In the present study, we have successfully synthesized manganese dioxide nanoparticles (MDONPs) using the leaves extract of Datura stramonium as reducing agents. The synthetic biological method was found highly efficient, low cost and eco-friendly. The formation of MDONPs was confirmed by visual colour change of solutions and UV-Visible spectroscopy. Biologically synthesized MDONPs have been characterized by using different analytical methods such as UV-Visible, Fourier transform infrared spectroscopy (FTIR), Powder X-ray diffraction (XRD), Field emission scanning electron microscopy (FESEM) and Energy-dispersive X-ray spectroscopy (EDS); all these techniques indicated the efficient formation of MDONPs. The antimicrobial activity of MDONPs was checked by using well diffusion method against Staphylococcus aureus, Proteus vulgaris, Salmonella typhi, Streptococcus mutans and Escherichia coli and MDONPs found powerful antimicrobial agents.