Implantable Patch of Oxidized Nanofibrillated Cellulose and Lysozyme Amyloid Nanofibrils for the Regeneration of Infarcted Myocardium Tissue and Local Delivery of RNA-Loaded Nanoparticles.

Biopolymeric implantable patches are popular scaffolds for myocardial regeneration applications. Besides being biocompatible, they can be tailored to have required properties and functionalities for this application. Recently, fibrillar biobased nanostructures prove to be valuable in the development of functional biomaterials for tissue regeneration applications. Here, periodate-oxidized nanofibrillated cellulose (OxNFC) is blended with lysozyme amyloid nanofibrils (LNFs) to prepare a self-crosslinkable patch for myocardial implantation. The OxNFC:LNFs patch shows superior wet mechanical properties (60 MPa for Young's modulus and 1.5 MPa for tensile stress at tensile strength), antioxidant activity (70% scavenging activity under 24 h), and bioresorbability ratio (80% under 91 days), when compared to the patches composed solely of NFC or OxNFC. These improvements are achieved while preserving the morphology, required thermal stability for sterilization, and biocompatibility toward rat cardiomyoblast cells. Additionally, both OxNFC and OxNFC:LNFs patches reveal the ability to act as efficient vehicles to deliver spermine modified acetalated dextran nanoparticles, loaded with small interfering RNA, with 80% of delivery after 5 days. This study highlights the value of simply blending OxNFC and LNFs, synergistically combining their key properties and functionalities, resulting in a biopolymeric patch that comprises valuable characteristics for myocardial regeneration applications.

A Guide to Polysaccharide-Based Hydrogel Bioinks for 3D Bioprinting Applications.

Three-dimensional (3D) bioprinting is an innovative technology in the biomedical field, allowing the fabrication of living constructs through an approach of layer-by-layer deposition of cell-laden inks, the so-called bioinks. An ideal bioink should possess proper mechanical, rheological, chemical, and biological characteristics to ensure high cell viability and the production of tissue constructs with dimensional stability and shape fidelity. Among the several types of bioinks, hydrogels are extremely appealing as they have many similarities with the extracellular matrix, providing a highly hydrated environment for cell proliferation and tunability in terms of mechanical and rheological properties. Hydrogels derived from natural polymers, and polysaccharides, in particular, are an excellent platform to mimic the extracellular matrix, given their low cytotoxicity, high hydrophilicity, and diversity of structures. In fact, polysaccharide-based hydrogels are trendy materials for 3D bioprinting since they are abundant and combine adequate physicochemical and biomimetic features for the development of novel bioinks. Thus, this review portrays the most relevant advances in polysaccharide-based hydrogel bioinks for 3D bioprinting, focusing on the last five years, with emphasis on their properties, advantages, and limitations, considering polysaccharide families classified according to their source, namely from seaweed, higher plants, microbial, and animal (particularly crustaceans) origin.

Bacterial Nanocellulose toward Green Cosmetics: Recent Progresses and Challenges.

In the skin care field, bacterial nanocellulose (BNC), a versatile polysaccharide produced by non-pathogenic acetic acid bacteria, has received increased attention as a promising candidate to replace synthetic polymers (e.g., nylon, polyethylene, polyacrylamides) commonly used in cosmetics. The applicability of BNC in cosmetics has been mainly investigated as a carrier of active ingredients or as a structuring agent of cosmetic formulations. However, with the sustainability issues that are underway in the highly innovative cosmetic industry and with the growth prospects for the market of bio-based products, a much more prominent role is envisioned for BNC in this field. Thus, this review provides a comprehensive overview of the most recent (last 5 years) and relevant developments and challenges in the research of BNC applied to cosmetic, aiming at inspiring future research to go beyond in the applicability of this exceptional biotechnological material in such a promising area.

Natural Polymers-Based Materials: A Contribution to a Greener Future.

