The aim of the study was to examine the antibiofilm activity of selected essential oils (EO): Lavandula angustifblia (LEO), Melaleuca alternifolia (TTO), Melissa officinalis (MEO) and some of their major constituents: linalool, linalyl acetate, alpha-terpineol, terpinen-4-ol. Biofilms were formed by Staphylococcus aureus ATCC 29213 and Escherichia coli NCTC 8196 on the surface of medical biomaterials (urinary catheter, infusion tube and surgical mesh). TTC reduction assay was used for the evaluation of mature biofilm eradication from these surfaces. Moreover, time-dependent eradication ofbiofilms preformed in polystyrene 96-well culture microplates was examined and expressed as minimal biofilm eradication concentration (evaluated by MTT reduction assay). TTO, alpha-terpineol and terpinen-4-ol as well as MEO, showed stronger anti-biofilm activity than LEO and linalool or linalyl acetate. Among the biomaterials tested, surgical mesh was the surface most prone to persistent colonization since biofilms formed on it, both by S. aureus and E. coli, were difficult to destroy. The killing rate studies of S. aureus biofilm treated with TTO, LEO, MEO and some of their constituents revealed that partial (50%) destruction of 24-h-old biofilms (MBEC50) was achieved by the concentration 4-8 x MIC after 1 h, whereas 2-4 x MIC was enough to obtain 90% reduction in biomass metabolic activity (MBEC90) after just 4 h of treatment. A similar dose-dependent effect was observed for E. coli biofilm which, however, was more susceptible to the action of phytochemicals than the biofilms of S. aureus. It is noteworthy that an evident decrease in biofilm cells metabolic activity does not always lead to their total destruction and eradication.