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Objectives: This study describes an assay applied to determine the pharmacokinetic parameters of acetylsalicylic acid following oral administration of 1000 mg of this drug in human. Materials and method: In the assay, a reversed-phase high-performance liquid chromatography method (column: Chromtech C8 3µm silica, 100×4.6 mm I.D.; mobile phase: acetonitrile/water: (5/95 v/v) buffer phosphate 0.05M pH 2.15 and acetonitrile/water: (25/75 v/v) buffer phosphate 0.05M pH 2.15; gradient mode; and UV detection at 230 nm) was used to simultaneously quantify acetylsalicylic acid and its main metabolites, including salicylic acid, gentisic acid, and salicyluric acid in urine, which was deproteinized with acetonitrile and centrifuged. Results: The method allows the determination of acetylsalicylic acid, salicylic acid, salicyluric acid and gentisic acid in urine as low as 13, 27.4, 21 and 9.3 µg/ml respectively with good precision (better than 5%). Conclusions: A fast and easy to perform reversed-phase HPLC method has been developed for the simultaneous determination of acetylsalicylic acid and its major metabolites in urine, in which no time-consuming extraction procedure is needed, simple mobile phases are used, and good precision and accuracy are achieved. The method can be further optimized and improved to be used to study the pharmacokinetic of acetylsalicylic acid. Keywords: Acetylsalicylic acid, high-performance liquid chromatography.

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