The application of green synthesis in nanotechnology is growing day by day. It's a safe and eco-friendly alternative to conventional methods. The current research aimed to study raw royal jelly's potential in the green synthesis of silver nanoparticles and their antibacterial activity. Royal jelly served as a reducing and oxidizing agent in the green synthesis technology of colloidal silver nanoparticles. The UV-Vis maximum absorption at ~ 430 nm and fluorescence emission peaks at ~ 487 nm confirmed the presence of Ag NPs. Morphology and structural properties of Ag NPs and the effect of ultrasound studies revealed: (i) the formation of polydispersed and spherical particles with different sizes; (ii) size reduction and homogeneity increase by ultrasound treatment. Antibacterial activity of different concentrations of green synthesized Ag NPs has been assessed on Gram-negative S. typhimurium and Gram-positive S. aureus, revealing higher sensitivity on Gram-negative bacteria.