High-performance liquid chromatographic assay with ultraviolet detection for quantification of dihydrofluorouracil in human lymphocytes: application to measurement of dihydropyrimidine dehydrogenase activity

The anticancer drug 5-fluorouracil (5FU) undergoes extensive biotransformat ion to 5-dihydrofluorouracil (5FUH2) by the enzyme dihydropyrimidine deshyd rogenase (DPD). A new HPLC method with direct UV detection for the determin ation of 5FUH2 in peripheral lymphocytes has been developed to detect DPD d eficiency in patients treated with 5FU-based therapy. The method has been s hown to be valid over the 5FUH2 concentration range of 1.14-37.88 nmol/ml. Optimal enzymatic conditions for DPD activity measurement were studied: inc ubation time, protein and 5FU concentrations. The assay was successfully cr oss-validated with the existing method using HPLC with radiochemical detect ion. (C) 2001 Elsevier Science B.V. All rights reserved.