[Determination of sildenafil and vardenafil in human plasma by high performance liquid chromatography coupled with liquid-liquid-liquid microextraction].

High performance liquid chromatography coupled with liquid-liquid-liquid microextraction was developed for the simultaneous determination of sildenafil and vardenafil in human plasma. The effects of extraction solvent, the volume of organic solvent, dropsize of acceptor phase, stirring rate and extraction time on the enrichment factors of analytes were investigated. The optimized experimental conditions, 300 microL toluene as the organic phase, 2 microL 0.2 mol/L HCl as the acceptor phase, 600 r/min of the stirring rate, and 40 min of the extraction time, were gotten. Under these conditions, high enrichment factors were obtained. The linear range of studied analytes was from 5 microg/L to 1.0 mg/L. The relative standard deviation was lower than 5%. The limits of detection were 1 microg/L for sildenafil and 0.5 microg/L for vardenafil at signal-to-noise ratio of 3. The method with little solvent consumption may provide high analyte preconcentration and excellent sample clean-up, and it is a sensitive and suitable method for simultaneous determination of the above two substances in human plasma.