Redox Tag Research Articles

Electrochemically Controlled ATRP for Cleavage-Based Electrochemical Detection of the Prostate-Specific Antigen at Femtomolar Level Concentrations.

As a single-chain glycoprotein with endopeptidase activity, the prostate-specific antigen (PSA) is valuable as an informative serum marker in diagnosing, staging, and prognosis of prostate cancer. In this report, an electrochemical biosensor based on the target-induced cleavage of a specific peptide substrate (PSA peptide) is designed for the highly selective detection of PSA at the femtomolar level, using electrochemically controlled atom transfer radical polymerization (eATRP) as a method for signal amplification. The PSA peptides, without free carboxyl sites, are attached to the gold surface via the N-terminal cysteine residue. The target-induced cleavage of PSA peptides results in the generation of carboxyl sites, to which the alkyl halide initiator α-bromophenylacetic acid (BPAA) is linked via the Zr(IV) linkers. Subsequently, the potentiostatic eATRP of ferrocenylmethyl methacrylate (FcMMA, as the monomer) leads to the surface-initiated grafting of high-density ferrocenyl polymers. As a result, a large amount of Fc redox tags can be recruited for signal amplification, through which the limit of detection (LOD) for PSA can be down to 3.2 fM. As the recognition element, the PSA peptide is easy to synthesize, chemically and thermally stable, and low-cost. Without the necessity of enzyme or nanoparticle labels, the eATRP-based amplification method is easy to operate and low-cost. Results also show that the cleavage-based electrochemical PSA biosensor is highly selective and applicable to PSA detection in complex biological samples. In view of these merits, the integration of the eATRP-based amplification method into cleavage-based recognition is believed to hold great promise for the electrochemical detection of PSA in clinical applications.

A reagentless electrochemical sensor for aflatoxin B1 with sensitive signal-on responses using aptamer with methylene blue label at specific internal thymine

Aptamer electrochemical sensors using immobilized aptamers with redox tag rely on the target binding-induced changes of current signal on electrode, offering advantages in operation convenience, no separation, rapidity, and sensitivity. Usually, the redox tag is placed on aptamer terminal, however, sometimes the terminal label may be insensitive to target-binding and fail to generate sensitive responses. The redox tag methylene blue (MB) labeled on different sites of aptamer may experience distinct changes in local environment, distance to electrode, or interactions with aptamer bases during affinity binding, which affect the current signal. Thus, it is possible to construct aptamer electrochemical sensors with sensitive and significant responses to targets by screening a series of sites (e.g., internal thymine T) of the aptamer and placing MB tag on a specific site of the aptamer. With this strategy, we successfully fabricated an electrochemical sensor on gold electrode for rapid, reagentless, and sensitive detection of aflatoxin B1 (AFB1), an important mycotoxin causing great health risks, by using a 26-mer DNA aptamer with MB on an internal T site (e.g., 18th T) and a thiol moiety at 5′ terminal. This sensor generated remarkable signal-on responses to AFB1, allowed a detection limit of 6 pM, and enabled detection of AFB1 in wine, milk and corn flour samples. This sensor can be well regenerated by rinsing with deionized water and reused, and shows good stability. This sensor and the demonstrated strategy are promising in wide applications.

Real-Time Electrochemical Measurement of N-Acetyl-ß-D-Glucosaminidase Activity, Using Redox-Tagged N-Acetyl-ß-D-Glucosaminide Hydrolysis as Proof-of-Principle

In-situ amperometry was used as a convenient non-optical tool for the assessment of the chitooligosaccharide-hydrolyzing activity of N-acetyl-$\beta $ -D- glucosaminidase from the marine Vibrio harveyi (VhGlcNAcase). Signal generation used a 3-mm-diameter boron-doped diamond (BDD) electrode for the anodic oxidation of 4-methylumbelliferone (4MU), which is released from the model substrate 4-methylumbelliferyl-N-acetyl-β-D-glucosaminide (GlcNAc-4MU) by enzymic hydrolysis. The recorded rise in 4MU electrode current reported the release of the redox label in real time and amperometric data analysis reproduced the Michaelis-Menten VhGlcNAcase activity plots derived from standard spectroscopic assays. Since it provides continuous readout, amperometry avoids the use of reaction-stopping chemicals and is also practical in turbid test solutions. Furthermore, the established electrochemical enzyme activity assay offers the chance to study novel substrate-attached redox tags and to explore assay miniaturization through implementation in microfluidic devices, enables portable assay development through the use of mini-potentiostats with smart phone/tablet connectivity and aids assay automation through combination with platforms for robotic electrochemistry.

