Analysis of plant nucleotide sugars by hydrophilic interaction liquid chromatography and tandem mass spectrometry
Analysis of plant nucleotide sugars by hydrophilic interaction liquid chromatography and tandem mass spectrometry
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DOI：
10.1016/j.ab.2013.11.026
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发表时间：
2014-03-01
影响因子：
2.9
通讯作者：
Heazlewood, Joshua L.
中科院分区：
文献类型：
--
作者：
Ito, Jun;Herter, Thomas;Heazlewood, Joshua L.
对植物细胞壁生物合成中涉及的复杂代谢过程的理解受到对许多相关化合物进行灵敏定量的困难的限制。亲水相互作用液相色谱是一种用于分析复杂生物提取物中的亲水代谢物的有用技术，并且构成了该方法量化植物细胞壁前体的基础。两性离子二氧化硅基固定相已用于从毫克量的叶组织中分离参与细胞壁生物合成的亲水性核苷酸糖。使用以选定的反应监测模式运行的串联质谱法来定量核苷酸糖。该方法具有高度可重复性，可定量低飞摩尔量的 12 核苷酸糖，线性响应高达四个数量级至数百皮摩尔。该方法还成功应用于分析两种模型植物物种的纯化叶提取物，其细胞壁糖组成存在差异，并表明 12 种核苷酸糖中的 6 种水平存在显着差异。植物核苷酸糖提取程序被证明具有良好的回收率和最小的基质效应。与目前使用的技术相比，该方法在应用于植物样品时可显着提高灵敏度。 (C) 2013 Elsevier Inc. 保留所有权利。
Understanding the intricate metabolic processes involved in plant cell wall biosynthesis is limited by difficulties in performing sensitive quantification of many involved compounds. Hydrophilic interaction liquid chromatography is a useful technique for the analysis of hydrophilic metabolites from complex biological extracts and forms the basis of this method to quantify plant cell wall precursors. A zwitterionic silica-based stationary phase has been used to separate hydrophilic nucleotide sugars involved in cell wall biosynthesis from milligram amounts of leaf tissue. A tandem mass spectrometry operating in selected reaction monitoring mode was used to quantify nucleotide sugars. This method was highly repeatable and quantified 12 nucleotide sugars at low femtomole quantities, with linear responses up to four orders of magnitude to several 100 pmol. The method was also successfully applied to the analysis of purified leaf extracts from two model plant species with variations in their cell wall sugar compositions and indicated significant differences in the levels of 6 out of 12 nucleotide sugars. The plant nucleotide sugar extraction procedure was demonstrated to have good recovery rates with minimal matrix effects. The approach results in a significant improvement in sensitivity when applied to plant samples over currently employed techniques. (C) 2013 Elsevier Inc. All rights reserved.