There is a growing interest of industry to replace synthetic chemicals by natural products with bioactive properties from plant origin. The aim of this study was to determine the chemical composition of European pennyroyal (Mentha pulegium) essential oil and to characterize the in vitro antioxidant and antimicrobial activities of its water (hot and cold) and ethanolic extracts and of the essential oil. The essential oil revealed menthone, pulegone and neo-menthol as the main constituents, comprising 35.9, 23.2 and 9.2% of the essential oil, respectively. The hot water extract exhibited the highest antioxidant activity and phenol content. In contrast, the extracts were not very effective to inhibit the growth of the seven foodborne spoilage and pathogenic bacteria tested, but the essential oil showed antibacterial activity against all bacterial strains. In conclusion, extracts and essential oil of M. pulegium from Mediterranean origin have huge potential as an alternative to chemical additives for the food industry.