Electrospinning of reconstituted silk fiber from aqueous silk fibroin solution

Silk nanofibers were successfully prepared by electrospinning an aqueous solution of regenerated silk fibroin (RSF, from Bombyx mori) with higher molecular weight. Many factors, such as concentration, electrical conductivity of the fibroin solution and applied electric field were found to influence the morphology of these nanofibers. The conformation of RSF nanofibers was transformed from random coil/helical to β-sheet after the post treatment with pure ethanol. Under optimal conditions, the as-spun non-woven mats achieved good mechanical properties. The apparent stress and strain at break were 11.1 ± 0.7 MPa and 10.2 ± 1.6%, respectively, which is important for the application of such a unique fibrous protein.     

丝素在离子液体中的溶解及再生丝素纤维的结构

以离子液体1-丁基-3-甲基咪唑氯盐（[BMIM]Cl）为溶剂,研究了丝素蛋白在[BMIM]Cl中的溶解情况,结果表明,温度对丝素在[BMIM]Cl中的溶解有较大影响,当温度〈90℃时,即使延长溶解时间,丝素也难以溶解,而当温度≥90℃时,随着温度升高,溶解速率明显加快。考虑到高温会使丝素降解,因此,选择100℃为丝素...     

On the strength of β-sheet crystallites of Bombyx mori silk fibroin

Silk fibroin, a natural multi-domain protein, has attracted great attention due to its superior mechanical properties such as ultra-high strength and stretchability, biocompatibility, as well as its versatile biodegradability and processability. It is mainly composed of β-sheet crystallites and amorphous domains. Although its strength is well known to be controlled by the dissociation of protein chains from β-sheet crystallites, the way that water as the solvent affects its strength and the reason that its theoretically predicted strength is several times higher than experimental measurement remain unclear. We perform all-atom molecular dynamics simulations on a β-sheet crystallite of Bombyx mori silk. We find that water solvent reduces the number and strength of hydrogen bonds between β-chains, and thus greatly weakens the strength of silk fibroin. By dissociating protein chains at different locations from the crystallite, we also find that the pulling strength for the interior chains is several times higher than that for the surface/corner chains, with the former being consistent with the theoretically predicted value, while the latter on par with the experimental value. It is shown that the weakest rupture strength controls the failure strength of silk fibre. Hence, this work sheds light on the role of water in the strength of silk fibroin and also provides clues on the origin of the strength difference between theory and experiment.     

Bio-inspired capillary dry spinning of regenerated silk fibroin aqueous solution

To biomimic the spinning process of silkworm or spider, a capillary spinning equipment was applied to spin regenerated silk fibroin (RSF) fibers from RSF aqueous solutions in air. This equipment exhibits a wide processing window for various RSF aqueous solutions. The effects of pH, metal ions, RSF concentration and spinning parameters on the spinnability of the spinning dope and the mechanical properties of the obtained fibers were investigated. As a result, spinning dopes with a pH from 5.2 to 6.9 have good spinnability, especially for the dope with a pH of 6.0 and a Ca²⁺ concentration of 0.3 M. The RSF concentration of this dope ranges from 44% to 48%. Under optimized conditions of our dry spinning experiments (L/D, 133; take-up speed, 30 mm/s), the obtained as-spun fiber has a breaking strength of 46 MPa, which can be improved up to 359 MPa after a preliminary post-drawing in 80 vol.% ethanol aqueous solution.     

The preparation of insoluble fibroin films induced by degummed fibroin or fibroin microspheres

Blending degummed fibroin (DF) or insoluble fibroin microspheres with concentrated fibroin solution, the insoluble films were obtained through drying the solution at 40–45 ^∘C. The conformation of silk fibroin films was analyzed by infrared spectrum and X-ray diffractometry. The results demonstrated that β-sheet conformation increased rapidly when the degummed silk or insoluble microspheres blended with fibroin, while the pure SF membrane was mainly composed of α/random coil conformation when the other conditions kept same. This suggested that fibroin microspheres and degummed fibroin could induce the formation of β-sheet crystal and the insoluble films, without methanol after-treatment, could be obtained at approximately room temperature. Although the fibroin films blending with DF had many protuberances, the films containing fibroin microspheres had the smooth surface and could be used effectively in biotechnological materials and biomedical application.     

