Silver nanoparticles (AgNPs) prepared by plants are simple, eco-friendly, and economical. In this study, Magnolia officinalis (MO) extract was applied to synthesize MO@AgNPs. Ultraviolet-visible (UV-vis) spectrum analysis indicated a peak at 440 nm. Most of the particles were spherical with sizes from 1 to approximately 60 nm based on transmission electron microscopy (TEM). X-ray diffraction (XRD) patterns showed a face-centered cubic crystal structure. The zeta value of MO@AgNPs was - 36.5 ± 0.6 mV, which was stable at 25 °C and 4 °C. Growth kinetic studies and the Kirby-Bauer diffusion method showed significant inhibitory activity on Candida albicans (ATCC 10231), Escherichia coli (ATCC BAA-2340), and Staphylococcus aureus (ATCC 25923); the minimum inhibitory concentrations (MIC) were 3, 9, and 9 μg/mL, and corresponding minimum bactericidal concentrations (MBC) were 5, 11, and 9 μg/mL, respectively. MO@AgNPs exhibited better antifungal activity compared to AgNPs prepared using sodium citrate. Further research revealed that MO@AgNPs increased the permeability of bacterial cell membranes. Moreover, the effect of MO@AgNPs on Candida albicans was significantly enhanced by blocking autophagy. The reactive oxygen species (ROS) induced by MO@AgNPs in Candida albicans was limited and may be related to its good antioxidant activity. Finally, MO@AgNPs have no significant cytotoxicity to the human liver LO2 cell line under 20 μg/mL.