The essential oil of Ajuga pseudoiva, collected from Tunisia, was analyzed using gas chromatography-mass spectroscopy. Thirty-two compounds accounting for 95.76% of the total oil were identified. Sesquiterpenes were found to be the most abundant components of A. pseudoiva oil. And they were mainly represented by viridiflorol (30.17%), germacrene B (9.26%) (alpha, beta and gamma)-eudesmol (8.11%) and aromadendrene (7.45%). The essential oil of A. pseudoiva showed radical scavengers activity (IC50 = 0.72 mg/mL) and displayed lipid peroxidation inhibitory activity (IC50 = 0.6 mg/mL). A. pseudoiva essential oil was also found to exhibit a dose-dependent ACE inhibitory activity with an IC50 value of 65.5 mu g/mL. Moreover, the antimicrobial activity of the essential oil was tested against 17 species of microorganisms, and the results obtained showed significant antibacterial activity against the Gram-positive and Gram-negative bacteria, with inhibition zones and minimal inhibitory concentration values of 14-32 mm and 84-137 mu g/mL and 6-21 mm and 105-336 mu g/mL, respectively. Higher activity was also found against several fungal strains. Crown Copyright (c) 2013 Published by Elsevier Ltd. All rights reserved.