In Brazil, the use ofEucalyptusis focused on the production of wood or pulp for the paper industry but without any general recovery of waste, with leaves and branches being left on the ground. One possibility is to use these residues as raw materials in the production of industrially relevant and value-added compounds such as essential oil. The aim of the present study was to investigate the chemical composition, yield, anti-inflammatory/antinociceptive activities, and acute toxicity in mice, as well as the antimicrobial effects of essential oils from the leaves of 7 varieties ofEucalyptusand hybrids againstEscherichia coli,Staphylococcus aureus,andCandida albicans. The extraction of oils was carried out using hydrodistillation, and they were analyzed by gas chromatography coupled to mass spectrometry. Urocam and Grancam were the plants that obtained the highest oil yield, with yields of 3.32 and 2.30%, respectively. The main chemical components identified in these plants were 1.8 cineole andα-pinene. The antinociceptive effect of the 7 oils (50 mg/kg, p.o.) was initially assessed in the acetic acid-induced writhing test. In this assay, a significant (p< 0.05) antinociceptive/anti-inflammatory effect was observed from 4 tested essential oils (E. benthamii, E. saligna,and the hybrids Urocam and Grancam) when compared to the vehicle-treated group. This effect was then confirmed in the formalin-induced paw licking test. No toxicological effects or alterations were observed in motor coordination after the administration of the studied oils to the animals. In the antimicrobial evaluation, the seven essential oils inhibited the growth ofS. aureus, E. coli, andC. albicansat different concentrations. Collectively, these results demonstrate that the essential oil from the leaves and branches ofEucalyptusspecies and varieties present potential biomedical applications and represent a source of antimicrobial and/or anti-inflammatory compounds.