Electrochemical sensing of concanavalin A using a non-ionic surfactant with a maltose moiety

To electrochemically detect concanavalin A (ConA), a new method was developed using mixed micelles between a non-ionic surfactant with a maltose moiety and electroactive daunomycin. The surfactants, in which the length of the alkyl chain was different, were n-decyl-β-d-maltoside, n-dodecyl-β-d-maltoside, and n-tetradecyl-β-d-maltoside. The measurement principle was due to the micelle breakdown caused by the binding between the ConA and maltose moieties. When ConA was combined with maltose moieties at a concentration of surfactant that was near the critical micelle concentration, the daunomycin that formed the micelles was moved to a solution from the micelles. As a result, the peak current of daunomycin increased as the concentration of ConA was increased. The mechanism was proposed using voltammetry, spectrometry, and gel filtration. The linear range using n-tetradecyl-β-d-maltoside was 2.0 × 10⁻⁹ to 8.0 × 10⁻⁸ M of ConA, and it was the most sensitive in the presence of the three surfactants. To examine whether selective binding took place, measurements with several proteins were carried out. The electrode responses of daunomycin were not influenced by the presence of 5.0 × 10⁻⁶ M protein. Furthermore, this method could be applied to the determination of ConA in a serum, and to the measurement of sugar chains that can be combined with ConA on the cell surface.     

Construction of a peptide with an electroactive daunomycin like a pendant arm to detect ovalbumin

In this study, a peptide-1 (RNRCKGTDVQAW) constructing lysozyme was conjugated with an electroactive daunomycin in order to voltammetrically detect ovalbumin (OVA). Hetero-bifunctional cross-linking agents with four kinds of ethylene chains in differing lengths were used to bind the peptide-1 and daunomycin. After a cross-linking agent had reacted with an amino group of daunomycin, the compound was introduced into the peptide to the cysteine residue in the peptide using a pendant arm. The OVA was sensed via a change in the electrode response of the daunomycin moiety, based on the binding between the peptide and the OVA. The adsorption of the peptide probe on the electrode increased with increases in the ethylene chain. The binding constants between the peptide probes and the OVA, however, did not depend on the length of the chain. This was because the ethylene chain influenced the binding. When the peptide and the daunomycin were bound using N-(6-maleimidocaproyloxy) sulfosuccinimide, the electrode response of the peptide probe was the most sensitive from among the four cross-linking agents. The calibration curve of the OVA using the peptide probe was linear and ranged from 1.5 × 10⁻¹¹ to 3.0 × 10⁻¹⁰ M. Furthermore, this method could be applied to the electrochemical sensing of the OVA in egg whites and in fetal bovine serum.     

A strategy for improving the sensitivity of molecularly imprinted electrochemical sensors based on catalytic copper deposition

A novel method to improve the sensitivity of molecularly imprinted polymer sensors was developed. Oxytetracycline (OTC), which was selected as the template molecule, was first rebound to the imprinted cavities. Gold nanoparticles were then labeled with the amino groups of OTC molecules via electrostatic adsorption and non-covalent interactions. Copper ions were catalytically reduced by the gold nanoparticles, and copper was deposited onto the electrode. The deposited copper was electrochemically dissolved, and its oxidative currents were recorded by differential pulse voltammetry (DPV). OTC could be determined indirectly within the concentration range of 3.0 × 10⁻¹⁰ to 1.5 × 10⁻⁷ mol L⁻¹ with a detection limit of 6.8 × 10⁻¹¹ mol L⁻¹.     

A selective modified carbon paste electrode for determination of cyanide using tetra-3,4-pyridinoporphyrazinatocobalt(II)

A chemically modified carbon paste electrode with 3,4-tetra pyridinoporphirazinatocobalt(II) (Co(3,4 tppa) was applied to the determination of free cyanide ion. The electrode has a linear range between 1.5 × 10⁻⁵ M and 1.0 × 10⁻² M with a Nernstian slope of 60 ± 1.5 mV/decade and its detection limit is 9 × 10⁻⁶ M. The response time of electrode is 5 min. The proposed electrode was applied successfully for the determination of cyanide in commercially available spring water. Some anions, such as SCN⁻, I⁻, Cl⁻, Br⁻ and oxalate that are usually serious interfering species for most of cyanide selective electrodes, did not have any interfering effect for this proposed electrode.     

