High‐Quality Covalently Grafting Hemoglobin on Gold Electrodes: Characterization,Redox Thermodynamics and Bio‐electrocatalysis

Herein, we report a versatile surface chemistry methodology to covalently immobilize ligands and proteins to self‐assembled monolayers (SAMs) on gold electrode. The strategy is based on two steps: 1) the coupling of soluble azido‐PEG‐amimo ligand with an alkynyl‐terminated monolayer via click reaction and 2) covalent immobilization hemoglobin (Hb) to the amine‐terminated ligand via carbodiimide reaction. Surface‐enhanced Raman scattering spectroscopy (SERS), atomic force microscopy (AFM), reflection absorption infrared spectroscopy (RAIR) and cyclic voltammetry are used to characterize the model interfacial reactions. We also demonstrate the excellent biocompatibility of the interface for Hb immobilization and reliable application of the proposed method for H₂O₂ biosensing. Moreover, the redox thermodynamics of the Fe³⁺/Fe²⁺ couple in Hb is also investigated.     

Synergistic microfluidic and electrochemical strategy to activate and pattern surfaces selectively with ligands and cells

We show a straightforward, flexible synergistic approach that combines microfluidics, electrochemistry, and a general immobilization strategy to activate regions of a substrate selectively for the precise immobilization of ligands and cells in patterns for a variety of cell-based assays and cell migration and cell adhesion studies. We develop microfluidic microchips to control the delivery of electrolyte solution to select regions of an electroactive hydroquinone SAM. Once an electrical potential is applied to the substrate, only the hydroquinone exposed to electrolyte solution within the microfluidic channels oxidizes to the corresponding quinone. The quinone form can then react chemoselectively with oxyamine-tethered ligands to pattern the surface. Therefore, this microfluidic/electrochemistry strategy selectively activates the surface for ligand patterning that exactly matches the channel design of the microfluidic channel. We demonstrate the ease of this system by first quantitatively characterizing the electrochemical activation and immobilization of ligands on the surface. Second, we immobilize a fluorescent dye to show the fidelity of the methodology, and third, we show the immobilization of biospecific cell adhesive peptide ligands to pattern cells. This is the first report that combines microfluidics/electrochemistry and a general electroactive immobilization strategy to pattern ligands and cells. We believe that this strategy will be of broad utility for applications ranging from fundamental studies of cell behavior to patterning molecules on a variety of materials for molecular electronic devices.     

An interfacial oxime reaction to immobilize ligands and cells in patterns and gradients to photoactive surfaces

We report a molecularly controlled interfacial chemoselective methodology to immobilize ligands and cells in patterns and gradients to self-assembled monolayers on gold. This strategy is based on reacting soluble ketone or aldehyde tethered ligands to surface-bound oxyamine alkeanethiols to generate a covalent oxime linkage to the surface. We characterize the kinetic behavior of the reaction on the surface with ferrocenecarboxaldehyde (FcCHO) as a model ligand. The precise extent of immobilization and therefore surface density of FcCHO on the SAM is monitored and determined by cyclic voltammetry, which shows a peudo-first-order rate constant of 0.13 min(-1). In order to generate complex surface patterns and gradients of ligands on the surface, we photoprotected the oxyamine group with nitroveratryloxycarbonyl (NVOC). We show that ultraviolet light irradiation through a patterned microfiche film reveals the oxyamine group and we characterize the rate of deprotection by immobilization of ketone containing redox active groups. Finally, we extend this strategy to show biospecific cell attachment of fibroblast cells by immobilizing ketone-GRGDS peptides in patterns. The interfacial oxime reaction is chemoselective and stable at physiological conditions (pH 7.0, 37 degrees C) and may potentially be used to install ligands on the surface in the presence of attached cells to modulate the cell microenvironment to generate dynamic surfaces for monitoring changes in cell behavior in real time.     

Electrochemical recognition of cations by bis(pyrrolo)tetrathiafulvalene macrocycles

[structure: see text] Tetrathiafulvalene redox-responsive ligands devoid of cis/trans isomerism containing the electroactive bis(pyrrolo[3,4-d])tetrathiafulvalene moiety and polyether subunits have been synthesized. One ligand exhibits high binding affinities for Pb2+ and Ba2+ cations as shown by independent methods (1H NMR, UV-vis spectroscopy, and cyclic voltammetry). The ability of this receptor to electrochemically recognize Pb2+ and Ba2+ is shown by cyclic voltammetry.     