A signal-on electrochemical aptasensor for rapid detection of aflatoxin B1 based on competition with complementary DNA

Aflatoxin B1 (AFB1) is the most toxic mycotoxin, causing harmful effects on human and animal health, and the rapid and sensitive detection of AFB1 is highly demanded. We developed a simple electrochemical aptasensor achieving rapid detection of aflatoxin B1 (AFB1). A short anti-AFB1 aptamer having a methylene blue (MB) redox tag at the 3′-end was immobilized on the surface of a gold electrode. In the absence of AFB1, a complementary DNA (cDNA) strand hybridized with the MB-labeled aptamer, causing MB apart from the electrode surface and low current of MB. In the presence of AFB1, AFB1 competed with the cDNA in the binding to the MB-labeled aptamer, and the aptamer-AFB1 binding caused formation of a hairpin structure, making the MB close to the electrode surface and current of MB increase. Under optimized conditions, we achieved detection of AFB1 over dynamic concentration range of 2 nM–4 μM by using this signal-on electrochemical aptasensor. This method only required a simple 5-min incubation of sample solution prior to rapid electrochemical sensing, more rapid than other electrochemical aptasensors. The sensor could be well regenerated and reused. This sensor allowed to detect AFB1 spiked in 20-fold diluted beer and 50-fold diluted white wine, respectively. It shows potential for detection of AFB1 in wide applications.

A Novel Target Receptor Design and Biosensing Platform Using RAFT Polymer-Based Molecular Templating

In this work, we propose a novel biosensing platform based on a single-chain molecularly templated polymer. We have utilized reversible addition-fragmentation chain transfer (RAFT) technique to synthesize a polymer-based target receptor consisting of N-isopropylacrylamide (NIPAM) as a backbone and methacrylic acid (MAA) and 4-vinylpyridine (4-VP) as functional monomers. In addition, vinylferrocene (VF) was incorporated into the polymer as a redox reporter. 4-nitrophenol (4-NP) was chosen as a model target for templating. The sensing platform was implemented by immobilizing the polymers on gold electrode surface, forming a brush-like polymer layer. As the target molecules bind to the polymer receptor, the polymer undergoes conformational change which can be detected via electrochemical methods. The proposed sensing platform is aimed to detect small species such as neurotransmitters and neurotoxins with high specificity. One of the major challenges with biochemical sensor is achieving selectivity in analyte detection and therefore, a highly selective target receptor is needed [1]. One strategy to achieving this goal is the technique known as molecular templating or molecularly imprinted polymers (MIP) [2] where the polymer receptors are synthesized in the presence of the ‘template’ molecule to capture its chemical and structural motifs. Traditionally, the molecularly imprinted polymers were highly crosslinked 3-dimensional structures with the target binding sites embedded within the bulk polymer. However, the challenge with this architecture is that the response time is slow (due to the analyte having to penetrate the polymer layer to find the receptors) and removing the bound target species is difficult, rendering this approach not suitable for real-time continuous monitoring applications. In our approach, we propose a single-chain templated polymer-based target receptor as a recognition element for chemical detection. We have chosen 4-nitrophenol (4-NP), one of the priority toxicants according to the US Environmental Protection Agency, as our model target analyte. The process for preparing the templated polymers is shown in Figure 1. Briefly, N-isopropylacrylamide (NIPAM), functional monomers, redox label, RAFT agent, initiator and the template monomers were added in a solvent for molecule assembly. Next, after removing the dissolved oxygen inside the mixed solution, the polymerization was initiated. Figure 2 illustrates the working principle of the proposed biosensing platform. After molecular assembly (a), The polymers are prepared via RAFT-polymerization (b) to ensure uniformity in the molecular weight, followed by removing the template from the polymer via dialysis (c). Once the templates are fully removed, the synthesized target receptors are attached onto the gold electrode, forming a brush-like monolayer assembly (d). During analyte detection, when the target binds to the molecular receptor, the conformation of the polymer changes which can be measured electrochemically using the redox tags (vinyl ferrocene) (e). The selective target rebinding is based on the weak interaction between the functional monomers in the polymer and 4-NP such as hydrogen bonding and the π-πstacking force (f). The immobilization of polymer on gold surface is confirmed using electrochemical impedance spectroscopy (EIS). As shown in Figure 3, the impedance is gradually increased with the incubation time, indicating successful attachment of the polymer on the gold surface. Figure 4 shows the process of determining the binding affinity of the templated polymers using gold nanoparticles (AuNPs) and UV-vis. Figure 5 (a) indicates that the MIP exhibits higher affinity toward 4-NP compared with the NIP. In Figure 5 (b), the selectivitymolecular templating is demonstrated by comparing wtih 3-nitrophenol (3-NP). In Figure 6, it is shown that, as the concentration increased, the charge transfer is promoted in the MIP due to the collapsing of the polymer upon target recognition. NIP, on the other hand, shows negligible response to the 4-NP, indicating no significant conformational change for the polymers. Figure 7 (a) summarizes the calibration curves for both MIP and NIP. The templated polymer-based sensor shows a linear response to 4-NP solution, indicating that the proposed target receptor and the sensing platform is capable of detecting 4-NP. Figure 7 (b) shows the calibration curves when the templated polymer-based sensor is exposed to either 4-NP or 3-NP, suggesting a promising specificity towards 4-NP. In summary, an electrochemical sensing platform has been successfully developed, and the selectivity of the 4-NP imprinted polymers has been confirmed. The results demonstrate that 4-NP imprinted polymer undergoes a conformation change during specific target recognition. We envision that the proposed sensing platform can potentially be used in other applications such as neurotransmitter detection and drug delivery monitoring.

