This study aimed to develop a direct infusion analytical platform for analyzing complex lipid mixtures using a solid-phase extraction (SPE) cartridge. The SPE cartridge was developed in-house and coupled to a contained-electrospray ionization (cESI) source, which enabled online fractionation of lipids followed by real-time mass spectrometry (MS) analysis of eluates. The SPE cartridge was packed with a silica sorbent. We used multiple solvents with varying polarities to sequentially elute and characterize different lipids. To stabilize the electrospray signal derived from nonpolar organic solvents, our SPE-cES-MS platform utilized a co-axial spray system that allowed an ammonium acetate containing lipase enzyme to be delivered from a second electrospray capillary. In its fullest implementation, the SPE-cESI-MS platform was applied to fractionate a sample mixture containing 18 different lipids using hexane, ethanol, and water for sequential elution from the silica sorbent. Together, the three elution steps enabled 93.0% of the lipids in the mixture to be detected, with overall improvement in signal intensities. In contrast, without the miniaturized SPE front-end, direct analysis of the 18-component lipid mixture resulted in the detection of 85.7% of the lipids. The solvents employed were optimized by evaluating their selectivity for polar and nonpolar lipids. The limits of detection associated with the elution of nonpolar lipids (e.g., trilinolein) from the polar silica surface were determined to be 0.62 µM and 0.84 µM for the elution of polar oleic acid from the same sorbent. This work represents a rapid and affordable analytical approach for analyzing complex lipid mixtures in situations where conventional chromatography cannot be used, including analysis on a portable mass spectrometer.
Probing non-covalent membrane protein-lipid interactions remains challenging due to the high hydrophobicity and aggregation propensity of membrane proteins. Here, we present a sheath liquid capillary electrophoresis-native top-down mass spectrometry (CE-nTDMS) platform that enables reproducible infusion of aquaporin Z-cardiolipin assemblies and preserves their non-covalent electrostatic interactions during top-down fragmentation. This workflow generated a diagnostic fragment ion that localizes a lipid-binding region within the protein backbone. Notably, charge-reduced precursor fragment ions of the lipid-bound assembly exhibited asymmetrical isotopic distributions following collisional activation, while corresponding apo precursors displayed symmetrical isotopic distributions. These findings demonstrate the utility of CE-nTDMS for reliable analysis of membrane protein-lipid assemblies and provide new insight into the behavior of lipid-bound fragments during collisional activation.
This study presents a sensing strategy based on reduction-boosted fluorescence of carbon dots for ALP detection, which exhibits a low limit of detection (LOD) of 0.65 U L-1 and a linear range of 5-70 U L-1. This assay can be used to detect ALP in human serum, showing great potential for early diagnosis.
京公网安备 11010802042870号
京ICP备2023020795号-1
扫码关注我们
求助内容：
应助结果提醒方式：