Collagen's triple helix structure is fundamental to the mechanical integrity of bodily tissues, and its stability is vital for optimal physiological function. While the stabilization mechanisms of simple, symmetric collagen homotrimers have been previously studied, the hydration dynamics of native-like collagen heterotrimers which comprise the vast majority of human collagens remain largely unexplored. In this study, we investigated the complex role of water in stabilizing heterotrimeric collagen model peptides (CMPs) using extensive molecular dynamics (MD) simulations. We examined structurally diverse AAB-type and ABC-type heterotrimers exhibiting varying experimental stability profiles to investigate how sequence asymmetry influences hydration organization and collagen stability. Our simulations revealed that water molecules dynamically organize around these complex structures, forming topological water networks (TWNs) via intermolecular hydrogen bonds. Quantitative analyses of peptide-water interactions, hydration-water dynamics, and normalized TWN counts, together with residue- and chain-resolved TWN characterization, demonstrated that hydration-shell organization is strongly sequence dependent and differs among heterotrimers with distinct thermal stabilities. In particular, the more thermally stable heterotrimers exhibited a greater propensity for hydration-shell water molecules to participate in cyclic TWNs, accompanied by reduced hydration-water mobility and favorable interchain hydrogen-bonding interactions. These findings extend our previous observations on collagen homotrimers and provide new atomistic insight into the sequence-dependent organization of hydration water surrounding collagen heterotrimers, suggesting that organized hydration water contributes to collagen stability in concert with interchain hydrogen bonding and hydration-water dynamics rather than acting as an independent stabilization mechanism.
Due to its dense crystalline structure and extensive hydrogen bonding network, cellulose exhibits limited reactive accessibility and reactivity, which severely restricts the reaction efficiency, degree of substitution, and uniformity of cellulose modifications such as carboxymethylation. To overcome these limitations, this study developed a novel glycerol-based pretreatment strategy to disrupt the hydrogen bonding network of bacterial cellulose (BC), thereby improving its reactive accessibility. The results confirmed that glycerol pretreatment significantly enhanced the reactivity of BC. The reaction time for preparing carboxymethyl cellulose (CMC) from glycerol-pretreated powdered BC was reduced from 2 to 1 h, representing a 100% improvement in reaction efficiency. In addition, glycerol pretreatment facilitated the exposure of hydroxyl groups in BC, thereby increasing the degree of substitution of CMC from 0.297 to 0.433 and from 0.427 to a peak value of 0.626, an approximately 46% enhancement. In addition, the CMC films prepared from glycerol-pretreated BC exhibited significantly improved light transmittance, reaching over 95% in the wavelength range of 300-1000 nm. This study provides theoretical support and technical guidance for the efficient production of CMC, expands its application potential in high-end fields, and promotes the high-value utilization of cellulose resources.
The biological function of intrinsically disordered proteins is frequently coupled to short linear motifs, which serve as protein binding sites. This goes often hand in hand with local transient structural element formation. The intrinsically disordered C-terminal region of the multi-site docking protein Grb2-associated binder 1 (Gab1) contains several well-characterized phosphotyrosine pairs, whereas the Tyr162 and Tyr183 epitopes have remained practically unstudied. Here, we combine computational prediction, circular dichroism and high-resolution NMR spectroscopy (chemical shift and relaxation analyses) to structurally characterize a Gab1 fragment containing residues 142-203. The here determined NMR structure shows an α-helix for residues Pro180 to Ile187, with Tyr183 positioned centrally, while all other parts of the fragment, including the region around Tyr162, are disordered. This helical conformation is maintained upon changes in pH, variation of peptide lengths and phosphorylation status. The inherent helix around Tyr183 distinguishes the local structural environment of Tyr162 and Tyr183 within the intrinsically disordered Gab1 tail and provides a structural framework for future studies on interaction partners and potential functional roles of this structural motif.
