Biohybrid Materials Research Articles

Development of Chitosan-Coated Electrospun Poly(3-hydroxybutyrate) Biohybrid Materials for Growth and Long-Term Storage of Bacillus subtilis

Numerous bacterial species can both suppress plant pathogens and promote plant growth. By combining these bacteria with stabilizing substances, we can develop biological products with an extended shelf life, contributing to sustainable agriculture. Bacillus subtilis is one such bacterial species, possessing traits that enhance plant growth and offer effective protection, making it suitable for various applications. In this study, we successfully incorporated B. subtilis into hybrid materials composed of poly(3-hydroxybutyrate) (PHB) fibers coated with chitosan film. The polymer carrier not only supports the normal growth of the bioagent but also preserves its viability during long-term storage. For that reason, the impact of chitosan molecular weight on the dynamic viscosity of the solutions used for film formation, as well as the resulting film’s morphology, mechanical properties, and quantity of incorporated B. subtilis, along with their growth dynamics was investigated. SEM was used to examine the morphology of B. subtilis, electrospun PHB, and PHB mats coated with chitosan/B. subtilis. The results from mechanical tests demonstrate that chitosan film formation enhanced the tensile strength of the tested materials. Microbiological tests confirmed that the bacteria incorporated into the hybrid materials grow normally. The conducted viability tests demonstrate that the bacteria incorporated within the electrospun materials remained viable both after incorporation and following 90 days of storage. Moreover, the prepared biohybrid materials effectively inhibited the growth of the plant pathogenic strain Alternaria. Thus, the study provides more efficient and sustainable agricultural solutions by reducing reliance on synthetic materials and enhancing environmental compatibility through the development of advanced biomaterials capable of delivering active biocontrol agents.

Selectivity of a Bioreceptor Element of the Sensor Based on an Organosilicon Material and Microorganism Cells

The yeast cells from Ogataea polymorpha, Blastobotrys adeninivorans, and Debaryomyces hansenii were immobilized on an organosilicon material by the sol-gel method, resulting in a hybrid biocatalyst. The catalytic activity of the immobilized mixture of microorganisms was evaluated using it as a bioreceptor element in a biosensor. The proposed biohybrid material, consisting of the yeast association immobilized on the organosilicate sol-gel matrix, exhibited broad substrate specificity. Thus, the immobilization of multiple yeast strains provides a heterogeneous catalyst able to biologically oxidize a wide range of organic compounds commonly found in wastewater and surface water.

P078 Structured biohybrid materials for the regeneration of the urogenital plexus, based on selective biofunctionalization of D3S-engineered silk fibers

P078 Structured biohybrid materials for the regeneration of the urogenital plexus, based on selective biofunctionalization of D3S-engineered silk fibers

Encapsulation of Bacillus subtilis in Electrospun Poly(3-hydroxybutyrate) Fibers Coated with Cellulose Derivatives for Sustainable Agricultural Applications.

One of the latest trends in sustainable agriculture is the use of beneficial microorganisms to stimulate plant growth and biologically control phytopathogens. Bacillus subtilis, a Gram-positive soil bacterium, is recognized for its valuable properties in various biotechnological and agricultural applications. This study presents, for the first time, the successful encapsulation of B. subtilis within electrospun poly(3-hydroxybutyrate) (PHB) fibers, which are dip-coated with cellulose derivatives. In that way, the obtained fibrous biohybrid materials actively ensure the viability of the encapsulated biocontrol agent during storage and promote its normal growth when exposed to moisture. Aqueous solutions of the cellulose derivatives-sodium carboxymethyl cellulose and 2-hydroxyethyl cellulose, were used to dip-coat the electrospun PHB fibers. The study examined the effects of the type and molecular weight of these cellulose derivatives on film formation, mechanical properties, bacterial encapsulation, and growth. Scanning electron microscopy (SEM) was utilized to observe the morphology of the biohybrid materials and the encapsulated B. subtilis. Additionally, ATR-FTIR spectroscopy confirmed the surface chemical composition of the biohybrid materials and verified the successful coating of PHB fibers. Mechanical testing revealed that the coating enhanced the mechanical properties of the fibrous materials and depends on the molecular weight of the used cellulose derivatives. Viability tests demonstrated that the encapsulated B. subtilis exhibited normal growth from the prepared materials. These findings suggest that the developed fibrous biohybrid materials hold significant promise as biocontrol formulations for plant protection and growth promotion in sustainable agriculture.

Monomer Composition as a Mechanism to Control the Self-Assembly of Diblock Oligomeric Peptide-Polymer Amphiphiles.

