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標題: 以毛細管電泳結合堆積線上濃縮方法分析γ-胺基丁酸及丙胺酸方法開發及其應用
On-line derivatization and stacking capillary electrophoretic analysis of γ-aminobutyric acid and alanine and its applications.
作者: 林雅品
Lin, Yar-Ping
關鍵字: Capillary electrophoresis;毛細管電泳;On-column derivatization;GABA;alanine;Stacking.;線上衍生化;胺基丁酸;丙胺酸
出版社: 化學系所
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γ-胺基丁酸( γ-Aminobutyric acid,簡稱GABA ) 在人體中是一種重要的神經傳導物質,具有降血壓、調節心律失常、調節激素分泌等作用。而在植物中,其主要作用是調節植物中碳氮比,植物中的麩胺酸( Glutamate )轉換成琥珀酸時便會生成GABA,所以此過程被稱為GABA分路(GABA shunt),而丙胺酸( Alanine )則是在GABA分路過程中的產物,故植物中也可測出丙胺酸。
本研究中利用鄰苯二甲醛(o-Phthalaldehyde,OPA)及2-硫基乙醇 (2-Mercaptoethanol,2-ME)與γ-胺基丁酸進行衍生化反應並生成螢光衍生物,再經由毛細管電泳/螢光偵測分析。由於鄰苯二甲醛的衍生化反應快速,故採用管柱內( in-capillary )衍生化,使樣品在管柱內直接衍生化並分離,於最佳衍生化條件下分析γ-胺基丁酸及丙胺酸,其線性濃度範圍介於0.05-5.0µM,線性相關係數分別為0.9995和0.9964,偵測極限各為4nM和20nM。
故利用鄰苯二甲醛(o-Phthalaldehyde,OPA)及2-硫基乙醇 (2-Mercaptoethanol,2-ME)與γ-胺基丁酸進行衍生化並結合線上濃縮,可以成功的將分析物γ-胺基丁酸定量。此方法簡單且快速,利用該方法來偵測市售含有γ-胺基丁酸成分之茶包及中藥會有良好的再現性與靈敏度。

An on-column derivatization capillary electrophoresis (CE) technique has been developed to determine γ-aminobutyric acid (GABA) and alanine (Ala) in plants. GABA and Ala were derivatized with o-phthaldialdehyde / 2-mercapto-ethanol (OPA/2-ME) reproducibly to fluorescence-labeled products, and then separated by CE and monitored with a high-sensitive fluorescence detector. The labeled derivatization and CE separation conditions were optimized. The labeled derivatization with 20mM of OPA and 26.67 mM of 2-ME (mole ratio=0.75) at pH 10 offered the best sensitive detection, and the separation with 30 mM sodium tetraborate buffer (pH 10) under 21 kV achieved good selectivity within 14 min. The detection is linear in the range of 0.05 - 5µM with correlation coefficients (γ2) above 0.9960 for both GABA and Ala with detection limits of 4 and 20 nM for GABA and Ala, respectively. The recoveries were 92.47% (3.58%RSD) and 92.25% (6.46% RSD) for five determinations of GABA and Ala, respectively. The contents of GABA and Ala in teas from different manufacture processes were detected.
The on-line stacking preconcentration was also used in the analysis of GABA and Ala in Chinese herbs, posterior to a dilution of extract solution for suppressing the matrix effect. With the on-line stacking at the optium condition, the linear range of GABA and Ala are from 5.0nM to 2.5µM with the correlation coefficients (r2) above 0.9992. The detection limit are 0.7nM and 0.8nM for GABA and Ala, respectively. By the in-capillary derivatization couple with stacking, it can be successful to detect the GABA and Ala in the food sample.
其他識別: U0005-1008200615590900
Appears in Collections:化學系所

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