DIRECT DETERMINATION OF TAMOXIFEN AND ITS 4 MAJOR METABOLITES IN PLASMA USING COUPLED-COLUMN HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY

A rapid, rugged and fully automated method has been developed for the determination of tamoxifen and its major metabolites in plasma. The sy stem is based upon an in-line extraction process combined with column switching to a coupled analytical column. The plasma sample is deprote inated by the addition of acetonitrile before injection onto a semi-pe rmeable surface (SPS) cyano guard column (1.0 x 0.46 cm I.D.). After w ashing the guard column briefly with water, the sample is eluted with a mobile phase composed of 35% acetonitrile in 20 mM potassium phospha te buffer (pH 3). The eluent is directed through a cyano analytical co lumn (25 x 0.46 cm I.D.) and a photochemical reactor where the analyte s are converted to highly fluorescent phenanthrene derivatives. Tamoxi fen, 4-hydroxytamoxifen, N-desdimethyltamoxifen, N-desmethyltamoxifen and tamoxifen-ol are eluted in that order at a flow-rate of 1.0 ml/min . The method has been validated for use in a clinical study utilizing tamoxifen in the treatment of recurrent cerebral astrocytomas.