Quantitative Electrochemical Analysis of Cathepsin Bactivity Using Carbon Nanofiber Nanoelectrode Arrays Withoptimized Peptide Substrate Length and Temperature

It is well known that the over-expression, increased activity and altered localization of many proteases are associated with tumor progression. Many clinical diagnoses target proteases as biomarkers. However, the quantitative analyses of the activity of tumor-related protease are challenging, because of the network interaction among the large set of proteases. Hence, there is a strong demand to develop a bioanalytical technique that can rapidly detect protease activities with high sensitivity and specificity. In this presentation, I will report an efficient electrochemical quantitative analyses method for detecting the cathepsin B activity by using a peptide-functionalized nanoelectrode array. The nanoelectrode array (NEA) was fabricated with vertically aligned carbon nanofibers (VACNFs), which serves as a unique electrochemical platform to enhance the protease detection. VACNFs of ~100-150 nm in diameter and ~5 mm in length are grown on chromium coated silicon wafer in uniform vertical alignment and are fully separated from each other to form a brush-like structure. With further processes to encapsulate the VACNFs in SiO2 by chemical vapor deposition (CVD) followed by mechanical polishing and reactive ion etching (RIE), ~100 nm long VACNF tips can be exposed above the SiO2 matrix, forming a stable NEA. By covalently attaching the exposed tip with proper peptide probes containing a ferrocene (Fc) redox tag at the distal, the VACNF NEA can be used to detect proteolysis with AC voltammetry. Such NEAs showed improved temporal resolution and reduced steric hindrance. As a result, reliable real-time proteolytic kinetics are recorded, from which the protease activity is quantitatively derived based on the heterogeneous Michalis-Menten model. In this work, we systematically investigated the cathepsin B activity with peptides of different length (including tetrapeptide, hexapeptide, and octapeptide) and at different temperatures (at 19.3, 29.5, 38.6, and 44.2 °C) to reveal their effects on the kinetic proteolysis rate by cathepsin B. The hexapeptide gave the highest proteolysis rate over tetrapeptide and octapeptide, as reflected by the smallest exponential decay time constant t. Meanwhile, 38.6 °C gave the highest proteolysis rate over the other 3 temperatures. At 38.6 °C, the optimized temperature, the limit of detection of cathepsin B activity and concentration has been determined to be 2.49 X 10-4 s-1 and 0.32 nM, respectively. Furthermore, the Fc-hexapeptide functionalized VACNF NEA shows very high selectivity, with essentially no measurable cross-reaction with 6.0 nM of other two cancer-related proteases, ADAM10 and ADAM17. This study demonstrates the potential to develop this method into a multiplex electronic chip for rapidly profiling the activity of up to 9 proteases simultaneously, which would facilitate accurate diagnoses of cancers and fast careening effective protease inhibitors as drug candidates. It would be a useful tool toward developing precision medicine.

