Immobilization Of Glucose Oxidase Research Articles

A high-power hybrid carbon nanotube/three-dimensional reduced graphene oxide glucose/O2 enzymatic biofuel cell

Long-lasting, high-performance enzymatic biofuel cells (EBFCs) are among the most promising candidates for renewable and green power generation. There are two main drawbacks to EBFCs: 1) short enzyme lifetime due to the enzyme's (protein) denaturation; and 2) low electron transfer efficiency as the enzyme's active site is deeply buried inside the non-conductive protein structure of the enzyme. In this study, a hybrid of carbon nanotubes (CNT) and three-dimensional graphene (3DG) was used to immobilize the glucose oxidase (GOx) on the surface of a glassy carbon electrode (GCE) to fabricate a modified bioanode in a glucose/O2 EBFC. This facile and low-cost CNT/3DG hybrid is a solution to the problems of instability, short lifespan, and poor electron transport in EBFCs. The exceptional electrocatalytic activity of the porous CNT/3DG hybrid is demonstrated by its effective immobilization of GOx and ability to collect energy from glucose oxidation by direct electron transfer (DET) while preserving the structure of the enzyme. The CNT acts like a tiny electrical wire that reaches and harvests the electron directly from the flavin adenine dinucleotide (FAD) (the redox center of GOx) and transfers it to the electrode surface, thus enhancing the performance of the EBFC. The immobilized GOx lifetime was increased to 210 days with a maximum power density of 253.36 µWcm-2 at 0.65 V. These results demonstrate the attractive capability of the CNT/3DG hybrid in the development of efficient biofuel cells and biosensors that employ enzymes in their structures.

Entrapment of glucose oxidase from Aspergillus niger ISL-09 in poly(acrylamide-co-acrylic acid) hydrogels for improved stability and catalytic efficiency towards industrial applications

The present study highlights the true potential of Aspergillus niger ISL-09 to produce glucose oxidase (GOx) and focuses on improving the catalytic properties of GOx by entrapping it in poly acrylamide-co-acrylic acid (AAm-co-AAc) hydrogels, fabricated by free-radical polymerization method. For this purpose, the optimum conditions for GOx activity were found to be: 15 g soybean meal (SBM) substrate, 20 mL moisture content, 96 h incubation period, and 1.5 mL of fungal spore suspension. The purified extracellular GOx was observed to have 32.4% yield, with 0.02 U/mg specific activity and approximately 80 kDa subunit molecular weight. The GOx was most active at pH 5.5 and 40 °C. The immobilization of GOx in poly (AAm-co-AAc) hydrogels led towards improved stability and catalytic efficiency, resulting in a 21.7% increase in activity compared to free enzyme. The study also examined the potential of GOx in the pharmaceutical and textile industries as Ca-gluconate producer and bleaching agent, respectively. The study concludes that A. niger ISL-09 is a promising source for GOx production under optimal conditions. Furthermore, the immobilization of GOx in poly (AAm-co-AAc) hydrogels can significantly improve its catalytic properties, making it suitable for different industrial applications. However, further scaling up is required for the better implementation in industry.

Multi-walled carbon nanotubes functionalized with a new Schiff base containing phenylboronic acid residues: applicationtothe development of a bienzymatic glucose biosensor using a response surface methodology approach.

Aninnovative supramolecular architectureis reportedfor bienzymatic glucose biosensing based on the use of a nanohybrid made of multi-walled carbon nanotubes (MWCNTs) non-covalently functionalized with a Schiff base modified with two phenylboronic acid residues (SB-dBA) as platform for the site-specific immobilization of the glycoproteins glucose oxidase (GOx) and horseradish peroxidase (HRP). The analytical signal was obtained from amperometric experiments at - 0.050V in the presence of 5.0 × 10-4M hydroquinone as redox mediator. The concentration of GOx and HRP and the interaction time between the enzymes and the nanohybrid MWCNT-SB-dBA deposited at glassy carbon electrodes (GCEs) were optimized through a central composite design (CCD)/response surface methodology (RSM). The optimal concentrations of GOx and HRP were 3.0mgmL-1 and 1.50mgmL-1, respectively, while the optimum interaction time was 3.0min. The bienzymatic biosensor presented a sensitivity of (24 ± 2) × 102 µA dL mg-1 ((44 ± 4) × 102 µA M-1), a linear range between 0.06mg dL-1 and 21.6mg dL-1 (3.1µM-1.2mM) (R2 = 0.9991),and detection and quantification limits of 0.02mg dL-1 (1.0µM) and 0.06mg dL-1 (3.1µM), respectively. The reproducibility for five sensors prepared with the same MWCNT-SB-dBA nanohybrid was 6.3%, while the reproducibility for sensors prepared with five different nanohybrids and five electrodes each was 7.9%. The GCE/MWCNT-SB-dBA/GOx-HRP was successfully used for the quantification of glucose in artificial human urine and commercial human serum samples.

