Structure Of Silk Fibroin Research Articles

Silk fibroin double-layer microneedles for the encapsulation and controlled release of triptorelin

A double-layer silk fibroin microneedles (SF-MNs) was proposed for the transdermal delivery of triptorelin. Two-step pouring and centrifugation were employed to prepare SF-MNs. Triptorelin was wrapped in MNs in the form of microcrystals with a size of ∼1 μm. β-sheet nanocrystals (the secondary structure of silk fibroin) were adjusted in content by methanol-vapor treatment to manipulate the characteristics of SF-MNs prepared with two concentrations of silk fibroin. The mechanical strength of MNs was measured and analyzed in proportion to the β-sheet content. The triptorelin in MNs could be released sustainedly in phosphate-buffered saline for 168 h, and the release amount decreased with increasing β-sheet content. The Ritger–Peppas equation was employed to fit the release data. A linear decreasing relationship was observed between the diffusion coefficient and increased β-sheet content. After administration to rats, SF-MNs exhibited long-term testosterone inhibition and maintained castration levels for ≥7 d. Manipulable mechanical properties and release behavior combined with biocompatibility and biodegradability render SF-MNs as viable long-term transdermal delivery devices for triptorelin.

Silk Fibroin Microneedle Patches for the Treatment of Insomnia.

As a patient-friendly technology, drug-loaded microneedles can deliver drugs through the skin into the body. This system has broad application prospects and is receiving wide attention. Based on the knowledge acquired in this work, we successfully developed a melatonin-loaded microneedle prepared from proline/melatonin/silk fibroin. The engineered microneedles’ morphological, physical, and chemical properties were characterized to investigate their structural transformation mechanism and transdermal drug-delivery capabilities. The results indicated that the crystal structure of silk fibroin in drug-loaded microneedles was mainly Silk I crystal structure, with a low dissolution rate and suitable swelling property. Melatonin-loaded microneedles showed high mechanical properties, and the breaking strength of a single needle was 1.2 N, which could easily be penetrated the skin. The drug release results in vitro revealed that the effective drug concentration was obtained quickly during the early delivery. The successful drug concentration was maintained through continuous release at the later stage. For in vivo experimentation, the Sprague Dawley (SD) rat model of insomnia was constructed. The outcome exhibited that the melatonin-loaded microneedle released the drug into the body through the skin and maintained a high blood concentration (over 5 ng/mL) for 4–6 h. The maximum blood concentration was above 10 ng/mL, and the peak time was 0.31 h. This system indicates that it achieved the purpose of mimicking physiological release and treating insomnia.

Investigation of chip formation mechanism in ultra-precision diamond turning of silk fibroin film

The importance of silk fibroin particles (size from tens of nanometres to hundreds of microns) as effective drug carriers has been well identified due to their superior mechanical strength, biocompatibility and biodegradability. They are usually fabricated by methods that have difficulties in controlling the size and shape to meet the required functions. In addition, using chemical agents in the fabrication process may result in biological risks and degradation of the fibroin structures. In this study, ultra-precision diamond turning (UPDT) technology is researched as an alternative chemical-free manufacturing approach to generate silk particles in the form of cutting chips with controllable size and shape. The machinability in UPDT of silk fibroin film (hereinafter referred to as silk film) was discussed in terms of specific cutting force and chip morphology. The material removal in UPDT of silk film is taken place in the ductile regime. Experimental results reveal that a rise in the cutting speed could significantly reduce the specific cutting force, while decreasing the depth of cut (DOC) could increase the specific cutting force because of the size effect. Moreover, a sharp point tool using a less than 2.5 μm/rev feed rate is preferred to generate helical silk chips. The formation mechanism of shear bands and serrated chips in UPDT of silk film was investigated with the aid of a finite element and smoothed particle hydrodynamics (FE-SPH) hybrid numerical model with Cowper-Symonds (CS) material equation. The material parameters of silk film were determined for the first time at p = 7 and D = 1140 s−1. The simulated specific cutting force with this set of material parameters is 49.0% smaller than the measured one. The simulated chip morphology (e.g. shear band spacing) was also in good agreement with the measured values. The simulation study shows that the shear band formation is done via two joining parts: one part propagates from the cutting edge to the free surface, another part initiates on the free surface and evolves towards the cutting edge. The chip segment is formed by a microcrack propagating from the free surface to the cutting edge. The underlying mechanism of formation of the serrated chip is found to have its roots in the hierarchical structure of silk fibroin.

