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A simple procedure for the determination of chlorophacinone in human serum by high-performance liquid chromatography is reported. After addition of a suitable internal standard, unknown samples were extracted at acid pH with ethyl acetate. The reconstituted methanolic extract was directly injected into the liquid chromatograph. Separation was performed with a Lichrospher C18 5-microns column with a methanol-phosphate buffer (85:15, v/v) mobile phase. The eluent was monitored with UV detection at 286 nm. The method was found to be sufficiently rapid, reproducible, selective, and sensitive for use in clinical and experimental pharmacokinetic studies. Application of this method to 20 intoxicated patients indicated that the risk of bleeding syndrome, which is always associated with this kind of intoxication, is no longer present when chlorophacinone levels drop to below 1 microgram/mL.