Determination of 4-hydroxyifosfamide in biological matrices by high-performance liquid chromatography

A high-performance liquid chromatographic method has been developed for the determination of 4-hydroxyifosfamide, a metabolite of ifosfamide, in plasma of cancer patients. The analyte is derivatized to 7-hydroxyquinoline, which can be detached fluorimetrically. The calibration graph is linear in the concentration range 0.05-25 microM, the limit of detection being 40 nM. Any inference from acrolein, another metabolite of ifosfamide, was ruled out. 4-Hydroxyifosfamide is very unstable in plasma and a stabilization procedure by adding citric acid has been developed. Thus treated, the samples were stable for 4 days. Analysis of a patient's plasma samples revealed that the 4-hydroxifosfamide concentration did not exceed 10 microM.