Mechanics and hierarchical structure transformation mechanism of wool fibers

Abstract

Various natural protein materials have hierarchical microscale and nanoscale structures that protect animals from suffering cold weather or external threats. Herein, we contribute an effective strategy for exploring the hierarchical structure transformation mechanism by stretching a single fiber and bundle fibers. Taking advantage of controllable stretching, the disulfide bonds and peptide chains were taken apart or reconstructed by the new crosslinking bonds inside the wool fiber. If the temperature, solution concentration and stretching velocity were appropriate, the strain was more than 230% and the microstructure transformation of a single fiber underwent α→β transition, disulfide bond breakage and macromolecule slippage occurred during the tensile process. The transformation mechanism was modeled and experimentally tested in wool bundle fibers after setting, suggesting that the stretching led to the transformation from α helix to β-pleated sheet, the breakage of disulfide bonds and the slippage of microfibrils. The availability of wool fibers provides many opportunities for bio-friendly cellular substrates and biosensor devices.

Keywords

wool fibers dynamic elastic–plastic behavior microstructure transformation mechanism anisotropy stretching

Wool fibers belong to a group of proteins that are composed of cortical and cuticular scales.¹ Disulfide bonds play an important role in improving protein stability and form a hierarchical structure of wool fibers. The cuticular scales are located on the outermost part of the fiber, which protects the inner part, the cortex, from the external environment, such as acids, alkalis and oxidizing agents, and also have a layered structure. The inner section, the cortex, is a complex polymer structure of alpha keratin, which provides its longitudinal mechanical properties.² Wool fibers have been developed in many fields, such as stretching slenderization, photolithography, catalytic, self-cleaning, biomedical and pharmaceutical.^3–17 The development of regenerated wool keratin utilizes micro-nano structures through keratin extracted methods owing to its good biocompatibility, robust mechanical properties and controllability.^18–21 Since the 1930s, researchers have intensively investigated the mechanical behavior of keratin-based fibers and the relationship between the structural change and the mechanical properties. As a result of experimental work, several two-phase structure models were proposed to interpret the shape of stress–strain curves, and to explore the correlation with the fiber structure and the mechanical behavior. The structure models included the Feughelman model,^22–26 Chapman/Hearle model^27–29 and Zahn model.³⁰ The Feughelman model put forward the two-phase model and was composed of microfibrils and a matrix. With the increase of humidity, the matrix swelled but the microfibrils did not swell. After that, the series-zone structure model was proposed to interpret the α→β transition in the yield region. In 1994, the model was improved in which the stretching caused the microfibrils to be close together and jam in the yield region. In the Chapman/Hearle model, it is thought that the microfibril was stretched and the matrix stress increased when the α→β transition occurred. The Zahn model suggested that the disulfide bonds formed between the microfibrils and the increase of modulus in the post-yield region was due to the role of disulfide bonds. Therefore, the Chapman/Hearle and Zahn models were developed based on the Feughelman series-zone structure model. It is worth noting that the Chapman/Hearle model introduced the stress–strain curve of the microfibril and matrix. The Zahn model indicated that a microfibril was divided into four α-helix segments instead of two kinds of microfibrils in the Feughelman model. Besides these deformation models, the stretching and computational studies of α keratin materials have been proposed to verify the deformation. Previous studies^14,31–36 revealed that stretching must involve the α-helix→β transition at low humidity and extension of chemical bonds in the peptide chain at high humidity. Pauling et al.³⁷ demonstrated that when the secondary structure of wool fiber was completely transformed from the α-helix to β-pleated sheet, the maximum stretching rate was 109% longer than its original length. In our previous study,^34,35 the breakage and reconstruction of some chemical bonds, especially hydrogen bonds and disulfide bonds, in wool fibers had been found by Raman spectral analysis and stress relaxation. Tsobkallo et al.³⁸ investigated the structural changes during the recovery process in stretched wool fibers by a Fourier transform infrared (FTIR) method, demonstrating the reorganization of the hydrogen bonds between the microfibrils and matrix owing to the influence of water content. Paquin and Colomban³⁶ suggested a quantitative change of the α-helix→β transition and the effect of water by a Raman study. It had been found that the α keratin materials would experience conformation transition from the α helix to β pleated sheets by molecular simulation under tensile loading. Chou and Buehler³⁹ simulated the mechanics of α keratin fibers, demonstrating that the system lost α helical structures and the disulfide bonds played a significant role in the mechanical properties of α keratin. Yu et al.⁴⁰ investigated the viscoelastic properties of α keratin fibers, including stress relaxation and creep, indicating the strain-rate sensitivity of the disulfide bonds. Zhmurov et al.⁴¹ and Ding et al.⁴² characterized α helix to β pleated sheets at the molecule level using biomolecular simulations. The analysis of the deformation of wool fibers in water and in dry air have been carried out in this study in order to explore the mechanics and hierarchical structure transformation mechanism of wool fibers, including viscoelasticity and microstructure transformation.

