High-throughput liquid chromatography-tandem mass spectrometry determination of bupropion and its metabolites in human, mouse and rat plasma using a monolithic column.
In the present work, a high-throughput LC/MS/MS method using a Chromolith RP-18 (50 mm x 4.6 mm) monolithic column was developed and partially validated for the determination of bupropion (BUP), an anti-depressant drug, and its metabolites, hydroxybupropion and threo-hydrobupropion (TB), in human, mouse, and rat plasma. A modern integrated liquid chromatograph and an LC/MS/MS system with a TurboIonSpray (TIS) interface were used for the positive electrospray selected reaction monitoring (SRM) LC/MS analyses. Spiked control plasma calibration standards and quality control (QC) samples were extracted by semi-automated 96-well liquid-liquid extraction (LLE) using ethyl acetate. A mobile phase consisting of 8mM ammonium acetate-acetonitrile (55:45, v/v) delivered isocratically at 5 ml/min, and split post-column to 2 ml/min directed to the TIS, provided the optimum conditions for the chromatographic separation of bupropion and its metabolites within 23s. The isotope-labeled D(6)-bupropion and D(6)-hydroxybupropion were used as internal standards. The method was linear over a concentration range of 0.25-200 ng/ml (bupropion and threo-hydrobupropion), and 1.25-1000 ng/ml (hydroxybupropion). The intra- and inter-day assay accuracy and precision were within 15% for all analytes in each of the biological matrices. The monolithic column performance as a function of column backpressure, peak asymmetry, and retention time reproducibility was adequately maintained over 864 extracted plasma injections.
Borges
Virginia
V
Analytical Toxicology, Department of Population Medicine and Diagnostic Sciences, Cornell University, 927 Warren Drive, Ithaca, NY 14850, USA.
Yang
Eric
E
Dunn
John
J
Henion
Jack
J
D000818:Animals; D016642:Bupropion / Q000097:blood*; D002853:Chromatography, Liquid / Q000295:instrumentation / Q000379:methods*; D006801:Humans; D013058:Mass Spectrometry / Q000295:instrumentation / Q000379:methods*; D051379:Mice; D051381:Rats; D012015:Reference Standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D016642:Bupropion