Most drugs and metabolites, both licit and illicit, are conjugated prior to excretion in urine or feces. Hydrolysis using beta-glucuronidase converts the glucuronide metabolites back to their free, or non-conjugated form, improving detection and increasing sensitivity for clinical of forensic assays. Evaluation of four beta-glucuronidase enzymes from abalone, red abalone, and two recombinant enzymes were evaluated to determine the most efficient hydrolysis of glucuronide metabolites and maximum recovery of non-conjugated compounds.
Hydrolysis experiments were performed using the new EVOLUTE® HYDRO solid phase extraction plate.The plate contains the Hydro frit technology system, which efficiently holds up aqueous sample and hydrolysis enzyme during incubation. This replaces the need for off-line hydrolysis, traditionally performed in a separate vial or well, and streamlines sample preparation by permitting hydrolysis on the SPE extraction plate.
This webinar presents results of the hydrolysis efficiency experiments, and LC-MS/MS results from extraction of urine samples fortified with a 96 compound panel of commonly prescribed and abused drugs by mixed mode solid phase extraction using in-well hydrolysis.
Stephanie J. Marin received her Ph.D. in chemistry from Arizona State University. She has expertise in sample preparation, liquid chromatography, and mass spectrometry. She has over 10 years of experience developing and validating clinical assays from her tenure at the ARUP Institute for Clinical and Experimental Pathology. She is the author of over 30 peer reviewed publications and book chapters and over 80 abstracts presented at national meetings.