Silk Dope Research Articles

The ratio of fibroin to sericin in the middle silk gland of Bombyx mori and its correlation with the extensional behavior of the silk dope.

Understanding how native silk spinning occurs is crucial for designing artificial spinning systems. One often overlooked factor in Bombyx mori is the secretion of sericin proteins. Herein, we investigate the variation in amino acid content at different locations in the middle silk gland (MSG) of B. mori. This variation corresponds to an increase in sericin content when moving towards the anterior region of the MSG, while the posterior region predominantly contains fibroin. We estimate the mass ratio of sericin to fibroin to be ~25/75 wt% in the anterior MSG, depending on the fitting method. Then, we demonstrate that the improvement in the extensional behavior of the silk dope in the MSG correlates with the increase in sericin content. The addition of sericin may decrease the viscosity of the silk dope, a factor associated with an increase in the spinnability of silk. We further discuss whether this effect could also result from other known physicochemical changes within the MSG.

Native-like Flow Properties of an Artificial Spider Silk Dope.

Recombinant spider silk has emerged as a biomaterial that can circumvent problems associated with synthetic and naturally derived polymers, while still fulfilling the potential of the native material. The artificial spider silk protein NT2RepCT can be produced and spun into fibers without the use of harsh chemicals and here we evaluate key properties of NT2RepCT dope at native-like concentrations. We show that NT2RepCT recapitulates not only the overall secondary structure content of a native silk dope but also emulates its viscoelastic rheological properties. We propose that these properties are key to biomimetic spinning and that optimization of rheological properties could facilitate successful spinning of artificial dopes into fibers.

Fine structure of the silk spinning system in the caddisworm, Hydatophylax nigrovittatus (Trichoptera: Limnephilidae)

Silk is produced by a variety of insects, but only silk made by terrestrial arthropods has been examined in detail. To fill the gap, this study was designed to understand the silk spinning system of aquatic insect. The larvae of caddis flies, Hydatophylax nigrovittatus produce silk through a pair of labial silk glands and use raw silk to protect themselves in the aquatic environment. The result of this study clearly shows that although silk fibers are made under aquatic conditions, the cellular silk production system is quite similar to that of terrestrial arthropods. Typically, silk production in caddisworm has been achieved by two independent processes in the silk glands. This includes the synthesis of silk fibroin in the posterior region, the production of adhesive glycoproteins in the anterior region, which are ultimately accumulated into functional silk dope and converted to a silk ribbon coated with gluey substances. At the cellular level, each substance of fibroin and glycoprotein is specifically synthesized at different locations, and then transported from the rough ER to the Golgi apparatus as transport vesicles, respectively. Thereafter, the secretory vesicles gradually increase in size by vesicular fusion, forming larger secretory granules containing specific proteins. It was found that these granules eventually migrate to the apical membrane and are exocytosed into the lumen by a mechanism of merocrine secretion.

The Importance of a Filament-like Structure in Aerial Dispersal and the Rarefaction Effect of Air Molecules on a Nanoscale Fiber: Detailed Physics in Spiders' Ballooning.

Many flying insects utilize a membranous structure for flight, which is known as a "wing." However, some spiders use silk fibers for their aerial dispersal. It is well known that spiders can disperse over hundreds of kilometers and rise several kilometers above the ground in this way. However, little is known about the ballooning mechanisms of spiders, owing to the lack of quantitative data. Recently, Cho et al. discovered previously unknown information on the types and physical properties of spiders' ballooning silks. According to the data, a crab spider weighing 20 mg spins 50-60 ballooning silks simultaneously, which are about 200 nm thick and 3.22 m long for their flight. Based on these physical dimensions of ballooning silks, the significance of these filament-like structures is explained by a theoretical analysis reviewing the fluid-dynamics of an anisotropic particle (like a filament or a high-slender body). (1) The filament-like structure is materially efficient geometry to produce (or harvest, in the case of passive flight) fluid-dynamic force in a low Reynolds number flow regime. (2) Multiple nanoscale fibers are the result of the physical characteristics of a thin fiber, the drag of which is proportional to its length but not to its diameter. Because of this nonlinear characteristic of a fiber, spinning multiple thin ballooning fibers is, for spiders, a better way to produce drag forces than spinning a single thick spider silk, because spiders can maximize their drag on the ballooning fibers using the same amount of silk dope. (3) The mean thickness of fibers, 200 nm, is constrained by the mechanical strength of the ballooning fibers and the rarefaction effect of air molecules on a nanoscale fiber, because the slip condition on a fiber could predominate if the thickness of the fiber becomes thinner than 100 nm.

