Abstract: To prepare a new type of multi-functional medical dressing, polyurethane (PU) nanofiber membranes loaded with different mass fractions of Nigella sativa seed extract (NSS) were prepared by electrospinning method to explore their application potential in wound dressing. The mechanical properties, hydrophilicity, swelling property, air permeability, antioxidant property, antibacterial property and cytotoxicity of the nanofiber membranes were characterized. The experimental results showed that the fiber diameter of PU/NSS nanofiber membranes increased with the increase of NSS mass fractions. The addition of NSS improved the hydrophilicity of PU nanofiber membrane, and the water contact angle decreased from (124.65±0.96)° to (29.90±0.49)°. The flavonoids active substances in NSS made PU/NSS nanofiber membranes have excellent antioxidant and antibacterial properties. The scavenging rate of 2, 2-diphenyl-1-picryl hydrazyl (DPPH) radical could be up to (59.98±0.86)%. The inhibition rates of E.coli and S.aureus were (99.08±0.38)% and (97.30±0.36)%, respectively. The cytotoxicity test results showed that PU/NSS nanofiber membranes had good biocompatibility. The PU/NSS nanofiber membranes prepared in this study are expected to be applied in the field of wound dressing.