Nanostructured Iron Oxide Platform for Impedimetric Cholesterol Detection

A nanostructured iron oxide (NanoFe₃O₄, particle size ca. 25 nm and roughness ca. 21 nm) film deposited onto a hydrolyzed indium‐tin‐oxide (ITO) coated glass plate has been used to immobilize cholesterol oxidase (ChOx) to fabricate an impedimetric cholesterol sensor. Electrochemical studies reveal that surface charged Fe₃O₄ nanoparticles provide better conformation for ChOx loading resulting in enhanced electron transfer between ChOx and the electrode. Impedimetric response studies of the ChOx/NanoFe₃O₄/ITO bioelectrode exhibit improved linearity (2.5–400 mg/dL), low detection limit (0.25 mg/dL), fast response time (25 s), high sensitivity (86 Ω/mg dL^?1/cm^?2) and a low value of the Michaelis‐Menten constant (K_m, 0.8 mg/dL) with a regression coefficient of 0.997.     

Polypyrrole/multiwalled carbon nanotubes‐based biosensor for cholesterol estimation

The nanocomposite electrode comprising of polypyrrole (PPY) and carboxy functionalized multiwalled carbon nanotubes (MWCNT) has been electrochemically fabricated onto indium–tin–oxide (ITO) electrode using p‐toluene sulfonic acid (PTS). Cholesterol oxidase (ChOx) and cholesterol esterase (ChEt) have been immobilized onto this PPY– MWCNT/ITO nanocomposite electrode using N‐ethyl‐N‐(3‐dimethylaminopropyl) carbodiimide and N‐hydroxy succinimide chemistry for estimation of esterified cholesterol. The ChEt–ChOx/PPY–MWCNT/PTS/ITO bioelectrode has been characterized using Fourier transform infrared spectroscopy, electrochemical techniques, and scanning electron microscope. This ChEt–ChOx/PPY–MWCNT/PTS/ITO nanobioelectrode has a response time of about 9 s, linearity of 4 × 10^?4 to 6.5 × 10^?3 M/l of cholesterol oleate concentration, K_m of 0.02 mM, and thermal stability of upto 45°C. This electrode exhibits improved biosensing characteristics compared with other total cholesterol electrodes reported in literature till date and can be used to estimate cholesterol in blood serum samples. Copyright © 2011 John Wiley & Sons, Ltd.     

Electrochemical Study of Chitosan‐SiO2‐MWNT Composite Electrodes for the Fabrication of Cholesterol Biosensors

We report a novel composite electrode made of chitosan‐SiO₂‐multiwall carbon nanotube (CHIT‐SiO₂‐MWNT) composite coated on the indium‐tin oxide (ITO) glass substrate. Cholesterol oxidase (ChOx) was covalently immobilized on the CHIT‐SiO₂‐MWNT/ITO electrode that resulted in a ChOx/CHIT‐SiO₂‐MWNT/ITO cholesterolactive bioelectrode. The CHIT‐SiO₂‐MWNT/ITO and ChOx/CHIT‐SiO₂‐MWNT/ITO electrodes were characterized with Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). The influence of various parameters was investigated, including the applied potential, pH of the medium, and the concentration of the enzyme on the performance of the biosensor. The cholesterol bioelectrode exhibited a sensitivity of 3.4 nA/ mgdL^?1 with a response time of five seconds. The biosensor using ChOx/CHIT‐SiO₂‐MWNT/ITO as the working electrode retained its original response after being stored for six months. The biosensor using ChOx/CHIT‐SiO₂‐MWNT/ITO as the working electrode showed a linear current response to the cholesterol concentration in the range of 50–650 mg/dL.     

