Jorge Tiago F, Florêncio Maria H, António Carla
Plant Metabolomics Laboratory, Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa (ITQB NOVA), Av. da República, 2780-157, Oeiras, Portugal.
Centro de Química e Bioquímica, Faculdade de Ciências, Universidade de Lisboa, Campo Grande, 1749-016, Lisbon, Portugal.
Methods Mol Biol. 2017;1631:279-293. doi: 10.1007/978-1-4939-7136-7_17.
Drought is a major limiting factor in agriculture and responsible for dramatic crop yield losses worldwide. The adjustment of the metabolic status via accumulation of drought stress-responsive osmolytes is one of the many strategies that some plants have developed to cope with water deficit conditions. Osmolytes are highly polar compounds, analysis of whcih is difficult with typical reversed-phase chromatography. Porous graphitic carbon (PGC) has shown to be a suitable alternative to reversed-phase stationary phases for the analysis of highly polar compounds typically found in the plant metabolome. In this chapter, we describe the development and validation of a PGC-based liquid chromatography tandem mass spectrometry (LC-MS) method suitable for the target analysis of water-soluble carbohydrates, such as raffinose family oligosaccharides (RFOs). We present detailed information regarding PGC column equilibration, LC-MS system operation, data analysis, and important notes to be considered during the steps of method development and validation.
干旱是农业中的一个主要限制因素，在全球范围内导致农作物产量大幅损失。通过积累干旱胁迫响应性渗透溶质来调节代谢状态，是一些植物为应对水分亏缺条件而发展出的众多策略之一。渗透溶质是高度极性的化合物，用典型的反相色谱法分析这些化合物很困难。多孔石墨化碳（PGC）已被证明是反相固定相的一种合适替代品，可用于分析植物代谢组中常见的高度极性化合物。在本章中，我们描述了一种基于PGC的液相色谱串联质谱（LC-MS）方法的开发和验证，该方法适用于对水溶性碳水化合物（如棉子糖家族寡糖，RFOs）进行靶向分析。我们提供了有关PGC柱平衡、LC-MS系统操作、数据分析以及方法开发和验证步骤中需考虑的重要注意事项的详细信息。
