Application of Multiple Reaction Monitoring in 2D-LC-MS/MS for the Simultaneous Quantification of UDCA and Internal Standards in Clinical Plasma Samples

This research focuses on the optimization and implementation of multiple reaction monitoring (MRM) in a 2D-LC-MS/MS system for the simultaneous quantification of Ursodoxycholic Acid (UDCA) and Valsartan (internal standard) in plasma samples. The analytical system incorporates dual chromatographic columns and electrospray ionization in negative mode, ensuring high specificity and sensitivity. The optimized chromatographic conditions, employing 0.2% formic acid and acetonitrile as mobile phases, achieved a total run time of less than 4 minutes. Calibration curves spanning 50–8000 ppb revealed excellent linearity, with a detection limit of 50 ppb. Validation adhered to EMEA and ICH M10 guidelines, confirming the method’s accuracy (98–102%), precision (RSD < 5%), and negligible matrix effects (MEF > 90%). Application to clinical samples demonstrated the method’s efficacy in high-throughput analysis, paving the way for broader adoption in pharmacokinetic and bioavailability studies. This study highlights the capability of MRM-enabled 2D-LC-MS/MS in delivering precise quantification of plasma biomarkers.