Freeze-Dryable, Stable, and Click-Reactive Nanoparticles Composed of Cello-oligosaccharides for Biomolecular Sensing.

Nanoparticles have been widely used as platforms for biomolecular sensing because of their high specific surface area and attractive properties depending on their constituents and structures. Nevertheless, it remains challenging to develop nanoparticulate sensing platforms that are easily storable without aggregation and conjugatable with various ligands in a simple manner. Herein, we demonstrate that nanoparticulate assemblies of cello-oligosaccharides with terminal azido groups are promising candidates. Azidated cello-oligosaccharides can be readily synthesized via the enzyme-catalyzed oligomerization reaction. This study characterized the assembled structures of azidated cello-oligosaccharides produced during the enzymatic synthesis and revealed that the terminal azidated cello-oligosaccharides formed rectangular nanosheet-shaped lamellar crystals. The azido groups located on the nanosheet surfaces were successfully exploited for antigen conjugation via the click chemistry. The resultant antigen-conjugated nanosheets allowed for the quantitative and specific detection of a corresponding antibody, even in 10% serum, owing to the antifouling properties of cello-oligosaccharide assemblies against proteins. It was found that the functionalized nanosheets were redispersible in water after freeze-drying. This remarkable characteristic is attributed to the well-hydrated saccharide residues on the nanosheet surfaces. Moreover, the antibody detection capability did not decline after the thermal treatment of the functionalized nanosheets in a freeze-dried state. Our findings contribute to developing convenient nanoparticulate biomolecular sensing platforms.

Sequential Approach for Water Purification Using Seashell-Derived Calcium Oxide through Disinfection and Flocculation with Polyphosphate for Chemical Pollutant Removal.

Safe water supply is usually inadequate in areas without water treatment plants and even in a city under emergency conditions due to a disaster, even though safe water is essential for drinking and other various purposes. The purification of surface water from a river, lake, or pond requires disinfection and removal of chemical pollutants. In this study, we report a water purification strategy using seashell-derived calcium oxide (CaO) via disinfection and subsequent flocculation with polyphosphate for chemical pollutant removal. Seashell-derived CaO at a concentration (2 g L-1) higher than its saturation concentration caused the >99.999% inactivation of bacteria, mainly due to the alkalinity of calcium hydroxide (Ca(OH)2) produced by hydration. After the disinfection, the addition of sodium polyphosphate at 2 g L-1 allowed for the flocculation of CaO/Ca(OH)2 particles with adsorbing chemical pollutants, such as Congo red, dichlorodiphenyltrichloroethane, di(2-ethylhexyl)phthalate, and polychlorinated biphenyls, for removing these pollutants; purified water was obtained through filtration. Although this purified water was initially highly alkaline (pH ∼ 12.5), its pH decreased into a weak alkaline region (pH ∼ 9) during exposure to ambient air by absorbing carbon dioxide from the air with the precipitating calcium carbonate. The advantages of this water purification strategy include the fact that the saturation of CaO/Ca(OH)2 potentially serves as a visual indicator of disinfection, that the flocculation by polyphosphate removes excessive CaO/Ca(OH)2 as well as chemical pollutants, and that the high pH and Ca2+ concentrations in the resulting purified water are readily decreased. Our findings suggest the usability of seashell-derived material-polymer assemblies for water purification, especially under emergency conditions due to disasters.

Interfacial jamming of surface-alkylated synthetic nanocelluloses for structuring liquids.

