Chitosan/gelatin‐based nanofibers display excellent biological performance in tissue engineering because of their biocompatible composition and nanofibrous structure with a high surface‐to‐volume ratio mimicking the native extracellular matrix. In this study, to save time and cost of experiments, a response surface methodology based on Box–Behnken design (BBD) is developed to predict the mean diameter of (chitosan:gelatin)/poly(vinyl alcohol) (PVA) nanofibers in three volume ratios of chitosan:gelatin by considering PVA percentage, applied voltage, and flow rate as input variables. The morphology and chemical composition of nanofibers are investigated through scanning electron microscopy (SEM) and Fourier transform infrared spectroscopy (FTIR), respectively. The optimum conditions to yield the minimum diameter of nanofibers with chitosan:gelatin ratios of 25:75, 50:50, and 75:25 are found and result in 165, 121, and 92 nm, respectively, which show good accordance with BBD estimated results. The tensile testing indicates that nanofibers containing higher ratio of chitosan:gelatin result in higher tensile stress and lower toughness and tensile strain. The water contact angle analysis (WCA) shows the appropriate hydrophilicity of crosslinked nanofibers. The MTT assay shows excellent cell viability and cell attachment of nanofibers for mouse fibroblast (L929) cells. The results indicate that optimum nanofibers are potent candidates for wound healing applications.
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