Liquid chromatography/tandem mass spectrometry sensitivity enhancement via online sample dilution and trapping: applications in microdosing and dried blood spot (DBS) bioanalysis

A simple online sample dilution, enrichment, and cleanup technique was developed for sensitive microdosing and dried blood spot (DBS) liquid chromatography/tandem mass spectrometric (LC/MS/MS) bioanalysis. Samples are diluted online with water and enriched in a trap column which is subsequently switched inline with the analytical column. Excellent lansoprazole (in acetonitrile) peak shape is maintained even with an 80‐µL injection. In comparison, similar chromatographic peaks were observed only when a small volume of the same solution, i.e., 1 µL, was injected on a regular high‐performance liquid chromatography (HPLC) system, where an injection of 5 µL resulted in severe peak fronting. A substantial enhancement in sensitivity is realized in the trapping mode using large injection volumes. The trap column is washed at the beginning and at the end of each injection with aqueous and organic solvent respectively to remove matrix components. This ultimately leads to reduction of matrix effects and mass spectrometer noise, thus facilitating the utilization of protein precipitation as the sample preparation for plasma samples. A lower limit of quantitation (LLOQ) of 0.5 pg/mL was demonstrated for lansoprazole in human plasma with a signal‐to‐noise (S/N) ratio of 13 using a 100 µL injection. Excellent intra‐day precision and accuracy were established for lansoprazole in human plasma with good linearity (R² > 0.999) from 0.5 to 500 pg/mL. This level of LLOQ makes LC/MS/MS a practical alternative for microdosing bioanalysis, where the dose is typically 100 times lower than the therapeutic dose. The same technique was applied to quantitate lansoprazole in human whole blood employing DBS technology. With a single 3‐mm punch, i.e. ～2 µL of whole blood or ～1 µL plasma, a LLOQ of 0.1 ng/mL showed sufficient S/N ratio (40) for lansoprazole when 75 µL of extract was injected. In all, the online sample dilution, cleanup, and enrichment technique demonstrated the practical utility of LC/MS/MS in microdosing and DBS bioanalysis. Copyright © 2010 John Wiley & Sons, Ltd.