This study aimed to synthesize palladium nanoparticles (PdNPs) using bioactive compounds from aqueous extracts of Quercus species (Quercus dalechampii, Quercus frainetto, and Quercus petraea) with potential biomedical applications. To optimize PdNPs biosynthesis, various parameters were explored, including the concentration of PdCl2, the extract-to-PdCl2 ratio, and the pH of the solution. The nanoparticles were characterized using ultraviolet/visible spectroscopy (UV/Vis), Fourier-transform infrared spectroscopy (FTIR), and dynamic light scattering (DLS). Total polyphenol content was measured using the Folin–Ciocâlteu method, while antioxidant capacity was evaluated through radical neutralization assays, including ABTS and DPPH, and through iron and copper reduction tests. Antimicrobial activity was tested against Gram-positive and Gram-negative bacteria, as well as Candida species. Phenolic compounds and flavonoids from the extracts were essential for the reduction in palladium ions and the stabilization of the nanoparticles. UV/Vis spectroscopy showed a distinct surface plasmon resonance peak, indicating the successful formation of PdNPs. FTIR analysis confirmed the interaction between the bioactive compounds and PdNPs, revealing characteristic peaks of phenolic groups. DLS analysis indicated a hydrodynamic diameter of 63.9 nm for QD-PdNPs, 48 nm for QF-PdNPs, and 63.1 nm for QP-PdNPs, highlighting good dispersion and stability in solution. Although the PdNPs did not exhibit strong antioxidant properties, they demonstrated selective antimicrobial activity, especially against Staphylococcus aureus and methicillin-resistant Staphylococcus aureus (MRSA). PdNPs also exhibited significant antifungal activity against Candida krusei, with a minimum inhibitory concentration (MIC) of 0.63 mg/mL, indicating their ability to compromise fungal cell integrity. This study contributes to developing eco-friendly biosynthesis methods for metallic nanoparticles and underscores the potential of PdNPs in various applications, including in the biomedical field.