Sol–gel derived nanoporous cerium oxide film for application to cholesterol biosensor

Cholesterol oxidase (ChOx) has been immobilized onto sol–gel derived nano-structured cerium oxide (NS-CeO₂) film deposited on indium-tin-oxide (ITO) coated glass substrate. Phase identification of sol–gel NS-CeO₂ film carried out using X-ray diffraction (XRD) yields reflection peak at 29.4° corresponding to (1 1 1) plane with oriented crystallite (34 nm) along c-axis normal to the substrate. Electrochemical studies reveal that NS-CeO₂ provides electroactive surface for the loading of ChOx and enhances electron transfer rate in the ChOx/NS-CeO₂/ITO bioelectrode. The low value of Michaelis–Menten constant (K_m) obtained as 2.08 mM indicates enhanced ChOx affinity to cholesterol. The observed results show application of sol–gel derived NS-CeO₂ for biosensing without any functionalization.     

Zinc oxide nanoparticles-chitosan composite film for cholesterol biosensor

Zinc oxide nanoparticles (NanoZnO) uniformly dispersed in chitosan (CHIT) have been used to fabricate a hybrid nanocomposite film onto indium-tin-oxide (ITO) glass plate. Cholesterol oxidase (ChOx) has been immobilized onto this NanoZnO-CHIT composite film using physiosorption technique. Both NanoZnO-CHIT/ITO electrode and ChOx/NanoZnO-CHIT/ITO bioelectrode have been characterized using Fourier transform-infrared (FTIR), X-ray diffraction (XRD), cyclic voltammetry (CV), scanning electron microscopy (SEM) and electrochemical impedance spectroscopy (EIS) techniques, respectively. The ChOx/NanoZnO-CHIT/ITO bioelectrode exhibits linearity from 5 to 300 mg dl⁻¹ of cholesterol with detection limit as 5 mg dl⁻¹, sensitivity as 1.41 × 10⁻⁴ A mg dl⁻¹ and the value of Michaelis-Menten constant (K_m) as 8.63 mg dl⁻¹. This cholesterol biosensor can be used to estimate cholesterol in serum samples.     

Polyaniline/Single‐Walled Carbon Nanotubes Composite Based Triglyceride Biosensor

Nanocomposite film comprising of polyaniline (PANI) and single walled carbon nanotubes (SWCNT) has been fabricated onto indium‐tin‐oxide (ITO) coated glass plate using electrophoretic technique. Co‐immobilization of glycerol dehydrogenase (GDH) and lipase (LIP) has been done via N‐ethyl‐N′‐(3‐dimethylaminopropyl) carbodiimide and N‐hydroxysuccinimide chemistry to explore its application for triglyceride (tributyrin) sensing. Response studies have been done using linear sweep voltammetry revealing that LIP‐GDH/PANI‐SWCNT‐TB/ITO bioelectrode can detect tributyrin in the range of 50 to 400 mg dL^?1 with low Michaelis–Menten constant of 1.138 mM, improved response time of 12 s, high sensitivity as 4.28×10^?4 mA mg^?1 dL and storage stability of about 13 weeks.     

Potential of Gelatin‐Zinc Oxide Nanocomposite as Ascorbic Acid Sensor

We report on the studies relating to fabrication of gelatin B (GB) and zinc oxide (ZnO) based nanocomposite (GB‐ZnO) film deposited on indium‐tin‐oxide (ITO) glass plate, and used for the immobilization of ascorbate oxidase (AsOx) which was further used for ascorbic acid (AA) detection. The structural and morphological studies of GB‐ZnO, and AsOx/GB‐ZnO/ITO bioelectrodes were carried out using XRD, SEM and FTIR techniques. This bioelectrode showed a broad range of linearity (5–500 mg/dL), low detection limit (1 mg/dL), higher sensitivity (0.106 µA mg/dLcm^?2) and low value of the apparent Michaelis? Menten constant (K_m^app=0.35 mg/dL) for AA. Efforts are being made to utilize this electrode for sensing AA in real samples in a bid to develop a strip based sensor.     