Diblock oligomeric peptide-polymer amphiphiles (PPAs) are biohybrid materials that offer versatile functionality by integrating the sequence-dependent properties of peptides with the synthetic versatility of polymers. Despite their potential as biocompatible materials, the rational design of PPAs for assembly into multichain nanoparticles remains challenging due to the complex intra- and intermolecular interactions emanating from the polymer and peptide segments. To systematically explore the impact of monomer composition on nanoparticle assembly, PPAs were synthesized with a random coil peptide (XTEN2) and oligomeric alkyl acrylates with different side chains: ethyl, tert-butyl, n-butyl, and cyclohexyl. Experimental characterization using electron and atomic force microscopies demonstrated that the tail hydrophobicity impacted accessible morphologies. Moreover, the characterization of different assembly protocols (i.e., bath sonication and thermal annealing) revealed that certain tail compositions provide access to kinetically trapped assemblies. All-atom molecular dynamics simulations of micelle formation unveiled key interactions and differences in core hydration, dictating the PPA assembly behavior. These findings highlight the complexity of PPA assembly dynamics and serve as valuable benchmarks to guide the design of PPAs for a variety of applications, including catalysis, mineralization, targeted sequestration, antimicrobial activity, and cargo transportation.

Sources and Extraction of Biopolymers and Manufacturing of Bio-Based Nanocomposites for Different Applications.

In the recent era, bio-nanocomposites represent an emerging group of nanostructured hybrid materials and have been included in a new field at the frontier of materials science, life sciences, and nanotechnology. These biohybrid materials reveal developed structural and functional features of great attention for diverse uses. These materials take advantage of the synergistic assembling of biopolymers with nanometer-sized reinforcements. Conversely, polysaccharides have received great attention due to their several biological properties like antimicrobial and antioxidant performance. They mainly originated in different parts of plants, animals, seaweed, and microorganisms (bacteria, fungi, and yeasts). Polysaccharide-based nanocomposites have great features, like developed physical, structural, and functional features; affordability; biodegradability; and biocompatibility. These bio-based nanocomposites have been applied in biomedical, water treatment, food industries, etc. This paper will focus on the very recent trends in bio-nanocomposite based on polysaccharides for diverse applications. Sources and extraction methods of polysaccharides and preparation methods of their nanocomposites will be discussed.

Signal‐Amplifying Biohybrid Material Circuits for CRISPR/Cas‐Based Single‐Stranded RNA Detection

AbstractThe functional integration of biological switches with synthetic building blocks enables the design of modular, stimulus‐responsive biohybrid materials. By connecting the individual modules via diffusible signals, information‐processing circuits can be designed. Such systems are, however, mostly limited to respond to either small molecules, proteins, or optical input thus limiting the sensing and application scope of the material circuits. Here, a highly modular biohybrid material is design based on CRISPR/Cas13a to translate arbitrary single‐stranded RNAs into a biomolecular material response. This system exemplified by the development of a cascade of communicating materials that can detect the tumor biomarker microRNA miR19b in patient samples or sequences specific for SARS‐CoV. Specificity of the system is further demonstrated by discriminating between input miRNA sequences with single‐nucleotide differences. To quantitatively understand information processing in the materials cascade, a mathematical model is developed. The model is used to guide systems design for enhancing signal amplification functionality of the overall materials system. The newly designed modular materials can be used to interface desired RNA input with stimulus‐responsive and information‐processing materials for building point‐of‐care suitable sensors as well as multi‐input diagnostic systems with integrated data processing and interpretation.

Assembly of Differently Sized Supercharged Protein Nanocages into Superlattices for Construction of Binary Nanoparticle-Protein Materials.

This study focuses on the design and characterization of binary nanoparticle superlattices: Two differently sized, supercharged protein nanocages are used to create a matrix for nanoparticle arrangement. We have previously established the assembly of protein nanocages of the same size. Here, we present another approach for multicomponent biohybrid material synthesis by successfully assembling two differently sized supercharged protein nanocages with different symmetries. Typically, the ordered assembly of objects with nonmatching symmetry is challenging, but our electrostatic-based approach overcomes the symmetry mismatch by exploiting electrostatic interactions between oppositely charged cages. Moreover, our study showcases the use of nanoparticles as a contrast enhancer in an elegant way to gain insights into the structural details of crystalline biohybrid materials. The assembled materials were characterized with various methods, including transmission electron microscopy (TEM) and single-crystal small-angle X-ray diffraction (SC-SAXD). We employed cryo-plasma-focused ion beam milling (cryo-PFIB) to prepare lamellae for the investigation of nanoparticle sublattices via electron cryo-tomography. Importantly, we refined superlattice structure data obtained from single-crystal SAXD experiments, providing conclusive evidence of the final assembly type. Our findings highlight the versatility of protein nanocages for creating distinctive types of binary superlattices. Because the nanoparticles do not influence the type of assembly, protein cage matrices can combine various nanoparticles in the solid state. This study not only contributes to the expanding repertoire of nanoparticle assembly methods but also demonstrates the power of advanced characterization techniques in elucidating the structural intricacies of these biohybrid materials.

