Immobilization of glucose oxidase in the regenerated silk fibroin membrane: Characterization of the membrane structure and its application to an amperometric glucose sensor employing ferrocene as electron shuttle

The structural changes of the regenerated silk fibroin membranes for immobilizing glucose oxidase were characterized with FT-IR spectra. The results of electronic absorption spectra and scanning electron microscopy illustrated that the glucose oxidase in the membranes existed in aggregates. The blend membranes exhibited an island-like structure, attributable to the incompatible and weak intermolecular interactions between regenerated silk fibroin and glucose oxidase. The glucose sensor was constructed by combining glucose oxidase immobilized in regenerated silk fibroin membrane with an Eastman-AQ-ferrocene modified electrode. The advantages of the mediator include high stability and the ability to use a less positive potential for increased selectivity.     

Immobilization of glucose oxidase on nonwoven fabrics with bombyx mori silk fibroin gel

The enzyme glucose oxidase (GOD) was immobilized on the nonwoven fabrics, which have excellent properties in diffusivity of substrates, mechanical strength, and handling, with Bombyx mori silk fibroin gel. The nonwoven fabrics of silk fibroin, viscose rayon, poly-ethyleneterephthalate, 6-nylon, and polypropylene with activated surface by fluoline treatment were used. The stabilities of GOD to heat or pH changes were much improved by the immobilization as well as the case of the GOD immobilized in the silk fibroin membrane. Among nonwoven fabrics, silk fibroin was the most excellent support material for the immobilization of GOD although all nonwoven fabrics used here are able to be used as the support materials. The increase of the sensitivity was observed when the glucose sensor was made with the GOD immobilized on nonwoven silk fabrics as four times compared with the case of the GOD immobilized in the silk fibroin membrane.     

Blend membrane of regenerated silk fibroin,poly(vinyl alcohol), and peroxidase and its application to a ferrocene-mediating hydrogen peroxide sensor

Before or after the blend membrane of regenerated silk fibroin (RSF), poly(vinyl alcohol) (PVA), and peroxidase is treated with ethanol, RSF, PVA, and peroxidase maintain their own structures. The conformational transition of RSF in the blend membrane is accomplished from the silk I structure to the silk II structure by ethanol treatment, which is used to immobilize peroxidase. A ferrocene-mediating sensor for H₂O₂ is made, which is based on the immobilization of peroxidase in the blend membrane of RSF and PVA. Performance and characteristics of the sensor were evaluated with respect to response time, detection limit, selectivity, and dependencies on temperature and pH as well as on operating and storage stability. © 1996 John Wiley & Sons, Inc.     

聚乳酸/丝素共混膜的制备及其性能

用分子量为10万的聚乳酸（PLLA）对丝素膜进行改性,研究不同的聚乳酸加入量对丝素膜性能的影响,对聚乳酸/丝素共混膜进行了一系列表征。万能电子试验机的测试结果表明,经聚乳酸改性后,丝素膜的断裂强度,断裂伸长率有了较大的改善,当加入聚乳酸占丝素质量为5%时,丝素膜的强度可达到27.1 MPa,伸长率达4.4%; 改性后的丝素膜的亲水性有一定程度降低,溶失率则明显减小,透汽透湿性也有所提高;红外光谱测试表明,改性后的丝素膜含有较多的β构象成分。     

再生丝素蛋白/羟基磷灰石多孔复合材料的制备

再生丝素蛋白具有良好的生物相容性,羟基磷灰石同时还具有成骨诱导性。通过将再生丝素蛋白制备形成丝素蛋白多孔材料,并在37℃下将其浸渍于模拟体液中可以制备再生丝素蛋白/羟基磷灰石多孔复合材料。扫描电镜研究发现在再生丝素蛋白多孔材料的孔隙中羟基磷灰石由针状晶体聚集而成,红外光谱和XRD等表征表明复合材料中羟基磷灰石以羰基取代的羟基磷灰石存在。制备的再生丝素蛋白/羟基磷灰石多孔材料有望作为骨组织修复材料使用。     

Silk fibroin/cellulose blend films: Preparation,structure, and physical properties

This article deals with the preparation and characterization of silk fibroin (Bombyx mori)/cellulose blend films. Following dissolution with a metal complex solution, the average molecular weight of silk fibroin slightly decreased. While cellulose was almost unaffected. After coagulation and washing, transparent films were obtained by blending fibroin and cellulose in all proportions. The crystalline structures of regenerated fibroin and cellulose were β-form and cellulose II, respectively, as shown by the characteristic x-ray diffraction profiles. Density values increased with cellulose content, though less than expected from a pure additive behavior. Moisture regain increased following the addition of a small amount of cellulose to silk fibroin. The mechanical properties showed that both strength and elongation at break of silk fibroin films were improved by blending with cellulose. IR spectra exhibited changes in the skeletal frequences of silk fibroin, suggesting the occurrence of intermolecular interactions between fibroin and cellulose through hydrogen bond formation. © 1995 John Wiley & Sons, Inc.     

