Production of High Performance Bioinspired Silk Fibers by Straining

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Production of high performance bioinspired silk fibers by Straining Flow Spinning Rodrigo Madurga, Alfonso M. Ganan-Calvo, Gustavo Ramón Plaza, Gustavo V. Guinea, Manuel Elices, and José PÉREZ-RIGUEIRO Biomacromolecules, Just Accepted Manuscript • DOI: 10.1021/acs.biomac.6b01757 • Publication Date (Web): 22 Feb 2017 Downloaded from http://pubs.acs.org on February 24, 2017

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Production of high performance bioinspired silk fibers by Straining Flow Spinning Rodrigo Madurga1,2, Alfonso M. Gañán-Calvo3, Gustavo R. Plaza1,2, Gustavo. V. Guinea1,2,4, Manuel Elices1,2, José Pérez-Rigueiro1,2,4* 1. Centro de Tecnología Biomédica. Universidad Politécnica de Madrid. 28223 Pozuelo de Alarcón (Madrid). Spain 2. Departamento de Ciencia de Materiales. ETSI Caminos, Canales y Puertos. Universidad Politécnica de Madrid. 28040. Madrid. Spain 3. Escuela Técnica Superior de Ingenieros. Universidad de Sevilla. 41092. Sevilla. Spain 4. Biomedical Research Networking Center in Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN), Madrid, Spain

*Corresponding author: [email protected] KEYWORDS: Regenerated fibers, fibroin, biomimetic, silk.

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ABSTRACT: In the last years there has been an increasing interest on bioinspired approaches for different applications, including the spinning of high performance silk fibers. Bioinspired spinning is based on the natural spinning system of spiders and worms and requires combining changes in the chemical environment of the proteins with the application of mechanical stresses. Here we present the novel Straining Flow Spinning (SFS) process and prove its ability to produce high performance fibers under mild, environmentally friendly conditions, from aqueous protein dopes. SFS is shown to be an extremely versatile technique which allows controlling a large number of processing parameters. This ample set of parameters allows fine-tuning the microstructure and mechanical behaviour of the fibers, which opens the possibility of adapting the fibers to their intended uses.


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Introduction Spinning of artificial silk fibers has constituted a challenge for the last two decades due to the difficulty of translating to the laboratory the subtle and versatile mechanisms with which Nature has endowed spinning glandular systems of spiders, silkworms and other arthropods. Materials scientists and engineers look with interest at silk fibers due to their extraordinary mechanical properties1, since the combination of high strain at breaking and tensile strength yields values of toughness that outperform those of high performance fibers (i.e. fibers with a work to fracture equal or higher than 50 MJ/m3) such as Nylon, Kevlar and natural silkworm silk2. Silkworm silk and spider silk have yet another interesting characteristic, since their structural proteins were found to be biocompatible3, so that they do not generate an inflammatory or allergenic response in the human body. Consequently, silk has arisen as one of the most promising materials over the last years. Unfortunately, the use of natural silk fibers is hampered by either scarcity (spider silks) or by the requirement of aggressive treatments that degrade the properties of the material4 (silkworm silk). Attempts for producing silk-based fibers have found that the production of high performance fibers is much more complicated than it was thought when proposed for the first time5. All spinning processes share a common principle by which the protein is initially dissolved in an adequate solvent to make the dope, and it is subsequently solidified through the interaction of the dope with a different medium. In this regard, the simplest spinning procedure is wet spinning, in which the dope enters into a coagulating bath that removes the solvent and leads to the formation of the fiber. Different authors have used alternative combinations for the fibroin solvent/coagulating bath couple, such as concentrated aqueous solution/saturated ammonium5, hexafluoroisopropanol/methanol6, 3 ACS Paragon Plus Environment

