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H/D Pooling of Benzoyl Derivatives: Internal Standard per Each Analyte for Lipidomic Quantitation by RP-UHPLC/MS

Authors: Peterka Ondřej | Jirásko Robert | Vaňková Zuzana | Idkowiak Jakub | Holčapek Michal

Year: 2022

Type of publication: ostatní - přednáška nebo poster

Page from-to: nestránkováno

Titles:

Language	Name	Abstract	Keywords
eng	H/D Pooling of Benzoyl Derivatives: Internal Standard per Each Analyte for Lipidomic Quantitation by RP-UHPLC/MS	Chemical derivatization of lipid functional groups can improve extraction efficiency, chromatographic separation, and sensitivity. Benzoyl chloride is a non-hazardous derivatization agent with high reactivity for several functional groups. The optimized benzoylation method increased the sensitivity 2 to 10-fold for almost all investigated lipid classes and even more than 100-fold for monoacylglycerols compared to the non-derivatization approach. Reversed-phase ultrahigh-performance liquid chromatography enables to resolve isomeric lipids, but for accurate quantitation a large number of internal standards must be used. H/D pooling means the mixing of non-labelled derivatives (H) and labelled derivatives (D) in the same ratio, which helps to easier identification and accurate quantitation. Our approach creates an internal standard per each derivatized lipid with the coelution of analytes and internal standards (ΔRT ± 0.1 min), ensuring the same matrix effect and ionization efficiency. Labeled derivatives are used as an internal standard mixture, producing the identical lipidomic profile and corresponding concentration for all analytes. The created doublets in the mass spectrum with known Δm/z facilitate identification and bring the next parameter for a high confidence identification. The method was validated and used for targeted quantitative analysis and lipidomic profiling of healthy controls and cancer patients.	derivatization; lipidomics; H/D pooling; reversed-phased UHPLC; human plasma