Immobilization Efficiency Research Articles

Functionalization of sawdust biochar using Mg-Fe-LDH and sodium dodecyl sulfonate enhanced its stability and immobilization capacity for Cd and Pb in contaminated water and soil

The increased contamination of potentially toxic element (PTE) has posed remarkable ecological risks to environment. Application of functionalized biochar for the remediation of PTE contaminated water and soils are of great concern, and effective strategies are urgently needed to enhance the removal capacity of biochar for PTE. As a novel surface modification technology, the effect of layered double hydroxides (LDH) and sodium dodecyl sulfonate (SDS) on the remediation capacity of biochar for PTE polluted soils and water remains unclear. Sawdust biochar (SB) was coated with Mg and Fe to synthesize the Mg-Fe-LDH functionalized biochar (MFB); thereafter, the MFB was mixed with SDS solution to synthesize the organic-Mg-Fe-LDH biochar (MSB). The potential of SB, MFB, and MSB for remediation of Cd and Pb contaminated soil and water was evaluated in terms of adsorption capacity, immobilization efficiency, and stability. Loading of Mg-Fe-LDH into SB, along with SDS treatment created a regular micro-nano hierarchical structure and enhanced the surface roughness, aromaticity, and hydrophobicity of MSB as compared to SB. MSB exhibited a significantly higher maximum adsorption capacity (mg g−1) for water Pb (405.2) and Cd (673.0) than MFB (335.9 for Pb and 209.0 for Cd) and SB (178.2 for Pb and 186.1 for Cd). MSB altered the soluble fraction of Cd/Pb to the residual fraction and thus significantly decreased their mobilization in soil. The higher removal/immobilization efficiency of MSB could be attributed to its alkalinity, and the enhanced synergistic interactions including surface precipitation, ion exchange, complexation, and hydrogen bonding. The resistance to carbon loss by H2O2, thermal recalcitrance index R50, and degree of graphitization in MSB were significantly improved compared to SB, indicating a more stable carbon fraction sequestered in MSB following aging in soil. These results indicate that MSB could be used for remediation of Cd and Pb contaminated soil and water.Graphical

Functionalization of titanium using KR-12 peptide: antibacterial and osteogenic properties

Titanium implants are widely used in orthopedic and dental applications owing to their biocompatibility and mechanical stability. Nonetheless, they encounter significant challenges pertaining to infection and poor osteointegration. We investigated the feasibility of two different coating strategies for titanium with the antimicrobial peptide KR-12: direct binding by way of carbodiimide chemistry and indirect attachment through a polydopamine (PDA) layer, comparing these approaches in a comprehensive study. This study assessed the immobilization efficiency, antibacterial efficacy, and osteogenic capability of both coating techniques. The findings indicated that PDA-mediated immobilization achieved greater KR-12 binding efficiency (36.80%) compared with direct covalent binding (24.08%) and enhanced antibacterial effectiveness against Staphylococcus aureus, with colony counts decreasing by 98% for the Ti-PDA-KR12 group and by 95% for the Ti-KR12 group compared with the control group. In vitro investigations utilizing rat bone marrow–derived mesenchymal stem cells demonstrated that PDA-KR12-coated surfaces markedly enhanced cell adhesion, proliferation, and osteogenic differentiation, as indicated by elevated alkaline phosphatase activity, osteocalcin synthesis, and upregulation of osteogenesis-associated gene markers. Our findings indicate that PDA-mediated peptide immobilization is a superior approach for augmenting both the antibacterial characteristics and osteogenic potential of titanium implants, presenting a possible method to enhance implant performance.