Enhancing the stability of anthocyanins extracts through adsorption into nanoclays – development of a smart biohybrid sensor for intelligent food packaging or as natural food additive/preservative

Anthocyanins (ACNs) are bioactive compounds sensible to high temperature, light and highly reactive to pH changes. These natural pigments present different colors according to the environment pH, which enables its use as bio-based sensor to control microbial quality of fresh perishable foodstuff. In this work, a biohybrid material (BH) composed of anthocyanins extract from jambolan fruit (Syzygium cumini) stabilized by adsorption onto montmorillonite (Mt) was developed, characterized and tested to monitor freshness of shrimp. Anthocyanins were recovered from extract using Mt as adsorbent at pH 1.5, 2.5, and 3.5 and temperatures of 10 and 20 °C. Langmuir and pseudo-second-order kinetic models were adjusted to explain these adsorption mechanisms. ACNs’ adsorption increased with decreasing pH (p > 0.05) and increasing temperature. At 20 °C and pH 1.5, 93 % of ACNs were recovered from extract (adsorption of 212.04 mg of ACNs/g of Mt). The obtained BH presented good stability and color changing capacity when exposed to ammonia vapor at pH varying from 1 to 13. Freshness of shrimp was monitored with the BH, validating its potential to be used as ammonia sensing or pH indicators in intelligent packaging.

Characterization of Functional Biohybrid Materials Based on Saccharomyces Cerevisiae Biomass

Biohybrid materials and engineered living materials (ELMs) are a dynamic field of research at the interface of material science and synthetic biology. Recently, microorganisms have been described as both functional and structural building blocks of such materials. Dry materials with bacteria as structural building block have previously not been investigated for their ability to retain catalytic activity. Herein, it is shown that Saccharomyces cerevisiae biomass can act as a structural building block in dry, macroscopic materials while simultaneously providing functionality. A benign method for the preparation of both coatings and free‐standing, flexible films is presented. Notably, free‐standing films can be prepared solely from biomass and plasticizers without the need for other additives. During the process, the integrity of the cells and the catalytic activity within is preserved, which is demonstrated by biocatalytic H2O2 degradation. The film‐forming properties of S. cerevisiae biomass and the influence of plasticizing and polymeric additives on the mechanical properties are investigated in detail and compared to related materials. The work presented is a first step toward new, mechanically strong microorganism‐based functional coatings and films.

Design of a Biohybrid Materials Circuit with Binary Decoder Functionality (Adv. Mater. 14/2024)

Design of a Biohybrid Materials Circuit with Binary Decoder Functionality (Adv. Mater. 14/2024)

ATP-Responsive Manganese-Based Bacterial Materials Synergistically Activate the cGAS-STING Pathway for Tumor Immunotherapy.

Stimulating the cyclic guanosine monophophate(GMP)-adenosine monophosphate (AMP) synthase (cGAS)-stimulator of interferon genes (STING) pathway is a crucial strategy by which bacteria activate the tumor immune system. However, the limited stimulation capability poses significant challenges in advancing bacterial immunotherapy. Here, an adenosine 5'-triphosphate (ATP)-responsive manganese (Mn)-based bacterial material (E. coli@PDMC-PEG (polyethylene glycol)) is engineered successfully, which exhibits an exceptional ability to synergistically activate the cGAS-STING pathway. In the tumor microenvironment, which is characterized by elevated ATP levels, this biohybrid material degrades, resulting in the release of divalent manganese ions (Mn2+) and subsequent bacteria exposure. This combination synergistically activates the cGAS-STING pathway, as Mn2+ enhances the sensitivity of cGAS to the extracellular DNA (eDNA) secreted by the bacteria. The results of the in vivo experiments demonstrate that the biohybrid materials E. coli@PDMC-PEG and VNP20009@PDMC-PEG effectively inhibit the growth of subcutaneous melanoma in mice and in situ liver cancer in rabbits. Valuable insights for the development of bacteria-based tumor immunotherapy are provided here.

