Exploring Acoustic Disruption of Lipid Assemblies for Mass Spectrometry

在化学系化学测量和成像项目的支持下，南佛罗里达大学的Theresa Evans-Nguyen和Venkat Bhethanabotla及其团队正在设计改进对细胞外信号至关重要的脂质组装的分析方法。离散脂质在这些组合中的作用尚未得到很好的理解。此外，由于脂质很容易聚集，甚至很难识别和定量这种组合的单个脂质成分。质谱（MS）提供了一个强大的分析工具，如果能够克服聚集的挑战，它可以提供大量有用的信息。为此，本项目侧重于利用受控声波破坏脂质聚集，从而促进质谱分析。本项目中开发的原则应可扩展到其他非特定组件，并可与当前的MS采样方法耦合。该研究涉及化学和工程领域的跨学科努力，并有望通过这两个领域加强STEM招聘计划。本研究的核心是在MS工作流程中使用新型表面声波（SAW）技术。在一种模式中，saw已经可以溶解称为外泌体的细胞外囊泡。此外，正如该团队先前所证明的那样，人们可以使用saw来减轻微流体传感器上非特异性结合造成的生物污垢。推而广之，saw不仅可以作为模型聚集体破坏双层膜，还可以进一步分解双层膜。在第二种模式中，SAW雾化（SAWN）对极性和非极性脂类都表现出独特的电离能力。通过耦合电晕放电（CD），南佛罗里达团队已经取得了与电喷雾电离（ESI）相当的效率。因此，使用SAWN-CD同时分析非极性胆固醇和极性磷脂具有ESI缺乏的特定能力。已知saw可以破坏细胞外囊泡并减轻非特异性生物污染。在这项工作中，SAW被用于将模型双层膜囊泡分解成单个脂质成分，并通过SAW雾化（SAWN），通过耦合电晕放电（CD）促进极性和非极性脂质的MS电离。具体目标是：(1)脂质体作为模型聚集体的配方，以便对saw进行系统评估；(2) SAW破坏参数对脂质组装分解的研究；3)研究SAW雾化参数，实现极性和非极性物质的同时电离。因此，通过这项研究，该团队的目标是开发一种可以应用于细胞外囊泡的方法，允许它们通过质谱直接采样，大大缩短了这种复杂组件的传统分析方法。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

With support from the Chemical Measurement and Imaging Program in the Division of Chemistry, Theresa Evans-Nguyen and Venkat Bhethanabotla and their groups at the University of South Florida are devising ways to improve analysis of lipid assemblies critical to extracellular signaling. The roles of discrete lipids in these assemblies are not well understood. Furthermore, because lipids aggregate so readily, it is difficult even to identify and quantitate the individual lipid constituents of such assemblies. Mass spectrometry (MS) offers a powerful analytical tool which could provide a great deal of useful information if aggregation challenges could be overcome. To that end, this project focuses on using controlled acoustic waves to disrupt lipid aggregation, thereby facilitating MS analysis. The principles developed in this project should be extendable to other non-specific assemblies and amenable to coupling with current MS sampling methodologies. The research involves a cross-disciplinary effort between the fields of chemistry and engineering, and is expected to strengthen STEM recruitment programs through both fields.At the heart of this research is the use of novel surface acoustic wave (SAW) technologies in the MS workflow. In one modality, SAWs can already lyse extracellular vesicles known as exosomes. Also, as has been previously demonstrated by the team, one can use SAWs to mitigate biofouling from non-specific binding on microfluidic sensors. By extension, SAWs may be feasibly tuned to not just disrupt but further disintegrate bilayer membranes as model aggregates. In a second modality, SAW nebulization (SAWN) exhibits a distinct ionization capacity for both polar and non-polar lipids. By coupling to corona discharge (CD), the South Florida team has achieved comparable efficiency to electrospray ionization (ESI). Thus, the employment of SAWN-CD for the simultaneous analysis of non-polar cholesterol and polar phospholipids comprises a specific capability for which ESI falls short. SAWs are known to effect disruption of extracellular vesicles and to mitigate non-specific biofouling. In this work, SAWs are used to disintegrate model bilayer membrane vesicles into individual lipid components and, through SAW nebulization (SAWN), to facilitate MS ionization of both polar and non-polar lipids by coupling to corona discharge (CD). Specific objectives are: (1) formulation of liposomes as model aggregations to enable a systematic evaluation of SAWs; (2) investigation of SAW disruption parameters to disintegrate lipid assemblies; and 3) investigation SAW nebulization parameters to achieve simultaneous ionization of polar and non-polar species. Thus, through this investigation, the team aims to develop a method that can be applied to extracellular vesicles, allowing them to be directly sampled by mass spectrometry, vastly short-cutting traditional analysis methods for such complex assemblies.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.