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Toxicological information

Repeated dose toxicity: oral

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Administrative data

Endpoint:
short-term repeated dose toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: data from peer - reviewed journals

Data source

Reference
Reference Type:
publication
Title:
Immunological studies on Amaranth, Sunset Yellow and Curcumin as food colouring agents in albino rats
Author:
Mohamed M. Hashem a, Attia H. Atta, Mahmoud S. Arbid , Somaia A. Nada, Gihan Farag Asaad
Year:
2010
Bibliographic source:
Food and Chemical Toxicology 48 (2010) 1581–1586

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)
Principles of method if other than guideline:
Repeated dose oral toxicity study of Amaranth in rats
GLP compliance:
not specified
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
Trisodium 3-hydroxy-4-(4'-sulphonatonaphthylazo)naphthalene-2,7-disulphonate
EC Number:
213-022-2
EC Name:
Trisodium 3-hydroxy-4-(4'-sulphonatonaphthylazo)naphthalene-2,7-disulphonate
Cas Number:
915-67-3
Molecular formula:
C20H14N2O10S3.3Na
IUPAC Name:
trisodium 3-hydroxy-4-(4'-sulphonatonaphthylazo)naphthalene-2,7-disulphonate
Constituent 2
Reference substance name:
Amaranth
IUPAC Name:
Amaranth
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder
Details on test material:
- Name of test material (as cited in study report): Amaranth dye (E123)
- Molecular formula (if other than submission substance): C20-H11-N2-O10-S3.3Na
C20-H14-N2-O10-S3.3Na
- Molecular weight (if other than submission substance): 604.4789 g/mole
- Substance type: Organic
- Physical state: Powder
- Impurities (identity and concentrations): No data available

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: National Research Center (Giza, Egypt)
- Age at study initiation: No data available
- Weight at study initiation: No data available
- Fasting period before study: No data available
- Housing: Animals were housed 10 per in steel mesh cages
- Diet (e.g. ad libitum): Commercial standard pellets fed enriched with barley and carrots, ad libitum
- Water (e.g. ad libitum): Tap water, ad libitum
- 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

Administration / exposure

Route of administration:
oral: unspecified
Vehicle:
not specified
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: No data available

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

VEHICLE
- Justification for use and choice of vehicle (if other than water): No data available
- Concentration in vehicle: 47 mg/kg bw/day
- Amount of vehicle (if gavage): No data available
- Lot/batch no. (if required): No data available
- Purity: No data available
Analytical verification of doses or concentrations:
not specified
Duration of treatment / exposure:
14 weeks
Frequency of treatment:
Daily
Doses / concentrations
Remarks:
Doses / Concentrations:
0 and 47 mg/kg bw/day
Basis:

No. of animals per sex per dose:
Total : 20
0 mg/kg bw/day: 10 female
47 mg/kg bw/day: 10 female
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Rats administered Amaranth 7.5 mg/kg b. wt. calculated for human and modified according to Paget and Barnes (1964) to be 47 mg/kg b. wt. throughout the experimental period.
- Rationale for animal assignment (if not random): Animals were assigned randomly to test group.
- Rationale for selecting satellite groups: No data available
- Post-exposure recovery period in satellite groups: No data available
- Section schedule rationale (if not random): No data available

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: No data available
- Time schedule: No data available
- Cage side observations checked in table [No.?] were included. No data available

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

BODY WEIGHT: Yes
- Time schedule for examinations: On day 14 and 28 day

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

FOOD EFFICIENCY: No data available
- 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 available

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

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

HAEMATOLOGY: Yes
- Time schedule for collection of blood: On day 14 and 28 day
- Anaesthetic used for blood collection: No data available
- Animals fasted: No data available
- How many animals: All 20 animals were tested.
- Parameters checked in table [No.?] were examined. TLC and Differential leucocytic count (Lymphocytes, Neutrophiles, Monocytes, Basophiles and Eosinophiles) were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: On day 14 and 28 day
- Animals fasted: No data available
- How many animals: All 20 animals were tested.
- Parameters checked in table [No.?] were examined. Mononuclear cell count, Total serum protein, Albumin, Globulins and A/G ratio were examined.

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

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

OTHER:
Organ weight: Spleen and Thymus gland were weighted.
Sacrifice and pathology:
GROSS PATHOLOGY: No data available

HISTOPATHOLOGY: No data available
Other examinations:
Delayed type hypersensitivity response and Serum protein fractionation were examined
Statistics:
Statistical analyses were performed by using one way ANOVA and SPSS version (9.0). Data were represented as mean ± SE at p ≤ 0.05

Results and discussion

Results of examinations

Clinical signs:
not specified
Mortality:
not specified
Body weight and weight changes:
no effects observed
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:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
not specified
Behaviour (functional findings):
not specified
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
not specified
Histopathological findings: non-neoplastic:
not specified
Details on results:
Mortality: No data available

Clinical signs: No data available

Body weight and weight gain: No significant effect on body weight gain of treated rats was observed as compared to control.

Food consumption and compound intake: No data available

Food efficiency: No data available

Water consumption and compound intake: No data available

Opthalmoscopic examination No data available

Haematology No significant effects on total leucocytes count were observed in treated rats however, a significant increase in lymphocyte and basophiles % and neutrophils and monocytes % were significantly decreased as compared to control.