Pore-selective immobilization of pH-sensitive polymer and glucose oxidase in the porous polyimide film for detection of glucose

Glucose selective oxidizing enzyme of glucose oxidase (GOD) was immobilized on the carboxyl group (–COOH) functionalized porous polyimide (PI) film (PI-GOD film). The –COOH functionalized film was fabricated by the modified breath figure method (BF) by casting the PI solution under humid conditions containing KOH. The amine group of GOD was covalently bonded to the PI-COOH film under catalytic conditions for the PI-GOD film. The PI-GOD film induced the color change of the bromothymol blue (BTB) indicator solution from green to yellow by adding glucose due to the oxidation of glucose by GOD to gluconic acid acting as an acidic source. Amine-terminated pH-sensitive polymethacrylic acid polymer (PMAA) was additionally immobilized to the PI-GOD film for the PI-GOD-PMAA film. The morphology of the pore surface of the PI-GOD-PMAA film was changed by the addition of glucose due to the coil-to-globule transition of the pH-sensitive polymer by the production of gluconic acid as a result of the reaction between glucose and GOD. The insulin adsorbed on the film was released by adding glucose due to the morphology change of the pH-sensitive polymer. The detection sensitivity of glucose was increased by the pore-selectively immobilized GOD acting as a micro-reactor.

Determining the kinetic and optimum characteristics of glucose oxidase immobilized on polyurethane

Abstract Objectives The primary aim of this study is to immobilize glucose oxidase (GOX) as a model enzyme using a simple and efficient method without the use of any toxic chemicals. This will enable the multiple reusability of GOX, thereby reducing the process cost. Furthermore, the objective is to determine the kinetic, optimum pH, and temperature parameters in comparison to soluble GOX. Methods For immobilization, GOX at a concentration of 5 mg/mL was mixed with an equal volume of hypol prepolymer in a beaker. Immobilization was achieved by the formation of amide bonds between –NH2 groups on the enzyme surface and the prepolymer. The activities of polyurethane foam (PUF)-immobilized and soluble GOX were monitored using horse radish peroxidase enzyme as a second enzyme. 2,2′-Azino-bis-3-ethylbenzothiazoline-6-sulfonic acid radical ABTS+ generated as a result of the enzymatic reactions was detected at 420 nm. Results The kinetic parameters for soluble GOX were Km=9.39 ± 1.13 mM and Vmax=9.14 ± 0.45 A.U sec−1 × 10−4, while for PUF-immobilized GOX Km=10.04 ± 1.30 mM and Vmax=9.09 ± 0.49 A.U sec−1 × 10−4. PUF-immobilized GOX displayed high performance during reuse experiments. PUF-immobilization led to the activity at higher temperatures and over a wide pH range compared to its soluble counterpart. Conclusions GOX was successfully immobilized for the first time using PUF. This technique can be used in the healthcare and food sectors as well as in biosensor applications. In the future, when successfully applied to many enzymes of industrial value, it could offer significant advantages.

