Simultaneous determination of unlabeled and deuterium-labeled indinavir in human plasma by high-performance liquid chromatography with tandem mass spectrometric detection

A method for the simultaneous determination of indinavir and its hexadeuterated analog (d(6)-indinavir) in human plasma is described. Isolation of the analytes and internal standard from plasma was achieved via liquid-liquid extraction with methyl-t-butyl ether. The analytes were chromatographed under reversed-phase conditions on a BDS-Hypersil C8 column. A Sciex API III+ tandem mass spectrometer equipped with a turbo ion-spray interface was used as the detector. Multiple reaction monitoring using the parent --> product ion combinations of m/z 614 --> 465, 620 --> 471 and 654 --> 505 were used to quantify indinavir, d(6)-indinavir, and internal standard, respectively. The method was validated, using 1-mL aliquots of plasma, in the concentration range in plasma of 1 to 200 ng/mL. Precision of the assay, as measured by the coefficient of variation, ranged from 0.9 to 4.3% and 0.9 to 6.2% for indinavir and d(6)-indinavir, respectively. Indinavir assay accuracy ranged from 95.8 to 105.0% of nominal, whereas the accuracy of the assay for d(6)-indinavir ranged from 97.4 to 104.0% of nominal. The assay was used to support a clinical study in which the stable isotope technique was used to determine the bioavailability of indinavir.