A simple, selective and sensitive LC-MS/MS assay has been developed for the determination of minocycline in human plasma. Plasma samples were prepared by protein precipitation, followed by chromatographic separation on a HyPURITY™ C8 (100 mm × 4.6 mm, 5 µm) column under isocratic conditions. The LOD and LLOQ of the method were 0.50 and 20.0 ng/ml, respectively. The intra-batch and inter-batch precision (% coefficient of variation) across quality control levels was less than 8.0%. For a set of incurred samples the percentage change in concentration was within ± 9.0%. The method was successfully applied to a bioequivalence study of 135 mg minocycline tablet formulation in 14 healthy Indian males. The reproducibility in the measurement of study data was demonstrated by incurred sample reanalysis.