A regenerable electrochemical sensor for electro-inactive cyclovirobuxine D detection in biological samples.

Based on the pKa determination of cyclovirobuxine D (CVB-D) using the method of potentiometry, we predicted the ionization state of CVB-D at physiological pH. Thus, by taking advantage of the ionization state and consequent non-covalent interactions between protonated CVB-D and deprotonated polymerized bromothymol blue (poly-BTB) under physiological conditions, we developed a simple and reusable electrochemical sensor that contains a poly-BTB/SWNT-modified electrode for electro-inactive CVB-D detection in biological fluids using poly-BTB as both the recognition unit and the electrochemical probe. Upon being immersed in the solution of CVB-D, the poly BTB-based electrode shows a current decrease due to the interaction-driven binding of CVB-D on the electrode surface. The current decrease in the electrochemical sensor toward CVB-D concentration shows a linear relationship in the dynamic ranges of 0.01-1 µM and 1-50 µM with a detection limit of 1.65 nM based on 3σ. The sensor can be easily regenerated through the removal of the binding of CVB-D from the electrode surface by highly negatively charged heparin, and it presents high repeatability with an RSD of less than 4.0% for seven measurements. In animal experiments, the electrochemical sensor was selective and sensitive for CVB-D determination in plasma and liver homogenates. The electrochemical sensor is readily accessible, robust, and cost-effective and holds good promise for more applications in biological and clinical fields associated with CVB-D using less technically demanding and simple operating procedures.

Integrating brucine with carbon nanotubes toward electrochemical sensing of hydroxylamine.

Based on the intrinsic electrochemical features of brucine integrated with carbon nanotubes (brucine/SWNTs), dimeric quinoid brucine was electrochemically generated by electroactivation of a brucine/SWNTs-modified GC electrode and used as a novel electrocatalyst for efficient electro-oxidation of hydroxylamine (HA). The electrocatalytic activity was investigated with cyclic voltammetry in the range pH 2.0 to pH 11.0, and the best electrocatalytic performance of the electrocatalyst was obtained at pH 10.0. By taking advantage of the electrocatalytic activity of the dimeric quinoid brucine toward HA, we have developed an electrochemical sensor for HA measurements based on a brucine/SWNTS-modified GC electrode using amperometry with the applied potential of + 0.1 V (vs. Ag/AgCl). Under the optimized conditions, the current response toward HA concentration shows a linear relationship in the dynamic ranges of 0.1-10 µM and 10-1000 µM with a detection limit of 0.021 µM based on the 3σ criterion. The sensor was used to assay HA in pharmaceuticals including hydroxyurea tablets and pralidoxime iodide injections with satisfactory results. The spike-and-recovery for samples of tap water (n = 9) and lake water (n = 9) was within 97.17-100.16%. Graphical abstract Schematic illustration of electrochemical sensing of hydroxylamine (HA) enabled by integrating brucine with single-walled carbon nanotube (brucine/SWNTs) based on electro-activation of brucine/SWNTs-modified GC electrode.

Online electrochemical systems for continuous neurochemical measurements with low-potential mediator-based electrochemical biosensors as selective detectors.

This study demonstrates a new strategy to develop online electrochemical systems (OECSs) for continuously monitoring neurochemicals by efficiently integrating in vivo microdialysis with an oxidase-based electrochemical biosensor with low-potential electron mediators to shuttle the electron transfer of the oxidases. By using thionine and xanthine oxidase (XOD) as examples of low-potential mediators and oxidases, respectively, we demonstrate that the use of low-potential mediators to shuttle the electron transfer of oxidases would offer a new approach to the development of oxidase-based biosensors with theoretical and technical simplicity. To construct the XOD-based biosensor, thionine was adsorbed onto carbon nanotubes and used to shuttle the electron transfer of XOD. The XOD-based biosensor was positioned into an electrochemical cell that was directly coupled with in vivo microdialysis to form an online electrochemical system (OECS) for continuous and selective measurements of the substrate of XOD (with hypoxanthine as an example). The OECS based on the low-potential mediators is highly selective against the species endogenously existing in the brain system, which is attributed to the low operation potential benefited from the low redox potentials of the mediators. Moreover, the OECS demonstrated here is stable and reproducible and could thus be envisaged to find some interesting applications in physiological and pathological investigations. This study essentially offers a new strategy to develop online electrochemical systems, which is of great importance in understanding the molecular basis of physiological and pathological events.

An indicator displacement assay-based optical chemosensor for heparin with a dual-readout and a reversible molecular logic gate operation based on the pyranine/methyl viologen.