Composite PEDOT-PSS based highly sensitive electrochemical sensors for sensing glucose from human saliva

Here we report two composites, especially two different 2D materials and Au nanoparticle composite with conducting polymer PEDOT-PSS as electrodes for highly sensitive, cost effective and reliable biosensors. Conducting polymer composite electrodes of AuNP/WS2/PEDOT-PSS and AuNP/GO/PEDOT-PSS were prepared by electrochemical deposition and were studied using various structural and electrochemical techniques. XRD, FTIR and SEM have been used for structural characterization which confirmed the elemental compositions of these composite electrodes. Electrochemical Impedance Spectroscopy (EIS) and Cyclic Voltammetry (CV) were used to examine the charge transfer kinetics and electroactive properties of the modified electrodes. Impedance spectra analysis had shown the reduced charge transfer resistance and diffusive property of AuNP/WS2/PEDOT-PSS electrodes compared to AuNP/GO/PEDOT-PSS. To demonstrate biosensing advantages of these electrodes, interaction between covalently immobilized glucose oxidase (GlOx) enzyme and D-glucose was monitored via chronoamperometry and CV. GlOx/AuNP/WS2/PEDOT-PSS/ITO yielded wider linear range from 0.74 to 440.67 µM towards the oxidation of glucose with high selectivity and lower limit of detection (LOD) of 1 µM. AuNP/GO/PEDOT-PSS/ITO composite electrode exhibited linearity in the range 3.84–373.33 µM and LOD 2.33 µM. Better sensitivity and wider linear range of GlOx/AuNP/WS2/PEDOT-PSS/ITO has been assigned to special layered structure as sheets formed by incorporation of WS2 to PEDOT-PSS. Validation of the glucose sensors has shown satisfactory results using real saliva samples. Study suggests use of conducting polymer, 2D layered material and metal nanoparticle composite electrodes based sensors offer wider linearity range and higher sensitivity for stable, reliable, cost- effective sensor electrodes which can be used easily in non-clinical diagnostics.

Developing Flexible Carbon Nitride Quantum Dots Decorated Polyaniline Nanocomposite Layer for a Non-Invasive Wearable Sweat Biosensor for Glucose Monitoring

Understanding dynamic biomarker fluctuations, particularly glucose, is vital for early diabetes therapy. Novel non-invasive wearable sweat biosensors are gaining appeal, enabling constant severity monitoring. Nevertheless, the key challenges are that multiplexed motions may result in undesirable metal cracking by the rigid electrocatalytic layer, leading to decreased sensitivity and stability. In this work, we designed carbon nitride quantum dots (CNQDs) decorated polyaniline (PANI) nanocomposite to realize flexible wearable biosensors with high detection accuracy. Upon the enhanced electron transfer by CNQDs, CNQDs/PANI nanocomposite offers greater sensitivity in H2O2 detection compared to pristine PANI, resulting in a sensitivity of 37.21 μA mM-1 cm-2. Therefore, after the glucose oxidase (GOx) immobilization, GOx/CNQDs/PANI-based biosensors is expected to show excellent performance for glucose detection in artificial sweat. Figure 1

Advancements in Glucose Monitoring: A Thin Film ZnO-Nanoflakes Based Highly Sensitive Wearable Biosensor for Noninvasive Sweat-Based Point-of-Care Monitoring for Diabetes