Experimental details

Materials and stretching treatment

Materials

Merino scoured wool fibers (average fiber diameter: 22.9 µm) were supplied by Shandong Nanshan Zhishang Sci-Tech Co., Ltd, Shandong, China. The fibers were scoured with sodium sulfite, zinc acetate and potassium persulfate, provided by Yonghua Chemical Co., Ltd. All chemicals were used as received.

Stretching treatment of the single wool fiber

The single fiber was pretreated with an aqueous sodium sulfite solution with the concentrations of 1% and 5% for 0.25, 1 and 3 min⁻¹ stretching rates at 30^oC, 50^oC and 75^oC. The number of tensile tests for each sample is more than 50. The treatment of wool fibers with sodium bisulfite results in the cleavage of disulfide bonds and forms cysteine residues according to the following formula

\begin{matrix} W - S - S - W + - HS {O_{3}}^{-} \\ W - SH + W - S - S {O_{3}}^{-} \end{matrix}

(1)

Stretching treatment of the wool bundle fibers

To characterize the microstructure transformation of the wool fibers, the wool bundle fibers were pretreated in the sodium bisulfite solution with the concentration of 5% for 2 minutes and then stretched to 30%, 50%, 80% and 110% longer than their original length in a 75^oC steaming chamber by a laboratorial stretching apparatus developed by the authors.

After the stretching process, the wool fibers were set with heavy metal ions to establish a stable crosslinking between the disulfide bonds in the fiber. The reaction formula is as follows

2 W - SH + Z n^{2 +} \to W - S - Zn - S - W + 2 H^{+}

(2)

where Zn²⁺ was introduced into the disulfide bonds between adjacent wool strands. Finally, the samples were rinsed three times with distilled water, and baked at 140^oC for 3 minutes in an oven.

For the further analysis of fiber structure, all five samples, that is, the one unstretched and four stretched wool fibers, were rinsed with petroleum ether and ethanol, prior to FTIR spectroscopic analysis, X-ray diffraction (XRD), birefringence measurement and tensile tests.

Measurements and characterization

Mechanical properties

To evaluate the mechanical properties of the wool bundle fibers, the stress–strain curves were obtained with a single fiber strength tester (Model XQ-1, China). The tensile tests of wool fibers were performed at a temperature of 20 ± 2^oC and relative humidity of 65 ± 2% with 10 mm gauge length.

Birefringence measurement

The birefringence measurement was conducted using a polarized light microscope to investigate the variation of orientation.

Fourier transform infrared spectra

FTIR spectra were recorded by transmission on a FTIR spectrometer (NicoletTM 5700, USA) to examine the molecular conformation of the wool fiber. For each measurement, 128 scans were performed with a resolution of 4 cm⁻¹ and a scope of 400–4000 cm⁻¹.

Transmission electron microscope

For transmission electron microscope (TEM) investigation, wool fibers were cut into smaller pieces (≈1 mm), washed three times in petroleum ether and ethanol and then stained with 1% aqueous osmium tetroxide for 5 days followed by 1% aqueous uranyl acetate, dehydrated and embedded in epoxy resin 812. Ultra-thin sections (≈90 nm) were obtained with a diamond blade. The sections were picked up and examined with a HITACHI H-800 electron microscope operating at 175 kV.

X-ray diffraction analysis

XRD analysis was performed by means of a D/max-2550 PC X-ray powder diffractometer (18 kW) using Cu Kα radiation, 0.1541 nm at room temperature.

Results and discussion

Hierarchical structure of wool fiber

Wool fibers are composed of a hierarchical structure, where two isolated right-handed α-helix chains form a left-handed coiled-coil (dimers) with disulfide bonds crosslinking. Then the dimers are crosslinked by the disulfide bonds via end-to-end and side-by-side patterns, as shown in Figure 1. The amorphous region is composed of sulfur-rich cysteine residues (11–17%) or high amounts of glycine, tyrosine and phenylalanine residues.⁴³ Moreover, these disulfide bonds can be formed inter- or intramolecular crosslinking within a single chain or between different peptide chains, which play an important role in protecting the stability of wool fibers. According to the literature, the microstructure of wool fibers remained intact and undamaged even though the 10 gL⁻¹ solution of NaOH was added and the temperature was heated to 120^oC, in which the cysteine residues were reduced.¹⁹ However, during the existence of reducing agents, the tensile strength of wool fibers will be decreased. The interaction can be quite complicated depending on the microstructure and the degree of crosslinking of disulfide bonds. Thus, exploring the microstructure changes and stretching mechanism of wool fibers under an external force, it is useful to understand the stability of wool fibers related to the design of new wool based on materials and structures. Herein, we discussed the structure properties relationships between the stress–strain curves in reducing solution for single wool fiber and after setting for bundle fibers. The discussion is organized as follows. Firstly, the stress–strain curves of a single fiber with different concentration, temperature and strain rate in sodium bisulfite solution are discussed in detail. Secondly, the microstructure transformation mechanism of setting wool fibers by stress–strain curves, FTIR polarization spectra and XRD was analyzed to verify the mechanism.

