Analytical Performance of Clay Paste Electrode and Graphene Paste Electrode-Comparative Study

The analytical performance of the clay paste electrode and graphene paste electrode was compared using square wave voltammetry (SWV) and cyclic voltammetry (CV). The comparison was made on the basis of a paracetamol (PA) determination on both working electrodes. The influence of pH and SWV parameters was investigated. The linear concentration ranges were found to be 6.0 × 10⁻⁷–3.0 × 10⁻⁵ and 2.0 × 10⁻⁶–8.0 × 10⁻⁵ mol L⁻¹ for clay paste electrode (ClPE) and graphene paste electrode (GrPE), respectively. The detection and quantification limits were calculated as 1.4 × 10⁻⁷ and 4.7 ×10⁻⁷ mol L⁻¹ for ClPE and 3.7 × 10⁻⁷ and 1.2 × 10⁻⁶ mol L⁻¹ for GrPE, respectively. Developed methods were successfully applied to pharmaceutical formulations analyses. Scanning electron microscopy and energy-dispersive X-ray spectroscopy were used to characterize ClPE and GrPE surfaces. Clay composition was examined with wavelength dispersive X-ray (WDXRF).     

Electrochemiluminescence Biosensor Based on PEDOT‐PSS‐ Graphene Functionalized ITO Electrode

A novel and sensitive electrochemiluminescence (ECL) method for ethanol biosensor was developed by co‐immobilizing the enzyme and ECL reagent Ru(bpy)₃²⁺ on the poly‐(3,4‐ethylene dioxythiophene) and polystyrene sulfonate functionalized graphene (PEDOT‐PSS‐G) nanocomposite film. Positively charged Ru(bpy)₃²⁺ could be immobilized effectively on the electrode surface with the negatively charged PSS and graphene, which provided a stable ECL platform for further modification with the enzyme. Moreover, the introduction of PEDOT and graphene can be acted as a conducting pathway to accelerate the electron transfer due to the high conductivity. Such biosensor combined enzymatic selectivity with the amplification of PEDOT‐PSS‐G performed well with a wide linear range, high sensitivity and good stability. The sensing platform was successfully applied to determine the amounts of alcohol in real samples.     

Bio-electrocatalysis of NADH and ethanol based on graphene sheets modified electrodes

Characterization and application of graphene sheets modified glassy carbon electrodes (graphene/GC) have been presented for the electrochemical bio-sensing. A probe molecule, potassium ferricyanide is employed to study the electrochemical response at the graphene/GC electrode, which shows better electron transfer than graphite modified (graphite/GC) and bare glassy carbon (GC) electrodes. Based on the highly enhanced electrochemical activity of NADH, alcohol dehydrogenase (ADH) is immobilized on the graphene modified electrode and displays a more desirable analytical performance in the detection of ethanol, compared with graphite/GC or GC based bio-electrodes. It also exhibits good performance of ethanol detection in the real samples. From the results of electrochemical investigation, graphene sheets with a favorable electrochemical activity could be an advanced carbon electrode materials for the design of electrochemical sensors and biosensors.     

Impedance Based Nanotoxicity Assessment of Graphene Nanomaterials at the Cellular and Tissue Level

The interest in graphene for biomedical applications has grown substantially in the past few years creating a need for biocompatibility testing. Biomedical engineering applications using graphene such as biosensing devices, microbial detection, disease diagnosis, and drug delivery systems are progressing rapidly, perhaps overlooking any possible hazards as graphene nanomaterials may interact with biological materials differently than other graphitic materials such as carbon nanotubes and fullerenes. As a potential application for graphene is drug delivery, the toxicity of graphene was tested against an in vitro model of the blood brain barrier (BBB) by measuring trans-endothelial-electrical resistance (TEER). A new approach in terms of electrical impedance sensing was also utilized to kinetically analyze the cytotoxicity of graphene nanomaterials towards the BBB model's individual components, rat astrocytes (CRL-2006) and mouse endothelial cells (CRL-2583), in real time by measuring the impedimetric response. Graphene showed little or no toxicity toward both individual cell types as the resistance measurements were similar to those of the control and further, graphene did not interrupt the integrity of the BBB model as a whole showing the biocompatibility of graphene and the broad potential of using these new nanomaterials for biomedical applications.     

