Simultaneous quantitation of etoposide and its catechol metabolite in human plasma using high-performance liquid chromatography with electrochemical detection

Etoposide, a highly active and widely used antineoplastic agent, is O-demet hylated to its active catechol metabolite. A high-performance liquid chroma tographic assay method for the simultaneous quantitation of etoposide and e toposide catechol in human plasma was established. Etoposide and etoposide catechol were extracted from plasma using chloroform and methanol followed by phase separation, evaporation of the organic phase, and reconstitution o f the residue. Chromatography was accomplished using a reversed-phase pheny l analytical column (390 mmx3.9 mm I.D.) with a mobile phase of 76.6% 25 mM citric acid-50 mM sodium phosphate (pH 2.4)-23.4% acetonitrile pumped isoc ratically at 1 ml/min with electrochemical detection. The limit of detectio n for etoposide was 1.2 nM and for etoposide catechol was 0.2 nM. The preci sion (CV) for etoposide ranged from 0.7 to 3% and for the catechol metaboli te from 1 to 6%; accuracy of predicted values ranged from 97 to 106% and 94 to 103%, respectively. The assay was linear from 0.1 to 10 mu M for etopos ide and from 0.005 to 0.5 mu M for etoposide catechol in plasma. Recovery o f etoposide and etoposide catechol ranged from 93 to 95% and 90 to 98%, res pectively. Stability of etoposide and etoposide catechol in human plasma co ntaining ascorbic acid stored at -70 degrees C for one year was demonstrate d. This assay procedure is suitable for evaluation of etoposide and etoposi de catechol pharmacokinetics in plasma following etoposide administration. (C) 1999 Elsevier Science B.V. All rights reserved.