A capacitive immunosensor for detection of cholera toxin

Contamination of food with biological toxins as well as their potential use as weapons of mass destruction has created an urge for rapid and cost effective analytical techniques capable of detecting trace amounts of these toxins. This paper describes the development of a sensitive method for detection of cholera toxin (CT) using a flow-injection capacitive immunosensor based on self-assembled monolayers. The sensing surface consists of monoclonal antibodies against the B subunit of CT (anti-CT), immobilized on a gold transducer. Experimental results show that the immunosensor responded linearly to CT concentrations in the range from 1.0 × 10⁻¹³ to 1.0 × 10⁻¹⁰ M under optimized conditions. The limit of detection (LOD) was 1.0 × 10⁻¹⁴ M. Two more analytical methods were employed for detection of CT using the same antibody namely, sandwich ELISA and surface plasmon resonance (SPR)-based immunosensor. The former had an LOD of 1.2 × 10⁻¹² M and a working range from 3.7 × 10⁻¹¹ to 2.9 × 10⁻¹⁰ M whereas, the later had an LOD of 1.0 × 10⁻¹¹ M and a linearity ranging from 1.0 × 10⁻⁹ to 1.0 × 10⁻⁶ M. These results demonstrate that the developed capacitive immunosensor system has a higher sensitivity than the other two techniques. The binding affinity of CT to the immobilized anti-CT was determined using the SPR-based immunosensor and an association constant (K_A) of 1.4 × 10⁹ M⁻¹ was estimated.     

Quantification of biogenic amines by microchip electrophoresis with chemiluminescence detection

A highly sensitive microchip electrophoresis (MCE) method with chemiluminescence (CL) detection was developed for the determination of biogenic amines including agmatine (Agm), epinephrine (E), dopamine (DA), tyramine, and histamine in human urine samples. To achieve a high assay sensitivity, the targeted analytes were pre-column labeled by a CL tagging reagent, N-(4-aminobutyl)-N-ethylisoluminol (ABEI). ABEI-tagged biogenic amines after MCE separation reacted with hydrogen peroxide in the presence of horseradish peroxidase (HRP), producing CL emission. Since no CL reagent was added to the running buffer, the background of the CL detection was extremely low, resulting in a significant improvement in detection sensitivity. Detection limits (S/N = 3) were in the range from 5.9 × 10⁻⁸ to 7.7 × 10⁻⁸ M for the biogenic amines tested, which were at least 10 times lower than those of the MCE–CL methods previously reported. Separation of a urine sample on a 7 cm glass/poly(dimethylsiloxane) (PDMS) microchip channel was completed within 3 min. Analysis of human urine samples found that the levels of Agm, E and DA were in the ranges of 2.61 × 10⁻⁷ to 4.30 × 10⁻⁷ M, 0.81 × 10⁻⁷ to 1.12 × 10⁻⁷ M, and 8.76 × 10⁻⁷ to 11.21 × 10⁻⁷ M (n = 4), respectively.     

Biosensor for multiplex detection of two DNA target sequences using enzyme-functionalized Au nanoparticles as signal amplification

Multiplex electrochemical detection of two DNA target sequences in one sample using enzyme-functionalized Au nanoparticles (AuNPs) as catalytic labels for was proposed. This DNA sensor was fabricated using a “sandwich” detection strategy, involving two kinds of capture probes DNA immobilized on glassy carbon electrode (GCE), and hybridization with target DNA sequences, which further hybridized with the reporter DNA loaded on the AuNPs. The AuNP contained two kinds of DNA sequences, one was complementary to the target DNA, while the other was noncomplementary to the target. The noncomplementary sequences were linked with horseradish peroxidase (HRP) and alkaline phosphatase (ALP), respectively. Enhanced detection sensitivity was obtained where the AuNPs carriers increased the amount of enzyme molecules per hybridization. Electrochemical signals were generated from the enzymatic products produced from the substrates catalyzed by HRP and ALP. Under optimal conditions, a 33-mer sequence could be quantified over the ranges from 1.5 × 10⁻¹³ to 5.0 × 10⁻¹² M with a detection limit of 1.0 × 10⁻¹³ M using HRP-AuNP as labels, and a 33-mer sequence could be quantified over the ranges from 4.5 × 10⁻¹¹ M to 1.0 × 10⁻⁹ M with a detection limit of 1.2 × 10⁻¹¹ M using ALP-AuNP as labels.     