Figure 1.

Hierarchical structure of wool fibers. Wool fibers feature two α-helix structures in which two dimers are connected by intramolecular and intermolecular disulfide bonds.

Dynamic elastic–plastic behavior of single wool fiber in reducing solution

To understand how wool fibers responded to stretching forces, we recorded the reaction stress as a function of the strain. A typical stress–strain curve of single wool fiber in the different concentrations of sodium bisulfite solution is shown in Figure 2. With the increase of stretching, two types of interactions must exist, namely strong disulfide (-S-S-) and weak hydrogen bonds (-N-H···O).³⁶ In the experiment, the wool fiber was immersed in 1% and 5% solution at 50^oC for 2 minutes, and then stretched. As illustrated in Figure 2, the stress–strain curve was divided into three distinct regions to facilitate the analysis of the mechanical response to the microfibril. For the stress–strain curve of 1% concentration, in the first region (from 0% to 5% strain), the stress increased linearly with strain until the angular point was reached, where the α -helix was stretched with a bond arrangement but without substantial structural change. In the second region, from 5% to 88%, the stress fluctuated around several repeat courses, named stress descending, stress flatting and stress ascending. It was found that the unfolding of the α-helix and the transition of the α-helix to β-pleated sheet occurred in this region. The inter and intra hydrogen bonds decreased and the α-helix unfolded to a β-pleated sheet structure. In addition, owing to the existence of sodium bisulfite solution, the disulfide bonds were prone to be broken and the tensile strength decreased. When the stretching energy applied to the wool fiber exceeded inter- and intramolecular interactions, the molecule chains started to slide along each other. As illustrated in Figure 2, the partial disulfide bonds were broken between different peptide chains, as well as within two α -helical chains, due to the role of sodium bisulfite. The breakages of disulfide bonds significantly caused more mobility within the peptide chains or two α-helical chains. Therefore, the stress in the fiber descended abruptly. After the breakage of the disulfide bonds, the peptide chains and two α -helical chains might slip and rearrange each other, which involved a stress flat. When the peptide chains or two α -helical chains were fully extended due to the breakage of partial disulfide bonds and the stress reached a balance, the stress was undertaken by many peptides and began to ascend. With the increase of stretching, the sodium bisulfite gradually penetrated into the wool fibers and more disulfide bonds and hydrogen bonds were broken. As a result, the fluctuation continued to appear with respect to the three repeat courses. Meanwhile, the α-helical domains started to extend and transited to β-pleated sheets. The α-helix was organized as a coiled coil, which was stabilized by the hydrogen bonds inside the helix chain, resulting in the chain twisting and a helical shape. The β-pleated sheets were held together by intermolecular hydrogen bonds. In the third region, stretching the covalent bonds in the polypeptide backbone of the wool fibers would lead to a rapid increase of stress as strain increased, such as with 1% concentration. The stress value was related to the dipping time, temperature and concentration. At 1% concentration, the breakage stress was 0.056 GPa and the breakage strain was about 107%. For 5% concentration, the wool fiber tended not to break and the strain was more than 230%, demonstrating that more disulfide bond breakage and slippage occurring. That is to say, if the concentration and temperature were appropriate, the crosslinking inside the fiber could be fully broken and the slippage would continue. In addition, the cleavage of disulfide bonds and the α→β transition imparted enhanced energy absorption capability to the structure owing to the increase of the stress–strain curve. It is worthy of note that the magnitude of disulfide bond rupture stress depended on the temperature and strain rate. To further understand the hierarchical structure transformation mechanism and the effect of disulfide crosslinks of single wool fiber in reducing solution, the α→β transition and disulfide bond breakage mechanism are schematically illustrated according to the experimental results of 5% solution, as shown in Figures 2(a)–(d). As shown in Figures 2(a)–(d), the breaking strain was predominately determined by the breakage percentage of the α-helix and disulfide bonds in wool fibers. When the elongation was large enough, as displayed in Figure 2(d), the disulfide bonds were cleaved, and the intermediate filament became more dominant. The α-helix in the intermediate filament was transformed to β sheets and then the disulfide bonds inside the dimers were broken, which caused the β sheets to retract to the α-helix. Moreover, the slippage of macromolecules gave rise to an increase in tensile strain. Therefore, the strain was more than 230%.