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hexaflluoroacetone hydrate/methanol7, formic acid/methanol8-10 . It has also been shown that the results can be improved by adding CaCl2 to the dope11. In particular, the usage of high concentration of fibroin in the dope has allowed spinning fibers with tensile properties comparable to those of natural silkworm silk12. Beside the conventional spinning techniques, an increasing knowledge of the basics of natural spinning has led to the development of biomimetic spinning processes. Fibroins accumulate in the silk glands at very high concentrations, despite their tendency to selfassembly13, 14. It is thought that a neutral pH and controlled ion concentration (Na+, K+, Ca2+ and Cl-) are essential to keep the fibroin as a soluble moiety15. Proteins might remain soluble due to their organization as liquid crystals16, as micelles17 or as a combination of both phases18. The acidification of the protein solution along the spinning duct19 prompts the self-assembly of the proteins into a new structure that can solidify after being subjected to mechanical shear stresses1, 20, 21. In this context, biomimetic spinning processes intend to modify the local environment of the dope by using a combination of coagulating fluids18, very often relying on microfluidic devices22. Here we present a new bioinspired approach for the spinning of regenerated silkworm silk fibers, called straining flow spinning (SFS)23. This new approach is inspired in the flow focusing technology24, that controls the flow of a fluid (in this case the dope) through its interaction with a second flowing fluid (focusing fluid)25, 26. As discussed below, SFS is shown to be a robust and versatile spinning technology that, in contrast to other existing methods, mimics the balanced conjunction of a mild chemical environment (ordinarily aqueous) and moderate shear forces found in natural spinning glands. As it will be shown below, the novel SFS procedure allows the spinning of high-performance regenerated silk 4 ACS Paragon Plus Environment

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fibers from aqueous dopes within a vast range of microstructures and tensile properties, and will help to the development of a new generation of high performance biomimetic fibers. Experimental procedure Preparation of the dope Silkworm (Bombyx mori) silk cocoons were degummed with water, in a ratio 1/50 (w/v), at 121oC during 50 minutes using an autoclave. The degummed silk was then dissolved to a concentration of 10% (w/v) in a 9.4 M LiBr solution at 60oC for 4 hours. Solving was followed by a dialysis step against deionized water using commercial Slide-A-Lyzer dialysis cassettes, with a molecular cut-off of 3.5 kDa, to remove the salts. The dialysis was performed for 3 days and water was changed every 8 hours. Removal of the salt was checked by measuring the electrical conductivity of water. The dialyzed fibroin solution was centrifuged (20 minutes at 5000 rpm) to remove any debris. CaCl2 was added to a final concentration of 1 M and centrifuged again under the same conditions. Finally, the solutions were concentrated by reverse dialysis for 15 hours. The reverse dialysis medium consisted of a PEG 8000 Da solution with CaCl2 1 M. PEG concentration was selected depending on the desired final fibroin concentration: for a fibroin final concentration of 8%, the PEG concentration was 10%; for a final fibroin concentration of 16%, PEG concentration was 20%. It was found that fibroin proteins are cleaved during the process27, and the average molecular weight of the proteins in the dope was found to be 90 kDa, as measured by polyacrylamide gel electrophoresis. Straining flow spinning setup


