A rapid and sensitive fluorimetric method for the determination of prenalterol hydrochloride is presented, based on the oxidation of the analyte with potassium hexacyanoferrate(III) in a slightly alkaline medium (pH 9.23). The different experimental parameters were carefully studied and incorporated into the procedure. The oxidation product exhibits a blue fluorescence with its emission maximum at 427 nm, and excitation maximum at 314 nm. Fluorescence intensity is a linear function of prenalterol hydrochloride concentration over the range of 0.2-3.6 microg/ml(-1) in the solution finally measured. The method was successfully applied to the determination of prenalterol hydrochloride in pharmaceutical formulations and biological fluids. A proposal for the reaction pathway is suggested.