ABSTRACT
A method for the quantification of mycophenolate mofetil (MMF, CellCept) in plasma using solid-phase extraction and HPLC is described here. A solution of internal standard is added to a 0.5-ml plasma aliquot. The resulting sample is treated with water and dilute HCl and applied to a C18 solid-phase extraction column. After a water wash, the MMF and internal standard are eluted with methanol-0.1 M citrate-phosphate buffer, pH 2.6 (80:20, v/v). A 20-microliters aliquot of the eluate is injected onto a C18 column (5 microns particle size, 150 x 4.6 mm I.D.) and eluted at ambient temperature with acetonitrile-0.05 M citrate-phosphate buffer, pH 3.6, containing 0.02 M heptanesulfonic acid (41:59, v/v). Quantification is achieved by UV detection at 254 nm. The method is reproducible, accurate and specific for MMF. Using 0.5 ml of plasma for analysis, the quantification limit is 0.400 microgram/ml and the range is 0.400-20 micrograms/ml. Based on the stability profile of MMF in plasma, it is recommended that blood samples collected following intravenous infusion be immediately stored on ice and that plasma be prepared rapidly, immediately stored frozen at -80 degrees C and analyzed within four months of collection.
Subject(s)
Chromatography, High Pressure Liquid/methods , Immunosuppressive Agents/blood , Mycophenolic Acid/analogs & derivatives , Chromatography, High Pressure Liquid/statistics & numerical data , Drug Stability , Humans , Hydrogen-Ion Concentration , Kinetics , Mycophenolic Acid/blood , Organ Transplantation , Quality Control , Sensitivity and SpecificityABSTRACT
A manual and an automated (Zymark PyTechnology robot) HPLC method for simultaneous determination of plasma mycophenolic acid (MPA) and its glucuronide conjugate (MPAG) are described here. Both methods are reproducible and accurate, and both are equivalent in all respects, including quantification limits (MPA, 0.100 microgram/ml; MPAG, 4.00 micrograms/ml), range (using 0.05-0.5 ml of plasma: MPA, 0.0500-20.0 micrograms/aliquot; MPAG, 2.00-200 micrograms/aliquot), precision, and accuracy. MPA and MPAG were stable under the conditions used with both methods. Results from aliquots of paired control samples, analyzed by the manual method over three years at six analytical laboratories, showed excellent agreement in precision and accuracy.
