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Metal–organic frameworks (MOFs), with large specific surface area and tunable porosity, have gained lots of attention for immobilizing enzymes. However, the intrinsic open channels of most reported MOFs are generally smaller than 2 nm, which significantly prevents the passage of enzymes, and the diffusion efficiency of substrates and products. Here we report a new hierarchical micro-mesoporous zeolitic imidazolate framework-8 (ZIF-8) with core-shell superstructure (HZIF-8) using colloidal hydrated zinc sulfate (ZnSO4•7H2O) as a soft template for enzyme immobilization. The ZnSO4•7H2O forms an aggregation of colloids due to the self-conglobation effect in methanol, which affords a soft template for the formation of HZIF-8. Cytochrome C (Cyt C) was immobilized in interior of HZIF-8 through entrapment during the formation of HZIF-8. The resultant immobilized Cyt C (Cyt C@HZIF-8) exhibited 4-fold and 3-fold higher activity than free Cyt C and Cyt C encapsulated in conventional microporous ZIF-8 (Cyt C@ZIF-8), respectively. Meanwhile, the Km value of Cyt C@HZIF-8 significantly decreased due to the presence of mesopores compared with Cyt C@ZIF-8, indicating enhanced substrate affinity. After 7 cycles, Cyt C@HZIF-8 still maintained 70% of its initial activity whereas Cyt C@ZIF-8 only retained 10% of its initial activity. Moreover, the obtained HZIF-8 showed outstanding performance in co-immobilization of multi-enzyme for the detection of glucose.

In this study, the surfactant activated lipase from Aspergillus oryzae was used to prepare magnetic hybrid nanoflowers by embedding Fe3O4 magnetic nanoparticles (MNPs) into hybrid nanoflowers (MhNF). Meanwhile, MNPs were integrated into the activated lipase hybrid nanoflowers through covalent cross-linking (cross-linked-MhNF). Activity recovery of the MhNF and cross-linked-MhNF was 190% and 174%, respectively. However, activity recovery of lipase hybrid nanoflowers (hNF) only retained 77%. Furthermore, the MhNF displayed higher Kcat/Km value than free lipase, indicating high catalytic efficiency of MhNF. Compared with free lipase, the MhNF exhibited high tolerance against methanol and storage stability. Furthermore, the MhNF could be easily recycled by magnet without obvious activity loss. After reusing for 10 cycles, the activated hNF only retained 26% activity of its original activity, while the MhNF still maintained 84% activity of its initial activity, indicating excellent reusability. In the reaction of producing biodiesel from soybean oil, the MhNF-catalyzed biodiesel yield reached 88%, while the free enzyme was only 69%. The yield of biodiesel catalyzed by MhNF was maintained at 76% even after 6 consecutive cycles.

In the recent past, numerous new types of nanostructured carriers, as support matrices, have been engineered to advance the traditional enzyme immobilization strategies. The current research aimed to develop a robust enzyme-based biocatalytic platform and its effective deployment in the industrial biotechnology sectors at large and catalysis area, in particular, as low-cost biocatalytic systems. Suitable coordination between the target enzyme molecules and surface pendent multifunctional entities of nanostructured carriers has led an effective and significant contribution in myriad novel industrial, biotechnological, and biomedical applications. As compared to the immobilization on planar two-dimensional (2-D) surface, the unique physicochemical, structural and functional attributes of nano-engineered matrices, such as high surface-to-volume ratio, surface area, robust chemical and mechanical stability, surface pendant functional groups, outstanding optical, thermal, and electrical characteristics, resulted in the concentration of the immobilized entity being substantially higher, which is highly requisite from applied bio-catalysis perspective. Besides inherited features, nanostructured materials-based enzyme immobilization aided additional features, such as (1) ease in the preparation or green synthesis route, (2) no or minimal use of surfactants and harsh reagents, (3) homogeneous and well-defined core-shell nanostructures with thick enzyme shell, and (4) nano-size can be conveniently tailored within utility limits, as compared to the conventional enzyme immobilization. Moreover, the growing catalytic needs can be fulfilled by multi-enzymes co-immobilization on these nanostructured materials-based support matrices. This review spotlights the unique structural and functional attributes of several nanostructured materials, including carbon nanotubes, graphene, and its derivate constructs, nanoparticles, nanoflowers, and metal-organic frameworks as robust matrices for laccase immobilization. The later half of the review focuses on the applied perspective of immobilized laccases for the degradation of emergent contaminants, biosensing cues, and lignin deconstruction and high-value products.

