Direct Plasma Analysis of Drug Compounds Using Monolithic Column Liquid Chromatography and Tandem Mass Spectrometry

Abstract

A monolithic silica column high-performance liquid chromatography−tandem mass spectrometry (HPLC−MS/MS) method has been developed for the high-speed direct simultaneous determination of a drug discovery compound and its major circulating metabolite (M-72) in rat plasma. This methodology makes use of flow programming and an alkyl-bonded silica rod column for fast macromolecule removal and chromatographic separation without the need for significant sample preparation. The matrix ionization suppression effect on the monolithic column HPLC−MS/MS system was investigated using the postcolumn infusion technique. After 200 plasma injections on a 50 × 4.6 mm monolithic silica column, consistent column efficiency of close to 39 000 theoretical plates/m and reproducible retention times for the analytes were observed. The apparent on-column recoveries of 12 test compounds in rat plasma samples were greater than 90%. The proposed fast direct plasma injection method was tested over a 3-day period with the interday coefficient of variation less than 15% for both analytes.