Phase separation in poly(vinyl alcohol)/gelatin blend systems

Blends of a commercial poly(vinyl alcohol) (a-PVA) derived from vinyl acetate and gelatin obtained from collagen were prepared by mixing aqueous solutions of both samples under various conditions. For the blend hydrogels, the depression of melting temperatures was observed at high PVA content. Similarly to the a-PVA/silk fibroin (SF) system, when the degrees of polymerization of PVA increased, the microphase separation region in a phase diagram of the blend films increased for the a-PVA/gelatin system. In the IR spectra of the blend films, no absorption bands corresponding to a conformational change of gelatin appeared. Comparison of the mechanical properties of the a-PVA/gelatin and a-PVA/SF systems, showed the interaction between PVA and gelatin molecules to be smaller than that between PVA and SF molecules. © 1999 Society of Chemical Industry     

Phase separation structure in poly(vinyl alcohol)/silk fibroin blend films

Blend films of a commercial poly(vinyl alcohol) (a-PVA) derived from vinyl acetate and silk fibroin (SF) obtained from degummed silk were prepared by mixing the aqueous solutions of both samples. A plain weave structure was recognized only in the blend films, whereas no structure was found for the superimposed films of both samples. The phase separation structure of the blend films was examined by microscopic observations elongation, tensile tests, and IR measurements. The microphase separation region increased with increase in the degree of polymerization of the PVA. In the IR spectra of the blend films with high PVA contents cast under certain conditions, the absorption peak attributed to the cross-β-form conformation of SF appeared strongly. Gelatin, a water-soluble and natural polymer, was also used for comparison with SF. The ternary phase diagram in an a-PVA/gelatin/H₂O system was obtained experimentally and the critical point was used to estimate the interaction parameter between PVA and gelatin molecules. The phase separation structure and the interaction between PVA and SF molecules were also discussed taking into consideration the results of the a-PVA/gelatin system. © 1998 SCI.     

Preparation of glucose oxidase membrane for the application of glucose sensor by plasma activation

Plasma activation is a method that takes the advantage of low temperature plasma to immobilize the bioactive materials. An active immobilized glucose oxidase membrane was obtained via plasma-initiated polymerization of acrylic acid. The obtained immobilized enzyme is very active and stable. After 100 tests, the response error of the glucose sensor is less than 5% and its linear detecting range is 0–300 ppm. A hydrophilic PP film treated by Ar or NH₃ plasma was used for the preparation of the immobilized glucose oxidase membrane. The obtained immobilized enzyme is also very active and stable. After 180 tests, the response error of the glucose sensor is less than 4% and its linear detecting range is 0–300 ppm. Furthermore, SEM was used to study the mophology of the glucose oxidase membranes obtained via both methods. And ESCA was used to analyze the plasm-initiated polymerization products and the plasma-treated PP films so as to obtain the optimum conditions for the immobilization of bioactive materials.     

Preparation of non-woven mats from all-aqueous silk fibroin solution with electrospinning method

In the present study, we successfully prepared non-woven mats from stable regenerated silk fibroin aqueous solution at high concentration. Scanning electronic microscope (SEM) was used to observe the morphology of the fibers. The structure of the fibers was characterized using Fourier transform infrared (FTIR), wide-angle X-ray diffraction (WAXD) and differential scanning calorimetry (DSC). The mechanical tests were also performed. In the as-spun fibers, silk fibroin was present in a random coil conformation, the stress and strain at break were 0.82 MPa and 0.76%, respectively, while after methanol treatment, the silk fibroin was transformed into a β-sheet-containing structure, the stress and strain at break increased to 1.49 MPa and 1.63%, respectively. This study provided an option for the electrospinning of silk fibroin without using organic solvent or blending with any other polymers, which may be important in tissue engineering scaffold preparation.     