Nanostructured materials constitute an interesting and novel class of support matrices for the immobilization of peroxidase enzymes. Owing to the high surface area, robust mechanical stability, outstanding optical, thermal, and electrical properties, nanomaterials have been rightly perceived as immobilization matrices for enzyme immobilization with applications in diverse areas such as nano-biocatalysis, biosensing, drug delivery, antimicrobial activities, solar cells, and environmental protection. Many nano-scale materials have been employed as support matrices for the immobilization of different classes of enzymes. Nanobiocatalysts, enzymes immobilized on nano-size materials, are more stable, catalytically robust, and could be reused and recycled in multiple reaction cycles. In this review, we illustrate the unique structural/functional features and potentialities of nanomaterials-immobilized peroxidase enzymes in different biotechnological applications. After a comprehensive introduction to the immobilized enzymes and nanocarriers, the first section reviewed carbonaceous nanomaterials (carbon nanotube, graphene, and its derivatives) as a host matrix to constitute robust peroxidases-based nanobiocatalytic systems. The second half covers metallic nanomaterials (metals, and metal oxides) and some other novel materials as host carriers for peroxidases immobilization. The next section vetted the potential biotechnological applications of the resulted nanomaterials-immobilized robust peroxidases-based nanobiocatalytic systems. Concluding remarks, trends, and future recommendations for nanomaterial immobilized enzymes are also given.

ZrCoCe getter films with thickness of ~ 2.3 μm were deposited on Si(100) wafers by direct current (DC) magnetron sputtering process. A 400-nm-thick Pd protection layer was then deposited on the as-deposited ZrCoCe film without exposure to atmosphere. Microstructure, surface morphology and surface chemical state of the films were analyzed. Moreover, hydrogen sorption properties were determined. The results show that the ZrCoCe film displays a cauliflower-like morphology and a porous columnar-like structure which is composed of nanocrystal grains. The Pd protection layer tightly adheres to the surface of the ZrCoCe film and efficiently prevents the oxidation of Zr under exposure to atmosphere. We find that the hydrogen sorption properties of the Pd-ZrCoCe film are significantly improved, in comparison with those of the as-deposited ZrCoCe film.

Multienzymatic conversion of carbon dioxide (CO2) into chemicals has been extensively studied. However, regeneration and reuse of co-factor are still the main problems for the efficient conversion of CO2. In this study, a nanoscale multienzyme reactor was constructed by encapsulating simultaneously carbonic anhydrase (CA), formate dehydrogenase (FateDH), co-factor (NADH), and glutamate dehydrogenases (GDH) into ZIF-8. In the multienzyme reactors, cationic polyelectrolyte (polyethyleneimine, PEI) was doped in the ZIF-8 by dissolving it in the precursors of ZIF-8. Co-factor (NADH) was anchored in ZIF-8 by ion exchange between PEI (positive charge) and co-factor (negative charge), and regenerated through GDH embedded in the ZIF-8, thus keeping high activity of FateDH. Activity recovery of FateDH in the multienzyme reactors reached 50%. Furthermore, the dissolution of CO2 in the reaction solution was increased significantly by the combination of CA and ZIF-8. As a result, the nanoscale multienzyme reactor exhibited superior capacity for conversion of CO2 to formate. Compared with free multienzyme system, formate yield was increased 4.6-fold by using the nanoscale multienzyme reactor. Furthermore, the nanoscale multienzyme reactor still retained 50% of its original productivity after 8 cycles, indicating excellent reusability.

