Non-targeted and targeted analysis of oxylipins in combination with charge-switch derivatization by ion mobility high-resolution mass spectrometry

Stefan Hellhake,Thorsten Benter,Pablo Steinberg,Nils Helge Schebb,Michael T Empl,Sven W Meckelmann,Walter Wißdorf,Kristina Rentmeister,Oliver J Schmitz

doi:10.1007/s00216-020-02795-2

Stefan Hellhake, Thorsten Benter + Show 7 more

Open Access

https://doi.org/10.1007/s00216-020-02795-2

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Abstract

Eicosanoids and other oxylipins play an important role in mediating inflammation as well as other biological processes. For the investigation of their biological role(s), comprehensive analytical methods are necessary, which are able to provide reliable identification and quantification of these compounds in biological matrices. Using charge-switch derivatization with AMPP (N-(4-aminomethylphenyl)pyridinium chloride) in combination with liquid chromatography ion mobility quadrupole time-of-flight mass spectrometry (LC-IM-QTOF-MS), we developed a non-target approach to analyze oxylipins in plasma, serum, and cells. The developed workflow makes use of an ion mobility resolved fragmentation to pinpoint derivatized molecules based on the cleavage of AMPP, which yields two specific fragment ions. This allows a reliable identification of known and unknown eicosanoids and other oxylipins. We characterized the workflow using 52 different oxylipins and investigated their fragmentation patterns and ion mobilities. Limits of detection ranged between 0.2 and 10.0 nM (1.0–50 pg on column), which is comparable with other state-of-the-art methods using LC triple quadrupole (QqQ) MS. Moreover, we applied this strategy to analyze oxylipins in different biologically relevant matrices, as cultured cells, human plasma, and serum.Graphical abstract

Highlights

Oxylipins are oxidation products of polyunsaturated fatty acids (PUFA) derived from the arachidonic acid (ARA) cascade which can act as lipid mediators [1, 2]
The second path of the ARA cascade involves the oxidation of PUFAs by different lipoxygenases (LOX), leading via hydroperoxy PUFA intermediates to leukotrienes and lipoxins as well as hydroxyl PUFAs such as 15hydroxyeicosapentaenoic acid (15-HEPE) [13,14,15]
Other ions commonly generated by electrospray ionization (ESI) such as adducts of H+, Na+, and K+ or neutral loss of water for the hydroxy PUFAs could not be observed with significant abundance

Summary

Introduction

Oxylipins are oxidation products of polyunsaturated fatty acids (PUFA) derived from the arachidonic acid (ARA) cascade which can act as lipid mediators [1, 2]. Three enzymatic pathways of the ARA cascade as well as non-enzymatic autoxidation lead to the formation of oxylipins [8, 9]. In the first enzymatic pathway, ARA or eicosapentaenoic acid (EPA) is converted by cyclooxygenases (COX) to different prostaglandins and thromboxanes such as prostaglandin D2 (PGD2) or prostaglandin E2 (PDE2) [1, 2, 12]. The second path of the ARA cascade involves the oxidation of PUFAs by different lipoxygenases (LOX), leading via hydroperoxy PUFA intermediates to leukotrienes and lipoxins as well as hydroxyl PUFAs such as 15hydroxyeicosapentaenoic acid (15-HEPE) [13,14,15].

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