Analytical approach to determining human biogenic amines and their metabolites using eVol microextraction in packed syringe coupled to liquid chromatography mass spectrometry method with hydrophilic interaction chromatography column

Analysis of biogenic amines (BAs) in different human samples provides insight into the mechanisms of various biological processes, including pathological conditions, and thus may be very important in diagnosing and monitoring several neurological disorders and cancerous tumors. In this work, we developed a simple and fast procedure using a digitally controlled microextraction in packed syringe (MEPS) coupled to liquid chromatography mass spectrometry (LC–MS) method for simultaneous determination of biogenic amines, their precursors and metabolites in human plasma and urine samples. The separation of 12 low molecular weight and hydrophilic molecules with a wide range of polarities was achieved with hydrophilic interaction chromatography (HILIC) column without derivatization step in 12 min. MEPS was implemented using the APS sorbent in semi-automated analytical syringe (eVol®) and small volume of urine and plasma samples, 50 µL and 100 μL, respectively. We evaluated important parameters influencing MEPS efficiency, including stationary phase selection, sample pH and volume, number of extraction cycles, and washing and elution volumes. In optimized MEPS conditions, the analytes were eluted by 3×50 μL of methanol with 0.1% formic acid. The chromatographic separation of analytes was performed on XBridge Amide™ BEH analytical column (3.0 mm×100 mm, 3.5 µm) using gradient elution with mobile phase consisting of phase A: 10 mM ammonium formate buffer in water pH 3.0 and phase B: 10 mM ammonium formate buffer in acetonitrile pH 3.0. The LC–HILIC–MS method was validated and, in optimum conditions, presented good linearity in concentration range within 10–2000 ng/mL for all the analytes with a determination coefficient (r2) higher than 0.999 for plasma and urine samples. Method recovery ranged within 87.6–104.3% for plasma samples and 84.2–98.6% for urine samples. The developed method utilizing polar APS sorbent along with polar HILIC column was applied for simultaneous bioanalysis of trace amounts of polar endogenous biogenic amines in real human urine and plasma samples.