Functionalized graphene oxide for the fabrication of paraoxon biosensors

There is an increasing need to develop biosensors for the detection of harmful pesticide residues in food and water. Here, we report on a versatile strategy to synthesize functionalized graphene oxide nanomaterials with abundant affinity groups that can capture histidine (His)-tagged acetylcholinesterase (AChE) for the fabrication of paraoxon biosensors. Initially, exfoliated graphene oxide (GO) was functionalized by a diazonium reaction to introduce abundant carboxyl groups. Then, N_α,N_α-bis(carboxymethyl)-l-lysine hydrate (NTA-NH₂) and Ni²⁺ were anchored onto the GO based materials step by step. AChE was immobilized on the functionalized graphene oxide (FGO) through the specific binding between Ni-NTA and His-tag. A low anodic oxidation potential was observed due to an enhanced electrocatalytic activity and a large surface area brought about by the use of FGO. Furthermore, a sensitivity of 2.23 μA mM⁻¹ to the acetylthiocholine chloride (ATChCl) substrate was found for our composite covered electrodes. The electrodes also showed a wide linear response range from 10 μM to 1 mM (R² = 0.996), with an estimated detection limit of 3 μM based on an S/N = 3. The stable chelation between Ni-NTA and His-tagged AChE endowed our electrodes with great short-term and long-term stability. In addition, a linear correlation was found between paraoxon concentration and the inhibition response of the electrodes to paraoxon, with a detection limit of 6.5 × 10⁻¹⁰ M. This versatile strategy provides a platform to fabricate graphene oxide based nanomaterials for biosensor applications.     

One‐Pot Fabrication of RGO–Ag3VO4 Nanocomposites by in situ Photoreduction using Different Sacrificial Agents: High Selectivity Toward Catechol Synthesis and Photodegradation Ability

Weak photon absorption and fast carrier kinetics in graphene restrict its applications in photosensitive reactions. Such restrictions/limitations can be overcome by covalent coupling of another photosensitive nanostructure to graphene, forming graphene‐semiconductor nanocomposites. Herein, we report one‐pot synthesis of RGO–Ag₃VO₄ nanocomposites using various sacrificial agents like ethanol, methanol, propanol and ethylene glycol (EG) under visible light illumination. The Raman spectral analysis and ¹³C MAS NMR suggest ethanol to be the best sacrificial agent among those studied. Thermal analysis studies, further, confirm the stability of the synthesized nanocomposite with ethanol as sacrificial agent. In view of this, the activity toward dye degradation was focused over the composites prepared via ethanol as sacrificial agent. It was observed and proved that cationic dyes could be degraded quantitatively and swiftly compared to anionic dyes (37.79%) in 1.5 h. This suggests that the surface of the nanocomposites is anionic as partial reduction takes place during synthesis process. In case of mixed dye degradation process, it was noticed that the presence of cationic dye doubles the degradation of anionic dye. The activity of these synthesized nanocomposites is more than five‐fold toward the phototransformation of phenol and photodegradation of textile dyes under visible light illumination.     

Regeneration of a Conjugated sp2 Graphene System through Selective Defunctionalization of Epoxides by Using a Proven Synthetic Chemistry Mechanism

