Expedient microdetermination of itraconazole and hydroxyitraconazole in plasma by high-performance liquid chromatography with fluorescence detection

The authors describe a useful and rapid micromethod for the analysis of itraconazole (ITZ) and its active metabolite hydroxyitraconazole (HIT) in human plasma. After a simple deproteinization of 100 muL plasma with acetonitrile, the drug, its metabolite, and an internal standard (IS, ketoconazole) were separated on an 8 mm x 10 cm NovaPak (Waters Associates; Milford, MA) C-18 4-mum particle-size radial compression cartridge. The compounds of interest were detected using a fluorescence detector with the excitation wavelength set at 260 nm and the emission at 365 nm. The mobile phase consisted of 420 mL water adjusted to a pH of 2.5 with phosphoric acid, 580 mL acetonitrile, and 100 muL triethylamine, which was delivered at a flow rate of 3.0 mL/min. This expedient and rugged method is being used to monitor therapeutic levels in bone marrow transplant recipients who are taking the drug for prophylaxis.