丝素/羧甲基壳聚糖共混膜的结构性能探讨

将含有甘油和戊二醛的丝素与羧甲基壳聚糖按一定比例混合,制得丝素/羧甲基壳聚糖共混膜,对共混膜的结构与性能进行了探讨。结果表明:随着羧甲基壳聚糖含量的增加,共混膜的透气率增大,加入交联剂戊二醛有效地改善了共混膜的力学性能,但其透气率有所降低;当丝素与羧甲基壳聚糖的质量比为4/1时,共混膜的断裂强度最大,力学性能较好,共混膜相容性较好,其断面光滑、致密。制备丝素/羧甲基壳聚糖共混膜的较佳条件为:丝素中的甘油质量分数为15%,戊二醛质量分数为0.075%,丝素与羧甲基壳聚糖质量比为4/1。     

Structural changes and their effect on mechanical properties of silk fibroin/chitosan blends

Silk fibroin/chitosan blend films were prepared by the solvent casting method. Miscibility between silk fibroin and chitosan was examined by dynamic mechanical thermal analysis. Structural changes of silk fibroin by the addition of chitosan were investigated by IR spectroscopy. The conformational transition of silk fibroin from random coil form to β‐sheet structure induced by blending with chitosan resulted in the increase of crystallinity and density of the blend films. The blend film containing 30 wt % chitosan exhibited a maximum increase in crystallinity and density. It was found that the tensile strength and initial tensile modulus of blend films were greatly enhanced with increasing the chitosan content and showed a maximum value at the composition of 30 wt % chitosan. © 1999 John Wiley & Sons, Inc. J Appl Polym Sci 74: 2571–2575, 1999     

Characterization of composite membrane of poly(vinyl alcohol) and regenerated silk fibroin for immobilization of horseradish peroxidase and an amperometric neckelocene-mediated sensor sensitive to hydrogen peroxide

Horseradish peroxidase (HRP) was effectively entrapped in a novel composite membrane of poly(vinyl alcohol) and regenerated silk fibroin. IR spectroscopy was employed to characterize the structure of the composite membrane and scanning electron microscopy was used to visualize the morphology of the composite membrane. HRP was utilized to amplify the amperometric response of the sensor by catalyzing reduction of hydrogen peroxide via nickelocene, an electron transfer mediator between immobilized HRP and a glassy carbon electrode. The influence of various experimental parameters such as pH, temperature and applied potential were explored for optimum analytical performance. The sensor possessed a variety of characteristics including good sensitivity, rapid response time and a low detection of limit of 0·1 μmol dm⁻³, which arose from the efficiency of electron transfer between immobilized HRP and the electrode via nickelocene.     

Structural and physical properties of silk fibroin/alginate blend sponges

Silk fibroin/alginate blend sponges were examined through IR spectroscopy, X‐ray diffractometry, and differential scanning calorimetry to determine the structural changes of silk fibroin. The effects of fibroin/alginate blend ratios on the physical and mechanical properties were investigated to discover the feasibility of using these blend sponges as biomedical materials such as wound dressings. The compressive modulus of silk fibroin was increased up to 30 kPa, from 7.1 kPa, by blending with alginate. Thermal crystallization behavior of fibroin induced by heat treatment was restricted by blending with alginate. In spite of that, the structural characteristics of fibroin were not changed by incorporation with alginate. © 2004 Wiley Periodicals, Inc. J Appl Polym Sci 93: 2174–2179, 2004     

Separation of alcohol–water mixture by pervaporation through a novel natural polymer blend membrane–chitosan/silk fibroin blend membrane

A novel natural polymer blend membrane, namely chitosan/silk fibroin blend membrane, was prepared. The selective solubility and the pervaporation properties of alcohol–water mixture were studied. The results showed that the membrane was water selective and the separation factor of ethanol–water mixture could be improved compared to pure chitosan membrane, when silk fibroin content in blend membrane was no more than 40 wt %. The blend membrane exhibited a best performance, (i.e., the water in permeate was large than 99 wt % when silk content was 20 wt % and the crosslinking agent–glutaraldehyde content was 0.5 mol %). The mechanism of improvement on pervaporation properties was explained by reducing the free volume and freeing hydrophilic groups of chitosan because of the strong intermolecular hydrogen bond forming between chitosan and silk fibroin in blend membrane. In addition, the influence of operation temperature and feed concentration as well as the pervaporation properties of isopropanol–water mixture were also studied. © 1999 John Wiley & Sons, Inc. J Appl Polym Sci 73: 975–980, 1999     