Green fluorescent protein (GFP) was originally obtained from Aequorea victoria and was successfully synthesized through recombinant technology. In this study, we revealed novel, fascinating insights into the properties of this well-known GFP by examining a traditional GFP separation process involving aqueous two-phase systems and several liquid chromatographs. We investigated the impact of organic solvents and salts on the conformation of GFP by measuring its fluorescence intensity and circular dichroism spectrum, and then proposed an optimized process to improve the yield of fluorescent GFP. We have noticed that fluorescent GFP migrates anomalously to ~37 kDa on non-heating SDS-PAGE due to retention of its compact β-barrel conformation, whereas the commonly reported 27 kDa migration corresponds to the fully heat-denatured, non-fluorescent form. Nevertheless, the most exciting discovery is that the purified GFP exhibited a remarkable ability to self-assemble into an intriguing fluorescent leaf-like structure upon evaporation-concentration under ambient conditions. To our knowledge, this represents the first direct visualization of such macroscopic fluorescent leaf-like assemblies formed by recombinant GFP, with fluorescence retained throughout the assembled morphology. This captivating phenomenon may have promising implications for protein-based nanomaterials and will inspire further exploration of protein self-assembly nanotechnology.
The impact of initial moisture content (1.5%-20%) on the ball-milling of pea starch was investigated across multiple structural levels. Scanning electron microscopy revealed moisture-dependent deformation mechanisms: plastic flow and agglomeration in high-moisture granules versus pseudo-brittle fracture in low-moisture samples. Despite these differences, XRD-detectable crystallinity was eliminated within 5 min of treatment, independent of initial moisture content. Fourier transform infrared spectroscopy confirmed the loss of short-range molecular order, including helical and hydrogen-bonded contributions. Gel permeation chromatography demonstrated more severe molecular degradation in low-moisture starch, suggesting a possible two-stage degradation behavior, with a rapid initial decrease of molar mass followed by a slower regime as the material becomes predominantly amorphous. Consequently, ball-milling enhanced solubility in both cold (30°C) and hot (90°C) water, increasing it from less than 1% and ~25% to 60%-70%, respectively. The increased cold-water solubility is explained by the loss of crystallinity. In contrast, the rise in hot-water solubility indicates a more profound structural disintegration. We propose that this is related to the mechanochemical degradation of amylopectin, which may suppress the formation of amylopectin-rich remnants during gelatinization and thereby promote dissolution.
Tissue patches are biomaterial-based structures designed to support the repair of damaged or functionally impaired tissues and are required to exhibit biocompatibility, mechanical integrity, and suitable surface characteristics. In this study, poly(vinyl alcohol) (PVA), chitosan (Chi), and hyaluronic acid (HA)-based composite films reinforced with zeolite (0-0.5% w/v) were developed and evaluated as potential tissue patch materials. The incorporation of zeolite significantly influenced the physicochemical and mechanical properties of the films. The elastic modulus decreased from 293.78 ± 64.47 N/mm2 for the zeolite-free film to 106.21 ± 9.50 N/mm2 at the highest zeolite content, indicating tunable flexibility. Water contact angle values increased from 46.09° to 67.23°, while maintaining overall hydrophilicity. The films exhibited rapid swelling behavior, reaching equilibrium within 30 min, and demonstrated controlled biodegradation with mass losses exceeding 75% after 42 days. Biological evaluations showed that all formulations maintained cell viability above 70%, with values ranging from 87.26% to 78.63%, and supported cell adhesion, confirming their biocompatible nature. The novelty of this study lies in demonstrating that low-concentration zeolite incorporation enables controlled tuning of mechanical, surface, and biological properties within a single PVA-Chi-HA system, without compromising biocompatibility. These findings highlight the potential of zeolite-reinforced composite films as multifunctional and customizable tissue patch candidates for tissue engineering applications.