TiO2 Photocatalysis in Aromatic "Redox Tag"-Guided Intermolecular Formal [2 + 2] Cycloadditions.

Since the pioneering work by Macmillan, Yoon, and Stephenson, homogeneous photoredox catalysis has occupied a central place in new reaction development in the field of organic chemistry. While heterogeneous semiconductor photocatalysis has also been studied extensively, it has generally been recognized as a redox option in inorganic chemistry where such "photocatalysis" is most often used to catalyze carbon-carbon bond cleavage and not in organic chemistry where bond formation is usually the focal point. Herein, we demonstrate that titanium dioxide photocatalysis is a powerful redox option to construct carbon-carbon bonds by using intermolecular formal [2 + 2] cycloadditions as models. Synergy between excited electrons and holes generated upon irradiation is expected to promote the overall net redox neutral process. Key for the successful application is the use of a lithium perchlorate/nitromethane electrolyte solution, which exhibits remarkable Lewis acidity to facilitate the reactions of carbon-centered radical cations with carbon nucleophiles. The reaction mechanism is reasonably understood based on both intermolecular and intramolecular single electron transfer regulated by an aromatic "redox tag". Most of the reactions were completed in less than 30 min even in aqueous and/or aerobic conditions without the need for sacrificial reducing or oxidizing substrates generally required for homogeneous photoredox catalysis.

TiO2 Photocatalysis in the Aromatic “Redox Tag”-Guided Intermolecular Formal [2 + 2] Cycloadditions

Light has the potential to serve as green and sustainable reagent to catalyze chemical reactions. Stemming from the pioneering work of Honda and Fujishima in 1972, titanium dioxide (TiO2) has occupied a central place in the research field of semiconductor photocatalysis. The excited electron/hole pair generated upon irradiation in aqueous aerobic conditions produces reactive oxygen species (ROSs) at the surface, which are account for subsequent redox transformations. They rapidly convert organic compounds into CO2 and H2O via carbon-carbon bond cleavage, therefore, semiconductor photocatalysis has generally been recognized as redox options in inorganic chemistry rather than organic chemistry where carbon-carbon bond formation is a focal point for extensive research efforts. While semiconductor photocatalysis is a heterogeneous process, a homogeneous alternative can be provided by the use of transition metal complexes, which is now well- known as “photoredox catalysis”. The early works by MacMillan, Yoon, and Stephonson have triggered the explosive growth in the general area of new reaction developments in this class. Arguably, heterogeneous semiconductor photocatalysis has taken a backseat to homogeneous photoredox catalysis in the field of organic chemistry. For TiO2 photocatalysis to be effective in carbon-carbon bond formation, the reactions must be carried out in dry anaerobic conditions in order not to produce ROSs that can damage organic compounds. We have been developing carbon-carbon bond formations by electrocatalysis in lithium perchlorate (LiClO4)/nitromethane (CH3NO2) electrolyte solution.1 Our reaction design involves the aromatic “redox tag” concept,2 that is, an aromatic ring that provides both the oxidant and the reductant in the overall net redox neutral carbon-carbon bond formations. In most cases, the use of LiClO4/CH3NO2 electrolyte solution is critical, which exhibits remarkable property as a Lewis acid to facilitate the reactions of carbon-centered radical cations with carbon nucleophiles, even unactivated alkenes. Our reaction development is also characterized by the importance of an intramolecular electron transfer process, which is recently presented by Melchiorre as an elegant “electron relay” mechanism. We anticipated that the use of LiClO4/CH3NO2 electrolyte solution would pave the way for carbon-carbon bond formations by TiO2 photocatalysis. It should also be noted that overall net redox neutral processes would be benefited from the involvement of both reductive and oxidative electron transfers potentially enabled by excited electron and hole. In this talk, the aromatic redox tag-guided intermolecular formal [2 + 2] cycloadditions3 by TiO2 photocatalysis will be discussed in details. (1) Imada, Y.; Yamaguchi, Y.; Shida, N.; Okada, Y.; Chiba, K. Chem. Commun. 2017, 53, 3960–3963. (2) (a) Okada, Y.; Chiba, K. Chem. Rev. 2018, accepted. (b) Okada, Y.; Yamaguchi, Y.; Ozaki, A.; Chiba, K. Chem. Sci. 2016, 7, 6387–6393. (c) Okada, Y.; Nishimoto, A.; Akaba, R.; Chiba, K. J. Org. Chem. 2011, 76, 3470–3476. (3) Chiba, K.; Miura, T.; Kim, S.; Kitano, Y.; Tada, M. J. Am. Chem. Soc. 2001, 123, 11314–11315. Figure 1

