十三種中草藥水萃物的抗氧化與抗發炎評估

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標題:	十三種中草藥水萃物的抗氧化與抗發炎評估 Evaluation of antioxidant anti-inflammatory of thirteen kinds of Chinese Herbal medicine extract
作者:	湯奇樺
貢獻者:	王柏森黃明星嘉南藥理科技大學：藥物科技研究所
關鍵字:	HPLC 抗發炎抗氧化石榴皮龍眼 Punica granatum Dimocarpus longan antioxidative anti-inflammation HPLC
日期:	2010
上傳時間:	2010-12-30 15:34:23 (UTC+8)
摘要:	近年來，隨著國人健康意識逐漸抬頭，中草藥植物的開發與運用，已成為重要的發展趨勢。本研究從數十種中草藥中，以總多酚含量測定及總黃酮含量測定篩選出富含多酚及類黃酮的材料，以龍眼葉、龍眼枝、龍眼殼、龍眼籽、龍眼乾、大葉千金拔(葉)、大葉千金拔(莖)、大葉千金拔(根)、石榴皮、蒲公英、蔓性千金拔、鵝仔草與白花蛇舌草等十三種為材料。研究分為管內與管外試驗，第一部分探討其DPPH自由基清除率、TEAC總抗氧化能力、還原力、亞鐵離子螯合能力、微脂粒氧化作用、Superoxide自由基清除能力與NO抑制能力等八個抗氧化平台找出較具潛力之藥材，進而建立此類藥材之HPLC化學成份指紋圖譜。第二部分為藉由巨噬細胞(RAW264.7)模式系統來評估中草藥萃取物對LPS誘導一氧化氮(nitric oxide, NO)生成率及分析胞內活性氧 (ROS)含量分析。研究結果顯示，在抗氧化分析方面，發現龍眼枝、龍眼葉及石榴皮有最佳的DPPH自由基清除率、TEAC總抗氧化能力、還原力、亞鐵離子螯合能力、微脂粒氧化作用、Superoxide自由基清除能力與NO抑制能力，在濃度為150 ppm時DPPH自由基清除率均超過95%；在濃度為100 ppm時TEAC總抗氧化能力均超過95%；在濃度為700 ppm時亞鐵離子螫合能力達80％以上；在濃度為150 ppm時其OD700值均超過0.522以上；於濃度為600 ppm時微脂粒氧化作用抑制效果均超過90％以上；在濃度600 ppm時Superoxide自由基清除能力均超過80%；在濃度250 ppm時NO抑制能力抑制效果均超過90%；在總多酚定量試驗，龍眼枝、龍眼葉及石榴皮每克萃取物含有相當於 253.9 mg、208.0 mg與148.0 mg之gallic acid。總類黃酮測定，石榴皮、鵝仔草及蒲公英之每克萃取物含有相當於70.7 mg、68.6 mg、51.0 mg之rutin。酪胺酸酶活性評估顯示，龍眼枝、龍眼葉及石榴皮在濃度2000 ppm時，抑制率均超過85%。龍眼枝、龍眼葉、石榴皮與龍眼殼HPLC的指紋圖譜，顯示含有gallic acid、(-)-epicatechin 、ellagic acid 、 ferulic acid、 quercetin與kaempferol等成份。在抗發炎試驗方面龍眼枝、龍眼籽與龍眼乾在濃度50 ppm 時時有超過50%的抑制效果。十三種中草藥水萃物對處理RAW 264.7 細胞ROS分析中，有五種樣品具有抑制ROS生成效果，依序為鵝仔草、白花蛇舌草、龍眼殼、龍眼葉與大葉千金拔(葉)。在LPS誘導細胞產生ROS方面有十種樣品具有抑制生成效果，依序為鵝仔草、蒲公英、蔓性千金拔、白花蛇舌草、石榴皮、龍眼殼、大葉千金拔(葉)、龍眼籽、龍眼乾與龍眼葉具有抑制ROS生成效果。 In recent years, people pay more attention on the health issues, therefore, the development and application of the Chinese herbals have become the world trend. In this study, 13 Chinese herbal medicine water extracts including the leaf of Dimocarpus longan(LDL), the branch of Dimocarpus longan(BDL), the shell of Dimocarpus longan(SHDL), the seed of Dimocarpus longan(SDL), the fruit Dimocarpus longan(FDL), the leaf of Flemingia macrophylla(LFM), the stem of Flemingia macrophylla(SFM), the root of Flemingia macrophylla(RFM), Punica granatum(PG), Taraxacum mongolicum(TM), Flemingia philippinensis(FP), Pterocypsela indica(PI) and Oldenlandia diffusa(OD) were conducted to determine their antioxidant and anti-inflammatory effects. In Part 1, we focused on the studies of the antioxidant activities, include the scavenging effect on α,α-diphenyl-β-picrylhydrazyl(DPPH) radicals, the trolox equivalent antioxidant capacity (TEAC), reducing power, ferrous ion chelating power, inhibition NO capacity, scavenging superoxide radical capacity and inhibitory effect on lipid peroxidation. The HPLC-UV finger print chromatogram and antityrosinase effects of the potent herbs were also measured. In part two, we investigated the effects of Chinese herbals, on RAW 264.7 macrophages production NO induced by LPS. The results show that all these herbal extracts demonstrate antioxidant abilities and in dose-dependent manner. BDL, LDL, and PG have the greatest DPPH radical scavenging effect. BDL, LDL, and PG have better antioxidative activity by TEAC. The ferrous ion chelating capacity and reducing power of the BDL, LDL, and PG have the best effect. Inhibition NO capacity of BDL, LDL, and PG have the better inhibition effect. Scavenging superoxide radical capacity of the BDL, LDL, and PG have the greatest superoxide radical scavenging effect. BDL, LDL, and PG have the better effect of inhibitory effect on lipid peroxidation. BDL, LDL, and PG had total polyphenol contents (253.9 mg、208.0 mg and 148.0 mg of gallic acid equivalent, respectively). PG, PI and TM had total flavonoid contents (70.7 mg、68.6 mg and 51.0 mg of rutin equivalent, respectively). BDL, LDL, and PG exhibit potent inhibitory effects on tyrosinase. A high performance liquid chromatography analysis revealed that phenolic acids and flavonoids, such as gallic acid, (-)-epicatechin , ellagic acid , ferulic acid, quercetin and kaempferol . We found that three extracts of Chinese herbs could inhibit nitric oxide generation induced by LPS by 50% or more in RAW 264.7 cells at a concentration of 50 µg/ml including BDL, SHDL and FDL has the best inhibitory effect. We found that five extracts of Chinese herbs could inhibit ROS production of RAW 264.7 cell including PI, OD, SHDL, LDL and LFM. That ten extracts of Chinese herbs could inhibit ROS production generations induced by LPS in RAW 264.7 cell including PI, TM, FP, OD, PG, SHDL, LFM, SDL, FDL and LDL.
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