The post-processing temperature or humidity can importantly control the secondary structure and characteristics of silk fibroin films.

Temperature and humidity (TH) are highly important factors that can control the secondary structure and characterization of silk fibroin (SF) biomaterials. In this study, the water stability, secondary structure, mechanical properties, surface morphology, and degradation of silk fibroin films (SFFs) with post-processing in different TH were investigated. Fourier transform infrared indicated that the SFF secondary structure did not change under low-relative humidity (RH) despite temperatures up to 180°C, while it transformed at 40°C with 100% RH in 10min. A film with a higher tensile strength (42.1 ± 8.2MPa) could be obtained after post-processing at 90°C/100% RH for 10min. While a film with higher ductility (elongation at break: 198.8 ± 31.8%) was generated after post-processing at 40°C/100% RH for 10min. Scanning electron microscope showed that the film presented a network structure of nanoparticles in series under certain TH post-treatment. Enzymatic hydrolysis proved that the SFFs containing a higher content of silk II structure degraded more slowly. Therefore, TH post-treatment is a relatively mild way to change the secondary structure and properties of SFFs, which can be widely used in loading drugs and maintaining the activity of drugs in SF biomaterials.

An Insight into Tunable Innate Piezoelectricity of Silk for Green Bioelectronics.

Commercially available bioelectronics account for significant percentage of e-waste, especially battery waste, that demand immediate intervention due to rising environmental concerns. Consumers are becoming increasingly aware and cautious of their contribution to carbon footprint on a regular basis. It has become imperative to adopt sustainability in every aspect of production of bioelectronics taking into consideration the growing market for wearable healthcare monitoring system. Green electronics is a relatively new concept gaining tremendous attention within the scientific and industrial community with the ultimate goal of employing organic, biodegradable, and self-sustainable system to replace the conventional inorganic battery-powered electronics. Silk is a green material that has been extensively explored for its use in functional electronics due to its tunable biodegradability and flexibility. Nevertheless, an intriguing property of Silk is its innate piezoelectricity. This review highlights the importance of crystal orientation and structure of Silk Fibroin to display piezoelectric response and documents possible strategies for its enhancement. It also provides insight into the possibility of using piezoelectric Silk as a piezoelectric sensor, actuator, and energy harvester to form self-powered hybrid systems for autonomous bioelectronics.

Regulating the mechanics of silk fibroin scaffolds promotes wound vascularization

Functional blood vessels are crucial to wound healing, and faster vascularization means faster tissue repair to some extent. Increasing numbers of pro-vascularization wound coverings are being developed and studied. Moreover, mechanical properties of the extracellular matrix can guide the behaviour of related cells to some degree. Studies have shown that the mechanical range of 1–7 kPa contributes to the differentiation of stem cells into endothelial cells and thus to the process of wound vascularization. Unfortunately, the regulatory mechanics of vascularizing wound coverings have been poorly studied. Silk fibroin (SF) has attracted much attention because of its good biocompatibility, degradability and adjustable mechanical properties. In this paper, silk scaffolds with mechanical properties of 2 kPa and 5.9 kPa were prepared by adjusting the mechanics of silk scaffolds in terms of freezing temperature and aligned structure. The mechanical properties of the 5.9 kPa aligned silk scaffold (ASS) showed good vascularization ability. By adjusting the intermediate conformation and physical structure of Silk fibroin (SF), the mechanical strength of the silk scaffold could be increased, enabling us to better understand the mechanical regulation mode. At the same time, the aligned structure of the aligned silk scaffold (ASS) promoted the migration and proliferation of cells related to wound repair to a certain extent. By adjusting the mechanical properties and physical structure of the material, an aligned silk scaffold with vascularization function was constructed, providing more possibilities for faster wound repair.

Structure of Silk I (Bombyx mori Silk Fibroin before Spinning) -Type II β-Turn, Not α-Helix.