Nanocelluloses derived from natural cellulose sources are promising sustainable nanomaterials. Previous studies have reported that nanocelluloses are strongly adsorbed onto liquid-liquid interfaces with the concurrent use of ligands and allow for the structuring of liquids, that is, the kinetic trapping of nonequilibrium shapes of liquids. However, the structuring of liquids using nanocelluloses alone has yet to be demonstrated, despite its great potential in the development of sustainable liquid-based materials that are biocompatible and environmentally friendly. Herein, we demonstrated the structuring of liquids using rectangular sheet-shaped synthetic nanocelluloses with surface alkyl groups. Synthetic nanocelluloses with ethyl, butyl, and hexyl groups on their surfaces were readily prepared following our previous reports via the self-assembly of enzymatically synthesized cello-oligosaccharides having the corresponding alkyl groups. Among the alkylated synthetic nanocelluloses, the hexylated nanocellulose was adsorbed and jammed at water-n-undecane interfaces to form interfacial assemblies, which acted substantially as an integrated film for structuring liquids. These phenomena were attributed to the unique structural characteristics of the surface-hexylated synthetic nanocelluloses; their sheet shape offered a large area for adsorption onto interfaces, and their controlled surface hydrophilicity/hydrophobicity enhanced the affinity for both liquid phases. Our findings promote the development of all-liquid devices using nanocelluloses.

Surface-mediated self-assembly of click-reactive cello-oligosaccharides for fabricating functional nonwoven fabrics.

Polymer fabrics are versatile materials used in various fields. Surface modification methods for hydrophobic polymer fibers have been developed to endow the materials with water wettability and functionality. Nevertheless, it remains a challenge to freely introduce functional groups to polymer fiber surfaces in a simple manner. Herein, we report the decoration of nonwoven fabric surfaces with azidated cello-oligosaccharide assemblies via molecular self-assembly. Cello-oligosaccharides with a terminal azido group were enzymatically synthesized and allowed to self-assemble in polyolefin, polyester, and vinylon nonwoven fabrics. It was found that the functional oligosaccharides formed bark-like assemblies on the nonwoven fiber surfaces, probably through heterogeneous nucleation. The hydrophilic oligosaccharide assemblies made the hydrophobic nonwoven surfaces water-wettable. Moreover, the azido group at oligosaccharide terminal was available for the post-functionalization of the modified nonwovens. In fact, an antigen was successfully conjugated to the modified nonwovens via the click chemistry. The antigen-conjugated nonwovens were useful for the specific and quantitative detection of a corresponding antibody. Our findings demonstrate the great potential of cello-oligosaccharide assembly for the functionalization of fabrics and other polymeric materials.

This study developed a novel and simple method for modifying surfaces of polymer nonwoven fabrics based on the self-assembly of azidated cello-oligosaccharides to fabricate water-wettable and click-reactive functional materials.

Distinguishing anti-PEG antibodies by specificity for the PEG terminus using nanoarchitectonics-based antibiofouling cello-oligosaccharide platforms.

The conjugation of poly(ethylene glycol) (PEG) to therapeutic proteins or nanoparticles is a widely used pharmaceutical strategy to improve their therapeutic efficacy. However, conjugation can make PEG immunogenic and induce the production of anti-PEG antibodies, which decreases both the therapeutic efficacy after repeated dosing and clinical safety. To address these concerns, it is essential to analyze the binding characteristics of anti-PEG antibodies to PEG. However, distinguishing anti-PEG antibodies is still a difficult task. Herein, we demonstrate the use of antibiofouling cello-oligosaccharide assemblies tethering one-terminal methoxy oligo(ethylene glycol) (OEG) ligands for distinguishing anti-PEG antibodies in a simple manner. The OEG ligand-tethering two-dimensional crystalline cello-oligosaccharide assemblies were stably dispersed in a buffer solution and had antibiofouling properties against nonspecific protein adsorption. These characteristics allowed enzyme-linked immunosorbent assays (ELISAs) to be simply performed by cycles of centrifugation/redispersion of aqueous dispersions of the assemblies. The simple assays revealed that the specific OEG ligand-tethering assemblies could distinguish anti-PEG antibodies to detect a specific antibody that preferentially binds to the methoxy terminus of the PEG chain with 3 repeating ethylene glycol units. Furthermore, quantitative detection of the antibodies was successfully performed with high sensitivity even in the presence of serum. The detectable and quantifiable range of antibody concentrations covered those required clinically. Our findings open a new avenue for analyzing the binding characteristics of anti-PEG antibodies in biological samples.

