UPLC–MS/MS for the Simultaneous Determination of Sildenafil and N-Desmethyl Sildenafil.

A sensitive high-performance liquid chromatography–tandem mass spectrometry method was established for the simultaneous determination of sildenafil and N -desmethyl sildenafil in human plasma. The protein precipitation was used for extraction and the gradient elution of the mobile phase A of water (containing 0.01% formic acid) and the mobile phase B of acetonitrile, and methanol (V:V = 1:1, containing 0.01% formic acid) was used for chromatographic separation on a C18 column. Quantification was performed by multiple reaction monitoring mode to monitor the precursor-to-product ion transitions of m/z 475.4 →  m/z 283.3 for sildenafil, m/z 461.4 →  m/z 283.2 for N -desmethyl sildenafil, m/z 483.3 →  m/z 108.1 for sildenafil-d8 (IS) and m/z 469.2 →  m/z 283.3 for N -desmethyl sildenafil-d8 (IS) at the positive ionization mode. The intra- and inter-day relative standard deviations were less than 6.8% and 4.1% for sildenafil and N -desmethyl sildenafil, respectively. Accuracy at four levels ranged from 93.1% to 115.9% for sildenafil and 95.6% to 112.5% for N -desmethyl sildenafil. The present method was sensitive and reliable for simultaneous quantification of sildenafil and its active metabolite and was successfully applied to a pharmacokinetic study of an oral low dose of sildenafil in Chinese healthy volunteers. [ABSTRACT FROM AUTHOR]
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