level of 300 mg/kg body weight of the test item (Step - I). Administration of the test item at 300 mg/kg did not result in any signs of toxicity and mortality at 24 hours after the dosing. As no mortality was observed at 24 hours after the dosing, three female animals were added to the study and treated with the same dose of 300 mg/kg of the test item (Step - II). Administration of the test item at 300 mg/kg did not result in any signs of toxicity and mortality after the dosing. No mortality was observed at 300 mg/kg dose group, hence additional three female animals were treated with the higher dose of 2000 mg/kg of the test item (Step - I). Administration of the test item at 2000 mg/kg resulted in diarrhea (black color stools) in all animals with onset at 2 hours and no mortality after the dosing. As no mortality were observed at 24 hours after the dosing, hence additional three female animals were treated with the higher dose of 2000 mg/kg of the test item (Step - II). Administration of the test item at 2000 mg/kg resulted in diarrhea (black color stools) in all animals with onset at 4 hours and no mortality after the dosing. All animals from 300 mg/kg and 2000 mg/kg dose groups survived through the study period of 14 days. Staining of the stool is attributed to the black color of the test item. Gross pathological examination did not reveal any abnormalities in animals from 300 mg/kg and 2000 mg/kg dose groups. Under the condition of this, it can be concluded that the lethal concentration value (LD50) for acute oral toxicity test was considered to be >2000 mg/kg bw, when female Sprague Dawley rats were treated with test chemical orally via gavage according to OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method). Hence, the test substance can be considered to be "not classified", as per the CLP criteria.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

> 2 000 mg/kg bw

Quality of whole database:

klimisch 2

Acute toxicity: via inhalation route

Link to relevant study records

Reference

Endpoint:

acute toxicity: inhalation

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

the study does not need to be conducted because exposure of humans via inhalation is not likely taking into account the vapour pressure of the substance and/or the possibility of exposure to aerosols, particles or droplets of an inhalable size

Acute toxicity: via dermal route

Link to relevant study records

Reference

Reference 1

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Justification for type of information:

Data is from experimental study report of read across substance

Qualifier:

according to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Principles of method if other than guideline:

This study was designed to determine the dermal LD50 of the test item (up to 2000 mg/kg) or to establish a non-lethal dose level of 2000 milligram of test item per kilogram of body weight.

GLP compliance:

yes

Test type:

standard acute method

Limit test:

yes

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: National Institute of Biosciences, Pune.
- Females (if applicable) nulliparous and non-pregnant: No data available
- Age at study initiation: Young adult male and female rats aged between 6 – 9 weeks were used.
- Weight at study initiation: The weight ranges of approximately 239.3 to 272.5 grams at initiation of dosing were used.
Body weights at the start :
Male
Mean : 270.14 g (= 100 %)
Minimum : 267.1 g (- 1.13 %)
Maximum : 272.5 g (+ 0.87 %)
Total No. of animals : 5
Female
Mean : 243.94 g (= 100 %)
Minimum : 239.3 g (- 1.90 %)
Maximum : 250.1 g (+ 2.53 %)
Total No. of animals : 5
- 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 and passed through the reverse osmosis membrane before use.
- Acclimation period: 5 days.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20.1 to 22.3 degree centigrade.
- Humidity (%): 55.7% to 59.6%.
- 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: 30-09-2016 to 15-10-2016

Type of coverage:

semiocclusive

Vehicle:

water

Remarks:

(Distilled water)

Details on dermal exposure:

TEST SITE
- Area of exposure: Trunk (dorsal surface and sides from scapular to pelvic area)
- % coverage: Approximately 10% of the total 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.

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 2000 mg/kg bw
- Constant volume or concentration used: No data available
- For solids, paste formed: Yes

VEHICLE
- Amount(s) applied (volume or weight with unit): No data available
- Concentration (if solution): No data available
- Lot/batch no. (if required): No data available
- Purity: No data available

Duration of exposure:

24 hours

Doses:

A single dose of 2000 mg of the test item per kilogram of body weight was administered to ten rats (five males and five females).

No. of animals per sex per dose:

10 (5/sex).