Improving the Tensile Properties of Wet Spun Silk Fibers Using Rapid Bayesian Algorithm.

Wet spinning of silkworm silk has the potential to overcome the limitations of the natural spinning process, producing fibers with exceptional mechanical properties. However, the complexity of the extraction and spinning processes have meant that this potential has so far not been realized. The choice of silk processing parameters, including fiber degumming, dissolving, and concentration, are critical in producing a sufficiently viscous dope, while avoiding silk's natural tendency to gel via self-assembly. This study utilized recently developed rapid Bayesian optimization to explore the impact of these variables on dope viscosity. By following the dope preparation conditions recommended by the algorithm, a 13% (w/v) silk dope was produced with a viscosity of 0.46 Pa·s, approximately five times higher than the dope obtained using traditional experimental design. The tensile strength, modulus, and toughness of fibers spun from this dope also improved by a factor of 2.20×, 2.16×, and 2.75×, respectively. These results represent the outcome of just five sets of experimental trials focusing on just dope preparation. Given the number of parameters in the spinning and post spinning processes, the use of Bayesian optimization represents an exciting opportunity to explore the multivariate wet spinning process to unlock the potential to produce wet spun fibers with truly exceptional mechanical properties.

Tensile properties of synthetic pyriform spider silk fibers depend on the number of repetitive units as well as the presence of N- and C-terminal domains

Spiders can spin seven different types of silk, some of which are well characterized, but studies on natural and synthetic pyriform silks are few. In this study, recombinant spidroins composed of one to three pyriform repeat units from Araneus ventricosus, in some cases flanked with non-repetitive N- and C-terminal domains (NT and CT), were produced and spun into continuous silk fibers using a wet-spinning process in organic solvents. All the fibers showed high and similar tensile strain (60–80%), but the Young's modulus, stress and toughness of fibers increased with increasing number of repeat units and in the presence of NT and CT as well. Systematic studies of the secondary structure contents of the different spinning dopes and spun fibers revealed no major differences between the different types of recombinant spidroins. This suggests that optimal tensile properties of artificial spider silks require the presence of several repetitive units as well as terminal domains and that secondary structure content of silk dope and fibers have limited correlation with mechanical behaviors.

Structure Water-Solubility Relationship in α-Helix-Rich Films Cast from Aqueous and 1,1,1,3,3,3-Hexafluoro-2-Propanol Solutions of S. c. ricini Silk Fibroin.

Silk fibroin (SF) produced by the domesticated wild silkworm, Samia cynthia ricini (S. c. ricini) is attracting increasing interest owing to its unique mechanical properties, biocompatibility, and abundance in nature. However, its utilization is limited, largely due to lack of appropriate processing strategies. Various strategies have been assessed to regenerate cocoon SF, as well as the use of aqueous liquid fibroin (LFaq) prepared by dissolution of silk dope obtained from the silk glands of mature silkworm larvae in water. However, films cast from these fibroin solutions in water or organic solvents are often water-soluble and require post-treatment to render them water-stable. Here, we present a strategy for fabrication of water-stable films from S. c. ricini silk gland fibroin (SGF) without post-treatment. Aqueous ethanol induced gelation of fibroin in the posterior silk glands (PSG), enabling its separation from the rest of the silk gland. When dissolved in 1,1,1,3,3,3-hexafluoro-2-propanol (HFIP), the SGF-gel gave a solution from which a transparent, flexible, and water-insoluble film (SGFHFIP) was cast. Detailed structural characterization of the SGFHFIP as-cast film was carried out and compared to a conventional, water-soluble film cast from LFaq. FTIR and 13C solid-state NMR analyses revealed both cast films to be α-helix-rich. However, gelation of SGF induced by the 40%-EtOH-treatment resulted in an imperfect β-sheet structure. As a result, the SGF-gel was soluble in HFIP, but some β-sheet structural memory remains, and the SGFHFIP as-cast film obtained has some β-sheet content which renders it water-resistant. These results reveal a structure water-solubility relationship in S. c. ricini SF films that may offer useful insights towards tunable fabrication of novel biomaterials. A plausible model of the mechanism that leads to the difference in water resistance of the two kinds of α-helix-rich films is proposed.

