Determination of WR-1065 in human blood by high-performance liquid chromatography following fluorescent derivatization by a maleimide reagent ThioGlo3

In order to improve the sensitivity and stability of human blood samples containing WR-1065 (i.e., active metabolite of the cytoprotective agent amifostine), a high-performance liquid chromatographic method was developed and validated using fluorescent derivatization with ThioGlo3. Using a sample volume of only 100 microl, the method was specific, sensitive (limit of quantitation=10 nM in deproteinized blood or 20 nM in whole blood), accurate (error < or = 3.2%) and reproducible (CV < or = 8.7%). In addition, the stability of WR-1065 in deproteinized and derivatized blood samples was assured for at least four weeks at -20 degrees C. This method should be particularly valuable in translating the kinetic-dynamic relationship of WR-1065 in preclinical models to that in cancer patients.