Department of Analytical and Environmental Chemistry and Szentágothai Research Centre, Faculty of Sciences, University of Pécs, Ifjúság útja 6., H-7624 Pécs, Hungary.
Institute of Bioanalysis, Medical School, University of Pécs, Szigeti út 12., H-7624 Pécs, Hungary.
Toxins (Basel). 2024 Jul 18;16(7):322. doi: 10.3390/toxins16070322.
The toxic nature of bacterial endotoxins is affected by the structural details of lipid A, including the variety and position of acyl chains and phosphate group(s) on its diglucosamine backbone. Negative-ion mode tandem mass spectrometry is a primary method for the structure elucidation of lipid A, used independently or in combination with separation techniques. However, it is challenging to accurately characterize constitutional isomers of lipid A extracts by direct mass spectrometry, as the elemental composition and molecular mass of these molecules are identical. Thus, their simultaneous fragmentation leads to a composite, so-called chimera mass spectrum. The present study focuses on the phosphopositional isomers of the classical monophosphorylated, hexaacylated -type lipid A. Collision-induced dissociation (CID) was performed in an HPLC-ESI-QTOF system. Energy-resolved mass spectrometry (ERMS) was applied to uncover the distinct fragmentation profiles of the phosphorylation isomers. A fragmentation strategy applying multi-levels of collision energy has been proposed and applied to reveal sample complexity, whether it contains only a 4'-phosphorylated species or a mixture of 1- and 4'-phosphorylated variants. This comparative fragmentation study of isomeric lipid A species demonstrates the high potential of ERMS-derived information for the successful discrimination of co-ionized phosphorylation isomers of hexaacylated lipid A.
细菌内毒素的毒性性质受脂质 A 的结构细节影响，包括其葡糖胺骨架上的酰基链和磷酸基团的种类和位置。负离子模式串联质谱是脂质 A 结构解析的主要方法，可单独使用或与分离技术结合使用。然而，由于这些分子的元素组成和分子量相同，直接通过质谱准确地表征脂质 A 提取物的结构异构体具有挑战性。因此，它们的同时碎裂会导致复合，即所谓的嵌合体质谱。本研究集中于经典单磷酸化、六酰化 - 型脂质 A 的磷酸化位置异构体。在 HPLC-ESI-QTOF 系统中进行了碰撞诱导解离 (CID)。能量分辨质谱 (ERMS) 用于揭示磷酸化异构体的独特碎裂谱。提出并应用了一种多级碰撞能的碎裂策略，以揭示样品的复杂性，无论其是否仅包含 4'-磷酸化物质还是 1-和 4'-磷酸化变体的混合物。对异构脂质 A 物质的这种比较碎裂研究表明，ERMS 衍生信息在成功区分六酰化脂质 A 的共电离磷酸化异构体方面具有很高的潜力。