Conformation transition of Bombyx mori silk protein monitored by time-dependent fourier transform infrared (FT-IR) spectroscopy: effect of organic solvent

The conformation transition from random coil and/or helix to β-sheet of silk protein is the most important step in the formation of silk fiber in nature as well as by artificial spinning. Time-dependent Fourier transform infrared (FT-IR) spectroscopy was used in this research to monitor such a conformation transition process induced by the organic solvents methanol, ethanol, propanol, isopropanol, and acetone. The kinetics of β-sheet formation of regenerated Bombyx mori silk fibroin in these organic solvents was obtained by the Δabsorbance-time curve from the time-dependent difference infrared spectra. The results showed that the conformation transition rate of silk fibroin was methanol > ethanol > acetone > propanol > isopropanol, which is in accordance with the polarity of these organic solvents. In connection with the mechanical properties and morphologies of regenerated silk fibers using these organic solvents as coagulation bath reported in the literature, we may conclude that the conformation transition rate of silk protein in the organic solvent is very important in wet-spinning to produce high-performance regenerated silk fibers.     

Influence of the protocol used for fibroin extraction on the mechanical properties and fiber sizes of electrospun silk mats

Silk fibroin (SF) was regenerated using three of the most common protocols described in the bibliography for the dissolution of raw SF (LiBr 9.3 M, CaCl₂ 50 wt.% or CaCl₂:EtOH:H₂O 1:2:8 in molar ratio). The integrity of regenerated SF in aqueous solution was analyzed by SDS-PAGE and different profiles of degradation were observed depending on the protocol used. This fact was found to affect also the aqueous solubility of the freeze dried protein. These different SFs were used to produce electrospun mats using SF solutions of SF 17 wt.% in 1,1,1,1′,1′,1′-hexafluoro-2-propanol (HFIP) and significant differences in fiber sizes, elongation and ultimate strength values were found. This work provides a global overview of the manner that different methods of SF extraction can affect the properties of electrospun SF-mats and consequently it should be considered depending on the use they are going to be made for.     

In vitro and in vivo research on using Antheraea pernyi silk fibroin as tissue engineering tendon scaffolds

In this paper, the feasibility of using Antheraea pernyi silk fibroin as tissue engineering tendon scaffold was investigated in vitro and in vivo, respectively, utilizing tenocytes and animal model. The animal model used here was an adult New Zealand White rabbit with a 15-mm gap defect in both sides of the Achilles tendon. The Achilles tendon defects in one side of hind legs were repaired using the braided A. pernyi silk fibroin scaffold in experimental group (n = 24), while the other side left untreated as negative group (n = 24). The recovery of the defect tendons were evaluated postoperatively at the 2nd, 6th, 12th, and 16th week using macroscopic, histological, immunohistochemical, scanning electron micrograph and biomechanical test techniques. In vitro results examined by scanning electron micrograph showed that A. pernyi silk fibroin promote the adhesion and propagation of the tenocytes. In vivo, at 16 weeks after implantation, morphological results showed that neo-tendons were formed, and bundles of collagen fibers in the neo-tendons were uniform and well oriented. Immunohistochemical results showed that collagen type in the regenerated tendons was predominantly type I. The maximum load of regenerated tendon at 16 weeks reached 55.46% of the normal tendon values. Preliminary, we concluded that A. pernyi silk fibroin promoted the recovery of Achilles tendon defect of rabbit and the application of A. pernyi silk fibroin as tissue engineering tendon scaffold is feasible.     

甲醇处理对再生丝素蛋白干法纺丝纤维结构及性能的影响

采用甲醉水溶液对初生的再生丝素蛋白干法纺丝纤维进行了后处理.研究表明,甲醉处理对再生丝的表面形态、结晶结构及机械性能有影响.甲醇处理前,初生丝蛋白纤维透明而有光泽;甲醇处理后,丝纤维呈现白色,光泽度下降,同时沿纤维轴方向出现了细小的条纹,并可观察到明显的双折射现象.甲醉处理还可使初生丝蛋白纤维中存在的无规卷曲或Silk...     