Electrochemical measurement of antibody-antigen recognition biophysics: Thermodynamics and kinetics of human chorionic gonadotropin (hCG) binding to redox-tagged antibodies

The thermodynamics and kinetics of antigen binding under diffusive conditions to an electrode surface modified with ferrocene-tagged antibodies is studied within this work, and realised experimentally for the case of human chorionic gonadotropin (hGC) as the antigen with monoclonal anti-hCG antibodies immobilised on an electrode surface via a molecular wire platform. The formation of the antigen-antibody complex is monitored through the blocking of the ferrocene voltammetry, thereby enabling the fractional coverage of the electrode binding sites to be unravelled as a function of time. It is found that, at low antigen concentrations, a Frumkin adsorption isotherm fits the data, with repulsive interactions between bound antigens playing a significant rôle, with an affinity constant that is an order of magnitude larger than in the case of an untagged antibody, suggesting that the chemical hydrophobicity of the redox tag may encourage stronger binding. Comparison of the experimental temporal data with relevant diffusion-adsorption models under activation control allows for the extraction of the kinetic parameters; at zero coverage, the rate constants for adsorption and desorption are, respectively, larger and smaller than the untagged antibody. The kinetic study enables the confirmation that this type of platform may be utilised for rapid (15 min) and quantitative electroimmunoassay. This is validated through proof-of-concept analytical measurements, yielding a limit of detection around 25 mIU mL−1 (corresponding to 2.7 ng mL−1) – a value used clinically for urine hCG measurements corresponding to around four weeks of gestational age.

Redox-Tag Processes: Intramolecular Electron Transfer and Its Broad Relationship to Redox Reactions in General.

Explosive growth in the use of open shell reactivity, including neutral radicals and radical ions, in the field of synthetic organic chemistry has been observed in the past decade, particularly since the advent of ruthenium complexes in 2008. These complexes generally induce single-electron transfer (SET) processes via visible-light absorption. Additionally, recent significant advancements in organic electrochemistry involving SET processes to provide open shell reactivity offer a complementary method to traditional polarity-driven reactions described by two-electron transfer processes. In this Review, we highlight the importance of intramolecular SET processes in the field of synthetic organic chemistry, which seem to be more elusive than the intermolecular versions, since they are net redox-neutral and thus cannot simply be regarded as oxidations or reductions. Such intramolecular SET processes can rationally be understood in combination with concomitant bond formations and/or cleavages, and are regulated by a structural motif that we call a "redox tag." In order to describe modern radical-driven reactions involving SET processes, we focus on a classical formalism in which electrons are treated as particles rather than waves, which offers a practical yet powerful approach to explain and/or predict synthetic outcomes.

Cascade toehold-mediated strand displacement along with non-enzymatic target recycling amplification for the electrochemical determination of the HIV-1 related gene

The authors describe a dual-signal electrochemical biosensor for highly sensitive determination of the HIV-1 related gene. This method is based on the application of cascaded toehold-mediated strand displacement reactions (TMSDRs) in combination with non-enzymatic target recycling amplification (TRA). A DNA machine with two TMSDRs was designed, and this resulted in reusable target and an output of two oligonucleotides, referred to as strand A (AS) labeled with the redox tag methylene blue (MB) and as untagged strand B (BS). A ferrocene (Fc)-modified signal probe (Fc-P1) is immobilized on the gold electrode surface by hybridizing with a thiolated probe (P2). The labeled AS causes the dissociation of Fc molecules and the gathering of MB molecules via strand displacement reaction. The target gene triggers TMSDRs and TRA. This leads to an increase in the distance changes between the redox tags and the gold electrode. The assay works in the 1 pM to 10 nM concentration range. On account of target recycling and dual recognition, the limit of detection is as low as 0.88 pM (at an S/N ratio of 3). The assay also has a remarkable selectivity which is ascribed to the use of both cascaded TMSDRs and dual recognition. In our perception, this assay represents a robust means of wide scope in that it may be applied to the detection of various kinds of nucleic acid even in complex samples.