Polyvinyl alcohol (PVA) is a synthetic polymer that is difficult to degrade in nature. In this study, we synthesized PVA-degrading enzymes (PVAase)-Cu3(PO4)2 hybrid nanoflowers by using crude PVAase from Bacillus niacini for PVA degradation. The influences of PVAase concentration, Cu2+ concentration, and incubation time on the nucleation and activity of the PVAase hybrid nanoflower were investigated. The maximal activity recovery of the PVAase hybrid nanoflower was approximately 85% at 0.25 mg/mL of PVAase, 0.36mM Cu2+, and 72 h incubation time. The optimum temperature and pH of PVAase did not change before and after immobilization. Compared with free PVAase, the PVAase hybrid nanoflower showed high thermal stability and storage stability. Additionally, the PVAase hybrid nanoflower displayed excellent reusability after eight cycles and promising PVA degradability, indicating its potential application in PVA degradation.

e-Poly-L-lysine (e-PL) is a natural antimicrobial cationic peptide. Antimicrobial activity of e-PL is closely related to its molecular weight (Mw). However, the antimicrobial mechanisms of e-PL with different Mws are still vague. In this study, Saccharomyces cerevisiae was used as the model system to analyze the mechanism from these three aspects: cell wall, cell membrane, and metabolism. The results showed that high-Mw e-PL (1-3 kDa and >3 kDa) and commercial e-PL product caused cell wall lesions, and significantly improved cell membrane permeability compared to low-Mw e-PL ( 3 kDa) and product displayed higher inhibition effect on the glycolysis pathway and tricarboxylic cycle than that of low-Mw e-PL (

Baker's yeast industries generate highly polluted effluents, especially the cell free broth (i.e., vinasse) characterized by high chemical oxygen demand, nitrogen, and salts. In this work, it was found that the residual by-products (i.e., ethanol and acetic acid) and salts in the vinasse severely inhibited the cell growth, which hindered the reuse of the vinasse for the production of Saccharomyces cerevisiae. Through optimizing a suitable control strategy, the productions of ethanol and acetic acid were eliminated. Then, a nanofiltration membrane (i.e., NF5) was preferred for preliminarily and simultaneously separating and concentrating valuable molecules (i.e., invertase, food grade proteins and pigments) in the vinasse, and the main fouling mechanism was cake layer formation. Subsequently, a reverse osmosis membrane (RO) was suitable to separate and concentrate salts in the NF5 permeate, where the membrane fouling was negligible. Finally, the RO permeate was successfully reused for the production of S. cerevisiae. In addition, without calculating the benefit from the recovery of the valuable molecules, the cost of the integrated process can be decreased by 59.8% compared with the sole triple effect evaporation. Meanwhile, the volume of the fresh water used in the fermentation process can be decreased by 68.8%. Thus, it is a sustainable process for the cleaner production of baker's yeast using the integrated fermentation and membrane separation process.

Biological metal-organic frameworks (bioMOFs) are a new subclass of the MOF family. In comparison with traditional MOFs, the bioMOFs are made of multifunctional biologically related ligands (bio-ligand) and metal ions. The bio-ligands confer biological compatibility for traditional MOFs, thus providing many opportunities for a wide array of biological applications. This review highlights the recent advances in the synthesis of bioMOFs comprising multifunctional bio-ligands and metal ions. These bio-ligands include nucleobases, amino acids, peptides, proteins, cyclodextrin, saccharides, and other biomolecules. Furthermore, the potential bio-applications of bioMOFs in several fields such as biomedicine, biosensing and bioimaging, antimicrobial applications, biomimetic catalysis, chiral separation, and environmental protection are also demonstrated.

