Deformation Drives Alignment of Nanofibers in Framework for

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Deformation Drives Alignment of Nanofibers in Framework for Inducing Anisotropic Cellulose Hydrogels with High Toughness Dongdong Ye, Qiaoyun Cheng, Qianlei Zhang, Yixiang Wang, Chunyu Chang, Liangbin Li, Haiyan Peng, and Lina Zhang ACS Appl. Mater. Interfaces, Just Accepted Manuscript • DOI: 10.1021/acsami.7b14900 • Publication Date (Web): 21 Nov 2017 Downloaded from http://pubs.acs.org on November 27, 2017

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ACS Applied Materials & Interfaces

Deformation Drives Alignment of Nanofibers in Framework for Inducing Anisotropic Cellulose Hydrogels with High Toughness Dongdong Ye,† Qiaoyun Cheng,† Qianlei Zhang,‡ Yixiang Wang,§ Chunyu Chang,†,* Liangbin Li,‡ Haiyan Peng,ǁ and Lina Zhang †,* †

College of Chemistry and Molecular Sciences, Wuhan University, Wuhan 430072, China.

‡

National Synchrotron Radiation Lab and College of Nuclear Science and Technology, University of

Science and Technology of China, Hefei 230026, China §

Department of Agricultural, Food and Nutritional Science, University of Alberta, Edmonton, Alberta,

T6G 2P5, Canada ǁ

Key Laboratory for Material Chemistry of Energy Conversion and Storage, School of Chemistry and

Chemical Engineering, Huazhong University of Science and Technology, Wuhan 430074, China

KEYWORDS: Cellulose hydrogel, Hierarchical structure, Alignment, Birefringence, Framework

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ABSTRACT Deformation driven alignment of molecules or nanofibers leading to anisotropy is a challenge in functional soft materials. Here, tough cellulose hydrogels which exhibited deformation-induced anisotropy are fabricated by reacting cellulose with a small amount of epichlorohydrin (EPI) in LiOH/urea solution and subsequent treating with dilute acid. The loosely crosslinked network that was obtained via chemical crosslinking of cellulose with EPI as a large framework maintained the elasticity of hydrogels, whereas nanofibers produced by the acid treatment formed physical crosslinked networks through hydrogen bonds which could efficiently dissipated mechanical energy. Meanwhile, the nanofibers also tended to form submicrobundles and participate in the formation of frameworks during the acid treatment. Under deformation, the nanofibers and submicrobundles in the physical networks synchronize easily to align with the large framework, generating the rapidly responsive birefringence behaviors with highly stable colors. Thus, the cellulose hydrogels possessing sensitively mechanoresponsive behavior could be utilized as a dynamic light switch and a soft sensor to accurately detect small external force, respectively. This work opens a novel pathway to construct tough and mechanoresponsive hydrogels via a green conversion of natural polysaccharide. INTRODUCTION It is well known that conventional hydrogels are extremely brittle and fragile after full swelling, which seriously limits their practical applications.1 Many efforts have been made to improve the mechanical performances of hydrogels, and various hydrogels with excellent mechanical properties, including topological hydrogels,2 nanocomposite hydrogels,3 double network hydrogels,4 dual-cross-linked hydrogels,5 and anisotropic hydrogels,6,7 have been sequentially reported. These hydrogels have very different ways to dissipate mechanical enenrgy, for example, the reinforcement mechanisms for double network hydrogels or dual-cross-linked hydrogels are the co-existence of weak network or crosslinks which can rupture to dissipate energy under deformation, and strong network or crosslinks to maintain the initial elastic configurations of hydrogels.8 Besides, hydrogels are required to exhibit biocompatibility based on biomimicry of the extracellular matrix for their basic and clinic applications ACS Paragon Plus Environment

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in tissue engineering, cell therapy, and biomedical research.9 Cellulose, as the most abundant renewable resource, which was previously thought to be impossible to dissolve in aqueous media, has been found to be soluble in alkali/urea aqueous solutions in our laboratory.10 Moreover, a series of biomedical materials with excellent biocompatibility have been constructed from the cellulose solution.11,12 Cellulose-based