Figure 2.

Schematic illustration of proposed stretching mechanism with wool fibers in 1% and 5% bisulfite sodium solution, including intra- and inter-chain hydrogen bonds and disulfide bonds.

Figure 3 illustrates the stress–strain curves of different temperatures and strain rates. There were evident changes in the initial modulus, yield stress, rupturing stress and strain levels during the stretching process. Lower temperature and higher strain rate resulted in greater strength and lower strain. The rupture stress and strain are 0.045 GPa and 108% at 30 ℃, and 0.056 GPa and 70% at 3 min⁻¹, respectively. Figure 3(a) demonstrates that the yield stress and the breakage stress were related to the disulfide bonds in the microfibril and matrix, respectively. The results indicated that the reducing agents broke not only the disulfide bonds crosslinking in the matrix, but also those among the microfibrils. Thus, the presence of the reducing agents enhanced the slippage of molecules. As the strain rate increased, the Young's modulus, yield stress and rupture strength in the stress–strain curves increased, suggesting that the structure of macromolecules was not enough to result in relaxation, as illustrated in Figure 3(b). The wool fiber had a yield stress change from 0.013 GPa (at 0.25 min⁻¹) to 0.019 GPa (at 3 min⁻¹), resulting in a decrease in the work of fracture. For Figure 3(a), the work of fracture was approximately 0.87 and 2.77 GPa at 75 ℃ and 30 ℃, respectively. The decrease of work with the increase of temperature resulted from slippage between the fibers, macrofibrils, microfibrils and dimers.

Figure 3.

Stress–strain curves of wool fibers at (a) different temperatures (30 ℃, 50 ℃ and 75 ℃) and (b) different pulling rates (0.25, 1 and 3 min⁻¹).

Microstructure transformation of the wool bundle fibers after setting

Inter and intra hydrogen bonds probe

FTIR spectral recordings of wool bundle fibers were carried out for various levels of stretching. As noted by Tsobkallo et al.,³⁸ the 2600–4000 cm⁻¹ region of the FTIR spectra gave information about the hydrogen bond interactions of wool fibers. The structure of the region was analyzed by curve fitting and performed as shown in Figure 4. When the wool fibers were stretched, with the breakage of intrahelical hydrogen bonds, the distance of the turns of the α-helix increased and transformed to β-pleated sheets. After the stretched wool fibers were set, the hydrogen bonds were reconstructed. According to Figures 4(a) and (b), we noticed that the changes of bonds were not found to vary as a function of extension.

Figure 4.

Fourier transform infrared spectra in the region 2600–4000 cm⁻¹ of (a) raw and (b) 110% wool fibers with component bands at wavenumber, cm⁻¹: 2925 (1 CH₃), 3065 (2 C=CH), 3180 (3 absorbed water in the amorphous phase), 3300 (4 NH), 3400 (5 free NH) and 3500 (6 OH).

