Studies on the Antioxidative Activities of Longan (Dimorcarpus longan Lour.) Flower Extracts
|Keywords:||龍眼花;抗氧化;Longan flower;antioxidant||Issue Date:||2005||Abstract:||
本實驗龍眼花之萃取方式為沸點下熱水迴流萃取及室溫下以95%乙醇、甲醇、乙酸乙酯與正己烷四種溶劑之攪拌萃取。再以七種抗氧化活性評估方法檢測此五種龍眼花不同溶劑之粗萃物，包括清除DPPH自由基、TEAC (Trolox equivalent antioxidant capacity)、氧自由基吸收能力 (Oxygen radical absorbance capacity, ORAC)、還原力、抑制微脂粒 (liposome) 過氧化、抑制銅離子誘導低密度脂蛋白 (low density lipoprotein, LDL) 氧化與以TGF-β1 (transforming growth factor-β1) 誘導肝癌細胞Hep 3B產生氧化損傷之保護效果。
實驗結果顯示，龍眼花水粗萃物於各抗氧化實驗中皆展現較高之抗氧化力，其清除DPPH自由基之EC50為3.69±0.16 μg/mL，TEAC值為1.22 mM Trolox equiv.，ORAC值為7.28 mM Trolox equiv.，還原力方面於50 μg/mL時A700 =1.84，抑制微脂粒過氧化之IC50為54.46±1.61 μg/mL，且延緩銅離子誘導LDL氧化之效果最佳，為同濃度下Trolox之0.62倍；以TGF-β1誘導肝癌細胞Hep 3B產生氧化損傷實驗中，其於100 μg/mL濃度時達約64.3±3.4%的回復率。整體而言抗氧化活性以水粗萃物最佳，其次為甲醇、乙醇與乙酸乙酯粗萃物，而正己烷粗萃物則幾乎無抗氧化效果。
進一步分析龍眼花不同溶劑粗萃物之抗氧化成分，結果顯示，水粗萃物中的總酚類化合物 (548.2±12.7 mg gallic acid equivalent / g of dry weight) 與没食子酸 (10.4 ± 0.3 mg gallic acid /g of dry weight) 含量最多，因此推測其抗氧化力可能來自於龍眼花中之酚類物質。再進ㄧ步將龍眼花水萃物經Sephadex LH-20管柱層析分離得到八個區分層，並進行抑制銅離子誘導LDL氧化測定，結果以fraction 7之100%甲醇沖提區分層之效果最佳。本研究結果顯示龍眼花中提供抗氧化性的有效成分可能為一些極性較高的酚類物質。
Recently, many chronic diseases such as cancer, atherosclerosis, and aging were found to be associated with oxidative damage. The imbalance between the concentration of reactive oxygen and nitrogen species and defense mechanisms of the body would cause oxidative damage, and antioxidants enhancement would reduce oxidative damage. Therefore, dietary supplementation of antioxidants was thought to be beneficial to health. The purpose of this study is to locate the most antioxidative solvent extract of Longan (Dimorcarpus longan Lour.) flower by various antioxidative assays, and to further investigate the most efficient antioxidative fraction after chromatographic separation by the assay of Cu2+-induced low density lipoprotein (LDL) oxidation.
First, the crude extracts were prepared by extracting Longan flower with boiling water and four solvents (95% ethanol, methanol, ethyl acetate and n-hexane) at room temperature. Then the five different solvent extracts of Longan flower were tested for various antioxidative assays, including DPPH free radical scavenging effect, Trolox equivalent antioxidant capacity (TEAC) assay, oxygen radical absorbance capacity (ORAC) assay, reducing power, inhibition of peroxidation in a liposome model system, Cu2+-induced oxidation of human LDL and recovery effect of TGF-β1 induced oxidative damage in Hep 3B cells.
The results of antioxidative assays revealed that the best effect was exhibited by the water extract, followed by methanol, ethanol, ethyl acetate and n-hexane extracts of Longan flower. The EC50 value of water extract in scavenging DPPH radicals was 3.69±0.16 μg/mL. Results of TEAC and ORAC assays revealed that water extract gave the highest TEAC value (1.22 mM Trolox equivalent) and ORAC value (7.28 mM Trolox equivalent). With regard to reducing power, at sample concentration of 50 μg/mL, water extract gave the best effect. Concerning inhibition of peroxidation in a liposome model system, the IC50 of water extract in inhibiting the peroxidation was 54.46±1.61 μg/mL, which was the lowest among all the extracts. As for the effect of Cu2+-induced oxidation of human LDL, water extract showed the best effect in delaying LDL oxidation. Finally, water extract gave the best recovery effect of TGF-β1 induced oxidative damage in Hep 3B cells at concentration of 100 μg/mL.
Water extract of Longan flower contained the most abundant amount of total polyphenol (548.2±12.7 mg gallic acid equivalent / g of dry weight) and gallic acid (10.4 ± 0.3 mg gallic acid /g of dry weight). According to the results of antioxidative assays, water extract showed the highest antioxidant capacity and we presumed that this could probably be related to the phenolic compounds. Then Sephadex LH-20 gel chromatography was employed to fractionate the water extract; eight fractions were obtained and tested for Cu2+-induced oxidation of human LDL. The result of antioxidative experiment revealed that fraction 7 showed the best effect in delaying LDL oxidation. We presumed that antioxidative capacity of Longan flower could probably be related to phenol compounds having higher polarity.
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