LIQUID-CHROMATOGRAPHY CHEMICAL-REACTION INTERFACE MASS-SPECTROMETRY AS AN ALTERNATIVE TO RADIOISOTOPES FOR QUANTITATIVE DRUG-METABOLISM STUDIES

Chemical reaction interface mass spectrometry (CRIMS) was coupled on-l ine with HPLC using a Vestee particle beam interface, A helium-assiste d nebulizer provided added stability with no loss in accuracy or preci sion as compared to the thermospray nebulizer at flow rates of up to 1 .0 mL/min using isocratic conditions, However, mass spectral response was found to be solvent-dependent for both the helium-assisted and the rmospray nebulizers, Postcolumn solvent addition of methanol eliminate d solvent-dependent decreases in mass spectral response. This allowed gradient HPLC: elutions to be performed. Under these conditions, the f low of solvent into the particle beam interface was 2.5 mL/min, so a c onventional thermospray nebulizer had to be used instead of the helium -assisted nebulizer. Experiments were conducted with the antianxiety a gent buspirone in order to validate the methodology. Metabolites from in vitro incubations of [N-15]/[C-14]buspironc with rat liver slices w ere analyzed by gradient LC/CRIMS and by gradient LC/[C-14] radioactiv ity counting. The response from LC/CRIMS analysis for individual metab olites was then compared with that obtained by LC/[C-14] radioactivity counting. An excellent correlation was observed between the two metho ds for metabolites with quite different HPLC characteristics, Thus, gr adient LC/CRIMS in combination with stable isotopes provides an altern ative to using radioisotopes for carrying out drug metabolism studies.