Graphene is a promising material capable of driving technological advancement. It is, however, a challenge to obtain pristine graphene in large quantities given the limitation of current synthetic methods. Among the numerous methods available, the chemical approach provides an optimistic outlook and has garnered much interest within the graphene community as a potential alternative. One of the most crucial steps of the chemical approach is the chemical reduction of graphene oxide as this dictates the final quality of the graphene sheets. In recent years, much of the focus has shifted to the usage of established reducing agents or oxygen removal reagents, frequently applied in organic chemistry, onto a graphene oxide platform. Herein, the selective removal of epoxide groups and subsequent regeneration of disrupted conjugated sp² system is highlighted, based on the synergistic effect of indium and indium(I) chloride. The morphological, structural, and electrical properties of the resulting graphene were fully characterized with X‐ray photoelectron, Fourier transform IR, solid‐state ¹³C NMR, and Raman spectroscopy; thermogravimetric analysis; scanning electron microscopy; and conductivity measurements. The as‐prepared graphene showed a tenfold increase in conductivity against conventional graphene treated with hydrazine reducing agent and demonstrated a high dispersion stability in ethanol. Moreover, the selective defunctionalization of the epoxide groups provides opportunities for potential tailoring of graphene properties for prospective applications.     

Facile hydrothermal growth graphene/ZnO nanocomposite for development of enhanced biosensor

Graphene/zinc oxide nanocomposite was synthesised via a facile, green and efficient approach consisted of novel liquid phase exfoliation and solvothermal growth for sensing application. Highly pristine graphene was synthesised through mild sonication treatment of graphite in a mixture of ethanol and water at an optimum ratio. The X-ray diffractometry (XRD) affirmed the hydrothermal growth of pure zinc oxide nanoparticles from zinc nitrate hexahydrate precursor. The as-prepared graphene/zinc oxide (G/ZnO) nanocomposite was characterised comprehensively to evaluate its morphology, crystallinity, composition and purity. All results clearly indicate that zinc oxide particles were homogenously distributed on graphene sheets, without any severe aggregation. The electrochemical performance of graphene/zinc oxide nanocomposite-modified screen-printed carbon electrode (SPCE) was evaluated using cyclic voltammetry (CV) and amperometry analysis. The resulting electrode exhibited excellent electrocatalytic activity towards the reduction of hydrogen peroxide (H₂O₂) in a linear range of 1–15 mM with a correlation coefficient of 0.9977. The sensitivity of the graphene/zinc oxide nanocomposite-modified hydrogen peroxide sensor was 3.2580 μAmM⁻¹ with a limit of detection of 7.4357 μM. An electrochemical DNA sensor platform was then fabricated for the detection of Avian Influenza H5 gene based on graphene/zinc oxide nanocomposite. The results obtained from amperometry study indicate that the graphene/zinc oxide nanocomposite-enhanced electrochemical DNA biosensor is significantly more sensitive (P < 0.05) and efficient than the conventional agarose gel electrophoresis.     

Multi‐walled Carbon Nanotubes Non‐covalently Functionalized with Polyarginine: A New Alternative for the Construction of Reagentless NAD+/Dehydrogenase‐based Ethanol Biosensor

This work reports for the first time the development of a reagentless enzymatic amperometric biosensor for ethanol based on the use of a glassy carbon electrode (GCE) modified with multi‐walled carbon nanotubes (MWCNTs) non‐covalently functionalized with polyarginine (Polyarg) as platform for the robust immobilization of alcohol dehydrogenase (ADH) and NAD⁺. The new strategy allows to obtain an integrated GCE/MWCNTs‐Polyarg/NAD⁺‐ADH ethanol biosensor with important advantages compared to the existing ethanol biosensors: avoids the external addition of the cofactor for each measurement, ensures a fast and sensitive quantification of ethanol due to the intimate interaction of the components, and allows the detection at considerably lower potentials due to the catalytic activity of the carbon nanostructures. These unique properties have made possible a very efficient ethanol quantification with a sensitivity of (1487±6) μA M^?1, detection limit of 0.65 μM, response time of 8 s, and reproducibility of 5.5 % with a very successful application for the quantification of ethanol in different commercial beverages.     