Nonenzymatic H2O2 Electrochemical Sensor Based on SnO2‐NPs Coated Polyethylenimine Functionalized Graphene

This paper demonstrated using polyethylenimine (PEI)‐functionalized graphene (Gr) incorporating tin oxide (SnO₂) hybrid nanocomposite as a platform for nonenzymatic H₂O₂ electrochemical sensor. The results of UV‐vis spectroscopy and X‐ray diffraction (XRD) confirmed the simultaneous formation of tin oxide (SnO₂) nanocomposite and reduction of graphene oxide (GO). Transmission electron microscopy (TEM) images showed a uniform distribution of nanometer‐sized tin oxide nanoparticles on the grapheme sheets, which could be achieved using stannous chloride (SnCl₂) complex instead of tin oxide as precursor. The electrochemical measurements, including cyclic voltammetry (CV) and amperometric performance (I‐t), showed that the PEI‐functionalized Gr supported SnO₂ (SnO₂‐PEI‐Gr) exhibited an excellent electrocatalytic activity toward the H₂O₂. The corresponding calibration curve of the current response showed a linear detection range of 9×10⁻⁶∼1.64×10⁻³ mol L⁻¹, while the limit of detection was estimated to be 1×10⁻⁶ mol L⁻¹. Electrochemical studies indicated that SnO₂ and functionalized Gr worked synergistically for the detection of H₂O₂.     

Direct Electrochemistry of Cholesterol Oxidase Immobilized on a Conducting Polymer: Application for a Cholesterol Biosensor

Direct electrochemistry of cholesterol oxidase (ChOx) immobilized on the conductive poly‐3′,4′‐diamine‐2,2′,5′,2″‐terthiophene (PDATT) was achieved and used to create a cholesterol biosensor. A well‐defined redox peak was observed, corresponding to the direct electron transfer of the FAD/FADH₂ of ChOx, and the rate constant (k_s) was determined to be 0.75 s^?1. Glutathione (GSH) covalently bonded with PDATT was used as a matrix for conjugating AuNPs, ChOx, and MP, simultaneously. MP co‐immobilized with ChOx on the AuNPs‐GSH/PDATT exhibited an excellent amperometric response to cholesterol. The dynamic range was from 10 to 130 μM with a detection limit of 0.3±0.04 μM.     

Nanocrystalline Tin‐Oxide Modified Electrodes and their Electrochemical Characterization

Nanocrystalline tin‐oxide particles were prepared as electrodes on the bases of ITO glass and AT‐cut quartz crystals (sputtered gold), respectively, and characterized for their electrochemical behavior. Experiments suggested that the SnO₂ particles could induce an energy barrier to the redox reactions taking place on the electrode surface. When the amount of SnO₂ exceeded ca. 10^?7 mol cm^?2, electrochemical activity demonstrated by the solution redox couples was entirely suppressed. Nevertheless, electrochemical impedance spectroscopic (EIS) measurements suggested that mutual communication between redox couples would still take place on the surface of SnO₂. For instance, although the CV curves of Fe(CN)₆^3‐/4‐ were completely blocked, the exchange current of Fe(CN)₆^3‐/4‐ could still flow through the tin‐oxide modified electrode, increasing with its concentration up to 40 mM. The propagation of electrons in the SnO₂ film was likely via a hopping mechanism. Electrochemical quartz microbalance (EQCM) measurements, in addition, suggested that a charge‐compensating cation (K⁺ or H⁺) uptake reaction may be induced as electrons were pumped to the Sn0₂ electrode, while, if electrons were removed, that could cause water desorption. Analysis based on the Frumkin adsorption isotherm showed the driving force behind the adsorption of water on SnO₂ is about ?2 kcal/mol. Nonetheless, the adsorbed water might face a competitive repulsion from acetonitrile when acetonitrile was used as the electrolyte medium.     

Horse radish peroxidase immobilized polyaniline for hydrogen peroxide sensor

Horse radish peroxidase (HRP) has been electrochemically entrapped into perchlorate (ClO) doped polyaniline (PANI) film deposited onto indium‐tin‐oxide (ITO) coated glass plate. This HRP‐PANI‐ClO/ITO bioelectrode characterized using Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), UV‐Visible spectroscopy, cyclic voltammetry (CV), and electrochemical impedance spectroscopy (EIS) techniques has been utilized for estimation of hydrogen peroxide (H₂O₂). This H₂O₂ sensor exhibits response time of 5 s, linearity from 3 to 136 mM, sensitivity as 0.5638 µA mM^?1 cm^?2 with linear regression of 0.985. The value of the Michaelis–Menten constant (K_m) has been obtained as 1.984 mM. Copyright © 2009 John Wiley & Sons, Ltd.     