hydrogels

have

attracted

much

attentions

due

to

their

biocompatibility,

biodegradability, low cost and hydrophilicity.13 To improve the strength of cellulose hydrogels, the sequential chemically and physically cross-linked cellulose hydrogels have been fabricated, exhibiting high stiffness and toughness.14 Materials, developed in the course of nature evolution like wood and crab shell, exhibit unique hierarchical architecture, which endows them with outstanding stiffness, toughness, and strength.15,16 Highly anisotropic wood-based materials have been constructed from the naturally structural anisotropy, on the basis of the nano- and micro-scale aligned fibers.17,18 The structure of regenerated silk fibroin under mechanical-tension can be manipulated to form patterned material with hierarchically oriented nano-, micro- and macrostructure.19 In the view of mentioned above, advanced functional materials with impressive properties are dominated by their hierarchical structure. Therefore, a new design that has never been considered for fabricating cellulose hydrogels can emerge, namely, utilizing “bottom-up” approarch in constructing hierarchical structure. In this work, we demonstrated that cellulose hydrogels with high thoughness and sensitive deformation-stimuli responsive behaviors could be constructed from a loosely chemical crosslinking network as large framework by adding tiny amount of epichlorohydrin (EPI) and a densely physical network containing nanofibers and submicrobundles via a green conversion of natural polysaccharide. The loose chemical network as a large framework could maintain the high toughness of hydrogel, whereas the dense physical cross-linked networks integrated inside framework were reversible, which could dissipate mechanical energy through the breaking of “sacrificial bonds” under large deformation for maintaining the high strength of hydrogel. The nanofibers (diameter < 100 nm) and submicrobundles (diameter < 1000 nm) in the physical network synchronized easily to align with chemical frameworks under deformation, leading to the rapidly responsive ACS Paragon Plus Environment


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birefringence behaviors with highly stable colors. The highly toughness and deformation-induced anisotropic soft materials are very important in sensing external stimulus. EXPERIMENTAL SECTION Materials. All of the reagents were used as received unless otherwise noted. The cellulose (cotton linter pulps), with the α-cellulose content beyond 95%, was provided by Hubei Chemical Fiber Group Ltd. (Xiangfan, China). The cotton linter pulps was used after totally dried with vacuum at 60 °C and without further purification. Its viscosity-average molecular weight in cadoxen was determined using an Ubbelohde viscometer at 25 °C to be 9.6×104 according to Mark-Houwink equation [η] (mLg-1) = 3.85×102

(Mw)0.76. LiOH·H2O, urea, concentrated sulfuric acid and epichlorohydrin (EPI) were purchased from

Sinophram Chemical Reagent Co., Ltd (Shanghai, China). Preparation of Tough Cellulose Hydrogels (TCHs). Transparent cellulose solution (6wt%) was prepared through dissolving cellulose (cotton linter pulps) in an aqueous 4.5wt% LiOH /15wt% urea solution which was pre-cooled to -12 °C (Figure S1a,b). Then, certain amounts of EPI were added drop wise into the bubble-free cellulose solution with stirring at 0 °C for homogeneous mixing and preliminary chemical crosslinking reaction. The transparent and viscous cellulose solution with different EPI/anhydroglucose unit (AGU) molar ratios were poured into hand-made molds after removing air bubbles via centrifugation, and kept at 5 °C for 8 h. Subsequently, the obtained cellulose gels were removed from the aforementioned molds and immersed into 5 wt% aqueous sulfuric acid pro-cooled at 5 °C for several minutes to terminate the chemical crosslinking reaction and simultaneously induce physical crosslinking. Finally, tough cellulose hydrogels were obtained after thorough washing with deionized water, and were coded as TCH-1 to TCH-7 according to the EPI/AGU molar ratios (Table 1). Loose chemical crosslinked cellulose hydrogel (LCH) as reference (EPI/AGU=1:1.33) was prepared using the same procedure expect the acid treatment. Water Content Measurement of Cellulose Hydrogel. Water content (Wc) of the hydrogel was calculated according to equation (1): ACS Paragon Plus Environment

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% =


× 100%

(1 )

Where Ws and Wd were the weight of the swollen hydrogel and dried gel, respectively. Atomic Force Microscopy (AFM) Measurements. The AFM images were recorded on the CypherTM S (Asylum Research) equipped with a buleDrive photothermic excitation option, which use not only the laser for detecting changes on the cantilever deflection, but also an additional blue laser to heat the cantilever of the probe to ensure a stable oscillation of the beam even in fluid environment. Besides, probe selection is one of the most important considerations in atomic force microscopy and successful AFM imaging starts with the probe. Accordingly, silicon nitride probes (SNL-10, BRUKER), with the average tip radius of 2 nm, spring constant of 0.06~0.35 N/m and resonance frequency of 18~65 kHz, were employed for scanning to investigate the hydrogels in a fluid environment. The experiment environment was maintained at ambient temperature (20 °C) and 100% humidity. Furthermore, we used probes (RTESP-300, BRUKER) with a high spring constant (~40 N/m) and a high resonance frequency (~300 kHz) in air environment to probe the surface morphologies of anhydrate hydrogels. All data of the images were analyzed using the AFM accessory software. Mechanical Tests. Mechanical measurements of hydrogels were characterized on a universal tensilecompressive instrument (Model 5576, INSTRON Instrument, USA) equipped with two flat-surface compression stages under compression test, pneumatic clamping under tensile strain and two kinds of load cells (50N and 1000N). For tensile measurements, hydrogel specimens with 60 mm long and 10 mm wide were stretched at a speed of 5 mm/min. The cylindrical hydrogel specimens with diameter of 10 mm and height of 20 mm were loaded between the two compression stages with the top stage applying uniaxial compression and release on the samples along the vertical direction at a speed of 5 mm/min. The cyclic loading-unloading tests, using the same specimens at equal test speed, were carried out by performing subsequent trials immediately after the initial loading. Nanoindentation

Measurements.

