The outstanding biodegradability, biocompatibility, affordability, and renewability of polylactic acid have made it a prominent biomaterial. Herein, an innovative, easy, and eco-friendly technique is used to prepare sodium polylactate (SP)-based nanofibers. Solution blowing spinning (SBS) was used to create fibrous mats of SP and polyvinyl alcohol (PVA). SBS's SP nanfibers were crosslinked using an aqueous solution of calcium chloride to produce moisture-resistant calcium polylactate nanofibrous spun mats. Both of UV-visible absorption spectra and transmission electron microscopy were utilized to study the produced zinc oxide (ZnO) nanoparticles (NPs) to indicate a diameter of around 15-23 nm with a high intensity absorption intensity at 370 nm. New polylactate copolymer was synthesized and characterized by infrared and NMR spectroscopic techniques. In order to prepare SP/PVA/ZnO nanocomposite nanofibers, various ZnO ratios were used. The morphologies of the composite nanofibers were investigated by infrared spectroscopy (FTIR), energy-dispersive X-ray analyzer, and scanning electron microscopy. The cytotoxicity tests of the prepared mat were studied by conducting experiments with L-929 cells at various time intervals. The prepared composite SP/PVA/ZnO nanofibers were subjected to cytotoxicity tests to determine their cytocompatibility. Results showed that those with ZnO concentrations between 0.5% and 2% were found to be less harmful than those with higher concentrations. A variety of bacterial species, including Bacillus pumilus and Staphylococcus aureus, as well as Klebseilla pneumoniae and Escherichia coli, were used to test the antibacterial properties of SP/PVA/ZnO spun mats. The ZnO NPs integrated in the SP/PVA fibrous mats were responsible for their antibacterial properties. After finding the appropriate concentration of ZnO that is least harmful while yet giving a satisfactory antibacterial activity, this biomaterial might be perfect for wound dressing applications. HIGHLIGHTS: New eco-friendly biodegradable sodium polylactate (SP) copolymer was synthesized. Zinc oxide nanoparticles (ZnO NPs) with a diameter of 15-23 nm were prepared. High antibacterial SP/PVA/ZnO fibers were prepared by solution blowing spinning. SP/PVA/ZnO nanofibers (180-220 nm) with various ratios of ZnO were presented. Cytotoxicity results showed that the cell viability decreases with increasing ZnO.