Effects of Ultraviolet Light Irradiation on Silk Fibroin Films Prepared under Different Conditions.

Silk fibroin (SF)-based materials are exposed to both natural and artificial ultraviolet (UV) light during preparation or administration. However, the effects of UV irradiation on SF films prepared under different conditions have not yet been described in detail. In this study, four SF films with different molecular weight (MW) distribution were fabricated using SF solutions, which were prepared by dissolving degummed SF for 0.5-24 h. We observed UV (365 nm) irradiation on SF films induced the increase of yellowness and absorbance at 310 nm of SF films, indicating the formation of new photo-products and di-tyrosine bonds by photo-oxidation. Due to di-tyrosine cross-links between SF chains, UV-irradiated SF films were not fully dissociated in urea solution. In addition to formation of new products, UV reduced the crystallinity of SF films by breaking hydrogen bonds of ß-sheet conformation. Unlike the UV-induced decomposition of physical interactions, UV did not affect the covalent bonds (i.e., peptide bonds). Through these experiments, we could expect that SF with higher MW was more susceptible and SF with lower MW was more resistant to UV-induced photo-oxidation and photo-degradation. These results provide useful information about UV-induced aging of SF-based materials under natural sunlight and UV irradiating conditions.

Photo-cross-linkable, insulating silk fibroin for bioelectronics with enhanced cell affinity.

Bioelectronic scaffolds that support devices while promoting tissue integration could enable tissue hybrids with augmented electronic capabilities. Here, we demonstrate a photo-cross-linkable silk fibroin (PSF) derivative and investigate its structural, electrical, and chemical properties. Lithographically defined PSF films offered tunable thickness and <1-µm spatial resolution and could be released from a relief layer yielding freestanding scaffolds with centimeter-scale uniformity. These constructs were electrically insulating; multielectrode arrays with PSF-passivated interconnects provided stable electrophysiological readouts from HL-1 cardiac model cells, brain slices, and hearts. Compared to SU8, a ubiquitous biomaterial, PSF exhibited superior affinity toward neurons which we attribute to its favorable surface charge and enhanced attachment of poly-d-lysine adhesion factors. This finding is of significant importance in bioelectronics, where tight junctions between devices and cell membranes are necessary for electronic communication. Collectively, our findings are generalizable to a variety of geometries, devices, and tissues, establishing PSF as a promising bioelectronic platform.

Effects of sterilization methods on the physical, chemical, and biological properties of silk fibroin membranes.

Silk fibroin has been widely explored for many biomedical applications, due to its biocompatibility and biodegradability. Sterilization is a fundamental step in biomaterials processing and it must not jeopardize the functionality of medical devices. The aim of this study was to analyze the influence of different sterilization methods in the physical, chemical, and biological characteristics of dense and porous silk fibroin membranes. Silk fibroin membranes were treated by several procedures: immersion in 70% ethanol solution, ultraviolet radiation, autoclave, ethylene oxide, and gamma radiation, and were analyzed by scanning electron microscopy, Fourier-transformed infrared spectroscopy (FTIR), X-ray diffraction, tensile strength and in vitro cytotoxicity to Chinese hamster ovary cells. The results indicated that the sterilization methods did not cause perceivable morphological changes in the membranes and the membranes were not toxic to cells. The sterilization methods that used organic solvent or an increased humidity and/or temperature (70% ethanol, autoclave, and ethylene oxide) increased the silk II content in the membranes: the dense membranes became more brittle, while the porous membranes showed increased strength at break. Membranes that underwent sterilization by UV and gamma radiation presented properties similar to the nonsterilized membranes, mainly for tensile strength and FTIR results.

Determination of multiphoton absorption of silk fibroin using the Z-scan technique.

The multiphoton absorption of silk fibroin at 810 nm was determined by open-aperture Z-scan. Three-photon absorption was confirmed at this wavelength and the three-photon cross section of silk fibroin was measured. Silk fibroin of varying molecular weight was tested and the cross section was found to increase exponentially with increasing molecular weight. The confirmation of a relatively large three-photon absorption cross section in silk will help lay the groundwork for future investigation of direct laser writing of three-dimensional structures within a silk matrix.

Silk fibroin/poly(vinyl alcohol) photocrosslinked hydrogels for delivery of macromolecular drugs.

