Highly Porous 3D Gold Enhances Sensitivity of Amperometric Biosensors Based on Oxidases and CuCe Nanoparticles

Metallic nanoparticles potentially have wide practical applications in various fields of science and industry. In biosensorics, they usually act as catalysts or nanozymes (NZs) and as mediators of electron transfer. We describe here the development of amperometric biosensors (ABSs) based on purified oxidases, synthesized nanoparticles of CuCe (nCuCe), and micro/nanoporous gold (pAu), which were electro-deposited on a graphite electrode (GE). As an effective peroxidase (PO)-like NZ, nCuCe was used here as a hydrogen-peroxide-sensing platform in ABSs that were based on glucose oxidase, alcohol oxidase, methylamine oxidase, and L-arginine oxidase. At the same time, nCuCe is an electroactive mediator and has been used in laccase-based ABSs. As a result, the ABSs we constructed and characterized were based on glucose, methanol, methyl amine, L-arginine, and catechol, respectively. The developed nCuCe-based ABSs exhibited improved analytical characteristics in comparison with the corresponding PO-based ABSs. Additionally, the presence of pAu, with its extremely advanced chemo-sensing surface layer, was shown to significantly increase the sensitivities of all constructed ABSs. As an example, the bioelectrodes containing laccase/GE, laccase/nCuCe/GE, and laccase/nCuCe/pAu/GE exhibited sensitivities to catechol at 2300, 5055, and 9280 A·M−1·m−2, respectively. We demonstrate here that pAu is an effective carrier of electroactive nanomaterials coupled with oxidases, which may be promising in biosensors

“Green” Prussian Blue Analogues as Peroxidase Mimetics for Amperometric Sensing and Biosensing

Prussian blue analogs (PBAs) are well-known artificial enzymes with peroxidase (PO)-like activity. PBAs have a high potential for applications in scientific investigations, industry, ecology and medicine. Being stable and both catalytically and electrochemically active, PBAs are promising in the construction of biosensors and biofuel cells. The “green” synthesis of PO-like PBAs using oxido-reductase flavocytochrome b2 is described in this study. When immobilized on graphite electrodes (GEs), the obtained green-synthesized PBAs or hexacyanoferrates (gHCFs) of transition and noble metals produced amperometric signals in response to H2O2. HCFs of copper, iron, palladium and other metals were synthesized and characterized by structure, size, catalytic properties and electro-mediator activities. The gCuHCF, as the most effective PO mimetic with a flower-like micro/nano superstructure, was used as an H2O2-sensitive platform for the development of a glucose oxidase (GO)-based biosensor. The GO/gCuHCF/GE biosensor exhibited high sensitivity (710 A M−1m−2), a broad linear range and good selectivity when tested on real samples of fruit juices. We propose that the gCuHCF and other gHCFs synthesized via enzymes may be used as artificial POs in amperometric oxidase-based (bio)sensors

Formaldehyde-sensitive conductometric sensors based on commercial and recombinant formaldehyde dehydrogenase

Novel formaldehyde-sensitive conductometric biosensors have been developed that are based on commercial bacterial formaldehyde dehydrogenase (FDH) from Pseudomonas putida and recombinant formaldehyde dehydrogenase (rFDH) from the yeast Hansenula polymorpha as the bio-recognition elements. The bio-recognition membranes have mono-layer architecture and consist of enzyme cross-linked with albumin and of the cofactors NAD (for FDH-based sensor) or NAD and glutathione (for rFDH-based sensor). This architecture of the biosensor allows the determination of formaldehyde without adding NAD and glutathione to the analyzed sample at every analysis and conducting measurements on the same transducer without cofactors regeneration since the bio-membrane contains it at high concentration (100 mM for NAD and 20 mM for glutathione). The response is linear in the range from 10 to 200 mM of formaldehyde concentration depending on the enzyme used. The dependence of the biosensor output signals on pH and buffer concentration as well as operational/storage stability and selectivity/specificity of the developed conductometric biosensors have been investigated. The relative standard deviation of the intra-sensor response did not exceed 4% and 10% for rFDH- and FDH-based sensors, respectively. The relative standard deviation of the inter-sensor response constituted 20% for both dehydrogenases used. The biosensors have been validated for formaldehyde detection in some real samples of pharmaceutical (Formidron), disinfectant (Descoton forte) and an industrial product (Formalin). A good correlation does exist between the concentration values measured by the conductometric biosensor developed in this work, an enzymatic method, amperometric biosensors developed earlier, and standard analytical methods of formaldehyde determination

Highly Sensitive Amperometric Biosensors Based on Oxidases and CuCe Nanoparticles Coupled with Porous Gold

Metallic nanoparticles are usually applied in biosensors as catalysts and/or mediators of electron transfer. We describe the development of amperometric biosensors (ABSs) based on oxidases and nanoparticles of CuCe (nCuCe). nCuCe, being an electro-active mediator and active peroxidase (PO) mimetic, was used as an H2O2-sensing platform in oxidase-based ABSs. ABSs for glucose, primary alcohols, methyl amine, catechol, and L-arginine, which are based on corresponding oxidases and nCuCe, were developed. These ABSs exhibited improved analytical characteristics in comparison with the appropriate bi-enzyme ABSs containing natural PO. Including electrodeposited porous gold in the chemo-sensing layer was shown to increase significantly the sensitivities of all constructed ABSs