Validated LC-MS/MS method for quantitation of total and free mycophenolic acid concentration and its application to a pharmacokinetic study in pediatric renal transplant recipients.

A simple and sensitive LC-MS/MS method was established to quantify total and free mycophenolic acid (MPA) plasma concentrations during immunosuppressive medication for pediatric renal transplantation. The chromatographic separation was performed with the Hypersil GOLD C column, using a mobile phase consisting of 0.1% formic acid in water and acetonitrile (60:40, v/v) at an isocratic flow rate of 0.4 ml/min. An Agilent 6420 triple quadrupole mass spectrometer was operated via a positive electrospray ionization interface using the transitions m/z 321.14 → 206.9 for MPA and m/z 324.15 → 209.9 for MPA-d3 (internal standard). The linearity was 0.1-50 μg/ml for total MPA and 0.0025-0.5 μg/ml for free MPA. The within-run and between-run precisions were all <5% and accuracy was within 96.23-107.63%. The validated method was successfully aspplied to a pharmacokinetic study in 28 pediatric renal recipients. The mean free fraction of MPA in our patients was 0.89% (ranging from 0.62 to 1.25%) and albumin level played a major role in the variability of free fraction of MPA, thus, in pediatric patients with hypoproteinemia, close free drug monitoring and dose adjustments should be considered to prevent toxicity.