Direct Comparison of Three Buckling-Based Methods to Measure the Elastic Modulus of Nanobiocomposite Thin Films.

To engineer tunable thin-film materials, the accurate measurement of their mechanical properties is crucial. However, characterizing the elastic modulus with current methods is particularly challenging for sub-micrometer thick films and hygroscopic materials because they are highly sensitive to environmental conditions and most methods require free-standing films which are difficult to prepare. In this work, we directly compared three buckling-based methods to determine the elastic moduli of supported thin films: (1) biaxial thermal shrinking, (2) uniaxial thermal shrinking, and (3) the mechanically compressed, strain-induced elastic buckling instability for mechanical measurements (SIEBIMM) method. Nanobiocomposite model films composed of cellulose nanocrystals (CNCs) and polyethyleneimine (PEI) were assembled using layer-by-layer deposition to control composition and thickness. The three buckling-based methods yielded the same trends and comparable values for the elastic moduli of each CNC-PEI film composition (ranging from 15 to 44 GPa, depending on film composition). This suggests that the methods are similarly effective for the quantification of thin-film mechanical properties. Increasing the CNC content in the films statistically increased the modulus; however, increasing the PEI content did not lead to significant changes. For the CNC-PEI system, the standard deviation of elastic moduli determined from SIEBIMM was 2-4 times larger than that for thermal shrinking, likely due to extensive cracking due to the different stress applied to the film when subjected to compression of a relaxed substrate versus the shrinking of a pre-strained substrate. These results show that biaxial thermal shrinking is a reliable method for the determination of the mechanical properties of thin films with a simple implementation and analysis and low sensitivity to small deviations in the input parameter values, such as film thickness or substrate modulus.

Multi-scale structuring of cell-instructive cellulose nanocrystal composite hydrogel sheets via sequential electrospinning and thermal wrinkling.

Structured hydrogel sheets offer the potential to mimic the mechanics and morphology of native cell environments in vitro; however, controlling the morphology of such sheets across multiple length scales to give cells consistent multi-dimensional cues remains challenging. Here, we demonstrate a simple two-step process based on sequential electrospinning and thermal wrinkling to create nanocomposite poly(oligoethylene glycol methacrylate)/cellulose nanocrystal hydrogel sheets with a highly tunable multi-scale wrinkled (micro) and fibrous (nano) morphology. By varying the time of electrospinning, rotation speed of the collector, and geometry of the thermal wrinkling process, the hydrogel nanofiber density, fiber alignment, and wrinkle geometry (biaxial or uniaxial) can be independently controlled. Adhered C2C12 mouse myoblast muscle cells display a random orientation on biaxially wrinkled sheets but an extended morphology (directed preferentially along the wrinkles) on uniaxially wrinkled sheets. While the nanofiber orientation had a smaller effect on cell alignment, parallel nanofibers promoted improved cell alignment along the wrinkle direction while perpendicular nanofibers disrupted alignment. The highly tunable structures demonstrated are some of the most complex morphologies engineered into hydrogels to-date without requiring intensive micro/nanofabrication approaches and offer the potential to precisely regulate cell-substrate interactions in a "2.5D" environment (i.e. a surface with both micro- and nano-structured topographies) for in vitro cell screening or in vivo tissue regeneration. STATEMENT OF SIGNIFICANCE: While structured hydrogels can mimic the morphology of natural tissues, controlling this morphology over multiple length scales remains challenging. Furthermore, the incorporation of secondary morphologies within individual hydrogels via simple manufacturing techniques would represent a significant advancement in the field of structured biomaterials and an opportunity to study complex cell-biomaterial interactions. Herein, we leverage a two-step process based on electrospinning and thermal wrinkling to prepare structured hydrogels with microscale wrinkles and nanoscale fibers. Fiber orientation/density and wrinkle geometry can be independently controlled during the electrospinning and thermal wrinkling processes respectively, demonstrating the flexibility of this technique for creating well-defined multiscale hydrogel structures. Finally, we show that while wrinkle geometry is the major determinant of cell alignment, nanofiber orientation also plays a role in this process.

Bioinspired Thermoresponsive Xyloglucan-Cellulose Nanocrystal Hydrogels.

