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毛细管电泳-电喷雾电离-质谱法测定氨基酸:不同蛋白质水解程序的评估

Determination of amino acids by capillary electrophoresis-electrospray ionization-mass spectrometry: an evaluation of different protein hydrolysis procedures.

作者信息

Simionato Ana V Colnaghi, Moraes Edgar Perin, Carrilho Emanuel, Tavares Marina Franco Maggi, Kenndler Ernst

机构信息

Institute of Chemistry, University of São Paulo, São Paulo, Brazil.

出版信息

Electrophoresis. 2008 May;29(10):2051-8. doi: 10.1002/elps.200700629.

Abstract

In this work, a CE equipment, online hyphenated to an IT MS analyzer by a linear sheath liquid interface promoting ESI, was used to develop a method for quantitative determination of amino acids. Under appropriate conditions (BGE composition, 0.8% HCOOH, 20% CH3OH; sheath liquid composition, 0.8% HCOOH, 60% methanol; V ESI, +4.50 kV), analytical curves of all amino acids from 3 to 80 mg/L were recorded presenting acceptable linearity (r >0.99). LODs in the range of 16-172 micromol/L were obtained. BSA, a model protein, was submitted to different hydrolysis procedures (classical acid and basic, and catalyzed by the H+ form of a cation exchanger resin) and its amino acid profiles determined. In general, the resin-mediated hydrolysis yields were overall similar or better than those obtained by classical acid or basic hydrolysis. The resulting experimental-to-theoretical BSA concentration ratios served as correction factors for the quantitation of amino acids in Brazil nut resin generated hydrolysates.

摘要

在本研究中,使用了一台通过促进电喷雾电离(ESI)的线性鞘液接口与离子阱质谱(IT MS)分析仪在线联用的毛细管电泳(CE)设备,开发了一种定量测定氨基酸的方法。在适当条件下(背景电解质(BGE)组成:0.8% 甲酸,20% 甲醇;鞘液组成:0.8% 甲酸,60% 甲醇;电喷雾电离电压(V ESI),+4.50 kV),记录了浓度范围为3至80 mg/L的所有氨基酸的分析曲线,呈现出可接受的线性关系(r >0.99)。检测限在16 - 172 μmol/L范围内。将牛血清白蛋白(BSA)这种模型蛋白进行不同的水解程序(经典的酸水解和碱水解,以及由阳离子交换树脂的H⁺形式催化水解),并测定其氨基酸谱。总体而言,树脂介导的水解产率总体上与经典酸水解或碱水解获得的产率相似或更高。所得的实验浓度与理论浓度的BSA比值用作巴西坚果树脂水解产物中氨基酸定量的校正因子。

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