Flow-injection chemiluminescence determination of formaldehyde in water

A modification of the Trautz-Schorigin reaction into a flow-injection analysis configuration is described. Different approaches were used at the optimization of chemiluminescence determination of formaldehyde in water based on the reaction of formaldehyde, gallic acid and hydrogen peroxide in an alkaline solution. Detection system with a 218 μl chemiluminescence cell was optimized by both a one-variable-at-a-time method, and a modified simplex method. A calibration graph is linear in the concentration range 4 × 10⁻⁸ to 1 × 10⁻⁵ M HCHO. The detection limit of formaldehyde for a signal-to-noise ratio of 3 is 4 × 10⁻⁸ M. The relative standard deviations for 15 repeated measurements of 1 × 10⁻⁶ and 5 × 10⁻⁶ mol l⁻¹ HCHO are 4.32 and 3.33%, respectively. The analysis time is 1.5 min. The method was applied to the determination of formaldehyde in urban rainwater. A comparison of results found by proposed method with those obtained by fluorimetric reference method provided a good agreement.     

Electrochemiluminescent disposable cholesterol biosensor based on avidin–biotin assembling with the electroformed luminescent conducting polymer poly(luminol-biotinylated pyrrole)

An electrochemiluminescent cholesterol disposable biosensor has been prepared by the formation of assembled layers on gold screen-printed cells. The detection layer is based on the electro-formation of new luminol copolymers with different synthesized biotinylated pyrroles prepared by click-chemistry, offering a new transduction layer with new electroluminescent properties on biosensors. The electrochemiluminescence (ECL) luminol copolymers are electroformed by cyclic voltammetry (five cycles) at pH 7.0 uses a10⁻³ M biotinylated pyrrole–luminol ratio of 1:10 in PBS buffer. With respect to the recognition layer, cholesterol oxidase was biotinylated by incubation with biotin vinyl sulfone, and immobilized on the copolymer by avidin–biotin interaction. The analytical signal of the biosensor is the ECL enzymatic initial rate working in chronoamperometric mode at 0.5 V excitation potential with 10 s between pulses at pH 9.5. The disposable device offers a cholesterol linear range from 1.5 × 10⁻⁵ M to 8.0 × 10⁻⁴ M with a limit of detection of 1.47 × 10⁻⁵ M and accuracy of 7.9% for 9.0 × 10⁻⁵ M and 14.1% for 2.0 × 10⁻⁴ M, (n = 5). Satisfactory results were obtained for cholesterol determination in serum samples compared to a reference procedure.     

Kinetic spectrophotometric determination of Tween 80

A novel kinetic spectrophotometic method for the determination of Tween 80 based on its interaction with 5(p-dimethylaminobenzylidene)rhodanine (PDR) in alkaline media is reported. The effect of variable on the rate of interaction of Tween 80 and PDR was investigated in order to establish the optimum conditions. The interaction was monitored spectrophotometically and change in absorbance (ΔA) of PDR at 464 nm at times of 30 and 270 s was used as an analytical parameter. Tween 80 can be measured in the range of 2.5×10⁻⁵ to 1.25×10⁻³ M with detection limit of 1.5×10⁻⁵ M. The relative standard deviation for eight replicate determinations of 2×10⁻⁴ and 1×10⁻³ M of Tween 80 solution was 4.08 and 3.88%, respectively. This method was used to determine Tween 80 in biscuit and multivitamin syrup.     

Flow-injection fluorometric quantification of pyruvate using co-immobilized pyruvate decarboxylase and aldehyde dehydrogenase reactor: Application to measurement of acetate, citrate and l-lactate

A flow-injection system for the quantification of pyruvate based on the coupled reactions of pyruvate decarboxylase (PDC) and aldehyde dehydrogenase (AlDH) was conceived and optimized. A co-immobilized PDC and AlDH reactor was introduced into the flow line. Sample and reagent (NAD⁺) were injected into the flow line by an open sandwich method and the increase of NADH produced by the immobilized-enzyme reactor was monitored fluorometrically at 455 nm (excitation at 340 nm). Linear relationships between the responses and concentrations of pyruvate were observed in the ranges of 2.0 × 10⁻⁵ to 1.5 × 10⁻³ M at the flow rate of 1.0 ml min⁻¹ and 5.0 × 10⁻⁶ to 1.0 × 10⁻³ M at the flow rate of 0.5 ml min⁻¹. The relative standard deviation for 10 successive injections was 0.95% at the 1.0 mM level. This FIA system for pyruvate was applied to the measurement of acetate, citrate and l-lactate.     

