Effect of the sterilization method on the properties of Bombyx mori silk fibroin films

We have compared the effects of different sterilization techniques on the properties of Bombyx mori silk fibroin thin films with the view to subsequent use for corneal tissue engineering. The transparency, tensile properties, corneal epithelial cell attachment and degradation of the films were used to evaluate the suitability of certain sterilization techniques including gamma-irradiation (in air or nitrogen), steam treatment and immersion in aqueous ethanol. The investigations showed that gamma-irradiation, performed either in air or in a nitrogen atmosphere, did not significantly alter the properties of films. The films sterilized by gamma-irradiation or by immersion in ethanol had a transparency greater than 98% and tensile properties comparable to human cornea and amniotic membrane, the materials of choice in the reconstruction of ocular surface. Although steam-sterilization produced stronger, stiffer films, they were less transparent, and cell attachment was affected by the variable topography of these films. It was concluded that gamma-irradiation should be considered to be the most suitable method for the sterilization of silk fibroin films, however, the treatment with ethanol is also an acceptable method.     

Near-infrared characterization on the secondary structure of regenerated Bombyx mori silk fibroin

Several advantages of the near-infrared (NIR) technique in the characterization of the secondary structure of regenerated Bombyx mori silk fibroin are demonstrated. Silk fibroin films with thicknesses ranging from 50 to 300 microm are suitable for the NIR measurement. The bands due to hydrogen-bonded water are independent in the NIR spectra and facilitate our investigation of the amide region. Analysis of the combination modes of amide groups in the NIR spectra could lead to a profile of conformation ratio of silk fibroin, which was supported by nuclear magnetic resonance (NMR) observations.     

Bombyx mori silk fibre and its composite: A review of contemporary developments

Application of natural fibres in composite plastics is gaining popularity in many industries and particularly the automotive industry. Using natural fibres in polymers provides many advantages over other techniques, and the application areas appear limitless. There is currently a shift of paradigm in the automotive industry towards a “green” outlook, due to consumers’ demand for more environmentally friendly vehicles. Natural fibres are biodegradable and are renewable natural source. These two characteristics are most important for disposal of components at end-life. They are recyclable and can be easily converted into thermal energy through combustion, without leaving residue. Among the natural fibres with proven potential application as reinforcement for polymers, Bombyx mori woven silk fibre is one that recently received special attention from researchers. B. mori silk fibre is one of the best fibres discovered in nature, providing high mechanical properties over frequently used natural fibres like sisal, jute, hemp and coir. This review presents a summary of B. mori woven silk fibre and its composite.     

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.     

湿热工艺对桑蚕丝纤维性能的影响

桑蚕丝(B.mor)是由天然的蚕丝蛋白组成,并可以被光、热、水和微生物等损坏。对湿热工艺诱导桑蚕丝纤维的结构和性能的变化进行了测定,观察发现,在150℃条件下将相对湿度从0%增加到70%会降低其拉伸强度。在一个成熟的环境中引入水可以促进水解降解。丝氨酸和酪氨酸的浓度由于相对湿度增加而增加,这导致了黄度指数的增加。蚕丝纤维的结晶结构没有显示出任何明显的变化,除了产生了少量的结晶副产物。     

Relationships between supercontraction and mechanical properties of spider silk

Typical spider dragline silk tends to outperform other natural fibres and most man-made filaments. However, even small changes in spinning conditions can have large effects on the mechanical properties of a silk fibre as well as on its water uptake. Absorbed water leads to significant shrinkage in an unrestrained dragline fibre and reversibly converts the material into a rubber. This process is known as supercontraction and may be a functional adaptation for the silk's role in the spider's web. Supercontraction is thought to be controlled by specific motifs in the silk proteins and to be induced by the entropy-driven recoiling of molecular chains. In analogy, in man-made fibres thermal shrinkage induces changes in mechanical properties attributable to the entropy-driven disorientation of 'unfrozen' molecular chains (as in polyethylene terephthalate) or the 'broken' intermolecular hydrogen bonds (as in nylons). Here we show for Nephila major-ampullate silk how in a biological fibre the spinning conditions affect the interplay between shrinkage and mechanical characteristics. This interaction reveals design principles linking the exceptional properties of silk to its molecular orientation.     

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.     