This study investigated the effects of directly incorporating untreated rosehip (RoHi) powder into electrospun poly(lactic acid)/poly(3-hydroxybutyrate-co-4-hydroxybutyrate) (PLA/P(3HB-co-4HB)) (70/30, w/w) blends, focusing on the structural, morphological, mechanical, wettability, and biological properties of resulting nanofibrous mats. The average fiber diameter of pure nanofibers was 776 nm, while RoHi addition resulted in diameters ranging from 468 nm (1% RoHi) to 1496 nm (5% RoHi). FTIR analysis confirmed that the characteristic functional groups of RoHi were preserved within the polymer matrix without chemical interaction. The water contact angle decreased from 136.45° for the pure mat to 116.11° for nanofibers containing 5% RoHi, leading to an increase in liquid adsorption capacity from 1.49% to 249.38%. Cytotoxicity results demonstrated that all samples were biocompatible, with the highest cell viability (148.91%) observed for nanofibers containing 1% RoHi. In vitro scratch assays revealed enhanced fibroblast migration for RoHi-containing nanofibers, particularly within the first 24 h, with the fastest wound closure observed at 3% and 5% RoHi contents. DPPH analysis showed increased antioxidant activity from 27.68% to 63.84%, whereas no antibacterial activity was detected against Staphylococcus aureus and Pseudomonas aeruginosa. Overall, RoHi-doped nanofibers exhibit promising potential as wound dressing materials considered suitable for use in chronic wounds due to their antioxidant activity and bio-responsive wound-healing performance.
Liraglutide, a glucagon-like peptide-1 receptor agonist, is widely used as a therapeutic macromolecule for the treatment of type 2 diabetes; however, its large-scale production is limited by high manufacturing costs and the frequent occurrence of closely related deletion impurities during synthesis. In the present study, we describe a novel, impurity-controlled, and industrially feasible synthetic strategy for liraglutide and its sequence-modified analogs. This method utilizes solution-phase incorporation of Pal-γ-Glu-OtBu in combination with a preassembled Boc-His (Boc)-Ala-Glu (OtBu)-OH tripeptide fragment, effectively minimizing the formation of des-His, des-Ala, and des-Glu impurities. The optimized protocol enabled the production of liraglutide and its analogs with consistent isolated yields and high chromatographic purity (> 95% by RP-HPLC), suitable for subsequent biological evaluation. The antidiabetic potential of liraglutide and three sequence-modified analogs, Lira (Trp-O25), Lira (desGly31), and Lira (Glu17), was evaluated using a Drosophila melanogaster model of high-sucrose diet induced diabetes. Among these, Lira (Glu17) exhibited the most pronounced metabolic improvements, significantly reducing free glucose (p < 0.001), trehalose (p < 0.001), and triglyceride levels (p < 0.001) when compared to diabetic controls. Furthermore, this analog effectively decreased lipid accumulation and reactive oxygen species in larval gut tissues and enhanced locomotor performance in both larva and adult flies. While Lira (Trp-O25) also demonstrated beneficial effects, Lira (desGly31) showed comparatively limited efficacy. Collectively, this study presents a cost-effective and impurity-controlled synthetic platform for liraglutide production and identifies Lira (Glu17) as a promising analog with enhanced antidiabetic activity, offering valuable insights for peptide manufacturing and GLP-1-based therapeutic development.
Chronic wounds require multifunctional dressings capable of simultaneously controlling infection, managing exudate, and promoting tissue regeneration. This study develops CuO nanoparticles incorporated alginate/chitosan hydrogel dressing reinforced onto nonwoven fabrics of pure cotton, pure hemp, and 50:50 cotton: hemp blend fabricated via full factorial design for wound dressing applications. The composites were characterized by surface morphology, chemical structure, mechanical properties, antibacterial activity, air permeability, and water absorbency. Morphological analysis confirms uniform hydrogel coating and successful CuO deposition. Mechanical strength and water absorbency increased with hydrogel concentration, with a maximum tensile strength of 115.61 N for the hemp reinforced sample at 2%. All CuO-loaded hydrogel composite dressings exhibited effective antibacterial performance. Although higher hydrogel concentrations reduce air permeability, blended substrates achieve a balanced combination of strength, breathability, and exudate management. This eco-friendly, cost-effective composite demonstrates strong potential for infection control for wound healing applications.