Hierarchical assembly of peptoids on MoS2

Bio-macromolecular assembly forms intricate scaffolds in living systems that allow them to biomineralize bone, enamel, and shell. Such bioinspired approaches have also been widely used in biohybrid materials synthesis. Recently, researchers have demonstrated functional bioelectronic structures by assembling biopolymers, including proteins and peptides, on van der Waals (VdW) materials for sensors and energy storage and harvesting. However, the hierarchical assembly of biopolymers on VdW materials has yet to be fully documented, because modulating the assembly architectures by controlling the energy landscape along with environmental stimuli, in which various assembly phases occur, remains challenging. In this study, we focused on peptoids, biomimetic polymers with properties similar to peptides but offering advantages such as greater side chain diversity, lower complexity due to elimination of backbone-backbone hydrogen binding, lower synthesis costs, and improved stability. Specifically, we designed two short peptoid sequences capable of assembling on MoS2. Our findings reveal diverse self-assembled phases, including monolayer hybrid films with high crystallinity, vesicle-like nanoparticles, lamellae, and multi-layer ribbons. We elucidated the assembly processes of these states and confirmed the occurrence of phase transitions between them by using in situ atomic force microscopy (AFM). The results highlight the critical roles of peptoid-peptoid, peptoid-solvent, and peptoid-MoS2 interactions. These insights significantly advance the understanding needed to design hierarchical biopolymer architectures at VdW material-solvent interfaces with potential implications for enhancing the performance of bioelectronic devices based on 2D vdW materials.

Supramolecular Self-Assembly of Proteins Promoted by Hybrid Polyoxometalates.

Controlling the formation of supramolecular protein assemblies and endowing them with new properties that can lead to novel functional materials is an important but challenging task. In this work, a new hybrid polyoxometalate is designed to induce controlled intermolecular bridging between biotin-binding proteins. Such bridging interactions lead to the formation of supramolecular protein assemblies incorporating metal-oxo clusters that go from several nanometers in diameter up to the micron range. Insights into the self-assembly process and the nature of the resulting biohybrid materials are obtained by a combination of Small Angle X-ray Scattering (SAXS), Transmission Electron Microscopy (TEM), and Dynamic Light Scattering (DLS), along with fluorescence, UV-vis, and Circular Dichroism (CD) spectroscopy. The formation of hybrid supramolecular assemblies is determined to be driven by biotin binding to the protein and electrostatic interactions between the anionic metal-oxo cluster and the protein, both of which also influence the stability of the resulting assemblies. As a result, the rate of formation, size, and stability of the supramolecular assemblies can be tuned by controlling the electrostatic interactions between the cluster and the protein (e.g., through varying the ionic strength of the solution), thereby paving the way toward biomaterials with tunable assembly and disassembly properties.

Design of a Biohybrid Materials Circuit with Binary Decoder Functionality.

Synthetic biology applies concepts from electrical engineering and information processing to endow cells with computational functionality. Transferring the underlying molecular components into materials and wiring them according to topologies inspired by electronic circuit boards has yielded materials systems that perform selected computational operations. However, the limited functionality of available building blocks is restricting the implementation of advanced information-processing circuits into materials. Here, a set of protease-based biohybrid modules the bioactivity of which can either be induced or inhibited is engineered. Guided by a quantitative mathematical model and following a design-build-test-learn (DBTL) cycle, the modules are wired according to circuit topologies inspired by electronic signal decoders, a fundamental motif in information processing. A 2-input/4-output binary decoder for the detection of two small molecules in a material framework that can perform regulated outputs in form of distinct protease activities is designed. The here demonstrated smart material system is strongly modular and can be used for biomolecular information processing for example in advanced biosensing or drug delivery applications.

PenTag, a Versatile Platform for Synthesizing Protein‐Polymer Biohybrid Materials

AbstractThe site‐specific and covalent conjugation of proteins on solid supports and in hydrogels is the basis for the synthesis of biohybrid materials offering broad applications. Current methods for conjugating proteins to desired targets are often challenging due to unspecific binding, unstable (noncovalent) coupling, or expensive and difficult‐to‐synthesize ligand molecules. Here, is presented PenTag, an approach for the bioorthogonal, highly specific, and covalent conjugation of a protein to its ligand for various applications in materials sciences. Penicillin‐binding protein 3 (PBP3) is engineered and shows that this protein can be used for the stable and spontaneous conjugation of proteins to dyes, polymers, or solid supports. PenTag as a crosslinking tool is applied for synthesizing stimuli‐responsive hydrogels or for the development of a biohybrid material system performing computational operations emulating a 4:2 encoder. Based on this broad applicability and the use of a small, cheap, and easy‐to‐functionalize ligand and a stable, soluble recombinant protein, is seen PenTag as a versatile approach toward biohybrid material synthesis.