The effects of cross-linking and anodic surface roughening on quinone polymer/carbon electrodes

A polymer in which anthraquinone-2-carbonyl groups were bound to polyethyleneimine was coated onto a glassy carbon electrode. Electrodes of this kind were studied using cyclic voltammetry and pH 7 aqueous solutions. At pH <10 only those quinone units in contact with the carbon surface are electroactive. It was shown that anodic surface roughening increased the limited number of electroactive groups in the polymer film and gave more stable activity and narrower voltammetric peaks. Above pH 10 redox propagation through the layer is more rapid but the anionic product desorbs. This desorption was inhibited by cathodically cross-linking a layer of mixed polymers on a polyethyleneimine backboned polymer containing fluorenone units as well as anthraquinone units.     

Electrochemistry of cobalt-porphyrin complexes in aqueous solution

The polarography and cyclic voltammetry of cobalt(III) hematoporphyrin was investigated in buffered aqueous solutions. In the absence of nitrogenous bases, which can act as axial ligands, the cobalt porphyrin is not electroactive. Polarographic waves are found in solutions containing pyridine and similar compounds. Both cobalt(III) and cobalt(II) hematoporphyrin can add two pyridine molecules, although only one ligand may react at lower concentrations. Adsorption problems were encountered.     

Characterisation of iron binding ligands in seawater by reverse titration

Here we demonstrate the use of reverse titration - competitive ligand exchange–adsorptive cathodic stripping voltammetry (RT-CLE–ACSV) for the analysis of iron (Fe) binding ligands in seawater. In contrast to the forward titration, which examines excess ligands in solution, RT-CLE–ACSV examines the existing Fe-ligand complexes by increasing the concentration of added (electroactive) ligand (1-nitroso-2-naphthol) and analysis of the proportion of Fe bound to the added ligand. The data manipulation allows the accurate characterisation of ligands at equal or lower concentrations than Fe in seawater, and disregards electrochemically inert dissolved Fe such as some colloidal phases. The method is thus superior to the forward titration in environments with high Fe and low ligand concentrations or high concentrations of inert Fe.     

A multivalent PyBox asterisk ligand

A discrete multivalent PyBox ligand was investigated. An efficient synthesis and the properties of 1 are reported (UV-visible, cyclic voltammetry, NMR, MALDI-Tof). It incorporates a sulfur-rich persulfurated benzene core which was compatible with a metal-catalyzed reaction in spite of donating and oxidizable sulfur atoms. Metal-catalysis with a persulfurated aromatic ligand was demonstrated for the first time in a model reaction: the Rh-enantioselective hydrosilylation of acetophenone. The interesting features were the reactivity and the enantiocatalytic behavior while varying the metal content. This work promotes new thoughts toward chiral supramolecular assemblies or metal nanoparticles stabilized with chiral multivalent ligands.     

Complexing ability of the versatile,redox-active, 3-[3-(diphenylphosphino)propylthio]-3',4,4'-trimethyl-tetrathiafulvalene ligand

The synthesis of a ligand containing as an electroactive core a tetrathiafulvalene moiety, 3-[3-(diphenylphosphino)propylthio]-3',4,4'-trimethyl-tetrathiafulvalene, is reported. Its versatile ability to act as a bidentate or a monodentate ligand, as demonstrated by the metal carbonyl complexes obtained, is described. The novel cis-Mo(CO)(4)(P-TTF)(2) 4 and cis-W(CO)(4)(P,S-TTF) 6 complexes have been characterized by X-ray diffraction analyses and cyclic voltammetry measurements. Within complex 4, no significant influence of the two electroactive ligands on the molybdenum center was detected, whereas, in complex 6, a weak influence of the TTF redox-active core can be observed on the redox behavior of the metal center.     

Cyclic voltammetric studies into the reduction of a series of tris(1,3-diketonato)iron(III) compounds in dichloromethane and dimethylsulphoxide

A series of seven tris(1,3-diketonato)iron(III) chelates were prepared and studied using cyclic voltammetry in dichloromethane and dimethylsulphoxide. In the former solvent only a single reduction wave is observed and is assigned to the Fe(III)-Fe(II) couple. Despite the metal-based redox chemistry the formal potential is strongly influenced by the substituent groups on the chelating ligands and can be linearly correlated with the sum of the Taft inductive parameters for these substituents. In dimethylsulphoxide the reduced monoanion [Fe(1,3-diketonate)₃]⁻ undergoes a following chemical reaction which is interpreted as the extrusion of one 1,3-diketonate ligand. That reaction is an equilibrium and the position of the equilibrium is observed to depend upon the electronic effect of the substituent groups on the chelating ligands. For the strongly withdrawing trifluoroacetylacetonate ligand the dissociation of that ligand is essentially quantitative.     