Control animals:

not specified

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Twice daily
- Necropsy of survivors performed: Yes
- Other examinations performed:
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 were 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, 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.

Statistics:

No data

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

Sex : Male
Group I -
Animal treated at the dose level of 2000 mg/kg: All animals survived through the study period of 14 days.

Sex : Female
Group I –
Animal treated at the dose level of 2000 mg/kg: All animals survived through the study period of 14 days.

Clinical signs:

other: 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. Sex : Female Group I - Animal treated at the dose level of 2000 mg/kg body weight did not resul

Gross pathology:

Gross pathological examination did not reveal any abnormalities in animals from 2000 mg/kg dose group.

Other findings:

- Other observations:
Evaluation of Dermal Reaction
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 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 skin reaction during the study period of 14 days.

Summary of Clinical Signs of Toxicity and Mortality

Test System : Sprague Dawley Rat

Sex : Male

Group  No.	Dose mg/kg	Observed Signs	Total Number of Animals	Animal Nos.	Period of signs in days  From - to	Mortality
I	2000	No clinical signs observed	5	1 - 5	0 - 14	0/5

 

Sex : Female 

Group  No.	Dose mg/kg	Observed Signs	Total Number of Animals	Animal Nos.	Period of signs in days  From - to	Mortality
I	2000	No clinical signs observed	5	6 - 10	0 - 14	0/5

 

Summary of Evaluation of Dermal Reaction

Test System : Sprague Dawley Rat

Sex : Male 

Group  No.	Dose mg/kg	Dermal Reaction	Total Number of Animals	Animal Nos.	Period of signs in days  From - to	Mortality
I	2000	No dermal reaction observed	5	1 - 5	0 - 14	0/5

Sex : Female 

Group  No.	Dose mg/kg	Dermal Reaction	Total Number of Animals	Animal Nos.	Period of signs in days  From - to	Mortality
I	2000	No dermal reaction observed	5	6 - 10	0 - 14	0/5

 

Mean Body Weight and Percent Body Weight Gain (g)

Test System : Sprague Dawley Rat

Sex : Male

Group No.	Dose (mg/kg body weight)		Body weight Day 0	Body weight Day 7	% body weight gain day 0-7	Body weight Day 14	% body weight gain day 7- 14	% body weight gain day 0- 14
I	2000	Mean	270.14	296.48	9.74	317.08	6.96	17.37
		± SD	2.10	7.20	1.82	6.26	1.34	1.56

 

Sex : Female

Group No.	Dose (mg/kg body weight)		Body weight Day 0	Body weight Day 7	% body weight gain day 0-7	Body weight Day 14	% body weight gain day 7- 14	% body weight gain day 0- 14
I	2000	Mean	243.94	255.88	4.89	267.40	4.50	9.61
		± SD	4.16	5.39	0.94	7.13	1.42	1.32

 

Summary of Gross Pathological Findings

 

Test System : Sprague Dawley Rat

Sex : Male

Group No.	Dose mg/kg	Animal Numbers	Animal Fate	Gross Pathological Findings
I	2000	1 - 5	TS	No abnormality detected

 

Sex : Female

Group No.	Dose mg/kg	Animal Numbers	Animal Fate	Gross Pathological Findings
I	2000	6 - 10	TS	No abnormality detected

TS = Terminal Sacrifice

Individual Animal -Clinical Signs of Toxicity and Mortality

Test System : Sprague Dawley Rat

Sex : Male 

Group No.	Dose mg/kg	Observed Signs	Total Number of Animals	Animal Nos.	Period of signs in days From - to	Mortality
I	2000	No clinical signs observed	5	1	0 - 14	0
				2	0 - 14	0
				3	0 - 14	0
				4	0 - 14	0
				5	0 - 14	0

 

Sex : Female

Group No.	Dose mg/kg	Observed Signs	Total Number of Animals	Animal Nos.	Period of signs in days From - to	Mortality
I	2000	No clinical signs observed	5	6	0 - 14	0
				7	0 - 14	0
				8	0 - 14	0
				9	0 - 14	0
				10	0 - 14	0

Individual Animal - Evaluation of Dermal Reaction

Test System : Sprague Dawley Rat

Sex : Male  

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
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
4	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
5	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

