Abstract: In this study, the antibacterial activity and mechanism of action of chlorogenic acid against bacteria were assessed. The data from minimum inhibitory concentration (MIC) values showed that chlorogenic acid effectively inhibited the growth of all tested bacterial pathogens, and the MIC values were ranging from 20 to 80 μg/mL. An investigation into action mode of chlorogenic acid against the pathogen indicated that chlorogenic acid significantly increased the outer and plasma membrane permeability, resulting in the loss of the barrier function, even inducing slight leakage of nucleotide. The leakage of cytoplasmic contents was also observed by electron micrographs. These results supported our hypothesis that chlorogenic acid bound to the outer membrane, disrupted the membrane, exhausted the intracellular potential, and released cytoplasm macromolecules, which led to cell death.