Natural polymers have emerged as promising candidates for the sustainable development of materials in areas ranging from food packaging and biomedicine to energy storage and electronics. In tandem, there is a growing interest in the design of advanced materials devised from naturally abundant and renewable feedstocks, in alignment with the principles of Green Chemistry and the 2030 Agenda for Sustainable Development. This review aims to highlight some examples of the research efforts conducted at the Research Team BioPol4fun, Innovation in BioPolymer-based Functional Materials and Bioactive Compounds, from the Portuguese Associate Laboratory CICECO-Aveiro Institute of Materials at the University of Aveiro, regarding the exploitation of natural polymers (and derivatives thereof) for the development of distinct sustainable biobased materials. In particular, focus will be given to the use of polysaccharides (cellulose, chitosan, pullulan, hyaluronic acid, fucoidan, alginate, and agar) and proteins (lysozyme and gelatin) for the assembly of composites, coatings, films, membranes, patches, nanosystems, and microneedles using environmentally friendly strategies, and to address their main domains of application.

Effect of the Micronization of Pulp Fibers on the Properties of Green Composites.

Green composites, composed of bio-based matrices and natural fibers, are a sustainable alternative for composites based on conventional thermoplastics and glass fibers. In this work, micronized bleached Eucalyptus kraft pulp (BEKP) fibers were used as reinforcement in biopolymeric matrices, namely poly(lactic acid) (PLA) and poly(hydroxybutyrate) (PHB). The influence of the load and aspect ratio of the mechanically treated microfibers on the morphology, water uptake, melt flowability, and mechanical and thermal properties of the green composites were investigated. Increasing fiber loads raised the tensile and flexural moduli as well as the tensile strength of the composites, while decreasing their elongation at the break and melt flow rate. The reduced aspect ratio of the micronized fibers (in the range from 11.0 to 28.9) improved their embedment in the matrices, particularly for PHB, leading to superior mechanical performance and lower water uptake when compared with the composites with non-micronized pulp fibers. The overall results show that micronization is a simple and sustainable alternative for conventional chemical treatments in the manufacturing of entirely bio-based composites.

Topical Drug Delivery Systems Based on Bacterial Nanocellulose: Accelerated Stability Testing.

Bacterial nanocellulose (BNC) membranes have enormous potential as systems for topical drug delivery due to their intrinsic biocompatibility and three-dimensional nanoporous structure, which can house all kinds of active pharmaceutical ingredients (APIs). Thus, the present study investigated the long-term storage stability of BNC membranes loaded with both hydrophilic and lipophilic APIs, namely, caffeine, lidocaine, ibuprofen and diclofenac. The storage stability was evaluated under accelerated testing conditions at different temperatures and relative humidity (RH), i.e., 75% RH/40 °C, 60% RH/25 °C and 0% RH/40 °C. All systems were quite stable under these storage conditions with no significant structural and morphological changes or variations in the drug release profile. The only difference observed was in the moisture-uptake, which increased with RH due to the hydrophilic nature of BNC. Furthermore, the caffeine-loaded BNC membrane was selected for in vivo cutaneous compatibility studies, where patches were applied in the volar forearm of twenty volunteers for 24 h. The cutaneous responses were assessed by non-invasive measurements and the tests revealed good compatibility for caffeine-loaded BNC membranes. These results highlight the good storage stability of the API-loaded BNC membranes and their cutaneous compatibility, which confirms the real potential of these dermal delivery systems.

Understanding the Structure and Dynamics of Nanocellulose-Based Composites with Neutral and ionic Poly(methacrylate) Derivatives using Inelastic Neutron Scattering and DFT Calculations.

Bacterial nanocellulose (BC)-based composites containing poly(2-hydroxyethyl methacrylate) (PHEMA), poly(methacroylcholine chloride) (PMACC) or poly(methacroylcholine hydroxide) (PMACH) were characterized by inelastic neutron scattering (INS) spectroscopy, combined with DFT (density functional theory) calculations of model systems. A reasonable match between calculated and experimental spectral lines and their intensities was used to support the vibrational assignment of the observed bands and to validate the possible structures. The differences between the spectra of the nanocomposites and the pure precursors indicate that interactions between the components are stronger for the ionic poly(methacrylate) derivatives than for the neutral counterpart. Displaced anions interact differently with cellulose chains, due to the different ability to compete with the O-H···O hydrogen bonds in cellulose. Hence, the INS is an adequate technique to delve deeper into the structure and dynamics of nanocellulose-based composites, confirming that they are true nanocomposite materials instead of simple mixtures of totally independent domains.