FTIR polarized spectra can provide useful information about the fiber structure changes of C=O in the amide I band by evaluating the secondary structure transition of wool fibers. In previous studies, the secondary structures of various keratin fibers, including the α-helix, β-pleated sheet, β-turns and random coils, were identified by FTIR and Raman spectra.^34,44 As reported, several treatments, such as heat, humidity and chemical methods, could facilitate the transition of α-keratin fibers from α-helix to β-pleated structures.^45–47 In addition, stretching produced an obvious crystallite and conformation on the α-helix and β-pleated sheets and the wool fibers were also investigated by XRD. Figure 5 presents the FTIR polarized spectra of wool fibers for parallel and perpendicular direction at different ratios. In Figure 5(a), we noticed that the maximum peak in the amide I group of raw wool fibers was observed at 1677, 1658, 1643 and 1627 cm⁻¹, which is usually attributed to β-turns, the α-helix, random coils and the β-pleated sheet, respectively. The bands observed at 1695 and 1612 cm⁻¹ can be attributed to the side chain amide I groups of glutamine, arginine and tyrosine, respectively. As expected, the bands were not found to vary as a function of stretching. When the wool fibers were stretched, the peaks gradually became unobvious and broader due to the role of secondary structure transformation. Moreover, with the increase of the stretching ratio, the location of absorbance peaks shifted to a low wavenumber. The results confirmed that the stretching process caused the conformation to extend and formed the β-pleated structure by hydrogen bonds. The conformation contents in the sample could be calculated from the deconvolution of amide I peaks, and the corresponding contents of the α-helix and β-pleated sheet are as indicated in Figure 5(b). As the stretching ratio increased, the α-helix content first decreased and then increased. Generally, for the α-helix structure, the C=O vibration direction was parallel with the wool fiber axis. However, the vibration direction of the β-pleated structure was perpendicular to the fiber axis. As shown in the parallel direction data, when the stretching ratio was less than 50%, the α-helix content reduced from approximately 42.78% in raw wool fibers to 25.34% in 50% stretched wool fibers due to the transformation from the α-helix to β structure. In contrast, the spectral contribution of the β-pleated content in the parallel direction showed almost no change at all. The results demonstrated that the FTIR polarization spectra were not sensitive to β-pleated content in the parallel direction. When the stretching ratio was higher than 50%, the α-helix content increased from 25.34% to 34.24% due to the disintegration or separation of α-helix structures from each other, according to Figure 5. However, the β-pleated content in the perpendicular band in Figure 5(b) was more obvious than that of the parallel band. It is worth noting that the β-pleated content had risen from approximately 13.59% (raw wool fiber) to 29.94% (80% stretching ratio). When the stretching ratio of wool fibers was 80%, the α-helix content was 16.72%. Moreover, we found a surprising result that when the stretching ratio was 50% and 80%, the α-helix content in the parallel direction was consistent with the β-pleated content in the perpendicular direction, as shown in Figure 5(b). The result showed that the α-helix structure directly transformed to a β-pleated structure when the stress–strain curve was in the yield region of the wool fibers. However, when the stretching ratio was 110%, the α-helix content (34.24%) and β-sheet content (15.80%) in the parallel direction were consistent with that of 80% (33.76% for α-helix and 15.81% for β-sheets). In contrast, the α-helix content contribution in the vertical direction was almost twice as large as 80%, suggesting that as the dimer composed of two α-helical chains scission increased, two separate α-helical chains were formed due to the cleavage of disulfide bonds in the dimers ends.

Figure 5.

Fourier transform infrared spectra and X-ray diffraction (XRD) of stretched wool fibers: (a) parallel band, vertical band; (b) α-helix and β-sheet content. The single bands of stretched wool fibers are attributed to corresponding secondary structures, according to previous studies⁴⁹: 1605–1615 cm⁻¹, aggregated strands; 1616–1637 cm⁻¹, β-sheets; 1638–1655 cm⁻¹, random coil; 1656–1662 cm⁻¹, α-helix; 1663–1695 cm⁻¹, β-turns. (c) XRD of different stretching ratio wool fibers. (d) The crystallite fraction from XRD.

To further quantify the α and β crystallite contents, XRD experiments were conducted and the results are indicated in Figures 5(c) and (d). The α crystallite peaks of wool fibers were around 9.3 °, 19.7 ° and 24.72 °. In contrast, the β crystallite peaks of wool fibers were located at 18.9 ° and 20.7 ° according to Figure 5(c). With the increase of the stretching ratio in the wool fibers, the maximum peak of wool fibers exhibited gradually a sharper and stronger diffraction peak at 2θ = 18.9 °, which is a typical characteristic pattern of the β crystallite structure. It is worth noting that the α and β crystallites were affected by the stretching ratios. The percentages of α crystallite in the stretched wool fibers were higher than those of the β crystallite structure. As illustrated in Figure 5(d), we noticed that the change trend of α and β crystallites was consistent with the parallel direction. Moreover, the peak area of the β crystallite structure increased gradually before the 80% stretching ratio. When the stretching ratio was 110%, the α crystallite was higher than the β crystallite, demonstrating the breakage of disulfide bonds inside the dimers and retraction of the β sheets to the α-helix. That is to say, when the bundle fibers were stretched, the secondary structure transformation was accelerated due to the existence of the synergistic effect between the fibers. Therefore, the actual percentage of the total α and β crystallites in wool fibers was higher than those of in parallel and vertical directions of the same stretched wool fibers. This indicated that the secondary structure of parallel or vertical direction did not cover all crystalline parts in wool fibers.