Variability of the mechanical properties of bone, and its evolutionary consequences

The relative variabilities (coefficient of variation (CV)) of 10 different mechanical properties of compact bone were determined from 2166 measurements. All measures of variability were made on a minimum of four specimens from any bone. Three pre-yield properties had a CV of about 12%. Six post-yield properties had CVs varying from 24 to 46%. Pre-yield properties increase as a function of mineral content, whereas post-yield properties decrease. These differences give insight into mechanical phenomena occurring at different stages during loading. Furthermore, the fact that some properties are more tightly determined than others has implications for the optimum values set by natural selection. This assertion is made more rigorous using a simple mathematical model for the evolutionarily optimal allocation in a trade-off where one property is imprecisely determined. It is argued that in general the optimum will be biased in favour of the more tightly determined properties than would be the case if all properties had the same CV.     

Enhanced mechanical properties of thermosensitive chitosan hydrogel by silk fibers for cartilage tissue engineering

Articular cartilage has limited repair capability following traumatic injuries and current methods of treatment remain inefficient. Reconstructing cartilage provides a new way for cartilage repair and natural polymers are often used as scaffold because of their biocompatibility and biofunctionality. In this study, we added degummed chopped silk fibers and electrospun silk fibers to the thermosensitive chitosan/glycerophosphate hydrogels to reinforce two hydrogel constructs which were used as scaffold for hyaline cartilage regeneration. The gelation temperature and gelation time of hydrogel were analyzed by the rheometer and vial tilting method. Mechanical characterization was measured by uniaxial compression, indentation and dynamic mechanical analysis assay. Chondrocytes were then harvested from the knee joint of the New Zealand white rabbits and cultured in constructs. The cell proliferation, viability, production of glycosaminoglycans and collagen type II were assessed. The results showed that mechanical properties of the hydrogel were significantly enhanced when a hybrid with two layers of electrospun silk fibers was made. The results of GAG and collagen type II in cell-seeded scaffolds indicate support of the chondrogenic phenotype for chondrocytes with a significant increase in degummed silk fiber–hydrogel composite for GAG content and in two-layer electrospun fiber–hydrogel composite for Col II. It was concluded that these two modified scaffolds could be employed for cartilage tissue engineering.     

Modifications of spider silk sequences in an attempt to control the mechanical properties of the synthetic fibers

Bacteria were genetically engineered to produce two spider silk protein variants composed of basic repeat units combining a flagelliform elastic motif ([GPGGX]₄) and a major ampullate silk strength motif ([linker/poly-alanine]. The secondary structures of the pure recombinant proteins in solution were determined by circular dichroism. The data presented suggest that the nature of the 5th and 10th amino acid (X) in the [GPGGX]₂ elastic motif and temperature have an impact on the amount of β-sheet structures present in the proteins. More specifically, increasing temperatures seem to be positively correlated with β-sheet formation for both proteins and this state is irreversible or reversible when both X (5th and 10th) in the elastic motif are hydrophilic or hydrophobic respectively. Moreover, each pure silk-like protein was able to spontaneously self-assemble into films from aqueous solutions. Two kinds of synthetic fibers were made by pulling fibers from these preassembled films as well as spinning fibers from each protein resolubilized in HFIP. The mechanical data show that the pulled fibers are far tougher than the spun fibers suggesting a better fiber organization. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Biosynthesis and characterization of typical fibroin crystalline polypeptides of silkworm Bombyx mori

We aimed to investigate the self-organization/self-assembly mechanisms of silkworm fibroin-based material. In the present study, for the first time, we designed and multimerized four DNA “monomer” sequences from structurally simple fibroin crystalline peptides or analog, [GAGAGX] (X = A, S, Y and V) to encode polypeptides [GAGAGX]₁₆ (eGA, eGS, eGY and eGV) using a “head-to-tail” construction strategy. Multimers were cloned into pGEX-KG and fusion proteins GST-[GAGAGX]₁₆ (KGA, KGS, KGY and KGV) were efficiently expressed in Escherichia coli. These fusion proteins were isolated and purified by GST affinity chromatography and confirmed by SDS–PAGE and Western blot analysis using antibody reactive to GST. The polypeptides were cleavaged from GST fusion proteins by digesting with thrombin enzyme. The composition of the four polypeptides was confirmed by composition analysis of amino acids, and their abilities to form β-sheet structure were determined by ThT fluorescence spectral analysis. The content of β-sheet among the four polypeptides followed the order: eGS > eGV > eGY > eGA.     