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The basic elements of a straining flow spinning system (SFS) are shown in Figure 1a. The experimental setup consists of: 1) two syringe pumps (Harvard Apparatus 11 Plus), one was equipped with a Becton-Dickinson 1 ml syringe for the dope and the other with a Becton-Dickinson 60 ml syringe for the focusing fluid. Both syringes had Luer-Lock connections. 2) The dope syringe was connected to a 1/16 FEP tube that ended in a fitting to the capillary with inner diameter of 150 µm. 3) This capillary crosses a PEEK tee connection with an inner diameter of 1.25 mm that connects the focusing syringe through a 1/16 FEP tube to the nozzle. The tubes and the capillary are connected so that the inner diameter of the dope circuit can decrease but never increase. The capillary remains aligned coaxially inside the nozzle with a copper coiled spring. All the tubing and connections were provided by IDEX Health and Science LLC. Spinning parameters The geometrical parameters were fixed to: diameter of the orifice of nozzle, D1 ~ 400 µm, inner diameter of the capillary, d1 = 150 µm, tapering angle at the end of the capillary, α = 90o, distance between the end of the capillary and the nozzle outlet, H = 3500 µm and distance between the take-up and the post-spinning drawing rollers, LPS = 60 cm. Two different dope compositions were chosen that differed in the fibroin concentration. Dopes were prepared in a 1 M CaCl2 solution and fibroin concentration of either 8 or 16% (w/v). Three different compositions were tested as coagulants: a mixture of ethanol and acetic acid 1 M in a ratio 80:20, a mixture of isopropanol and acetic acid 1 M in a ratio 80:20, and a PEG 8000 solution in water with a polymer concentration of 30% (w/v). In all cases, the coagulating bath had the same composition as the focusing fluid. 6 ACS Paragon Plus Environment

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Figure 1. Scheme of the straining flow spinning process and fibers spun with this technique. (a) Schematic representation of the straining flow spinning technique including its main elements. The inset shows a detail of the capillary-nozzle system. (b), Image of the dope jet exiting the capillary. The outlet of the nozzle would be located on the right out of


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the field of the micrograph. (c), Representative SEM images of the fracture of SFS spun fibers. Scale bar = 2 µm. The hydrodynamic parameters comprise Qd, Qf and VR1: flow rate of the dope, flow rate of the focusing fluid and take-up roller velocity, respectively. When applied, the post-spinning roller velocity VR2 must also be considered. The flow rate of the dope was fixed in all experiments to Qd= 5 µl/min. The flow rate of the focusing fluid, Qf, has a strong influence in the production speed of the dope due to the acceleration that it exerts in the dope jet. Therefore, the focusing fluid was adjusted in each experiment so that the spinning was performed continuously, i.e. without either breaking the fiber or being accumulated in the coagulating bath. The take-up roller velocity was fixed in all experiments to VR1=3 m/min. In the experiments that included the post-spinning drawing step, the velocity of the postspinning roller was adjusted to the maximum velocity that did not induce the breaking of the fiber. The four different combinations of spinning conditions discussed in this work are summarized in Table 1. Table 1. Values of the parameters for the four spinning processes presented in this work.

Code

[SF] (%)

Coagulant

Qf (ml/min)

VR2 (m/min)

DR

8-PEG 8 PEG 30% 0.5 16-Et 16 Ethanol:Acetic acid 1M (80:20) 2.5 16-Et-2.4X 16 Ethanol:Acetic acid 1M (80:20) 2.5 7.3 2.4 16-Iso-3.5X 16 Isopropanol:Acetic acid 1M (80:20) 0.6 10.6 3.5 Silk fibroin concentration ([SF]), focusing fluid flow rate (Qf), velocity of the postspinning roller (VR2) and drawing ratio (DR) defined as the ratio between the velocity of the post-spinning roller, VR2, and that of the take-up roller, VR1. The spinning parameters that are not included in this table were maintained constant as explained in the experimental section.


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Characterization of the regenerated fibers The microstructure of the regenerated silkworm silk fibers was analysed by infrared spectroscopy. Spectra were obtained using a Nicolet iS5 spectrometer equipped with an iD5 ATR accessory. Prior to the measurement, a background spectrum was obtained by applying a constant pressure to the clean diamond crystal and, subsequently, the background spectrum was subtracted from the spectrum of the sample. Each IR spectrum was obtained from a bundle of fibers folded into a small ball and pressed against the diamond window at constant pressure. All spectra were the result of averaging 64 measurements in the range from 550 to 4000 cm-1 with a resolution of 4 cm-1. Samples were mounted on aluminium foil frames with a base length of 20 mm and fixed with ethylcyanocrylate for tensile testing. Before starting the tensile test, the diameter of the fibers was measured with an optical microscope (Leica DMI 3000B, objective magnification of 40X). Then the aluminium frame was fixed to the upper and lower grips of an Instron 4411 tensile testing machine. The lower grip rested on a precision balance (Precisa XT220A, resolution 0.1 mg) that was used for measuring force as described elsewhere2, 28. After being mounted in the testing machine and determining the zero load point (the point in which the fiber is tight but no force is exerted on it), the initial length of the fiber, L0, was measured with a calliper. Tensile tests were performed at a constant speed of 1 mm/min either in air (nominal environmental conditions of 25oC and 35% relative humidity) or immersed in water at 25 ºC. Stresses were calculated from the diameter measured from the optical micrographs considering a circular cross-sectional area.