A novel non-invasive electrochemical biosensing device for in situ determination of the alcohol content in blood by monitoring ethanol in sweat

A non-invasive, passive and simple to use skin surface based sensing device for determining the blood's ethanol content (BAC) by monitoring transdermal alcohol concentration (TAC) is designed and developed. The proposed prototype is based on bienzyme amperometric composite biosensors that are sensitive to the variation of ethanol concentration. The prototype correlates, through previous calibration set-up, the amperometric signal generated from ethanol in sweat with its content in blood in a short period of time. The characteristics of this sensor device permit determination of the ethanol concentration in isolated and in continuous form, giving information of the BAC of a subject either in a given moment or its evolution during long periods of time (8 h). Moreover, as the measurements are performed in a biological fluid, the evaluated individual is not able to alter the result of the analysis. The maximum limit of ethanol in blood allowed by legislation is included within the linear range of the device (0.0005–0.6 g L⁻¹). Moreover, the device shows higher sensitivity than the breathalyzers marketed at the moment, allowing the monitoring of the ethanol content in blood to be obtained just 5 min after ingestion of the alcoholic drink. The comparison of the obtained results using the proposed device in the analysis of 40 volunteers with those provided by the gas chromatographic reference method for determination of BAC pointed out that there were no significant differences between both methods.     

Electrochemical Determination of Dopamine Using a Poly(3,4-Ethylenedioxythiophene)-Reduced Graphene Oxide-Modified Glassy Carbon Electrode

Poly(3,4-ethylenedioxythiophene) was deposited on a reduced graphene oxide-decorated glassy carbon electrode through an electrochemical polymerization. The resulting glassy carbon electrode-reduced graphene oxide-poly(3,4-ethylenedioxythiophene) electrode was applied as an electrochemical biosensor for the determination of dopamine in the presence of ascorbic acid and uric acid. The material deposited on glassy carbon electrode was investigated in terms of morphology and structural analysis. The comparison of electrochemical behavior of the glassy carbon electrode-reduced graphene oxide-poly(3,4-ethylenedioxythiophene) electrode with the glassy carbon electrode-graphene oxide, glassy carbon electrode-reduced graphene oxide, and glassy carbon electrode-poly(3,4-ethylenedioxythiophene) electrodes exhibited high electrocatalytic activity for dopamine detection. Electrochemical kinetic parameters of glassy carbon electrode-reduced graphene oxide-poly(3,4-ethylenedioxythiophene), including the charge transfer coefficient α (0.49) and electron transfer rate constant k_s (1.04), were determined and discussed. The glassy carbon electrode-reduced graphene oxide-poly(3,4-ethylenedioxythiophene) electrode was studied for the determination of dopamine by differential pulse voltammetry and exhibited a linear range from 19.6 to 122.8?µM with a sensitivity of 3.27?µA?µM^?1?cm^?2 and a detection limit of 1.92?µM. The developed biosensor exhibited good selectivity toward dopamine with high reproducibility and stability.     

Tannic Acid Modified Electrochemical Biosensor for Glucose Sensing Based on Direct Electrochemistry

A novel glucose biosensor was constructed through the immobilization of glucose oxidase (GOx) on gold nanoparticles (Au NPs) deposited, and chemically reduced graphene oxide (rGO) nanocomposite. In the synthesis, tannic acid (TA) was used for the reduction of both graphene oxide, and Au³⁺ to rGO, and Au NPs, respectively. Also, by harnessing the π‐π interaction between graphene oxide and TA, and protein‐TA interaction, a novel nanocomposite for the fabrication of a third generation biosensor was successfully constructed. Upon the oxidation of TA to quinone, which is easily reducible at the negative potential range, enhanced electron transfer was obtained. The cyclic voltammetry (CV) results demonstrated a pair of well‐defined and quasi‐reversible redox peaks of active site molecule of GOx. The biosensor exhibited a linear response to glucose concentrations varying from 2 to 10 mM with a sensitivity of 18.73 mA mM⁻¹ cm⁻². The fabricated biosensor was used for the determination of glucose in beverages.     