Bioactive Phytochemicals from Wild Arbutus unedo L. Berries from Different Locations in Portugal: Quantification of Lipophilic Components.

The lipophilic composition of wild Arbutus unedo L. berries, collected from six locations in Penacova (center of Portugal), as well as some general chemical parameters, namely total soluble solids, pH, titratable acidity, total phenolic content and antioxidant activity was studied in detail to better understand its potential as a source of bioactive compounds. The chemical composition of the lipophilic extracts, focused on the fatty acids, triterpenoids, sterols, long chain aliphatic alcohols and tocopherols, was investigated by gas chromatography-mass spectrometry (GC-MS) analysis of the dichloromethane extracts. The lipophilic extractives of the ripe A. unedo berries ranged from 0.72% to 1.66% (w/w of dry weight), and consisted mainly of triterpenoids, fatty acids and sterols. Minor amounts of long chain aliphatic alcohols and tocopherols were also identified. Forty-one compounds were identified and among these, ursolic acid, lupeol, α-amyrin, linoleic and α-linolenic acids, and ß-sitosterol were highlighted as the major components. To the best of our knowledge the current research study provides the most detailed phytochemical repository for the lipophilic composition of A. unedo, and offers valuable information for future valuation and exploitation of these berries.

Wood inspired biobased nanocomposite films composed of xylans, lignosulfonates and cellulose nanofibers for active food packaging.

The growing concerns on environmental pollution and sustainability have raised the interest on the development of functional biobased materials for different applications, including food packaging, as an alternative to the fossil resources-based counterparts, currently available in the market. In this work, functional wood inspired biopolymeric nanocomposite films were prepared by solvent casting of suspensions containing commercial beechwood xylans, cellulose nanofibers (CNF) and lignosulfonates (magnesium or sodium), in a proportion of 2:5:3 wt%, respectively. All films presented good homogeneity, translucency, and thermal stability up to 153 °C. The incorporation of CNF into the xylan/lignosulfonates matrix provided good mechanical properties to the films (Young's modulus between 1.08 and 3.79 GPa and tensile strength between 12.75 and 14.02 MPa). The presence of lignosulfonates imparted the films with antioxidant capacity (DPPH radical scavenging activity from 71.6 to 82.4 %) and UV barrier properties (transmittance ≤19.1 % (200-400 nm)). Moreover, the films obtained are able to successfully delay the browning of packaged fruit stored over 7 days at 4 °C. Overall, the obtained results show the potential of using low-cost and eco-friendly resources for the development of sustainable active food packaging materials.

Switchable adhesive films of pullulan loaded with a deep eutectic solvent-curcumin formulation for the photodynamic treatment of drug-resistant skin infections.

Antimicrobial photodynamic therapy (aPDT) is a potent tool to surpass the global rise of antimicrobial resistance; still, the effective topical administration of photosensitizers remains a challenge. Biopolymer-based adhesive films can safely extend the residence time of photosensitizers. However, their wide application is narrowed by their limited water absorption capacity and gel strength. In this study, pullulan-based films with a switchable character (from a solid film to an adhesive hydrogel) were developed. This was accomplished by the incorporation of a betaine-based deep eutectic solvent (DES) containing curcumin (4.4 µg.cm-2) into the pullulan films, which tuned the films' skin moisture absorption ability, and therefore they switch into an adhesive hydrogel capable of delivering the photosensitizer. The obtained transparent films presented higher extensibility (elongation at break up to 338.2%) than the pullulan counterparts (6.08%), when stored at 54% of relative humidity, and the corresponding hydrogels a 4-fold higher adhesiveness than commercial hydrogels. These non-cytotoxic adhesives allowed the inactivation (∼5 log reduction), down to the detection limit of the method, of multiresistant strains of Staphylococcus aureus in ex vivo skin samples. Overall, these materials are promising for aPDT in the treatment of resistant skin infections, while being easily removed from the skin.

Hydrogel Bioinks of Alginate and Curcumin-Loaded Cellulose Ester-Based Particles for the Biofabrication of Drug-Releasing Living Tissue Analogs.