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Assuming that fiber volume remains constant during stretching, as reported for spider silks29, true stress, σ, and true strain, ε, were calculated as:

ε = Ln

σ=

L L0

F L =F A A0 L0

where A0 and L0 are, respectively, the initial area and length of the sample, A and L are their instantaneous values and F the applied force. To obtain a more detailed analysis of the surface and morphology of the material, some fibers were observed with a field emission scanning electron microscope (FESEM), Zeiss Auriga, at 5 kV with a current of 0.06 nA. The samples were fixed to the sample holders with double-sided carbon tape and metallized with gold before the observation. Wet-stretching, supercontraction and recovery Wet stretching is a simple procedure for controlling and enhancing the mechanical performance of silk fibers developed by the authors30, 31. It, consists of stretching a fiber immersed in water up to a selected strain, then allowing it to relax and –after fixing its ends– let it dry overnight . In this work samples were wet-stretched up to a value of 80% the strain at breaking in water. The existence of supercontraction was assessed through recovery tests32, in which a fiber was strained in air up to a given deformation and then allowed to contract freely in water and to dry overnight. Subsequently, the fiber was stretched again in air up to a higher value


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of strain and then immersed in water for free contraction. Three loading-unloading steps were applied to the fibers up to true strains of 10%, 18% and 26%. Results Straining Flow Spinning The SFS process (Figure 1), which is introduced in this work, represents an extremely versatile spinning technique that is especially suited –but not limited– to the production of silk-based fibers, where a delicate equilibrium between chemical changes and mechanical stretching must be achieved. By its configuration, SFS mimics the two basic processes that simultaneously occur along the duct during natural spinning in spiders and worms, namely elongational flow and a continuous modification of the chemistry of the dope. SFS allows controlling a wide range of processing parameters, and an initial screening (data not shown) established the feasibility of SFS for producing fibers under a broad range of conditions. This initial screening also showed the robustness of the technique, since it was found that by optimizing the processing conditions up to 1000 m could be continuously spun. In this work, the length was limited by the maximum volume of focusing fluid used (60 ml). SFS processing parameters can be grouped in three categories: geometrical, hydrodynamic and chemical, and they control the mechanisms that lead to the initial formation steps of the fiber in the region between the capillary tip and the nozzle (inset in Figure 1a). One of the key points of SFS is the control exerted on the dope jet by the convergent axisymmetric coflow of the focusing fluid, which produces a pressure drop that pulls the dope and induces its straining (Figure 1b). This feature is shared with the flow focusing technique24, on which the SFS process is inspired. 11 ACS Paragon Plus Environment

