Aims: To measure antibacterial activity of the semi-synthetic flavonoid 3-O-octanoyl-(-)-epicatechin and investigate the mechanism of action. Methods and Results: MICs determined by the broth microdilution method were 50 mu g ml(-1) for beta-lactam sensitive and resistant Staphylococcus aureus, and 100 mu g ml(-1) for vancomycin sensitive and resistant enterococci. In time-kill studies, 100 mu g ml(-1) 3-O-octanoyl-(-)-epicatechin reduced colony forming unit numbers of antibiotic sensitive and methicillin-resistant Staph. aureus below detectable levels within 120 min. Bacterial aggregation was not observed when cells exposed to 3-O-octanoyl-(-)-epicatechin were examined by light microscopy. It was also shown that 50 mu g ml(-1) 3-O-octanoyl-(-)-epicatechin is capable of reducing colony forming unit numbers of high cell density Staph. aureus populations by 80-fold within 60 min incubation, and inducing leakage of 50% of their internal potassium within just 10 min. Conclusions: 3-O-Octanoyl-(-)-epicatechin is active against Gram-positive bacteria, has bactericidal activity against both antibiotic sensitive and resistant strains, and is likely to exert its primary antibacterial effect by damaging the cytoplasmic membrane. Significance and Impact of the Study: 3-O-Octanoyl-(-)-epicatechin has significant antibacterial activity and additional structural modification and/or formulation studies may allow this to be potentiated.