We have reported an optical indicator displacement assay (IDA) for heparin with a UV-vis absorbance and fluorescence dual-readout based on pyranine/methyl viologen (MV2+). Upon introducing heparin, pyranine/MV2+ shows a clearly observable increase in UV-vis absorbance and a turn-on of the fluorescence signal. We have demonstrated that the ionic nature of buffers significantly affects the pyranine displacement and the zwitterionic HEPES was most suitable for heparin sensing. After careful screening of experimental conditions, the pyranine/MV2+-based optical chemosensor exhibits a fast, sensitive, and selective response toward heparin. It shows dynamic linear concentration of heparin in the ranges of 0.1-40 U·mL-1 and 0.01-20 U·mL-1 for the absorptive and fluorescent measurements, respectively, which both cover the clinically relevant levels of heparin. As with the animal experiments, the optical chemosensor has been demonstrated to be selective and effective for heparin level qualification in rat plasma. The chemosensor is readily accessible, cost-effective, and reliable, which holds a great promise for potential application on clinical and biological studies. Furthermore, this IDA system can serve as an IMPLICATION logic gate with a reversible and switchable logical manner.

Online electrochemical measurements of Ca2+ and Mg2+ in rat brain based on divalent cation enhancement toward electrocatalytic NADH oxidation.

This study describes a novel electrochemical approach to effective online monitoring of electroinactive Ca(2+) and Mg(2+) in the rat brain based on the current enhancement of divalent cations toward electrocatalytic oxidation of NADH. Cyclic voltammetry for NADH oxidation at the electrodes modified with the polymerized film of toluidine blue O (TBO) reveals that the current of such an electrocatalytic oxidation process is remarkably enhanced by divalent cations such as Ca(2+) and Mg(2+). The current enhancement is thus used to constitute an electrochemical method for the measurements of Ca(2+) and Mg(2+) in a continuous-flow system with the polyTBO-modified electrode as the detector. Upon being integrated with in vivo microdialysis, the electrochemical method is successfully applied in investigating on cerebral Ca(2+) and Mg(2+) of living animals in two aspects: (1) online simultaneous measurements of the basal levels of Ca(2+) and Mg(2+) in the brain of the freely moving rats by using ethyleneglcol-bis(2-aminoethylether) tetraacetic acid (EGTA) as the selective masking agent for Ca(2+) to differentiate the net current responses selectively for Ca(2+) and Mg(2+); and (2) online continuous monitoring of the cerebral Mg(2+) following the global ischemia by using Ca(2+)-masking agent (i.e., EGTA) to completely eliminate the interference from Ca(2+). Compared with the existing methods for the measurements of cerebral Ca(2+) and Mg(2+), the method demonstrated here is advantageous in terms of its simplicity both in instrumentation and in the experimental procedures and near real-time nature, and is thus highly anticipated to find wide applications in understanding of chemical events involved in some physiological and pathological processes.

Noncovalent attachment of NAD+ cofactor onto carbon nanotubes for preparation of integrated dehydrogenase-based electrochemical biosensors.

This study describes a facile approach to the preparation of integrated dehydrogenase-based electrochemical biosensors through noncovalent attachment of an oxidized form of beta-nicotinamide adenine dinucleotide (NAD(+)) onto carbon nanotubes with the interaction between the adenine subunit in NAD(+) molecules and multiwalled carbon nanotubes (MWCNTs). X-ray photoelectron spectroscopic and cyclic voltammetric results suggest that NAD(+) is noncovalently attached onto MWCNTs to form an NAD(+)/MWCNT composite that acts as the electronic transducer for the integrated dehydrogenase-based electrochemical biosensors. With glucose dehydrogenase (GDH) as a model dehydrogenase-based recognition unit, electrochemical studies reveal that glucose is readily oxidized at the GDH/NAD(+)/MWCNT-modified electrode without addition of NAD(+) in the phosphate buffer. The potential for the oxidation of glucose at the GDH/NAD(+)/MWCNT-modified electrode remains very close to that for NADH oxidation at the MWCNT-modified electrode, but it is more negative than those for the oxidation of glucose at the MWCNT-modified electrode and for NADH oxidation at a bare glassy carbon electrode. These results demonstrate that NAD(+) molecules stably attached onto MWCNTs efficiently act as the cofactor for the dehydrogenases. MWCNTs employed here not only serve as the electronic transducer and the support to confine NAD(+) cofactor onto the electrode surface, but also act as the electrocatalyst for NADH oxidation in the dehydrogenase-based electrochemical biosensors. At the GDH/NAD(+)/MWCNT-based glucose biosensor, the current is linear with the concentration of glucose being within a concentration range from 10 to 300 microM with a limit of detection down to 4.81 microM (S/N = 3). This study offers a facile and versatile approach to the development of integrated dehydrogenase-based electrochemical devices, such as electrochemical biosensors and biofuel cells.