Zeolitic imidazole frameworks (ZIFs) with tunable pore sizes and high surface areas have recently used as an effective support for immobilizing enzymes. However, the instability in the aqueous acidic environment has limited their practical applications in some cases. In this work, we develop a novel catalase/ZIFs composite with mesoporous silica shell (mSiO2@CAT/ZIFs) via co-precipitation, and controlled self-assembly of silanes. During preparation, the cetyltrimethylammonium bromide induced the formation of the mesostructured silica layer on the outer surface of CAT/ZIFs. The resultant mSiO2@CAT/ZIFs exhibited high activity recovery (92%). Compared with the conventional CAT/ZIFs and free CAT, mSiO2@CAT/ZIFs exhibited excellent acid resistance. For example, after 30 min in acetate buffer solution (pH 3.0), the CAT/ZIFs and free CAT almost lost activity whereas the mSiO2@CAT/ZIFs still retained 35% of original activity. Meanwhile, the thermostability of the mSiO2@CAT/ZIFs was enhanced significantly compared with conventional CAT/ZIFs. In addition, the mSiO2@CAT/ZIFs displayed excellent storage stability, and retained 60% of its initial activity after 15 days storage period. Furthermore, the mSiO2@CAT/ZIFs could maintain 70% of its initial activity after 8 continuous uses, demonstrating superior reusability than the free CAT and CAT/ZIFs. These results demonstrated that the mSiO2@CAT/ZIFs are potential for practical applications even in the acidic environment.

Enzyme catalysis has been attracting increasing interest in the past twenty years. Nevertheless, most reports concerning enzyme catalysis have been carried out using single enzymes. Recent years, multiple enzyme cascade reactions have a significant role for the production of many compounds at an industrial level because they permit to perform very complex reactions. Especially, the development of coimmobilized multienzymatic systems is increasingly driven by economic and environmental constraints that provide an impetus to develop alternatives to conventional multistep synthetic methods. Up to now, process optimization and novel strategies of coimmobilized multienzymatic systems hardly have been reviewed. In this review, we focus on some recent novel techniques in preparing co-immobilized multienzymatic systems and the up-to-date advances in the application of multienzymatic systems. Moreover, we also discuss the improvements that co-immobilization multienzymatic systems offer enzymes such as reusability, catalytic activity, and stability.

Bacterial cellulose (BC) has numerous excellent properties but the absence of antibacterial activity restricts its applications in biomedical field. Therefore, in order to introduce the antibacterial characteristics into BC; herein, a facile method for incorporation of ZnO nanoparticles (ZnO-NPs) is presented. BC films were first immersed in zinc nitrate solution, followed by treating with NaOH solution, the BC loaded ZnO nanocomposite films were dried by a sheet former instrument at 80 °C for 20 min. The obtained BC/ZnO nanocomposites were characterized by different techniques. XRD results showed the hexagonal wurtzite structure of ZnO-NPs while FE-SEM results displayed the particle size of ZnO-NPs was ranging from 70 to 100 nm. Thermogravimetric study revealed the thermal stability of nanocomposite films. The nanocomposite exhibited photocatalytic activity and revealed 91% degradation of methyl orange (MO) under UV-irradiation within 2 h. Moreover, the nanocomposites demonstrated significant UV-blocking properties and showed antibacterial activities against tested Gram-positive and Gram-negative bacterial strains. This work provides a simple and novel method for the synthesis of BC/ZnO nanocomposite as a functional biomaterial.

Background As a natural antibacterial cationic peptide, ε-poly-l-lysine (ε-PL) is applied as a food preservative. However, the mechanism of ε-PL against Staphylococcus aureus (S. aureus) has not been elucidated. Especially, its antimicrobial mechanism at the metabolomics has not been yet thoroughly described. Results This work aimed at clarifying the antibacterial activity and mechanism of ε-PL against S. aureus. Effects of ε-PL with different concentration on cell morphology, cell wall, and membrane integrity were investigated. Furthermore, the effect of ε-PL on metabolite properties of S. aureus was also studied. The results revealed that ε-PL disrupted the cell wall and membrane integrity of treated cells. ε-PL induced the structural change of peptidoglycan in cell wall, causing cell wall more fragile. Meanwhile, the permeability of the S. aureus cell membrane was increased by ε-PL. More importantly, ε-PL with different concentration could cause different effects on metabolic pathways of S. aureus. ε-PL with high concentration could directly restrain the central carbon metabolism. However, ε-PL with low concentration could only inhibit the glycolytic pathway. Conclusion These results showed that the antimicrobial mechanism of ε-PL against S. aureus was a synergistic action.

