Untargeted Lipidomic Profiling of Dry Blood Spots Using SFC-HRMS

Abstract

Lipids are essential cellular constituents that have many critical roles in physiological functions. They are notably involved in energy storage and cell signaling as second messengers, and they are major constituents of cell membranes, including lipid rafts. As a consequence, they are implicated in a large number of heterogeneous diseases, such as cancer, diabetes, neurological disorders, and inherited metabolic diseases. Due to the high structural diversity and complexity of lipid species, the presence of isomeric and isobaric lipid species, and their occurrence at a large concentration scale, a complete lipidomic profiling of biological matrices remains challenging, especially in clinical contexts. Using supercritical fluid chromatography coupled with high-resolution mass spectrometry, we have developed and validated an untargeted lipidomic approach to the profiling of plasma and blood. Moreover, we have tested the technique using the Dry Blood Spot (DBS) method and found that it allows for the easy collection of blood for analysis. To develop the method, we performed the optimization of the separation and detection of lipid species on pure standards, reference human plasma (SRM1950), whole blood, and DBS. These analyses allowed an in-house lipid data bank to be built. Using the MS-Dial software, we developed an automatic process for the relative quantification of around 500 lipids species belonging to the 6 main classes of lipids (including phospholipids, sphingolipids, free fatty acids, sterols, and fatty acyl-carnitines). Then, we compared the method using the published data for SRM 1950 and a mouse blood sample, along with another sample of the same blood collected using the DBS method. In this study, we provided a method for blood lipidomic profiling that can be used for the easy sampling of dry blood spots.

Dried blood spots in clinical lipidomics: optimization and recent findings

“…The separation of oxidized lipids as well as the study of positional isomers can benefit from SFC-MS approaches due to the properties of this lipidomic approach [66]. From the eligible studies, only one study using SFC-MS matched the selection criteria and focused on method development and detection of complex lipids [37]. It allowed the identification from DBS of 500 lipid species from several classes of lipids, such as phospholipids, sphingolipids, free fatty acids, sterols, and fatty acyl-carnitines.…”

“…Considering the identification of DBS lipid species, different lipid classes were analysed and identified, such as FA, PUFA [20-23, 26, 29, 36, 38], or acylcarnitines [17,18], mainly analysed by GC-MS with a few studies by LC-MS. Other classes of complex lipids like sphingolipids [27,35], PL, Chol, TG, prenols, CER, and sterols [13,15,19,24,25,28,37,39] were analysed either by LC-MS, DI-MS, or SFC-MS. However, only one study solely reported the identification of several variables of the lipid fraction, not specifying lipid classes or species (Table 2) [16].…”

“…SFC-MS was used to develop a new analytical strategy for a high-throughput and comprehensive lipidomic analysis [37]. The researchers developed and optimized a new method that allowed the separation of 17 classes of lipids by SFC-MS-Q-TOF.…”

Tricky Isomers—The Evolution of Analytical Strategies to Characterize Plasmalogens and Plasmanyl Ether Lipids

“…Generally, normal phase chromatography and hydrophilic interaction liquid chromatography (HILIC) separate lipids in a lipid class-dependent manner, while in reversed-phase chromatography a lipid species-specific separation behavior is facilitated. Both principles are widely applied in lipidomic studies as described by Harrieder et al (2022 ), while promising methods utilizing supercritical fluid chromatography for separation are under development ( Wolrab et al, 2020 ; Le Faouder et al, 2021 ) that should enable class-wise separation of plasmanyl and plasmenyl lipid species, which in comparison with strategies refined for DI methods (necessary to correct for type-II isotopic effects) should allow high-throughput lipidomics with plasmalogen resolution on a whole lipidome scale.…”

Mass Spectrometry-Based Zebrafish Toxicometabolomics: A Review of Analytical and Data Quality Challenges

“…Some classes will be overlooked, while others will be enriched [ 61 ]. Depending on the goal of the study, the extraction can also be divided into “global” metabolomics [ 56 ], polar (primary) metabolomics [ 62 ], and lipidomics [ 63 ], which can be further subdivided and tailored for less abundant lipid classes, such as polyunsaturated fatty acids [ 64 ], steroids [ 65 ], or the epilipidome [ 66 ].…”