Immobilization of Paenibacillus polymyxa with biopolymers to enhance the production of 2,3-butanediol

BackgroundPaenibacillus polymyxa, is a Gram-positive, plant growth promoting bacterium, known for producing 98% optically pure 2,3-butanediol, an industrially valuable chemical for solvents, plasticizers and resins. Immobilization of Paenibacillus polymyxa has been proposed to improve the cell stability and efficiency of the fermentation process, reduce contamination and provide easy separation of butanediol in the culture broth as compared to conventional bioprocesses. This research aimed to explore the potential of Paenibacillus polymyxa with immobilization technique to produce 2,3-butanediol.ResultsWe investigated different immobilization methods with natural biopolymers like alginate, chitosan and carrageenan-chitosan-based immobilization. These methods were further investigated for their immobilization efficiency and yield in 2,3-butanediol production. Carrageenan-chitosan beads enabled a higher cell concentration and demonstrated superior cell retention to calcium-alginate-chitosan beads. Carrageenan-chitosan immobilization preserved 2,3-butanediol production in bacteria and increased the product formation rate.ConclusionCarrageenan-chitosan immobilization enables non-pathogenic Paenibacillus polymyxa to be a capable 2,3-butanediol producer with increased product formation rate, which has not been previously reported. This novel strategy offers promising alternative to traditional fermentation processes using pathogenic strains and can be further applied in co-cultivations for metabolite production, wastewater management and bioremediation.

Rational design of redox active metal organic frameworks for mediated electron transfer of enzymes.

The efficient immobilization of redox mediators remains a major challenge in the design of mediated enzyme electrode platforms. In addition to stability, the ability of the redox-active material to mediate electron transfer from the active-site buried enzymes, such as flavin adenine dinucleotide-dependent glucose dehydrogenase (FADGDH) and lactate oxidase (LOx), is also crucial. Conventional immobilization techniques can be synthetically challenging, and immobilized mediators often exhibit limited durability, particularly in continuous operation. Here, we design a novel redox-active cobalt-based metal-organic framework (raMOF) obtained via the partial ligand substitution of 2-methylimidazole (MeIm) with a 1,2-naphthoquinone-4-sulfonate (NQSO) redox probe, as a promising platform for high-performance enzyme electrodes. This nanostructured raMOF, combined with multi-walled carbon nanotubes (CNTs), provided a high current density of up to 2.06 mA cm-2 during enzymatic reactions and maintained remarkable operational stability, retaining 100% of its current over 54 hours. This stability far exceeded that of adsorbed NQSO on CNTs, which experienced a complete loss of the initial current, highlighting the significant advantage of the raMOF-based platform for high-performance enzyme electrodes.

Immobilization of Glycosyltransferase into a Hydrophilic Metal-organic Framework for Efficient Biosynthesis of Chondroitin Sulfate.

Chondroitin sulfate (CS) is a structurally complex anionic polysaccharide widely used in medical, cosmetic and food applications. Enzymatic catalysis is an important strategy for synthesizing CS with uniform chain lengths and well-defined structures. However, the industrial application of glycosyltransferases is hindered by limitations such as low expression yields, poor stability, and challenges in reuse. We developed a mild and rapid one-step synthetic method for the efficient immobilization of chondroitin synthase (KfoC). The resulting KfoC@ZIF-90 composite exhibits high catalytic activity, thermal stability, and pH adaptability. Notably, KfoC@ZIF-90 exhibited 5-fold enhanced thermal stability at 40°C and retained 86% relative activity at pH 10, while also maintaining 90% activity in organic solvents, surpassing the performance of free KfoC. Molecular docking analysis revealed that encapsulating KfoC within hydrophilic ZIF-90 reduced its binding energy to the substrate, thereby improving catalytic performance. Furthermore, KfoC@ZIF-90 can be easily separated from the reaction solution by centrifugation, simplifying product isolation and purification while enabling enzyme reuse. These attributes significantly enhance operability and reduce processing costs, making enzymatic CS synthesis more feasible for industrial applications.

Direct Printing of an Electrochemical Device and Its Interface with Paper for Uric Acid Detection in Human Sweat.

