Analysis of 5-fluorouracil in plasma and urine by high-performance liquid chromatography

A relatively simple, sensitive and rapid high-performance liquid chromatographic method is described for measuring the anticancer drug 5-fluorouracil (5-FU) in human plasma and urine. The procedure includes liquid-liquid extraction using ethyl acetate-methanol (95:5) and preparative column chromatography to separate 5-FU from constituents normally occurring in these biological samples. The columns contained a specially modified form of diatomaceous earth, which requires no pre-conditioning washes. Reversed-phase high-performance liquid chromatography was performed on a C18 column (70 mm x 4.6 mm I.D.) with a mobile phase of water-methanol (95:5) and ultraviolet detection (268 nm). The overall recovery from plasma and urine was 91 and 94%, respectively, at the concentration of 50 ng/ml. The determination limit of the assay for 5-FU was 10 ng/ml of plasma and urine. Concentrations of 5-FU between 10 and 500 ng/ml were measured in plasma and urine with a relative standard deviation of 6.8%. In order to evaluate the procedure, plasma and urine samples from three patients treated with 5-FU by continuous intravenous perfusion, were investigated.