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Herein, a novel magnetic biocomposite (Fe3O4@Au-S1/S2) was applied to analyze thrombin. The Fe3O4@Au-S1/S2 consisted of Fe3O4@Au nanoparticles (Fe3O4@Au NPs) as carriers for magnetic separation and magnetic field-induced self-assembly, thiolated complementary strand (S1) anchored based on Au-S bond and thrombin binding aptamer (S2) as a recognition element. As a redox indicator, methylene blue (MB) can be adsorbed to DNA anchored on the surface of Fe3O4@Au NPs by electro-static interaction. In the absence of thrombin, MB were adsorbed on double-stranded DNA (S1/S2) which anchored on Fe3O4@Au NPs and a high electrochemical signal of MB was recorded by Differential pulse voltammetry. Conversely, the complementary strand (S1) exposed after thrombin competitively bonded with aptamer. The introduction of Pb2+-dependent DNAzyme (S3) split S1 at specific rA site, resulting in the significantly decreased adsorption capacity of MB. Thus, the thrombin detection could be recorded by monitoring the electrochemical signal reduction of MB through incubation of thrombin with S3. This method exhibited a high sensitivity toward thrombin with a broad linear range from 5 pmol L−1 to 5 nmol L−1 and a limit of detection of 1.8 pmol L−1.

Signal amplification is very important for electrochemical immunoassays. We report an enzyme-free electrochemical immunosensor based on an electrochemical-chemical-chemical (ECC) redox cycle advanced (RCA) signal amplification strategy for the ultrasensitive detection of Carcinoembryonic antigen (CEA). In this scheme, CeO2/Au NPs@SiO2 is connected with ferrocene as the redox indicator, and the detection buffer is composed of the reducing agent hydroquinone (HQ) and tris(2-carboxyethyl) phosphine (TCEP). First, ferrocenecarboxylic acid (FcA) is oxidized to FcA+ on the electrode. Then, HQ reduces FcA+ to FcA to trigger the cyclic reaction of the inner ring. Second, the oxidation product of HQ is catalyzed by TCEP. The product is reduced to HQ again to complete the cyclic reaction of the outer ring, so the entire cyclic reaction forms a closed loop. The system realizes the high-efficiency regeneration of the electroactive material Fc, thereby ensuring the full amplification of the electrical signal. The results show that the immunosensor exhibits good analytical performance, the detection range is 0.01 pg/mL-80 ng/mL, the detection limit is 0.0037 pg/mL, and the immunosensor has excellent selectivity and stability and performs well in the detection of actual samples. This strategy provides a new method for the early screening of CEA.

A new redox oligomer “oligo-methoxy-phenyl-acetonitrile” (Fc-acid-OMPA) modified with ferrocene groups and carboxylic acids as functional groups in side chains, has been synthesized. The redox indicator has been used for monitoring the electrochemical behavior. The functional groups give the flexibility for direct covalent attachment of biomolecules. The electrochemical properties of the redox oligomer film deposited on gold electrodes have been studied by cyclic voltammetry (CV), which showed a rate of electron transfer of 6.43 s−1. The oligomer has been studied as a transducer for electrochemical DNA sensing and for this purpose the acid functional group of Fc-acid-OMPA was attached with the DNA probe of hepatitis C bearing amino group in 5′ position through amid link. The efficiency of DNA attachment on the oligomer film has been analyzed by X-Ray Photoelectron Spectrometry (XPS) and FT-IR spectroscopy. Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) have been used to analyze the biosensor construction and DNA detection. A wide linear range of detection from 1 fM to 100 pM have been demonstrated with a limit of detection (LOD) of 0.2 fM. The biosensor has showed an appreciated sensitivity to PCR samples of genomic DNA from Mycobacterium tuberculosis, and has been able to detect a single mutation which confers resistance of M. tuberculosis to rifampicin drug.

