| Summary: | Olive leaves are rich in phenolic compounds, which give them antioxidant properties that are associated with a lower incidence of disease. Therefore, the aim of this work was to determine the phenolic content, antioxidant activity, and toxicity of the aqueous extracts of olive leaves of the main Spanish and Greek cultivated and wild genotypes. For these purposes, ‘Picual’ and ‘Arbequina’ leaves from Spain and ‘Koronoeiki’ and ‘Kalamon’ leaves from Greece were collected, as were wild olive leaves from both countries. The aqueous extracts of these genotypes were analyzed by HPLC-DAD, and the DPPH·, ABTS·<sup>+</sup> Folin–Ciocalteu, and Microtox<sup>®</sup> methods were also used. ‘Picual’ had the highest oleuropein values, followed by wild olive leaves from both countries and ‘Arbequina’. The latter was reflected in the antioxidant activity measured by DPPH· and ABTS·<sup>+</sup>, which positioned the leaves of ‘Arbequina’, ‘Picual’, and the wild genotypes as having the most antioxidant activity. As expected, these leaves also had the highest total phenol content, as measured by Folin–Ciocalteu. Regarding the inhibition of the bioluminescence of <i>Aliivibrio fischeri</i> of the aqueous leaf extracts measured by Microtox<sup>®</sup>, the EC50<sub>15</sub> ranged between 11.82 and 82.50 mg/mL, demonstrating similar behavior to other herbal infusions.
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