Researchers are becoming more interested in the eco-friendly bio-synthesis of vanadium oxide nanoparticles because they could be used in many areas of science and technology. The Punica granatum peel extract was used to prepare vanadium oxide nanoparticles (V2O5-NPs) by treating VOSO4.H2O with it. The synthesized particles was characterized using SEM, FTIR, and UV visible spectroscopy. The synthesized particles and extract were then tested for their ability to kill bacteria and fungi by testing them against species like Escherichia coli, Pseudomonas florescens, Bacillus subtilis, Staphylococcus aureus, Pseduomonousaliginoza, Candida albicans, Rhizopus oryzae, Aspergillus Niger, Aspergillus Parasiticus, and Aspergillus flavus. The comparison of antibacterial and antifungal activity against microbes was carried out and it was found from the growth curves of bacteria and fungi that the vanadium oxide nanoparticles showed more zone of inhibition than the Punica granatum peel extract.