Antibacterial Synthetic Nanocelluloses Synergizing with a Metal-Chelating Agent.

Antibacterial materials composed of biodegradable and biocompatible constituents that are produced via eco-friendly synthetic strategies will become an attractive alternative to antibiotics to combat antibiotic-resistant bacteria. In this study, we demonstrated the antibacterial properties of nanosheet-shaped crystalline assemblies of enzymatically synthesized aminated cellulose oligomers (namely, surface-aminated synthetic nanocelluloses) and their synergy with a metal-chelating antibacterial agent, ethylenediaminetetraacetic acid (EDTA). Growth curves and colony counting assays revealed that the surface-aminated cellulose assemblies had an antibacterial effect against Gram-negative Escherichia coli (E. coli). The cationic assemblies appeared to destabilize the cell wall of E. coli through electrostatic interactions with anionic lipopolysaccharide (LPS) molecules on the outer membrane. The antibacterial properties were significantly enhanced by the concurrent use of EDTA, which potentially removed metal ions from LPS molecules, resulting in synergistic bactericidal effects. No antibacterial activity of the surface-aminated cellulose assemblies was observed against Gram-positive Staphylococcus aureus even in the presence of EDTA, further supporting the contribution of electrostatic interactions between the cationic assemblies and anionic LPS to the activity against Gram-negative bacteria. Analysis using quartz crystal microbalance with dissipation monitoring revealed the attractive interaction of the surface-aminated cellulose assembly with LPS Ra monolayers artificially produced on the device substrate.

Nanospiked paper: Microfibrous cellulose materials nanostructured via partial hydrolysis and self-assembly.

Nanocelluloses, such as cellulose nanofibers and nanocrystals, are sustainable nanomaterials that are generally extracted from natural raw materials in a top-down manner. These nanomaterials and their assemblies are facilitating new applications of biopolymers. However, creating nanostructures from conventional cellulosic materials including paper and cloth remains challenging. Herein, we report an approach for bottom-up nanostructuring of conventional microfibrous cellulose materials via a molecular self-assembly strategy. As a precursor cellulose material, paper was allowed to swell with aqueous phosphoric acid for the partial dissolution and hydrolysis of cellulose while maintaining its microfibrous structure. The generated cello-oligosaccharides in a dissolved state started to self-assemble upon adding water as a coagulant, resulting in nanospike-like assemblies on the microfiber surfaces. The resultant nanospiked papers were found to serve as a precursor for synthesizing silver nanoparticle-cellulose composites with bactericidal activities. Our findings promote the development of cellulose-based functional materials with nanostructures designed via molecular self-assembly.

Biofilm Degradation by Seashell-Derived Calcium Hydroxide and Hydrogen Peroxide.

Microbial cells and self-produced extracellular polymeric substances assembled to form biofilms that are difficult to remove from surfaces, causing problems in various fields. Seashell-derived calcium hydroxide, a sustainable inorganic material, has shown high bactericidal activity even for biofilms due to its alkalinity. However, its biofilm removal efficacy is relatively low. Herein, we report a biofilm degradation strategy that includes two environmentally friendly reagents: seashell-derived calcium hydroxide and hydrogen peroxide. A biofilm model of Escherichia coli was prepared in vitro, treated with calcium hydroxide-hydrogen peroxide solutions, and semi-quantified by the crystal violet stain method. The treatment significantly improved biofilm removal efficacy compared with treatments by calcium hydroxide alone and hydrogen peroxide alone. The mechanism was elucidated from calcium hydroxide-hydrogen peroxide solutions, which suggested that perhydroxyl anion and hydroxyl radical generated from hydrogen peroxide, as well as the alkalinity of calcium hydroxide, enhanced biofilm degradation. This study showed that concurrent use of other reagents, such as hydrogen peroxide, is a promising strategy for improving the biofilm degradation activity of seashell-derived calcium hydroxide and will contribute to developing efficient biofilm removal methods.

