Detection Of Nicotinamide Adenine Dinucleotide Research Articles

Optimization of 1H-MRS methods for large-volume acquisition of low-concentration downfield resonances at 3 T and 7 T.

This goal of this study was to optimize spectrally selective 1H-MRS methods for large-volume acquisition of low-concentration metabolites with downfield resonances at 7 T and 3 T, with particular attention paid to detection of nicotinamide adenine dinucleotide (NAD+) and tryptophan. Spectrally selective excitation was used to avoid magnetization-transfer effects with water, and various sinc pulses were compared with a band-selective, uniform response, pure-phase (E-BURP) pulse. Localization using a single-slice selective pulse was compared with voxel-based localization that used three orthogonal refocusing pulses, and low bandwidth refocusing pulses were used to take advantage of the chemical shift displacement of water. A technique for water sideband removal was added, and a method of coil channel combination for large volumes was introduced. Proposed methods were compared qualitatively with previously reported techniques at 7 T. Sinc pulses resulted in reduced water signal excitation and improved spectral quality, with a symmetric, low bandwidth-time product pulse performing best. Single-slice localization allowed shorter TEs with large volumes, enhancing signal, whereas low-bandwidth slice-selective localization greatly reduced the observed water signal. Gradient cycling helped remove water sidebands, and frequency aligning and pruning individual channels narrowed spectral linewidths. High-quality brain spectra of NAD+ and tryptophan are shown in 4 subjects at 3 T. Improved spectral quality with higher downfield signal, shorter TE, lower nuisance signal, reduced artifacts, and narrower peaks was realized at 7 T. These methodological improvements allowed for previously unachievable detection of NAD+ and tryptophan in human brain at 3 T in under 5 min.

Highly-selective optical filter for NADH fluorescence detection in multiphoton microscopy

Colorectal cancer (CRC) is a pressing global health concern, emphasizing the need for early detection tools. In this study an optical filter for precise detection of nicotinamide adenine dinucleotide (NADH) fluorescence via two-photon excitation fluorescence (TPEF) was developed. Fabricated with silicon dioxide and titanium dioxide thin films in a Fabry-Perot structure, the filter achieved a peak transmittance of about 95% at 483 nm, with a 12 nm full-width at half maximum. TPEF measurements using a tailored setup and NADH liquid phantoms underscored the filter's significance in selectively capturing NADH fluorescence while mitigating interference from other fluorophores. This work marks a substantial stride towards integrating multiphoton microscopy into conventional colonoscopy, enabling non-invasive, objective optical biopsy for colorectal tissue analysis. Further refinements of the experimental setup are imperative to advance tissue differentiation and enhance CRC diagnosis.

Facile fabrication of electrochemically reduced graphene oxide/polythionine-methylene blue and its use as a platform for detection of nicotinamide adenine dinucleotide in the artificial urine sample

Here we report on a facile, rapid, selective, and stable electrochemical sensing platform for nicotinamide adenine dinucleotide (NADH) based on polythionine-methylene blue/electrochemically reduced graphene oxide modified glassy carbon electrode (PTH-MBDES-ERG/GCE). The ERG was directly formed on a glassy carbon electrode (ERG/GCE) through one-step electrochemical reduction of GO in PBS solution (10 mM, pH 7.4). In a PBS electrolyte, respectively, containing 50% (v/v) three different deep eutectic solvents (DES), thionine and methylene blue were polymerized on the surface of ERG/GCE. The electrochemical properties of the polymers polymerized in different electrolytes were investigated. The PTH-MBDES-ERG composite has a higher performance in the electrocatalytic oxidation of NADH than that of PTH-MBDES. The as-prepared PTH-MBDES-ERG/GCE demonstrated excellent direct electrocatalytic oxidation toward NADH, providing a favorable platform for electron transfer from NADH to the electrode. The sensor displayed a wide linear range from 1.52 μM to 3333.33 μM with a limit of detection of 0.51 nM. The interference from the ascorbic acid and uric acid was negligible. The prepared sensor was further tested for the determination of NADH in artificial urine samples, showing the PTH-MBDES-ERG/GCE formed by electropolymerization have promising biomedical applications.

RETRACTED: Covalent functionalization mediated anchoring of gold nanoparticles on graphene oxide surface: A unique approach for electrode preparation

Abstract Now a days, the electrochemical approach is employed in various technological applications ranging from sensing to electrochemical energy storage/conversion. However, the efficiency of the process strictly depends on the type of electrode material and process of electrode fabrication. Here, graphene oxide surface is functionalized using 3-(mercaptopropyl)-trimethoxy silane (MPTS) and attached to polycrystalline gold surface (AuE) at room temperature. Thereafter, a hybrid electrode (AuE/FGO-AuNP) has been fabricated by anchoring the gold nanoparticles (Au NPs) onto the functionalized surface by simple dipping into the colloidal Au NPs solution under stirring condition. After the systematic characterization, the fabrication of the hybrid electrode has been validated by collecting the cyclic voltammetry and Nyquist impedance in a redox system (ferrocyanide/ferricyanide solution). Interestingly, the AuE/FGO-AuNP shows an efficient sensing platform for the oxidative detection of Nicotinamide adenine dinucleotide (NADH), an important co-enzyme. From this preliminary observation, it has been presumed that this process of electrode fabrication will be a potential approach for future electrochemical applications.

