Abstract: OBJECTIVE RP-HPLC method detection for simultaneous determination of Mycophenolic acid (MPA), its phenol glucuronide metabolite (MPAG) and acyl-MPAG (AcMPAG) in human plasma was established. METHODS The plasma samples were prepared with protein-precipitating reagent, and the supernatant was eluted on Zorbax column (250 mm×4.6 mm, 5 mm) with 20 mmol×L^-1 NaH₂PO₄ buffer (pH 3.0, adjusted with 20% phosphoric acid) and methanol (45∶55) at 304 nm. The column temperature was 45 ℃, and the flow rate was 1.2 mL×min^-1. RESULTS The assay was linear within the range of 0.2-50 mg×mL^-1 for MPA (r=0.999 7), 2.8-531 mg×mL^-1 for MPAG(r=0.999 9), and 0.3-24 mg×mL^-1 for AcMPAG(r=0.999 4). Mean absolute recovery of MPA and its metabolites was more than 80%. The average recoveries of MPA, MPAG and AcMPAG were 94%.0–101.4%, 98.4%–101.9% and 96.1%–104.2 %, respectively. The RSD of within-day and between-day were all lower than 15%. CONCLUSION The method described is sensitive, reproducible, and will be useful in TDM or pharmacokinetic studies of MPA.