A displacement-controlled

nanoindenter

machine

(Piuma

Nanoindenter by Optics11, Netherlands) equipped with a controller, an optical fiber and a spherical ACS Paragon Plus Environment


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probe (Optics, Netherlands) was used to investigate the force-displacement curves on surface of hydrogels in the fluid environment. In this work, two kinds of probes with cantilever stiffness of 3.32 N/m and 45.5N/m, corresponding with tip radius of 46 mm and 25 mm, were used to test LCH and TCH, respectively. The effective Young’s modulus was calculated using the Oliver & Pharr theory from the slope of the initial portion between 65% and 85% of the unloading curve. The grid scan was 10×10 points, and the point-to-point pitch is 20 µm.

Figure 1. Fabrication and orientation of TCH. (a-c) Schematic diagram of cellulose hierarchical networks and the orientation within deformed cellulose hydrogel: a) construction of loosely chemically crosslinked cellulose network as framework through reacting cellulose by adding tiny amount of EPI in cellulose solution; b) formation of hierarchical structure including cellulose submicrobundles and nanofibers after rapid dilute acid treatment; and c) synchronized orientation of both physical and chemical networks under external force. (d-f) Representative AFM height images of: d) cellulose chains and aggregates in dilute cellulose solution with the concentration of 0.005 mg/mL; e) TCH; and f) deformed TCH.

Morphology Measurement. Scanning electron microscopy (SEM) images were taken with the field emission scanning electron microscopy (Zeiss, SIGMA, Germany) using an accelerating voltage of 5 kV. The hydrogels with certain degree of tensile or compressive deformation, were frozen in liquid nitrogen, snapped immediately, and freeze-dried to remove the water content without causing significant ACS Paragon Plus Environment

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structural change to the hydrogels. Both surface and cross-section of dried specimens were coated with gold vapor, observed, and photographed. X-ray Measurements. Wide angle X-ray scattering (WAXS) and small angle X-ray scattering (SAXS) profiles were obtained at the beamline BL16B1 of Shanghai Synchronization Radiation Facility (SSRF). The wavelength was 0.124 nm, and a CCD X-ray detector (MAR CCD 165, 2048 x 2048 pixels with pixel size of 80 µm) was employed to collect data. The sample-to-detector distances were calibrated to 152.2 mm by yttrium sesquioxide (Y2O3) for WAXS, and 2540 mm by beef rendon for SAXS, respectively. The data acquisition time was 30s for both the WAXS and SAXS patterns, and the two dimensional scattering images were analyzed with Fit2D software from European Synchronization Radiation Facility. The variable orientation parameter of hydrogels was achieved in a miniaturization stretching equipment. The corresponding WAXS profiles were normalized to beam intensity and corrected through subtracting the background scattering from the air and inner water because the signal arising from inner water was overlapped with that of cellulose to have great influence on identification of cellulose. SAXS data were corrected for background scattering through subtracting contribution from air and sample holder. Nano-tomography Experiment. The nano-tomography experiment was performed using the full field transmission hard X-ray microscopy experiment on the beamline 4W1A of the Beijing Synchrotron Radiation Facility. Briefly, an elliptically shaped capillary condenser was used to focus the incident Xrays onto the hydrogel specimen, and a phase ring was placed in the back focal plane of the objective to provide the Zernike phase contrast. A zone-plate objective magnified images of hydrogel in a 1024 x 1024 charge coupled camera. In this work, phase contrast imaging mode with the photon energy of 8 keV, view field of 60 µm x 60 µm and a spatial resolution of 100 nm were selected. Combined with computerized tomography, the spatial information of cellulose hydrogel was obtained in 3D space. RESULTS AND DISCUSSION Formation and Structure of Tough Cellulose Hydrogels. Cellulose solution was prepared in LiOH/urea aqueous system at low temperature. In this system, an inclusion complex associated with celluACS Paragon Plus Environment


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lose chains and alkali-urea-water clusters generated through hydrogen-bonding at low temperature, leading to the cellulose dissolution.20 Especially, the alkali-urea sheath around the cellulose complex could be broken at elevated temperatures and concentrations, or dilute acidic condition, leading to the aggregation of

Figure 2. Morphology of deformed TCHs. (a) AFM phase images of TCH-5 with certain extents of tensile deformation. (b-c) SEM images and corresponding magnified SEM images of TCH-5 with certain extents of tensile deformation. cellulose chains in a parallel pattern to form nanofibers.21 The aggregation behavior of the cellulose macromolecules in solution was characterized with atomic force microscopy (AFM) and dynamic light scattering (DLS) (Figure S1c, d). The results demonstrated that the cellulose chains easily aggregated in a parallel pattern to form nanofibers with an increase of cellulose concentration. The dissolved cellulose mainly existed in form of nanofibers in the concentrated cellulose solution, making it possible to conACS Paragon Plus Environment

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struct loosely cross-linked gel through chemical crosslinking of cellulose with EPI as well as physical networks consisted of nanofibers. Thus, cellulose hydrogels with hybrid crosslinking networks were fabricated, and were coded as TCH-1 to TCH-7 according to the EPI/AGU molar ratios (Table 1).