Hydrogels are three-dimensional polymer networks widely used in biomedical applications as drug delivery and tissue engineered scaffolds to effectively repair or replace damaged tissue. In this paper we demonstrate a newly synthesized cytocompatible and drug releasing photo-crosslinked hydrogel based on poly(vinyl alcohol) methacrylate and silk fibroin which possesses tailorable structural and biological properties. The initial silk fibroin content was 0%, 10%, 20%, 30%, 40% and 50% with respect to the weight of poly(vinyl alcohol) methacrylate. The prepared hydrogels were characterized with respect to morphology, crystallinity, stability, swelling, mass loss and cytotoxicity. FITC-dextrans of different molecular weights were chosen as model drugs molecules for release studies from the hydrogels. The hydrogels containing different silk fibroin percentages showed differences in pore size and distribution. X-ray diffraction analysis revealed that amorphous silk fibroin in poly(vinyl alcohol) methacrylate is crystallized to ß-sheet secondary structure upon gelation. The sol fraction increased with increasing fibroin concentration in the co-polymer gel (from 18% to 45%), although the hydrogel extracts were non-cytotoxic. Similarly, the addition of silk fibroin increased water uptake by the gels (from 7% to 21%). FITC-dextran release from the hydrogels was dependent on the silk fibroin content and the molecular weight of encapsulated molecules. The study outlines a newer type of photo-crosslinked interpenetrating polymer network hydrogel that possess immense potential in drug delivery applications.

Structural evolution of regenerated silk fibroin under shear: combined wide- and small-angle x-ray scattering experiments using synchrotron radiation.

The structural evolution of regenerated Bombyx mori silk fibroin during shearing with a Couette cell has been studied in situ by synchrotron radiation small- and wide-angle x-ray scattering techniques. An elongation of fibroin molecules was observed with increasing shear rate, followed by an aggregation phase. The aggregates were found to be amorphous with beta-conformation according to infrared spectroscopy. Scanning x-ray microdiffraction with a 5 microm beam on aggregated material, which had solidified in air, showed silk II reflections and a material with equatorial reflections close to the silk I structure reflections, but with strong differences in reflection intensities. This silk I type material shows up to two low-angle peaks suggesting the presence of water molecules that might be intercalated between hydrogen-bonded sheets.

The ESR signals in silk fibroin and wool keratin under both the effect of UV-irradiation and without any external effects and the formation of free radicals.

ESR studies have been done on natural and UV-irradiated silk fibroins and wool keratins at the temperature range of -196 degrees C to 20 C. The intensities of ESR signals obtained from the irradiated samples at -196 C remarkably increase with respect to those of natural samples. While the signals mainly consist of triplet peaks at -196 C. a doublet arises around the room temperatures. For the first time, at room temperature without any external effect the complicated ESR spectra of fibrous proteins (wool keratin and silk fibroin) whose components are as follows have been observed: (1) (for white wool keratin) a central doublet with deltaHm = 1.1 mT and g = 2.0075; deltaHm = 5mT and g = 2.1911; (2) a wide peak with deltaHm approximately 66 mT and g approximately 2.1575; (3) the 'sulfur' peak given in the literature with deltaHm = 2.2 mT and g = 2.0218; (4) the signal with deltaHm = 0.6 mT and g = 2.0065, and for silk fibroin, (a) a very wide signal with deltaHm approximately 70 mT and g approximately 2.084; (b) a very sharp signal with deltaHm approximately 1.1 mT and g approximately 2.01; and (c) relatively narrower signal with deltaHm approximately 5 mT and g approximately 2.336. It has been shown by recombination kinetic method that 30-50% of the free radicals formed by UV-irradiation do not undergo recombination up to 220 degrees C and 15 degrees C for silk libroin and wool keratin, respectively, even they keep their concentration constant for long period of time (weeks, months, even longer). In this article, considering above-mentioned results, the mechanism of signals observed in natural wool keratin and silk fibroin without any external effects is examined. We can briefly explain the role of the subject of the article, by considering fibrous proteins and some applications of the reactions by free radical occurring in these proteins tinder the effects of different factors in medicine and biology and the important role of oxidation and the other kinds of degradations on these processes. as well as the significant applications of ESR investigations on comprehending the processes by free radical.