Electroactive metal-organic frameworks (MOFs) with large surface area and manipulatable structural properties show promise as a new type of signal probe for electrochemical biosensing application. In this work, an electroactive Ni-MOF, assembled by the redox-active ligands 4,4′,4″-Tricarboxytriphenylamine (H 3 TCA), a triphenylamine derivatives, as the electroactive source and magnetic ordered Ni 4 O 4 clusters as electronic transport nodes, is first designed and applied for electrochemical aptasensing of thrombin (Tb). The designed Ni-MOF probe realizes a stable and sensitive electrochemical signal output based on simple sandwich-type aptasensing because the high-content TCA active sites and good magnetic ordered intermediate of Ni 4 O 4 clusters are periodically arranged in well-defined porous structure of the MOF. The Ni-MOF probe assembled by redox-active ligand presents the high stability and can be directly applied in electrochemical aptasensor, avoiding any post-modification and the addition of redox mediators. As a result, the constructed electrochemical aptasensor shows a wide linear relationship for Tb from 0.05 pM to 50 nM and a detection limit of 0.016 pM ( S / N = 3). Furthermore, the proposed aptasensor is successfully applied to analysis of target Tb in real serum sample with satisfactory results. The present work indicates that fabricating a redox-active organic molecule in functionalized MOFs offer a feasible strategy to design high-stable electroactive MOFs for construction of electrochemical biosensors with simplicity, high selectivity and sensitivity.

An electrochemical biosensor capable of indirect detection of DNA damage induced by any one of the three endocrine-disrupting compounds (EDCs) – bisphenol A (BPA), 4-nonylphenol (NP) and 4- t -octylphenol (OP), has been researched and developed. The methylene blue (MB) dye was used as the redox indicator. The glassy carbon electrode (GCE) was modified by the assembled dsDNA/graphene oxide-chitosan/gold nano-particles to produce a dsDNA/GO-CS/AuNPs/GCE sensor. It was characterized with the use of electrochemical impedance spectroscopy (EIS), cyclic voltammetry (CV) and scanning electron microscopy (SEM) techniques. The loading/release of the MB dye by the dsDNA/GO-CS/AuNPs film was investigated, and the results showed that the process was reversible. Based on this, the sensor was used to measure the difference between the loading capabilities of intact and damaged dsDNA in the films. The sensor was then successfully applied to detect DNA damage electrochemically. The differential pulse voltammetry (DPV) peak current ratio for MB, observed before and after DNA damage, increased linearly in the presence the BPA, NP or OP compounds; the treatment range was 10–60 min, and the respective damage rates were 0.0069, 0.0044 and 0.0031 min −1 , respectively. These results were confirmed by the binding constants: 2.09 × 10 6 M −1 (BPA-DNA), 1.28 × 10 6 M −1 (NP-DNA) and 9.33 × 10 5 M −1 (OP-DNA), all of which were obtained with the use of differential pulse stripping voltammetry (DPSV).

The paper concerns mainly the modified Gran methods, considered as extrapolative standard addition methods. Particularly, the approximation: [Formula: see text] where xi congruent with 2, is applied for the needs of the modified Gran I methods, in accurate and simplified versions, denoted as MG(I)A and MG(I)S methods. These methods are compared with Gran II methods, in original (G(II)) and modified accurate (MG(II)A) versions. All the methods were tested with use of results obtained from simulated potentiometric titrations. The effects involved with a pre-assumed slope vartheta(*)-value for a redox indicator electrode and in errors in potential (E) and volume (V) readouts on the error in equivalence volume (V(eq)) value thus determined is considered. It is recognized that MG(I)A and MG(II)A appear to be the best methods applicable for V(eq) determination among ones considered hitherto. Moreover, the MG(I)S method is the best choice for determination of the true slope theta(*)-value for the redox indicator electrode.

The electrochemical generation of quinoid intermediates and their subsequent reaction with sulphydryl thiols (RSH) have been investigated as a method by which the latter can be quantified. Nucleophilic addition of the thiol species to the electrogenerated quinoid structure leads to the production of a chemically reduced adduct whose subsequent re-oxidation at the electrode leads to an amplification of the oxidative current. This can be correlated to the concentration of thiol present within the medium. The influence of indicator composition on the nature of the analytical response has been assessed through examining the electrochemical properties of 15 derivatives and the validity of the proposed reaction pathway corroborated by comparison of the voltammetric response with computer simulations. Interference by other electroactive species (ascorbate, urate, tyrosine) and other physiological constituents (cystine, lysine) has also been assessed. © 2001 Elsevier Science B.V. All rights reserved.