 

 

Sex : Female  

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
6	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
7	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
8	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
9	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
10	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

Individual Animal - Body Weight and Percent Body Weight Gain (g)

Test System : Sprague Dawley Rat

Sex    : Male

Group : I

Dose  : 2000 mg/kg body weight 

Animal No.	Body weight Day 0	Body weight Day 7	% body weight gain day 0-7	Body weight Day 14	% body weight gain day 7- 14	% body weight gain day 0- 14
1	267.1	286.2	7.15	309.2	8.04	15.76
2	269.2	292.7	8.73	314.1	7.31	16.68
3	270.4	297.5	10.02	320.1	7.60	18.38
4	271.5	302.2	11.31	316.2	4.63	16.46
5	272.5	303.8	11.49	325.8	7.24	19.56

 

Sex    : Female

Group : I

Dose  : 2000 mg/kg body weight 

Animal No.	Body weight Day 0	Body weight Day 7	% body weight gain day 0-7	Body weight Day 14	% body weight gain Day 7- 14	% body weight gain day 0- 14
6	239.3	250.9	4.85	262.1	4.46	9.53
7	241.3	254.7	5.55	260.0	2.08	7.75
8	243.5	251.5	3.29	265.5	5.57	9.03
9	245.5	258.4	5.25	272.4	5.42	10.96
10	250.1	263.9	5.52	277.0	4.96	10.76

Individual Animal - Gross Pathological Findings

Test System : Sprague Dawley Rat

Sex    : Male

Group : I

Dose  : 2000 mg/kg body weight 

Animal No.	Fate	Gross Pathological Findings
1	TS	No abnormality detected
2	TS	No abnormality detected
3	TS	No abnormality detected
4	TS	No abnormality detected
5	TS	No abnormality detected

 

Sex    : Female

Group : I

Dose  : 2000 mg/kg body weight 

Animal No.	Fate	Gross Pathological Findings
6	TS	No abnormality detected
7	TS	No abnormality detected
8	TS	No abnormality detected
9	TS	No abnormality detected
10	TS	No abnormality detected

TS = Terminal sacrifice

Interpretation of results:

other: Not classified

Conclusions:

It was concluded that the acute dermal median lethal dose (LD50) of test chemical, when administered to male and female Sprague Dawley rats was considered to be greater than 2000 mg/kg body weight. Thus, according to CLP criteria for acute toxicity rating for the chemicals,the test chemical can be classified under the category "Not Classified".

Executive summary:

A study was designed and conducted to determine the acute dermal toxicity profile of test chemical in Sprague Dawley rats. The study was performed according to OECD 402 Guidelines.

A single dose of 2000 mg/kg was administered to ten rats (five males and five females). Animals were observed for clinical signs, mortality, until sacrifice.Onset, duration and severity of any sign were recorded. The clinical signs and mortality observations were 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, respiratory, circulatory system and behavior pattern. Dermal reaction was observed daily for study period of 14 days. 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). No gross abnormalities were observed in animals sacrificed terminally hence, no histopathology was performed. 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 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. 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. It was concluded that the acute dermal median lethal dose (LD50) of test chemical, when administered to male and female Sprague Dawley rats was considered to be greater than 2000 mg/kg body weight. Thus, according to CLP criteria for acute toxicity rating for the chemicals,the test chemical can be classified under the category "Not Classified".

Reference 2

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Remarks:

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:

guideline study with acceptable restrictions

Justification for type of information:

Data is from read across substance experimental study report.

Qualifier:

according to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Principles of method if other than guideline:

Acute Dermal Toxicity of Disodium 5-acetylamino-4-hydroxy-3-(phenylazo)naphthalene-2,7-disulphonate (Cas No: 3734-67-6) in Rat.

GLP compliance:

yes

Test type:

standard acute method

Limit test:

yes

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source of test material: Sustainability Support Services (Europe) AB, Sweden
- Lot/batch No.of test material: 0009
- Expiration date of the lot/batch: 4/7/2021
- Purity test date: No data

RADIOLABELLING INFORMATION (if applicable)
- Radiochemical purity: No data
- Specific activity: No data
- Locations of the label: No data
- Expiration date of radiochemical substance: No data

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Ambient Temperature
- Stability under test conditions: No data
- 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: Test item was moistened with distilled water before application.
- 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) No data

OTHER SPECIFICS:
Safety Precautions: Safety precautions included use of protective clothing, gloves, masks and eye protection (glasses).

