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An HPLC method has been developed for the simultaneous determination of the toxic Aconitum alkaloids, aconitine, mesaconitine and hypaconitine in blood and urine samples. The samples were initially subjected to solid phase extraction using Oasis(R) MCX cartridges, and the alkaloids were separated on an XTerra(TM) RP18, column, gradient-elluted with acetonitrille: ammonium hydrogen carbonate buffer. Calibration curves were linear in the range 2.75-550 ng for aconitine and hypaconitine, and 3-600 ng for mesaconitine: the limit of detection was 0.1 ng (signal-to-noise ratio of 3) for each alkaloid. The described analysis proved to be sensitive, rapid and economical, and will be applied in the identification and determination of these alkaloids in forensic and therapeutic drug monitoring. Copyright (C) 2004 John Wiley Sons, Ltd.