Figure 3. Appearance of deformed TCHs. (a, c, e) Photographs of TCHs under various deformation (e.g., bending, twisting and knotting) observed under bright filed. (b, d, f) Iridescent birefringence patterns of deformed TCHs under polarized light. Scar bar: 1 cm. Figure 1 shows the schematic diagram of formation of TCH, and representative AFM height images of cellulose solution and hydrogels, respectively. Firstly, loosely cross-linked gels were synthesized by chemical crosslinking of cellulose with a small amount of EPI, where the EPI/AGU molar ratio was accurately controlled to achieve tunable crosslinking density (Figure 1a; Table 1). It was noted that the cross-linking density was measured to be very low, ranging from 72.92 to 322.22 mol m-3 (Table S1). Subsequently, the chemical crosslinked gels were immersed in dilute sulfuric acid aqueous solution, in which the naked cellulose molecules could align sufficiently in parallel manner to form nanofibers and submicrobundles, constituting a dense physical crosslinking network (Figure 1b). Thus, the TCH was consisted of the loosely irreversible chemical crosslinking and densely reversible physical crosslinking networks (Figure 1e). The chemical crosslinking reaction was confirmed by the solid state

13

C NMR

(Figure S2). Moreover, the signal of C4 at 89 ppm (crystalline) for physically cross-linked cellulose hydrogel shifted to ~84 ppm (amorphous) and showed a large loss of resolution, indicating that the degree ACS Paragon Plus Environment


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of cystallinity of cellulose hydrogel greatly decreased after chemical crosslinking. Besides, with an increase of EPI content, the peak C2,3,5 became broad, demonstrating that the chemical crosslinking density in TCH increased, leading to the lower mobility of carbons of D-glucose units.22 The crystallinity of various hydrogels were characterized with the X-ray diffraction patterns (Figure S3). A strong peak at 20.0°, which corresponds to the (110) reflection of cellulose II crystallite, appeared in TCH, suggesting an increase of crystallinity. It has been reported that the substantial lateral aggregation of nanofibers into submicrobundles led to the formation of cellulose crystallite hydrate.23 Besides, the crystallinity of TCHs decreased as the chemical crosslinking density of hydrogels increased from TCH-1 to TCH-7, indicating that both EPI content and acid treatment played the important roles in the tuning of the crystallinity degree of TCHs. Interestingly, LCH with chemical crosslinking was fragile and brittle, which easily cracked under compression and could not withstand stretching (Figure S4a-c). However, TCHs exhibited high strength and excellent toughness after hybrid crosslinking, which could undergo the compression of a finger, and tensile deformation (Figure S4d-f). These results revealed that the physically cross-linked networks significantly improved the mechanical properties of the cellulose hydrogels. Whereas, the loose chemical crosslinking like as framework maintained the high toughness of hydrogel under large deformation (Figure 1c, f). Table 1.Water content and mechanical properties of cellulose hydrogels.

Sample

Molar ratios

wc

Tensile Test

Compressive Test

Εn

W σc εc Εc -3 EPI/AGU w/w% MPa % MPa MJ m MPa % MPa MPa LCH 1:1.33 98.2 0.07 59 0.03 0.01 0.09 77 0.01 0.04 TCH-1 1:9.28 88.5 2.26 119 4.09 1.29 4.97 72 0.81 1.58 TCH-2 1:4.64 90.1 1.76 120 2.13 0.97 5.74 81 0.36 1.41 TCH-3 1:3.09 91.8 1.50 134 0.99 0.84 6.32 82 0.48 1.20 TCH-4 1:2.32 91.5 1.32 150 0.89 0.91 6.25 87 0.25 0.95 TCH-5 1:1.86 91.3 1.23 160 0.81 0.88 5.08 86 0.20 0.92 TCH-6 1:1.55 91.5 0.69 139 0.48 0.45 3.03 84 0.18 0.67 TCH-7 1:1.33 89.2 0.45 116 0.26 0.23 1.98 84 0.05 0.23 wc, water content of the hydrogels; σt, εt and Εt, tensile strength, elongation at break, and Young’s modulus under tension; W, the fracture energy, a parameter that expresses one aspect of toughness; σc, εc and Εc, stress at fracture, fracture stain, and modulus under compression. Εn, the effective Young’s modulus under nanoindentation measurement.