The orientation degree of wool fibers has a significant effect on their mechanical properties. The oriented structure of wool bundle fibers was detected by the XRD and birefringence method in two directions: one perpendicular to the axis of the wool fiber and the other parallel with it. The variation of the preferred orientation and birefringence values of the wool fibers is shown in Figure 6. It should be noted that the stretching strengthened the preferred orientation and birefringence values in the range from 0% to 110%, leading to an improvement in the anisotropy. The improvement in anisotropy might be explained by the hypothesis that the stretching increased the orientation degree between the molecules. Without stretching, the preferred orientation and birefringence values were minimal, and with the addition of the stretching ratio, the transformation from α-helix to β-pleated sheet and the breakage of disulfide bonds increased and the intermediate filament became gradually close to form a much higher orientation structures. The TEM micrographs of 80% stretched wool fibers are shown in Figure 6(c). According a study by Bruce Fraser and Parry,⁴⁸ the filaments observed in the assembly were placed in a hexagonal lattice, in which the microfibrils were separated by the matrix and the diameter of the microfibril was nearly the same. When the wool fiber was stretched, the intermediate microfibril was squeezed by the surrounding microfibrils and elongated along the stretching direction, as illustrated in Figure 6(c). Obviously, the stretching led to a closer packing of all microfibrils within any given cross-section. In addition, the microfibrils were composed of different thickness. The results indicated that during loading, some of the weak disulfide bonds broke and the microfibril was stretched and elongated. Some of strong disulfide bonds remained intact until the stress was reached, in which the microfibrils remained relatively stable. Meanwhile, slippage of molecule chains or microstructural elements and α→β transformation occurred. As shown in Figure 6, cracks were formed between the cuticle and the cortex after stretching, indicating slippage between the molecules.

Figure 6.

X-ray diffraction scan of preferred orientation and birefringence ((a) and (b)) and transmission electron microscope (TEM) micrographs of 80% stretched wool fibers in the cortex (c). The TEM image illustrates the microfibril packing in the microfibril.

Tensile properties and strain modulus after setting wool fibers

The tensile response of wool fibers was studied and compared, and the results of stress–strain curves are shown in Figure 7. It was evident that the original wool fibers, as shown in Figure 7(a), had three distinct regions, that is, Hookean, yield and post-yield. At the beginning, in the Hookean region, the wool fiber displayed linear elastic behavior before yield points, in which these α-helices experienced a reversible bond angle rearrangement. Beyond the yield points, the microstructure was transformed from the α-helix to the β-pleated sheet. Meanwhile, the stress–strain curve in this region indicated a very slow increase in stress. In the last region, with the increase of stress, the covalent bonds of the wool backbone were further stretched until breakage. In Figure 7(b), wool fibers with 30% stretching ratio were still exhibited in the three regions, indicating that the α-helix structure still existed in wool fibers. Meanwhile, for 50% wool fibers, the stress–strain curves showed two or three regions, as illustrated in Figure 7(c). Moreover, the curves of three regions were in the majority. The curves of two regions were in the no-yield region, which were relatively similar to the tensile curves of silk, with the microstructure composed of the β crystals. This was due to the α→β transformation during the stretching. In Figure 7(d), for the 80% stretching ratio, the stress–strain curves of two regions were in the majority and the breakage elongation decreased to 20–30%. This indicated that β crystals were in the majority in the 80% wool fibers. For 110% wool fibers, the stress–strain curves were only divided into two distinct regions, demonstrating that most of the α-helices in wool had been transformed to the β-pleated sheets, as shown in Figure 7(e). Therefore, with the increase of the stretching ratio, the breaking strength and Young's modulus increased, but the elongation decreased according to Figure 7(f). The reason why the breaking strength and Young's modulus increased and elongation decreased was due to the higher β-sheet structure in the stretching wool fibers, as indicated in the FTIR results and illustrated in Figure 5. The Young's modulus and tensile strength increased from 2.88 to 4.51 GPa and 0.15 to 0.21 GPa, respectively. In contrast, the breaking strain obviously decreased from 46.67% to 15.67% for the raw wool fiber with 110% stretching ratio. This could be because the stretching wool fibers were rich in β-sheets, which could increase the intermolecular crosslinking during the setting process. As illustrated by Figures 6(b) and (c), the stretching facilitated the increase of orientation degree and the closer packing of microfibrils in wool fiber, which gave rise to increase of β-sheet crystals. However, the extendibility of the stretching wool fibers was to a large extent decided by the α-helix. In other words, the α-helix was flexible and more like a spring to contribute to the elongation, while the β-sheet was rigid and provided strength. Thus, with the increase of the stretching ratio in wool fibers, the content of the breaking strain β-sheet increased while α-helix content dropped.

Figure 7.

Tensile stress–strain curves of (a) raw, (b) 30%, (c) 50%, (d) 80% and (e) 110% wool fibers; (f) the tensile characteristic value from the stress–strain curves.