原子力显微镜下蚕丝及蜘蛛丝的微观结构

利用原子力显微镜研究了蚕丝丝素、蜘蛛牵引丝及其内外层包卵丝的微观结构.研究表明,丝素和包卵丝纤维的纵向表面都有成丝过程中液态丝蛋白流动而形成的清晰的构槽和条纹,在低速下自然分泌的牵引丝的表面皮层相对比较细腻,而垂直下落蜘蛛在高速下分泌的牵引丝具有和丝素纤维比较相似的微观结构特征.这些丝纤维的断面内都分布有大量微细的原纤,形状基本为圆形,其中三种蜘蛛丝的微纤维直径相似,而丝素纤维内的微纤维要粗得多.     

Molecular cloning of Bombyx mori cytochrome P450 gene and its involvement in fluoride resistance

To investigate the effects of fluorosis on development and gene expression profiles of silkworm, highly resistant silkworm strain 441, and highly susceptible silkworm strain 440 were treated with 200 ppm fluoride (designated as 440F and 441F) and water (designated as 440DZ and 441DZ). Fluorotic silkworm showed body color and behavior changes. Statistical analysis indicated that growth index of 440F was lower than 440DZ, 441DZ, and 441F. The mortality of 440F was higher than others. Fluorescent differential display enabled us to obtain a differentially expressed cDNA. Bioinformatics analyses indicated that it belonged to cytochrome P450 family, denoted Bmcyp306a1, which contained seven exons and six introns. Phylogenetic tree showed BmCYP306A1 had high homology with Manduca sexta’ P450 protein. Expression analysis indicated that Bmcyp306a1 was exclusively expressed in 441DZ and 441F and was down-regulated under fluoride treatment. The tissue-specific expression indicated Bmcyp306a1 had high-expression level in midgut and ovary in 441F. The data revealed that there was obvious dose–effect and times relationship with the pathological changes and gene expression. Expression profiles of Bmcyp306a1 suggested that P450 gene was crucial to physiological modification and might be involved in fluoride resistance.     

蜘蛛丝力学性能的多变性

为进一步探索蜘蛛丝的成丝机理,研究了不同个体大小,不同生存方式,不同环境湿度的大腹圆蛛牵引丝的拉伸性能。结果表明,蜘蛛对其丝纤维的结构和性能具有自我调控能力,随着环境,自身条件,生存方式等成丝条件的变化,蜘蛛牵引丝的力学性能也会发生相应的变化。     

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.     

Variability in atmospheric light-scattering properties with altitude

The altitudinal variability in angular scattering properties of the troposphere and stratosphere was measured with a balloonborne polar nephelometer. An analysis of interdependent experimental parameters provides information on the scattering phase function and inferred size distribution of aerosol particles as a function of altitude. These aerosol characteristics are extractable from the scattering dissymmetry index and its relationship to spectral dispersion in turbidity and the aerosol phase function. The problem of the correlation between backscatter and extinction is addressed from the perspectives of in situ observations.     

Mechanical properties and structure of silkworm cocoons: A comparative study of Bombyx mori,Antheraea assamensis,Antheraea pernyi and Antheraea mylitta silkworm cocoons

As a protective shell against environmental damage and attack by natural predators, the silkworm cocoon has outstanding mechanical properties. In particular, this multilayer non-woven composite structure can be exceptionally tough to enhance the chance of survival for silkworms while supporting their metabolic activity. Peel, out-of-plane compression and nano-indentation tests and micro-structure analysis were performed on four types of silkworm cocoon walls (domesticated Bombyx mori, semi-domesticated Antheraea assamensis and wild Antheraea pernyi and Antheraea mylitta silkworm cocoons) to understand the structure and mechanical property relationships. The wild silkworm cocoons were shown to be uniquely tough composite structures. The maximum work-of-fracture for the wild cocoons (A. pernyi and A. mylitta) was approximately 1000 J/m², which was almost 10 times the value for the domesticated cocoon (Bombyx mori) and 3 ~ 4 times the value for the semi-domesticated cocoon (A. assamensis). Calcium oxalate crystals were found to deposit on the outer surfaces of the semi-domesticated and wild cocoons. They did not show influence in enhancing the interlaminar adhesion between cocoon layers but exhibited much higher hardness than the cocoon pelades.     

Variability in mechanical performance of G-10cr cryogenic-grade insulating laminates

An assessment is made of the variability in cryogenic mechanical performance of insulating laminates produced by five manufacturers to a common component and procedure specification. Results at 295 K and 76 K indicate that the specification is adequate to ensure the desired degree of product uniformity.