Due to their unique physicochemical characteristics, palladium nanoparticles (Pd-NPs) have shown tremendous promise in biological applications. The biosynthesis of Pd-NPs employing Saudi propolis has been designed to be environmental, fast, controlled, and cost-effective. The formation and stability of biosynthesized Pd-NPs by Saudi propolis extract were proved by ultraviolet–visible spectrophotometry (UV-Vis), Fourier-transform infrared spectroscopy (FT-IR), and Zeta potential analysis. Transmission electron microscopy (TEM), scanning electron microscopy (SEM), and X-ray diffraction (XRD) findings show that the average particle size of Pd-NPs is between 3.14 and 4.62 nm, which is in quantum scale. The Saudi propolis enhanced the antimicrobial activity against B. subtilis, S. aureus, E. coli, K. pneumoniae, and C. albicans. Pd-NPs show effective anticancer activity against ductal carcinoma (MCF-7) with IC50 of 104.79 µg/mL.