In this study, carbonic anhydrase (CA, EC 4.2.1.1) molecules were embedded into metal-organic frameworks (MOFs) via co-precipitation (CA@ZIF-8), and then these CA@ZIF-8 nanocomposites were encapsulated in the poly(vinyl alcohol) (PVA)-chitosan (CS) hydrogel networks to prepare CA@ZIF-8-PVA-CS composite hydrogels (PVA/CS/CA@ZIF-8) with high activity, stability, and reusability. The immobilization efficiency of CA was greater than 70%, suggesting the high immobilization efficiency. The prepared PVA/CS/CA@ZIF-8 composite membranes displayed excellent higher stability against a high temperature, denaturants, and acid than free CA and CA@ZIF-8. Furthermore, these membranes exhibited an excellent performance for CO2 capture. The amount of calcium carbonate obtained by PVA/CS/CA@ZIF-8 hydrogel membranes was 20- and 1.63-fold than free CA and CA@ZIF-8 composites, respectively. Furthermore, the hydrogel membranes exhibited superior reusability and mechanical strength. The hydrogel membrane maitained 50% of its original activity after 11 cycles. However, CA@ZIF-8 completely lost activity. These results indicated that the PVA/CS/CA@ZIF-8 membranes can be efficiently applied to capture CO2 sequestration.

Metal-organic frameworks (MOFs) have been emerged as a promising support for immobilizing enzymes owing to the tunable porosity, high surface area, and structural diversity. However, most of these possess nanometer size and small pores, which are difficult to recover them from the reaction medium and present low immobilization efficiency and protein loading capacity, and high substrate diffusion limitations. Herein, a novel magnetic amino-functionalized zeolitic imidazolate framework-8 (ZIF-8) with 3D highly ordered macroporous structure was synthesized using the assembled polystyrene (PS) nanosphere monoliths as a template. Subsequently, catalase (CAT) molecules were immobilized on the surface of macroporous magnetic ZIF-8 and inside the macropores by precipitation, covalent binding and cross-linking. The resultant immobilized CAT showed high immobilization efficiency (58%) and protein loading capacity (29%), leading to 500% higher activity than the immobilized CAT on ZIF-8 (CAT/ZIF-8). Meanwhile, the immobilized CAT could be easily recovered with a magnet without obvious activity loss. The traditional CAT/ZIF-8 lost its activity after 6 cycles, whereas, the immobilized CAT retained 90% activity of its initial activity after reusing for 8 cycles, indicating excellent reusability. In conclusion, this study provides a facile and efficient approach to immobilize enzymes on/in MOFs with enhanced activity and excellent recyclability.

The Phenylketonuria (PKU) is an inborn defect of phenylalanine (Phe) metabolism, in which Phe accumulated in the blood causing alterations at the central nervous system. Therefore, the detection of PKU is very important for the early diagnosis of PKU patients. However, existing tests for PKU are time-consuming and require high-resource laboratories. In this study, a novel paper-based biosensor based on phenylalnine ammonia lyase (PAL) hybrid nanoflowers was constructed that provides a semi-quantitative output of the concentration of Phe from urine samples. PAL@Ca3(PO4)2 hybrid nanoflowers (PAL@NF) were first prepared using PAL and Ca2+. Synthesis conditions of the PAL@NF on the formation of the PAL@NF were optimized. The PAL@NF exhibited 90% activity recovery under optimal condition. Compared with free PAL, the PAL@NF displayed good storage stability and increased tolerance to proteolysis. After five consecutive operating cycles, the PAL@NF still retained 73% of its initial activity, indicating excellent reusability. Furthermore, the paper-based biosensor was able to detect Phe concentration in urine samples, and exhibited good linearity to the Phe concentrations in the range from 60 to 2400 μM and the response time was only about 10 min. Therefore, the paper-based biosensor can be a promising candidate as a biosensor for the detection of PKU.