3D bioprinting is a versatile technique that allows the fabrication of living tissue analogs through the layer-by-layer deposition of cell-laden biomaterials, viz. bioinks. In this work, composite alginate hydrogel-based bioinks reinforced with curcumin-loaded particles of cellulose esters (CEpCUR) and laden with human keratinocytes (HaCaT) are developed. The addition of the CEpCUR particles, with sizes of 740 ± 147 nm, improves the rheological properties of the inks, increasing their shear stress and viscosity, while preserving the recovery rate and the mechanical and viscoelastic properties of the resulting fully cross-linked hydrogels. Moreover, the presence of these particles reduces the degradation rate of the hydrogels from 26.3 ± 0.8% (ALG) to 18.7 ± 1.3% (ALG:CEpCUR_10%) after 3 days in the culture medium. The 3D structures printed with the ALG:CEpCUR inks reveal increased printing definition and the ability to release curcumin (with nearly 70% of cumulative release after 24 h in PBS). After being laden with HaCaT cells (1.2 × 106 cells mL-1), the ALG:CEpCUR bioinks can be successfully 3D bioprinted, and the obtained living constructs show good dimensional stability and high cell viabilities at 7 days post-bioprinting (nearly 90%), confirming their great potential for application in fields like wound healing.

Nanofibrillated cellulose/gellan gum hydrogel-based bioinks for 3D bioprinting of skin cells.

The development of suitable bioinks is an important research topic in the field of three-dimensional (3D) bioprinting. Herein, novel hydrogel-based bioinks composed of nanofibrillated cellulose (NFC) and gellan gum (GG) in different NFC/GG mass proportions (90:10, 80:20, 70:30, and 60:40) were developed and characterized. The increase in the content of GG, as well as its combination with NFC, enhanced their rheological properties, increasing both storage (G') and loss (G") moduli and the G' recovery capacity of the hydrogels (from 70.05 ± 3.06 % (90:10) to 82.63 ± 1.21 % (60:40)), as well as their mechanical properties, increasing the compressive stiffness and stress from 114.02 ± 10.93 Pa (90:10) to 337.16 ± 34.03 Pa (60:40) and from 18.27 ± 1.32 kPa (90:10) to 47.17 ± 3.59 kPa (60:40), respectively. The hydrogels were non-cytotoxic against human keratinocyte cells (HaCaT), with cell viabilities above 70 % for up to 72 h. The hydrogel 60:40 was loaded with HaCaT cells (3 × 106 cells mL-1) and bioprinted. The cell viability was maintained elevated until day 7 (90 ± 3 %) after bioprinting. These results highlight that the combination of these two biopolymers was a good strategy for the development of novel hydrogel-based bioinks for extrusion 3D bioprinting applications.

Dissolvable Carboxymethylcellulose Microneedles for Noninvasive and Rapid Administration of Diclofenac Sodium.

The aim of this study is to prepare dissolvable biopolymeric microneedle (MN) patches composed solely of sodium carboxymethylcellulose (CMC), a water-soluble cellulose derivative with good film-forming ability, by micromolding technology for the transdermal delivery of diclofenac sodium salt (DCF). The MNs with ≈456 µm in height displayed adequate morphology, thermal stability up to 200 °C, and the required mechanical strength for skin insertion (>0.15 N needle-1 ). Experiments in ex vivo abdominal human skin demonstrate the insertion capability of the CMC_DCF MNs up to 401 µm in depth. The dissolution of the patches in saline buffer results in a maximum cumulative release of 98% of diclofenac after 40 min, and insertion in a skin simulant reveals that all MNs completely dissolve within 10 min. Moreover, the MN patches are noncytotoxic toward human keratinocytes. These results suggest that the MN patches produced with CMC are promising biopolymeric systems for the rapid administration of DCF in a minimally invasive manner.

Injectable Nanocomposite Hydrogels of Gelatin-Hyaluronic Acid Reinforced with Hybrid Lysozyme Nanofibrils-Gold Nanoparticles for the Regeneration of Damaged Myocardium.

