High-Pressure Liquid Chromatographic Analysis of Eflornithine in Serum

Eflornithine [2-(difluoromethyl)-DL-ornithine monohydrochloride, monohydrate; DFMO] is an ornithine decarboxylase inhibitor used to treat Pneumocystis carinii pneumonia (PCP) in AIDS patients unresponsive or intolerant to conventional drug therapy. A reversed-phase HPLC method utilizing precolumn dansylation is described which permits the analysis of DFMO in serum of AIDS patients in support of pharmacokinetic studies. Norvaline (2-amino valeric acid) was added as the internal standard and the sample was extracted with three portions of ice-cold 80% ethanol. The supernatants were evaporated in a vacuum oven at 50 degrees C, and the residue was dissolved in base and derivatized at 25 degrees C for 4 h in the dark using dansyl chloride in acetone. Dilute base was then added and 25 to 50 microL was injected into the HPLC. Chromatography was performed on a C-8, 15-cm column with a linear gradient from a 95:5 solution of 10 mM pH 4 acetate buffer and THF, to a 90:10 solution of acetonitrile and THF over a period of 28 min. Detection was by UV at 330 nm, and DFMO and norvaline eluted at 14.7 and 17.7 min, respectively. The method was linear over a range of 5 to 950 micrograms/mL, and replicate analyses of a 25-micrograms/mL control specimen produced a between-run coefficient of variation of 6%. No interferences were encountered in a variety of patient and normal serum blanks.