In the contemporary global landscape, diabetes is a prevalent and concerning presence that contributes to noteworthy mortality rates. The etiology of this condition involves a subtle interaction between insulin deficiency and elevated blood sugar levels which results in a complex metabolic disorder. The harmful consequences of this physiological abnormality become evident through the increased probability of kidney failure, heart attack, chronic liver diseases, and a pernicious ocular affliction leading to blindness. Against this backdrop, the necessity for prompt, accurate, and continuous monitoring of glucose concentrations with any form of flexible sensor becomes notably evident for type I and type II diabetes patients, pregnant women with diabetes, athletes, and high-risk persons. Sweat, an often-overlooked physiological excretion, is a reservoir of vital biological information due to its accessibility and analyzability. It is an advantageous matrix for glucose monitoring, surpassing blood, saliva, and tears. Its non-invasive nature offers a patient-friendly alternative to conventional blood sampling. The continuous, real-time nature of sweat extraction facilitates dynamic monitoring without the discomfort associated with invasive methods. This inherent advantage positions sweat as a promising matrix for glucose sensing, underscoring its potential as a valuable tool in diabetes management. It overcomes the limitations posed by other bodily fluids like saliva and tears in terms of accessibility, patient comfort, and reflective accuracy. The assimilation of nanomaterials into biosensing technologies heralds a paradigm shift, conferring heightened sensitivity upon these mechanisms. Noteworthy among these innovations are flexible wearable biosensors leveraging zinc oxide (ZnO) thin films, offering promising avenues for the discernment of glucose levels within sweat for healthcare monitoring. ZnO nanoflakes (ZnO-NFs) distinguish themselves by endowing flexible biosensors with unparalleled sensitivity, reliability, continuous monitoring capabilities, and non-invasiveness from zinc oxide nanoparticles incorporated graphene-carbon nanotube hybrid (GR-CNT-ZnO) and ergocalciferol-multi walled carbon nanotubes based (ERGOc-MWCNTd/Nf) traditional glucose sensors. The distinctive advantages of ZnO-NFs over bulk materials are multifaceted. Notably, the polarized (0001) plane orientation inherent in ZnO-NFs amplifies enzyme immobilization, thereby augmenting sensing performance. The elevated isoelectric point (9.5) of ZnO-NFs facilitates the immobilization of biomolecules, obviating the need for an ancillary binding layer. A single-step and efficacious sonochemical approach has been used to synthesize a thin layer of ZnO-NFs virtually on any substrate. The electrochemical flexible glucose biosensor was fabricated through the judicious immobilization of glucose oxidase (GOx) on ZnO nanoflakes, possessing a mere 20nm thickness and synthesized adeptly on an Au-coated stretchable polyethylene terephthalate (PET). The sensitivity of the flexible biosensor was determined by 29.97 μA/decade/cm² and the minimum detection limit is 100 μM. The coefficient of determination of the designed flexible electrode is found 97.76%, which is significantly linear. The designed PET/Au/ZnO-NFS/GOx flexible biosensors showed good repeatability for different sample concentrations. The enzymatic flexible sensor also showed good repeatability for three sample concentrations. The reproducibility data for two identical flexible sensors were found within the LoA of the Bland-Altman plot with a statistically significant p-value of 0.08 derived from the student t-test. Moreover, the flexible sensor exhibited notable repeatability, reproducibility, and remarkable stability conducive to the precise quantification of glucose. The application of this flexible wearable sensor has been validated through its utilization in human sweat samples, yielding results that align favorably with conventional methodology.

Synthesis and comparison of the performance of two different water-soluble phthalocyanine based electrochemical biosensor

Herein, a comparative study between novel water-soluble phthalocyanine-based biosensors was performed for the application of glucose sensing. For this purpose, two different copper (II) and manganese (III) phthalocyanines and their water-soluble derivatives were synthesized, and then their role as a supporting material for enzyme immobilization was evaluated by comparing their sensor performances. Two different phthalocyanine (AP-OH2-MnQ (MnPc) and AP-OH2-CuQ (CuPc)) were tested using electrochemical biosensor with immobilized glucose oxidase (GOx). To the best of our knowledge, the related water-soluble phthalocyanine-based glucose biosensors were attempted for the first time, and the developed approach resulted in improved biosensor characteristics. The constructed biosensors GE/MnPc/GOx and GE/CuPc/GOx showed good linearity between 0.003–1.0 mM and 0.05–0.4 mM, respectively. The limit of detection was estimated at 0.0026 mM for the GE/MnPc/GOx and 0.019 mM for the GE/CuPc/GOx. KMapp and sensitivity values were also calculated as 0.026 mM and 175.043 µAmM−1 cm−2 for the GE/MnPc/GOx biosensor and 0.178 mM and 117.478 µAmM−1 cm−2 for the GE/CuPc/GOx biosensor. Moreover, the fabricated biosensors were successfully tested to detect glucose levels in beverages with high recovery results. The present study shows that the proposed water-soluble phthalocyanines could be a good alternative for quick and cheap glucose sensing with improved analytical characteristics.