This study aimed to prepare six different metronidazole-loaded film formulations by 3D printing for periodontal treatment. Chitosan (CH) and alginate (ALG) gels were prepared separately and printed layer-by-layer, varying polymer type and number of layers. Polyelectrolyte complex (PEC) gels were prepared by mixing CH and ALG (1:1) at different polymer concentrations to investigate the effect of complexation strength on film performance. Films were characterized by thickness measurements, drug content analysis, Micro-CT and SEM imaging, swelling, adhesion, antimicrobial activity, release kinetics, and cumulative drug release. Increasing ALG layers significantly enhanced swelling and accelerated metronidazole release, whereas CH-dominant films exhibited lower swelling and slower release. Overall, PEC films modified the drug release profile. Layer-by-layer printing promoted more uniform diffusion, whereas stronger polyelectrolyte complexation slowed drug release due to increased structural irregularity. All films were confirmed to exhibit microbiological activity. Micro-CT analysis in conjunction with SEM enabled precise evaluation of the layered film structure produced by 3D printing and detailed assessment of layer definition, internal heterogeneity, and their relationship with swelling and drug release behaviour. These findings suggest that tailoring the structural design via polymer layering or complexation represents a promising strategy for local drug delivery systems in periodontal therapy.
Incorporating ferulic acid (FA) into a polymeric hydrogel impairs its bioactive properties. The synthesis used in this study involved radical-mediated surface grafting of nanocellulose for 24 h, using 2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO) to oxidize cellulose nanofibers (oCNF) and phosphorylated cellulose nanofibers (pCNF), resulting in total polyphenol contents of 30.6 and 6.3 mg/g, respectively. Spectroscopic analyses of oCNF included solid-state high-power decoupling with magic-angle spinning nuclear magnetic resonance (13C HPDEC/MAS NMR), Fourier transform infrared (FTIR) spectroscopy, and X-ray photoelectron spectroscopy (XPS), confirming that FA was successfully grafted onto the backbone through ester linkages on the nanofibril surface. Rheology testing and morphological analysis via transmission electron microscopy (TEM) further supported these findings. TEM imaging revealed that the grafting process preserved the nanoscale morphology, yielding fibrils with an average diameter of approximately 28.2 ± 9.3 nm. Key aspects affecting FA grafting include the colloidal stability of the CNF suspension, evaluated by zeta potential, and surface activation via oxidation. In this study, FA was grafted onto oCNF through a radical-induced reaction to enhance their functional performance, enabling the development of functional and sustainable hydrogels with high antioxidant capacity (85.62% ± 1.65%).
Multilayer films of bovine serum albumin (BSA) and chondroitin sulfate (CS) were assembled on gold substrates at pH 4.2 using the layer-by-layer (LbL) method, and their structural evolution was investigated via surface plasmon resonance (SPR) spectroscopy. To elucidate the role of adsorption kinetics in multilayer buildup, BSA adsorption times of 15, 30, 60, and 120 min (for each BSA adsorption cycle) were examined, while the CS adsorption time was kept constant at 15 min. Analysis of SPR reflectivity curves revealed two distinct growth regimes for all cases, an initial exponential followed by a linear regime. Longer BSA adsorption time promoted deeper chain infiltration, extending the exponential region and shifting the exponential-to-linear transition to higher bilayer numbers. In parallel, evaluation of the average dielectric constant of the polymer layer enabled quantification of the film volume fraction, which initially increased linearly before reaching a saturation plateau. The evolution of the adsorbed mass was inferred from the combined analysis of film thickness and volume fraction, revealing two distinct growth cases depending on the adsorption time. These cases reflect different relationships between the transition from exponential to linear growth and the saturation of the volume fraction. These findings demonstrate that BSA adsorption time is a key kinetic parameter governing chain diffusion, film compaction, and overall multilayer architecture. The results provide insight into the physicochemical mechanisms that dictate biopolymer film growth and offer a route for tuning the structural and optical properties of protein-polysaccharide coatings.
