SIMULTANEOUS DETERMINATION OF MYCOPHENOLIC-ACID AND ITS GLUCURONIDE IN HUMAN PLASMA USING A SIMPLE HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY PROCEDURE

We describe a reversed-phase HPLC method for determination of total my cophenolic acid (MPA), its free concentration (MPAf), and the glucuron ide metabolite (MPAG), based on simple sample preparation and gradient elution chromatography. The compounds were quantified in parallel by absorbance at 254 nm and 215 nm in the internal standard mode. Lineari ty was verified up to 50 mg/L for MPA and up to 500 mg/L for MPAG (r > 0.999). Detection limits at 215 and 254 nm were, respectively, 0.01 an d 0.03 mg/L for MPA, and 0.03 and 0.1 mg/L for MPAG. The recovery of M PA was 95-106%;recovery of MPAG was 96-106%. The imprecision (CV) for MPA (0.2-25 mg/L) was <8.4% (254 nm) and <4.4% (215 nm) within day (n = 12) and <9.2% (254 nm) and <6.2% (215 nm) between days (n = 12). The imprecision for MPAG (10-250 mg/L) was <4.9% (254 nm) and <3.4% (215 nm) within day, and <6.1% (254 nm) and <5.9% (215 nm) between days. Fo r quantification of MPAf, 100 mu L of ultrafiltrate was applied direct ly to the column. The detection limit was 0.005 mg/L at 215 nm and 0.0 15 mg/L at 254 nm. In the range between 18-210 mu g/L, the within-day CVs were <11.8% (n = 12) and the between-day CVs were <15.8% (n = 12).