Thermoresponsive hydrogels present unique properties, such as tunable mechanical performance or changes in volume, which make them attractive for applications including wound healing dressings, drug delivery vehicles, and implants, among others. This work reports the implementation of bioinspired thermoresponsive hydrogels composed of xyloglucan (XG) and cellulose nanocrystals (CNCs). Starting from tamarind seed XG (XGt), thermoresponsive XG was obtained by enzymatic degalactosylation (DG-XG), which reduced the galactose residue content by ∼50% and imparted a reversible thermal transition. XG with native composition and comparable molar mass to DG-XG was produced by an ultrasonication treatment (XGu) for a direct comparison of behavior. The hydrogels were prepared by simple mixing of DG-XG or XGu with CNCs in water. Phase diagrams were established to identify the ratios of DG-XG or XGu to CNCs that yielded a viscous liquid, a phase-separated mixture, a simple gel, or a thermoresponsive gel. Gelation occurred at a DG-XG or XGu to CNC ratio higher than that needed for the full surface coverage of CNCs and required relatively high overall concentrations of both components (tested concentrations up to 20 g/L XG and 30 g/L CNCs). This is likely a result of the increase in effective hydrodynamic volume of CNCs due to the formation of XG-CNC complexes. Investigation of the adsorption behavior indicated that DG-XG formed a more rigid layer on CNCs compared to XGu. Rheological properties of the hydrogels were characterized, and a reversible thermal transition was found for DG-XG/CNC gels at 35 °C. This thermoresponsive behavior provides opportunities to apply this system widely, especially in the biomedical field, where the mechanical properties could be further tuned by adjusting the CNC content.

Xyloglucan Structure Impacts the Mechanical Properties of Xyloglucan-Cellulose Nanocrystal Layered Films-A Buckling-Based Study.

Interactions between polysaccharides, specifically between cellulose and hemicelluloses like xyloglucan (XG), govern the mechanical properties of the plant cell wall. This work aims to understand how XG molecular weight (MW) and the removal of saccharide residues impact the elastic modulus of XG-cellulose materials. Layered sub-micrometer-thick films of cellulose nanocrystals (CNCs) and XG were employed to mimic the structure of the plant cell wall and contained either (1) unmodified XG, (2) low MW XG produced by ultrasonication (USXG), or (3) XG with a reduced degree of galactosylation (DGXG). Their mechanical properties were characterized through thermal shrinking-induced buckling. Elastic moduli of 19 ± 2, 27 ± 1, and 75 ± 6 GPa were determined for XG-CNC, USXG-CNC, and DGXG-CNC films, respectively. The conformation of XG adsorbed on CNCs is influenced by MW, which impacts mechanical properties. To a greater degree, partial degalactosylation, which is known to increase XG self-association and binding capacity of XG to cellulose, increases the modulus by fourfold for DGXG-CNC films compared to XG-CNC. Films were also buckled while fully hydrated by using the thermal shrinking method but applying the heat using an autoclave; the results implied that hydrated films are thicker and softer, exhibiting a lower elastic modulus compared to dry films. This work contributes to the understanding of structure-function relationships in the plant cell wall and may aid in the design of tunable biobased materials for applications in biosensing, packaging, drug delivery, and tissue engineering.

Dynamically Evolving Surface Patterns through Light-Triggered Wrinkling Erasure.

For many applications, it is imperative that changes in polymer surface topography, especially periodic patterns, can be triggered on command by a well-defined remote signal. In this contribution, we report a light-induced cascade of changes in wrinkling wavelengths on thin polymer layers supported by an elastomeric substrate under tensile stress. Through the applied supramolecular design, the effect of varying the ratio of light-active and light-passive components can be easily assessed, and it is shown that both the cascade type as well as the rate of the progress of the dynamic light-induced changes can be tuned by this ratio as well as by the light intensity. Furthermore, for the reported phenomena to occur, nominally only every 20th polymer repeat unit needs to be occupied by a chromophore, which makes the conversion of the sub-nanometer photoisomerization reaction into 10 µm scale changes of periodic surface patterns extremely efficient.

Hydrazide-Derivatized Microgels Bond to Wet, Oxidized Cellulose Giving Adhesion Without Drying or Curing.

Hydrazide-derivatized poly(N-isopropylacrylamide-co-acrylic acid) microgels gave strong adhesion to wet, TEMPO oxidized, regenerated cellulose membranes without a drying or heating step. Adhesion was attributed to hydrazone covalent bond formation with aldehyde groups present on the cellulose surfaces. This is one of only three chemistries we have found that gives significant never-dried adhesion between wet cellulose surfaces. By contrast, for cellulose joints that have been dried and heated before wet testing, the hydrazide-hydrazone chemistry offers no advantages over standard paper industry wet strength resins. The design rules for the hydrazide-microgel adhesives include: cationic microgels are superior to anionic gels; the lower the microgel cross-link density, the higher the adhesion; longer PEG-based hydrazide tethers offer no advantage over shorter attachments; and, adhesion is independent of microgel diameter. Many of these rules were in agreement with predictions of a simple adhesion model where the microgels were assumed to be ideal springs. We propose that the unexpected, high cohesion between neighboring microgels in multilayer films was a result of bond formation between hydrazide groups and residual NHS-carboxyl esters from the preparation of the hydrazide microgels.