Using a laser-scribed (direct printing) technique, we have fabricated an enzymeless, mediatorless, and paper-interfaced electrochemical device (P-LSG) for uric acid detection on a flexible polyimide sheet. Various paper substrates were investigated, and it was found that Whatman filter paper-1 is promising to obtain the best electrochemical signals at the small volume of electrolyte, i.e., 20 μL. Furthermore, the Whatman filter paper-1 was modified with gold nanoparticles (AuNPs) to improve the electrocatalytic activity of the P-LSG device. The fabricated AuNP-modified P-LSG biosensor exhibited excellent electrocatalytic activity for the detection of uric acid over a wide range of 10 to 750 μM, with sensitivity of ∼0.214 μA μM-1 cm-2, and a limit of detection of ∼1.4 μM. The sensor was further validated by using the UHPLC-ESI-MS/MS technique, and the observed percentage recovery was less than 10%. This work opens the window to modified paper substrates with various nanomaterials to improve the sensing parameters. The electrolyte storage capacity and rich chemistry of paper additionally provide an efficient immobilization platform for biorecognition elements to diagnose other metabolites. Furthermore, it has the potential to analyze the volume of small samples (like sweat, tears, urine, etc.) using paper to develop noninvasive wearable biosensors.

Exploiting unique NP1 interface: Oriented immobilization via electrostatic and affinity interactions in a tailored PDA/PEI microenvironment enhanced by concanavalin A.

Enzyme immobilization techniques are crucial for enhancing enzyme stability and catalytic efficiency. Traditional methods such as physical adsorption and simple covalent binding often fail to maintain enzyme activity and stability. In this study, an innovative multi-level immobilization strategy was proposed to achieve efficient targeted immobilization of nuclease P1 (NP1) by fine-tuning the surface microenvironment. Molecular simulation results revealed that the distinctive electrostatic distribution and the specific placement of basic amino acids, such as lysine, on the NP1 surface caused dopamine to preferentially adsorb on areas away from NP1's active site. This selective adsorption facilitated the directed immobilization of NP1, while the positively charged environment generated by the co-deposited surface further enhanced NP1's adsorption capacity. This multilevel modification was found to significantly optimize the physicochemical environment of the immobilized surface through surface characterization and enzymatic testing. This strategy greatly improves enzyme activity (3590.0 U/mg), stability, and reusability (70% after 10 cycles). In particular, NP1 on this surface exhibited an optimal Michaelis constant (Km) of 34.0mM and a maximum reaction rate of 5.5mMmin-1, demonstrating the remarkable effect of the modification strategy in enhancing the enzyme catalytic performance. The present study provides an efficient and stable immobilization platform for enzyme catalytic applications by precisely modulating the surface microenvironment and the oriented immobilization strategy, which has an important potential for practical applications. This stable and reusable NP1 platform allows for efficient DNA/RNA cleavage, facilitating its application in industrial biocatalysis, biomedical enzyme-based processes, and biosensors.

SpyFixer enables efficient site-specific immobilization for protein-protein interaction analysis and antibody purification.

Traditional methods of protein immobilization often result in activity loss due to random coupling. This study introduces SpyFixer, a variant of SpyCatcher that achieves over 99% efficient site-specific protein immobilization. We applied SpyFixer on two platforms: bio-layer interferometry (BLI) for protein-protein interaction analysis and epoxy agarose resin for antibody purification. Using human growth hormone (hGH) and the Z domain of Protein A as model proteins, we demonstrated that SpyFixer enables efficient, site-specific immobilization on BLI sensors, yielding reproducible kinetic data with lower variability than conventional methods. Additionally, we developed a cost-effective strategy for antibody purification utilizing SpyFixer-modified resin, which exhibited remarkable capture efficiencies exceeding 90%, elution efficiencies over 70%, and purities over 90% for human immunoglobulin G (hIgG) from complex samples, including bacterial lysates, human serum, and recombinant fermentation broth. The resin's loading capacity surpassed 200mg/mL, and no significant activity loss was observed after 20 regeneration cycles. This study further advances the potential of Spy chemistry in biotechnological applications.

