Laboratory of Biochemistry, Wageningen University, Wageningen, The Netherlands.
Magn Reson Chem. 2011 Dec;49 Suppl 1:S55-60. doi: 10.1002/mrc.2833.
In many metabolomics studies, metabolite identification by mass spectrometry (MS) often is hampered by the lack of good reference compounds, and hence, NMR information is essential for structural elucidation, especially for the very large group of secondary metabolites. The classical approach for compound identification is to perform time-consuming and laborious HPLC fractionations and purifications, before (re)dissolving the molecules in deuterated solvents for NMR measurements. Hence, a more direct and easy purification protocol would save time and efforts. Here, we propose an automated MS-guided HPLC-MS-solid phase extraction-NMR approach, which was used to fully characterize flavonoid structures present in crude tomato plant extracts. NMR spectra of plant metabolites, automatically trapped and purified from LC-MS traces, were successfully obtained, leading to the structural elucidation of the metabolites. The MS-based trapping enabled a direct link between the mass signals and NMR peaks derived from the selected LC-MS peaks, thereby decreasing the time needed for elucidation of the metabolite structures. In addition, automated 1H NMR spectrum fitting further speeded up the candidate rejection process. Our approach facilitates the more rapid unraveling of yet unknown metabolite structures and can therefore make untargeted metabolomics approaches more powerful.
在许多代谢组学研究中，质谱（MS）对代谢物的鉴定常常受到缺乏良好参考化合物的阻碍，因此，NMR 信息对于结构阐明是必不可少的，特别是对于非常大的次生代谢物组。化合物鉴定的经典方法是在（重新）将分子溶解在氘代溶剂中进行 NMR 测量之前，进行耗时且费力的 HPLC 分级和纯化。因此，更直接和简单的纯化方案将节省时间和精力。在这里，我们提出了一种自动化 MS 引导的 HPLC-MS-固相萃取-NMR 方法，该方法用于充分表征粗番茄植物提取物中的类黄酮结构。成功获得了从 LC-MS 痕迹中自动捕获和纯化的植物代谢物的 NMR 谱，从而对代谢物的结构进行了阐明。基于 MS 的捕获能够在从所选 LC-MS 峰衍生的质量信号和 NMR 峰之间建立直接联系，从而减少了解析代谢物结构所需的时间。此外，自动 1H NMR 光谱拟合进一步加快了候选物的排除过程。我们的方法促进了对未知代谢物结构的快速揭示，因此可以使非靶向代谢组学方法更加强大。
