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A combination process of using pure water for electrospinning and electron beam (EB)-assisted cross-linking is proposed as a dual green route to obtain strengthened silk fibroin (SF)-based nonwoven nanofibers toward value-added healthcare or biomedical products. The first challenge is to optimize the electrospinning process using SF/poly(ethylene oxide) (PEO) in a pure water solution for the fabrication of nonwoven nanofibers. Another technological challenge is to stabilize the dissoluble SF/ PEO nonwoven nanofibers through a cross-linking network created by a free radical reaction using EB irradiation. Herein, we optimized SF/PEO combination and the electrospinning parameters for the fabrication of SF-PEO nanofibers. Under mild conditions using an aqueous solution of SF-PEO and tunable electrospinning parameters, elongated, cylindrical-shaped, and smooth SF-PEO nanofibers with a diameter of 169 ± 5 nm were obtained. The stability of the SF-PEO nanofibers could be tuned by EB irradiation with the absorbed doses in the range of 10−100 kGy. Aiming at well-defined characteristics of nanofibers, the morphology, gel fraction, kinetics of swelling behavior, chemical and packing structure, thermal and mechanical properties, cell cytotoxicity, and proliferation were characterized and relatively clarified with the influencing parameters. It is important to note that EB irradiation with a reliable sterilization dose of 25 kGy was an optimum dose for cross-linking. Upon cross-linking the SF-PEO nanofibers, the dissolution of the nanofibers reduced, they became more stable, and their mechanical properties significantly improved with an increase in ductility by ∼22-fold. The cross-linked SF-PEO nanofibers promoted cell proliferation and proved nontoxic to skin fibroblast cells (L929), and they are thus appropriate for healthcare and biomedical applications. This study shows the potential of using 3G approaches, i.e., green renewable polymers, green electrospinning, and a green EB irradiation process, for sustainable chemistry for biomedical engineering applications.

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