Cover Feature: Copper and Nickel Complexes of Oxamate−Phenol Containing Ligands: A Structural Dichotomy in Oxidized Species (Eur. J. Inorg. Chem. 15/2023)

The copper and nickel complexes of two tetradentate ligands derived from bis(aminophenol) and bis(phenol) architectures connected by an oxamate linker were isolated. Depending on the metal and ligand, the complex is isolated with either an intact (deprotonated) ligand ( 1²⁻ ), one-electron oxidized ligand ( 2⁻ ) or quinone form ( 3 ). Surprisingly, the Mannich base is easier to oxidize than the amidophenol derivatives. The complexes were characterized by X-ray diffraction, cyclic voltammetry, UV-Vis-NIR and EPR spectroscopies. Complex 1 shows two reversible oxidation waves assigned to the successive iminosemiquinone/aminophenolate redox systems. Complex 2⁻ shows an intense NIR feature, as well as an EPR signal at g_iso=2.043, consistent with a metallic contribution to the main ligand radical SOMO. Complex 3 shows the typical feature of an isolated Cu(II) complex. Spectro-electrochemistry coupled to DFT calculations demonstrate a ligand-centered oxidative redox chemistry for all the complexes.     

Simple voltammetric approach for characterization of two-step surface electrode mechanism in protein-film voltammetry

Many enzymes embedding multivalent metal ions or quinone moieties as redox-active centres undergo electrochemical transformation via two successive electron transfer steps. If electrochemical features of such redox enzymes are analyzed with “protein-film voltammetry”, one frequently meets a challenging reaction scenario where the two electron transfers take place at the same formal potential. Under such conditions, one observes voltammogram with a single oxidation-reduction pattern hiding voltammetric features of both redox reactions. By exploring some aspects of the two-step surface EECrev mechanism one can develop simple methodology under conditions of square-wave voltammetry to enable recognizing and characterizing each electron transfer step. The method relies on the voltammetric features of the second electron transfer, which is coupled to a follow-up chemical reaction. The response of the second electron transfer step shifts to more positive potentials by increasing the rate of the chemical reaction. The proposed methodology can be experimentally applied by modifying the concentration of an electrochemically inactive substrate, which affects the rate of the follow-up chemical reaction. The final voltammetric output is represented by two well-separated square-wave voltammetric peaks that can be further exploited for complete thermodynamic and kinetic analysis of the EECrev mechanism.

     

Total synthesis of eustifolines A-D and glycomaurrol via a divergent Diels-Alder strategy

The Diels-Alder reaction between a quinone monoimine and cyclic diene allows for the construction of substituted carbazoles in a regiospecific manner. This methodology has sucessfully been employed in a divergent strategy, culminating in the synthesis of eustifolines A-D and glycomaurrol.     

Self-assembled monolayers of a hydroquinone-terminated alkanethiol onto gold surface. Interfacial electrochemistry and Michael-addition reaction with glutathione

An electroactive self-assembled monolayer (SAM) was fabricated by covalent attachment of a novel hydroquinone-terminated dodecanethiol onto the gold surface and its electrochemical behavior was investigated using cyclic voltammetry and electrochemical impedance spectroscopy. The capability of the designed SAM in immobilization of organic molecules onto the gold surface was studied utilizing the Michael-addition as a model reaction. The results obtained from cyclic voltammetry, electrochemical impedance and grazing incidence Fourier transform infrared (GI-FTIR) spectroscopy revealed that, upon applying an anodic potential to the Au-SAM electrode system in the presence of glutathione, the electrochemically generated p-quinone participated in a Michael-addition reaction with glutathione and the corresponding Michael adduct was formed at the solid–liquid interface. The kinetic parameters were then derived for this interfacial Michael-addition reaction.     

Application of a Carbon Paste Electrode Modified with a Schiff Base Ligand to Mercury Speciation in Water

A carbon paste electrode, modified with benzylbisthiosemicarbazone is used for mercury speciation in water samples. Mercury ion is selectively accumulated on the electrode surface at open circuit and its analysis was performed by cyclic voltammetry or square‐wave voltammetry (SWV). A detection limit of 8 μg L^?1 (3σ) was found for 15 min of accumulation using SWV as measurement technique. The effect of several metallic ions and organic substances on voltammetric signal is examined. For speciation purposes, a ligand competition methodology between ligands in solution and electrode is used. Model mercury complexes are characterized as a function of their dissociation kinetics. The method was applied to mercury speciation in water samples from the Jarama River in Madrid.     

