ABSTRACT
In the past few years, high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) has matured to a true alternative to antibody-based immunoassays in routine therapeutic drug monitoring. In transplantation medicine, mass spectrometry-based assessment of immunosuppressant drug levels is considered a gold standard diagnostic procedure. We describe a fast state-of-the-art routine online solid-phase extraction (SPE) HPLC-MS/MS analysis platform that allows monitoring of cyclosporine A, tacrolimus, sirolimus and everolimus from 50-microl aliquots of EDTA whole blood specimens within 3.4 min total analysis time. Sample purification is done by offline protein precipitation followed by two automated chromatographic separation steps. Mass spectrometry-based analyte quantification relies on selected reaction monitoring experiments. The assay underwent complete validation and performance evaluation studies and performs very well in several international proficiency testing schemes. In daily routine, it allows reporting of about 75 patient sample results per work shift with a typical total individual sample turnaround time of less than 3 h.
Subject(s)
Chromatography, High Pressure Liquid/methods , Cyclosporine/blood , Immunosuppressive Agents/blood , Sirolimus/analogs & derivatives , Sirolimus/blood , Tacrolimus/blood , Tandem Mass Spectrometry/methods , Everolimus , HumansABSTRACT
Novel DHODH inhibitors were developed based on a previously described series by introduction of heteroatoms into the cyclopentene ring and hydroxyl groups attached to it. Also, the hydrophobic biphenyl side chain was replaced with benzyloxy phenyl groups. Activities on human, rat, and mouse enzymes indicate a species specificity of these inhibitors.
