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Quantification of azithromycin in human plasma by liquid chromatography tandem mass spectrometry: application to a bioequivalence study.

A specific, sensitive and rapid liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) method has been developed and validated for the determination of azithromycin in human plasma. After deproteinizing the plasma sample with methanol, azithromycin and internal standard (IS: roxithromycin) were separated using a mobile phase comprised of acetonitrile : ammonium acetate buffer (50 mM, containing 0.05% acetic acid)=85:15 on a Hypersil GOLD C18 column (50 mm×2.1 mm ID, dp 1.9 μm). Detection was performed with a tandem mass spectrometer by selective reaction monitoring (SRM) through electrospray ionization. Target ions were monitored at [M+H]+ m/z 749.5→591.5 and 837.7→679.5 in positive electrospray ionization (ESI) mode for azithromycin and IS respectively. Linearity was established for the range of concentrations 2-800 ng/mL with a coefficient of correlation (r) of 0.9996. The lower limit of quantification (LLOQ) was identifiable and reproducible at 2.0 ng/mL. Both intra- and inter-batch standard deviations were less than 15%. The validated method was successfully applied to study the comparative bioavailability of azithromycin for suspension in test vs. reference in healthy Chinese volunteers through the statistical comparison of pharmacokinetic parameters obtained with the two formulations.