σt

εt

Εt


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Figure 4. X-ray scattering results of TCH-5 hydrogels with different stretching strains. (a-b) 2D WAXS patterns of TCH-5 hydrogels with different stretching strains (0% and 130%). (c) Azimuthalintegrated intensity distribution curves of WAXS patterns, where 0o represents the perpendicular direction. (d-e) 2D SAXS patterns of TCH-5 hydrogels with different stretching strains (0% and 130%). (f) Azimuthal-integrated intensity distribution curves of SAXS patterns, where 0o represents the perpendicular direction.

AFM and SEM technologies were used to investigate the morphology of hydrogels (Figure S5). Obviously, LCH exhibited submicro- and nano-porous architecture on the surface and inner micropores (~2 µm) (Figure S5a). The pores on the surface of TCH became larger, and hierarchical structure constituted of nanofibers and submicrobundles could be observed on the surface and cross-section (Figure S5b). Besides, the diameter of cellulose nanofibers and pore sizes gradually reduced with the decrease dosage of EPI (Figure S5c, d; Table 1). These results indicated that the cellulose nanofibers constituted the physical networks through reversible hydrogen bonds, depending on the EPI content and acid treatment. The formation of the nanofibers and submicrobundles in the cellulose hydrogel networks was further confirmed by the full field transmission hard X-ray microscopy (TXM) (Movie S1). Therefore, ACS Paragon Plus Environment


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Figure 5. Mechanical properties of cellulose hydrogels. (a) Tensile stress-strain curves of LCH and TCHs. (b) Corresponding tensile stress-strain tests of LCH and TCHs during loading-unloading cycles of varying maximum stretching. (c) Representative tensile stress-strain curves of TCH-5 hydrogels during 10 times loading-unloading cycles at 40%, 100%, 140% and 160% strain, respectively. (d) Compressive stress-strain curves of LCH and TCHs. (e) Corresponding compressive stress-strain tests of LCH and TCHs during loading-unloading cycles of varying maximum stretching. (f) Representative compressive stress-strain tests of TCH-4 hydrogels during 10 times loading-unloading cycles at 50%, 60%, 70% and 80% strain, respectively. (g) Force-displacement curves of LCH and TCHs from a displacement-controlled nanoindenter machine (Piuma Nanoindenter by Optics11, Netherlands). (h-i) Representative effective Young’s modulus 3D map of LCH and TCH-5 from nano-indentation measurements, respectively. The grid scan was 10 × 10 points, and the point-to-point pitch is 20 µm.

the chemical networks as relatively lager framework, whereas the physical networks containing nanofibers and submicrobundles linked through strong hydrogen bonding interaction were integrated inside the chemical framework, jointly constituting the hierarchical architecture. To better understand the oriACS Paragon Plus Environment

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ented structure of the deformed cellulose hydrogels under external force, the hydrogels were stretched gradually and treated immediately by freeze-drying for AFM and TEM observations. Figure 2 displays the AFM and TEM images of the deformed TCH-5 specimens with tensile strains (ε) of 0%, 80%, 120%, and 160%, respectively. Interestingly, the cellulose submicrobundles were oriented along the direction of external force, and the orientation trend was gradually obvious with an increase of tensile strain, as shown in Figure 2a, b. Furthermore, the hierarchical structure in TCHs was also orientable during the compressive process (Figure S6). The cellulose hydrogel with layered porous structure appeared after compressing, indicating that the rearrangement and orientation of the cellulose nanofibers and submicrobundles occurred in the deformed hydrogels. As shown in Figure 2c, there were many broken networks in TCH-5 at strain of 160%, suggesting that the reversible physical networks in the hierarchical networks were fractured under strong external force. Thus, the reversible physical networks could efficiently dissipate mechanical energy through the breaking of “sacrificial bonds”, contributing to the toughness of materials. Deformation-Induced Anisotropy and Orientation Degree of Cellulose Hydrogels. Interestingly, TCHs could undergo multiform mechanical deformations (e.g., bending, twisting and knotting) and exhibit anisotropy, namely iridescent birefringence patterns under polarized light (Figure 3).The birefringence phenomenon in the deformed hydrogels induced by external force revealed that an anisotropic architecture appeared. The change of birefringence colors in hydrogels could be observed during the continuous deformation (Movie S2). Moreover, the birefringence patterns could be well reserved by drying the deformed hydrogel specimen (Figure S7). The orientation degree of the deformed cellulose hydrogels was captured with X-ray scattering techniques on a synchronization radiation facility. Figures 4a and S8 illustrate wide angle X-ray scattering (WAXS) images of TCH-5 specimen under different tensile strains. Without stretching, TCH-5 exhibited a nearly uniform diffraction pattern at all azimuthal angles on the (110) scattering plane, indicating that no orientation occurred in the native hydrogel. As it was gradually stretched from 30% to 130% (strain), the equatorial arcs became more and more apparent, indicating the alignment of hierarchical structure along the drawing direction. The WAXS patterns menACS Paragon Plus Environment