Evaluation of Electrochemical Sensor Using Microfluidic System

This paper presents the characterization and testing of electrochemical sensor using microfluidic system. Various geometric patterns were laser cut into the platinum working electrode of a biosensor. In this work, a microfluidic chamber was designed that allows phosphate buffer solution (PBS) to flow across the sensor, using a peristaltic pump, while varying the concentration of hydrogen peroxide. The amperometric characterization of the electrochemical sensor with 25, 50, 75, and 100[Formula: see text][Formula: see text]m perforation and 75[Formula: see text][Formula: see text]m spacing showed the highest sensitivity. This result was to be expected the purpose of patterning the sensors was to provide a 3-dimensional structure to the planar electrode in order for the enzyme, glucose oxidase, to be immobilized. Future work will include selecting one of the patterns for immobilization of glucose oxidase allowing us to realize a fully functional glucose sensor.

Enzymes on chemical gardens: Chemobrionics-based electrochemical biosensor

The relationship between biological materials and the immobilization matrix is a critical point in attaching biomolecules onto transducers in the preparation of biosensors. Various chemical molecules are used to conjugate biological compounds in the design of biofunctional surfaces. Among them, chemobrionics (CBCs) are good alternatives as an immobilization matrix to improve performance parameters such as linear range, limit of detection (LOD), stability, reproducibility, etc. Here, CBCs are used to develop enzymatic biosensor, and glucose oxidase (GOx) is selected as a model enzyme. Firstly, CBC is functionalized using (3-Aminopropyl)triethoxysilane (APTES) to obtain free amine groups for immobilization of GOx via covalent bonds. The synthesized CBC-APTES is used as a matrix for the conjugation of GOx using glutaraldehyde as a crosslinker. The proposed CBC-APTES/GOx is formed on the glassy carbon electrode, and its performance is checked for glucose detection. The linear range is 0.025–1.25 mM with an LOD of 0.024 mM glucose. After finding no effect of some potential interference on the current response, CBC-APTES/GOx is successfully applied to analyze glucose in artificial samples such as serum, urine, sweat, and saliva. The objective here is to outline the integration of CBC in biosensor preparation, which has been successfully carried out for the first time in the literature.

A Mn-porphyrinic metal–organic framework immobilizing glucose oxidase for combined photodynamic/chemodynamic/starvation therapy

A Mn-porphyrinic metal–organic framework immobilizing glucose oxidase for combined photodynamic/chemodynamic/starvation therapy

A Glucose Oxidase-Curcumin Composite Nanoreactor for Multimodal Synergistic Cancer Therapy.

Glucose oxidase (GOx) selectively oxidizes β-d-glucose into gluconic acid and hydrogen peroxide; thus, it has emerged as a promising anticancer agent by tumor starvation and oxidative therapy. Here, we developed a nanoscale platform or "nanoreactor" that incorporates GOx and the bioactive natural product curcumin (CUR) to achieve a multimodal anticancer nanocomposite. The composite nanoreactor was formed by loading CUR in biodegradable polymeric nanoparticles (NPs) of poly(ethylene glycol)-b-poly(ε-caprolactone) (PEG-PCL). Prime-coating of the NPs with an iron(III)-tannic acid complex enabled facile immobilization of GOx on the NP surface. The NPs were monodisperse with a hydrodynamic diameter of 122 nm and a partially negative surface charge. The NPs were also associated with an excellent CUR loading efficiency and sustained release up to 96 h, which was accelerated by surface-immobilized GOx and followed supercase II transport. Viability assays were conducted on two model cancer cell lines, MCF-7 and MDA-MB-231 cells, as well as human dermal fibroblasts as a representative normal cell line. The assays revealed significantly improved potency of CUR in the composite nanoreactor, with up to 6000- and 1280-fold increase in MCF-7 and MDA-MB-231 cells, respectively, and lower toxicity toward normal cells. The NPs were also able to promote intracellular reactive oxygen species (ROS) generation and dissipation of the mitochondrial membrane potential, providing important clues on the mechanism of action of the nanoreactor. Further investigation of caspase-3 activity revealed that the nanoreactor had no effect or inhibited caspase-3 levels, signifying a caspase-independent mechanism of inducing apoptosis. Our findings present a promising nanocarrier platform that combines therapeutic agents with distinct mechanisms of action acting in synergy for more effective cancer therapy.

