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In this research project, a high-performance liquid chromatography (HPLC) method was developed for the determination of acyclovir (ACV) in plasma. The plasma samples, recharged with acyclovir and in presence of 5’,-N-methylcarboxyamidoadenosine (MECA) as an internal standard, were purified using a solid-phase extraction technique with Waters Oasis HLB columns. The separation of the components from the extract was carried out in a LiChrospher 100 RP-18 column for further ultraviolet detection at a wavelength range of 250-260 run. The mobile phase composition was 18% acetonitrile, sodium dodecylsulphate. 5 mM and phosphate buffer at pH 2.6 with an analysis time of 13 min per sample. The average retention time for acyclovir was of 5.0 min and for the internal standard 11.2 min. The calibration curve was linear ranging between 0.05 and 1.80 mug/ml. The detection limit was 0.006 mug/ml with a quantification limit of 0.020 mug/ml. The ACV recuperation percentage for 250 mul of plasma was between 94.7 and 109.7% with a coefficient of variation not higher than 5.2%. This method was developed and validated for use in bioavailability and bioequivalence studies. (C) 2003 Elsevier Science B.V. All rights reserved.