Simultaneous measurement of proline and related compounds in oak leaves by high-performance ligand-exchange chromatography and electrospray ionization mass spectrometry for environmental stress studies
Simultaneous measurement of proline and related compounds in oak leaves by high-performance ligand-exchange chromatography and electrospray ionization mass spectrometry for environmental stress studies
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DOI：
10.1016/j.chroma.2008.12.030
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发表时间：
2009-02-13
影响因子：
4.1
通讯作者：
Guignard, Cedric
中科院分区：
文献类型：
--
作者：
Oufir, Mouhssin;Schulz, Nadine;Guignard, Cedric
采用高效配体交换液相色谱（HPLEC）和质谱联用技术，以N-乙酰脯氨酸为内标物，对干旱胁迫下橡树叶片中的脯氨酸、羟脯氨酸、甲基脯氨酸、甜菜碱和胡芦巴碱等胁迫相关物质进行同时分析。使用Aminex HPX-87 C色谱柱分析甲醇/氯仿/水浸提液，并通过电喷雾电离质谱（ESI-MS）的正离子模式在单离子监测（SIM）模式下进行特异性定量。在一天内的研究中，N-乙酰脯氨酸添加到橡树叶提取物中的回收率范围为85.2%至122.1%。标准校准曲线在0.3125 ~ 10 μ mol L-1范围内呈良好的线性关系，胡芦巴碱的相关系数最低为0.99961。这种替代程序的优点，以前公布的方法相比，使用荧光或安培检测，是同时和直接检测的的crosslinkedprotectants在一个单一的色谱运行，最小的样品制备，良好的特异性和减少的定量限，范围从0.1到0.6 μ mol L-1。五十六天的水分亏缺导致叶片游离脯氨酸水平增加（2.4倍，P
A mass spectrometer was coupled to high-performance ligand-exchange liquid chromatography (HPLEC) for simultaneous analysis of stress associated solutes such as proline, hydroxyproline, methylproline, glycine betaine and trigonelline extracted from leaves of drought stressed oaks and an internal standard namely N-acetylproline. Methanol/chloroform/water extracts were analyzed using an Aminex HPX-87C column and specifically quantified by the positive ion mode of an electrospray ionisation-mass spectrometry (ESI-MS) in single ion monitoring (SIM) mode. The recovery of N-acetyl proline added to oak leaf extracts ranged from 85.2 to 122.1% for an intra-day study. Standard calibration curves showed good linearity in the measured range from 0.3125 to 10 mu mol L-1 with the lowest correlation coefficient of 0.99961 for trigonelline. The advantages of this alternative procedure, compared to previously published methods using fluorescence or amperometric detections, are the simultaneous and direct detection of osmoprotectants in a single chromatographic run, a minimal sample preparation, a good specificity and reduced limits of quantification, ranging from 0.1 to 0.6 mu mol L-1. Fifty-six days of water deficit exposure resulted in increased foliar free proline levels (2.4-fold, P