3-Acetylpyridine On-Tissue Paternò–Büchi Derivatization Enabling High Coverage Lipid C═C Location-Resolved MS Imaging in Biological Tissues

As the predominant phospholipids in mammalian cells, phosphatidylcholines (PCs) have been demonstrated to play a crucial role in a multitude of vital biological processes. Research has highlighted the significance of the diversity in PC isomers as instigators of both physiological and pathological responses, particularly those with variations in the position of double bonds within their fatty chains. Profiling these PC isomers is paramount to advancing our understanding of their biological functions. Despite the availability of analytical methods utilizing high-resolution secondary mass spectrometry (MS 2 ) fragmentation, a novel approach was imperative to facilitate large-scale profiling of PC isomers while ensuring accessibility, facility, and reliability. In this study, an innovative strategy centered around structure-driven predict-to-hit profiling of the double bond positional isomers for PCs was meticulously developed, employing negative reversed-phase liquid chromatography−multiple reaction monitoring (RPLC-MRM). This novel methodology heightened the sensitivity. The analysis of rat lung tissue samples resulted in the identification of 130 distinct PC isomers. This approach transcended the confines of available PC isomer standards, thereby broadening the horizons of PC-related biofunction investigations. By optimizing the quantitation reliability, the scale of sample analysis was judiciously managed. This work pioneers a novel paradigm for the exploration of PC isomers, distinct from the conventional methods reliant on high-resolution mass spectrometry (HRMS). It equips researchers with potent tools to further explore the biofunctional aspects of lipids.

Section: Discussionmentioning

confidence: 99%

Novel Structure-Driven Predict-to-Hit Strategy for PC Double Bond Positional Isomer Identification Based on Negative LC-MRM Analysis

Tie,

Cui,

Zhang

et al. 2024

“…The strategy of chemical derivatization-coupled MS/MS bypasses the reaction required inside the mass analyzer. On-tissue or online derivatization was explored by a PB reaction, − epoxidation, and singlet-oxygen reaction for MSI applications. These methods improve the sensitivity for detection of phospholipids and CC location isomers; however, due to a lack of capability to identify the chain composition, these methods cannot allocate CC locations onto the fatty acyl chain (e.g., PC 16:0_18:1( n -9) or PC 16:0/18:1).…”

Section: Introductionmentioning

confidence: 99%

MS³ Imaging Enables the Simultaneous Analysis of Phospholipid C═C and sn-Position Isomers in Tissues

Guo,

Cao,

Fan

et al. 2024

Mass spectrometry (MS) imaging of lipids in tissues with high structure specificity is challenging in the effective fragmentation of position-selective structures and the sensitive detection of multiple lipid isomers. Herein, we develop an MS 3 imaging method for the simultaneous analysis of phospholipid C�C and sn-position isomers by on-tissue photochemical derivatization, nanospray desorption electrospray ionization (nano-DESI), and a dual-linear ion trap MS system. A novel laser-based sensing probe is developed for the real-time adjustment of the probe-to-surface distance for nano-DESI. This method is validated in mouse brain and kidney sections, showing its capability of sensitive resolving and imaging of the fatty acyl chain composition, the snposition, and the C�C location of phospholipids in an MS 3 scan. MS 3 imaging of phospholipids has shown the capability of differentiation of cancerous, fibrosis, and adjacent normal regions in liver cancer tissues.

“…Contributing to the structural diversity, more than 48 000 unique lipids are involved in the LIPID MAPS Structure Database, a considerable amount of which are isomers and isobars . Unsaturated lipids often exist in isomeric structures that differ only in the position of carbon–carbon double bonds (CC), the stereochemistry of CC ( cis / trans ) or stereospecific numbering ( sn ) positions, fulfilling different biological roles. , They are increasingly recognized as physiologically active molecules and promising lipid markers. , Recent works revealed that disruption of lipid homeostasis is associated with many disorders such as cancer, − diabetes, − and Alzheimer’s disease. − …”

Section: Introductionmentioning

confidence: 99%

Aza-Prilezhaev Aziridination-Enabled Multidimensional Analysis of Isomeric Lipids via High-Resolution U-Shaped Mobility Analyzer–Mass Spectrometry

Li,

Wang,

Guo

et al. 2024

Unsaturated lipids constitute a significant portion of the lipidome, serving as players of multifaceted functions involving cellular signaling, membrane structure, and bioenergetics. While derivatization-assisted liquid chromatography tandem mass spectrometry (LC-MS/MS) remains the gold standard technique in lipidome, it mainly faces challenges in efficiently labeling the carbon−carbon double bond (C�C) and differentiating isomeric lipids in full dimension. This presents a need for new orthogonal methodologies. Herein, a metal-and additive-free aza-Prilezhaev aziridination (APA)-enabled ion mobility mass spectrometric method is developed for probing multiple levels of unsaturated lipid isomerization with high sensitivity. Both unsaturated polar and nonpolar lipids can be efficiently labeled in the form of N−H aziridine without significant side reactions. The signal intensity can be increased by up to 3 orders of magnitude, achieving the nM detection limit. Abundant site-specific fragmentation ions indicate C�C location and sn-position in MS/MS spectra. Better yet, a stable monoaziridination product is dominant, simplifying the spectrum for lipids with multiple double bonds. Coupled with a U-shaped mobility analyzer, identification of geometric isomers and separation of different lipid classes can be achieved. Additionally, a unique pseudo MS 3 mode with UMA-QTOF MS boosts the sensitivity for generating diagnostic fragments. Overall, the current method provides a comprehensive solution for deep-profiling lipidomics, which is valuable for lipid marker discovery in disease monitoring and diagnosis.