Visible-Light-Activated Dual-Pathway Strategy for Comprehensive Lipid Isomer Analysis

Phospholipids are essential components of biological membranes. Their structural diversity, due to the presence of sn-positions, C = C locations and geometric configurations, significantly influences their biological functions. The isomerization of unsaturated lipid isomers plays a crucial role in bacterial responses to environmental stimuli. However, in-depth mechanistic studies have been lacking effective analytical tools. Here, we developed a visible light-activated cycloaddition and isomerization reaction between methyl benzoylformate and unsaturated lipids, combined with liquid chromatography-mass spectrometry, to establish a comprehensive method for resolving lipid isomers. Subsequently applying this method to investigate bacterial stress response mechanisms, we identified unique sn-position, C = C location and cis/trans isomers conversion patterns in Pseudomonas under high-temperature stress. This reveals the critical role of lipid isomers in bacterial responses to environmental stress. This study not only enables comprehensive identification of lipid isomers but also provides an effective tool and key insights for investigating bacterial stress response mechanisms.