Biopolymeric injectable hydrogels are promising biomaterials for myocardial regeneration applications. Besides being biocompatible, they adjust themselves, perfectly fitting the surrounding tissue. However, due to their nature, biopolymeric hydrogels usually lack desirable functionalities, such as antioxidant activity and electrical conductivity, and in some cases, mechanical performance. Protein nanofibrils (NFs), such as lysozyme nanofibrils (LNFs), are proteic nanostructures with excellent mechanical performance and antioxidant activity, which can work as nanotemplates to produce metallic nanoparticles. Here, gold nanoparticles (AuNPs) were synthesized in situ in the presence of LNFs, and the obtained hybrid AuNPs@LNFs were incorporated into gelatin-hyaluronic acid (HA) hydrogels for myocardial regeneration applications. The resulting nanocomposite hydrogels showed improved rheological properties, mechanical resilience, antioxidant activity, and electrical conductivity, especially for the hydrogels containing AuNPs@LNFs. The swelling and bioresorbability ratios of these hydrogels are favorably adjusted at lower pH levels, which correspond to the ones in inflamed tissues. These improvements were observed while maintaining important properties, namely, injectability, biocompatibility, and the ability to release a model drug. Additionally, the presence of AuNPs allowed the hydrogels to be monitorable through computer tomography. This work demonstrates that LNFs and AuNPs@LNFs are excellent functional nanostructures to formulate injectable biopolymeric nanocomposite hydrogels for myocardial regeneration applications.

Advanced Nanocellulose-Based Materials: Production, Properties, and Applications.

Natural polymers, such as polysaccharides and proteins, are being extensively utilized as substrates to create advanced materials [...].

Cellulose and protein nanofibrils: Singular biobased nanostructures for the design of sustainable advanced materials.

Polysaccharides and proteins are extensively used for the design of advanced sustainable materials. Owing to the high aspect ratio and specific surface area, ease of modification, high mechanical strength and thermal stability, renewability, and biodegradability, biopolymeric nanofibrils are gaining growing popularity amongst the catalog of nanostructures exploited in a panoply of fields. These include the nanocomposites, paper and packaging, environmental remediation, electronics, energy, and biomedical applications. In this review, recent trends on the use of cellulose and protein nanofibrils as versatile substrates for the design of high-performance nanomaterials are assessed. A concise description of the preparation methodologies and characteristics of cellulosic nanofibrils, namely nanofibrillated cellulose (NFC), bacterial nanocellulose (BNC), and protein nanofibrils is presented. Furthermore, the use of these nanofibrils in the production of sustainable materials, such as membranes, films, and patches, amongst others, as well as their major domains of application, are briefly described, with focus on the works carried out at the BioPol4Fun Research Group (Innovation in BioPolymer based Functional Materials and Bioactive Compounds) from the Portuguese associate laboratory CICECO-Aveiro Institute of Materials (University of Aveiro). The potential for partnership between both types of nanofibrils in advanced material development is also reviewed. Finally, the critical challenges and opportunities for these biobased nanostructures for the development of functional materials are addressed.

Bioactive Bacterial Nanocellulose Membranes Enriched with Eucalyptus globulus Labill. Leaves Aqueous Extract for Anti-Aging Skin Care Applications.

Bacterial nanocellulose (BNC) membranes, with remarkable physical and mechanical properties, emerged as a versatile biopolymeric carrier of bioactive compounds for skin care applications. In this study, BNC membranes were loaded with glycerol (as plasticizer and humectant agent) and different doses (1-3 µg cm-2) of an aqueous extract obtained from the hydro-distillation of Eucalyptus globulus Labill. leaves (HDE), for application as sheet facial masks. All membranes are resistant and highly malleable at dry and wet states, with similar or even better mechanical properties than those of a commercial BNC mask. Moreover, the HDE was found to confer a dose-dependent antioxidant activity to pure BNC. Additionally, upon 3 months of storage at 22-25 °C and 52% relative humidity (RH) or at 40 °C and 75% RH, it was confirmed that the antioxidant activity and the macroscopic aspect of the membrane with 2 µg cm-2 of HDE were maintained. Membranes were also shown to be non-cytotoxic towards HaCaT and NIH/3T3 cells, and the membrane with 2 µg cm-2 of HDE caused a significant reduction in the senescence-associated ß-galactosidase activity in NIH/3T3 cells. These findings suggest the suitability and potential of the obtained membranes as bioactive facial masks for anti-aging applications.