Facile fabrication of the porous three-dimensional regenerated silk fibroin scaffolds

In the present work, we report a new facile method to fabricate porous three-dimensional regenerated silk fibroin (RSF) scaffolds through n-butanol- and freezing-induced conformation transition and phase separation. The effects of RSF concentration, freezing temperature and n-butanol addition on the microstructure, the secondary structures of silk fibroin and apparent mechanical properties of the RSF scaffolds were investigated by SEM, ¹³C CP-MAS NMR spectra and mechanical testing, respectively. By adjusting the RSF concentration and n-butanol addition, the pore size of the scaffold could be controlled in the range from of 10 μm to 350 μm with 84%–98% of porosity. The tensile strength of the wet scaffold reached the maximum of 755.2 ± 33.6 kPa when the concentration of RSF solution was increased to 15% w/w. Moreover, post-treatment with ethanol further induced conformation transition of RSF from random coil or helix to β-sheet. The porous scaffolds prepared by this facile and energy-saving method with good biocompatibility will have great potential for application in tissue engineering.     

Fabrication and characterization of fibroin solution and nanoparticle from silk fibers of Bombyx mori

The present investigation aims to study the effect of degumming time on the structural property of silk fiber obtained by silk cocoons of Bombyx mori, followed by preparation of the regenerated silk fibroin (RSF) solution which can be subsequently molded into silk nanoparticles. Silk fibers degummed with different media at different time intervals were investigated for the degumming loss and were characterized using Ffourier transform infrared (FTIR), differential scanning calorimetry (DSC), x-ray diffraction (XRD), and scanning electron microscopy (SEM). Maximum degumming was observed when the fibers were treated with sodium carbonate for 60 min. SEM and atomic force microscopy (AFM) images of RSF solution showed aggregation of silk globules resulting in formation of solvated macrochains and giving it an appearance of island-like morphology. Blank silk nanoparticles prepared from the RSF solution showed a smooth and spherical surface devoid of any adhesion using SEM, AFM, and transmission electron microscopy (TEM). The prepared silk nanoparticles may further be explored for loading drug entities and targeting.     

Analysis of structure and properties of biodegradable regenerated silk fibroin fibers

In this paper, the molecular weight change of native silk fibroin fibers when they are dissolved in neutral salt solution, and the relationships of structural change of the regenerated SF fibers with their mechanical properties and degradability have been studied. The results shows that the mechanical properties of regenerated SF fibers are lower than those of native SF fibers, but the biodegradability is raised.     

丝素蛋白柔性电子器件材料的研究进展

丝素蛋白材料凭借良好的生物相容性、可控生物降解性、再生形貌多样性等已被制成柔性电子器件在电子领域进行了应用研究.本文首先综述不同溶解方法对蚕丝再生材料制备的影响,同时对丝素蛋白材料的(微球、膜、纤维、凝胶、支架等)制备方法、材料性能进行分析,最后总结了近年来丝素蛋白基柔性电子材料的应用研究进展.尽管已有研究表明可获得各...     

Silver nanoparticles incorporated electrospun silk fibers

We present a simple and mass-producible method of incorporating silver nanoparticles on the surface of electrospun silk non-woven membranes for the fabrication of antimicrobial wound dressings. Nanofibrous silk membranes with fiber diameters of 460 +/- 40 nm were electrospun from an aqueous Bombyx mori fibroin solution. The electrospun membranes incorporating silver nanoparticles were prepared by dipping the membranes in aqueous silver nitrate (AgNO3) solution (0.5 or 1.0 wt%) followed by photoreduction. Field emission scanning and transmission electron microscopy showed that silver nanoparticles were generated on the electrospun silk fibroin nanofibers as well as inside them. The interaction between the silver nanoparticles and amide groups in the silk fibroin molecules was characterized using X-ray photoelectron spectroscopy.     