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: National Institute of Biosciences, Pune.
- Females (if applicable) nulliparous and non-pregnant: [no]
- Age at study initiation: Young adult male and female rats aged between 6 – 9 weeks were used.
- Weight at study initiation: The weight ranges of approximately 211.4 to 246.3 grams at initiation of dosing were used.
Body weights at the start :
Male
Mean : 240.62 g (= 100 %)
Minimum: 236.8 g (- 1.59 %)
Maximum: 246.3 g (+ 2.36 %)
Total No. of animals : 5
Female
Mean : 216.02 g (= 100 %)
Minimum: 211.4 g (- 2.14 %)
Maximum: 220.6 g (+ 2.12 %)
Total No. of animals : 5
- Fasting period before study: No data
- 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 and passed through the reverse osmosis membrane before use.
- Acclimation period: 5 days.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): Room temperature was maintained at 20.1 to 21.9 degree centigrade.
- Humidity (%): Room humidity was maintained at 55.5% to 59.2%.
- 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: No data

Type of coverage:

occlusive

Vehicle:

unchanged (no vehicle)

Details on dermal exposure:

TEST SITE
- Area of exposure: Dorsal surface and sides from scapular to pelvic area.
- % coverage: Approximately 10% of the total 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.

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 2000 mg/kg bw
- Constant volume or concentration used: No data
- For solids, paste formed: No data

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 exposure:

24 hours

Doses:

A single dose of 2000 mg of the test item per kilogram of body weight was administered to ten rats (five males and five females).

No. of animals per sex per dose:

10 (5/sex).

Control animals:

yes

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Twice daily
- Necropsy of survivors performed: Yes
- Other examinations performed:
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 were 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, 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).

Statistics:

No data

Preliminary study:

No data

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Remarks on result:

other: No mortality was observed

Mortality:

Sex : Male
Group I -
Animal treated at the dose level of 2000 mg/kg: All animals survived through the study period of 14 days.

Sex : Female
Group I –
Animal treated at the dose level of 2000 mg/kg: All animals survived through the study period of 14 days.

Clinical signs:

other: 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. Sex : Female Group I - Animal treated at the dose level of 2000 mg/kg body weight did not resul

Gross pathology:

Gross pathological examination did not reveal any abnormalities in animals from 2000 mg/kg dose group.

Other findings:

- Other observations:
Evaluation of Dermal Reaction
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 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 skin reaction during the study period of 14 days.

Table No. I

 

Summary of Clinical Signs of Toxicity and Mortality

Test System : Sprague Dawley Rat

Sex : Male

Group  No.	Dose mg/kg	Observed Signs	Total Number of Animals	Animal Nos.	Period of signs in days  From - to	Mortality
I	2000	No clinical signs observed	5	1 - 5	Day 0 - Day 14	0/5

 

 

Sex : Female

Group  No.	Dose mg/kg	Observed Signs	Total Number of Animals	Animal Nos.	Period of signs in days  From - to	Mortality
I	2000	No clinical signs observed	5	6 - 10	Day 0 - Day 14	0/5

 

   

Table No. II

 

Summary of Evaluation of Dermal Reaction

Test System : Sprague Dawley Rat

Sex : Male 

Group  No.	Dose mg/kg	Dermal Reaction	Total Number of Animals	Animal Nos.	Period of signs in days  From - to	Mortality
I	2000	No dermal reaction observed	5	1 - 5	Day 0 - Day 14	0/5

 

 

Sex : Female

 

Group  No.	Dose mg/kg	Dermal Reaction	Total Number of Animals	Animal Nos.	Period of signs in days  From - to	Mortality
I	2000	No dermal reaction observed	5	6 - 10	Day 0 - Day 14	0/5

 

 

Table No.III

 

Mean Body Weight and Percent Body Weight Gain (g)

