Quantification of free mycophenolic acid by high-performance liquid chromatography-atmospheric pressure chemical ionisation tandem mass spectrometry

To facilitate the investigation of free mycophenolic acid concentrations we developed a high-performance liquid chromatography tandem mass spectrometr y method using indomethacin as an internal standard. Free drug was isolated from plasma samples (500 mul) using ultrafiltration, The analytes were ext racted from the ultrafiltrate (200 mul) using C-18 solid-phase extraction. Detection was by selected reactant monitoring of mycophenolic acid (m/z 318 .9-->190.9) and the internal standard (m/z 356.0-->297.1) with an atmospher ic pressure chemical ionisation interface. The total chromatographic analys is time was 12 min. The method was found to be linear over the range invest igated, 2.5-200 mug/l (r>0.990, n=6). The relative recovery of the method f or the control samples studied (7.5, 40.0 and 150 mug/l) ranged from 95 to 104%. The imprecision of the method, expressed in terms of intra- and inter -day coefficients of variation, was <8 and <9%, respectively. Further, anal ysis of pooled patient plasma produced an intra-day imprecision of 6.6%. Th e signal-to-noise ratio at the limit of quantification (2.5 mug/l) was appr oximately 5:1. The mean absolute recovery (n=6) of mycophenolic acid and th e internal standard were 76.0+/-13.5% and 86.0+/-9.1%, respectively. The me thod reported provides an accurate and precise quantification of free mycop henolic acid over a wide analytical range and thus can be used for routine monitoring and pharmacokinetic studies. (C) 2000 Elsevier Science B.V. All rights reserved.