Facile colorimetric detection of as(V) in Rice with immobilized acid phosphatase on hollow metal-organic frameworks hybrid with peroxidase-like activity

Quantitative analysis of As(V) in rice is of great significance for food safety and heavy metal pollution control. Here, a facile colorimetric method for As(V) detection was constructed by using immobilized acid phosphatase (ACP) in hollow metal-organic frameworks hybrid. Metalloporphyrin and gold nanoparticles modified hollow zeolite imidazole framework-8 [Au/HZIF-8@TCPP(Fe)], named AuHT, was chosen here as ACP immobilizing carrier with peroxidase-like activity. Firstly, the morphology, structure, immobilization efficiency and catalytic ability of obtained AuHT@ACP were fully characterized. Then, based on the inhibition of As(V) on immobilized ACP and cascade reaction mediated by ACP and AuHT, a colorimetric biosensor was established with excellent simplicity. After comprehensive validation, this colorimetric method presented the advantages of wide linear range (10.0–1000.0 μg/L), low LOD (4.0 μg/L), nice accuracy (recovery of 93.7–109.6 %) and good selectivity. Finally, this method was applied to visual detection of As(V) in rice samples with different varieties.

Immobilization of lipase on mesoporous silica nanocarriers for efficient preparation of phytosterol esters

Phytosterol esters (PEs) are favored for their good solubility, high bioavailability, and various health benefits. Mesoporous silica (MS) nanocarriers were synthesized and utilized to immobilize free lipase AYS “Amano” (from Candida rugosa) for further preparing PEs. The optimal immobilization conditions were an initial enzyme content of 160 mg/mL, pH of 7.0, temperature of 30 °C, and adsorption time of 6 h, resulting in a protein loading of 42.3 mg/g and an immobilization efficiency of 73.4 %. Compared to commercial carriers, MS carriers demonstrated superior immobilization efficiency and phytosterol conversion rates when soybean oil was used as the PEs source for the transesterification. The phytosterol conversion rate reached 97.7 % in a solvent system with optimal reaction conditions. After 30 days of storage, the immobilized lipase retained 55.7 % of its activity, and it had 63.0 % retention activity after 10 reuse cycles. This study significantly enhanced the lipase's phytosterols conversion and reusability, supporting sustainable and efficient industrial production of PEs.

Extracellular Lipase from Pseudomonas aeruginosa SB-37: Production by Solid State Fermentation, Immobilization, and Characterization

Native lipase is still promising as an industrial biocatalyst. This study aimed to investigate the production of native local lipase using solid state fermentation (SSF) methods, immobilization the lipase by Ca-alginate entrapment, and characterization based on substrate preferences. To obtain high lipase production using SSF methods, we optimized the type of agro-wastes substrates, fermentation time, oil induction percentage and volume of preculture percentage. The optimal condition for lipase production via solid-state fermentation involved a 7% (v/v) preculture of Pseudomonas aeruginosa SB-37, utilizing palm kernel meal as the substrate, supplemented with 6% (v/w) oil induction (soybean oil:tween 80 = 75:25) at 50 °C for 24 h. This gave a lypolitic activity value of 2 U/gds (gram dry weight substrates). Since the protein profile of extracellular lipase has a few protein bands, we perform direct immobilization on crude protein supernatant. Immobilization by Ca-alginate entrapment results in loading capacity and recovery activity values of 86.84% and 148%, respectively. The immobilized lipase retained 92% activity until four batch repetition and keep 40% activity at tenth batch. The highest hydrolytic activity of immobilized lipase was 0.9 U/g gel on the pNP_8 substrates. While the highest transesterification activity was observed with acetonitrile solvent and substrates of pNP_8 and isopropanol with the activity value at 0.6 U/g gel. This present study emphasized the feasibility of producing lipase as a biocatalysts using economical agro-industrial wastes and efficient immobilization using entrapment method. Copyright © 2024 by Authors, Published by BCREC Publishing Group. This is an open access article under the CC BY-SA License (https://creativecommons.org/licenses/by-sa/4.0).

Spherical Nucleic Acid Probes on Floating-Gate Field-Effect Transistor Biosensors for Attomolar-Level Analyte Detection.

