The incorporation of carbohydrate polymers is one of the most efficient strategies to reinforce protein matrices for electrospinning application. In the present work, a basil seed gum (BSG)-reinforced whey protein isolate (WPI) was developed via electrospinning for the co-encapsulation of zinc oxide nanoparticles (ZnONPs) and curcumin (CU). The physicochemical attributes of the nanofiber samples could be controlled by varying the BSG mixing ratio. The Field emission scanning electron microscopy images showed bead-free morphology of WPI/BSG/ZnONPs/CU nanofibers with average fiber diameter of around 362 ± 41 nm. The formation of new H₂ bonds after introduction of BSG and active components was corroborated by Fourier-transform infrared spectroscopy. The nanofibers loaded with ZnONPs/CU displayed improved surface hydrophobicity and high potential for hampering colon cancer cells in vitro. The results proved that the proposed electrospun structures were thermally stable and composed by homogenous nanofibers of high bactericide properties, thus representing promising structures suitable for various biomedical applications.