In recent years, there has been increasing interest in developing functional bioink systems that better replicate biological and biochemical microenvironments while maintaining high print fidelity and cell viability. This study reports the development and characterization of gelatin-chitosan-based bioinks reinforced with hexagonal boron nitride nanosheets (BNNSs) for extrusion-based 3D bioprinting. Structural, chemical, and morphological analyses demonstrated the successful incorporation of BNNSs without the formation of new chemical bonds, while minor shifts in amide bands indicated enhanced hydrogen bonding and physical interactions. Rheological studies revealed that gelatin concentration was the primary factor governing viscosity, whereas BNNSs provided a composition-dependent reinforcing effect. Notably, at a low shear rate (0.001 s-1), viscosity increased from 2865 Pa·s for low concentration gelatin bioinks to 9322 Pa·s for BNNS-containing high concentration formulations, representing more than a threefold increase. In contrast all formulations exhibited low viscosities below 3 Pa·s at 100 s-1, confirming favorable extrusion behavior. Viscoelastic analysis further showed lower tan δ values for high gelatin content formulations, indicating elastic-dominant behavior and improved shape retention at printing temperatures. BNNS incorporation slightly reduced the glass transition temperature by approximately 10°C while preserving blend compatibility, contributing to enhanced thermal responsiveness and more uniform temperature distribution during printing. Printability analysis demonstrated that BNNSs improved shape fidelity in high-viscosity formulations, yielding printability index (Pr) values close to unity (Pr ≈ 0.94) and stable filament formation. Swelling and degradation studies showed that BNNS-containing 7.5% gelatin scaffolds exhibited reduced swelling and retained approximately 70% structural integrity after 14 days, whereas low-polymer formulations underwent rapid degradation. Cell viability assessments confirmed improved fibroblast adhesion and proliferation on BNNS-containing scaffolds. The incorporation of BNNSs improves the rheological and thermal characteristics of the gelatin-chitosan bioinks and positively influences cell response. These findings suggest that BNNS-containing formulations provide a more stable and thermally responsive printing platform.
With rapid development of material science, application of lignocellulose and agricultural wastes to produce green and functional packaging materials has received much interest. Herein, tobacco stem cellulose nanocrystals (TC) were firstly prepared via facile and mild formic acid/choline chloride (FA/ChCl) deep eutectic solvent (DES) treatment. Systematic comparison of the TC nanomaterials with conventional cotton cellulose nanocrystals (CNCs) were performed from nanoscale to the applicable functions of the prepared films. Tobacco stem showed notable delignification after the FA/ChCl treatment with well-maintaining of xylan matrix, which induced more sphere (with average width of ~32 nm) and shorter nanocrystals after homogenization. The treatment also caused a series of crystalline structural variations, including reduced crystallinity and crystallite size, favorable for promoting molecular flexibility of TC. Vacuum-assisted drying induced the fabrication of the film materials. Even though TC films had less visible iridescence than CNCs films, they exhibited satisfying mechanical strength promotion, with breakage of ~30 MPa, and the elongation of ~14%, mainly due to the reduced rigidity, crystallinity and remaining xylan matrix. Therefore, the present study provided a novel approach to controllable fabricating high-quality cellulose nanocrystals for film materials fabrication, as well as a facile strategy for high-value added utilization of tobacco stem biomass.
