Search Results (246)

A new approach in amperometric enzyme electrodes production based on all-electrochemically assisted procedures will be described. Enzyme (glucose oxidase) immobilization was performed by in situ co-crosslinking of enzyme molecules through electrophoretic protein deposition, assuring enzyme immobilization exclusively onto the transducer surface (Pt electrode). Analogously, the poor selectivity of the transducer was dramatically improved by the electrosynthesis of non-conducting polymers with built-in permselectivity, permitting the formation of a thin permselective film onto the transducer surface, able to reject common interferents usually found in real samples. Since both approaches required a proper and distinct electrochemical perturbation (a pulsed current sequence for electrophoretic protein deposition and cyclic voltammetry for the electrosynthesis of non-conducting polymers), an appropriate coupling of the two all-electrochemical approaches was assured by a thorough study of the likely combinations of the electrosynthesis of permselective polymers with enzyme immobilization by electrophoretic protein deposition and by the use of several electrosynthesized polymers. For each investigated combination and for each polymer, the analytical performances and the rejection capabilities of the resulting biosensor were acquired so to gain information about their sensing abilities eventually in real sample analysis. This study shows that the proper coupling of the two all-electrochemical approaches and the appropriate choice of the electrosynthesized, permselective polymer permits the easy fabrication of novel glucose oxidase biosensors with good analytical performance and low bias in glucose measurement from typical interferent in serum. This novel approach, resembling classical electroplating procedures, is expected to allow all the advantages expected from such procedures like an easy preparation biosensor, a bi-dimensional control of enzyme immobilization and thickness, interferent- and fouling-free transduction of the electrodic sensor and, last but not the least, possibility of miniaturization of the biosensing device. Full article

We develop a modular, wireless, solar- and battery-powered system for detecting chlorpyrifos (Lorsban^TM 2.5% DP) in water using electrochemical impedance spectroscopy (EIS). The system integrates a Raspberry Pi Zero 2W for data processing, Python-based software (version 3.12.2), and a solar charge manager to power all components via a lithium-ion battery and solar panel. A commercial EmStat Pico Module and an amperometric biosensor with acetylcholinesterase (AChE) detect chlorpyrifos. Nine water samples with varying concentrations were tested using a 20 Hz–200 kHz frequency sweep and 15 mV excitation. Bode plots and statistical analyses confirmed statistically significant impedance variation as a function of chlorpyrifos concentration, validating the system as a portable, sensitive, and effective tool for environmental monitoring. Full article

The non-monotonic behavior of amperometric enzyme-based biosensors under uncompetitive and noncompetitive (mixed) substrate inhibition is investigated computationally using a two-compartment model consisting of an enzyme layer and an outer diffusion layer. The model is based on a system of reaction–diffusion equations that includes a nonlinear term associated with non-Michaelis–Menten kinetics of the enzymatic reaction and accounts for the partitioning between layers. In addition to the known effect of substrate inhibition, where the maximum biosensor current differs from the steady-state output, it has been determined that external diffusion limitations can also cause the appearance of a local minimum in the current. At substrate concentrations greater than both the Michaelis–Menten constant and the uncompetitive substrate inhibition constant, and in the presence of external diffusion limitation, the transient response of the biosensor, after immersion in the substrate solution, may follow a five-phase pattern depending on the model parameter values: it starts from zero, reaches a global or local maximum, decreases to a local minimum, increases again, and finally decreases to a steady intermediate value. The biosensor performance is analyzed numerically using the finite difference method. Full article

Amperometric biosensors (ABSs) and enzymatic biofuel cells (BFCs) share several fundamental principles in their functionality, despite serving different primary purposes. Both devices rely on biorecognition, redox reactions, electron transfer (ET), and advanced electrode materials, including innovative nanomaterials (NMs). ABSs and BFCs, utilizing microbial oxidoreductases in combination with electroactive NMs, are both efficient and cost-effective. In the current study, several laboratory prototypes of BFCs have been developed with bioanodes based on yeast flavocytochrome b₂ (Fcb₂) and alcohol oxidase (AO), and a cathode based on fungal laccase. For the first time, BFCs have been developed featuring anodes based on Fcb₂ co-immobilized with redox NMs on a glassy carbon electrode (GCE), and cathode-utilizing laccase combined with gold–cerium–platinum nanoparticles (nAuCePt). The most effective lactate BFC, which contains gold–hexacyanoferrate (AuHCF), exhibited a specific power density of 1.8 µW/cm². A series of BFCs were developed with an AO-containing anode and a laccase/nAuCePt/GCE cathode. The optimal configuration featured a bioanode architecture of AO/nCoPtCu/GCE, achieving a specific power density of 3.2 µW/cm². The constructed BFCs were tested using lactate-containing food product samples as fuels. Full article

