We assessed the quorum sensing (QS) inhibitory impact of sesamol against the foodborne bacterium Pseudomonas aeruginosa. At concentrations ranging from 50 to 200 μg/mL, sesamol significantly inhibited the production of virulence factors such as protease, elastase, pyocyanin, rhamnolipid, and chemotaxis, and improved the susceptibility of bacterial and biofilm cells to colistin. Integrated transcriptomics, metabolomics, and docking analyses indicated that exposure to sesamol destroyed the QS system and down-regulated the expressions of genes encoding virulence and antioxidant enzymes. The down-regulation of genes encoding antioxidant enzymes intensified oxidative stress, as demonstrated by the enhancement of reactive oxygen species and H2O2. The enhanced oxidative stress changed the components of the cell membrane, improved its permeability, and ultimately enhanced the susceptibility of bacterial and biofilm cells to colistin. Moreover, exposure to sesamol also led to the disorder of amino acid metabolism and energy metabolism, eventually attenuating the pathogenicity of P. aeruginosa. These findings indicated that sesamol can function as a potent anti-virulence agent to defend against food spoilage caused by P. aeruginosa.