D&C Red No.7

CAS No. 5281-04-9

3-Hydroxy-4-[(4-methyl-2-sulfophenyl)azo]-2-naphthalene
carboxylic acid calcium salt

3-ヒドロキシ-4-［（4-メチル-2-スルフォフェニル）アゾ］-2-ナフタレン
カルボン酸カルシウム塩

Molecular formula: C18H12N2O6S・Ca Molecular weight: 424.25

ABSTRACT

With regard to repeat dose toxicity, males that received 300 mg/kg or more showed significantly decreased serum calcium and phosphorus levels. The 1000 mg/kg males further showed significantly decreased potassium and total cholesterol levels, and increased chloride and GOT levels. The relative kidney weights in the 1000 mg/kg males significantly increased, and the thymus weights in the 100 and 1000 mg/kg females significantly decreased. Histopathological lesions suggesting effects of D&C Red No.7 included an increased incidence of renal tubule regeneration in males that received 300 mg/kg or more, and the increased incidence of renal tubule regeneration and necrotic or foamy tubular epithelial cells in females that received 100 mg/kg or more. These lesions increased in severity and incidence in a dose-dependent manner. NOELs for repeat dose toxicity are considered to be 100 mg/kg/day for males and less than 100 mg/kg/day for females. In terms of reproductive and developmental toxicity, no treatment-related adverse effects were noted. The NOEL for reproductive/developmental toxicity is considered to be 1000 mg/kg/day.

D&C Red No.7 was not mutagenic in Salmonella typhimurium TA100, TA98, TA1535, TA1537 and Escherichia coli WP2 uvrA. Neither structural nor numerical chromosomal aberrations were induced in CHL/IU cells up to the concentration going 50% cell growth inhibition or the limit concentration of 5 mg/ml, in the absence or presence of an exogenous metabolic activation system.

SUMMARIZED DATA FROM THE STUDIES

1. Repeat Dose and Reproductive/Developmental Toxicity 1)

Purity	:	98 %
Test species/strain	:	Rat/Crj:CD (Sprague-Dawley)
Test method	:	OECD Combined Repeat Dose and Reproductive/Developmental Toxicity Screening Test
Route	:	Oral (Gavage)
Doses	:	100, 300, 1000 mg/kg/day
Number of animals	:	Males, 13; Females, 13/group
Vehicle	:	5% Gum Arabic solution
Administration period	:	Males, 42 days Females, from 14 days prior to mating to Day 3 of lactation
Terminal kill	:	Males, Day 43 Females, Day 4 of lactation
GLP	:	Yes

　Test results:

<Repeat dose toxicity>

All animals survived to the end of the study. No clinical findings indicative of chemical toxicity were observed; red-stained feces of dosed animals were due to the color of D&C Red No.7. The mean body weight gain and food consumption in both sexes of dosed groups were comparable to those in the control groups throughout the study. No biologically significant changes in hematological parameters were noted in any dosed male groups. Males that received 300 mg/kg or more showed significantly decreased serum calcium and phosphorus levels. Further, significant decreases in serum potassium and total cholesterol levels, and significant increases in chloride and GOT levels were shown in the males that received 1000 mg/kg. Males receiving 1000 mg/kg showed significant increases in relative kidney weights, and females that received 100 or 1000 mg/kg showed decreases in thymus weights in comparison with the controls. No other significant differences in organ weights were observed in either the males or the females. On the histopathological examination, predominant alterations suggesting effects of D&C Red No.7 in dosed rats were observed in the kidney. The lesions included an increased incidence of renal tubular epithelium regeneration in males receiving 300 mg/kg or more, and this and necrotic or foamy tubular epithelial cells in females that received 100mg/kg or more. These lesions increased in severity and incidence in a dose-dependent manner.

NOELs for repeat dose toxicity are considered to be 100 mg/kg/day for males and less than 100 mg/kg/day for females.

<Reproductive and developmental toxicity>

No treatment-related adverse effects were detected.

The NOEL is considered to be 1000 mg/kg/day.

2. Genetic Toxicity

2-1 Bacterial test 1)

Purity	:	98%
Test species/strains	:	S.typhimurium TA100, TA1535, TA98, TA1537, E. coli WP2 uvrA
Test method	:	Guideline for Screening Mutagenicity Testing of Chemicals (Japan)
Procedures	:	Plate incorporation method (including azo dye method)
Solvent	:	DMSO
Positive controls	:	-S9, AF-2 (TA100, WP2, TA98), sodium azide (TA1535) and 9-aminoacridine (TA1537) +S9, 2-aminoanthracene (all strains), trypane blue (TA100, TA98, azo reduction method)
Dosage	:	0, 312.5, 625, 1250, 2500, 5000μg/plate
S-9	:	Rat liver, induced with phenobarbital and 5,6-benzoflavone, Syrian Hamster liver, not induced(azo reduction method)
Plates/test	:	3
Number of replicates	:	2
GLP	:	Yes

　Test results:

Minimum concentration of test substance at which toxicity was observed:

No toxicity was observed up to a concentration of 5000 μg/plate with or without metabolic activation.

Genetic effects:

S. typhimurium TA100, TA1535, TA 98, TA1537

	+	?	-
with metabolic activation	[ ]	[ ]	[*]
without metabolic activation	[ ]	[ ]	[*]

E. coli WP2 uvrA

with metabolic activation	[ ]	[ ]	[*]
without metabolic activation	[ ]	[ ]	[*]

2-2. Non-bacterial in vitro test (chromosomal aberration test) 1)

Purity	:	About 98%
Type of cell used	:	Chinese hamster CHL/IU cells
Test method	:	Guidelines for Screening Mutagenicity Testing of Chemicals(Japan)
Solvent	:	0.5% Carboxymethyl cellulose sodium
Positive controls	:	-S9, Mitomycin C +S9, Cyclophosphamide
Doses	:	-S9 (continuous treatment): 0, 0.09, 0.19, 0.37 mg/ml -S9 (short-term treatment): 0, 1.3, 2.5, 5.0 mg/ml +S9 (short-term treatment): 0, 1.3, 2.5, 5.0 mg/ml
S-9	:	Rat liver, induced with phenobarbital and 5,6-benzoflavone
Plates/test	:	2
GLP	:	Yes

　Test results:

Lowest concentration producing cytogenetic effects in vitro:

without metabolic activation (continuous treatment ): > 0.37 mg/ml

without metabolic activation (short-term treatment): > 5.0 mg/ml

with metabolic activation (short-term treatment): > 5.0 mg/ml

Genotoxic effects:

	clastogenicity			polyploidy
	+	?	-	+	?	-
without metabolic activation:	[ ]	[ ]	[*]	[ ]	[ ]	[*]
with metabolic activation:	[ ]	[ ]	[*]	[ ]	[ ]	[*]

1)	The tests were performed by the Hatano Research Institute, Food and Drug Safety Center, 729-5 Ochiai, Hadano-shi, Kanagawa 257, Japan. Tel +81-463-82-4751 Fax +81-463-82-9627