Field-effect transistor (FET) sensors are attractive for the label-free detection of target biomolecules, offering ultrahigh sensitivity and a rapid response. However, conventional methods for modifying biomolecular probes on sensors often involve intricate and time-consuming procedures that require specialized training. Herein, we propose a simple and versatile approach to functionalize floating-gate (FG) FET sensors by exploiting the strong binding ability of polyvalent interactions and the three-dimensional structure of densely functionalized spherical nucleic acids (SNAs). Crucially, the SNAs can be easily deposited onto a dielectric layer under mild conditions, ensuring stable immobilization of the probes. Further, the SNAs show efficient and robust immobilization on various dielectric layers including Y2O3, Ta2O5, and HfO2, forming conjugates that resist denaturation by various agents. By modifying the DNA sequence within the SNAs, we achieved highly sensitive FG-FET biosensors for DNA, adenosine triphosphate, and viral nucleic acids at the attomolar level. For clinical samples detection, unamplified enterovirus 71 RNA at levels as low as 0.13 copies μL-1 was detected within 100 s. Moreover, the sensor attained 100% accuracy for analyte detection in both positive and negative samples. Our findings provide a general and simple method for fabricating FET-based biochemical sensors and demonstrate that the SNA-modified FG-FET biosensor is a versatile and reliable integrated platform for ultrasensitive biomarker detection.

An Electrochemical Oxidation and Intercalation Strategy for Iodide Removal Using LDHs.

Radioactive iodide harms the ecosystem and human health, necessitating its immobilization to mitigate aquatic iodine pollution. Layered double hydroxides (LDHs), a family of 2D clays with intercalated anions and controllable interlayer structures, are technologically and economically viable adsorbents to eliminate various anion pollutants. However, LDHs exhibit an extremely low affinity toward iodide species. Here, an electrochemical oxidation and intercalation strategy is developed to successfully remove and immobilize iodide using LDHs. The iodide can be transformed into I2/I3 - and subsequent IO3 - ions on NiFe-LDH/CC electrode at 0.6 and 1.0V (vs Ag/AgCl), with an iodine adsorption capacity of 962 and 355mg g-1, respectively. I2/I3 - are adsorbed on the surface of NiFe-LDH with a weak interaction, while IO3 - inserts into the interlayer of LDH with a high affinity due to the strong hydrogen bonding and coordination between IO3 - and Ni atoms. This LDH-based electrochemical removal strategy also has high adaptability to a low iodide concentration, wide pH range, and outstanding ion selectivity. This study can lay the foundation for efficient iodide elimination and immobilization from radioactive liquids.

Machine Learning-Assisted SERS Reveals the Biochemical Signature of Enhanced Protein Secretion from Surface-Modified Magnetic Nanoparticles.

This study introduces a novel investigation of the interaction between Komagataella phaffii cells and iron oxide-based magnetic nanoparticles (Fe3O4 MNPs) via protein secretion and machine learning (ML)-assisted surface-enhanced Raman scattering (SERS). For the first time, we produced Fe3O4, Fe3O4@PEG, Fe3O4@PEI10kDa, and Fe3O4@PEI25kDa MNPs by a one-pot coprecipitation reaction. The addition of polymers to the reaction conditions significantly affected the shape, surface charge, size, and size distribution of the MNPs. The surface modification of MNPs is effectively accomplished using polyethylenimine (PEI), and the ζ-potential values of the MNPs exceed +25 mV under the NH4OH control. The homogeneity of MNPs synthesized with NH4OH is more pronounced according to transmission electron microscopy (TEM) pictures. All MNPs exhibited excellent immobilization efficiency (>92%) when we used 250 ppm Fe-containing MNP solutions. Smaller MNPs uniformly encapsulated the surface of K. phaffii cells, whereas larger MNPs exhibited irregular accumulation. K. phaffii cells exhibited excellent viability in all MNP solutions at up to 1000 ppm of Fe concentrations. Finally, the highest recombinant azurin protein secretion rate was obtained in Fe3O4@PEI10kDa MNP-immobilized cells (about 1.3 times). The ML-assisted SERS analysis revealed that MNP interactions with K. phaffii cells were mediated by proteins such as mannoproteins and membrane transporter proteins as well as N-acetylglucosamine (i.e., chitin). These findings revealed the effect of the size and surface properties of MNPs on the immobilization of K. phaffii cells and the enormous potential of magnetic immobilization for protein secretion.

Ultrasensitive Organic Photoelectrochemical Transistor Biosensor Based on DNA as Nanocarrier for Efficient Immobilization of Signal Probe with Pimping Background.

