利用毛細管微胞電動層析法線上偵測類黃酮抗氧化能力

Direct On-line analysis of flavonoids and coesponsive antioxidative capacities by micellar electrokinetic chromatography.

韓尚祐(San-Yu Han)

指導教授 ： 吳立真

國立暨南國際大學/科技學院/應用化學系/碩士(2010年)

摘要

黃酮類的抗氧化性對於人體健康有諸多益處。目前已有相關偵測方法評估各式抗氧化物質之抗氧化能力，例如 ABTS+, DPPH, TEAC, ORAC等。這些方法多為離線式測量，若要測量混和物中各成分之抗氧化能力，便遇到瓶頸。此外，黃酮類中常見帶電相同，但具極性差異之結構衍生物，造成分離效果不佳。而微胞電動力層析法則可克服此缺點，同時亦可提高偵測極限。因此，為達到線上偵測抗氧化物之抗氧化能力。本實驗使用自由ABTS+後衍生系統搭配微胞電動層析(Micellar electrokinetic chromatography; MEKC)分析具自由基抓取能力之抗氧化物。本實驗之設計分為兩大部分，前半段為抗氧化物質之毛細管微胞電動力層析，先進行成分(例如，黃酮類)分離，之後再串聯ABTS+流洗系統檢測其清除ABTS+自由基抗氧化能力。並在毛細管後衍生系統探討不同緩衝液濃度、pH值、SDS濃度等變因,在最佳化條件下，ABTS+試劑成功與微胞電動力層析結合。結果顯示，在緩衝溶液[borate-HCl]=10mM，pH=9，電壓為20kV之條件下，可有效分離quercetin、rutin、hesperetin、kaempferol、catechin、apigenin 、naringenin、genistein八種抗氧化物，並在ABTS+流洗系統為[ABTS]=7mM加入2.45mM氧化劑K2S2O8的條件下同時偵測其抗氧化能力。此外，本系統之結果與一般離線方法比較，靈敏度較高。

關鍵字

微胞電動層析 ； 黃酮類 ； 抗氧化能力 ； 線上偵測 ； 後衍生

並列摘要

It is well known that the bioactivities of flavonoids benefit human health in several ways. One of the most characterized bioactivities of flavonoids is their antioxidative capacity. Various approaches have been developed to evaluate this property, including ABTS+, DPPH, TEAC, and ORAC analysis. The requirements of these approaches may differ markedly. Thus, they can be applied to measure the antioxidative capacity of flavonoids with different polarity. However, these approaches can determine the total antioxidative capacity of a single molecule or mixtres, but not simultaneously analyzing a single component of a mixture. Herein, we developed a system to measure the total and individual antioxidative capacity of mixtures at once. A micelle electrokinetic chromatography (MEKC) capillary electrophoresis system coupling with ABTS+ analysis was employed to meet the requirements. The system was constituted of two sections. The fist section was to separate analytes by MEKC (detecting at A254nm). The second section was coupling to a flow injection system to inject ABTS+ solution to oxidize the analytes for the following determination at A734nm. Eight falvonoids (quercetin, rutin, hesperetin, kaempferol, catechin, apigenin, naringenen, genistein) was used as standards to measure their total and individual antioxidative capacity. The factors for optimization of the system included BGE concentration, pH, and SDS concentrations. Our results indicated that these eight flavonoids were successfully separated and the total and individual antioxidative capacities were determined with a correlation coefficient of 0.98 to their off-line measurements. The optimal conditions were 10mM borate-Hcl buffer at pH 9.0, 20kV. Moreover, the sensitivity of this system was better than the off-line counterparts.

並列關鍵字

MEKC ； ABTS ； flavonoid ； antioxidative ability

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