Test System : Sprague Dawley Rat

Sex : Male

Group No.	Dose (mg/kg body weight)		Body weight Day 0	Body weight Day 7	% body weight gain day 0-7	Body weight Day 14	% body weight gain day 7- 14	% body weight gain day 0- 14
I	2000	Mean	240.62	261.48	8.65	284.90	8.96	18.38
		± SD	3.89	8.12	1.68	8.58	0.70	1.92

 

 

 

Sex : Female

Group No.	Dose (mg/kg body weight)		Body weight Day 0	Body weight Day 7	% body weight gain day 0-7	Body weight Day 14	% body weight gain day 7- 14	% body weight gain day 0- 14
I	2000	Mean	216.02	228.50	5.76	238.72	4.46	10.48
		± SD	3.57	6.69	1.39	8.08	0.65	1.96

 

Table No.IV

 

Summary of Gross Pathological Findings

Test System : Sprague Dawley Rat

 

Sex : Male

Group No.	Dose mg/kg	Animal Numbers	Animal Fate	Gross Pathological Findings
I	2000	1 - 5	TS	No abnormality detected

 

Sex : Female

Group No.	Dose mg/kg	Animal Numbers	Animal Fate	Gross Pathological Findings
I	2000	6 - 10	TS	No abnormality detected

 TS = Terminal Sacrifice

 

Interpretation of results:

other: Not classified

Conclusions:

It was concluded that the acute dermal median lethal dose (LD50) of Disodium 5-acetylamino-4-hydroxy-3-(phenylazo)naphthalene-2,7-disulphonate supplied by Sustainability Support Services (Europe) AB, Sweden, when administered to male and female Sprague Dawley rats was found to be greater than 2000 mg/kg body weight. Thus by considering the CLP criteria for acute toxicity rating for the chemicals, it infers that Disodium 5-acetylamino-4-hydroxy-3-(phenylazo)naphthalene-2,7-disulphonate does not exhibits acute toxicity by the dermal route.

Executive summary:

The study now reported was designed and conducted to determine the acute dermal toxicity profile of Disodium 5-acetylamino-4-hydroxy-3-(phenyl azo)naph thalene-2,7-disulphonate in Sprague Dawley rats. 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. It was concluded that the acute dermal median lethal dose (LD₅₀) of Disodium 5-acetylamino-4-hydroxy-3-(phenylazo)naphthalene-2,7-disulphonate supplied by Sustainability Support Services (Europe) AB, Sweden, when administered to male and female Sprague Dawley rats was found to be >2000 mg/kg body weight. Thus by considering the CLP criteria for acute toxicity rating for the chemicals, it infers that Disodium 5-acetylamino-4-hydroxy-3- (phenylazo)naphthalene-2,7-disulphonate does not exhibits acute toxicity by the dermal route.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

> 2 000 mg/kg bw

Quality of whole database:

K2

Additional information

Acute oral toxicity:

1. The reported study was designed and conducted to determine the acute oral toxicity profile of the given test chemical in Sprague Dawley rats. Initially, three female animals were treated at the dose level of 300 mg/kg body weight of the test item (Step - I). Administration of the test item at 300 mg/kg did not result in any signs of toxicity and mortality at 24 hours after the dosing. As no mortality was observed at 24 hours after the dosing, three female animals were added to the study and treated with the same dose of 300 mg/kg of the test item (Step - II). Administration of the test item at 300 mg/kg did not result in any signs of toxicity and mortality after the dosing. No mortality was observed at 300 mg/kg dose group, hence additional three female animals were treated with the higher dose of 2000 mg/kg of the test item (Step - I). Administration of the test item at 2000 mg/kg resulted in diarrhea (black color stools) in all animals with onset at 2 hours and no mortality after the dosing. As no mortality were observed at 24 hours after the dosing, hence additional three female animals were treated with the higher dose of 2000 mg/kg of the test item (Step - II). Administration of the test item at 2000 mg/kg resulted in diarrhea (black color stools) in all animals with onset at 4 hours and no mortality after the dosing. All animals from 300 mg/kg and 2000 mg/kg dose groups survived through the study period of 14 days. Staining of the stool is attributed to the black color of the test item. Gross pathological examination did not reveal any abnormalities in animals from 300 mg/kg and 2000 mg/kg dose groups. Under the condition of this, it can be concluded that the lethal concentration value (LD50) for acute oral toxicity test was considered to be >2000 mg/kg bw, when female Sprague Dawley rats were treated with test chemical orally via gavage according to OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method). Hence, the test substance can be considered to be "not classified", as per the CLP criteria.