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The chemical environment of the dope is modified through its interaction with the focusing fluid inside the nozzle and, additionally, it may be further modified through the interaction of both fluids with the coagulating bath. Chemical modifications of the dope may consist of variations in the pH, removal of water molecules, ion exchange between the dope and the other fluids or a combination of all of these mechanisms. In concurrence with the chemical modifications, shear stresses are initially induced by the elongational flow transmitted by the focusing fluid to the dope, and can be controlled by the geometry of the capillary-nozzle system and flow rates of the dope and the focusing fluid. Additional stresses are induced on the dope when traversing the nozzle outlet. In contrast to other spinning techniques, SFS allows a significant uncoupling between the instantaneous chemical environment of the dope and the stresses to which the dope proteins are subjected. As a first demonstration of its full potential, the SFS technique has been applied to the spinning of high performance regenerated silkworm silk fibers. Based on an initial screening, silk protein concentrations of 8% and 16% were used for the production of high performance fibers. CaCl2 was added, since calcium ions had been reported to exert a stabilizing effect on the native fibroin solution33 and help increasing its solubility in water. The chemistry of the focusing fluids (which in this work always coincides with the composition of the coagulating bath) also relied on the previous screening and comprises three different compositions: Polyethylene glycol (PEG) 30% (w/v), as representative of the group of purely dehydrating fluids, and ethanol-acetic acid and isopropanol-acetic acid blends, as representatives of the acidifying coagulants (although a dehydrating effect due to the presence of the alcohols is also expected). In all cases, the use of harsh chemicals, such


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as hexafluoroisopropanol or methanol, was avoided. Additionally, the possibility of subjecting the fibers to a post-spinning drawing in air during spinning was included. The four spinning conditions of the SFS processes presented in this work are summarized in Table 1. The processes differ in (1) the concentration of silk proteins in the dope, (2) the composition of the focusing fluid and (3) in the inclusion or not of an in-air post-spinning drawing step. The geometrical and hydrodynamic parameters were kept constant in all the processes. These conditions were selected to demonstrate the versatility of the SFS technique, since it allows controlling the microstructure and the properties of fibers obtained from the same protein dope under very different processing conditions. In the following, morphology, microstructure and mechanical performance of the spun fibers are reviewed. Fiber morphology Figure 1c shows representative images of the fibers obtained. In all cases, regenerated silk fibers could be spun continuously. Diameters in the range 8-13 microns depending on the processing parameters were obtained with high reproducibility, which is a pre-requisite for any spinning technique. For each spinning conditions the diameters, D, measured were: 8PEG, D=12.2 ± 0.8 µm; 16-Et, D=10.7± 0.2 µm; 16-Et-2.4X, D=8.4 ± 0.7 µm and 16-Iso3.5X, D= 9.0 ± 0.5 µm. Small irregularities were found in 8-PEG samples and are supposed to be due to the presence of PEG adherences on the surface of the fiber. Fiber microstructure


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The possibility of modifying the microstructure of the fiber by controlling the processing parameters was assessed by FTIR-ATR spectroscopy. Figure 2a compares the amide I peak of the four types of fibers analysed, since this peak contains information on the secondary structure of the proteins. A significant difference between the sample 8-PEG and the other samples is apparent. Although all samples show a maximum in the region assigned to the β-sheet structure (1621-1637 cm-1)34-36, the peak of the 8-PEG sample is slightly displaced to higher wavenumbers. This displacement has been previously assigned to a decrease in the content of aggregated strands, which absorb in the region from 1610-1620 cm-1

37, 38

. The

aggregated strands correspond to regions of the protein chains in an extended conformation and very closely aligned with neighbouring chains, so that the formation of very strong intermolecular hydrogen bonds is favoured38. The decrease in the content of aggregated strands in the PEG-8 sample might be associated with an increase in the random coil conformation, which appears as a shoulder at approximately 1647 cm-1 37.


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Figure 2. Microstructural analysis and tensile properties of the regenerated fibers. (a), Amide I peaks of the ATR-FTIR spectra of regenerated silk fibers produced by straining flow spinning. The positions of the contributions from the main secondary structures are indicated on the Figure. (b-d), Representative true stress-true strain curves for the four types of fibers (b) as-spun fibers tested in air (the inset shows the detail of the curves at low values of strain), (c) as spun fibers tested in water and (d) wet stretched fibers tested in air. All tests in air were performed under nominal conditions, T=25oC and RH=40%. The breaking point of each sample is indicated on the Figure. Samples are identified as: 8-PEG