Optical chemical sensing schemes exploiting redox properties of Prussian Blue/Prusian White film system are reported. Transparent, thin film composed of Prussian Blue and N-substituted polypyrrole was chemically deposited on non-conducting polyester foil. The process of the film formation is induced and accelerated by UV radiation. The film is robust, crack-free and strongly adheres to the plastic support. The film was used as a receptor part of a flow-through detector cell. All spectrophotometric measurements were performed at the wavelength of 720 nm, i.e. at the absorption maximum of the Prussian Blue film. The Prussian Blue/Prussian White film sensor system exhibits excellent operational stability and long shelf-life. The Prussian Blue film is shown to be suitable for optical determination of selected strong reductants. Also potential application of the film for optical determination of alkali cations involved in the redox process is shown. In situ generated Prussian White film is mechanically stable, however, can be easily oxidized to Prussian Blue form. This feature of the Prussian White film can be applied for optical determination of various oxidants. It was also shown that the presented detector cell is useful for optical determination of redox species in non-transparent, turbid samples. The Prussian Blue/Prussian White film redox system offers a new approach to near-infrared optical sensing applications.

The redox indicator, Ferroin, tris(1,10-phenanthroline) iron(II) was successfully incorporated into the perfluorosulfonated ionomer membrane Nafion®. The redox properties of the optode membrane were studied under batch and flow conditions using acidified bromate or cerium(IV) solutions as oxidants and ascorbic acid or sodium ascorbate solutions as reductants. The experiments under flow conditions were performed in a computer-controlled sequential flow injection system incorporating a flow-through measuring cell where the optode membrane was mounted. Optical fibre technology was used to monitor the membrane absorbance with time. Some loss of sensitivity was observed after a number of redox cycles of the optode membrane. This was attributed to the dissociation of the Ferroin in the presence of H+ ions that entered the optode membrane from the acidified oxidising solutions and to the formation of a binuclear Fe(III)/1,10-phenanthroline complex when Ferroin was oxidised. Both undesirable effects were suppressed by the addition of excess 1,10-phenanthroline to the membrane composition.

The reversible redox couples (Fe(III)/Fe(II); Ce(IV)/Ce(III); Fe(CN) 6 3– /Fe(CN) 6 4− ; I 2 /I − ; Br 2 /Br − and VO 3 − /VO 2+ ) were studied as indicating redox systems for biamperometric determination of paracetamol in a flow injection assembly. Considering the selectivity of assayed systems against excipients and antioxidants that typically are presented in pharmaceutical formulations (e.g. sucrose, glucose, ascorbic acid and hydrogen sulphite) the systems Fe(III)/Fe(II) and VO 3 − /VO 2+ were selected for the determination of the drug. The sample was injected into a carrier which merged with the oxidant stream where the drug was oxidised by excess oxidant. The observed signal (current) showed a linear dependence on the concentration of reduced species. The calibration graphs were linear over the range 0.1–7 μg ml −1 of paracetamol for iron(III) and up to 20 μg ml −1 for metavanadate, with a relative standard deviation of 1.6 and 1.5% (5 μg ml −1 paracetamol; n = 20) and a sample throughput of 149 and 79 h −1 for iron(III) and metavanadate, respectively. Under flow optimal chemical and conditions for determining paracetamol both indicating systems were tested for different families compounds of pharmaceutical interest (phenothiazines, sulphonamides, nitrocompounds, epinephrine derivates, tetracyclines and pyridine derivates).

A new redox indicator reaction between Cu 2+ , Mn 2+ and SO 2− 3 in the presence of 2,2′-bicinchoninic acid is investigated. Optimum reaction conditions are determined. The influence of nonaqueous solvents on the reaction rate is studied. It is shown that in the presence of isopropyl alcohol the sensitivity of sulfite determination increases up to 0.3 μg/ml. Random errors lie within the limits allowable at the determination of microconcentrations. To raise the sensitivity of the determination of sulfite in some inorganic salts (KCl, KNO 3 , CsNO 3 ), this impurity was concentrated by the method of low-temperature directed crystallization. The metrological characteristics of the developed technique are presented.