A Highly Sensitive and Selective Label-free Electrochemical Biosensor with a Wide Range of Applications for Bisphenol A Detection

A label-free DNA-based electrochemical biosensor owning high sensitivity and selectivity has been established for detecting bisphenol A in a wide range of applications. Coupling the high electrochemical performance of graphene oxide-thionine-Au nanomaterial with the specific binding capacity of the aptamers to BPA, the monitoring of trace amount of BPA was realized, the detection limit was 3.3 pg ⋅ mL⁻¹ with strong anti-interference. Besides, using molecular docking, it was found that BPA binds to the bases DC-49, DC-51, DG-52, DG-53 and DA-63 on the aptamer via hydrogen bonding and π-π stacking interactions. Finally, the biosensor had been successfully applied in different real samples.     

Thin Films and Composites Based on Graphene for Electrochemical Detection of Biologically‐relevant Molecules

Graphene is one of the most studied materials ever, owing to its exceptional electronic, mechanical and thermal properties, which allow for many different types of application. In this review, we shall concentrate on the use of graphene and derivatives for electrochemical sensors and biosensors, where emphasis is placed on the importance of surface functionalization as this permits synergistic combinations with other nanomaterials and biomolecules. In addition to describing recent advances in graphene‐based electroanalytical applications, we discuss a few examples of their use in detecting small biomolecules and in immunosensing for a few diseases using films and composites. Also discussed are the possible methods for mass production of graphene, which is key to low‐cost biosensors for implantable devices and portable systems in point‐of‐care diagnosis.     

Gold nanostar-enhanced surface plasmon resonance biosensor based on carboxyl-functionalized graphene oxide

A new high-sensitivity surface plasmon resonance (SPR) biosensor based on biofunctional gold nanostars (AuNSs) and carboxyl-functionalized graphene oxide (cGO) sheets was described. Compared with spherical gold nanoparticles (AuNPs), the anisotropic structure of AuNSs, which concentrates the electric charge density on its sharp tips, could enhance the local electromagnetic field and the electronic coupling effect significantly. cGO was obtained by a diazonium reaction of graphene oxide (GO) with 4-aminobenzoic acid. Compared with GO, cGO could immobilize more antibodies due to the abundant carboxylic groups on its surface. Testing results show that there are fairly large improvements in the analytical performance of the SPR biosensor using cGO/AuNSs-antigen conjugate, and the detection limit of the proposed biosensor is 0.0375 μg mL⁻¹, which is 32 times lower than that of graphene oxide-based biosensor.     

Effect of Enzyme and Cofactor Immobilization on the Response of Ethanol Oxidation in Zirconium Phosphate Modified Biosensors

Two different self‐contained ethanol amperometric biosensors incorporating layered [Ru(phend)₂bpy]²⁺‐intercalated zirconium phosphate (ZrP) as the mediator as well as yeast‐alcohol dehydrogenase (y‐ADH) and its cofactor nicotinamide adenine dinucleotide (NAD⁺) were constructed to improve upon a design previously reported where only this mediator was immobilized in the surface of a modified electrode. In the first biosensor, a [Ru(phend)₂bpy]²⁺‐intercalated ZrP modified carbon paste electrode (CPE) was improved by immobilizing in its surface both y‐ADH and NAD⁺ using quaternized Nafion membrane. In the second biosensor, a glassy carbon electrode was modified with [Ru(phend)₂bpy]²⁺‐intercalated ZrP, y‐ADH, and NAD⁺ using Nafion as the holding matrix. Calibration plots for ethanol sensing were constructed in the presence and absence of ZrP. In the absence of ZrP in the surface of the modified glassy carbon electrode, leaching of ADH was observed as detected by UV‐vis spectrophotometry. Ethanol sensing was also tested in the presence and absence of ascorbate to measure the selectivity of the sensor for ethanol. These two ethanol biosensors were compared to a previously reported one where the y‐ADH and the NAD⁺ were in solution, not immobilized.     

