This paper describes a high-performance liquid chromatography method for the determination of atenolol in human plasma. Atenolol and the internal standard, metoprolol, were extracted from plasma by using a liquid-liquid extraction method. The method was developed on an Ace C18 reverse-phase column using a mobile phase of methanol-water (50:50, v/v) containing 0.1 trifluoroacetic acid. The calibration curve was linear within the concentration range of 5-150 ng/mL. Intra-day and inter-day precision values for atenolol in plasma were less than 6.1, and accuracy (relative error) was better than 5.5. The mean recovery of atenolol was 98.4 for plasma. The limits of detection and quantification of atenolol were 1.5 and 5 ng/mL, respectively. Also, this assay was successfully applied to six patients with hypertension who had been given an oral tablet of 50 mg atenolol. © 2012 The Author [2012]. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissionsoup.com.