Currently, the evolution of green chemistry in the synthesis of nanoparticles (NPs) with the usage of plants has captivated a great response. In this study, in vitro plantlets and callus of Silybum marianum were exploited as a stabilising agent for the synthesis of zinc oxide (ZnO) NPs using zinc acetate and sodium hydroxide as a substitute for chemical method. The contemporary investigation defines the synthesis of ZnO NPs prepared by chemical and bio-extract-assisted methods. Characterisation techniques such as X-ray diffraction, scanning electron microscopy, Fourier transform infrared spectroscopy and energy dispersive X-ray were used to confirm the synthesis. Although chemical and bio-assisted methods are suitable choices for NPs synthesis, the bio-assisted green assembly is advantageous due to superior stability. Moreover, this report describes the antibacterial activity of the synthesised NPs against standard strains of Klebsiella pneumonia and Bacillus subtilis.