An UHPLC-QE-Orbitrap-MS Method for Accurate Quantification of Short-Chain Fatty Acids in Serum From an Older Chinese Population

Abstract

Short-chain fatty acids (SCFAs), such as acetic acid, propionic acid, lactic acid, β-hydroxybutyric acid, and crotonic acid, play key biological roles and are also strongly associated with the maintenance of health and the development of age-related diseases. However, an accurate method for SCFAs detection in human serum is lacking. Herein, we developed an UHPLC-QE-Orbitrap MS method based on 3-nitrophenylhydrazine derivatization in negative electrospray ionization through parallel reaction monitoring mode for the simultaneous detection of 11 SCFAs in the serum, and the analysis was performed on an Agilent Proshell 120 EC-C₁₈ column (2.1 mm × 100 mm, 2.7 μm). Three pairs of isomers—isobutyric and butyric acid, isovaleric and valeric acid, and isocaproic and caproic acid—were completely separated in 20 min in a single run. Our method exhibited satisfactory linearity (R > 0.99) for all analytes, and both intra-day and inter-day accuracies (73.74% to 127.9%) and precisions ( < 21%) were acceptable for most targeted compounds. The extraction recoveries ranged from 90.80% to 111.7%, and the internal standard-normalized matrix effects were 74.43%–116.9%. This method was successfully applied to a cohort of 1021 older Chinese individuals. Our results may further the understanding of the metabolic phenotypes associated with SCFAs in other populations.