Tilted fiber Bragg grating sensor based on surface plasmon resonance and electrospinning for glucose detection

A tilted fiber Bragg grating (TFBG) sensor based on surface plasmon resonance (SPR) with electrospinning nanofibers was proposed for the precise detection of glucose. In this design, a 50 nm thick gold film was deposited on the surface of a 10° TFBG. Polyacrylonitrile (PAN) nanofibers were wrapped around the sensor through electrospinning, subsequently serving as a scaffold for the immobilization of glucose oxidase (GOD). Within the dynamic range of 100–500 mg/dL glucose concentrations, the sensor exhibited good response and worked effectively. The experimental results show that the sensitivity and limit of detection (LOD) of the sensor are 12.42 pm/(mg/dL) and 100 mg/dL. Furthermore, the structural characterization was carried out using atomic force microscopy (AFM), scanning electron microscopy (SEM), and Fourier transform infrared spectrometer (FTIR). This sensor exhibits outstanding stability and high sensitivity, providing a new method for diabetes detection.

Smartphone-enabled flow injection amperometric glucose monitoring based on a screen-printed carbon electrode modified with PEDOT@PB and a GOx@PPtNPs@MWCNTs nanocomposite

This study presents a modified screen-printed carbon electrode (SPCE) to determine glucose in a custom-built flow injection system. The biosensor was constructed by immobilizing glucose oxidase on porous platinum nanoparticles decorated on multi-walled carbon nanotubes (GOx@PPtNPs@MWCTNs). The fabrication of the biosensor was completed by coating the GOx@PPtNPs@MWCTNs nanocomposite on an SPCE modified with a nanocomposite of poly(3,4-ethylenedioxythiophene) and Prussian blue (GOx@PPtNPs@MWCTNs/PEDOT@PB/SPCE). The fabricated electrode accurately measured hydrogen peroxide (H2O2), the byproduct of the GOx-catalyzed oxidation of glucose, and was then applied as a glucose biosensor. The glucose response was amperometrically determined from the PB-mediated reduction of H2O2 at an applied potential of −0.10 V in a flow injection system. Under optimal conditions, the developed biosensor produced a linear range from 2.50 μM to 1.250 mM, a limit of detection of 2.50 μM, operational stability over 500 sample injections, and good selectivity. The proposed biosensor determined glucose in human plasma samples, achieving recoveries and results that agreed with the hexokinase-spectrophotometric method (P > 0.05). Combining the proposed biosensor with the custom-built sample feed, a portable potentiostat and a smartphone, enabled on-site glucose monitoring.

Self-propelled enzyme-controlled IR-mesoporous silica Janus nanomotor for smart delivery

Here, we report the preparation of a novel Janus nanoparticle with opposite Ir and mesoporous silica nanoparticles through a partial surface masking with toposelective modification method. This nanomaterial was employed to construct an enzyme-powered nanomachine with self-propulsion properties for on-command delivery. The cargo-loaded nanoparticle was provided with a pH-sensitive gate and unit control at the mesoporous face by first attaching boronic acid residues and further immobilization of glucose oxidase through reversible boronic acid esters with the carbohydrate residues of the glycoenzyme. Addition of glucose leads to the enzymatic production of H2O2 and gluconic acid, being the first compound catalytically decomposed at the Ir nanoparticle face producing O2 and causing the nanomachine propulsion. Gluconic acid leads to a pH reduction at the nanomachine microenvironment causing the disruption of the gating mechanism with the subsequent cargo release. This work demonstrates that enzyme-mediated self-propulsion improved release efficiency being this nanomotor successfully employed for the smart release of Doxorubicin in HeLa cancer cells.