Cellulose derived from ascidians (tunicates) is distinguished from plant-based counterparts by its marine origin, with high crystallinity, and complex hierarchical architecture. However, quantitative structure-property relationships governing its performance in bioplastic applications remain underexplored. Here, cellulose isolated from three ascidian species Ascidia sp. (T1), Herdmania cf. pallida (T2), and Ascidia sydneiensis (T3) was systematically characterized. X-ray diffraction reveals crystallinity indices (CrI) of 48% (T1) and 60% (T2, T3), the latter approaching values reported for highly ordered systems such as bacterial cellulose. Thermogravimetric analysis demonstrates species-dependent thermal stability, with maximum degradation temperatures of 345°C (T1) versus 400°C-401°C (T2, T3). Notably, T2 and T3 exhibit thermal behavior comparable to microcrystalline and bacterial cellulose, despite CrI values lower than those systems, indicating that hydrogen-bonding density and microfibrillar order govern thermal resilience. Scanning electron microscopy reveals distinct microfibrillar architectures, ranging from highly branched networks to compact laminar structures, which govern water interaction and mechanical response. Water absorption varies markedly by species: T1 and T3 absorb 2200-2400 wt% within 10 min, consistent with their branched, open fibrillar morphologies, whereas T2 absorbs only 1200 wt%, reflecting a compact lamellar microstructure that restricts water diffusion. Hydrolytic degradation after 28 days in neutral water remains minimal across all samples, confirming exceptional resistance to hydrolytic scission under mild conditions. Bioplastics fabricated from these celluloses exhibit tensile strengths of 1-4 MPa, directly correlating with microstructural packing. Collectively, these results establish that ascidian cellulose is the combination of thermal stability up to 400°C, tunable water affinity (1200-2400% absorption), and hydrolytic resistance (1-9% loss over 28 days) arises from species-specific interactions between crystallinity, hydrogen bonding, and microfibrillar architecture. This positions ascidian-derived cellulose as a distinct marine macromolecular scaffold for sustainable bioplastics where controlled water interaction and structural durability are required. In general, it is established the relationship between the biological origin, hierarchical structure, and macroscopic properties of tunicate cellulose, highlighting its potential as a marine-derived macromolecular building block suitable for sustainable bioplastics applications.
This study reports the fabrication and comprehensive characterization of multicomponent silver-doped zinc oxide (Ag/ZnO)-poly(vinyl alcohol)/alginate(PVA/Alg) biocomposite hydrogels enriched with clove oil (CO), prepared via a scalable and solvent-free freeze-thaw (F-T) process. The F-T treatment generated physically cross-linked hydrogel networks with preserved structural integrity and favorable mechanical performance. Morphological analysis revealed a homogeneous and interconnected porous architecture at low CO contents, while contact-angle measurements confirmed hydrophilic surfaces with composition-dependent wettability. X-ray diffraction and Fourier transform infrared spectroscopy verified the successful incorporation of Ag/ZnO and CO into the PVA/Alg matrix without disrupting the overall polymer network. Barrier performance was strongly governed by CO content. Increasing CO loading reduced water vapor transmission through the formation of hydrophobic domains, while oxygen permeability reached a maximum at 1 wt.% CO, highlighting tunable mass transport behavior. The hydrogels exhibited rapid swelling, sustained hydration stability for up to 72 h, and a composition-dependent CO release behavior. Collectively, these findings elucidate clear structure-processing-property relationships in Ag/ZnO-incorporated PVA/Alg hydrogels containing CO and demonstrate their promise as multifunctional materials for further investigation in wound dressing applications.
Hydrogels based on thiol-ene step-growth chemistry have gained increased attention due to their superior properties over the currently standard materials based on chain growth polymerization. In the thiol-ene reaction, a crosslinker with at least two thiol groups is necessary for network formation. Many currently used crosslinkers exhibit cytotoxic potential, are non-biodegradable, or involve toxic chemicals and relatively complicated procedures in their synthesis, thus hindering their broader application. As an alternative, the use of a protein (fibrinogen gamma chain, FGG) recombinantly expressed in Escherichia coli was investigated. The FGG is part of the multimeric fibrinogen involved in hemostasis. This protein complex is stabilized by disulfide crosslinking. This presence of cysteines in the sequence makes FGG a promising candidate as a thiol donor in thiol-ene reactions. It was shown for the first time, that a cysteine-containing protein expressed in E. coli was capable of forming hydrogels with norbornene functionalized gelatin. An increase in FGG concentration led to higher gel stiffness and a decrease in the swelling ratio. Furthermore, the material exhibited cell adhesive properties and biocompatibility. Overall, a proof-of-principle could be achieved, opening up the use of recombinant proteins without further modifications as crosslinkers in thiol-ene based hydrogels, providing a cost-effective, safe, and scalable material source.