A Facile Measurement for Monitoring Dragline Silk Dope Concentration in Nephila pilipes upon Spinning.

In spite of all the efforts towards deciphering the silk spinning process of spiders, the underlying mechanism is yet to be fully revealed. In this research, we designed a novel approach that allowed us to quantitatively evaluate the concentration change of silk dope during the liquid-to-solid spinning process of the orb-weaver Nephila pilipes. As a prior characterization of the optimal silking conditions, we first gauged the influence of silking-rate, ranging from 1.5 to 8.0 m/min, on dragline silk diameters and silk tensile strengths obtained from the spiders. Next, to evaluate the liquid content of the silk dope, the major ampullate gland was dissected and the concentration of the sac portion was measured by thermogravimetric analysis (TGA). The solid content of the dragline fibers leaving the spinneret was investigated by calculating the ratio of collected dried silk to the weight loss of the spider recorded in situ upon spinning. As the results indicate, the tensile strength and diameter of the spun dragline fibers were 800–1100 MPa and 8–11 μm, respectively. The liquid content of silk stored in the major ampullate sac (50.0 wt%) was significantly lower than that of silk leaving the spinnerets (80.9–96.1 wt%), indicating that a liquid supplying mechanism might be involved during the spinning process. This reveals, for the first time, quantitative evidence in support of the lubricative hypothesis proposed formerly, namely that a liquid coating layer is supplemented to compensate for silking resistance during the spinning process of a spider. The spigot, at the exit of the spinneret, is speculated to serve as a valve-like controller that regulates the lubrication process along with fiber formation. Taken together, these findings provide understanding of the physiological functions in the spider spinning process and could further shed some light on the future biomimetic development of silk material fabrication.

Combination of Amorphous Silk Fiber Spinning and Postspinning Crystallization for Tough Regenerated Silk Fibers.

An artificial spinning system using regenerated silk fibroin solutions is adopted to produce high-performance silk fibers. In previous studies, alcohol-based agents, such as methanol or ethanol, were used to coagulate silk dope solutions, producing silk fiber with poor mechanical properties compared with those of native silk fibers. The alcohol-based coagulation agents induce rapid β-sheet crystallization of the silk molecules, which inhibits subsequent alignment of the β-sheet crystals. Here, we induce gradual β-sheet formation to afford adequate β-sheet alignment similar to that of native silk fiber. To this aim, we developed an amorphous silk fiber spinning process that prevents fast β-sheet formation in silk molecules by using tetrahydrofuran (THF) as a coagulation solvent. In addition, we apply postdrawing to the predominantly amorphous silk fibers to induce β-sheet formation and orientation. The resultant silk fibers showed a 2.5-fold higher extensibility, resulting in 1.5-fold tougher silk fibers compared with native Bombyx mori silk fiber. The amorphous silk fiber spinning process developed here will pave the way to the production of silk fibers with desired mechanical properties.

Injectable and release-controlled hydrogel based on xanthan gum and silk fibroin

The natural process of silk spinning covers a fascinating versatility of aggregate states, ranging from colloidal solutions through hydrogels to solid systems. The transition among these states is controlled by a carefully orchestrated process in vivo. Major players within the natural process include the control of spatial pH throughout passage of the silk dope, the composition and type of ions, and fluid flow mechanics within the duct, respectively. The function of these input parameters on the spinning process is reviewed before detailing their impact on the design and manufacture of silk based drug delivery systems (DDS). Examples are reported including the control of hydrogel formation during storage or significant parameters controlling precipitation in the presence of appropriate salts, respectively. The review details the use of silk fibroin (SF) to develop liquid, semiliquid or solid DDS with a focus on the control of SF crystallization, particle formation, and drug–SF interaction for tailored drug load.

Biomimetic spinning of artificial spider silk from a chimeric minispidroin.

Herein we present a chimeric recombinant spider silk protein (spidroin) whose aqueous solubility equals that of native spider silk dope and a spinning device that is based solely on aqueous buffers, shear forces and lowered pH. The process recapitulates the complex molecular mechanisms that dictate native spider silk spinning and is highly efficient; spidroin from one liter of bacterial shake-flask culture is enough to spin a kilometer of the hitherto toughest as-spun artificial spider silk fiber.

