Rapid and simple determination of mycophenolic acid in human plasma by ion-pair RP-LC with fluorescence detection

Mycophenolic acid (MPA) is an immunosuppressive drug given as the prodrug o f mycophenolate mofetil (MR IF). In order to investigate the pharmacokineti cs of MPA, a simple, specific, sensitive and reliable method has been estab lished for the quantitative determination of MPA in plasma from renal trans plant recipients. The method involves a single-step protein precipitation p rocedure and a specific determination by ion-pair reversed-phase high-perfo rmance liquid chromatography (HPLC) with fluorescence detection. Separation was achieved on a Cls column (150 x 4.6 mm, 5 mum) with a mobile phase com posed of berate buffer (pH 10.0; 50 mM) - acetonitrile - tetrabutylammonium bromide (200 mM) (75.25:1, v/v/v). The fluorescence detector was set at 31 0 (excitation) and 430 nm (emission). Following protein precipitation with ice-cold acetonitrile, clear supernatants (50 mul) were injected into the H PLC system. The retention time of MPA was approximately 4.5 min. The HPLC r un time was 8 min. The assay was linear in concentration range 0.2-20.0 mug /ml for MPA in human plasma. Precision of the assay in the concentration ra nge examined was from 0.89 to 3.21% for the intra-assay run and from 3.01 t o 4.35% for the inter-assay run. A limit of detection was 0.05 mug/ml at a signal-to-noise ratio of 3. This validated method was then applied to the d etermination of MPA concentrations in renal transplant recipients after ora l administration of 0.75 g of MMF. (C) 2001 Elsevier Science B.V. All right s reserved.