Sodium 4-(4-(2-hydroxynaphthalenylazo)phenylazo)benzenesulphonate

Repeated dose toxicity: oral

Administrative data

Endpoint:

chronic toxicity: oral

Type of information:

experimental study

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 publication

Data source

Reference

Reference Type:

publication

Title:

Haematological Changes Induced By Feeding A Common Food Colour, Metanil Yellow, In Albino Mice

Author:

O.M. Prasad and Padam B. Rastogi

Year:

1983

Bibliographic source:

Toxicology Letters, 16 (1983) 103-107

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: Refer below principle

Principles of method if other than guideline:

Chronic oral toxicity study was performed to determine the oral toxic nature of metanil yellow upon repeated exposure

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Sodium 3-(p-anilinophenylazo)benzenesulphonate

EC Number:

209-608-2

EC Name:

Sodium 3-(p-anilinophenylazo)benzenesulphonate

Cas Number:

587-98-4

Molecular formula:

C18H14N3O3S.Na

IUPAC Name:

sodium 3-[(4-anilinophenyl)diazenyl]benzenesulfonate

Test material form:

solid

Details on test material:

- Name of test material: Ext. D&C Yellow No. 1
- IUPAC name: sodium 3-[(4-anilinophenyl)diazenyl]benzenesulfonate
- Molecular formula: C18H14N3O3SNa
- Molecular weight: 375.383 g/mol
- Substance type: organic

Specific details on test material used for the study:

- Name of test material: Ext. D&C Yellow No. 1
- IUPAC name: sodium 3-[(4-anilinophenyl)diazenyl]benzenesulfonate
- Molecular formula: C18H15N3O3SNa
- Molecular weight: 375.383 g/mol
- Substance type: organic
- Physical state: No data
- Purity: No data available
- Impurities (identity and concentrations): no data available

Test animals

Species:

mouse

Strain:

other: Albino

Details on species / strain selection:

No data

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: No data
- Age at study initiation: 4-6 weeks
- Weight at study initiation: 20-25 g
- Fasting period before study: No data
- Housing: No data
- Diet (e.g. ad libitum): pelleted lab chow (Hindustan Lever Ltd., Bombay, India)
- Water (e.g. ad libitum): Water ad libitum
- Acclimation period: No data

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 24°C ±1°C
- Humidity (%):No data
- Air changes (per hr): No data
- Photoperiod (hrs dark / hrs light): No data

IN-LIFE DATES: From: To: No data

Administration / exposure

Route of administration:

oral: feed

Details on route of administration:

No data

Vehicle:

other: Feed

Remarks:

pelleted lab chow (Hindustan Lever Ltd., Bombay, India)

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS: Metanil yellow was mixed with pelleted lab chow (Hindustan Lever Ltd., Bombay, India) at dose levels of 0, 100, 500 or 3000 mg/Kg bw

DIET PREPARATION
- Rate of preparation of diet (frequency): No data
- Mixing appropriate amounts with (Type of food): No data
- Storage temperature of food: No data

VEHICLE
- Justification for use and choice of vehicle (if other than water): pelleted lab chow (Hindustan Lever Ltd., Bombay, India)
- Concentration in vehicle: 0, 100, 500 or 3000 mg/Kg bw
- Amount of vehicle (if gavage): No data
- Lot/batch no. (if required): No data
- Purity: No data

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

No data

Duration of treatment / exposure:

180 days

Frequency of treatment:

Daily

Doses / concentrations

Remarks:

0, 100, 500 or 3000 mg/Kg bw

No. of animals per sex per dose:

Total: 320
0 mg/Kg bw: 40 males and 40 females
100 mg/Kg bw: 40 males and 40 females
500 mg/Kg bw: 40 males and 40 females
3000 mg/Kg bw: 40 males and 40 females

Control animals:

yes, concurrent vehicle

Details on study design:

No data

Positive control:

No data

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Throughout the experiment
- Cage side observations checked in table [No.?] were included. General condition of the animals was observed

DETAILED CLINICAL OBSERVATIONS: No data
- Time schedule: No data

BODY WEIGHT: No data
- Time schedule for examinations: No data

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): No data
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No data
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No data

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No data
- Time schedule for examinations: No data

OPHTHALMOSCOPIC EXAMINATION: No data
- Time schedule for examinations: No data
- Dose groups that were examined: No data