Urea as new stabilizing agent for imipenem determination: electrochemical study and determination of imipenem and its primary metabolite in human urine

Imipenem shows a fast chemical conversion to a more stable imin form (identical to that of biochemical dehydropeptidase degradation) in aqueous solutions and stabilizing agents used avoid its electrochemical study and determination.The aim of this work is the proposal of urea as stabilizing agent which allows the electrochemical study of imipenem and the proposal of electrochemical methods for the determination of imipenem and its primary metabolite (M1) in human urine samples. Electrochemical studies were realized in phosphate buffer solutions over pH range 1.5-8.0 using differential-pulse polarography, DC-tast polarography, cyclic voltammetry and adsorptive stripping voltammetry. In acidic media, a non-reversible diffusion-controlled reduction involving a two steps mechanism which involves one electron and one proton in the first step and two electrons and two protons in the second step occurs and the mechanism for the reduction was suggested.A differential-pulse polarographic method for the determination of imipenem in the concentration range 3.2 × 10⁻⁶ to 2 × 10⁻⁵ M (0.95-3.4 mg/L) and its primary metabolite in the concentration range 1.4 × 10⁻⁶ to 10⁻⁴ M (0.43-26.1 mg/L) with detection limits of 9.6 × 10⁻⁷ M (0.28 μg/L imipenem) and 4.3 × 10⁻⁷ M (0.14 μg/L M1) was proposed. Also, a method based on controlled adsorptive pre-concentration of imipenem on the hanging mercury drop electrode followed by voltammetric measure, allows imipenem determination in the concentration range 1.8 × 10⁻⁸ to 1.2 × 10⁻⁶ M (5.42-347 μg/L) with a detection limit of 5.4 × 10⁻⁹ M (1.63 μg/L). The proposed methods have been used for the direct determination of the analytes in a pharmaceutical formulation and human urine.     

Time measurement-visual analysis of l-cysteine using the autocatalytic sodium sulfite/hydrogen peroxide reaction system and its application to length detection-flow analysis

Trace amounts of l-cysteine can function as a trigger, i.e., reaction initiator, in the autocatalytic sodium sulfite/hydrogen peroxide reaction system. Rapidly changing of pH after induction time is visually confirmed by color changing of bromothymol blue in this autocatalytic reaction. Based on this finding, μg L⁻¹ levels of l-cysteine were measured over time using the autocatalytic reaction system. The determination range using the above method was 5.0 × 10⁻⁸-2.5 × 10⁻⁶ M, the detection limit (3σ) was 1.8 × 10⁻⁸ M (1.94 μg L⁻¹), and the relative standard deviation was 2.41% at an l-cysteine concentration of 5 × 10⁻⁷ M (n = 5). This method was also applied to length detection-flow injection analysis. The determination range for the flow injection analysis was 2.0 × 10⁻⁷-1.0 × 10⁻⁵ M. The detection limit (3σ) was 1.4 × 10⁻⁷ M (17.0 μg L⁻¹), and the relative standard deviation was 0.91% at an initial l-cysteine concentration of 10⁻⁶ M (n = 5).     

Electrochemiluminescence biosensor for the assay of small molecule and protein based on bifunctional aptamer and chemiluminescent functionalized gold nanoparticles

