Department of Pharmacy, Analytical Biochemistry Group, University of Groningen , Antonius-Deusinglaan 1, Building Code XB20, level 6, Groningen, 9713 AV, The Netherlands.
Anal Chem. 2014 May 20;86(10):5089-100. doi: 10.1021/ac500810r. Epub 2014 May 5.
The identification of unknown compounds remains to be a bottleneck of mass spectrometry (MS)-based metabolomics screening experiments. Here, we present a novel approach which facilitates the identification and quantification of analytes containing aldehyde and ketone groups in biological samples by adding chemical information to MS data. Our strategy is based on rapid autosampler-in-needle-derivatization with p-toluenesulfonylhydrazine (TSH). The resulting TSH-hydrazones are separated by ultrahigh-performance liquid chromatography (UHPLC) and detected by electrospray ionization-quadrupole-time-of-flight (ESI-QqTOF) mass spectrometry using a SWATH (Sequential Window Acquisition of all Theoretical Fragment-Ion Spectra) data-independent high-resolution mass spectrometry (HR-MS) approach. Derivatization makes small, poorly ionizable or retained analytes amenable to reversed phase chromatography and electrospray ionization in both polarities. Negatively charged TSH-hydrazone ions furthermore show a simple and predictable fragmentation pattern upon collision induced dissociation, which enables the chemo-selective screening for unknown aldehydes and ketones via a signature fragment ion (m/z 155.0172). By means of SWATH, targeted and nontargeted application scenarios of the suggested derivatization route are enabled in the frame of a single UHPLC-ESI-QqTOF-HR-MS workflow. The method's ability to simultaneously quantify and identify molecules containing aldehyde and ketone groups is demonstrated using 61 target analytes from various compound classes and a (13)C labeled yeast matrix. The identification of unknowns in biological samples is detailed using the example of indole-3-acetaldehyde.
未知化合物的鉴定仍然是基于质谱（MS）的代谢组学筛选实验的瓶颈。在这里，我们提出了一种新的方法，通过向 MS 数据中添加化学信息，促进了生物样品中含醛和酮基团的分析物的鉴定和定量。我们的策略基于快速自动进样针内衍生化与对甲苯磺酰基腙（TSH）。所得的 TSH 腙通过超高效液相色谱（UHPLC）分离，并通过电喷雾电离-四极杆飞行时间（ESI-QqTOF）质谱仪，采用 SWATH（顺序窗口采集所有理论片段离子谱）数据非依赖性高分辨率质谱（HR-MS）方法进行检测。衍生化使小、电离性差或保留的分析物适用于反相色谱和电喷雾电离在两种极性下。带负电荷的 TSH 腙离子在碰撞诱导解离时还表现出简单且可预测的碎裂模式，这使得通过特征碎片离子（m/z 155.0172）能够进行未知醛和酮的化学选择性筛选。通过 SWATH，在单个 UHPLC-ESI-QqTOF-HR-MS 工作流程的框架内，可实现所建议衍生化途径的靶向和非靶向应用场景。该方法同时定量和鉴定含醛和酮基团的分子的能力通过来自各种化合物类别的 61 个目标分析物和（13）C 标记酵母基质得到证明。通过吲哚-3-乙醛的实例详细说明了生物样品中未知物的鉴定。