The activities of native and immobilized alcohol oxidases (AOD) originating from Pichia pastoris, Hansenula sp., Candida boidinii and Peniophora gigantea were compared with regard to the oxidation of various higher alcohols. Except Candida boidinii , the immobilized enzymes were applied in a flow-injection analysis system and investigated as a function of the various operation variables. During the oxidation O 2 is consumed which concentration was measured with an amperometric detector. At the same time H 2 O 2 is produced, the concentration of which is used for the oxidation of a redox indicator (ABTS) catalysed by peroxidase and measured with a photometer. The amperometric method is simpler and more economical, but less sensitive. The photometric method consumes the dissolved redox indicator and peroxidase, but is highly sensitive especially for higher alcohols. The concentrations of n -propanol, n -butanol and n -pentanol can be measured with satisfactory sensitivity with immobilized alcohol oxidases originating from Pichia pastoris and Peniophera gigantea in a one molar potassium phosphate buffer (pH 7.5). The concentrations of isopropanol, octanol, dodecanol and benzylalcohol can be measured only when larger than 1 g l −1 .

By using three equations, viz., the Nernst equation, the acid-base dissociation constant equation and the equation for the Kubelka-Munk function for reflectance, an equation describing the redox behaviour of the immobilised indicator 2,6-dichloroindophenol has been formulated. After determination of the acid dissociation constant of the reduced form of the indicator, theoretical responses were calculated and compared to the experimental responses. The results indicate that the redox equilibrium involves a one-electron transfer in the pH range 2 to 5.

According to the current demands of Green Analytical Chemistry and regarding the need for lower reagent consumption with improved analytical performance, an automatic methodology with a flow-through optosensor incorporating solid-phase spectrophotometric detection was developed. The sensor used in this work was based on the redox state of thionine whose oxidized form is blue and reduced form is colorless with monitoring carried out at 621 nm. This redox indicator was immobilized on gel beads and subsequently packed into a flow-through cell. It was then assembled into a sequential injection system and was shown to be an excellent alternative to monitor enzymatic redox reactions automatically as the redox catalysis is performed by glucose dehydrogenase. This enzyme is a representative dehydrogenase enzyme and uses NAD(+) as cofactor, promoting the oxidation of glucose to glucono-lactone and reduction of NAD(+) to NADH. The produced NADH promotes color depletion on the surface of the sensor. The calibration graph for glucose was linear between 5.74 x 10(-4) and 2.00 x 10(-3) mol L(-1) and detection limit was 1.72 x 10(-4) mol L(-1). Glucose concentration in different samples including sera, salines, perfusion solutions, powder for preparing oral solutions and solutions for hemodialysis was determined. The method proved to be reproducible with a RSD < 5% for glucose determinations.

This study describes the application of the modified Gran I and II methods for determination of equivalence volume in redox titrations and for calibration, i.e. for determination of the correct slope of the redox indicator electrodes, which appears to be a major problem in using these electrodes. The four new versions of the Gran II method, denoted as MG(II)B, MG(II)C, MG(II)D and MG(II)E methods, in which the problem of calibration and determination of equivalence volume are considered simultaneously are suggested and compared with MG(I)S and MG(II)A methods known hitherto. All the methods are experimentally tested and discussed.

A sensitive kinetic fluorimetric system is proposed for the determination of hydrogen peroxide produced by enzymatic oxidation of cholesterol, l -amino acids and linoleic acid. 2-Hydroxynaphthaldehyde thiosemicarbazone (HNTS) is oxidized by hydrogen peroxide or hydroperoxides in an ammoniacal medium in a Mn(II)-catalyzed reaction to give a fluorescent product (λex = 390 nm, λem = 450 nm). The lowest concentration of hydrogen peroxide determined is 50 pmol. Cholesterol was determined in egg yolk, cod liver oil and horse serum. The ranges of concentration for substrates were 0.33–3.74 μM cholesterol; 0.3–10 μM, 0.6–15 μM and 0.75–10 μM, for l -leucine, l -phenyalanine and l - serine, respectively; and 15–150 μM linoleic acid.

A volumetric method is proposed for the rapid determination of small amounts of uranium. The hexavalent uranium is reduced to the quadrivalent state by chromous chloride, the excess of which is destroyed by air oxidation. A low potential redox indicator is used to follow the reduction and air oxidation. Ferric sulphate is added to the solution and the resulting ferrous ion titrated with dichromate solution.

Theoretical expressions are derived for the color transition of variamine blue B base used as a redox indicator in the compleximetric titration of iron (III) with EDTA , and photometric and visual methods of end-point location are discussed. The addition of iron(II) favored the elimination of the interference of some metals. The effect of added iron(II) is illustrated and quantitatively accounted for.