Abstract
In vitro metabolic stability studies are performed routinely in drug discovery to determine the rate of metabolism as well as the metabolic fate of compounds. These studies are labor intensive, involving incubation of the compound with a biological matrix, sampling at various timepoints, stopping the reaction, and sample preparation for analysis. All of these steps involve manual pipetting in the conventional method.
 
An automated method for in-vitro metabolism studies is reported here. The method significantly reduces the time and manual labor required and has other advantages, such as better reproducibility and unattended operation.
 
This method utilizes an autosampler custom configured with cooling and incubation capabilities. The autosampler is programmed to directly inject incubation samples at set timepoints onto an online extraction column. The extracted sample then enters an analytical column for separation and ultimately the mass spectrometer for detection. The injection has the dual function of stopping the reaction and starting the analysis on the LC/MS. This method was used for the metabolic stability study of a prodrug in plasma and liver S9 fractions of five different species. The stability data from the automated method were similar to those obtained using the conventional method. The potential for this method to increase throughput of metabolic stability studies in drug discovery is demonstrated.

Background
Fran Lai received her M.S. in Chemistry from Iowa State University, with a thesis on ion exchange chromatography. She also studied mass spectrometry at the University of Nebraska under Dr. Michael Gross. She has been an analytical chemist for over 20 years and is currently a Consultant in LC/MS/MS Bioanalytical. Her career started at Syntex Pharmaceuticals (now "Roche Palo Alto") and Biogen in Boston, then she developed a deep interest in analytical instrumentation and worked for chromatography instrumentation companies including Waters, Varian, Thermo, and Dionex, in the areas of applications development and technical marketing. She obtained further education in mass spectrometry and started working as a GC/MS chemist at HMT Technology, and later as an LC/MS bioanalyst at Genentech. During her 7 years at Genentech, she acquired experience in in-vivo and in-vitro studies for small molecule drug discovery, while applying her analytical skills in LC/MS/MS bioanalyses. One day she realized that in-vitro or metabolism experts and analytical instrumentation experts seem to come from different planets! As someone who was knowledgeable in both areas, she saw a need that the metabolism scientists had that the instrumentation expert could provide for. After several months' work, she came up with an innovative automated online system for incubation and analysis for in-vitro studies.