Hibiscus sabdariffa LINN. (roselle) is widely cultivated in tropical areas and its red persistent calyx is the major component possessing a sour taste that is used as beverage and food colorants. It contains many chemical constituents including alkaloids, L-ascorbic acid, anisaldehyde, anthocyanin, bcarotene, b-sitosterol, citric acid, cyanidin-3-rutinoside, delphinidin, galactose, gossypetin, hibiscetin, mucopolysaccharide, pectin, protocatechuic acid, polysaccharide, quercetin, stearic acid and wax. As a traditional medicine, it is claimed to be effective against kidney stones and urinary bladder stones.1,2) It is also used for its antibacterial, antifungal, hypocholesterolemic, antispasmodic and antihypertensive actions. [3][4][5][6] Numerous studies have linked the elevation of plasma low-density lipoprotein (LDL)-cholesterol (LDL-C) level with the increased incidence of atherosclerotic events. LDL particles undergo extensive lipid peroxidation, resulting in generation of oxidized LDL and formation of atherosclerotic lesions. [7][8][9][10][11] Predictably, antioxidants such as a-tocopherol and probucol have been found to reduce LDL oxidation. [12][13][14][15] Various antioxidant constituents have been found in the calyx of Hibiscus sabdariffa LINN., including hibiscus anthocyanin, quercetin, L-ascorbic acid and protocatechuic acid (PCA). Antioxidant effects of roselle extracts have been investigated in many experimental models, [16][17][18][19] however the antioxidant effects of Hibiscus sabdariffa LINN. on LDL oxidation have so far not been fully determined. The present study was designed to quantitatively investigate the antioxidant effect of roselle on the oxidative modification of LDL induced by CuSO 4 in vitro by monitoring the formation of conjugated dienes and the formation of thiobarbituric acid reactive substances (TBARs).
MATERIALS AND MATHODS
Plant Material and ExtractsThe dried calyxes of roselle were blended to a fine powder and one kilogram of powders was extracted with 1 l of water and filtered through filter paper by suction. The filtrates were pooled and dried at 50°C in a rotary evaporator. The average yield of the extract was 45.0%. The extracts were stored in a dark, moisture-free container at 4°C. The antioxidant effect of PCA, a polyphenolic compound, has been investigated and found to exhibit potent action in primary cultures of rat hepatocytes induced by tert-butylhydroperoxide (t-BHP).18) Therefore, in the present experiment, PCA content in the calyx of roselle was used for the standardization of batches; 25 mg of dried calyx extracts contained 10.98 mg of PCA. For this experiment, 150 mg of the crude extracts were dissolved in 1 ml of distilled water.Reagents Disodium ethylene diamine was purchased from Mallinckrodt Inc. (St. Louis, MO, U.S.A.) and sodium chloride, sodium carbonate and sodium hydroxide from Merck (Haar, Germany). Diethylether was purchased from BDH laboratory supplies (Poole, England). Thiobarbituric acid, potassium bromide, disodium phosphate, sodium phosphate, sod...