Biosensor for total cholesterol estimation using N-(2-aminoethyl)-3-aminopropyltrimethoxysilane self-assembled monolayer

Cholesterol oxidase (ChOx), cholesterol esterase (ChEt), and horseradish peroxidase (HRP) have been co-immobilized covalently on a self-assembled monolayer (SAM) of N-(2-aminoethyl)-3-aminopropyltrimethoxysilane (AEAPTS) deposited on an indium–tin–oxide (ITO) glass surface. These enzyme-modified (ChOx-ChEt-HRP/AEAPTS/ITO) biosensing electrodes have been used to estimate cholesteryl oleate from 10 to 500 mg dL⁻¹. The sensitivity, K _m value, and shelf-life of these ChEt-ChOx-HRP/AEAPTS/ITO biosensing electrodes have been found to be 124 nA mg⁻¹ dL, 95.098 mg dL⁻¹ (1.46 mmol L⁻¹), and ten weeks, respectively. The ChEt-ChOx-HRP/AEAPTS/ITO bio-electrodes have been used to estimate total cholesterol in serum samples. Figure Covalent immobilization of enzymes onto AEAPTS/ITO surface using EDC/NHS chemistry Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Electrophoretically deposited polyaniline nanotubes based film for cholesterol detection

Polyaniline nanotube (PANI-NT) based films have been fabricated onto indium-tin-oxide (ITO) coated glass plates via electrophoretic technique. These PANI-NT/ITO electrodes have been utilized for covalent immobilization of cholesterol oxidase (ChOx) using glutaraldehyde (Glu) as cross-linker. Structural, morphological and electrochemical characterization of PANI-NT/ITO electrode and ChOx/Glu/PANI-NT/ITO bioelectrode have been done using FT-IR spectroscopy, SEM, electrochemical impedance spectroscopy and cyclic voltammetry techniques. Response studies of the ChOx/Glu/PANI-NT/ITO bioelectrode have been carried out using both linear sweep voltammetry and UV-Visible spectrophotometry. The results of the biosensing studies reveal that this bioelectrode can be used to detect cholesterol in wide detection range of 25-500 mg/dL with high sensitivity of 3.36 mA mg(-1) dL and fast response time of 30 s at pH 7.4. This bioelectrode exhibits very low value of Michaelis-Menten constant of 1.18 mM indicating enhanced interactions between cholesterol and ChOx immobilized onto this nanostructured PANI matrix.     

Peptide Nucleic Acid Immobilized Biocompatible Silane Nanocomposite Platform for Mycobacterium tuberculosis Detection

We have prepared nanocomposite films comprising of 3‐glycidoxypropyltrimethoxysilane (GOPS) and iron‐oxide (Fe₃O₄) onto indium‐tin‐oxide (ITO) glass plate for covalent immobilization of 21‐mer peptide nucleic acid (PNA). These films have been characterized using contact angle, atomic force microscopy (AFM), electrochemical techniques. The electrochemical response of the GOPS/ITO and Fe₃O₄‐GOPS/ITO electrodes has been investigated by hybridization with complementary, non‐complementary and one‐base mismatch using methylene blue as electrochemical indicator. The PNA/Fe₃O₄‐GOPS/ITO bioelectrode exhibits improved specificity and detection limit (0.1 fM) as compared to that of the PNA‐GOPS/ITO bioelectrode (0.1 pM). This PNA/Fe₃O₄‐GOPS/ITO electrode can be utilized for detection of hybridization with the complementary sequence in sonicated M. tuberculosis genomic DNA within 90 s of hybridization time.     

