Salmonella enterica, a food- and water-borne pathogen, triggers food poisoning and enteric infections. The effectiveness of antibiotics against Salmonella infections is decreasing due to bacterial resistance. Developing novel antimicrobial agents is crucial and urgent. Here, we screened 550 natural compounds and found that isopropyl paraben (IPPB) effectively inhibited Salmonella enterica serovar Typhimurium (ST) invasion of host cells without impacting bacterial growth, reducing the risk of developing bacterial resistance. Further investigation revealed that IPPB bound to the regulatory protein HilD and inhibited the transcription of the type III protein secretion system 1 (T3SS-1) regulatory genes hilD, hilC and rtsA. This interference blocked the production of T3SS-1 effectors. Importantly, IPPB exerted certain therapeutic effects on both G. mellonella larvae and murine models infected with ST and improved the ST-induced gut microbiota dysbiosis in mice. These findings suggest that IPPB has potential as a novel antimicrobial agent, targeting the T3SS-1 of Salmonella.