The modification of starches by cationization is a fundamental physicochemical process aimed at improving their physicochemical properties and expanding their industrial applications. Traditionally, this modification is associated with long duration, high energy consumption, and waste generation. This article proposes a method based on reactive extrusion (REX) as a sustainable alternative for modifying corn (Zea mays everta) and chayote roots (Sechium edule) or chinchayote starches. A single-screw extruder was utilized to assess the effects of temperature on the degree of substitution (DS) and the functional and structural properties of the modified starches. Glycyltrimethylammonium chloride (GTAC) was used as a cationizing agent in both methods, REX and conventional cationization (CT), at a concentration of 3%. The results indicate that extrusion can produce starches with a DS equivalent to that obtained by CT in the case of corn starch (0.21-0.23). Rheometry shows a decrease in the viscosity peaks due to the pre-gelatinization process. Calorimetry showed a decrease in enthalpy and an increase in tractability temperature for the REX-modified starches due to the temperature and shear to which they were subjected. The spectroscopic technique showed the incorporation of GTAC into the starch structure. The results of physicochemical characterization show that the REX is identified as a viable alternative to CT, offering a faster, more energy-efficient, and environmentally friendly process. The effectiveness of REX in altering the physicochemical properties of starch suggests its potential for innovative industrial applications, such as water treatment or the production of biodegradable materials.
This study introduces a biomimetic dual-protein semi-interpenetrating polymer network (semi-IPN) platform integrating collagen (C) and the phosphorylated globular protein ovalbumin within a bioactive polyurethane (PU) cross-linked matrix. Unlike previously reported collagen-PU-polysaccharide systems, the structural incorporation of a second protein phase enables cooperative regulation of mineral nucleation, interfacial charge distribution, and biological response. Three polysaccharides-starch (CA-A), carboxymethyl cellulose (CA-CMC), and xanthan gum (CA-GX)-were incorporated to modulate network architecture and functionality. Ovalbumin reduced gelation time (t1/2≈30 min), while polysaccharide chemistry governed crosslink density, swelling behavior, crystallinity, and degradation kinetics. CA-GX achieved the highest crosslinking degree (~63%), whereas CA-A exhibited pronounced swelling (~1125%). CA-CMC developed dendritic fibrillar domains with enhanced semicrystallinity, resembling extracellular matrix organization. The semi-IPN structure, confirmed by urea linkage formation, improved viscoelastic strength (G'≈420 Pa) and thermal stability (Tmax≈380°C). All scaffolds were cytocompatible and supported fibroblast, monocyte, and bone-marrow-derived cell metabolism. The dual-protein architecture contributed to hemocompatibility, regulated platelet adhesion, selective antibacterial activity (stronger against Gram-negative bacteria), modulation of inflammatory markers (reduced TNF-α expression), and significant in vitro mineralization, with CA-CMC promoting nearly a 200% increase in carbonated hydroxyapatite deposition in simulated body fluid. By integrating structural collagen fibrils with a mineralization-active globular protein within a tunable polysaccharide-PU framework, this work establishes a multifunctional hybrid scaffold platform capable of coordinated soft- and hard-tissue regenerative responses beyond single-protein semi-IPN systems.
This review explores the multifaceted role of nanocellulose, comprising cellulose nanofibers (CNFs), cellulose nanocrystals (CNCs), and bacterial nanocellulose (BNC), in the design and fabrication of scaffolds for biomedical applications. Structural insights into nanocellulose reveal its capacity to mimic the extracellular matrix (ECM), providing a conducive environment for cell adhesion, proliferation, and differentiation. Modification strategies such as surface functionalization, grafting with bioactive molecules, and incorporation of therapeutic agents further enhance its biological performance and targeted functionality. Innovative scaffold fabrication techniques, including 3D printing, electrospinning, freeze-drying, gas foaming, and cell sheet engineering, are discussed in the context of tailoring scaffold architecture to meet the mechanical and biological requirements of various tissues. The integration of nanocellulose into composite materials and its synergy with other polymers and biomolecules demonstrate great potential in addressing complex tissue regeneration challenges. Biomedical applications cover a wide range of tissues, including skin, bone, cartilage, muscle, nerve, and vascular systems, highlighting the versatility of nanocellulose-based scaffolds. Nanocellulose stands as a key biomaterial in the development of next-generation, eco-friendly scaffolds that align with the principles of sustainability and regenerative medicine.