HAEMATOLOGY: Yes
- Time schedule for collection of blood: After 180 days
- Anaesthetic used for blood collection: No data
- Animals fasted: No data
- How many animals: No data
- Parameters checked in table [No.?] were examined. Total erythrocyte count (TEC), Differential leukocyte count (DLC)- neutrophils, lymphocytes, monocytes, eosinophils, basophils, hemoglobin, erythrocyte sedimentation rate (ESR), mean corpuscular volume (MCV), MCG, MCHC, Heinz bodies, platelet count (PC), PCV, Bleeding time (BT), coagulation time (CT)

CLINICAL CHEMISTRY: No data
- Time schedule for collection of blood: No data
- Animals fasted: No data
- How many animals: No data
- Parameters checked in table [No.?] were examined. No data

URINALYSIS: No data
- Time schedule for collection of urine: No data
- Metabolism cages used for collection of urine: No data
- Animals fasted: No data
- Parameters checked in table [No.?] were examined. No data

NEUROBEHAVIOURAL EXAMINATION: No data
- Time schedule for examinations: No data
- Dose groups that were examined: No data
- Battery of functions tested: sensory activity / grip strength / motor activity / other: No data

OTHER: No data

Sacrifice and pathology:

GROSS PATHOLOGY: Yes, organs were removed for gross pathology

HISTOPATHOLOGY: No data

Other examinations:

No data

Statistics:

No detailed data available

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

General condition was unaffected at dose levels of 100 and 500 mg/Kg bw except at 3000 mgKg bw

Mortality:

not specified

Body weight and weight changes:

not specified

Food consumption and compound intake (if feeding study):

not specified

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not specified

Ophthalmological findings:

not specified

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

100 mg/Kg bw: No abnormalilties were observed

500 mg/Kg bw: No abnormalilties were observed

3000 mg/Kg bw: Statistical analysis of various values exhibited significant decrease in TEC and Hb and increase in ESR, MCV and MCH suggesting normochromic macrocytic anaemia.

TLC, PC, PCV, MCHC, BT and CT, however, remained normal in both male and female animals.

A marked increase in the number of lymphocytes and macrocytes and a decrease in the number of neutrophils and eosinophils suggesting the occurrence of chronic lymphocytic leukaemia.

Heinz bodies began to appear in the blood after 3 months feeding in mice fed metanil yellow at 3.0 g/kg body weight and were observed in 80-90% erythrocytes at the end of the experiment.

Blood showed marked anisocytosis, poikilocytosis, stomatocytes, schistocytes and a large number of target cells, all normochromic in character. An
overwhelming preponderance of mature lymphocytes with scant cytoplasm, larger and denser clumps of nuclear chromatin and accentuation of parachromatin were also observed, which were suggestive of chronic lymphocytic leukaemia.

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

Mice fed metanil yellow at 3000 mg/kg bw showed hepatomegaly and splenomegaly at autopsy.

Neuropathological findings:

not specified

Histopathological findings: non-neoplastic:

not specified

Histopathological findings: neoplastic:

not specified

Other effects:

not specified

Details on results:

Clinical signs and mortality
Clinical signs: General condition was unaffected at dose levels of 100 and 500 mg/Kg bw except at 3000 mgKg bw

Mortality: No data

Body weight and weight gain: No data

Food consumption and compound intake: No data

Food efficiency: No data

Water consumption and compound intake: No data

Opthalmoscopic examination: No data

Haematology:
100 mg/Kg bw: No abnormalilties were observed

500 mg/Kg bw: No abnormalilties were observed

3000 mg/Kg bw: Statistical analysis of various values exhibited significant decrease in TEC and Hb and increase in ESR, MCV and MCH suggesting normochromic macrocytic anaemia.

TLC, PC, PCV, MCHC, BT and CT, however, remained normal in both male and female animals.

A marked increase in the number of
lymphocytes and macrocytes and a decrease in the number of neutrophils and eosinophils suggesting the occurrence of chronic lymphocytic leukaemia.

Heinz bodies began to appear in the blood after 3 months feeding in mice fed metanil yellow at 3.0 g/kg body weight and were observed in 80-90% erythrocytes at the end of the experiment.

Blood showed marked anisocytosis, poikilocytosis, stomatocytes, schistocytes and a large number of target cells, all normochromic in character. An
overwhelming preponderance of mature lymphocytes with scant cytoplasm, larger and denser clumps of nuclear chromatin and accentuation of parachromatin were also observed, which were suggestive of chronic lymphocytic leukaemia.

Clinical chemistry: No data

Urinanalysis: No data

Neurobehaviour: No data

Organ weights: No data

Gross pathology: Mice fed metanil yellow at 3000 mg/kg bw showed hepatomegaly and splenomegaly at autopsy.