Recently, considerable attention has been paid to Bombyx mori silk fibroin by a range of scientists from polymer chemists to biomaterial researchers because it has excellent physical properties, such as strength, toughness, and biocompatibility. These appealing physical properties originate from the silk fibroin structure, and therefore, structural determinations of silk fibroin before (silk I) and after (silk II) spinning are a key to make wider applications of silk. There are discrepancies about the silk I structural model, i.e., one is type II β-turn structure determined using many solid-state and solution NMR spectroscopies together with selectively stable isotope-labeled model peptides, but another is α-helix or partially α-helix structure speculated using IR and Raman methods. In this review, firstly, the process that led to type II β-turn structure by the authors was introduced in detail. Then the problems in speculating silk I structure by IR and Raman methods were pointed out together with the problem in the assignment of the amide I band in the spectra. It has been emphasized that the conformational analyses of proteins and peptides from IR and Raman studies are not straightforward and should be very careful when the proteins contain β-turn structure using many experimental data by Vass et al. In conclusion, the author emphasized here that silk I structure should be type II β-turn, not α-helix.

Study on Non-knotted Barbed Suture with Silk Fibroin Structure

Study on Non-knotted Barbed Suture with Silk Fibroin Structure

Chemical, Thermal, Time, and Enzymatic Stability of Silk Materials with Silk I Structure.

The crystalline structure of silk fibroin Silk I is generally considered to be a metastable structure; however, there is no definite conclusion under what circumstances this crystalline structure is stable or the crystal form will change. In this study, silk fibroin solution was prepared from B. Mori silkworm cocoons, and a combined method of freeze-crystallization and freeze-drying at different temperatures was used to obtain stable Silk I crystalline material and uncrystallized silk material, respectively. Different concentrations of methanol and ethanol were used to soak the two materials with different time periods to investigate the effect of immersion treatments on the crystalline structure of silk fibroin materials. X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), Raman scattering spectroscopy (Raman), Scanning electron microscope (SEM), and Thermogravimetric analysis (TGA) were used to characterize the structure of silk fibroin before and after the treatments. The results showed that, after immersion treatments, uncrystallized silk fibroin material with random coil structure was transformed into Silk II crystal structure, while the silk material with dominated Silk I crystal structure showed good long-term stability without obvious transition to Silk II crystal structure. α-chymotrypsin biodegradation study showed that the crystalline structure of silk fibroin Silk I materials is enzymatically degradable with a much lower rate compared to uncrystallized silk materials. The crystalline structure of Silk I materials demonstrate a good long-term stability, endurance to alcohol sterilization without structural changes, and can be applied to many emerging fields, such as biomedical materials, sustainable materials, and biosensors.

Rapid Synthesis of Silk-Like Polymers Facilitated by Microwave Irradiation and Click Chemistry.

Silk is a natural fiber that surpasses most man-made polymers in its combination of strength and toughness. Silk fibroin, the primary protein component of silk, can be synthetically mimicked by a linear copolymer with alternating rigid and soft segments. Strategies for chemical synthesis of such silk-like polymers have persistently resulted in poor sequence control, long reaction times, and low molecular weights. Here, we present a two-stage approach for rapidly synthesizing silk-like polymers with precisely defined rigid blocks. This approach utilizes solid-phase peptide synthesis to create uniform oligoalanine "prepolymers", followed by microwave-assisted step-growth polymerization with bifunctional poly(ethylene glycol). Multiple coupling chemistries and reaction conditions were explored, with microwave-assisted click chemistry yielding polymers with Mw ∼ 14 kg/mol in less than 20 min. These polymers formed antiparallel β-sheets and nanofibers, which is consistent with the structure of natural silk fibroin. Thus, our strategy demonstrates a promising modular approach for synthesizing silk-like polymers.

Chain-folded lamellar structure and dynamics of the crystalline fraction of Bombyx mori silk fibroin and of (Ala-Gly-Ser-Gly-Ala-Gly)n model peptides