Improving the Processability and Performance of Micronized Fiber-Reinforced Green Composites through the Use of Biobased Additives.

Green composites made of bioplastics reinforced with natural fibers have gained considerable attention over recent years. However, the use of natural fibers in composites usually compromise some key properties, such as the impact strength and the processability of the final materials. In the present study, two distinct additives, namely an epoxidized linseed oil (ELO) and a sugar-based surfactant, viz. GlucoPure® Sense (GPS), were tested in composite formulations of poly(lactic acid) (PLA) or poly(hydroxybutyrate) (PHB) reinforced with micronized pulp fibers. Both additives showed a plasticizing effect, which led to a decrease in the Young's and flexural moduli and strengths. At the same time, the elongation and flexural strain at break were considerably improved on some formulations. The melt flow rate was also remarkably improved with the incorporation of the additives. In the PHB-based composites, an increment of 230% was observed upon incorporation of 7.5 wt.% ELO and, in composites based on PLA, an increase of around 155% was achieved with the introduction of 2.5 wt.% GPS. ELO also increased the impact strength to a maximum of 29 kJ m-2, in formulations with PLA. For most composites, a faster degradation rate was observed on the formulations with the additives, reaching, in the case of PHB composites with GPS, a noteworthy weight loss over 75% under burial testing in compost medium at room temperature.

Deep Eutectic Solvent Formulations and Alginate-Based Hydrogels as a New Partnership for the Transdermal Administration of Anti-Inflammatory Drugs.

The transdermal administration of nonsteroidal anti-inflammatory drugs (NSAIDs) is a valuable and safer alternative to their oral intake. However, most of these drugs display low water solubility, which makes their incorporation into hydrophilic biopolymeric drug-delivery systems difficult. To overcome this drawback, aqueous solutions of bio-based deep eutectic solvents (DES) were investigated to enhance the solubility of ibuprofen, a widely used NSAID, leading to an increase in its solubility of up to 7917-fold when compared to its water solubility. These DES solutions were shown to be non-toxic to macrophages with cell viabilities of 97.4% (at ibuprofen concentrations of 0.25 mM), while preserving the anti-inflammatory action of the drug. Their incorporation into alginate-based hydrogels resulted in materials with a regular structure and higher flexibility. These hydrogels present a sustained release of the drug, which is able, when containing the DES aqueous solution comprising ibuprofen, to deliver 93.5% of the drug after 8 h in PBS. Furthermore, these hydrogels were able to improve the drug permeation across human skin by 8.5-fold in comparison with the hydrogel counterpart containing only ibuprofen. This work highlights the possibility to remarkably improve the transdermal administration of NSAIDs by combining new drug formulations based on DES and biopolymeric drug delivery systems.

Alginate-Lysozyme Nanofibers Hydrogels with Improved Rheological Behavior, Printability and Biological Properties for 3D Bioprinting Applications.

In this study, alginate nanocomposite hydrogel bioinks reinforced with lysozyme nanofibers (LNFs) were developed. Alginate-LNF (A-LNF) suspensions with different LNF contents (1, 5 and 10 wt.%) were prepared and pre-crosslinked with 0.5% (w/v) CaCl2 to formulate A-LNF inks. These inks exhibit proper shear-thinning behavior and good recovery properties (~90%), with the pre-crosslinking step playing a crucial role. A-LNF fully crosslinked hydrogels (with 2% (w/v) CaCl2) that mimic 3D printing scaffolds were prepared, and it was observed that the addition of LNFs improved several properties of the hydrogels, such as the morphology, swelling and degradation profiles, and mechanical properties. All formulations are also noncytotoxic towards HaCaT cells. The printing parameters and 3D scaffold model were then optimized, with A-LNF inks showing improved printability. Selected A-LNF inks (A-LNF0 and A-LNF5) were loaded with HaCaT cells (cell density 2 × 106 cells mL-1), and the cell viability within the bioprinted scaffolds was evaluated for 1, 3 and 7 days, with scaffolds printed with the A-LNF5 bioink showing the highest values for 7 days (87.99 ± 1.28%). Hence, A-LNF bioinks exhibited improved rheological performance, printability and biological properties representing a good strategy to overcome the main limitations of alginate-based bioinks.