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(blue curve), 16-Et (solid black curve), 16-Et-2.4X (dashed black curve) and 16-Iso-3.5X (dashed red curve). Mechanical properties of the regenerated fibers The tensile characterization of the fibers further illustrates the wide range of properties that can be exhibited by fibers produced with the SFS technique by varying the processing parameters. Figure 2b shows the true stress- true strain curves of the fibers tested in air. Large differences are observed both in the overall shape of the curves and in the values of the main mechanical parameters. Thus, PEG-8 samples show a large value of strain at breaking, ε~1.3, that is comparable to that of maximum supercontracted spider dragline silk39. However, and in contrast to the values of the tensile strength reached by natural silkworm silk (≈ 500 MPa), these samples reach modest values of tensile strength (σ~80 MPa). The combination of the tensile strength and strain at breaking in PEG-8 samples yields a value of work to fracture of Wf~45 MJ/m3, which is comparable to that of natural silkworm silk27 and to the authors’ knowledge, is the highest value of toughness reported for a regenerated silk fiber without post-spinning drawing. It is worth stressing that the comparable values of work to fracture found in natural silk and PEG-8 regenerated samples are the result of alternative combinations of the values of tensile strength and strain at breaking. From Figure 2b it is also apparent that fibers coagulated using alcohols may show very different tensile behaviours despite of their concurring infrared spectra. Samples 16-Et and 16-Et-2.4X show a brittle behaviour (inset in Figure 2b) when tested in air, while samples


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16-Iso-3.5X show a ductile behaviour with values of tensile strength and strain at breaking of ε~0,4 and σ~120 MPa, that yield a value of work to fracture of Wf~32 MJ/m3. Previous results on regenerated fibers40 have shown that the properties of brittle fibers, such as Et-16 and 16-Et-2.4X samples, may be improved by stretching the fibers in water (i.e. after being subjected to a wet stretching process). Although the ultimate origin of this enhanced behavior is not clear, it is assumed to be related with an increased conformational freedom of the protein chains. Conformational freedom would be increased as a result of the reduction of the number of entanglements between the chains41 that occurs when the chains undergo relative displacements when stretched in water. This increased conformational freedom, in turn, would allow the re-organization of the proteins upon subsequent stretching as shown from the behavior of electrospun regenerated silk fibers42. From an experimental point of view a first hint to the feasibility of this treatment is provided by a significant increase in the strain at breaking of the fibers when tested in water. The tensile properties of the fibers tested in water are shown in Figure 2c, where a large increase in ductility of samples Et-16 and 16-Et-2.4X is noticed. The results obtained from tensile tests in water suggested that the properties of the fibers might be modified through a wet-stretching process, as described in the Methods Section. The true stress-true strain curves of the wet-stretched fibers are shown in Figure 2d. The tensile properties of the as spun initially brittle fibers Et-16 and 16-Et-2.4X are improved considerably, both in stresses and strains at breaking, yielding values of work to fracture of Wf~60 MJ/m3. In contrast, modest variations are observed in the 16-Iso-3.5X and 8-PEG samples. The main tensile parameters of fibers are summarized in Table 2.


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Table 2. Mechanical parameters of the different samples including natural silkworm silk for comparison. Number of samples (n), elastic modulus (E), tensile strength (σu), strain at breaking (εu) and work to fracture (Wf). The values represent the mean ± standard deviation.

Fiber

n

σu (MPa)

Natural silkworm silk27 8-PEG 8-PEG (Wet-stretched) 16-Et 16-Et (Wet-stretched) 16-Et-2.4X 16-Et-2.4X (Wet-stretched) 16-Iso-3.5X 16-Iso-3.5X (Wet-stretched)

4 2 5 2 5 3 5 3

500 ± 30 86 ± 8 101 ± 8 52 ± 2 190 ± 20 70 ± 6 250 ± 20 112 ± 11 330 ± 20

Ei (GPa)

Wf (MJ/m3)

