Chamberlain Casey A, Hatch Marguerite, Garrett Timothy J
Department of Pathology, Immunology and Laboratory Medicine, University of Florida, Gainesville, FL 32610, USA.
Metabolites. 2021 May 11;11(5):308. doi: 10.3390/metabo11050308.
Paper spray ionization mass spectrometry (PSI-MS) is a direct MS analysis technique with several reported bacterial metabolomics applications. As with most MS-based bacterial studies, all currently reported PSI-MS bacterial analyses have focused on the chemical signatures of the cellular unit. One dimension of the bacterial metabolome that is often lost in such analyses is the exometabolome (extracellular metabolome), including secreted metabolites, lipids, and peptides. A key component of the bacterial exometabolome that is gaining increased attention in the microbiology and biomedical communities is extracellular vesicles (EVs). These excreted structures, produced by cells in all domains of life, contain a variety of biomolecules responsible for a wide array of cellular functions, thus representing a core component of the bacterial secreted metabolome. Although previously examined using other MS approaches, no reports currently exist for a PSI-MS analysis of bacterial EVs, nor EVs from any other organism (exosomes, ectosomes, etc.). PSI-MS holds unique analytical strengths over other commonly used MS platforms and could thus provide an advantageous approach to EV metabolomics. To address this, we report a novel application representing, to our knowledge, the first PSI-MS analysis of EVs from any organism (using the human gut resident as the experimental model, a bacterium whose EVs were never previously investigated). In this report, we show how we isolated and purified EVs from bacterial culture supernatant by EV-specific affinity chromatography, confirmed and characterized these vesicles by nanoparticle tracking analysis, analyzed the EV isolate by PSI-MS, and identified a panel of EV-derived metabolites, lipids, and peptides. This work serves as a pioneering study in the field of MS-based EV analysis and provides a new, rapid, sensitive, and economical approach to EV metabolomics.
纸喷雾电离质谱法（PSI-MS）是一种直接质谱分析技术，已有多项报道将其应用于细菌代谢组学研究。与大多数基于质谱的细菌研究一样，目前所有报道的PSI-MS细菌分析都集中在细胞单元的化学特征上。在这类分析中经常被忽略的细菌代谢组的一个维度是胞外代谢组（细胞外代谢组），包括分泌的代谢物、脂质和肽。细菌胞外代谢组的一个关键组成部分是细胞外囊泡（EVs），它在微生物学和生物医学领域越来越受到关注。这些由生命各领域的细胞产生的分泌结构含有多种负责广泛细胞功能的生物分子，因此代表了细菌分泌代谢组的核心组成部分。尽管此前已使用其他质谱方法进行过研究，但目前尚无关于细菌EVs的PSI-MS分析的报道，也没有关于任何其他生物体（外泌体、微囊等）的EVs的报道。与其他常用的质谱平台相比，PSI-MS具有独特的分析优势，因此可以为EV代谢组学提供一种有利的方法。为了解决这个问题，我们报告了一项新的应用，据我们所知，这是对任何生物体的EVs进行的首次PSI-MS分析（以人体肠道细菌作为实验模型，该细菌的EVs此前从未被研究过）。在本报告中,我们展示了如何通过EV特异性亲和色谱从细菌培养上清液中分离和纯化EVs，通过纳米颗粒跟踪分析对这些囊泡进行确认和表征，用PSI-MS分析EV分离物，并鉴定了一组源自EV的代谢物、脂质和肽。这项工作是基于质谱的EV分析领域的开创性研究，为EV代谢组学提供了一种新的、快速、灵敏且经济的方法。