Preparation of regenerated Antheraea yamamai silk fibroin film and controlled‐molecular conformation changes by aqueous ethanol treatment

The methods of dissolving Antheraea yamamai (A. yamamai) silk fibroin fibers and preparing the regenerated silk fibroin were studied. The molecular conformations of A. yamamai silk fibroin film treated by different concentration ethanol solution were investigated by XRD, FTIR, and Raman spectra. The results indicated that A. yamamai silk fibroin fibers could be completely dissolved in dense lithium thiocyanate solution at about 35°C for 1 h. The initial regenerated film consisted of α‐helix and random coil components. The aqueous ethanol treatment of the film resulted in significant increase in β‐sheet component and improvement of water resistance of the film. This effect was strongly dependent on ethanol concentration, and 60–80% (w/w) ethanol was most effective. © 2009 Wiley Periodicals, Inc. J Appl Polym Sci, 2010     

丝素蛋白/聚丙烯酸共混膜仿生合成羟基磷灰石

将丝素蛋白(SF)与聚丙烯酸(PAA)共混,制备丝素蛋白/聚丙烯酸(SF/PAA)共混膜;然后将此共混膜进行改性及矿化处理后,放置于(37±0.5)℃人体仿生液中24h,诱导合成丝素蛋白/羟基磷灰石(SF/HA)复合材料.利用傅里叶红外(FTIR)、X-射线衍射(XRD)、环境扫描电镜(ESEM)以及X射线能谱(EDX...     

用二维FTIR研究纳米TiO2改性丝素蛋白聚集态结构转变

采用溶胶-凝胶方法制备纳米TiO₂复合丝素膜。UV和SEM测试结果表明,该丝素膜中纳米TiO₂均匀分散在丝素中,TiO₂粒径约为80 nm;同时采用一维红外光谱、二维红外相关光谱对纯丝素膜及复合丝素膜结构进行表征。结果表明,随着纳米TiO₂的生成,丝素蛋白中弱氢键缔合的N—H键以及自由的N—H键发生断裂及重组,生成了强氢键;丝素分子从无序状态转变为有序排列,同时无规线团构象及α螺旋构象向β折叠构象发生转变,最后促使丝素蛋白的结晶构象从Silk Ⅰ转变为Silk Ⅱ。     

凝固条件对纤维素/丝素蛋白纤维相形态及性能的影响

蛋白质纤维具有光滑柔顺、透气吸湿等优点,然而天然蛋白纤维产量有限。再生蛋白纤维的制备通常采用与其它成纤高分子接枝或共混的方法,有利于提高再生蛋白纤维的断裂强度。选用同为天然高分子的纤维素为基体,以共溶剂溶解纤维素与蛋白质,进而纺丝成形制得力学性能满足要求的纤维素/丝素蛋白共混纤维。为了探究凝固剂组成对纤维素/丝素蛋白共混纤维相形态及性能的影响,选用水、乙醇、乙醇/1-丁基-3-甲基咪唑氯盐([BMIM]Cl)等作为凝固剂。研究发现:乙醇作为凝固剂时,纤维素与丝素蛋白能很好地同时凝固;而当在乙醇凝固浴中加入适量的[BMIM]Cl径向均匀分散。通过对凝固剂组成的调控能有效提升纤维的力学强度。     

Effects of superfine silk protein powders on mechanical properties of wet‐spun polyurethane fibers

Mechanical measurements were employed to investigate the effects of three types of superfine silk protein powder on tensile strength, elongation, and elasticity of wet‐spun Pellethane^® 2363‐80AE polyurethane (PU) fiber. These superfine silk protein powders included undegummed silk (with both native silk fibroin and sericin, water insoluble), native silk fibroin (with native silk fibroin only, water insoluble), and regenerated silk fibroin (with regenerated silk fibroin only, water soluble) in powder form. Experimental data derived from the mechanical measurements illustrated that the miscibility between the PU and regenerated silk fibroin were superior to that between PU and the other two silk proteins. This may be attributed to the similar chemical structure and microphase separation of PU and regenerated silk fibroin with lower molecular weight than native silk fibroin. This preliminary work may provide some information for biomimetic processing of silk‐inspired PU biofibers, which combine elasticity of synthetic PU with biofunction of natural silk fibroin for special biomedical applications. © 2009 Wiley Periodicals, Inc. J Appl Polym Sci, 2009