| [Objectives] Using organic solvents in the preparation of silk fibroin (SF) solution limits the further biomedical applications of SF. In this project, a high concentration of SF aqueous solution suitable for electrospun is obtained from natural silk by means of the high molecular weight water soluble polymer dialysis. Then, SF fibrous membrane is fabricated via electrospinning technique after the process parameters are optimized. To screen the appropriate biomimetic mineralization method for SF membrane, inorganic depositions formed on the surface of the membranes through three different systems are explored. This study provides an experimental basis for the construction of SF scaffolds and expands the sources of material for bone replacement in tissue engineering.[Methods] Firstly, ultrafine silk fibroin fibrous membranes are prepared by electrospinning and then process influences on the diameters and morphology of fibers such as spinning solution concentration(20%-40%) and voltage(14kV-22kV) are investigated according to the electrospinning theory and the analysis of scanning electron microscopy (SEM) with the aim to get the optimum process parameter. At the same time, it discusses changes of fibers structure and properties after the75%ethanol vapor treatment. Secondly, SF fibrous membranes are soaked in10×simulated body fluid(10×SBF), supersaturated calcification solution(SCS), alternative soaking solutions(ASS) three different biomimetic solutions for studying their mineralization. All of the obtained deposition is analyzed with Scanning electron microscope-Energy dispersive spectroscopy (SEM-EDX), Fourier Transform infrared spectroscopy (FTIR), and X-ray diffraction (XRD) so as to get an understanding of different process influences on different results of mineralization.[Results](1) By using the high molecular weight water-soluble polymer dialysis and the electrospinning technique comprehensively, SF electrospinning membranes are successfully prepared from the SF solution used water as a solvent in order to maintain the good biocompatibility of SF.(2) Silk fibroin micron fibers can be prepared when the spinning solution concentration is from20%to40%. Fiber diameter represents highly remarkable linear relation to spinning solution concentration, increasing with solution concentration; decreases firstly and then increases with its voltage.(3) The best process parameter for SF is concentration33%, voltage18kV, polar distance15cm, flow of1.0ml/h. The fiber has the narrower diameter distribution and the diameter reaches1.826μm.(4) The structure of fibers is markedly changed to β-sheet conformation after the75%ethanol vapor treatment and they become insoluble in water.(5) Through10x SBF, SCS and ASS three mineral methods, apatite can be formed on the surface of SF membranes, and the deposition amount increases with the extension of immersion. Meanwhile, different methods of mineralization have different effects on mineral crystals morphology. Compared with the other two methods mentioned above, alternative soaking solution can form needle-like mineral crystals with the length of about320nm, smallest, stable shaped, evenly distributed.(6) Deposition obtained by means of ASS is mainly nano-carbonated hydroxyapatite, which is consistent with the hydroxyapatite crystals in bone.(7) ASS method is an effective approach to the mineralization of silk fibroin.[Conclusions] In this work, SF fibrous membranes which have three-dimentional structure and high surface area to volume ratio are successfully prepared from SF aqueous solution by electrospinning. Furthermore, calcium and phosphorus begin deposition, nucleation and growth on SF fibrous membranes surface and become nano-sized bone-like HA eventually. Therefore, the scaffold with both advantages of SF and HA may be a hopeful biomaterial used in bone tissue engineering. |
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