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建立了微流控芯片毛细管柱后扩散衍生激光诱导荧光检测氨基酸的方法。利用微流控芯片的二维平面结构特征,在分离通道末端增加支通道,通过扩散法引入柱后衍生试剂,避免了电压引入法对分离通道流型的影响,因而提高了分离效率。考察了支通道长度、衍生试剂液面高度、检测点位置对衍生结果的最优条件,考察了衍生试剂引入方法、催化剂种类、缓冲溶液种类对检测结果的影响。用20 mmol/L硼砂-NaOH(pH=10)溶液作为电泳缓冲溶液,与柱后衍生试剂1.0 mmol/L NDA+8.0 mmol/L 2-ME+35 mmol/L硼砂(pH 10.0)的30%(V/V)甲醇溶液反应,精氨酸、苯丙氨酸、天冬酰胺、脯氨酸、丙氨酸、甘氨酸在不加任何添加剂的情况下可达到基线分离。本法用于板蓝根药材中主要游离氨基酸的分离检测,相对标准偏差小于4.4%(n=5),回收率为92.3%~98.6%。所测板蓝根药材中精氨酸和脯氨酸含量分别为14.97,8.02 mg/g。
A method for the determination of amino acids by microfluidic chip capillary column post-diffusion derivatization laser induced fluorescence was established. Using the two-dimensional planar structure of the microfluidic chip, branch channels are added at the end of the separation channel, and post-column derivatization reagents are introduced by the diffusion method to avoid the influence of the voltage introduction method on the flow channel of the separation channel, thus improving the separation efficiency. The optimal conditions for the derivatization results of the branch channel length, the height of the derivatization reagent and the position of the detection point were investigated. The effects of introduction methods, catalyst types and buffer solution types on the test results were investigated. A 20% aqueous solution of 20 mmol / L borax-NaOH (pH = 10) was used as electrophoresis buffer to react with 30% of post-column derivatization reagent 1.0 mmol / L NDA + 8.0 mmol / L 2-ME + 35 mmol / L borax (pH 10.0) (V / V) methanol solution, arginine, phenylalanine, asparagine, proline, alanine and glycine can be baseline separated without any additives. The method was applied to the determination of main free amino acids in Radix isatidis. The relative standard deviations (RSDs) were less than 4.4% (n = 5) and the recoveries were 92.3% ~ 98.6%. The contents of arginine and proline in Radix isatidis were 14.97 and 8.02 mg / g, respectively.