Histopathology: No data

Effect levels

Key result

Dose descriptor:

NOAEL

Effect level:

500 mg/kg bw (total dose)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

clinical signs

gross pathology

haematology

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

Table I Haematological Findings in Male Mice Fed with Metanil Yellow at 0.0 and 3.0 G/Kg Body Weight Concentrations

No.	Experiment	SI Unit	0 mg/Kg bw	3000 mg/Kg bw
1	Total erythrocyte count (TEC)	10^12/L	6.13±0.45	3.86 ± 0.70
2	Differential leukocyte count (DLC) Neutrophils Lymphocytes Monocytes Eosinophils Basophils	10^9/L	5.67 ± 0.23 3.28 ± 0.16 0.44 ± 0.02 0.05 ± 0.01 0.00 ± 0.00	0.44 ± 0.16 9.10±0.44 0.65±0.03 0.01 ± 0.00 0.01 ± 0.00
3	Haemoglobin (Hb)	g/dL	16.29 ± 1.30	13.64 i 0.55
4	Erythrocyte sedimentation rate (ESR)	mm/h	1.81±0.25	4.18 ± 0.75
5	Mean corpuscular volume (MCV)	FI	11.77 ± 1.77	139.88±10.00
6	Mean corpuscular haemoglobin (MCH)	Pg	24.94±2.88	35.41±0.79
7	Heinz bodies	% of erythrocytes	0.00±0.00	89.25±2.50

Mean± Standard error 

 

Table II Haematological Findings in Female Mice Fed with Metanil Yellow at 0.0 and 3.0 G/Kg Body Weight Concentrations

No.	Experiment	SI Unit	0 mg/Kg bw	3000 mg/Kg bw
1	Total erythrocyte count (TEC)	10^12/L	5.26±0.37	3.49 ± c0.21
2	Differential leukocyte count (DLC) Neutrophils Lymphocytes Monocytes Eosinophils Basophils	10^9/L	5.14 ± 0.28 3.09 ± 0.19 0.40 ± 0.04 0.04 ± 0.01 0.00 ± 0.00	0.32 ± 0.11 9.20 ± 0.54 0.69 ± 0.06 0.01 ± 0.00 0.01 ± 0.00
3	Haemoglobin (Hb)	g/dL	13.54 ± 1.10	10.50 ± 1.00
4	Erythrocyte sedimentation rate (ESR)	mm/h	2.21 ± 0.30	4.31 ± 0.25
5	Mean corpuscular volume (MCV)	FI	74.10 ± 5.40	151.86 ± 4.20
6	Mean corpuscular haemoglobin (MCH)	Pg	25.76 ± 2.97	33.23 ± 0.47
7	Heinz bodies	% of erythrocytes	0.00 ± 0.00	81.75 ± 2.25

Mean± Standard error

 

Applicant's summary and conclusion

Conclusions:

The No Observed Adverse Effect level (NOAEL) for the test compound sodium 3-[(4-anilinophenyl)diazenyl]benzenesulfonate (metanil yellow) is determined to be 500 mg/Kg bw upon repeated exposure for 180 days to male and female albino mice by oral intubation route.

Executive summary:

Chronic oral toxicity study was performed to determine the oral toxic nature of sodium 3-[(4-anilinophenyl)diazenyl]benzenesulfonate (metanil yellow) upon repeated exposure for 180 days. The test compound was force fed to male and female albino mice by oral intubation at a dose levels of 0, 100, 500 or 3000 mg/Kg bw.

 

The animals were observed for general condition, changes in hematological parameters and gross pathology.

 

General condition was unaffected at dose levels of 100 and 500 mg/Kg bw except at 3000 mgKg bw. No change was observed by the 100 and 500 mg/Kg bw but feeding of this colour at the rate of 3000 mg/kg body weight led to certain changes in the haematological values. Total erythrocyte count (TEC) and Hb had decreased whereas ESR, MCV, and mean corpuscular haemoglobin (MCH) had increased. These facts suggested the occurrence of normochromic macrocytic anaemia. Differential leukocyte count (DLC) showed marked increase in the number of lymphocytes and monocytes and decrease in the number of neutrophils and eosinophils. Heinz bodies were observed in 80-90% erythrocytes. Other haematological values like total leukocyte count (TLC), platelet count (PC), PCV, MCHC, bleeding time (BT) and coagulation time (CT) were normal at 3000 mg/Kg bw.

 

Hence, the No Observed Adverse Effect level (NOAEL) for the test compound metanil yellow is 500 mg/Kg bw.