An electrochemiluminescence (ECL) biosensor for simultaneous detection of adenosine and thrombin in one sample based on bifunctional aptamer and N-(aminobutyl)-N-(ethylisoluminol) functionalized gold nanoparticles (ABEI-AuNPs) was developed. A streptavidin coated gold nanoparticles modified electrode was utilized to immobilize biotinylated bifunctional aptamer (ATA), which consisted of adenosine and thrombin aptamer. The ATA performed as recognition element of capture probe. For adenosine detection, ABEI-AuNPs labeled hybridization probe with a partial complementary sequence of ATA reacted with ATA, leading to a strong ECL response of N-(aminobutyl)-N-(ethylisoluminol) enriched on ABEI-AuNPs. After recognition of adenosine, the hybridization probe was displaced by adenosine and ECL signal declined. The decrease of ECL signal was in proportion to the concentration of adenosine over the range of 5.0 × 10⁻¹²–5.0 × 10⁻⁹ M with a detection limit of 2.2 × 10⁻¹² M. For thrombin detection, thrombin was assembled on ATA modified electrode via aptamer–target recognition, another aptamer of thrombin tagged with ABEI-AuNPs was bounded to another reactive site of thrombin, producing ECL signals. The ECL intensity was linearly with the concentration of thrombin from 5 × 10⁻¹⁴ M to 5 × 10⁻¹⁰ M with a detection limit of 1.2 × 10⁻¹⁴ M. In the ECL biosensor, adenosine and thrombin can be detected when they coexisted in one sample and a multi-analytes assay was established. The sensitivity of the present biosensor is superior to most available aptasensors for adenosine and thrombin. The biosensor also showed good selectivity towards the targets. Being challenged in real plasma sample, the biosensor was confirmed to be a good prospect for multi-analytes assay of small molecules and proteins in biological samples.     

Ultrasensitive determination of silver ion based on synchronous fluorescence spectroscopy with nanoparticles

Based on the characteristics of synchronous fluorescence spectroscopy (SFS), a new method with high sensitivity and selectivity was developed for rapid determination of silver ion with functional cadmium sulphide (CdS) nanoparticles as a fluorescence probe. When Δλ (λ_em − λ_ex) = 215 nm, maximum synchronous fluorescence is produced at 304 nm. Under optimal conditions, functional cadmium sulphide displayed a calibration response for silver ion over a wide concentration range from 0.8 × 10⁻¹⁰ to 1.5 × 10⁻⁸ mol L⁻¹. The limit of detection was 0.4 × 10⁻¹⁰ mol L⁻¹ and the relative standard deviation of seven replicate measurements for the lowest concentration (0.8 × 10⁻¹⁰ mol L⁻¹) was 2.8%. Compared with several fluorescence methods, the proposed method had a wider linear range and improved the sensitivity. Furthermore, the concentration dependence of the synchronous fluorescence intensity is effectively described by a Langmuir-type binding isotherm.     

An electrochemical sensor prepared by sonochemical one-pot synthesis of multi-walled carbon nanotube-supported cobalt nanoparticles for the simultaneous determination of paracetamol and dopamine

Multi-walled carbon nanotubes (MWCNTs) functionalized by cobalt nanoparticles were obtained using a single step chemical deposition method in an ultrasonic bath. The composite material was characterized using scanning electron microscopy (SEM) and energy dispersive X-ray analysis (EDX). The electroactivity of the cobalt-functionalized MWCNTs was assessed in respect to the electrooxidation of paracetamol (PAR) and dopamine (DA). It was found that the carbon nanotube supported cobalt nanoparticles have significantly higher catalytic properties. The proposed electrode has been applied for the simultaneous determination of PAR and DA. The modified electrode could resolve the overlapped voltammetric waves of PAR and DA into two well-defined voltammetric peaks with peak to peak separation of about 203 mV. On the other hand, the presence of potential drug interfering compounds AA and UA did not affect the voltammetric responses of PAR and DA. The current of oxidation peaks showed a linear dependent on the concentrations of PAR and DA in the range of 5.2 × 10⁻⁹–4.5 × 10⁻⁷ M (R² = 0.9987) and 5.0 × 10⁻⁸–3.0 × 10⁻⁶ M (R² = 0.9999), respectively. The detection limits of 1.0 × 10⁻⁹ M and 1.5 × 10⁻⁸ M were obtained for PAR and DA, respectively. The proposed electrode showed good stability (peak current change: 4.9% with and RSD of 2.6% for PAR; 5.5% with and RSD of 3.0% for DA over 3 weeks), reproducibility (RSD 2.3% for PAR and RSD 1.5% for DA), repeatability (RSD 2.25% for PAR and RSD 2.50% for DA) and high recovery (99.7% with an RSD of 1.3% for PAR; 100.8% with an RSD of 1.8% for DA). The proposed method was successfully applied to the determination of PAR and DA in pharmaceuticals.     