Recent Progress on Heparin-Protamine Particles for Biomedical Application.

Biomolecules are attractive building blocks with self-assembly ability, structural diversity, and excellent functionality for creating artificial materials. Heparin and protamine, a clinically relevant pair of biomolecules used in cardiac and vascular surgery, have been shown to coassemble into particulate polyelectrolyte complexes in vitro. The resulting heparin-protamine particles exhibit adhesive properties that enable advantageous interactions with proteins, cells, and various other substances and have been employed as functional materials for biomedical applications. In this review article, we summarize recent progress in research on the use of heparin-protamine particles as drug carriers, cell adhesives, and cell labels. Studies have demonstrated that heparin-protamine particles are potentially versatile in biomedical fields from drug delivery and regenerative medicine to plastic surgery.

Recent Progress in the Development of Disinfectants from Scallop Shell-Derived Calcium Oxide for Clinical and Daily Use.

The current pandemic of novel coronavirus disease (COVID-19) has highlighted the importance of disinfectants. As a raw material for next-generation disinfectants, scallop shell-derived calcium oxide (CaO) has been revealed to exhibit significant virucidal and microbicidal activities and is compatible with living tissues and the environment. This minireview summarizes recent progress in the development of disinfectants from scallop shell-CaO, focusing especially on studies of clinical and daily use applications. We describe the preparation, basic characteristics, and virucidal and microbicidal activities of scallop shell-CaO disinfectants. Furthermore, their applications in the disinfection of contaminated masks and the treatment of infected wounds are briefly introduced.

Self-assembly of cellulose for creating green materials with tailor-made nanostructures.

Inspired by living systems, biomolecules have been employed in vitro as building blocks for creating advanced nanostructured materials. In regard to nucleic acids, peptides, and lipids, their self-assembly pathways and resulting assembled structures are mostly encoded in their molecular structures. On the other hand, outside of its chain length, cellulose, a polysaccharide, lacks structural diversity; therefore, it is challenging to direct this homopolymer to controllably assemble into ordered nanostructures. Nevertheless, the properties of cellulose assemblies are outstanding in terms of their robustness and inertness, and these assemblies are attractive for constructing versatile materials. In this review article, we summarize recent research progress on the self-assembly of cellulose and the applications of assembled cellulose materials, especially for biomedical use. Given that cellulose is the most abundant biopolymer on Earth, gaining control over cellulose assembly represents a promising route for producing green materials with tailor-made nanostructures.

Efficacy of Bioshell Calcium Oxide Water as Disinfectants to Enable Face Mask Reuse.

Bioshell calcium oxide (BiSCaO) is derived from scallop shells and after heat treatment exhibits broad microbicidal activity. BiSCaO Water is a disinfectant prepared by collecting the aqueous layer after adding BiSCaO powder to water, is colorless and transparent, and has a pH of 12.8. We compared the utility of commercially available BiSCaO Water, ethanol, sodium hypochlorite, hypochlorous acid and hydrogen peroxide solutions as sterilization agents to enable the reuse of surgical and N95 face masks. The microbicidal efficacy of each disinfectant was evaluated using pieces of surgical and N95 face masks contaminated with normal bacterial flora. The results suggest that BiSCaO Water has excellent disinfection activity toward contaminated polypropylene masks and has minimal adverse effect on the structure of non-woven masks.

Structured liquids with interfacial robust assemblies of a nonionic crystalline surfactant.