Organic photoelectrochemical transistor (OPECT) assays are mainly focused on the improvement of photoactive materials at the gating interface, which leads to an enhanced initial signal alongside significant background noise. This phenomenon can cause considerable deviations and impose limits on target detection. In this study, we achieved sensitive and low-background detection of alkaline phospholipase (ALP) activity using single-stranded DNA (ssDNA) as a nanocarrier to effectively immobilizing CdS quantum dots (QDs) sensitizing UiO-66. Specifically, ssDNA-labeled CdS QDs could be connected to UiO-66 via Zr-O-P bonds, while sodium tripolyphosphate (STPP) interfered with the binding of CdS QD-ssDNA to UiO-66 due to an occupancy effect. The hydrolysis of STPP, catalyzed by ALP, diminished this occupancy effect, allowing for increased binding of CdS QDs-ssDNA to UiO-66 and thereby enhancing the responsive signal. The competitive dynamic between STPP and ssDNA allowed for controlled binding of CdS QDs-ssDNA photosensitizers on UiO-66, thus realizing sensitive detection of ALP. This organic integration of biological modulation and signal amplification in OPECT has led to the creation of a novel sensing platform for detecting ALP concentrations ranging from 0.005 to 100 U·L-1, and the detection limit is 0.0012 U·L-1, thus paving the way for innovative approaches to sensitively monitor ALP activity.

PET Foils Functionalized with Reactive Copolymers as Adaptable Microvolume ELISA Spot Array Platforms for Multiplex Serological Analysis of SARS-CoV-2 Infections.

Microvolume ELISA platforms have become vital in diagnostics for their high-throughput capabilities and minimal sample requirements. High-quality substrates with advanced surface properties are essential for these applications. They enable both efficient biomolecule immobilization and antifouling properties, which are critical for assay sensitivity and specificity. This study presents PET-based microvolume ELISA spot arrays coated with amine- and DBCO-reactive copolymers MCP-2 and Copoly Azide. The platforms were designed for the sensitive and specific detection of specific antibodies such as COVID-19 biomarkers. Supporting robust attachment of the SARS-CoV-2 nucleoprotein (NP), these arrays outperform traditional approaches. It was demonstrated that covalent attachment methods proved more efficient than passive adsorption, together with the reduction of non-specific binding. Analytical performance was verified with classical ELISA and real-time Surface Plasmon Resonance (SPR) analysis. It enables sensitive detection of IgG and IgA antibodies, including IgG subclasses, in human serum. Clinically, the platform achieved 100.0% sensitivity and 92.9% specificity for anti-NP antibody detection in COVID-19-positive and negative samples. Additionally, DNA-directed immobilization extended the platform's utility to multiplex serological measurements. These findings underscore the potential of PET-based microvolume ELISA arrays as scalable, high-throughput diagnostic tools suitable for detecting multiple biomarkers in a single assay and easily integrated into microfluidic devices.

Cofactor immobilization for efficient dehydrogenase driven upgrading of xylose

In this study, the coupled immobilization on nanosilica of xylose dehydrogenase and alcohol dehydrogenase was accomplished with high efficiency, which was 90 %. Moreover, immobilization of the cofactors oxidized and reduced nicotinamide adenine dinucleotide, i.e., NAD+ and NADH, on silica material was examined and the impact on the effectiveness of the process was determined. The highest efficiency of NAD+ immobilization was found to be 56 %, which was obtained after 24 h of immobilization at 30 °C, pH 7 For NADH, the best immobilization efficiency was 53 % which was achieved after 24 h at 25 °C, pH 7. The KM and Vmax values were determined for various configurations of the biocatalytic systems showing, as expected, that immobilization of the enzymes decreased the catalytic rate (Vmax) and slightly increased the KM, but verifying that the immobilization of the cofactors did not significantly affect the kinetics, but would enable high conversion, and potentially continued enzymatic reaction. The use of the system configuration with co-immobilized enzymes, immobilized NAD+ and immobilized NADH thus allowed for obtaining over 90 % efficiency of xylose conversion in one batch, which was significantly higher than the systems with single free or only one immobilized cofactor. Using UV-Vis measurements, it was confirmed that effective cofactor regeneration occurred in the systems with immobilized components thus allowing for sustained enzyme catalyzed upgrading of xylose to xylonic acid.

