Determination of psychoactive substances in one microliter plasma using a novel 3D printing microfluidic paper-based column coupled to liquid chromatography-mass spectrometry

Abstract

Paper spray ionization is a simple method to analyze samples by mass spectrometry (MS) without sample preparation. However, using paper substrates results in limited performance due to low lifetime, low reproducibility, severe analyte diffusion, and poor chromatography. To address these issues, a stereolithography 3D printed microfluidic paper-based column coupled to liquid chromatography-MS (µPC-LC-MS) was developed. The fabrication process created well-defined microchannels inside a single paper substrate in less than 1 s. The µPC-LC-MS significantly extended the lifetime of the substrate, up to 50 min per run with continuous gradient elution at a flow rate of 20 µL min-1 from regular LC pumps. A single µPC can be reused up to 5 cycles with an RSD of 9.5 % without signal reduction, overloading, or interference. Separation of three psychoactive substances/metabolites (methamphetamine, 7-aminonitrazepam, and morphine-3-glucuronide) was achieved with peak resolution from 0.50 to 1.21, high sensitivity (LOD between 0.5 and 1 ng mL-1) and accuracy (87.0–109.1 %), linearity from 5 to 500 ng mL-1 in both water and plasma, and coefficient of determination (R2) from 0.993 to 1.000 using only 1 µL plasma. The µPC-LC-MS demonstrated robustness and sensitivity, making it a promising tool for bioanalysis based on paper-based microfluidics.