Simultaneous determination of udenafil and its active metabolite, DA-8164, in human plasma and urine using ultra-performance liquid chromatography-tandem mass spectrometry: Application to a pharmacokinetic study

A rapid, sensitive, and simple ultra-performance liquid chromatography-tandem mass spectrometry (UPLC/MS/MS) method for the determination of udenafil and its active metabolite, DA-8164, in human plasma and urine using sildenafil as an internal standard (IS) was developed and validated. Udenafil, DA-8164 and IS from a 100 μL aliquot of biological samples were extracted by protein precipitation using acetonitrile. Chromatographic separation was carried on an Acquity UPLC BEH C₁₈ column (50 × 2.1 mm, i.d., 1.7 μm) with an isocratic mobile phase consisting of acetonitrile and containing 0.1% formic acid (75:25, v/v) at flow rate of 0.4 mL/min, and total run time was within 1 min. Detection and quantification was performed by the mass spectrometer using multiple reaction-monitoring mode at m/z 517 → 283 for udenafil, m/z 406 → 364 for DA-8164 and m/z 475 → 100 for IS. The assay was linear over a concentration range of 1-600 ng/mL with a lower limit of quantification of 1 ng/mL in both human plasma and urine. The coefficient of variation of this assay precision was less than 13.7%, and the accuracy exceeded 92.0%. This method was successfully applied for pharmacokinetic study after oral administration of udenafil 100 mg to healthy Korean male volunteers.