Schröter Jenny, Fülöp Annabelle, Hopf Carsten, Schiller Jürgen
Faculty of Medicine, Institute for Medical Physics and Biophysics, Leipzig University, Härtelstr. 16-18, 04107, Leipzig, Germany.
Center for Biomedical Mass Spectrometry and Optical Spectroscopy (CeMOS), Mannheim University of Applied Sciences, Paul-Wittsack-Strasse 10, 68163, Mannheim, Germany.
Anal Bioanal Chem. 2018 Mar;410(9):2437-2447. doi: 10.1007/s00216-018-0926-9. Epub 2018 Feb 14.
Unequivocal assignment of phospholipid peaks in complex mixtures is difficult if only the m/z values but no tandem mass spectrometry (MS/MS) data are available. This is usually the case for matrix-assisted laser/desorption ionization time-of-flight (MALDI-TOF) MS imaging experiments and the analysis has normally to be performed without prior separation. Another problem might be the often matrix-induced loss of one methyl group in phosphatidylcholine (PC) species, which makes them detectable as negative ions becoming isomers of some phosphatidylethanolamines (PEs). Selected lipid mixtures of known compositions were investigated by negative ion MALDI-TOF MS and various imaging experiments. In addition to common matrices such as 2,5-dihydroxybenzoic acid (DHB) and 9-aminoacridine (9-AA), different binary matrices, including 2,5-dihydroxyacetophenone (2,5-DHAP) as matrix additive to DHB, were tested to probe their performance in both ionization modes. Beside artificial PC and PE mixtures of known compositions, egg yolk and liver extracts as well as cryosections from liver and pancreas tissue were selected as biologically relevant systems. The majority of the binary MALDI matrices used here leads to the loss of a methyl group from PC in the negative ion mode, which makes the clear identification of PE species ambiguous. However, this problem does not apply if a mixture of DHB and 2,5-DHAP is used. Therefore, the application of DHB/2,5-DHAP as matrix is a simple method to unequivocally identify PEs even in complex mixtures and tissue sections as negative ions and without the necessity to separate the individual lipid classes prior to MS detection. Graphical abstract Many common MALDI matrices (such as 9-AA) induce the loss of a methyl group from PC rendering the PC detectable as negative ion. These ions (m/z 744.6 in the upper trace) represent isomers of typical PE species. It will be shown that this problem can be avoided if mixtures between DHB and 2,5-DHAP are applied. At these conditions, POPC is exclusively detectable as a matrix adduct with DHB (at m/z 912.6, lower trace) and does not interfere with PE. This approach can also be used in MALDI MS imaging.
如果仅提供质荷比（m/z）值而没有串联质谱（MS/MS）数据，那么在复杂混合物中明确磷脂峰的归属是很困难的。基质辅助激光解吸电离飞行时间质谱（MALDI-TOF）成像实验通常就是这种情况，并且分析通常必须在没有预先分离的情况下进行。另一个问题可能是磷脂酰胆碱（PC）类物质中经常会出现基质诱导的一个甲基的丢失，这使得它们作为负离子被检测到时成为一些磷脂酰乙醇胺（PE）的异构体。通过负离子MALDI-TOF质谱和各种成像实验研究了已知组成的选定脂质混合物。除了常见的基质如2,5-二羟基苯甲酸（DHB）和9-氨基吖啶（9-AA）外，还测试了不同的二元基质，包括作为DHB基质添加剂的2,5-二羟基苯乙酮（2,5-DHAP），以探究它们在两种电离模式下的性能。除了已知组成的人工PC和PE混合物外，还选择了蛋黄和肝脏提取物以及肝脏和胰腺组织的冷冻切片作为生物学相关系统。这里使用的大多数二元MALDI基质会导致负离子模式下PC的一个甲基丢失，这使得PE类物质的明确鉴定变得模糊。然而，如果使用DHB和2,5-DHAP的混合物，这个问题就不会出现。因此，应用DHB/2,5-DHAP作为基质是一种简单的方法，可以即使在复杂混合物和组织切片中也能明确地将PE鉴定为负离子，并且无需在质谱检测之前分离各个脂质类别。图形摘要许多常见的MALDI基质（如9-AA）会导致PC的一个甲基丢失，使PC作为负离子被检测到。这些离子（上迹线中m/z 744.6）代表典型PE类物质的异构体。结果表明，如果应用DHB和2,5-DHAP之间的混合物，这个问题可以避免。在这些条件下，1-棕榈酰-2-油酰磷脂酰胆碱（POPC）仅作为与DHB的基质加合物被检测到（在m/z 912.6，下迹线），并且不会干扰PE。这种方法也可用于MALDI质谱成像。