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tioned above were obtained by removing the backgrounds of water and air (Figure S9). From the corresponding intensity-azimuthal angle curves (Figure 4b and S8b), the full width at half maximum (FWHM) sharply decreased with the stretching process of hydrogel, proving that the orientation of the cellulose nanofibers and submicrobundles along the stretching direction occurred, as described in Figure 1c. To quantify the orientation degree of the stretching-deformed hydrogels, the Herman’s orientation parameter (fc) was calculated as the following equations =

(2)

< cos >=

$/

&

!"#

(3)

$/ & '!"#

where ( was azimuthal angle, and * (( ) was the 1-D intensity distribution along with the azimuthal angle after the subtraction of the background intensity. The average is calculated by integrating the intensity of specific 2θ diffraction peak along the ( , using the aforementioned equation. The fc of stretched TCH-5 specimen rose gradually to 0.81 with an increase of tensile strain to 130% (Figure S8c). After completely eliminating the influence of water, the dried gels which reserved the orientated structure were also monitored by the wide-angle X-ray diffraction (Figure S10). The anhydrous hydrogels exhibited a similar increasing tendency in orientation parameters along with the augmented tensile deformation, further confirming the high orientation of the hierarchical structure in the stretchingdeformed hydrogels. Small angle X-ray scattering (SAXS) was also used to characterize the structure of hydrogel under various tensile strains (Figure 4d-f; Figure S11, 12). For TCH-5 specimen without deformation, the uniform intensity rings appeared in SAXS patterns, reflecting an isotropic structure (Figure 4d). With an increase of deformation of hydrogel under uniaxial tensile stress, a sharp elongated equatorial streak appeared, showing anisotropy in ring intensity (Figure 4e; Figure S11a). It was revealed that the stressinduced orientation of the cellulose submicrobundles occurred along the stress direction, resulting in an anisotropic structure. The FWHM of azimuthal-integrated intensity distribution curves gradually narrowed with the increasing deformation of hydrogel (Figures 4f, S11b, c), which were consistent with the ACS Paragon Plus Environment

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results of WAXS. The orientation parameters of cellulose hydrogels (fc) were also calculated according to equations (2) and (3). As tensile strain increased from 0% to 130%, fc rose from 0.06 to 0.89 (Figure S11d). Furthermore, the morphology changes of hydrogels under compression deformation were investigated by SAXS from two directions (the beam was perpendicular or parallel to the compression direction) (Figure S12). When the direction of X-ray was perpendicular to the compression deformation, ovalization of SAXS patterns could be observed, due to the alignment and orientation of the cellulose submicrobundles. The fc values increased progressively from 0.0053 to 0.92 with an increase of the compressive strain from 0% to 85%, consistent with the SEM results (Figure S6). However, no obvious orientation behavior was found when the direction of X-ray was parallel to the compression direction, and fc values slightly increased from 0.014 to 0.098 as compressive strain increased from 0% to 85%. These results demonstrated strongly a radial orientation of hierarchical structure in the hydrogels. Thus, we could conclude that the movable cellulose nanofibers and submicrobundles integrated inside the large framework were easily synchronized with the networks to align along the stretching direction of hydrogels, leading to their anisotropic architecture. In our findings, the large chemical cross-linked framework played an important role in synchronizing alignment and orientation of the hierarchical networks in the hydrogels. Mechanical Properties of Cellulose Hydrogels. The stress-strain tests of LCH and TCHs were in detail conducted (Figure 5, S13; Table 1). The tensile strength and fracture energy of LCH were only 0.07 MPa and 0.01 MJ m-3 with an elongation at break of 59% (Figure S13a; Table 1). Remarkably, the fracture stress and fracture energy of TCH-7 reached 0.45 MPa and 0.23 MJ m-3, which were 5 times and 23 times higher than those of LCH, respectively. Besides, the ruptured strain of TCH-7 was 116%, which was 2 times that of LCH (Figure 5a and Table 1). The results further confirmed that the TCHs with the incorporation of physical networks containing nanofibers displayed a fulfilling improvement of both fracture stress and ruptured strain, compared with LCH. The results demonstrated that the cellulose nanofibers and submicrobundles in the physical networks reinforced significantly the TCHs. Besides, the ACS Paragon Plus Environment