Artificial enzymes or nanozymes (NZs) are gaining significant attention in biotechnology due to their stability and cost-effectiveness. NZs can offer several advantages over natural enzymes, such as enhanced stability under harsh conditions, longer shelf life, and reduced production costs. The booming interest in NZs is likely to continue as their potential applications expand. In our previous studies, we reported the “green” synthesis of copper hexacyanoferrate (gCuHCF) using the oxidoreductase flavocytochrome b₂ (Fcb₂). Organic–inorganic micro-nanoparticles were characterized in detail, including their structure, composition, catalytic activity, and electron-mediator properties. An SEM analysis revealed that gCuHCF possesses a flower-like structure well-suited for concentrating and stabilizing Fcb₂. As an effective peroxidase (PO) mimic, gCuHCF has been successfully employed for H₂O₂ detection in amperometric sensors and in several oxidase-based biosensors. In the current study, we demonstrated the uniqueness of gCuHCF that lies in its multifunctionality, serving as a PO mimic, a chemosensor for ammonium ions, a biosensor for L-lactate, and exhibiting perovskite-like properties. This exceptional ability of gCuHCF to enhance fluorescence under blue light irradiation is being reported for the first time. Using gCuHCF as a PO-like NZ, novel oxidase-based sensors were developed, including an optical biosensor for L-arginine analysis and electrochemical biosensors for methanol and glycerol determination. Thus, gCuHCF, synthesized via Fcb₂, presents a promising platform for the development of amperometric and optical biosensors, bioreactors, biofuel cells, solar cells, and other advanced devices. The innovative approach of utilizing biocatalysts for nanoparticle synthesis highlights a groundbreaking direction in materials science and biotechnology. Full article

Electrochemical enzyme biosensors are extensively utilized in clinical analysis and environmental monitoring, yet achieving effective enzyme immobilization while maintaining high activity remains a challenge. In this work, we developed a flow-through enzyme biosensor system using a 3D-printed flow-through electrochemical cell fabricated from commercially available poly (lactic acid). After modification with thiacalixarene-functionalized oligo (lactic acids) (OLAs), the material enabled efficient immobilization of uricase on the inner surface of a replaceable reactor of the cell. Swelling and hydrolytic stability of OLAs in cone, partial cone, and 1,3-alternate conformations were studied, with 1,3-alernate conformation demonstrating superior stability and enzyme immobilization performance. The use of OLAs enhanced immobilization efficiency by over 30% and protected the reactor from swelling, hydrolytic degradation, and enzyme loss. The biosensor was validated for amperometric uric acid determination, with a screen-printed carbon electrode modified with carbon black and Prussian Blue. This modification reduced the cathodic potential for uric acid detection to –0.05 V. The biosensor exhibited a linear detection range of 10 nM to 30 μM with a detection limit of 7 nM, and it performed effectively in artificial urine and synthetic blood plasma. The novel cell design, featuring easy assembly and low-cost replaceable parts, makes this biosensor a promising candidate for routine clinical analysis and other practical applications. Full article

Enzyme-based portable amperometric biosensors are precise and low-cost medical devices used for rapid cancer biomarker screening. Sarcosine (Sar) is an ideal biomarker for prostate cancer (PCa). Because human serum and urine contain complex interfering substances that can directly oxidize at the electrode surface, rapid Sar screening biosensors are relatively challenging and have rarely been reported. Therefore, highly sensitive and selective amperometric biosensors that enable real-time measurements within <1.0 min are needed. To achieve this, a chitosan–polyaniline polymer nanocomposite (CS–PANI NC), a carrier for dispersing mesoporous carbon (MC), was synthesized and modified on a screen-printed carbon electrode (SPCE) to detect hydrogen peroxide (H₂O₂). The sarcosine oxidase (SOx) enzyme-immobilized CS–PANI–MC-2 ternary NCs were referred to as supramolecular architectures (SMAs). The excellent electron transfer ability of the SMA-modified SPCE (SMA/SPCE) sensor enabled highly sensitive H₂O₂ detection for immediate trace Sar biomarker detection. Therefore, the system included an SMA/SPCE coupled to a portable potentiostat linked to a smartphone for data acquisition. The high catalytic activity, porous architecture, and sufficient biocompatibility of CS–PANI–MC ternary NCs enabled bioactivity retention and immobilized SOx stability. The fabricated biosensor exhibited a detection limit of 0.077 μM and sensitivity of 8.09 μA mM⁻¹ cm⁻² toward Sar, demonstrating great potential for use in rapid PCa screening. Full article