A dense graphene monolith with poloxamer prefunctionalization enabling aqueous redispersion to obtain solubilized graphene sheets

The realization of good aqueous dispersibility of commercial graphene products composed of exfoliated graphene sheets is of significance for downstream applications. However, the tap density of commercial graphene powder is quite low (0.03–0.1 kg/m³), meaning that 1 kg graphene powder occupies about 10–30 m³ in volume during transportation. And, the available content of commercial graphene dispersion/slurry in aqueous medium cannot exceed 5 wt%, although the density is high (≈ 1050 kg/m³). In this work, a graphene monolith was prepared by oven-drying of graphene sheets prefunctionalized with poloxamer surfactants. Our graphene monoliths not only have a high density (1500 kg/m³) and high graphene content (≈ 10 wt%), but also a full capability to be completely redispersed (≈ 100%) in water by bath sonication to obtain solubilized graphene sheets, whose lateral size and thickness are unchanged compared to as-exfoliated ones. Moreover, a simple empirical method was proposed to predict the redispersion capability of graphene monoliths using different poloxamers by contact angle measurements. Our results provide a universal approach to make exfoliated graphene-based products with better downstream availability and lower transportation cost.     

Bioelectrocatalytic Activity of Immobilized Alcohol Oxidase on 4‐(pyrrole‐1‐yl) Benzoic Acid Modified Carbon Nanotubes

A novel bioelectrocatalytic system was prepared by immobilizing alcohol oxidase (AOx) onto multiwalled carbon nanotubes (MWCNT) modified with 4‐(pyrrole‐1‐yl) benzoic acid (PyBA). Functional carboxylic groups from PyBA create covalent amide linkages with amine groups from the enzyme molecule and provide an anchor for the effective immobilization of AOx improving the stability of the whole system. The immobilized enzyme displayed a pair of reversible redox peaks of flavin adenine dinucleotide (FAD) cofactor with the formal potential E^0’=?0.451 V. The response showed a surface‐controlled electrode process with the heterogeneous electron transfer rate constant k_s=2.7 s^?1. Under aerobic conditions AOx(FADH₂) can be oxidized to AOx(FAD) by oxygen, which then reacts with ethanol decreasing the cathodic response, which could be used for ethanol detection with a high sensitivity 13.1 μA mM^?1 cm^?2. The lack of bioactivity towards ethanol in anaerobic conditions suggests the presence of two types of AOx molecules in the system: active with oxygen maintaining the direct electron transfer feature and not active without a redox mediator, due to the deeply embedded FAD cofactor. The polarization curve showed that the electrooxidation current of ethanol appears at ?410 mV and reaches 2.0 µA cm^?1 at ?300 mV. In this case, the bioactivity of AOx to ethanol can be observed offering promising solution for the development of mediatorless systems for application to biosensors and biofuel cells.     

Electrocatalytic Oxidation of Glucose by the Glucose Oxidase Immobilized in Graphene‐Au‐Nafion Biocomposite

Graphene was successfully prepared and well separated to individual sheets by introducing  SO₃⁻. XRD and TEM were employed to characterize the graphene. UV‐visible absorption spectra indicated that glucose oxidase (GOx) could keep bioactivity well in the graphene‐Au biocomposite. To construct a novel glucose biosensor, graphene, Au and GOx were co‐immobilized in Nafion to further modify a glassy carbon electrode (GCE). Electrochemical measurements were carried out to investigate the catalytic performance of the proposed biosensor. Cyclic voltammograms (CV) showed the biosensor had a typical catalytic oxidation response to glucose. At the applied potential +0.4 V, the biosensor responded rapidly upon the addition of glucose and reached the steady state current in 5 s, with the present of hydroquinone. The linear range is from 15 μM to 5.8 mM, with a detection limit 5 μM (based on the S/N=3). The Michaelis‐Menten constant was calculated to be 4.4 mM according to Lineweaver–Burk equation. In addition, the biosensor exhibits good reproducibility and long‐term stability. Such impressive properties could be ascribed to the synergistic effect of graphene‐Au integration and good biocompatibility of the hybrid material.