Mass Spectrometry Method for Carbohydrate Structure

复杂的碳水化合物是基本上所有生命系统的组成部分，在生物系统中起着关键作用。 各种各样的生物过程和识别事件。然而，它们的详细结构决定仍然存在， 难虽然质谱法可以提供关于碳水化合物结构的重要信息，但确定碳水化合物的结构是一个非常重要的步骤。 由较大碳水化合物结构的断裂产生的单个单糖离子的立体化学是 一般不可能。而且，通过质谱法分析异构寡糖的混合物仍然很困难 因为基本电荷/质量比本身不能物理分辨异构体结构。因此， 该项目的目的是开发一种分析仪器，将显着提高结构测定的能力， 复杂的碳水化合物，使异构体结构被解析。 拟议的仪器将基于离子阱质谱和离子迁移率的原则 光谱法在其最终形式中，仪器将具有耦合到双栅极的电喷雾电离（ESI）源。 离子迁移谱仪（IMS）。双栅离子迁移谱仪，根据大小和形状分离离子， 质量，将用于选择特定的寡糖异构体以注入离子阱质谱仪（ITMS）。 流动性选择的寡糖的脉冲将在ITMS中浓缩，然后碰撞诱导 解离以产生所选糖单体的离子。然后将这些单体注入第二个， 串联IMS以鉴定单体离子的立体化学和异头构型。分离和 将单一寡糖引入ITMS中，并解离和鉴定单糖， 复杂寡糖的完整结构分析是可能的。该计划的关键步骤是 通过在离子阱中的碎裂产生单糖和通过IMS分离单糖。因此，在本发明中， 该项目的R21阶段建议开发用于寡糖片段化的新型MS方法， 用于分离寡糖以及寡糖的立体异构体和端基异构体对的新颖IMS方法， 单糖离子。在R33阶段，将建造一个IMS-IT-IMS仪器，对其进行表征， 用于阐明寡糖结构。 本赠款申请中提出的整体工具有可能产生以前从未有过的数据， 这类重要而神秘的生物化合物，将为新兴领域提供一种新的工具， 结构糖组学。到目前为止，大多数天然存在的寡糖的详细化学结构已经 无法单独通过质谱法进行鉴定。该项目的成功完成将提供一个工具， 不仅将质谱的分析能力扩展到分析同分异构的单糖和寡糖， 通常也可用于分析所有类型的分子，其中可能存在关于可能的异构体的不确定性。 混合物的变体或较大聚合物分子的异构单体成分。

Complex carbohydratesare integral, functioning components of essentially all living systems, playing critical roles in an enormous variety of biological processes and recognition events. Yet, their detailed structural determination remains difficult. While mass spectrometry can provide important information about carbohydrate structures, a determination of the stereochemistry of individual monosaccharide ions that arise by fragmentation of larger carbohydrate structures is not generally possible. Also, analysis of mixtures of isomeric oligosaccharides by mass spectrometry remains difficult because the fundamental charge/mass ratio alone does not physically resolve isomeric structures. Thus, the goal of this project is to develop an analytical instrument that will markedly improve capabilities in the structural determination of complex carbohydrates by enabling isomeric structures to be resolved. The proposed instrument will be based on the principles of both ion trap mass spectrometry and ion mobility spectrometry. In its final form, the instrument will have an electrospray ionization (ESI) source coupled to a double gate ion mobility spectrometer (IMS). The double gate IMS, separating ions on the basis of size and shape rather than mass, will serve to select specific oligosaccharide isomers for injection into an ion trap mass spectrometer (ITMS). Pulses of the mobility selected oligosaccharide will be concentrated in the ITMS followed by collision induced dissociation to generate ions of selected sugar monomers. These monomers will then be injected into a second, tandem IMS to identify the stereochemistry and anomeric configuration of monomer ions. By the separation and introduction of single oligosaccharides into the ITMS with dissociation and identification of the monomeric sugars, complete structural analysis of complex oligosaccharides may be possible. The critical steps in this plan are the production of monosaccharides by fragmentation in the ion trap and the separation of monosaccharides by IMS. Thus, the R21 phase of the project proposes to develop novel MS methods for the fragmentation of oligosaccharides and novel IMS methods for the separation of oligosaccharides as well as stereoisomeric and anomeric pairs of monosaccharide ions. In the R33 phase an IMS-IT-IMS instrument will be constructed, characterized and demonstrated for the elucidation of oligosaccharide structures. The overall instrument proposed in this grant application has the potential to generate data never before possible on this important and enigmatic class of biological compounds and will provide a novel tool for the emerging field of Structural Glycomics. To date, the detailed chemical structures of most naturally occurring oligosaccharides have eluded identification solely by mass spectrometry. Successful completion of this project will provide a tool which will not only extend the analytical power of mass spectrometry to analysis of isomeric mono- and oligosaccharides, but will also be generally useful for analysis of all types of molecules where ambiguity may exist concerning possible isomeric variants in mixtures or isomeric monomeric constituents of larger polymeric molecules.