Solid-state NMR is a powerful analytical technique to determine the composite structure of Bombyx mori silk fibroin (SF). In our previous paper, we proposed a lamellar structure for Ala-Gly copolypeptides as a model of the crystalline fraction in Silk II. In this paper, the structure and dynamics of the crystalline fraction and of a better mimic of the crystalline fraction, (Ala-Gly-Ser-Gly-Ala-Gly)n (n = 2–5, 8), and 13C selectively labeled [3-13C]Ala-(AGSGAG)5 in Silk II forms, were studied using structural and dynamical analyses of the Ala Cβ peaks in 13C cross polarization/ magic angle spinning NMR and 13C solid-state spin-lattice relaxation time (T1) measurements, respectively. Like Ala-Gly copolypeptides, these materials have lamellar structures with two kinds of Ala residues in β-sheet, A and B, plus one distorted β-turn, t, formed by repetitive folding using β-turns every eighth amino acid in an antipolar arrangement. However, because of the presence of Ser residues at every sixth residue in (AGSGAG)n, the T1 values and mobilities of B decreased significantly. We conclude that the Ser hydroxyls hydrogen bond to adjacent lamellar layers and fix them together in a similar way to Velcro®.

Processing Strategies to Obtain Highly Porous Silk Fibroin Structures with Tailored Microstructure and Molecular Characteristics and Their Applicability in Water Remediation

The present work reports on the control of silk fibroin (SF) porous structures performance through various processing methods. The study includes the analysis of two dissolving techniques (CaCl2/H2O/EtOH ternary and LiBr/H2O binary solutions), three regeneration methods (gelation, lyophilization and gas foaming) and one post-processing (EtOH). In all the cases, followed steps lead to SF structures with porosity values above 94% and large surface areas. Also, results about samples microstructure, secondary organization, crystallinity and water behavior, reveal a direct correlation between processing and SF properties.Thanks to the achieved progress, the SF varying porous structures were evaluated for metalloids (As5+ and As3+) and heavy metals (Cr6+ and Cr3+) adsorption, observing a direct relationship between samples processing and ionic species adsorption ability.Thus, it is shown that the control of the properties of SF based porous structures through processing, represents a suitable and ecofriendly approach for the development of bio-based materials for environmental applications.

Silk as a biodegradable resist for field-emission scanning probe lithography

The patterning of silk allows for manufacturing various structures with advanced functionalities for optical and tissue engineering and drug delivery applications. Here, we propose a high-resolution nanoscale patterning method based on field-emission scanning probe lithography (FE-SPL) that crosslinks the biomaterial silk on conductive indium tin oxide (ITO) promoting the use of a biodegradable material as resist and water as a developer. During the lithographic process, Fowler-Nordheim electron emission from a sharp tip was used to manipulate the structure of silk fibroin from random coil to beta sheet and the emission formed nanoscale latent patterns with a critical dimension (CD) of ∼50 nm. To demonstrate the versatility of the method, we patterned standard and complex shapes. This method is particularly attractive due to its ease of operation without relying on a vacuum or a special gaseous environment and without any need for complex electronics or optics. Therefore, this study paves a practical and cost-effective way toward patterning biopolymers at ultra-high level resolution.

THE EFFECT OF SELENIUM ON THE SUPRAMOLECULAR STRUCTURE AND THERMAL CHARACTERISTICS OF FIBROIN BOMBYX MORI L

The enrichment of the molecular structure of silk fibroin by selenium atom led to an increase in the branching of fibroin macromolecule. As a result, the amorphous fraction of fibroin microfiber increases which leads to an increase in the strength characteristic of the silk thread. At the same time, this change in the supramolecular structure of fibroin involving a selenium atom has enabled us to study the two-modification mechanism for crystallizing fibroin microfibers. Based on studies on the use of temperature-gravimetric and X-ray diffraction (XRD) analysis and relative changes in the proportion of amorphous and crystalline regions, we came to the conclusion that fibroin microfibrils consist mainly of extended crystallites CSC – crystallites with the stretched chains. They alternate along the axis of microfiber with amorphous layers, the sizes of which are smaller than the sizes of the folded crystallites (CFC). Therefore, CFC cannot be located in amorphous layers between the CSC along the microfiber’s axis. As a result, the ability to fold branched sections of the macromolecule decreases, that is, CFC decreases. This increases the proportion of amorphous areas of microfibers of the fibroin. In the model, which is proposed by author non-crystallized in the form of CSC, segments of a macromolecule, on the sides of the central core and attached to it, restored their crystal structure (CFC) – with folded conformation of chains.

Direct Observation of Native Silk Fibroin Conformation in Silk Gland of Bombyx mori Silkworm.