A facile fluorescent "turn-off" method for sensing paraquat based on pyranine-paraquat interaction.

Development of a technically simple yet effective method for paraquat (PQ) detection is of great importance due to its high clinical and environmental relevance. In this study, we developed a pyranine-based fluorescent "turn-off" method for PQ sensing based on pyranine-PQ interaction. We investigated the dependence of analytical performance of this method on the experimental conditions, such as the ion strength, medium pH, and so on. Under the optimized conditions, the method is sensitive and selective, and could be used for PQ detection in real-world sample. This study essentially provides a readily accessible fluorescent system for PQ sensing which is cheap, robust, and technically simple, and it is envisaged to find more interesting clinical and environmental applications.

Rapid-scan time-resolved FT-IR spectroelectrochemistry studies on the electrochemical redox process.

The electron transfer to or from molecules containing multiple redox centers has been extensively investigated. Rapid scan time-resolved FT-IR-RAS spectroelectrochemistry was used to investigate the electron-transfer mechanism in this report. The electron transfer of two typical compounds, 1,4-benzoquinone and 1,4-bis(2-ferrocenylvinyl)benzene, was examined with this method. Although the two compounds show two-electron transfer in the redox process, 1,4-benzoquinone exhibits two single electron waves while 1,4-bis(2-ferrocenylvinyl)benzene exhibits a single wave in cyclic voltammetric experiments. The IR absorption of the intermediate, BQ*- and p-(Fc-CH=CH)+2-benzene, at 1506 and 1589 cm(-1), respectively, appeared and disappeared on the experimental time scale in the oxidation and reduction process was observed. In the oxidation process of the p-(Fc-CH=CH)2-benzene molecule, one Fc was oxidated to Fc+ first and the electron-withdrawing ability of Fc+ was stronger than that of Fc, which resulted in the D-pi-A structure and the band at 1589 cm(-1) becoming visible. Then as the oxidation continues, the other Fc was oxidated to Fc+ too, which resulted in the reforming of the symmetry of the benzene ring A-pi-A, so the band at 1589 cm(-1) disappeared. Similar phenomenon can be elucidated in the reduction process but the configuration type changed from A-pi-A to D-pi-A and finally to D-pi-D. Hence, not only 1,4-benzoquinone but also 1,4-bis(2-ferrocenylvinyl)benzene show two consecutive one-electron processes. In addition, it is observed that the existing time of the electrochemical reaction intermediate (BQ*- and p-(Fc-CH=CH)+2-benzene) is prolonged at low temperatures due to slow reaction kinetics.

A non-oxidative electrochemical approach to online measurements of dopamine release through laccase-catalyzed oxidation and intramolecular cyclization of dopamine.

A new electrochemical approach to selective online measurements of dopamine (DA) release in the cerebral microdialysate is demonstrated with a non-oxidative mechanism based on the distinct reaction properties of DA and the excellent biocatalytic activity of laccase. To make the successful transition of the distinct sequential reaction properties of DA from a conceptual determination protocol to a practical online analytical system, laccase enzyme is immobilized onto magnetite nanoparticles and the nanoparticles are confined into a fused-silica capillary through an external magnetic field to fabricate a magnetic microreactor. The microreactor is placed in the upstream of the thin-layer electrochemical flow cell to efficiently catalyze the oxidation of DA into its quinonoid form and thereby initialize the sequential reactions including deprotonation, intramolecular cyclization, disproportionation and/or oxidation to finally give 5,6-dihydroxyindoline quinone. The electrochemical reduction of the produced 5,6-dihydroxyindoline quinone at bare glassy carbon electrode is used as the readout for the DA measurement. The laccase-immobilized microreactor is also found to catalyze the oxidation of ascorbic acid (AA) and 3,4-dihydroxyphenylacetic acid (DOPAC) into electroinactive species and, as such, to eliminate the great interference from both species. Moreover, the successful transition of the mechanism for DA detection from the conventional oxidative electrochemical approach to the non-oxidative one substantially enables the measurements virtually interference-free from physiological levels of uric acid, 5-hydroxytryptamine, norepinephrine, and epinephrine. The current response is linear with DA concentration within a concentration range from 1 to 20 microM with a sensitivity of 3.97 nA/microM. The detection limit, based on a signal-to-noise ratio of 3, is calculated to be 0.3 microM. The high selectivity and the good linearity as well as the high stability of the online method make it very potential for continuous monitoring of cerebral DA release in physiological and pathological processes.