Advancements in Glucose Monitoring: A Thin Film ZnO-Nanoflakes Based Highly Sensitive Wearable Biosensor for Noninvasive Sweat-Based Point-of-Care Monitoring for Diabetes

In the contemporary global landscape, diabetes is a concerning presence that contributes to noteworthy mortality rates. The etiology of this condition involves a subtle interaction between insulin deficiency and elevated blood sugar levels. Against this backdrop, the necessity for prompt, accurate, and continuous noninvasive monitoring of glucose concentrations becomes notably evident. The electrochemical flexible glucose biosensor was fabricated by immobilizing glucose oxidase (GOx) on ZnO nanoflakes, possessing a 20nm thickness, and synthesized on an Au-coated stretchable PET film. The sensitivity of the flexible biosensor was determined by 29.97μA/decade/cm², and the minimum detection limit was 1μM. The coefficient of determination was 97.76%, which is significantly linear.

Magnetic Nanoparticle Support with an Ultra-Thin Chitosan Layer Preserves the Catalytic Activity of the Immobilized Glucose Oxidase.

Here, we developed magnetically recoverable biocatalysts based on magnetite nanoparticles coated with an ultra-thin layer (about 0.9 nm) of chitosan (CS) ionically cross-linked by sodium tripolyphosphate (TPP). Excessive CS amounts were removed by multiple washings combined with magnetic separation. Glucose oxidase (GOx) was attached to the magnetic support via the interaction with N-hydroxysuccinimide (NHS) in the presence of carbodiimide (EDC) leading to a covalent amide bond. These steps result in the formation of the biocatalyst for D-glucose oxidation to D-gluconic acid to be used in the preparation of pharmaceuticals due to the benign character of the biocatalyst components. To choose the catalyst with the best catalytic performance, the amounts of CS, TPP, NHS, EDC, and GOx were varied. The optimal biocatalyst allowed for 100% relative catalytic activity. The immobilization of GOx and the magnetic character of the support prevents GOx and biocatalyst loss and allows for repeated use.

Intensification of a biocatalytic oxidation under fine bubble aeration in a rotating bed reactor

The utilization of oxidases suffers from insufficient oxygen availability due to its low solubility in gas-liquid multiphase systems. To overcome this limitation, oxygen is continuously supplied to the reaction medium during the course of the reaction, however, it causes significant gas consumption. In this study, a novel experimental set-up was established by installing a sintered frit porous sparger in the SpinChem® rotating bed reactor. Covalently immobilized glucose oxidase on epoxy-functionalized carriers was used in the rotating bed reactor with an activity yield of the immobilization of over 98%. For fine bubble aeration, the volumetric mass transfer coefficient kLa (67.3 h−1) tripled compared to macrobubble aeration (22.5 h−1), while the volume-specific aeration rate remained constant. A maximum reaction yield of 96.5% was reached. The reaction rate was improved by a factor of 12.9 for the fine bubble aeration compared to macrobubble aeration under identical conditions. With fine bubble aeration, the oxygen consumption was reduced by 87.5% over macrobubble aeration, establishing the same enzyme-specific reaction rates at a comparable kLa. Validation was carried out in repetitive batches, achieving the same glucose oxidase specific activity. This study demonstrates the advantages of fine bubble aeration and presents promising results for addressing the sustainable biotransformation processes.

A smartphone-enabled colorimetric platform based on enzyme cascade amplification strategy for detection of Staphylococcus aureus in milk

Staphylococcus aureus is a pathogenic bacterium contaminating milk and dairy foods causing food poisoning and foodborne pathogens. In this work, a smartphone-enabled enzyme cascade-triggered colorimetric platform was constructed using a cascade bio-nanozyme formed by immobilized glucose oxidase (GOx) on Fe3O4@Ag for rapid detection of S. aureus. Benefiting from reasonable experimental design, a bio-nanozyme cascade-triggered reaction was achieved through H2O2 produced by GOx oxidation of glucose, followed by in situ catalysis of 3,3',5,5'-tetramethylbenzidine (TMB) by the inherent peroxidase-like activity of Fe3O4@Ag to produce color signals. Staphylococcus aureus detection could be performed through naked-eye observation and smartphone measurement, and the developed assay can achieve quantitative and qualitative detection of S. aureus. The on-site nanoplatform had satisfactory specificity and sensitivity with a low detection limit of 6.9 cfu·mL-1 in 50 min. Moreover, the nanoplatform has good practicality in the detection of S. aureus in milk samples. Therefore, the assay has potential application prospects in food safety inspection.