Linking naturally and unnaturally spun silks through the forced reeling of Bombyx mori

The forced reeling of silkworms offers the potential to produce a spectrum of silk filaments, spun from natural silk dope and subjected to carefully controlled applied processing conditions. Here we demonstrate that the envelope of stress–strain properties for forced reeled silks can encompass both naturally spun cocoon silk and unnaturally processed artificial silk filaments. We use dynamic mechanical thermal analysis (DMTA) to quantify the structural properties of these silks. Using this well-established mechanical spectroscopic technique, we show high variation in the mechanical properties and the associated degree of disordered hydrogen-bonded structures in forced reeled silks. Furthermore, we show that this disorder can be manipulated by a range of processing conditions and even ameliorated under certain parameters, such as annealing under heat and mechanical load. We conclude that the powerful combination of forced reeling silk and DMTA has tied together native/natural and synthetic/unnatural extrusion spinning. The presented techniques therefore have the ability to define the potential of Bombyx-derived proteins for use in fibre-based applications and serve as a roadmap to improve fibre quality via post-processing.

Effect of Sodium Carbonate Concentrations on the Formation and Mechanism of Regenerated Silk Fibroin Nanofibers by Electrospinning

Degumming is the first process for the preparation of all silk‐based products. In this paper, effect of sodium carbonate concentrations for silk degumming on the formation of electrospun silk fibroin nanofibers was investigated and the reason for the silk electrospinning process was explained for the first time by differences from the microstructure of regenerated silk fibroin. With increasing the sodium carbonate concentration, microstructure both in the aqueous solutions and in the electrospinning solutions transformed from nanofibrils to nanoparticles, leading to obvious changes on rheological property; electrospinning solutions with nanofibrils behaved like the native silk dope and owned remarkably higher viscosity than the solutions with nanoparticles showing very low viscosity. More interestingly, nanofibrils favored the formation of silk nanofibers with ease, and even nanofibers could be electrospun at concentration 2%. However, nanoparticles were completely unable to generate nanofibers at high spinning concentration 8%. Importance of sodium carbonate concentrations is heavily emphasized for impacting the microstructure types and further influencing the electrospinning performance of regenerated silk. Hence, sodium carbonate concentrations provide a controllable choice for the preparation of silk‐based electrospun biomaterials with desired properties.

Effect of molecular weight and concentration on crystallinity and post drawing of wet spun silk fibroin fiber

Wet spun silk fibroin (SF) filaments have attracted considerable attention because of their potential in biotechnological applications including surgical sutures, tissue engineering and wound dressing. Although the molecular weight (MW) of polymers is one of key factors affecting the wet spinnability of dope along with the structural characteristics and properties of wet spun filament, no related study has been conducted. In this study, regenerated SFs with different MWs and concentrations were prepared by wet spinning. The effects of the SF concentration and MW on 1) wet spinnability and rheology of silk dope solution and 2) crystallinity index and post drawing performance of wet spun silk filament were examined. Their relationships were also investigated. The rheological measurements showed that an 80 mPa·s viscosity is needed to obtain a continuous wet spun SF filament. As the MW of SF increased, the peak position of the maximum draw ratio shifted to a lower SF concentration with a concomitant increase in the maximum draw ratio value at the peak. Interestingly, the crystallinity index obtained from Fourier transform infrared spectroscopy (FTIR) revealed a similar trend to the maximum draw ratio suggesting that the post drawing ability is strongly affected by the quantity of short-ordered crystalline regions in wet spun SF filaments. On the other hand, X-ray diffraction did not detect any crystallinity change in the SF filament produced from the formic acid solvent system. It was concluded that MW strongly affected the dope solution viscosity and the crystallinity index from FTIR and these determined the fiber formation of dope and post drawing performance of fiber.

Effects of degumming conditions on electro-spinning rate of regenerated silk

Electro-spun silk webs are potentially good candidates as tissue engineering scaffolds owing to their good bio- and cyto-compatibility. However, the low fabrication rate of electro-spun silk mats has been one of the obstacles in the mass production of such nanofibrous silk mats in applications to the biomedical field. In this study, the effects of degumming ratio and silk concentration on the electro-spinning process were investigated by using regenerated silk with different residual sericin contents and different silk concentrations in terms of the morphology and structure of the electro-spun silk web. The rate of production of electro-spun silk mats could be increased by approximately 5 fold at a degumming ratio of 19.5%. The electro-spinning rate of silk was affected by two main factors: (1) dope solution viscosity and (2) degumming ratio of silk. The conductivity of the silk dope solution, however, had little effects on the electro-spinning of regenerated silk. A constant spun fiber morphology was observed within the electro-spinning rate range (0.3–1.4ml/h). Fourier transform infrared spectroscopy showed that partial β-sheet crystallization occurred during electro-spinning. The molecular conformation was relatively unaffected by the electro-spinning rate of silk.