To understand the natural silk spinning mechanism, synchrotron Fourier transform infrared (S-FTIR) microspectroscopy was employed in this study to monitor the conformation changes of silk protein in the silk gland of Bombyx mori silkworm. The ultrahigh brightness of S-FTIR microspectroscopy allowed the imaging of the silk gland with micrometer-scale spatial resolution. Herein, tissue sections of a silk gland, including cross-section slices and longitudinal-section slices, were characterized. The results obtained clearly confirm that the conformation of the silk fibroin changes gradually along the silk gland from the tail to the spinneret. In the middle silk gland, silk fibroin mainly contains random coil/helix conformation. When it comes to the spinneret through the anterior silk gland, the content of β-sheet increases, but the content of random coil/helix instead reduces gradually. Further, the β-sheet distribution in the cross-section of the anterior silk gland was imaged using S-FTIR mapping technique. The results show that the structural distribution of the silk fibroin in cross-section is uniform without significant shell-core structure, which implies that the primary driving force to induce the conformation transition of silk fibroin from random coil/helix to β-sheet during the spinning process is elongational flow of silk fibroin in the silk gland and not the shear force between the silk fibroin and the lumen wall of silk gland. These direct pieces of evidence of silk fibroin structure in the silk gland would definitely promote a deeper understanding of the natural spinning process.

The extraction and measurement of nickel metal ion in crab, shellfish and rice samples using magnetic silk fibroin – EDTA ligand and furnace atomic absorption spectrometry

The main aim of the present study was the measurement of nickel metal ion in the real samples of crab, oyster and rice by the designed magnetic nano adsorbent silk fibroin-EDTA ligand (SF-Fe3O4-EDTA). Due to the structure of silk fibroin (possessing lots of functional groups which are suitable for attachment of ligands and high surface area), it was used in the structure of fabricated nano-adsorbent. To follow the fabrication processes of the magnetic nano-adsorbent, different techniques of fourier-transform infrared spectroscopy (FTIR), ultraviolet–visible spectroscopy (UV–Visible), vibrating sample magnetometer (VSM), and field emission scanning electron microscopy (FE-SEM) were used. The optimization processes were performed with the chemometric method of response surface modeling with sufficient accuracy and precision. Using this chemometric method, the optimum values of pH, absorption time, the concentration of nano-adsorbent and temperature were calculated to be 6, 21 min, 4 mg L−1 and 28 °C, respectively. Due to the magnetic nature of the constructed nano-adsorbent, a magnet bar was used to separate the nano-adsorbent from the solution and then inject to the furnace atomic absorption device. Using the magnetic nano-adsorbent of silk fibroin-EDTA ligand and furnace atomic absorption a detection limit of 0.0017 µg L−1 and a linear range of 0.0030–5.0 µg L−1 for determination of nickel metal ion were obtained. The determination of nickel metal ion in the crab tissue, oyster tissue and rice samples were performed and the obtained results revealed the successful applicability of the designed method for determination of nickel metal ion in the real samples.

RETRACTED ARTICLE: Glyoxal modification mediates conformational alterations in silk fibroin: Induction of fibrillation with amyloidal features

Silkwormsilk protein fibroin is widely exploited to develop novel silk-based biomaterials due to its stable β-sheet structure, providing high crystallinity and tensile strength. The polymorphic behaviour of silk fibroin provides a window to modulate its structural transitions during self-assembly for different functional outcomes. Most studies are therefore mainly focused on formation of well-developed β-sheet structure and self-assembly of silk fibroin which are regulated by many parameters. Glyoxal, a highly reactive α-oxoaldehyde, reacts with different proteins to form advanced glycation end products (AGEs) following Maillard-like reaction. Considering the significance of protein modification by glyoxal-derived AGEs, in the present study the effect of glyoxal (250, 500 and 1000 μM) on the structure of silk fibroin has been investigated. CD and fluorescence studies reveal that higher concentrations of the α-oxoaldehyde induce considerable alterations of secondary and tertiary structure of the protein leading to aggregation following incubation with for 3 weeks. The aggregates exhibit fibrillar morphology with amyloidal nature as evident from SEM, FTIR and XRD experiments. The findings highlight that glycationinduced modification can be a possible approach for modulating the conformation of the silk protein which may be relevant in connection to clinical, biomedical or synthetic biology based applications.