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effects of chemical crosslinking on the mechanical properties of TCHs were also evaluated. Interestingly, the tensile strength of the TCHs increased from 0.45 MPa to 2.26 MPa with a decrease of the EPI/AGU molar ratio from 1:1.33 for TCH-7 to 1:1.98 for TCH-1. Usually, the mechanical strength of hydrogel networks can be enhanced by an increase of crosslinking density. However, in our findings, the cellulose hydrogels with relatively less chemical crosslinking allowed the higher mobility of cellulose nanofibers, which facilitated the formation of denser physical networks composed by hydrogen bonds linked crystallinity and nanofibers during acid treatment, leading to the better mechanical performance. The loading-unloading tensile tests of cellulose hydrogels are shown in Figures 5b and S11b. It was revealed that the enhanced mechanical properties of TCHs was attributed to the formation of hierarchical structure as well as better energy dissipation during the repeated loading-unloading tests (Figure 5c, S11c). For example, the dissipated energy of LCH was 1.03 kJ m-3 at the strain 50%, whereas TCH-5 could dissipate energy as much as 31.08 kJ m-3 at strain 50% and 601.86 kJ m-3 at strain 160%. These results further demonstrated that the physical networks in TCHs could efficiently dissipate energy through breaking of the hydrogen bonds under deformation. Moreover, the compressive stress-strain curves and loading-unloading compressive tests of LCH and TCHs were examined in detail (Figure 5d-f; S11d-f). TCH-5 exhibited the highest compressive strength (~6.32MPa) at the compressive strain of 87%, whereas LCH could only endure a maximum compressive stress of about 0.09 MPa at the ruptured strain of 77%. The compressive dissipation energy of TCH-4 at the strain of 60% was 114.51 kJ m-3, and 529.03 kJ m-3 at the strain of 80%, which was far higher than that of LCH (1.34 kJ m-3). In the view of above results, the mechanical properties of cellulose hydrogels were improved significantly as a result of the unique hierarchical network structure. Furthermore, the fracture stress, ruptured strain, and dissipated energy of TCHs were tunable by varying the EPI/AGU molar ratio. The Young’s modulus of TCHs calculated by tensile and compressive tests was 0.26~4.09 MPa and 0.05~0.81 MPa, respectively (see Table 1). To further understand the effective Young’s modulus (Εn) and its distribution on the hydrogels, nanoindentation technology was employed according to Oliver & ACS Paragon Plus Environment

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Pharr compliance method.24 Figure 5g shows the load-displacement curves of various cellulose hydrogel specimens. The stiffness of the TCH hydrogels evidently increased with the decrease of EPI/AGU molar ratio according to the slope of force-displacement curves. Εn was calculated by the slope of the initial portion of unloading curve according to equation (4): +, = // .

-

(4)

01234 156738

Where S is the slope of the initial portion of unloading curve reflecting the elastic unloading stiffness, r is the spherical indenter radius, and hmax depth respectively. Uniform distribution of Εn on LCH was observed with the average En value of 0.04 MPa, whereas the En value of TCH-7 rose to 0.23 MPa with uniform distribution (Figure 6h, S14). After slightly decreasing the EPI/AGU molar ratio, En of TCH-5 increased to 0.92 MPa (Figure 5i). Εn of TCHs ranged from 0.23 to 1.58 MPa increased with a decrease of EPI/AGU molar ratio. The trend was well consistent with those from tensile and compressive tests (see Table 1). Even if the water contents of TCHs were around 90%, TCHs still exhibited remarkable mechanical properties, including large elongation, high strength and high toughness, which stemmed from

the

hierarchical

structure

in

hydrogels.

Mechano-Responsive Behaviors of Cellulose Hydrogel. When the stretched hydrogel was placed between crossed polarizers and rotated from 0° to 90° (Figure 6a), the laser power passed through hydrogel increased and then decreased. Moreover, the laser power increased with the increase of stretching strain of hydrogels. These results further confirmed that the oriented hierarchical structure existed in the stretched hydrogels, leading to anisotropy (Figure 6b). The apparent birefringence of the stretched hydrogel could be observed at the rotation angle of 45°, but birefringence disappeared rapidly after removal of the stretching force from hydrogel (Figure 6c), indicating a sensitively mechano-responsive behavior. Similarly, the birefringence colors of the TCH-5 occurred under compressive deformation and disappeared after removing force, proving the speedily mechano-responsive behavior of hydrogels (Figure 7a; Movie S3). Interestingly, the birefringence colors of TCH hydrogels responded quickly with repeated loading-unloading experiment with a compressive strain of 5% for 1000 times, and immediately disACS Paragon Plus Environment


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appeared as soon as removal of external force, revealing stable mechano-responsive properties of the hydrogel (Figure 7b, c; Movie S4). To quantify the influence of external force on the properties of hydrogels, the mechano-responsive behavior of hydrogel under compression was detected by monitoring the variation of He-Ne laser power with polarizing laser detector (Figure 7d). Figure 7e and Movie S5 show typical on/off switching behavior of cellulose hydrogel when the polarized laser passed through hydrogel with/without deformation. Without external force, cellulose hydrogels exhibited isotropic structure, so the polarized laser could penetrate hydrogel and analyzer. The laser power was about 0.28 mW, indicating that the switch was on. Under the deformation, the cellulose nanofibers and submicrobundles aligned and oreinted rapidly in the network, leading to anisotropy. The polarized laser was then disturbed in the anisotropic networks and the depolarized laser was collected by analyzer.

Figure 6. (a) Schematic illustration of angle variation of stretched hydrogel between the crossed polarizers. (b) The depolarized laser power as a function of rotation angle (0°~90°) of stretched hydrogels with various tensile strain from 0 to 160%. (c) Polarized optical microscopy images of stretched cellulose hydrogel and normal cellulose hydrogel with the rotation angle of 45°. Scar bar: 500 µm.

