Analysis of unsaturated fatty acids by supercritical fluid chromatography tandem mass spectrometry coupled with online Patern`o-Bchi reaction

Carbon double bond (C--C) positions in unsaturated fatty acids (UFAs) can be identified based on Patern`o-Buchi (PB) reaction coupled with mass spectrometry (MS). However, achieving a high conversion rate and confusing mass spectrum are still challenging. In this study, an online PB reaction coupled with supercritical fluid chromatography-mass spectrometry (SFC-MS), using a homemade flow microreactor, was established to analyze C--C bonds in UFAs, and benzaldehyde (BA) was proposed as the optimized reagent. Benefiting from this setup, both prototype and product ions of FAs can be rapidly captured at the same retention time, significantly enhancing the accuracy of lipid identification, especially when dealing with complex compounds. Furthermore, a high C--C bond conversion rate was achieved, ranging from 53 to 67 %. Additionally, this strategy demonstrated excellent relative quantification abilities, which were based on the ratios of either the sum of diagnostic ions or the individual diagnostic ions. This novel approach was proposed and applied for the specific recognition of free FA in human plasma, which successfully provides quantitative information about UFA isomers. This strategy is believed to hold promise for the analysis of unsaturated lipid isomers in complex biological samples.