Surface properties and cytocompatibillity of silk fibroin films cast from aqueous solutions in different concentrations

Silk fibroin film (SFF) has been widely used in biomaterials. SFF is usually prepared from a regenerated silk aqueous solution and its properties depend remarkably on the preparation conditions. However, the effect of the silk fibroin concentration (C ₀) on the SFF surface properties as well as the cytocompatibility has rarely been investigated. In this work we prepared a series of Bombyx mori SFFs by casting SF aqueous solutions with the concentration from 10° to 10² mg/mL on TCPS substrate at 60°C. The test results of atomic force microscopy, attenuated total reflection Fourier transform infrared and contact angles analysis showed that the film surface roughness and β-sheet structure increased with the increase of C ₀, whereas the surface hydrophilicity increased with the decrease of C ₀. The in vitro clotting time measurement results revealed that the SFFs prepared from the thinner solution showed a longer APTT (activated partial thromboplastin time) and TT (thrombin time). The results of microscopy and MTT assay also revealed that cell adhesion and growth were enhanced on the SFF cast from lower C ₀ for fibroblasts. In contrast, endothelial cells showed a similar behavior on all those films that were prepared from the solution in different concentrations.     

A novel electrospun silk fibroin/hydroxyapatite hybrid nanofibers

A novel electrospinning of silk fibroin/hydroxyapatite hybrid nanofibers with different composition ratios was performed with methanoic acid as a spinning solvent. The silk fibroin/hydroxyapatite hybrids containing up to 30% hydroxyapatite nanoparticles could be electrospun into the continuous fibrous structure. The electrospun silk fibroin/hydroxyapatite hybrid nanofibers showed bigger diameter and wider diameter distribution than pure silk fibroin nanofibers, and the average diameter gradually increased from 95 to 582 nm. At the same time, the secondary structure of silk fibroin/hydroxyapatite nanofibers was characterized by X-ray diffraction, Fourier transform infrared analysis, and DSC measurement. Comparing with the pure silk fibroin nanofibers, the crystal structure of silk fibroin was mainly amorphous structure in the hybrid nanofibers. X-ray diffraction results demonstrated the hydroxyapatite crystalline nature remained as evidenced from the diffraction planes (002), (211), (300), and (202) of the hydroxyapatite crystallites, which was also confirmed by Fourier transform infrared analysis. The thermal behavior of hybrid nanofibers exhibited the endothermic peak of moisture evaporation ranging from 86 to 113 °C, and the degradation peak at 286 °C appeared. The SF/HAp nanofibers mats containing 30% HAp nanoparticles showed higher breaking tenacity and extension at break for 1.1688 ± 0.0398 MPa and 6.55 ± 1.95%, respectively. Therefore, the electrospun silk fibroin/hydroxyapatite hybrid nanofibers should be provided potentially useful options for the fabrication of biomaterial scaffolds for bone tissue engineering.     

Studies on spinning and rheological behaviors of regenerated silk fibroin/<Emphasis Type="Italic">N-</Emphasis>methylmorpholine-<Emphasis Type="Italic">N-</Emphasis>oxide·H<Subscript>2</Subscript>O solutions

In this paper, N-Methylmorpholine-N-Oxide (NMMO) was used as solvent for regenerated silk fibroin (RSF). Concentrated solutions with RSF content from 10 to 25 wt% were obtained by dissolving RSF film into NMMO monohydrate. The spinning and rheological properties of RSF/NMMO·H₂O solutions were studied by a simple piston type single filament spinning apparatus and HAAKE RS150L rheometer, respectively. The results are discussed to understand the influences of shear rate, temperature and RSF concentration on the rheological behaviors of RSF/NMMO·H₂O and to choose appropriate process condition for spinning. Spinning was performed using a dry-wet spinning process. The regenerated silk fibre was prepared successfully and the tenacity can reach to 3.07cN/dtex.     

