A Sensitive LC-MS/MS Method for the Determination of Afatinib in Human Plasma and Its Application to a Bioequivalence Study.

A high performance liquid chromatography-tandem mass spectrometry assay for the determination of afatinib (AFT) in human plasma was established. A simple sample preparation of protein precipitation was used and separation was achieved on a C18 column by the gradient mixture of mobile Phase A of water (containing 0.1% ammonia) and the mobile Phase B of acetonitrile and water (V:V = 95:5, containing 0.2% ammonia). The multiple reaction monitoring mode was used to monitor the precursor-to-production transitions of m/z 486.2 â†’ m/z 371.4 for AFT and m/z 492.2 â†’ m/z 371.3 for AFT-d6 (internal standard) at positive ionization mode. The calibration curve ranged from 0.100 to 25.0 ng·mL-1 and the correlation coefficient was greater than 0.99. The intra- and inter-batch precision was less than or equal to 10.0%. Accuracy determined at four concentrations was in the range of 92.3-103.3%. In summary, our method was sensitive, simple and reliable for the quantification of AFT and was successfully applied to a bioequivalence study.

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 â†’ m/z 283.3 for sildenafil, m/z 461.4 â†’ m/z 283.2 for N-desmethyl sildenafil, m/z 483.3 â†’ m/z 108.1 for sildenafil-d8 (IS) and m/z 469.2 â†’ 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.

An improved LC-MS/MS method for determination of cinacalcet in human plasma and its application to the evaluation of food intake effect on the pharmacokinetics of cinacalcet in healthy volunteers.

A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the determination of cinacalcet in human plasma was developed and validated. This assay was based on liquid-liquid extraction and cinacalcet-d4 was used as an internal standard (IS). Separation was achieved on a C18 column by the mobile phase A of water (containing 0.1% formic acid) and the mobile phase B of acetonitrile-water (95:5, v/v) (containing 0.2% formic acid) with gradient elution. Quantification was done using multiple reaction monitoring mode to monitor the precursor-to-product ion transitions of m/z 358.2 → m/z 155.2 for cinacalcet and m/z 362.3 → m/z 155.0 for IS at positive ionization mode. The calibration curve was established over the range 0.05-20.0 ng/mL and the correlation coefficient was >0.99. The intra- and inter-day relative standard deviations were <5.8%. Accuracy determined at four concentrations ranged between 96.0 and 106.0%. This method was successfully applied to a pharmacokinetic description of oral dose of cinacalcet and the significant effect of food intake on the pharmacokinetics of cinacalcet was first demonstrated in Chinese healthy volunteers.