Aptamer‐based Biosensor Developed to Monitor MUC1 Released by Prostate Cancer Cells

AbstractElectrochemical aptasensors can detect cancer biomarkers such as mucin 1 (MUC1) to provide point‐of‐care diagnosis that is low‐cost, specific and sensitive. Herein, a DNA hairpin containing MUC1 aptamer was thiolated, conjugated with methylene blue (MB) redox tag, and immobilized on a gold electrode by self‐assembly. The fabrication process was characterized by scanning electron microscopy, X‐ray spectroscopy analysis and electrochemistry techniques. The results evidenced a stable and sensitivity sensor presenting wide linear detection range (0.65–110 ng/mL). Therefore, it was able to precisely detect MUC1 production patterns in normal (RWPE‐1) and prostate cancer cells (LNCaP and PC3). The biosensor has ability to detect MUC1 in complex samples being an efficient and useful platform for cancer diagnosing in early stages and for physiological applications such as cancer treatment monitoring.

Integrated signal probe based aptasensor for dual-analyte detection

For the multi-analyte detection, although the sensitivity has commonly met the practical requirements, the reliability, reproducibility and stability need to be further improved. In this work, two different aptamer probes labeled with redox tags were used as signal probe1 (sP1) and signal probe2 (sP2), which were integrated into one unity DNA architecture to develop the integrated signal probe (ISP). Comparing with the conventional independent signal probes for the simultaneous multi-analyte detection, the proposed ISP was more reproducible and accurate. This can be due to that ISP in one DNA structure can ensure the completely same modification condition and an equal stoichiometric ratio between sP1 and sP2, and furthermore the cross interference between sP1 and sP2 can be successfully prevented by regulating the complementary position of sP1 and sP2. The ISP-based assay system would be a great progress for the dual-analyte detection. Combining with gold nanoparticles (AuNPs) signal amplification, the ISP/AuNPs-based aptasensor for the sensitive dual-analyte detection was explored. Based on DNA structural switching induced by targets binding to aptamer, the simultaneous dual-analyte detection was simply achieved by monitoring the electrochemical responses of methylene blue (MB) and ferrocene (Fc) This proposed detection system possesses such advantages as simplicity in design, easy operation, good reproducibility and accuracy, high sensitivity and selectivity, which indicates the excellent application of this aptasensor in the field of clinical diagnosis or other molecular sensors.