Tetrathiafulvalene hydrazone an efficient precursor of various chelating electroactive ligands

Novel bidentate electroactive ligands containing one or two tetrathiafulvalene (TTF) cores as redox active unit have been synthesized thanks to the condensation of various carbonyl derivatives with TTF hydrazone. The electron donating ability of these redox active ligands determined by cyclic voltammetry is described together with the investigations of their molecular structures by X-ray diffraction studies. The chelating ability of these ligands has been exemplified through the coordination to molybdenum carbonyl fragment or the complexation to difluoroboron moiety.     

Multi-enzyme anode composed of FAD-dependent and NAD-dependent enzymes with a single ruthenium polymer mediator for biofuel cells

A multi-enzyme electrode composed of FAD-dependent and NAD-dependent enzymes was fabricated using a poly-ruthenium complex (PAHA–Ru), which has two 1,10-phenanthroline-5,6-dione molecules as ligands. PAHA–Ru was used to immobilize FAD-dependent glucose dehydrogenase (FAD–GDH) onto an electrode and to examine PAHA–Ru containing the quinone moieties as an electron mediator. In cyclic voltammetry measurements of the FAD–GDH modified electrode in the presence of D-glucose, a catalytic current was obtained, which indicated electron transfer from FAD–GDH to PAHA–Ru. Our previous study has reported that PAHA–Ru with the quinone ligands also works as a mediator for NADH oxidation on an NAD-dependent alcohol dehydrogenase (NAD–ADH) modified electrode. Hence, FAD–GDH and NAD–ADH were co-immobilized with PAHA–Ru to make a multi-enzyme electrode. Using this multi-enzyme electrode as an anode, catalytic currents were observed in D-glucose solution, ethanol solution, and a mixed D-glucose and ethanol solution. The catalytic current in the mixed solution was greater than the currents obtained in the single substrate solutions, indicating bioelectrocatalysis reactions by the two enzymes and the single mediator in the mixed solution. Thus, we demonstrated that PAHA–Ru modified electrode enables selection of enzymes and their substrates from a wider range for enzymatic biofuel cells.     

Evaluation of Binding Between Electroactive Biotin Derivative and Streptavidin Immobilized on Chitin Film

Evaluation of the streptavidin‐biotin binding at the surface of chitin film was carried out with voltammetry. Immobilization of streptavidin was attempted to the protonated chitin film, based on an electrostatic interaction that hardly causes any change in the protein structure. The streptavidin‐biotin binding was estimated from changes in the electrode response of biotin labeled with an electroactive compound. Although the response of daunomycin as an electroactive compound did not change at an electrode covered with streptavidin/chitin film, the response of the labeled biotin decreased. This observation shows that streptavidin is immobilized on the chitin film and the biotin binds with immobilized streptavidin. Consequently, it was clear that the chitin film is useful as a reaction field for protein‐ligand binding. Generally, a binding event between protein and its ligand in the living body occurs on the cell surface. The electrochemical evaluation of protein‐ligand binding on a natural polysaccharide like chitin membrane surface is important.     

A metal-complex-tolerant CuAAC 'click' protocol exemplified through the preparation of homo- and mixed-metal-coordinated [2]rotaxanes

A series of mono- and bis-metallated [2]rotaxanes has been prepared using a CuAAC 'click' protocol that is compatible with metal-coordinated building blocks and ligands; the methodology provides a general means for appending a metal ion or complex to an organic scaffold via Cu(I)-catalysed 'click' chemistry, even when the molecule contains redox-active or kinetically labile metals or vacant ligand sites.     

Electrocatalytic Reduction of Chromium(VI) by Thionin: Electrochemical Properties and Mechanistic Study

The electrochemical properties of aqueous thionin (an electroactive water soluble dye) of pH 1–12 were investigated by cyclic voltammetry at a boron doped diamond(BDD) electrode. A well defined reversible redox couple was observed in acidic, neutral and alkaline solutions. The standard potential and kinetic parameters for thionin were obtained by fitting experimental cyclic voltammograms to those generated by the DigiSim program. The electrogenerated reduced form of thionin has been used as an efficient organic catalyst for the reduction of Cr(VI) at a BDD electrode immersed in aqueous media. The cyclic voltammetry measurements indicate that an electrocatalytic process occurs, where electrochemically generated thionin reduced species (Leucothionin) is oxidized by Cr(VI) back to the parent thionin species via a EC' reaction mechanism. The determination of catalytic rate constant (K_cat) was accomplished again by fitting experimental cyclic voltammograms with simulated ones.