HYPOTHESIS: The structuring of liquids, that is, the kinetic trapping of nonequilibrium shapes of liquid-liquid interfaces, shows great promise for various applications, especially all-liquid devices. The strategies reported thus far to stabilize such unstable states include interfacial jamming of large colloidal particles and interfacial coassembly of elaborate molecules and/or nanoparticles. However, the structuring of liquids using a simple molecular surfactant has not been sufficiently demonstrated. We hypothesized that a surfactant with strong intermolecular interactions would form interfacial assemblies that behave substantially as solid particles for the structuring of liquids. EXPERIMENTS: n-Octyl cello-oligosaccharide, a novel nonionic surfactant developed recently was explored as a candidate because of the ability of cello-oligosaccharides to form robust crystalline assemblies. Interfacial assembly of the nonionic crystalline surfactant was investigated for various water-organic solvent interfaces via pendant drop tensiometry and emulsification. FINDINGS: The crystalline surfactant was found to self-assemble and form a crystalline monolayer at water-organic solvent interfaces, allowing arrested shape changes of the liquid-liquid interfaces. Irregular-shaped liquid droplets were successfully created under various solution conditions, such as various organic solvents for the oil phase and the water phase with high ionic strengths and harsh pH values.

Safety of Concentrated Bioshell Calcium Oxide Water Application for Surface and Skin Disinfections against Pathogenic Microbes.

Immediately post-production, commercially available bioshell calcium oxide (BiSCaO) water is colorless, transparent, and strongly alkaline (pH 12.8), and is known to possess deodorizing properties and broad microbicidal activity. However, BiSCaO Water may represent a serious safety risk to the living body, given the strong alkalinity. This study aimed to investigate the safety of BiSCaO Water for use as an antiseptic/disinfectant despite concerns regarding its high alkalinity. The change over time in pH of BiSCaO Water was measured during air contact (stirring BiSCaO Water in ambient air). When sprayed on metal, plastic, wood piece, paper, and skin surfaces, the pH of BiSCaO Water decreased rapidly, providing a white powder coating upon drying. Scanning electron microscopy images, energy dispersive X-ray elemental mapping, and X-ray diffractograms showed that the dried powder residues of BiSCaO Water were composed primarily of calcium carbonate. These results suggested that BiSCaO Water is a potent reagent that may overcome the obstacles of being strongly alkaline, making this material appropriate for use in disinfection against pathogenic microbes.

pH-Triggered Self-Assembly of Cellulose Oligomers with Gelatin into a Double-Network Hydrogel.

Multicomponent systems for self-assembled molecular gels provide huge opportunities to generate collective or new functions that are not inherent in individual single-component gels. However, gelation tends to require careful and complicated procedures, because, among a myriad of kinetically trapped structures related to the degree of mixing of multiple components over a wide range of scales, from molecular level to macroscopic scale, a limited number of structures that exhibit the desired function need to be constructed. This study presents a simple method for the construction of double-network (DN) hydrogels with improved stiffness composed of crystalline cellulose oligomers and gelatin. The pH-triggered self-assembly of cellulose oligomers leads to the formation of robust networks composed of crystalline nanofibers in the presence of dissolved gelatin, followed by cooling to allow for the formation of soft gelatin networks. The resultant DN hydrogels exhibit improved stiffness; the improvement in gel stiffness with double networking is comparable to that of previously reported DN hydrogels produced via a time-consuming enzymatic reaction.

Correction: Biocatalytic oligomerization-induced self-assembly of crystalline cellulose oligomers into nanoribbon networks assisted by organic solvents.

[This corrects the article DOI: 10.3762/bjnano.10.173.].

Confined Reduced Graphene Oxides as a Platform for DNA Sensing in Solutions Crowded with Biomolecules.

Promising systems for simple biomolecular sensing rely on the adsorption of probe DNAs onto graphene oxides (GOs). However, in biological samples crowded with nontarget biomolecules, the systems suffer from nonspecific displacement of the adsorbed probe DNAs by those crowding biomolecules, which is a major obstacle for practical application. Herein, we demonstrate DNA sensing in solutions crowded with biomolecules, such as concentrated protein solutions and even serum, using reduced GOs (rGOs) confined within robust networks of crystalline cellulose oligomers. The confinement of GOs was conducted with concomitant reduction in one step via the enzymatic cellulose oligomer synthesis. The confined rGOs could adsorb probe DNAs more tightly than could GOs due to higher hydrophobicity while being prevented from aggregation even in water by the confinement effect, allowing for sensing in solutions crowded with biomolecules. The sensitivity was comparable to that of the original GO-based sensing in clean buffer solutions. This study paves the way for simple biomolecular sensing in biological samples.