Conclusions

In summary, we determined that the stretching of single wool fibers served as an excellent approach for exploring the microstructure transformation mechanism. This study demonstrated the effective investigation of wool fibers by secondary transition, disulfide bond crosslinking and macromolecule slippage. When the treatment condition was suitable, the strain of a single fiber is over 2.3 times. To unveil the underlying mechanism of the stretching process, the microstructure of the wool bundle fibers was investigated and the hierarchical structure was found. The model was built by transformation from the α-helix to the β-pleated sheet, the breakage of disulfide bonds and the slippage of microfibrils. Our results offer new mechanistic, structural insights into the complex dynamic behavior of stretched wool fibers and an understanding of the unique role of coiled coil in fibrous proteins.

Footnotes

Declaration of conflicting interests

The authors received no financial support for the research, authorship, and/or publication of this article.

Funding

The authors received no financial support for the research, authorship and/or publication of this article.

ORCID iD

Hongling Liu

References

Church

Corino

Woodhead

. The analysis of Merino wool cuticle and cortical cells by Fourier transform Raman spectroscopy. Biopolymers 1997; 42: 7–17.

Fougere

Vargiolu

Pailler-Mattei

, et al. Study of hair topography modification by interferometry. Wear 2009; 266: 600–604.

Zhu

Zeng

Meng

, et al. Using wool keratin as a basic resist material to fabricate precise protein patterns. Adv Mater 2019; 31: 1900870.

Liu

, et al. Observation of luminescent gold nanoclusters using one-step syntheses from wool keratin and silk fibroin effect. Eur Polym J 2018; 99: 1–8.

Xing

Shi

Zhao

, et al. Mesoscopic-functionalization of silk fibroin with gold nanoclusters mediated by keratin and bioinspired silk synapse. Small 2017; 13: 1702390.

Eder

Amini

Fratzl

. Biological composites—complex structures for functional diversity. Science 2018; 362: 543.

Huang

Restrepo

Jung

J-Y

, et al. Multiscale toughening mechanisms in biological materials and bioinspired designs. Adv Mater 2019; 31: 1901561.

Puglia

Ceccolini

Fortunati

, et al. Effect of processing techniques on the 3D microstructure of poly (l-lactic acid) scaffolds reinforced with wool keratin from different sources. J Appl Polym Sci 2015; 132: 42890.

Zhang

Nie

Zhang

, et al. Quantitative change in disulfide bonds and microstructure variation of regenerated wool keratin from various ionic liquids. ACS Sustain Chem Eng 2017; 5: 2614–2622.

10.

Tsobkallo

Aksakal

Darvish

. Recovery process in stretched wool fibers. J Macromol Sci Phys 2010; 49: 495–505.

11.

Bin

Tian

, et al. In situ synthesis of gold nanoparticles on wool powder and their catalytic application. Materials (1996–1944) 2017; 10: 295.

12.

Xin

Weilin

Wenbin

, et al. Thermoplastic film from superfine wool powder. Fibre Text East Eur 2009; 17: 82–86.

13.

Pakdel

Daoud

Wang

. Self-cleaning and superhydrophilic wool by TiO2/SiO2 nanocomposite. Appl Surf Sci 2013; 275: 397–402.

14.

Church

Corino

Woodhead

. The effects of stretching on wool fibres as monitored by FT-Raman spectroscopy. J Mol Struct 1998; 440: 15–23.

15.

Chen

Xing

, et al. Wool keratin and silk sericin composite films reinforced by molecular network reconstruction. J Mater Sci 2018; 53: 5418–5428.

16.

Motaghi

Eskandarnejad

Montazer

, et al. Mechanical slenderising of coarse wool fibre and determination of its characteristics with FTIR and Raman spectroscopy. J Text Inst 2012; 103: 8–18.

17.

Cao

Billows

. Crystallinity determination of native and stretched wool by X-ray diffraction. Polym Int 1999; 48: 1027–1033.

18.

Rajabinejad

Zoccola

Patrucco

, et al. Physicochemical properties of keratin extracted from wool by various methods. Text Res J 2017; 88: 2415–2424.

19.

Shavandi

Silva

Bekhit

, et al. Keratin: dissolution, extraction and biomedical application. Biomater Sci 2017; 5: 1699–1735.

20.

Ramya

Thangam

Madhan

. Comparative analysis of the chemical treatments used in keratin extraction from red sheep's hair and the cell viability evaluations of this keratin for tissue engineering applications. Proc Biochem 2020; 90: 223–232.

21.

Shavandi

Bekhit

AE-DA

Carne

, et al. Evaluation of keratin extraction from wool by chemical methods for bio-polymer application. J Bioactive Compatib Polym 2016; 32: 163–177.

22.

