SAMPLE CLEANUP AND HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHIC TECHNIQUESFOR MEASUREMENT OF WHOLE-BLOOD RAPAMYCIN CONCENTRATIONS

Determination of rapamycin (RAPA), a potential adjunct to cyclosporin immunosuppressive therapy in solid organ transplantation, presents a c hallenge because of the agent's unusual chemical properties. RAPA is n ot readily extracted from biological matrices, is light-sensitive, and exists in numerous isomeric forms. Using liquid-liquid extraction tec hniques with tert.-butyl methyl ether followed by ethanol, one achieve s 96% recovery of RAPA and 70% recovery of the internal standard, beta -estradiol-3-methyl ether, from human whole blood. Subsequent high-per formance liquid chromatography (HPLC) utilizing two heated reversed-ph ase C18 columns in tandem and an 85% methanol-water mobile phase with ultraviolet detection at 276 nm provides measurement of RAPA from 1-ml samples of human whole blood. The chromatography requires less than 4 0 min per sample. The assay exhibits less-than-or-equal-to 10% standar d error of the mean and less-than-or-equal-to 12% coefficient of varia tion over the concentration range 2-100 ng/ml. The method has been tes ted using pharmacokinetic profiles from renal transplant recipients re ceiving bolus intravenous RAPA infusions.