 

2. An acute oral toxicity study was performed to access the toxicity potential of the read across substance Brilliant Black PN (CAS no. 2519-30-4, E.C. no.: 219-746-5. The test substance was administered in 10 male and female mice of ICI Alderley Park Strain 1 SPF strain via oral route. The test substance was administered at a dose of 2000mg/kg. The treated animals were further observed for clinical signs of toxicity and mortality. The LD50 value with 95% confidence limits were calculated according to LItchfield & Wilcoxon. No clinical signs of toxicity or mortality was observed in any of the animal at the dose of 2000mg/kg. Therefore, the LD 50 value of Brilliant Black PN was found to be >2000 mg/kg for mice. Hence, the test substance was considered to be "not classified", as per the CLP criteria.

 

3. A simple acute oral toxicity experiment was conducted prior to Comet assay in male DDY mice. The test was performed to access the toxicity potential of the read across substance Amaranth dye (CAS no.: 915-67-3). Male ddY mice were obtained from Japan SLC Co., Shizuoka, Japan, at 7 weeks of age and used after 1 week of acclimatization. They were fed commercial pellets MF (Oriental Yeast Industries Co., Tokyo, Japan) and tap water ad libitum throughout the acclimatization period and the experiment. The animal room was kept at 20–24 °C with a 12 h light – dark cycle. In order to set appropriate doses for the assay, we determined approximate LD50 by simple acute toxicity experiments on four–five animals. When no death was observed at 2000 mg/kg, the LD50 was defined as >2000 mg/kg. The acute oral LD50 in male DDY mice was determined to be greater than 2000 mg/kg after dosing with test chemical. Hence, the test substance was considered to be not classified as per the CLP criteria.

Acute Inhalation Toxicity : 

Waiver

Acute Dermal Toxicity :

1. A study was designed and conducted to determine the acute dermal toxicity profile of test chemical in Sprague Dawley rats. The study was performed according to OECD 402 Guidelines.

A single dose of 2000 mg/kg was administered to ten rats (five males and five females). Animals were observed for clinical signs, mortality, until sacrifice.Onset, duration and severity of any sign were recorded. The clinical signs and mortality observations were 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, respiratory, circulatory system and behavior pattern. Dermal reaction was observed daily for study period of 14 days. 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). No gross abnormalities were observed in animals sacrificed terminally hence, no histopathology was performed. 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 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. 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. It was concluded that the acute dermal median lethal dose (LD50) of test chemical, when administered to male and female Sprague Dawley rats was considered to be greater than 2000 mg/kg body weight. Thus, according to CLP criteria for acute toxicity rating for the chemicals,the test chemical can be classified under the category "Not Classified".

 

2.The study now reported was designed and conducted to determine the acute dermal toxicity profile of Disodium 5-acetylamino-4-hydroxy-3-(phenyl azo)naph thalene-2,7-disulphonate in Sprague Dawley rats. 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. It was concluded that the acute dermal median lethal dose (LD₅₀) of Disodium 5-acetylamino-4-hydroxy-3-(phenylazo)naphthalene-2,7-disulphonate supplied by Sustainability Support Services (Europe) AB, Sweden, when administered to male and female Sprague Dawley rats was found to be >2000 mg/kg body weight. Thus by considering the CLP criteria for acute toxicity rating for the chemicals, it infers that Disodium 5-acetylamino-4-hydroxy-3- (phenylazo)naphthalene-2,7-disulphonate does not exhibits acute toxicity by the dermal route.

 

Justification for classification or non-classification

Thus, based on the above-summarised studies of test chemical and its structurally similar read across substances, it can be concluded that LD50 value is >2000 mg/kg bw for acute oral and dermal studies. Thus, comparing this value with the criteria of CLP regulation, test chemical cannot be classified for acute oral and dermal toxicity.