New Biocide Based on Tributyltin(IV) Ferulate-Loaded Halloysite Nanotubes for Preserving Historical Paper Artworks.

Combining biologically active compounds with nanocarriers is an emerging and promising strategy for enhancing the activities of molecules while reducing their levels of toxicity. Green nanomaterials have recently gained momentum in developing protocols for treating and preserving artifacts. In this study, we designed a functional biohybrid material by incorporating tributyltin(IV) ferulate (TBT-F) into halloysite nanotubes (HNTs), generating a new formulation called HNT/TBT-F. The primary objective was to develop a formulation with robust antimicrobial properties and reinforcing features for treating paper with artistic and historical value. To characterize HNT/TBT-F, assess the HNT's loading capacity, and investigate the TBT-F release kinetics from the nanotubes, various analytical techniques, including UV-Vis and infrared spectroscopies, thermogravimetry, and microscopy analysis, were employed. Furthermore, we evaluated the antimicrobial potential of TBT-F and HNT/TBT-F against Kocuria rhizophila, a bacterial strain known for its opportunistic behavior and a cause of artifact biodeterioration. HNT/TBT-F exhibited a significantly stronger bactericidal effect than TBT-F alone against K. rhizophila cells growing planktonically or those forming a biofilm. This enhanced performance could relate to the confinement of TBT-F within the nanotubes, which likely improved its physical-chemical stability and increased the local concentration of TBT-F upon contact with the bacterial cells. Additionally, we evaluated the mechanical properties of a paper treated with HNT/TBT-F, assessing any potential alterations in its color. The findings of this study highlight the favorable attributes of the HNT/TBT-F formulation and its potential for developing protocols aimed at consolidating and preserving culturally significant paper objects.

"3-in-1" Hybrid Biocatalysts: Association of Yeast Cells Immobilized in a Sol-Gel Matrix for Determining Sewage Pollution.

This study presents a novel ″3-in-1″ hybrid biocatalyst design that combines the individual efficiency of microorganisms while avoiding negative interactions between them. Yeast cells of Ogataea polymorpha VKM Y-2559, Blastobotrys adeninivorans VKM Y-2677, and Debaryomyces hansenii VKM Y-2482 were immobilized in an organosilicon material by using the sol-gel method, resulting in a hybrid biocatalyst. The catalytic activity of the immobilized microorganism mixture was evaluated by employing it as the bioreceptor element of a biosensor. Optical and scanning electron microscopies were used to examine the morphology of the biohybrid material. Elemental distribution analysis confirmed the encapsulation of yeast cells in a matrix composed of methyltriethoxysilane (MTES) and tetraethoxysilane (TEOS) (85 and 15 vol %, respectively). The resulting heterogeneous biocatalyst exhibited excellent performance in determining the biochemical oxygen demand (BOD) index in real surface water samples, with a sensitivity coefficient of 50 ± 3 × 10-3·min-1, a concentration range of 0.3-31 mg/L, long-term stability for 25 days, and a relative standard deviation of 3.8%. These findings demonstrate the potential of the developed hybrid biocatalyst for effective pollution monitoring and wastewater treatment applications.

Cellulose nanofibers infused with pomegranate gold nanoparticles display antibacterial activity

This study investigated the antibacterial activity of cellulose nanofibers (CNFs) impregnated with gold nanoparticles (AuNPs) functionalized with pomegranate extract. The CNFs were obtained from cellulose extracted from coconut fiber. The pomegranate-AuNPs were characterized by UV-Vis, transmission electron microscopy (TEM), dynamic light scattering (DLS) and inductively coupled plasma mass spectrometry (ICP-MS). The cytotoxicity was analyzed by an MTT assay in murine fibroblasts (L929) and human keratinocytes (HaCaT). CNFs impregnated with pomegranate-AuNPs were characterized by TEM, energy dispersive X-ray (EDX) analysis, ICP-MS and DLS, and the antibacterial activity was analyzed against the gram-positive strains: Staphylococcus aureus and Enterococcus faecalis, and the gram-negative strains: Escherichia coli and Pseudomonas aeruginosa. In this study, we show that CNFs impregnated with pomegranate-AuNPs form a stable, non-cytotoxic material with expressive antibacterial activity, as demonstrated by a reduction of between 60% and 99% in the number of S. aureus, E. coli and P. aeruginosa colonies. Therefore, the biohybrid material developed here, which is both inexpensive and easy to produce, shows considerable potential for antibacterial and industrial applications.

Nanostructured biohybrid material with wide-ranging antiviral action

Nanostructured biohybrid material with wide-ranging antiviral action