DETERMINATION OF UNBOUND ETOPOSIDE CONCENTRATION IN ULTRAFILTERED PLASMA BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY WITH FLUOROMETRIC DETECTION

Etoposide is a highly protein bound drug, and monitoring the concentra tion of free drug could help individualize dosage in oncological patie nts. The cost and difficulty of the standard techniques (equilibration dialysis) has hampered the monitoring of free drugs. We describe a si mple HPLC method for the measurement of free etoposide concentration i n plasma. Sample preparation involves the ultrafiltration of plasma by a Centrifree device for 30 min at 2000 g and extraction with chlorofo rm. The isocratic separation is performed with a mu Bondapak phenyl an alytical column. Fluorimetric detection is used (288-328 nm excitation and emission wavelengths). Linearity of the calibration curve is exce llent between 0.05 and 1 mu g/ml. Accuracy and precision are reported at the concentrations 0.06 and 0.4 mu g/ml: within-run accuracy is 10% and 6.2%, respectively; between-run accuracy is less than or equal to 1%; within-run coefficients of variation (C.V.) are 10.6 and 5.0%; be tween-run C.V. are 11.6 and 6.8% respectively. The range of the assay is 0.05 to 1 mu g/ml. The feasibility of the technique has been tested in 7 patients treated with oral etoposide for hepatocarcinoma (mean p rotein binding 91%). We found no interference from endogenous substanc es, co-administered drugs (alizapride, furosemide, ranitidine) and oth er antineoplastic agents (doxorubicine, idarubicine, vinblastine, vino relbine).