The laser power was only 0.02 mW, indicating that the switch was off. The quickly responsive switching of cellulose hydrogel under small deformation (Strian: 5%) could make it have potential application as a soft tactile sensor to accurately detect small external force in tissue engineering. CONCLUSION


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Tough cellulose hydrogels were fabricated successfully using an ingenious strategy, where dense physical crosslinked networks consisted of nanofibers and submicrobundles was integrated inside the relatively large framework formed by loose chemical crosslinking, leading to the hierarchical architecture. Thus,

Figure 7. Mechano-responsive properties of TCH-5 hydrogels. (a) Photographs of cellulose hydrogel during the compressive process between crossed polarizers. Scar bar: 1 cm. (b) Color changes of cellulose hydrogel during successive stressed-release process for 1000 times with a strain of 5%. Scar bar: 1 cm. (c) Compressive stress-strain curves of hydrogel under 1000 loading-unloading cycles with a strain of 5%. (d) Schematic diagram for detection of laser power passed through the mechano-responsive of hydrogel. (e) The on/off switching performance of cellulose hydrogel.



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the densely physical networks could not only improve significantly the mechanical performance of the hydrogels, but dissipate efficiently mechanical energy through the reversible breaking of the strong hydrogen bonding interaction, whereas the chemical networks as framework could maintain the high toughness of hydrogel. The dense physical crosslinked networks were permeated in the relatively large framework, leading to easy synchronizing behavior of the double networks in the cellulose hydrogels. Under deformation, the densely physical networks synchronized easily to align and orient, leading to the rapidly mechano-responsive birefringence behaviors. The color of the deformed hydrogels was highly stable, even though the loading-unloading cycles were repeated for 1000 times. This work provided a novel strategy for developing tough polysaccharide-based hydrogel materials with deformation-induced anisotropy for tissue engineering applications. ASSOCIATED CONTENT Supporting Information The Supporting Information is available free of charge on the ACS Publications website at DOI:xxxx. Appearance of cotton linter pulps, cellulose solution and cellulose hydrogels; properties of cellulose dilute solution; solid state 13C NMR measurements; X-ray diffraction measurements; crosslinking density and mechanical properties of LCHs; morphology of TCHs; morphology of compressed TCHs; photographs of dried TCH specimens; WAXS and SAXS results of deformed TCHs and dried TCHs. AUTHOR INFORMATION Corresponding Authors Email: [email protected] (L. Z.) Email: [email protected] (C. C.) ORCID: Lina Zhang: 0000-0003-3890-8690 ACS Paragon Plus Environment

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Chunyu Chang: 0000-0002-3531-5964 Notes The authors declare no competing financial interest. ACKNOWLEDGMENTS This work was supported by the Major Program of Natural Science Foundation of China (21334005), the Major International (Regional) Joint Research Project of National Natural Science Foundation of China (21620102004), and Hubei Province Science Foundation for Youths (2015CFB499). REFERENCES 1. Sun, T. L.; Kurokawa, T.; Kuroda, S.; Ihsan, A. B.; Akasaki, T.; Sato, K.; Haque, M. A.; Nakajima, T.; Gong, J. P. Physical Hydrogels Composed of Polyampholytes Demonstrate High Toughness and Viscoelasticity. Nat. Mater. 2013, 12, 932-937. 2. Okumura, Y.; Ito, K. The Polyrotaxane Gel: A Topological Gel by Figure-of-Eight Cross-Links. Adv. Mater. 2001, 13, 485-487. 3. Tamesue, S.; Ohtani, M.; Yamada, K.; Ishida, Y.; Spruell, J. M.; Lynd, N. A.; Hawker, C. J.; Aida, T. Linear Versus Dendritic Molecular Binders for Hydrogel Network Formation with Clay Nanosheets: Studies with ABA Triblock Copolyethers Carrying Guanidinium Ion Pendants. J. Am. Chem. Soc. 2013, 135, 15650-15655. 4. Gong, J. P.; Katsuyam, Y.; Kurokawa, T.; Osada, Y. Double-network Hydrogels with Extremely High Mechanical Strength. Adv. Mater. 2003, 15, 1155-1160. 5. Sun, J. -Y.; Zhao, X. H.; Illeperuma, W. R. K.; Chaudhuri, O.; Oh, K. H.; Mooney, D. J.; Vlassak, J. J.; Suo, Z. Highly Stretchable and Tough Hydrogels. Nature 2012, 489, 133-136. 6. Zhou, J.; Du, X. W.; Gao, Y.; Shi, J. F.; Xu, B. Aromatic–aromatic Interactions Enhance Interfiber Contacts for Enzymatic Formation of a Spontaneously Aligned Supramolecular Hydrogel. J. Am. Chem. Soc. 2014, 136, 2970-2973. ACS Paragon Plus Environment


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Deformation Drives Alignment of Nanofibers in Framework for

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