High sensitive determination of zinc with novel water-soluble small molecular fluorescent sensor

A high sensitive method of quantitative analysis for the determination of zinc in the nutrition supplements has been developed by using a novel water-soluble fluorescent sensor HQ3: (8-pyridylmethyloxy-2-methyl-quinoline). Under the optimized condition of 67 mM phosphate buffer, pH 7.4, and 5% (v/v) DMSO, the zinc concentration showed good linear relationship with fluorescence intensity in the range of 7.5 × 10⁻⁸ to 2.5 × 10⁻⁵ M with the detection limit of 1.5 × 10⁻⁸ M. HQ3 exhibited high selectivity to zinc comparing with other metal ions except for cadmium. The developed analytical method was successfully used for determining the content of zinc in a real sample of zinc gluconate solution of Sanchine.     

A thin-layer contactless conductivity cell for detection in flowing liquids

The design is described of a thin-layer contactless conductivity detector suitable for liquid chromatography and flow-injection analysis. Its principal analytical parameters have been determined using a potassium chloride solution: the linear dynamic range extends from 7.5 × 10⁻⁶ to 1.5 × 10⁻² S m⁻¹, corresponding to the KCl concentration range from 0.5 to 1000 μM, the limit of detection equals 3.5 × 10⁻⁶ S m⁻¹ (0.2 μM KCl), the detection repeatability, expressed in terms of the relative standard deviation, amounts to 1.13% and the detection volume is 0.6 μL. The detector was applied to detection of ionic compounds, benzoic, lactic and octanesulfonic acids, and sodium capronate, after their separation by liquid chromatography in a Biospher PSI 100C 18 columns using a 60% aqueous acetonitrile mobile phase. The frequency characteristics of the detector are reasonably theoretically described on the basis of a simple model which is commonly used in the field of contactless impedance detectors.     

Raman sensitivity enhancement for aqueous absorbing sample using Teflon-AF 2400 liquid core optical fibre cell

The compatibility Teflon-AF 2400 liquid core optical fibre with resonance Raman spectroscopy (RRS-LCOF) was used to detect aqueous biomolecules. The maximum sensitivity enhancement factor for concentrations greater than the detection limit in a conventional cell was 10, and detection limit reduction of about 1000-fold have been achieved for the measurement of aqueous absorbing sample using Teflon-AF 2400 fibre Raman cell compared to the conventional cell. We were able to collect spectra of 2.5 × 10⁻⁹ and 2.5 × 10⁻¹⁰ M aqueous β-carotene using 16.2 mW of laser power and 10 s integration time. This volume of a 2.5 × 10⁻¹⁰ M aqueous solution corresponds to only 1.5 fmol or 830 fg of β-carotene. The results of this preliminary study indicate that RRS-LCOF has potential in bioanalytical and biomedical applications.     

Selective recognition of ovalbumin using a molecularly imprinted polymer

Micro-contact imprinting has been used to form thin-film molecular imprints of ovalbumin (OVA) in polymers supported on glass slides. Thermocalorimetric data was used to optimise the choice of functional monomer and cross-linker to maximise selectivity and minimise non-specific recognition.A polymer comprising polyethyleneglycol 400 dimethacrylate (95 vol.%) and methacrylic acid (5 vol.%) showed both maximum recognition for OVA when made as a molecularly imprinted polymer (MIP), and minimal recognition when made as a non-imprinted, i.e. control polymer. OVA rebinding to the molecularly imprinted polymer, from a buffered 2 µM OVA solution, was 1.55 × 10^{− 11} mol cm^{− 2}, while the control polymer showed 10-fold less re-binding, i.e. 0.154 × 10^{− 11} mol cm^{− 2}.Experiments in which human serum albumin (HSA), conalbumin, ovomucoid or lysozyme, were re-bound to the polymers, either as single proteins or in competition with OVA, showed them to have low affinity for the polymer formulation used. Of the competing proteins examined, in non-competitive binding experiments, HSA showed the greatest affinity 0.45 × 10^{− 11} mol cm^{− 2} for the OVA imprinted polymer. In two protein competition experiments, i.e. with OVA and a competing protein present at equal concentrations (2 µM), OVA binding to the OVA imprinted polymer was in all cases significantly greater than that of the competitor.