Sensitive liquid chromatography assay with ultraviolet detection for a new phosphodiesterase V inhibitor, DA-8159, in human plasma and urine

A sensitive high-performance liquid chromatographic (HPLC) method with ultraviolet absorption detection (292nm) was developed and validated for the determination of the new phosphodiesterase V inhibitor, DA-8159 (DA), in human plasma and urine. A single step liquid-liquid extraction procedure using ethyl ether was performed to recover DA and the internal standard (sildenafil citrate) from 1.0ml of biological matrices combined with 200μl of 0.1M sodium carbonate buffer. A Capcell Pak C₁₈ UG120 column (150mmx4.6mm I.D., 5μm) was used as a stationary phase and the mobile phase consisted of 30% acetonitrile and 70% 20mM potassium phosphate buffer (pH 4.5) at a flow rate of 1.0ml/min. The lower limit for quantification was 5ng/ml for plasma and 10ng/ml for urine samples. Within- and between-run accuracy and precision were ≤15 and ≤10%, respectively, in both plasma and urine samples. The recovery of DA from human plasma and urine was greater than 70%. Separate stability studies showed that DA is stable under the conditions of analysis. This validated assay was used for the pharmacokinetic analysis of DA during a phase I, rising dose study.