This work presents the development of an amperometric biosensor for detecting aspartate aminotransferase (AST) activity in biological fluids using a platinum disk electrode as the working transducer. Optimal concentrations of substrates (aspartate, α-ketoglutarate) and the coenzyme (pyridoxal phosphate) were determined to ensure efficient biosensor operation. A semi-permeable poly-m-phenylenediamine membrane was applied to enhance selectivity against electroactive interferents. The biosensor demonstrated good stability (storage, continuous operation, and production reproducibility) and analytical performance (sensitivity 8.56 nA/min for 50 U/L AST, LOD 1 U/L, linear range 1–110 U/L). Testing with real samples showed a high correlation (R = 0.989) with spectrophotometric analysis, supporting its potential for further applications. Full article

We report the development of amperometric biosensors (ABSs) employing flavocytochrome b₂ (Fcb₂) coupled with nanoparticles (NPs) of noble metals on graphite electrode (GE) surfaces. Each NPs/GE configuration was evaluated for its ability to decompose hydrogen peroxide (H₂O₂), mimicking peroxidase (PO) activity. The most effective nanoPO (nPO) was selected for developing ABSs targeting L-lactate. Consequently, several Fcb₂/nPO-based ABSs with enhanced sensitivity to L-lactate were developed, demonstrating mediated ET between Fcb₂ and the GE surface. The positive effect of noble metal NPs on Fcb₂-based sensor sensitivity may be explained by the synergy between their dual roles as both PO mimetics and electron transfer mediators. Furthermore, our findings provide preliminary data that may prompt a re-evaluation of the mechanism of L-lactate oxidation in Fcb₂-mediated catalysis. Previously, it was believed that L-lactate oxidation via Fcb₂ catalysis did not produce H₂O₂, unlike catalysis via L-lactate oxidase. Our initial research revealed that the inclusion of nPO in Fcb₂-based ABSs significantly increased their sensitivity. Employing other PO mimetics in ABSs for L-lactate yielded similar results, reinforcing our hypothesis that trace amounts of H₂O₂ may be generated as a transient intermediate in this reaction. The presence of nPO enhances the L-lactate oxidation rate through H₂O₂ utilization, leading to signal amplification and heightened bioelectrode sensitivity. The proposed ABSs have been successfully tested on blood serum and fermented food samples, showing their promise for L-lactate monitoring in medicine and the food industry. Full article

With improvements in medical environments and the widespread use of smartphones, interest in wearable biosensors for continuous body monitoring is growing. We developed a wearable multiplexed bio-sensing system that non-invasively monitors body fluids and integrates with a smartphone application. The system includes sensors, readout circuits, and a microcontroller unit (MCU) for signal processing and wireless communication. Potentiometric and amperometric measurement methods were used, with calibration capabilities added to ensure accurate readings of analyte concentrations and temperature. Laser-induced graphene (LIG)-based sensors for glucose, lactate, Na⁺, K⁺, and temperature were developed for fast, cost-effective production. The LIG electrode’s 3D porous structure provided an active surface area 16 times larger than its apparent area, resulting in enhanced sensor performance. The glucose and lactate sensors exhibited high sensitivity (168.15 and 872.08 μAmM⁻¹cm⁻², respectively) and low detection limits (0.191 and 0.167 μM, respectively). The Na⁺ and K⁺ sensors demonstrated sensitivities of 65.26 and 62.19 mVdec⁻¹, respectively, in a concentration range of 0.01–100 mM. Temperature sensors showed an average rate of resistance change per °C of 0.25%/°C, within a temperature range of 20–40 °C, providing accurate body temperature monitoring. Full article

The development of biosensors for determining the most diverse biomolecules is a constant focus of many research groups. There is a latent need to propose sensors that combine portability, simple measurements, and good analytical performance. Here, we propose an electrochemical immunosensor that is fully portable and energy-independent for diagnosing antibodies against SARS-CoV-2 (the virus that causes COVID-19). Initially, disposable screen-printed carbon electrodes (SPEs) were covered by gold microblobs (AuMBs), which were synthesized amperometrically from Au³⁺ ions. Then, the SPE-AuMBs were coated with cysteamine, which allowed the N-hydroxysuccinimide-activated SARS-CoV-2 antigen (spike protein) to be immobilized. The antigen-activated electrode was used to detect COVID-19 antibodies from current measurements obtained by differential pulse voltammetry. The AuMBs synthesis time was optimized, and the presence of gold structures improved the electrochemical responses of the SPE. It was possible to quantitatively determine antibodies in the concentration range of 0.25 to 10 µg mL⁻¹. This range includes concentrations found in biological fluids from patients at any stage of the disease. An analysis took approximately the same time as traditional rapid nasal tests (20 min) and costed less, considering all the steps necessary to prepare a disposable antigen-functionalized SPE. Full article