Optimization of the conditions for immobilization of crude pectinase on chitin using ultrasound and glutaraldehyde and its application in grape juice clarification

The current research focuses on optimizing the immobilization of crude pectinase, produced by solid-state fermentation, onto chitin using ultrasound and glutaraldehyde as a crosslinking agent for grape juice clarification. A Box-Behnken design was used to model the effects of ultrasound time (5–15 min), ultrasound amplitude (20 – 60 %), and water-to-support ratio for ultrasound application (16−24) on immobilized crude pectinase activity and immobilization efficiency. Optimal conditions, comprising ultrasound time of 10 min, ultrasound amplitude of 20 %, and water-to-support ratio for ultrasound application of 24, yielded an immobilized crude pectinase activity of 2.19 IU/mL and 74 % immobilization efficiency. The immobilized crude pectinase showed improved kinetic properties and retained 69 % activity after three use cycles. Both free and immobilized crude pectinase enhanced juice clarity, reducing sugar, and TSS while lowering viscosity.

Harnessing Lignin Nanoparticles for Sustainable Enzyme Immobilization: Current Paradigms and Future Innovations.

Lignin, a vital plant component, is key in providing structural integrity and is the second most abundant biopolymer in nature. The growing interest in sustainable and efficient biocatalysis has driven the exploration of lignin nanoparticles (LNPs) as a promising platform for enzyme immobilization. Given lignin's abundance and structural role in plants, converting it into nanoparticles offers a potential eco-friendly alternative to traditional supports. This comprehensive review explores recent advancements in using LNPs for enzyme immobilization, focusing on loading techniques, immobilization efficiency, enzyme activity levels, and various factors that affect the performance of enzymes immobilized on LNPs. The review also addresses the primary challenges associated with enzyme immobilization on LNPs and discusses future innovations in this field. Adopting eco-friendly immobilization platforms based on LNPs is expected to have broad applications in industries like food, pharmaceuticals, animal feed, and detergents. However, there is still potential to customize LNPs further and develop novel immobilization techniques to leverage their benefits fully. By understanding the properties and advantages of these nanostructured lignin supports, researchers can design and create innovative nanocatalysts for various industrial applications.

Enhancing the Stability and Anticancer Activity of Escherichia coli Asparaginase Through Nanoparticle Immobilization: A Biotechnological Perspective on Nano Chitosan.

There is a shortage in the experimental research directly comparing the effectiveness of different nanoparticles in boosting asparaginase (ASNase) activity. This study assessed the impact of various nanoparticles on enhancing ASNase activity, stability, and anticancer effects through immobilization. Escherichia coli ASNase was immobilized on different nanoparticles, and its efficiency was measured. The research included analyzing the enzyme's secondary structure, stability, activity at different temperatures, kinetic parameters, shelf life, and activity in blood serum. The anticancer efficacy was determined by measuring the IC50. The study also investigated the anticancer mechanisms by examining the enzyme's toxicity on cancer cells, focusing on apoptosis indicators like nuclear intensity, membrane permeability, mitochondrial membrane permeability, and cytochrome c release. Among the tested nanoparticles, nano chitosan yielded the best improvements. ASNase immobilized on nano chitosan reached 90% immobilization efficiency fastest among the studied nanoparticles, achieving this within 72 h, whereas other nanoparticles took 120 h. Immobilization modified ASNase's secondary structure by increasing alpha helices and reducing random coils, with nanochitosan and magnetic iron oxide showing the most pronounced effects. Immobilized ASNase exhibited enhanced activity, stability across temperature (widest with nanochitosan, 25-65 °C), and a broader optimal pH range compared to the free enzyme, with a Km of 1.227 mM and a Vmax of 454.54 U/mg protein. Notably, the nano-chitosan-immobilized ASNase retained over 85% of its activity after 9 months of storage and maintained high activity in blood serum. This improved stability and activity translated into the highest anticancer activity (Lowest IC50) and was more effective than doxorubicin in disrupting cancer cell structures.