Carbon Nanotubes‐Based Amperometric Cholesterol Biosensor Fabricated Through Layer‐by‐Layer Technique

A carbon nanotubes‐based amperometric cholesterol biosensor has been fabricated through layer‐by‐layer (LBL) deposition of a cationic polyelectrolyte (PDDA, poly(diallyldimethylammonium chloride)) and cholesterol oxidase (ChOx) on multi‐walled carbon nanotubes (MWNTs)‐modified gold electrode, followed by electrochemical generation of a nonconducting poly(o‐phenylenediamine) (PPD) film as the protective coating. Electrochemical impedance measurements have shown that PDDA/ChOx multilayer film could be formed uniformly on MWNTs‐modified gold electrode. Due to the strong electrocatalytic properties of MWNTs toward H₂O₂ and the low permeability of PPD film for electroacitve species, such as ascorbic acid, uric acid and acetaminophen, the biosensor has shown high sensitivity and good anti‐interferent ability in the detection of cholesterol. The effect of the pH value of the detection solution on the response of the biosensor was also investigated. A linear range up to 6.0 mM has been observed for the biosensor with a detection limit of 0.2 mM. The apparent Michaelis‐Menten constant and the maximum response current density were calculated to be 7.17 mM and 7.32 μA cm^?2, respectively.     

Rapid and Sensitive Electrochemical Detection of Carbaryl Based on Enzyme Inhibition and Thiocholine Oxidation Mediated by a Ruthenium(III) Complex

In the electrochemical detection method for pesticides that measures their inhibitory effects on acetylcholinesterases (AChEs), the direct electrooxidation of the enzyme product (thiocholine, SCh) is slow at conventional electrodes. To overcome this limitation, an electron mediator is required to lower the applied potential and facilitate the transfer of electrons between the enzyme product and electrode. In this study, [Ru(NH₃)₅py]³⁺ is introduced as an electron mediator in inhibition‐based pesticide detection. To obtain a better signal‐to‐background ratio, [Ru(NH₃)₅py]³⁺, which undergoes a fast outer‐sphere reaction, is combined with low‐electrocatalytic indium‐tin‐oxide (ITO) electrodes at which many interfering species undergo slow redox reactions. AChE is immobilized onto an avidin‐modified ITO electrode via the direct adsorption of avidin onto ITO followed by the biospecific binding of biotinylated AChE to the avidin. SCh is generated from acetylthiocholine by AChE. Subsequently, SCh converts [Ru(NH₃)₅py]³⁺ to [Ru(NH₃)₅py]²⁺, which is then oxidized at the ITO electrode. This procedure allows the sensitive detection of carbaryl at a low applied potential of 0.15 V vs Ag/AgCl. The calculated detection limit for carbaryl is approximately 0.3 pM. This simple and sensitive pesticide sensor is thus very promising and should be extendable to the onsite environmental monitoring of other pesticides.     

A Single Drop Fabrication of the Cholesterol Biosensor Based on Synthesized NiFe2O4NPs Dispersed on PDDA‐CNTs