This paper describes the design and development of a novel electrochemical biosensor for measuring glycerol in wine. Our initial detailed studies were aimed at deducing the optimum conditions for biosensor operation by conducting hydrodynamic voltammetric and amperometric studies. The resulting voltammograms revealed a maximum electrocatalytic current at 0.0 V vs. Ag/AgCl, which we used for all further studies. We also examined the effect of pH (8–10) on the amperometric responses of different glycerol concentrations over a range of 0.04 to 0.20 mM. Based on our findings, we propose that pH 9 would be suitable as the supporting electrolyte for further studies with the amperometric biosensor. The biosensor was constructed by immobilising 10 units of GLDH and 660 μg NAD⁺ onto the MB-SPCE surface using glutaraldehyde (GLA) as a cross-linking agent. Calibration studies were performed with glycerol over the 1.0–7.5 mM concentration range. Chronoamperometry was the electrochemical technique chosen for this purpose as it is convenient and can be performed with only 100 μL of sample directly deposited onto the biosensor’s surface. In the current study, we observed linear calibration plots with the above standard solutions using current measurements at a selection of sampling times along the chronoamperograms (30–340 s). We have evaluated the glycerol biosensor by carrying out an analysis of commercially available red wine. Overall, these findings will form a platform for the development of novel rapid technology for point-of-test evaluation of glycerol in the production and quality control of wine. Full article

Electrochemical biosensors are critical in advancing biomedical and pharmaceutical therapies because of their adaptability and cost-effectiveness. Voltammetric and amperometric sensors are of particular interest. These sensors typically consist of a specialized tip or biorecognition element and a transducer that converts biological data into readable signals. Efficient biosensor materials are essential for addressing health emergencies, with coordination polymers (CPs) and metal–organic frameworks (MOFs) showing promise. Functionalization strategies are necessary to enhance the usability of pristine MOFs, owing to issues such as low conductivity. The integration of conductive polymers with MOFs has resulted in the development of highly efficient biosensors. Both enzymatic and nonenzymatic biosensors are used for analyte detection; nonenzymatic approaches are gaining popularity owing to their durability and accuracy. MOFs and CPs have been applied in sensitive electrochemical biosensors to detect fatal brain tumors such as glioblastomas (GBM). These biosensors demonstrate enhanced selectivity and sensitivity, highlighting the potential of MOFs and CPs in advancing electrochemical biosensor technology for both in vivo and in vitro applications. Full article

L-Lactate is an important bioanalyte in the food industry, biotechnology, and human healthcare. In this work, we report the development of a new L-lactate electrochemical biosensor based on the use of multiwalled carbon nanotubes non-covalently functionalized with avidin (MWCNT-Av) deposited at glassy carbon electrodes (GCEs) as anchoring sites for the bioaffinity-based immobilization of a new recombinant biotinylated lactate oxidase (bLOx) produced in Escherichia coli through in vivo biotinylation. The specific binding of MWCNT-Av to bLOx was characterized by amperometry, surface plasmon resonance (SPR), and electrochemical impedance spectroscopy (EIS). The amperometric detection of L-lactate was performed at −0.100 V, with a linear range between 100 and 700 µM, a detection limit of 33 µM, and a quantification limit of 100 µM. The proposed biosensor (GCE/MWCNT-Av/bLOx) showed a reproducibility of 6.0% and it was successfully used for determining L-lactate in food and enriched serum samples. Full article

Galactose monitoring in individuals allows the prevention of harsh health conditions related to hereditary metabolic diseases like galactosemia. Current methods of galactose detection need development to obtain cheaper, more reliable, and more specific sensors. Enzyme-containing amperometric sensors based on galactose oxidase activity are a promising approach, which can be enhanced by means of their inclusion in a redox polymer coating. This strategy simultaneously allows the immobilization of the biocatalyst to the electroactive surface and hosts the electron shuttling units. An additional deposition of capping polymers prevents external interferences like ascorbic or uric acid as well as biofouling when measuring in physiological fuels. This work studies the protection effect of poly(2-methacryloyloxyethyl phosphorylcholine-co-glycidyl methacrylate (MPC) and polyvinylimidazole-polysulfostyrene (P(VI-SS)) when incorporated in the biosensor design for the detection of galactose in human plasma. Full article