Quantitation of β-lapachone and 3-hydroxy-β-lapachone in human plasma samples by reversed-phase high-performance liquid chromatography

β-Lapachone is an o-naphthoquinone found to have in vitro cytotoxicity in cancer cells, type I human immunodeficiency virus, and fungi. Analytical methods for evaluating β-lapachone in biological fluids using high-performance liquid chromatography (HPLC) have not been published. The reversed-phase HPLC method described in this report utilizes liquid extraction of a 0.5-ml plasma sample with average recoveries of 67 ± 10.8% and 70 ± 10.3% for β-lapachone and 3-hydroxy-β-lapachone, respectively. Sensitivity of the assay using ultraviolet (UV) detection at 256 nm is 15 ng ml^-1 from a 100 μl injection. Plasma standards for β-lapaehone and 3-hydroxy-β-lapachone are linear with no significant difference in slope between the compounds. The retention times of 2.7 min for 3-hydroxy-β-lapachone and 5.9 min for β-lapachone result in a clean separation permitting use of the same assay procedure without modification for both compounds. This assay offers the advantage that either β-lapachone or 3-hydroxy-β-lapachone can serve as the internal standard, depending on which compound is being analyzed.