Shotgun lipidomics: multidimensional MS analysis of cellular lipidomes

Abstract

Shotgun lipidomics, comprised of intrasource separation, multidimensional mass spectrometry and computer-assisted array analysis, is an emerging powerful technique in lipidomics. Through effective intrasource separation of predetermined groups of lipid classes based on their intrinsic electrical propensities, analyses of lipids from crude extracts of biologic samples can be directly and routinely performed. Appropriate multidimensional array analysis of lipid pseudomolecular ions and fragments can be performed leading to the identification and quantitation of targeted lipid molecular species. Since most biologic lipids are linear combinations of aliphatic chains, backbones and head groups, a rich repertoire of multiple lipid building blocks present in discrete combinations represent experimental observables that can be computer reconstructed in conjunction with their pseudomolecular ions to directly determine the lipid molecular structures from a lipid extract. Through this approach, dramatic increases in the accessible dynamic range for ratiometric quantitation and discrimination of isobaric molecular species can be achieved without any prior column chromatography or operator-dependent supervision. At its current state of development, shotgun lipidomics can analyze over 20 lipid classes, hundreds of lipid molecular species and more than 95% of the mass content of a cellular lipidome. Thus, understanding the biochemical mechanisms underlying lipid-mediated disease states will be greatly facilitated by the power of shotgun lipidomics.

Alkaline methanolysis of lipid extracts extends shotgun lipidomics analyses to the low-abundance regime of cellular sphingolipids

“…Either neutral-loss (NL) scanning or precursor-ion (PI) scanning at the mass or m/z ratio of a fragment, respectively, can be performed to "isolate" a given class or group of lipids from which individual lipid molecular species can be identified through multidimensional array analysis. Each of these fragments represents a building block of the class or the group of lipids [13].…”

“…Finally, quantitation by shotgun lipidomics is performed in a two-step procedure [13,21,22]. First, the abundant and nonoverlapping molecular species of a class are quantified by comparison with a preselected internal standard of the class after 13 C deisotoping [4,14] from a survey scan.…”

“…One of the major analytical platforms in current lipidomics practice is multidimensional massspectrometry (MS) 1 -based shotgun lipidomics [13][14][15]. This platform has now evolved into a mature technology that includes a series of simple steps such as multiplexed extractions, *Corresponding author.…”

“…Although lipidomics has emerged as a distinct field only within the past few years [1][2][3], numerous new discoveries and/or advances have already been made [3][4][5][6][7][8][9][10][11][12]. Its essential roles in identifying the biochemical mechanisms of lipid metabolism, investigating the functions of an individual gene of interest, identifying novel biomarkers, and evaluating drug efficacy, among others, are becoming increasingly recognized.One of the major analytical platforms in current lipidomics practice is multidimensional massspectrometry (MS) 1 -based shotgun lipidomics [13][14][15]. This platform has now evolved into a mature technology that includes a series of simple steps such as multiplexed extractions, *Corresponding author.…”

“…Either neutral-loss (NL) scanning or precursor-ion (PI) scanning at the mass or m/z ratio of a fragment, respectively, can be performed to "isolate" a given class or group of lipids from which individual lipid molecular species can be identified through multidimensional array analysis. Each of these fragments represents a building block of the class or the group of lipids [13].Finally, quantitation by shotgun lipidomics is performed in a two-step procedure [13,21,22]. First, the abundant and nonoverlapping molecular species of a class are quantified by comparison with a preselected internal standard of the class after 13 C deisotoping [4,14] from a survey scan.…”