Isotopic dilution method for bile acid profiling reveals new sulfate glycine-conjugated dihydroxy bile acids and glucuronide bile acids in serum

An ultrahigh performance liquid chromatography tandem mass spectrometry method (UHPLC-MS/MS) was developed for the determination of 41 target and 8 additional bile acids isomers (BAs) in biological fluids. BAs were analysed by solid-phase extraction on 50 mu A biofluid-aliquots, followed by a properly optimised 27 min-chromatographic run. The method provided high sensitivity (limits of detection 0.0002-0.03 mu M, limits of quantitation 0.0007-0.11 mu M), linearity (R-2>0.99) and precision (relative standard deviations <= 16%). A strategy of scheduled/unscheduled injections of real samples together with neutral loss (80 Da and 176 Da) scans allowed us to find additional bile acid isomers not a priori included in the method, while high resolution full scan and MS/MS fragmentation analysis confirmed their structural adherence to the bile acid family. Moreover this is the first study quantifying four sulfate glycine conjugated-dihydroxy bile acid isomers, independently of the diet and postprandial time. Application to a dietary intervention kinetic study confirmed the existence of possible metabotypes amongst the study population (n = 20). A trend differentiating males from females was observed suggesting that serum samples from women contained smaller amounts of certain bile acids. (C) 2019 The Authors. Published by Elsevier B.V.