A cholesterol biosensor based on cholesterol oxidase‐poly(diallyldimethylammonium chloride)‐carbon nanotubes‐nickel ferrite nanoparticles (ChOx‐PDDA‐CNTs‐NiFe₂O₄NPs) solution is easily fabricated by using a single dropping step on a glassy carbon electrode (GCE) surface. This technique is an alternative way to reduce complexity, cost and time to produce the biosensor. The uniformly dispersed materials on the electrode surface enhance the catalytic reaction of cholesterol oxidase and electron transfer from the oxidation of hydrogen peroxide in the system. The nickel ferrite nanoparticles were synthesized by co‐precipitation and calcination at various temperatures. These nanoparticles were then characterized using field emission scanning electron microscopy (FE‐SEM), energy‐dispersive X‐ray spectroscopy (EDX), Fourier transform infrared spectroscopy (FTIR), cyclic voltammetry (CV) and X‐ray diffraction (XRD). The synthesized material calcined at 700 °C was well defined and presented the octahedral metal stretching with cubic NiFe₂O₄NPs phase. In cyclic voltammetric study, the ChOx‐PDDA‐CNTs‐NiFe₂O₄NPs/GCE showed 0.43 s⁻¹ charge transfer rate constant (K _s), 7.79×10⁻⁶ cm² s⁻¹ diffusion coefficient value (D ), 0.13 mm² electroactive surface area (A _e) and 3.58×10⁻⁸ mol cm⁻² surface concentration ( ). This modified electrode exhibits stability in term of percent relative standard deviation (%RSD=0.62 %, n=10), reproducibility (%RSD=0.81, n=10), high sensitivity (25.76 nA per mg L⁻¹ cm⁻²), linearity from 1 to 5,000 mg L⁻¹ (R²=0.998) with a low detection limit (0.50 mg L⁻¹). Its Michaelis‐Menten constant (K _m) was 0.14 mM with 0.92 μA maximum current (I _max) and demonstrated good selectivity without the effects of electroactive species such as ascorbic acid, glucose and uric acid. The cholesterol biosensor was successfully applied to determine cholesterol levels in human blood samples, showing promise due to its simplicity and availability.     

Functionalized Gold Nanoparticles – Octadecylamine Hybrid Langmuir‐Blodgett Film for Enzyme Sensor

Mediator free enzyme sensor has been fabricated by covalently immobilizing cholesterol oxidase (ChOx) onto 11‐mercaptoundecanoic acid functionalized gold nanoparticles (MUDA‐AuNPs) – octadecylamine (ODA) hybrid Langmuir–Blodgett film. The cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) studies reveal that MUDA‐AuNP/ODA LB film has good affinity for ChOx and provides favorable microenvironment for direct electron transfer between enzyme and electrode. Interference free estimation of cholesterol has been realized at 0.3 V with linear range from 25 to 500 mg/dL, detection limit of 23.38 mg/dL, sensitivity of 1.085 μA mM^?1 and response time of 20 s at pH 7.0.     

Transition‐Metal‐Catalyzed Oxidation of Metallic Sn in NiO/SnO2 Nanocomposite

It is well accepted that metallic tin as a discharge (reduction) product of SnO_x cannot be electrochemically oxidized below 3.00 V versus Li⁺/Li⁰ due to the high stability of Li₂O, though a similar oxidation can usually occur for a transition metal formed from the corresponding oxide. In this work, nanosized Ni₂SnO₄ and NiO/SnO₂ nanocomposite were synthesized by coprecipitation reactions and subsequent heat treatment. Owing to the catalytic effect of nanosized metallic nickel, metallic tin can be electrochemically oxidized to SnO₂ below 3.00 V. As a result, the reversible lithium‐storage capacities of the nanocomposite reach 970 mAh g^?1 or above, much higher than the theoretical capacity (ca. 750 mAh g^?1) of SnO₂, NiO, or their composites. These findings extend the well‐known electrochemical conversion reaction to non‐transition‐metal compounds and may have important applications, for example, in constructing high‐capacity electrode materials and efficient catalysts.     

Hemin Functionalized Multiwalled Carbon Nanotubes as a Matrix for Sensitive Electrogenerated Chemiluminescence Cholesterol Biosensor

Based on hemin‐MWCNTs nanocomposite and hemin‐catalyzed luminol‐H₂O₂ reaction, a sensitive electrogenerated chemiluminescence (ECL) cholesterol biosensor was proposed in this paper. Firstly, hemin‐MWCNTs was prepared via π–π stacking and modified on the surface of GCE. Subsequently, cholesterol oxidase (ChOx) was adsorbed on the modified electrode to achieve a cholesterol biosensor. Hemin‐MWCNTs nanocomposite provided the electrode with a large surface area to load ChOx, and endowed the nanostructured interface on the electrode surface to enhance the performance of biosensor. The biosensor responded to cholesterol in the linear range from 0.3 µM to 1.2 mM with a detection limit of 0.1 µM (S/N=3).