Radical Cation Diels-Alder By Electrocatalyst

In recent years, redox reactions with single electron transfer have received significant attention from many chemists. Organo-catalysts and photo-catalysts have been growing as a promising approach to electron path-ways. The electrochemical method is also playing roles as one of the most straightforward processes. With this method, we designed intermolecular carbon-carbon bond forming reactions, in which electron is the key reagent and the catalytic amount of electricity is enough for the full conversion. For example, radical cation Diels-Alder reaction with aromatic "redox tag" is in accordance with it. In this reaction, the chain cycle is the key factor to realize the significant decrease of consumed electricity. Our success leads to the development of the environmentally friendly and extreme high productivity synthetic method to produce the compound which has useful chemical structure for natural compounds and pharmaceuticals. Our investigation commenced with the reaction between trans-anethole (1) and isoprene (2), using electrocatalytic approach. In the reaction, it is understood well that the radical cation (3 ・ +) is generated, accompanying by chain process (Fig. 1). From cyclic voltammetry investigation, it is indicated that 1 (Ep ox = 1.07 V vs. Ag/AgCl) is anodically oxidized selectively in the presence of 2 (Ep ox = 1.83 V vs Ag/AgCl). Based on this, we demonstrate that the aromatic radical cation, which is given by the intermolecular [4 + 2] cycloaddition between radical cation 1 ・ + and 2, gives rise to a radical cation chain cycles and completion of the reaction with the catalytic amount of electricity. Here, we envisioned that the aromatic ring of 1might work as a redox tag to accomplish an electrocatalytic Diels-Alder reaction. Hence, we carried out anodic oxidation of 1in the presence of 2 equiv. of 2 at 1.00 V vs. Ag/AgCl. Beyond our expectation, only 0.1 F/mol of electricity was sufficient to make the reaction completed in excellent yield. This result suggests that the aromatic radical cation 3 ・ + was reduced predominantly by 1, comparing the reduction at the electrodes. Furthermore, the investigation of Cyclic Voltammetry shows that the oxidation peak of Diels-Alder adduct (3) is 1.40 V vs Ag/AgCl, which is sufficiently higher than that of 1. It can explain that radical cation 3 ・ + would oxidize 1 preferably, leading to the chain propagation. Based upon the mechanistic comprehension, we confirmed the applicable range of substrates, which are dienophile and diene, in the electrocatalytic Diels-Alder reaction. It showed that the moderate range of dienes was available, but the scope of styrene structures was limited. To obtain deeper insight into the mechanism, we conducted density functional theory (DFT) calculations. The calculation results demonstrate that the Dies-Alder reaction is proceed in stepwise mechanism. Besides, we speculated that the combination of electrolyte and solvent had an important role in this reaction. Although we have tried to use a variety of electrolytes and solvents, we have not found better media than LiClO4 / Nitromethane which leads this reaction to the excellent yield and faraday efficiency. Considering that using tetrabutylammonium perchlorate did not work in this reaction, we assume that Li+ has the important role. With the investigation of 7Li NMR and the heat of solution, it is implied that the solvation of ions is the key factor to achieve the electrocatalytic C-C bond forming reaction. Then, we turned out interest to how much we could improve the electro-energy efficiency. As we demonstrated above, only 0.1 F/mol, was enough to complete the reaction. However, we postulated that it still has room to be encouraged. We screened several factors; type of cell, type of electrodes, electrode’s surface area, reaction temperature, concentration of starting materials, concentration of electrolyte, etc… Surprisingly, it turned out that the concentration of substrates had the significant impact on the efficient production. Based upon the optimized condition, we have succeeded in that only 0.015 F/mol realized the full conversion. It is demonstrated by GC-MS monitoring that from 0.005 F/mol to 0.01 F/mol, the average number of chain cycle is 106. This optimized condition has the potential of having a big impact on chemical and pharmaceutical production. Our electrochemical approach realizes the environmentally friendly and extreme high productive synthetic method; requiring only a small amount of solvent and electrolyte, high electro-energy efficiency and recyclable. Taking advantage of these points, we demonstrate that we can scale up the production of the compounds that have useful chemical structure for natural compounds and pharmaceuticals. It is noted that electro chemical approach is promising for not only green but also economical chemistry. Figure 1

Thrombin aptasensor enabled by Pt nanoparticles-functionalized Co-based metal organic frameworks assisted electrochemical signal amplification

In this work, a Pt nanoparticles-functionalized Co-based metal organic frameworks (PtNPs@Co(II)MOFs@PtNPs) was synthesized and applied in electrochemical aptasensor for thrombin (TB) detection. First, the Co(II)MOFs@PtNPs were prepared via the mixed solvothermal method, which consists of inner Pt nanoparticles (PtNPs) encapsulated by aminofunctionalized Co(II)MOFs materials. Following that, additional PtNPs were adsorbed on the surface of Co(II)MOFs@PtNPs, resulting in the formation of PtNPs@Co(II)MOFs@PtNPs nanocomposite. The PtNPs@Co(II)MOFs@PtNPs nanocomposites with a large surface area were implimented as nanocarriers to immobilize a mass of TBA II for the formation of the TBA II bioconjugates that could be captured onto the electrode surface by sandwich-type format. Moreover, the PtNPs@Co(II)MOFs@PtNPs nanocomposites could directly use as redox tags for charge-generating and electron-transporting with the electron transfer from Co(II) to Co(III). Furthermore, in the presence of H2O2, the PtNPs@Co(II)MOF@PtNPs could effectively catalyze H2O2 oxidation with improvement electron transfer of redox probe, resulting in electrochemical signal amplification. Based on the above superior advantages, TB was determined in the concentration range from 0.1pM to 50nM with a detection limit of 0.33fM. Furthermore, the excellent sensitivity and selectivity can be easily established for quantitative analysis of other analytes.

