The large water holding capacity of Basil Seed (Ocimum basilicum L.) Mucilage (BSM) gives it potential to produce a valuable polymer for water holding applications such as wound dressing. The objective of this research was to prepare a natural-based antibacterial wound dressing from BSM by freeze-drying. Various contents of zinc oxide nanoparticles (ZnO-NP) were incorporated as an antibacterial agent. BSM hydrogel sponge showed considerable porosity and degree of swelling. From FTIR analysis, hydrogen bond and electrostatic interaction between BSM molecules and ZnO-NP were confirmed. SEM images revealed an interconnecting open-cell structure of pores in the BSM hydrogel sponge with a good distribution of ZnO-NP. Moreover, increase in ZnO-NP content improved the mechanical properties (stress at maximum load 8.9 MPa, Young's modulus 151 MPa and strain at maximum load 51%), thermal properties, water retention capacity and antibacterial activity. Cytotoxicity and cell adhesion studies of BSM hydrogel sponge indicated non-cytotoxicity and non-adherent nature of the sponge.