| Epidemiological researches reveal that diet custom is closely correlating to several chronic diseases incidence, such as cancer, cardiovascular, hepatopathy disease, etc. In vivo and in vitro experiments indicate dietary flavonoids comprise an important role in defense and therapy of these chronic diseases. Legumes have been cultivated for thousands of years, and played an important role in the traditional diets of many regions throughout the world. Beans have long been recognized for their protein content and more recently have been noted for their soluble-fiber content, but in general there has been relatively little research and discussion about other nutritional attributes of legumes. Since isoflavones in soybeans were found to possess a myriad of biological effects to protect against several chronic diseases such as atherosclerosis, cancer, osteoporosis,etc., other nonsoy legumes had been taken to notice. Especially, flavonoids in legumes were emphasized for their antioxidative properties.Beans are widespreadly cultivated in China, which commonly consumed for dietary food. However, no work was carried out to research the biological effects ofcommonly consumed legumes systematically. Many of the varieties of beans that are commonplace today are even unknown until relatively recent times. In this study, a " food nutrient" plan cooperating with Germany is carried out. Eight common Chinese edibile legumes were selected, namely. Mungbean (Vigna radiata (L.) Wilczek), Adzuki bean (Phaseolus angular is Wight.) , Soybean (Glycine max (L.) Merr.), Lima bean (Phaseolus lunatus L.), Broadbean ( Viciafaba L.). pea (Pisum sativum L.), Common bean 1 (Phaseolus vulgaris L.), Common bean I I (Phaseolus vulgaris van humilis Alief) . To expand the potential for assessment of dietary intake of legumes, and their relationships to pathologic processes, main compounds of selected legumes were analyzed systematically in this study. Meanwhile, antioxidative activities of eight legume extracts were evaluated by analyzing their free radical scavenging capacities. Furthermore, cytoprotective effects of these extracts on Fe +-induced oxidative damaged cultured rat primary hepatocytes are also investigated.1. Legume extraction, main constituents separation and identificationEight extracts were acquired by ethanol extracting followed with polyamide enrichment and freeze-drying ultimately in Chinese edible legumes. Preliminary identification of UV-scanning. Thin-layer chromatography (TLC) and HCL/Mg reaction suggested that extracts contain flavonoids. Main compounds were separated by HPLC methods and identificated by mass spectrometer coupled to a HPLC system and NMR-spectra analysis. Several flavonoids were found in each legume extracts.2. Antioxidative and cytoprotective effects of Chinese edible legume ethanol extracts on Fe2+-induced oxidative damage in primary cultured rat hepatocytesAntioxidative activities evaluated by Photochenii revealed that adzuki beanextract possesed the strongest antioxidative activity followed with mung bean, common bean II, soybean, common bean I, broad bean, pea and lima bean extract. Cytoprotective effects of extracts on Fe2+-induced oxidative damage in primary cultured rat hepatocytes were assessed by MTT reduction and ALT leakage assays. The results indicated that 100 jomol/L FeSO4 exposure for 16 h induced acute cytotoxicity significantly (P<0.05) . Comparing with untreated group, MTT assay results decreased to 35%, ALT leakage increased about 4 flod. Hepatocytes pretreated with extract of mungbean, adzuki bean, common bean I and common bean II increased MTT reduction significantly in comparison to group treated with FeSO4 only (PO.05) . ALT leakage of hepatocytes was also suppressed by pretreated with extract of mungbean, adzuki bean, common bean I and common bean II at proper concentration (P<0.05) . Furthermore, hepatocytes treated with extracts above increased cell survival ability evidently in 24 h. Taken together, results indicated that cytoprotective effects of legume extracts on Fe2+-treated in primary cultured rat hepatocytes might arise from their antioxidative activities.3. Antioxidative effects of main compounds obtained from Chinese legume extractsAntioxidative activities evaluated by Photochemr in vitro revealed that myricetin rutinoside possesed strongest antioxidative activity among compounds obtained from Chinese legume extracts. 1 nmol myricetin rutinoside was tantamount to 5.7 nmol ascorbic acid equivalents.4. Cytoprotective effects of myricetin rutinoside on Fe2+-induced oxidative damage in primary cultured rat hepatocytesPrimary cultured rat hepatocytes pretreated with 50. 100 umol/L myricetin rutinoside increased MTT reduction 1.62 and 2.04 flod in comparison to grouptreated with FeS04 only (PO.01) . ALT leakage of primary cultured rat hepatocytes pretreated with 20, 100 u.mol/L myricetin rutinoside was also decreased to 30.7±3.0 U/L and 23.7±3.7 U/L by comparing with 47.7±8.8 U/L in group treated with FeSO4 only (P<0.05) . Furthermore, MDA decreased to 44% in oxidative damaged primary cultured rat hepatocytes pretreated with 50 u.mol/L myricetin rutinoside compared with Fe +-treated group.5. Effects of myricetin rutinoside on SOD, GSH-Px activity and mRNA expression of Fe2+-induced oxidative damage in primary cultured rat hepatocytesOxidative damaged primary cultured rat hepatocytes pretreated with 50, 100 u.mol/L myricetin rutinoside SOD activity about 1.56 and 1.57 flod in comparison to group treated with FeSO4 only (PO.05) . GSH-Px activity increased 1.64, 1.63 and 2.36 flod in group treated with 20, 50, 100 u,mol/L myricetin rutinoside respectively.The SOD mRNA expression in Fe2+-treated rat hepatocytes was about 49% of control group. SOD mRNA expression in hepatocytes pretreated with 10, 50. 100 umol/L myricetin rutinoside was 60%, 75%, 118%, respectively. Group pretreated with 100 u.mol/L myricetin rutinoside was about 2.4 fold over control in mRNA expression. The GSH-Px mRNA expression in rat hepatocytes treated with FeSO4 was about 63% of control group. GSH-Px mRNA expression in hepatocytes pretreated with 10, 50, lOOuM myricetin rutinoside was 106%, 93%. 114%, respectively. Group pretreated with 100 u.mol/L myricetin rutinoside was about 1.8 fold over control in mRNA expression. These results indicated that myricetin rutinoside increased the activity of SOD and GSH-Px in oxidative damaged primary cultured rat hepatocytes relating to up-regulae SOD and GSH-Px mRNA expression.Conclusion:1. It was concluded that flavonoids were the main compounds in eight Chinese legume extracts. Especially, myricetin rutinoside obtained from adzuki bean was not reported.2. Eight legume extracts have in different strength with the ranked order of adzuki bean > mung bean > common bean II > soybean > common bean I > broad bean > pea > lima bean.3. Extracts obtained from mung bean, adzuki bean, common bean I and common bean II possesed cytoprotective effects in Fe2+-induced oxidative damaged primary cultured hepatocytes. Comparing antioxidative capacity in vitro with cytoprotective effects of extracts , a close positive correlation was found between them.4. Antioxidative activities evaluated by Photochemr in vitro revealed that myricetin rutinoside obtained from adzuki bean possesed strongest free radical scavenging capability among compounds obtained from Chinese legume extracts.5. Cytoprotective mechanisms of myricetin rutinoside in oxidative damaged cultured rat primary hepatocytes was to increase the elimination of free radical and SOD, GSH-Px activity.6. RT-PCR results indicated that increase of SOD and GSH-Px activity in oxidative damaged cultured rat primary hepatocytes might arise from up-regulating the mRNA expression.
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