Study on the structure of SF fiber mats electrospun with HFIP and FA and cells behavior

Bombyx mori silk fibroin (SF) fiber mats were prepared by electrospinning with the solvent of hexafluoroisopropanol (HFIP) and formic acid (FA). The average diameters of SF fiber mats observed by SEM were 2.0 and 0.3 μm when different solvent, HFIP and FA, were used. Fourier transform infrared and X-ray diffraction were employed to study the secondary structure of the SF fiber mats; the results showed that the electrospin solvent not only affect the secondary structure of as-spun SF fiber mats, but also indirectly affect the structure transition of SF fiber mats post-treatment with ethanol. And the SF fiber mats electrospun with FA showed more β-sheet structure before and after ethanol treatment. The differential thermal analysis curve indicated that the solvent of HFIP or FA had a weak effect on the thermal properties of SF fiber mats. To assay the cytocompatibility and cell behavior on the SF fiber mats, cell attachment, spreading, and proliferation of normal human epidermal fibroblasts (NHEF) seeded on the scaffolds was studied. The results indicated that the SF fiber mats support NHEF attachment and growth on SF fiber mats in vitro, and no difference between the SF fiber mats electrospun with HFIP and FA was observed. In this article, a desired morphology and secondary structure of SF fiber mats could be prepared by choosing different electrospinning solvent.     

静电纺再生丝素纳米纤维:纤维直径分布与力学性能

采用静电纺丝的方法研制了再生丝素纳米纤维(ERSF)膜,纤维直径为50～1000nm.将脱胶后的桑蚕丝溶解在摩尔比为1:2:8 的60℃CaCl2/CH3CH2OH/H2O三元体系中,将该溶液冷冻干燥后溶解在98%的甲酸中得到再生丝素溶液,对其进行静电纺丝.研究了不同纺丝条件下,静电纺再生丝素纤维的直径分布.研究发现:在一定的电压和喷丝头与接收屏的距离(C-D)下,7wt%是具有良好可纺性的临界浓度.纤维的直径随着溶液浓度的增加而增大,随着C-D的增加而减小,并且在C-D较大时可以获得较均匀的纤维.电压是另一个影响纤维直径的重要因素,当电压高于某一数值时,可以纺得细而均匀的纳米级再生丝素纤维.在9wt%,12cm C-D and 15KV 的纺丝条件下,80%的纤维直径在50～150nm之间.由于所纺得的再生丝素纤维膜在水中会产生收缩,因此用甲醇和丙酮对其进行处理.力学性能是影响纤维膜实际使用的重要性能,我们测定和分析了静电纺再生丝素纤维膜处理前后的力学性能.     

化学交联改性丝素蛋白/海藻酸钠纤维的制备与性能

利用聚乙二醇二缩水甘油醚(PEGDE)化学交联丝素蛋白(SF)、以CaCl_2交联海藻酸钠(SA),采用分步交联的方法制备了SF/SA双网络型复合纤维,通过DV-C型数显黏度计、FTIR、XRD、电子单纤维强力仪、核磁共振变温弛豫分析仪、SEM等对纺丝溶液表观黏度和交联改性SF/SA复合纤维的结构性能进行了表征,研究了交联剂PEGDE添加量对纺丝溶液的表观黏度和交联改性SF/SA复合纤维的力学性能、交联度、微观形貌、吸湿性能和保湿性能等的影响。结果表明,随着PEGDE含量的增加,纺丝溶液的表观黏度呈先下降后增加的趋势,加入适量的交联剂可以降低纺丝溶液的表观黏度,增加纤维的可纺性;纤维的断裂强度、断裂伸长率和交联度呈先增加后减少的趋势,PEGDE与SF的质量比为3∶1时,交联改性SF/SA复合纤维的断裂强度和交联度达到最大值,分别为2.34cN/dTex和55.38%;交联改性SF/SA复合纤维的交联度越大,纤维表面沟槽结构越密集,纤维内部结构越紧密均匀,SF与SA的相容性越好;交联改性后,SF/SA复合纤维吸湿性能和保湿性能得到提高,吸湿平衡时间提前。