Electrochemical detection of Nanog in cell extracts via target-induced resolution of an electrode-bound DNA pseudoknot

Nanog is among the most important indicators of cell pluripotency and self-renew, so detection of Nanog is critical for tumor assessment and monitoring of clinical prognosis. In this work, a novel method for Nanog detection is proposed by using electrochemical technique based on target-induced conformational change of an electrode-bound DNA pseudoknot. In the absence of Nanog, the rigid structure of the pseudoknot will minimize the connection between the redox tag and the electrode, thus reducing the obtained faradaic current. Nevertheless, the Nanog binding may liberate the flexible single-stranded element that transforms the DNA pesudokont into DNA hairpin structure due to steric hindrance effect, thus making the electrochemical tag close to the electrode surface. Consequently, electron transfer can be enhanced and very well electrochemical response can be observed. By using the proposed method, Nanog can be determined in a linear range from 2nM to 25nM with a detection limit of 163 pM. Furthermore, the proposed method can be directly used to assay Nanog not only in purified samples but also in complex media (cell extracts), which shows potential applications in Nanog functional studies as well as clinical diagnosis in the future.

Nanorod Aspect Ratios Determined by the Nano‐Impact Technique

AbstractThe in situ electrochemical sizing of individual gold nanorods is reported. Through the combination of electrochemical dissolution and the use of a surface‐bound redox tag, the volume and surface area of the nanorods are measured, and provide the aspect ratio and the size of the nanorods. Excellent independent agreement is found with electron microscopy analysis of the nanorods, establishing the application of nano‐impact experiments for the sizing of anisotropic nanomaterials.

Nanorod Aspect Ratios Determined by the Nano-Impact Technique.

The in situ electrochemical sizing of individual gold nanorods is reported. Through the combination of electrochemical dissolution and the use of a surface-bound redox tag, the volume and surface area of the nanorods are measured, and provide the aspect ratio and the size of the nanorods. Excellent independent agreement is found with electron microscopy analysis of the nanorods, establishing the application of nano-impact experiments for the sizing of anisotropic nanomaterials.

Electrocatalytic Diels-Alder Reactions Assisted By Aromatic Redox Tag

Although the Diels-Alder reaction is one of the most classic organic transformations, it is still in the front line of carbon-carbon bond formation toolbox and has been producing modern research topics in synthetic chemistry. Especially, the reaction mechanism has been studied intensively and also expanding to a wide variety of the combination of diene and dienophile. The reaction partners must be electronically matched, namely, electron-rich and electron-deficient substrates are required. In order to overcome this limitation, electron transfer strategy can be used. Typically, the reaction between both an electron-rich diene and dienophile are electronically mismatched, however, one electron oxidation by chemical oxidants or photosensitizers can produce a radical cation of one component, which is then trapped by the other component. We have been developing electrocatalytic intermolecular carbon-carbon bond formations in lithium perchlorate/nitromethane electrolyte solution. We have designed the reaction based on an aromatic “redox tag” concept, that is, the aromatic ring which provides both as an oxidant and a reductant in an overall electrochemical transformation. Since radicals, ions, and radical ions are all significantly stabilized by an aromatic ring and the redox potentials can be fine-tuned by the number and/or position of substituents, programmed electron transfer events could be achieved. These results in hand, we herein demonstrate electrocatalytic Diels-Alder reactions based on an aromatic redox tag strategy. Initially, we tried to optimize the reaction conditions by using trans-anethole and isoprene as a model reaction partner. These substrates are both relatively electron-rich, therefore, no reaction occurred without an electron transfer. In this context, we found that only a catalytic amount of electrical input was able to complete the reaction, even in high concentration conditions. We also found that the reaction partners were able to be designed by an expected manner by using an aromatic redox tag concept. In the presentation, the synthesis and electrochemical details will be discussed.

Dual-Target Electrochemical Biosensing Based on DNA Structural Switching on Gold Nanoparticle-Decorated MoS2 Nanosheets.

A MoS2-based electrochemical aptasensor has been developed for the simultaneous detection of thrombin and adenosine triphosphate (ATP) based on gold nanoparticles-decorated MoS2 (AuNPs-MoS2) nanocomposites. Two different aptamer probes labeled with redox tags were simultaneously immobilized on an AuNPs-MoS2 film modified electrode via Au-S bonds. The aptamers presented structural switches with the addition of target molecules (thrombin and ATP), resulting in methylene blue (MB) far from or ferrocene (Fc) close to the electrode surface. Therefore, a dual signaling detection strategy was developed, which featured both "signal-on" and "signal-off" elements in the detection system because of the target-induced structure switching. This proposed aptasensor could simultaneously determine ATP and thrombin as low as 0.74 nM ATP and 0.0012 nM thrombin with high selectivity, respectively. In addition, thrombin and ATP could act as inputs to activate an AND logic gate.