Biocatalytic oligomerization-induced self-assembly of crystalline cellulose oligomers into nanoribbon networks assisted by organic solvents.

Crystalline poly- and oligosaccharides such as cellulose can form extremely robust assemblies, whereas the construction of self-assembled materials from such molecules is generally difficult due to their complicated chemical synthesis and low solubility in solvents. Enzyme-catalyzed oligomerization-induced self-assembly has been shown to be promising for creating nanoarchitectured crystalline oligosaccharide materials. However, the controlled self-assembly into organized hierarchical structures based on a simple method is still challenging. Herein, we demonstrate that the use of organic solvents as small-molecule additives allows for control of the oligomerization-induced self-assembly of cellulose oligomers into hierarchical nanoribbon network structures. In this study, we dealt with the cellodextrin phosphorylase-catalyzed oligomerization of phosphorylated glucose monomers from ᴅ-glucose primers, which produce precipitates of nanosheet-shaped crystals in aqueous solution. The addition of appropriate organic solvents to the oligomerization system was found to result in well-grown nanoribbon networks. The organic solvents appeared to prevent irregular aggregation and subsequent precipitation of the nanosheets via solvation for further growth into the well-grown higher-order structures. This finding indicates that small-molecule additives provide control over the self-assembly of crystalline oligosaccharides for the creation of hierarchically structured materials with high robustness in a simple manner.

Temperature-Directed Assembly of Crystalline Cellulose Oligomers into Kinetically Trapped Structures during Biocatalytic Synthesis.

Crystalline polysaccharides, such as cellulose and chitin, can form superior assemblies in terms of physicochemical stability and mechanical properties. However, their use as molecular building blocks for self-assembled materials is rare, possibly because each crystalline polysaccharide has its own unique monomer unit, preventing molecular design for controlling the self-assembly. Herein, we demonstrate the temperature-directed assembly of crystalline cellulose oligomers into kinetically trapped structures, namely, precipitated nanosheets, nanoribbon network hydrogels, and dispersed nanosheets (in descending order of temperature). It was found that enzymatically synthesized cellulose oligomers self-assembled in situ into those structures depending on the synthetic temperatures. Mechanistic studies suggested that the formation of the nanoribbon networks and the dispersed nanosheets at lower temperatures were driven by synergy between the decreased hydrophobic effect and the simultaneously induced self-crowding effect. Furthermore, nanoribbon network formation was exploited for the construction of cellulose oligomer-based hybrid gels with colloidal particles. Our findings promote the development of robust self-assembled materials composed of crystalline polysaccharides with highly ordered nano-to-macroscale structures.

Assembly of reduced graphene oxides into a three-dimensional porous structure via confinement within robust cellulose oligomer networks.

The assembly of nanomaterials into a networked superstructure is a strategy used to construct macroscopic porous materials having the functional properties of nanomaterials. However, because nanomaterials generally prefer densely packed assembled states owing to their high surface energies, the construction of a fine porous structure is still a challenge. In this study, we demonstrate the assembly of reduced graphene oxides (rGOs) into a fine porous structure via confinement within robust cellulose oligomer networks. The confinement of rGOs within cellulose oligomer networks was achieved in one step via the enzymatic synthesis of cellulose oligomers. When the resultant cellulose oligomer gels confining rGOs were reduced by hydrogen iodide, the robust cellulose oligomer networks served as a confinement space for rGOs, preventing excessive aggregation of the rGOs and thus encouraging their assembly into a fine porous structure. Electrochemical measurements revealed that the porous rGO materials could act as electrode materials for supercapacitors. Our strategy based on simple physical confinement will allow for the creation of functional porous materials with excellent nanomorphologies from various nanomaterials.