Abstract: Using stems of Senna occidentalis as raw material, four kinds of extracts were obtained through ultrasonic ethanol extraction and subsequent extracted with different polar solvents. The antibacterial activities of these four extracts against Staphylococcus aureus, Candida albicans, and Acinetobacter baumannii were tested using the filter paper diffusion method. It was found that the chloroform extract exhibited the best overall antibacterial effect. Further separation and purification of the chloroform extract was achieved through chromatography to obtain component 1, component 2, and component 3. Mass spectrometry analysis was used to identify the main antibacterial compounds in component 1 with significant antibacterial activity. The results indicated that component 1, which had relatively low polarity, showed significant antibacterial effects. In component 1, 22 compounds were detected, including 7 flavonoids, 13 anthraquinones, 1 polyphenol, and 1 coumarin. Three individual anthraquinone compounds, namely emodin, chrysophanol, and physcion, were selected for determination of the minimum inhibitory concentration(MIC) and minimum bactericidal concentration(MBC). The results showed that the MIC and MBC values of these three compounds against A. baumannii and S. aureus were lower than those of the chloroform extract. Among them, emodin, chrysophanol, and physcion exhibited the strongest inhibition activity against S. aureus, with MIC values of 0.2 g/L and MBC values of 0.39 g/L. Physcion showed the best overall inhibitory effect and might be one of the main compounds responsible for the antibacterial activity in S. occidentalis.