Silks: Properties and uses of natural and designed variants

Silks: Properties and uses of natural and designed variants

Structure of silk by raman spectromicroscopy: From the spinning glands to the fibers

Raman spectroscopy has long been proved to be a useful tool to study the conformation of protein-based materials such as silk. Thanks to recent developments, linearly polarized Raman spectromicroscopy has appeared very efficient to characterize the molecular structure of native single silk fibers and spinning dopes because it can provide information relative to the protein secondary structure, molecular orientation, and amino acid composition. This review will describe recent advances in the study of the structure of silk by Raman spectromicroscopy. A particular emphasis is put on the spider dragline and silkworm cocoon threads, other fibers spun by orb-weaving spiders, the spinning dope contained in their silk glands and the effect of mechanical deformation. Taken together, the results of the literature show that Raman spectromicroscopy is particularly efficient to investigate all aspects of silk structure and production. The data provided can lead to a better understanding of the structure of the silk dope, transformations occurring during the spinning process, and structure and mechanical properties of native fibers.

Plasticity in major ampullate silk production in relation to spider phylogeny and ecology.

Spider major ampullate silk is a high-performance biomaterial that has received much attention. However, most studies ignore plasticity in silk properties. A better understanding of silk plasticity could clarify the relative importance of chemical composition versus processing of silk dope for silk properties. It could also provide insight into how control of silk properties relates to spider ecology and silk uses.We compared silk plasticity (defined as variation in the properties of silk spun by a spider under different conditions) between three spider clades in relation to their anatomy and silk biochemistry. We found that silk plasticity exists in RTA clade and orbicularian spiders, two clades that differ in their silk biochemistry. Orbiculariae seem less dependent on external spinning conditions. They probably use a valve in their spinning duct to control friction forces and speed during spinning. Our results suggest that plasticity results from different processing of the silk dope in the spinning duct. Orbicularian spiders seem to display better control of silk properties, perhaps in relation to their more complex spinning duct valve.

Folding process of silk fibroin induced by ferric and ferrous ions

Bombyx mori silk fiber has useful mechanical properties largely due to a high content of ordered β-sheet crystallites separated by non-crystalline spacers. Metallic ions present in the silk dope in nature could affect the β-sheet content. In this work, we used solid-state 13C NMR, EPR and Raman spectroscopy to investigate how the ferric/ferrous ions affect the folding process of the silk fibroin. NMR and Raman results indicate that ferric and ferrous ions have different effects on the secondary structure of silk fibroin. Ferric ions can induce a conformation change from helix to β-sheet form in silk fibroin when their concentration exceeds a critical value, while ferrous ions cannot. EPR results indicate that the ferric ions bound with silk fibroin have a high-spin state ( S = 5/2) with g-value of g 1 = 1.950, g 2 = 1.990 and g 3 = 1.995, zero-field splitting interaction D of 1.2–2 cm −1, and symmetric character of E/ D = 1/3, resulting in an effective g-value of g′ = 4.25. The hydrophilic spacer GTGSSGFGPYVAN(H)GGYSGYEYAWSSESDFGT in the heavy chain of silk fibroin is likely to be involved in the binding of ferric ions, and His, Asn and Tyr residues are considered as the potential binding sites.

Natural and unnatural silks

Natural silk is an important biopolymer with huge potential as it combines superb mechanical properties with environmentally sensitive production methods. Native silk dope taken straight from the gland can easily and without chemical assistance be drawn into strong fibres. Artificial silk fibres, on the other hand, rely on spinning dopes typically ‘reconstituted’ from natural silk fibres by strong chaotropic agents. Such fibres do not form readily, and often require chemical post-spin treatment for stabilisation. In addition these fibres tend to be brittle, and so far have been unable to match native fibres. Here we present novel rheometric data to argue that native and reconstituted silkworm silk dope differ in kind, not just in degree. While native silks behave like typical molten polymers, reconstituted silks do not. We conclude that rheology provides a powerful tool in the quest to learn from the Nature's polymer fibre technology.