Recent advances in silk-based wearable sensors

In recent years, wearable electronics has received extensive attention, providing new opportunities for implementing health monitoring, human disease diagnosis and treatment, and intelligent robotics. Sensor is one of the key components of wearable electronics. Silk (Bombyx Mori) material shows unique features including high yield, excellent tensile strength (0.5–1.3 GPa) and toughness ((6–16) × 10<sup>4</sup> J/kg), good biocompatibility, programmable/controllable biodegradability, novel dielectric properties, and various material formats. With the rapid development of biomaterials and related manufacturing technologies, advanced silk-based materials have been studied and applied to wearable sensors. Here, we firstly introduce the five-level structure of silk fibroin from bottom to top and characteristics of silk-based advanced materials, and then review the research progress of silk-based advanced materials in wearable sensors in recent years, including mechanical sensors, electrophysiological sensors, temperature sensors and humidity sensors. The working mechanism, structure and performance of different sensors, the role of silk proteins in them, and their applications in health monitoring are discussed and summarized. Finally, the challenges and future prospects of silk-based wearable sensors in practical applications are put forward.

Influence of alcohol treatments on properties of silk-fibroin-based films for highly optically transparent coating applications.

Thin films of silk fibroin were prepared by solvent evaporation from calcium chloride/ethanol aqueous solution. The influence of alcohol treatments on thermal, mechanical and optical properties of silk-fibroin-based film is presented. To understand the conformal structure of the alcohol-treated silk fibroin film, the IR spectral decomposition method is employed. The optical properties especially the optical transparency, haze and fluorescence emission of alcohol-treated silk fibroin film is systematically investigated together with the conformal structure to understand the effect of the fibril such as the beta-sheet influencing the optical properties. Monohydric alcohol treatment increased beta-turn content in the regenerated silk fibroin structure. These affected the amount of light diffusion and scattering within silk-fibroin films. With alcohol-treatment, all the silk-fibroin films exhibit exceptional optical transparency (>90%) with different levels of optical haze (2.56–14.17%). In particular, ethanol-treated silk-fibroin films contain the highest content of beta-turns (22.8%). The ethanol-treated silk-fibroin films displayed a distinct interference of oscillating crests and troughs in the UV-Vis transmittance spectra, thereby showing the lowest optical haze of 2.56%. In contrast, the silk-fibroin films treated with methanol and propanol exhibit the highest (14.17%) and second-highest (10.29%) optical transmittance haze, respectively. The beta-turn content of the silk-fibroin films treated with methanol is the lowest (20.5%). These results show the relationship between the beta-turn content and optical haze properties. The results manifestly provide a method to manufacture exceptional optically transparent silk-fibroin films with adjustable light diffusion and scattering which can be designed to meet specific applications with the potential to provide UV-shielding protection via monohydric alcohol treatment.

Biomimetic sulfated silk nanofibrils for constructing rapid mid-molecule toxins removal nanochannels

Biomacromolecules and their assemblies exhibit the great potential for biomimetic construction of novel and functional nanomaterials. In this work, sulfated silk nanofibrils (SSNFs) were firstly prepared with special designed sulfated silk fibroin molecule by a facile self-assembly technique, which were incorporated into the poly (vinyl alcohol) (PVA) hydrogel. Then they were together coated on the electrospun polyacrlonitrile (PAN) nanofibrous scaffold to fabricate a novel PVA/SSNFs/PAN thin film nanofibrous composite (TFNC) membranes for hemodiafiltration. Combining the results from filtration and hemodiafiltration tests, it could be deduced that the formation of nanogaps between PVA matrix and SSNFs provided more nanochannels for water and uremic toxins to transport rapidly through the functional layer. Especially the incorporation of SSNFs into PVA hydrogel layer endowed PVA/SSNFs TFNC hemodiafiltration membranes with rapid mid-molecule toxins removal channels. PVA/SSNFs-5 could clear 65.7% of lysozyme, also maintaining 85.0% clearance of urea and 94.7% interception of BSA after 4 h of dialysis. In addition, the blood compatibility of PVA/SSNFs TFNC membranes was also enhanced after the introduction of biomimetic SSNFs due to the heparin-like structure of sulfated silk fibroin (SSF). The biomimetic assembly route demonstrated here represents a green and efficient way for designing various functional nanomaterials in bio-nanotechnologies.