Feughelman

. A two-phase structure for keratin fibers. Text Res J 1959; 29: 223–228.

23.

Feughelman

Haly

. The mechanical properties of wool keratin and its molecular configuration. Kolloid-Zeitschrift 1960; 168: 107–115.

24.

Feughelman

. 40—The mechanical properties of set wool fibres and the structure of keratin. J Text Inst Trans 1960; 51: T589–T600.

25.

Feughelman

. A model for the mechanical properties of the α-keratin cortex. Text Res J 1994; 64: 236–239.

26.

Feughelman

. Natural protein fibers. J Appl Polym Sci 2002; 83: 489–507.

27.

Hearle

JWS

Chapman

. On polymeric materials containing fibrils with a phase transition. I. General discussion of mechanics applied particularly to wool fibers. J Macromol Sci B 1968; 2: 663–695.

28.

Chapman

. A mechanical model for wool and other keratin fibers. Text Res J 1969; 39: 1102–1109.

29.

Hearle

. A critical review of the structural mechanics of wool and hair fibres. Int J Biol Macromol 2000; 27: 123–138.

30.

Wortmann

F-J

Zahn

. The stress/strain curve of α-keratin fibers and the structure of the intermediate filament. Text Res J 1994; 64: 737–743.

31.

Cao

. Is the α-β transition of keratin a transition of α-helices to β-pleated sheets? Part I. In situ XRD studies. J Mol Struct 2000; 553: 101–107.

32.

Kreplak

Doucet

Briki

. Unraveling double stranded α-helical coiled coils: an x-ray diffraction study on hard α-keratin fibers. Biopolymers 2001; 58: 526–533.

33.

Cao

. Is the α–β transition of keratin a transition of α-helices to β-pleated sheets. II. Synchrotron investigation for stretched single specimens. J Mol Struct 2002; 607: 69–75.

34.

Liu

. Study of the structure transformation of wool fibers with Raman spectroscopy. J Appl Polym Sci 2007; 103: 1–7.

35.

Liu

Jin

. Modeling the stress-relaxation behavior of wool fibers. J Appl Polym Sci 2008; 110: 2078–2084.

36.

Paquin

Colomban

. Nanomechanics of single keratin fibres: a Raman study of the α-helix →β-sheet transition and the effect of water. J Raman Spectr 2007; 38: 504–514.

37.

Pauling

Corey

Branson

. The structure of proteins; two hydrogen-bonded helical configurations of the polypeptide chain. Proc Natl Acad Sci USA 1951; 37: 205–211.

38.

Tsobkallo

Aksakal

Darvish

. Recovery process in stretched wool fibers. J Macromol Sci B 2010; 49: 495.

39.

Chou

C-C

Buehler

. Structure and mechanical properties of human trichocyte keratin intermediate filament protein. Biomacromolecules 2012; 13: 3522–3532.

40.

Yang

André Meyers

. Viscoelastic properties of α-keratin fibers in hair. Acta Biomaterialia 2017; 64: 15–28.

41.

Zhmurov

Kononova

Litvinov

, et al. Mechanical transition from α-helical coiled coils to β-sheets in fibrin(ogen). J Am Chem Soc 2012; 134: 20396–20402.

42.

Ding

Borreguero

Buldyrey

, et al. Mechanism for the α-helix to β-hairpin transition. Protein Structure Funct Bioinformat 2003; 53: 220–228.

43.

Wang

Yang

McKittrick

, et al. Keratin: structure, mechanical properties, occurrence in biological organisms, and efforts at bioinspiration. Progr Mater Sci 2016; 76: 229–318.

44.

Liu

Zhao

. Structural changes in slenderized yak hair induced by heat–humidity conditions using Raman spectroscopy. J Mol Struct 2013; 1037: 57–62.

45.

Kreplak

Doucet

Dumas

, et al. New aspects of the α-helix to β-sheet transition in stretched hard α-keratin fibers. Biophys J 2004; 87: 640–647.

46.

Kreplak

Franbourg

Briki

, et al. A new deformation model of hard α-keratin fibers at the nanometer scale: implications for hard α-keratin intermediate filament mechanical properties. Biophys J 2002; 82: 2265–2274.

47.

Yang

Wang

, et al. Structure and mechanical behavior of human hair. Mater Sci Eng C 2017; 73: 152–163.

48.

Bruce Fraser

Parry

DAD

. Macrofibril assembly in trichocyte (hard α-) keratins. J Struct Biol 2003; 142: 319–325.

49.

Kaplan

Cebe

. Determining beta-sheet crystallinity in fibrous proteins by thermal analysis and infrared spectroscopy. Macromolecules 2006; 39: 6161–6170.