Clinical chemistry: Increased number of circulating mononuclear cells in peripheral blood but no effect on Total serum protein, albumin and globulin concentration were observed in treated rats as compared to control.

Urinanalysis: No data available

Neurobehaviour: No data available

Organ weights No significant effect on relative Spleen and Thymus gland weight were observed in treated rats as compared to control.

Gross pathology: No data available

Histopathology: No data available

Details on results: Decrease in neutrophiles and monocytes percent could affect the integrity of the cellular immune response.

Delayed hyper sensitivity:
Significantly decreased in oedema % of the paw were observed in treated rats as compared to control.

Serum protein fractionation:
Increases in density of albumin band and no effect on density of globulin region were observed in treated rats as compared to control.

Effect levels

Dose descriptor:
NOAEL
Effect level:
47 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
female
Basis for effect level:
other: No effect on body weight, hematology, clinical chemistry, Organ weight and humoral immune response

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

Effect of Amaranth in daily dose of 47 mg/kg b. wt., respectively, for 2 weeks before and 2 weeks after sensitization with sheep RBC’s on body weight and relative organ (mean ± SE, n = 10).

Groups

Body weight (g)

Spleen (% of body weight)

Thymus gland (% of body weight)

Control

216.0 ± 1.71

0.45 ± 0.04

0.22 ± 0.02

Amaranth

229.4 ± 14. 8

8 0.46 ± 0.04

0.18 ± 0.01

* Significant at p 6 0.05compared to control.

Effect of Amaranth in daily dose of 47 mg/kg b. wt., respectively, for 2 weeks before and 2 weeks after sensitization with sheep RBC’s on the total (TLC) and differential leucocytic (DLC) count in albino rats (mean ± SE, n = 10).

Groups

TLC(103/mm3)

Differential leucocytic count (%)

Control

35.84 ± 1.5

 

61.7 ± 3.51

35 ± 1.73

3.3 ± 1.04

1 ± 0.12

1.2 ± 0.15

Amaranth

32.03 ± 2.1

68.72±3.07*

26.33 ± 2.52*

2.5 ± 0.85*

1.3 ± 0.09*

1.25 ± 0.09

 

 

 

 

 

 

 

* Significant p 6 0.05 compared to control.

Effect of Amaranth in daily dose of 47 mg/kg b. wt., respectively, for 14 days before and 2 weeks after sensitization with sheep RBC’s on number of circulating mononuclear cells from peripheral blood samples in albino rats (mean ± SE, n = 10).

Groups

Mononuclear cell count X103/mm3

Control

19.77 ± 1.21

Amaranth

24.74 ± 1.19***

* Significant p 6 0.05 compared with control.

** Significant increase.

*** Significant decrease.

Effect of Amaranth in daily dose 47 mg/ kg b. wt. for 4 weeks on delayed hypersensitivity reaction induced by intradermal injection of 0.025 X 109SRBC/ml into right hind foot pad in albino rats in the 30thday (mean ± SE, n = 10).

Groups

Paw volume

% of oedema

 

Before injection

After injection

 

Control

0.86 ± 0.06

33.75 ± 2.81

33.75 ± 2.81

Amaranth

0.91 ± 0.03

1.25 ± 0.05

26.68 ± 2.29*

* Significant p 6 0.05 compared with control.

Effect of Amaranth in daily doses of 47 mg/kg b. wt., respectively, for 2 weeks before and 2 weeks after sensitization with sheep RBC’s on total serum protein, albumin and globulin (g%) and albumin/ globulin ratio in albino rats (mean ± SE, n = 10).

Groups

Total serum

protein (g/dl)

Albumin

(g/dl)

Globulins

(g/dl)

A/G ratio

Control

6.89 ± 0.2

4.17 ± 0.2

2.63 ± 0.07

1.6 ± 0.11

Amaranth

6.44 ± 0.42

4.08 ± 0.34

2.2 ± 0.15

1.88 ± 0.17

Applicant's summary and conclusion

Conclusions:
NOAEL was considered to be 47 mg/kg bw/day when Sprague Dawley female rats were treated with Amaranth dye (E123)
Executive summary:

In a repeated dose oral toxicity study, Sprague Dawley female rats were treated with Amaranth dye (E123) in the concentration of 0 and 47 mg/kg bw/day for 4 weeks.On day 14 animals were injected intraperitoneally with 1 ml/rat of 10% sheep RBCs suspension to test thedelayed type hypersensitivity response in rats.No significant effect on body weight gain was observed in treated rats as compared to control. Nosignificant effects on total leucocytes count were observed in treated rats however, a significant increase in lymphocyte and basophiles % and significant decreased in neutrophils and monocytes % were observed in treated rats as compared to control. Decrease in neutrophiles and monocytes percent could affect the integrity of the cellular immune response but not thehumoral immune response.Similarly, Increased number of circulating mononuclear cells in peripheral blood was observed but no effect on Total serum protein, albumin and globulin concentration were observed in treated rats as compared to control. In addition, Significant decreased in oedema % of the paw and increases in density of albumin band and no effect on density of globulin region were observed in treated rats as compared to control. No significant effect on relative Spleen and Thymus gland weight were observed in treated rats as compared to control. Therefore, NOAEL was considered to be47mg/kg bw/day whenSprague Dawley female rats were treated withAmaranth dye (E123) orally for 4 weeks.