15 ± 3 15 ± 2 124 ± 5 1.0 ± 0.3 17 ± 3 3±2 1.3 ± 0.1 4.1 ± 0.1 50 ± 10 4±1 1.70 ± 0.01 4.1 ± 0.1 30 ± 10 9±4 37 ± 4 4.3 ± 0.1 22 ± 3 11 ± 3

60 ± 7 37 ± 4 13 ± 3 0.36 ± 0.03 60 ± 20 0.6 ± 0.1 60 ± 20 32 ± 5 50 ± 10

εu (%)

Lastly, the feasibility of SFS for producing fibers that show supercontraction and are able to recover their mechanical properties after an arbitrary loading history was assessed. Supercontraction43 is a characteristic feature of spider silk which implies the existence of a ground state to which the material can revert upon immersion in water. Despite natural silkworm silk does not exhibit supercontraction, it was found that this potentially useful property could be imparted to regenerated silkworm silk fibers40. Figures 3a and 3b show the true stress-true strain curves for wet stretched samples 16-Iso-3.5X and 16-Et-2.4X tested in air after being subjected to repeated cycles of stretching in air and free contraction in water, as explained in the Methods Section. Both types of fibers are shown to recover their initial mechanical properties after every recovery step, which is the defining property of supercontraction.


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Figure 3. Recovery tests of regenerated fibers. (a) Recovery of samples 16-Iso-3.5X. (b) Recovery of samples 16-Et-2.4X. Solid and dashed lines in each plot are used to distinguish between two different fibers spun under the same processing conditions. Black curves are for the first cycle, blue curves for the second cycle, red curves for the third cycle and green curves for the fourth cycle. The light blue curve at zero stress indicates immersion in water allowing the fiber to supercontract. The fibers were allowed to dry overnight between subsequent cycles. Conclusion The results presented in this work show that SFS allows spinning under a wide range of conditions in terms of the geometric, hydrodynamic and chemical parameters that define the process, which represents a uniquely ample set of degrees of freedom. Besides, the microstructure and tensile properties of the fibers can be tuned by controlling the processing parameters. In particular, it was found that SFS allows generating high performance regenerated silk fibers with different secondary structures. It is shown that


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high performance can be obtained both in as spun fibers (even without a post-spinning drawing step) or, alternatively, in fibers after being subjected to a wet stretching process. Lastly, supercontracting regenerated silkworm silk fibers can be easily produced with this technique. In conclusion, SFS appears as an extremely versatile technique that allows producing high performance silk-based fibers with varying microstructures and tensile properties under very mild processing conditions. Besides, the principles of the technique allow its scaling up to an industrial level. The possibility of obtaining fibers within a wide range of properties opens the possibility of adapting the material to its intended use, akin to as spiders proceed when spinning silk fibers in nature.


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ACKNOWLEDGMENTS Straining Flow Spinning was developed in Universidad Politécnica de Madrid and Universidad de Sevilla with the assistance of Ingeniatrics Tecnologías S.L. (Sevilla, Spain), to which the SFS technology is licenced.

ABBREVIATIONS SFS, Straining Flow Spinning; PEG, Polyethylene glycol; Et, Ethanol; Iso, Isopropanol.


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Author Contributions R.M. performed the experimental work. G.V.G., G.R.P. and M.E. contributed to the data analysis and reviewed the manuscript. J.P.-R. and A.M.G.-C. proposed the experiments. The main text was written by R.M. and J.P.-R.


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Funding Sources The work was funded by Ministerio de Economía y Competitividad (Spain) through project MAT2012-38412-C02-01, by Fundación Marcelino Botín and by Banco Santander through its Santander Universities Global Division.


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Conflict of interest statement Patent application of the Straining Flow Technology (SFS) was filed on December 18th, 2015. The technology is licenced to Ingeniatrics Tecnologías S.L. (Seville, Spain)


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Table of Contents


Production of High Performance Bioinspired Silk Fibers by Straining

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