Enhanced Electrochemiluminescence in a Microwell Bipolar Electrode Array Prepared with an Optical Fiber Bundle

AbstractThe creation of microwells on an electrode surface has been used to improve the electrode performance. Herein, we report the fabrication of a microwell bipolar electrode array (MBEA) by selectively etching a fiber core and coating a thin gold layer on one pole of optical fiber bundle. The remote electrochemiluminescence (ECL) sensing platform is developed with the combination of the MBEA and ECL, such that light emission is generated at the distal face of the optical fiber bundle and detected at its proximal face. The resulting microwells in the MBEA can enhance the ECL generation, and produce a bell‐like ECL emission, for which higher light intensity is measured in the center of the fiber core than around its periphery. Then, it is revealed in theoretical simulations that the parabolic‐shaped bottom in the microwell induces the strong overlapped diffusion fields in its center and promotes the generation of ECL there. Finally, the application of MBEA is demonstrated in the detection of nicotinamide adenine dinucleotide (NADH).

A trifunctional split dumbbell probe coupled with ligation-triggered isothermal rolling circle amplification for label-free and sensitive detection of nicotinamide adenine dinucleotide

The nicotinamide adenine dinucleotide (NAD+) is an important small biomolecule that participates in a variety of physiological functions, and it has been regarded as a potential biomarker for disease diagnosis and a promising target for disease treatment. The conventional methods for NAD+ assay often suffer from complicated procedures, expensive labeling, poor selectivity, and unsatisfactory sensitivity. Herein, we develop a label-free and sensitive method for NAD+ assay based on the integration of a trifunctional split dumbbell probe with ligation-triggered isothermal rolling circle amplification (RCA). We design a trifunctional split dumbbell probe that can act as a probe for NAD+ recognition, a template for RCA reaction, and a substrate for SYBR Green I binding. In the presence of target NAD+, it can serve as a cofactor to active E. coli DNA ligase which subsequently catalyzes the ligation of split dumbbell probe to form a circular template for RCA reaction, generating numerous dumbbell probe amplicons which can be easily and label-free monitored by using SYBR Green I as the fluorescent indicator. Due to the high fidelity of NAD+-dependent ligation and high amplification efficiency of RCA amplification, this method exhibits high sensitivity with a detection limit of 85.6 fM and good selectivity with the capability of discriminating target NAD+ from its analogs. Moreover, this method can be applied for accurate and sensitive detection of NAD+ in complex biological samples and cancer cells, holding great potential in NAD+-related biological researches and clinical diagnosis.

Dual-signal from sandwich structural sensing interface for NADH electrochemical sensitive detection

Sensitive and selective detection of nicotinamide adenine dinucleotide (NADH) is of great importance since it plays crucial roles in the living organism. In this work, a novel sandwich structural electrochemical sensor was fabricated by the electropolymerization of 3-aminophenylboronic acid (APBA) on reduced graphene oxide modified electrode for nicotinamide adenine dinucleotide (NADH) detection. Based on boronate affinity, NADH can be anchored on the sensing interface through the covalent bond between the cis-diol group of NADH and the boric acid of APBA. Moreover, due to the polyphenolic hydroxyl structure, the anchored NADH can further be bound with ferrocene boric acid (FcBA) in solution via the second boronate affinity. Thus, a dual-signal aroused from the oxidation peak currents of NADH and FcBA were employed for the detection of NADH with improved sensitivity, resulting in a wide linear range of 5.0 × 10−8 −1.0 × 10−5 mol/L and a low limit of detection of 2.6 × 10−8 mol/L (S/N = 3). In addition, double boronate affinity endowed the sensor with excellent selectivity to NADH. With the good repeatability and stability, the prepared sensor was employed to determine NADH in human blood serum samples with satisfactory results.

Electrochemical determination of glutamic pyruvic transaminase using a microfluidic chip

The activity of glutamic pyruvic transaminase (GPT) is an important clinical evidence for some acute diseases such as acute hepatopathy and myocardial infarction. Thus, there is a demand for rapid determination of GPT in small formats at point-of-need. Herein, we describe a novel method of electrochemical determination of GPT with microfluidic technique. GPT activity was indirectly determined via the electrochemical (EC) detection of nicotinamide adenine dinucleotide (NADH) produced from the GPT transdeamination reaction. A type of microfluidic chip was developed, in which a passive mixer comprising 100 sub-ribs and a three-electrode strip for EC were integrated. To verify the response to NADH, a series of NADH concentrations varying from 19 µM to 5 mM were calibrated with cyclic voltammetry within the microfluidic chip. And a linear relationship with R 2 0.9982 between the peak current and the concentration of NADH was obtained. Then, the GPT activity was determined using the chips containing and not containing a ribs-type mixer. And a linear relationship which contained two sections between the GPT activity and the peak current was obtained. The chip with a ribs-type mixer exhibited the sensitivity of 0.0341 μA U−1 L in the range of 10–50 U L−1 and 0.0236 μA U−1 L in the range of 50–250 U L−1. And the detection limit of the chip with a ribs-type mixer was 9.25 U L−1. The complete detection process of GPT activity within the microfluidic chip was realized, and the time-consuming problem was remarkably improved too.

Detection of cerebral NAD+ in humans at 7T.

To develop 1 H-based MR detection of nicotinamide adenine dinucleotide (NAD+ ) in the human brain at 7T and validate the 1 H results with NAD+ detection based on 31 P-MRS. 1 H-MR detection of NAD+ was achieved with a one-dimensional double-spin-echo method on a slice parallel to the surface coil transceiver. Perturbation of the water resonance was avoided through the use of frequency-selective excitation. 31 P-MR detection of NAD+ was performed with an unlocalized pulse-acquire sequence. Both 1 H- and 31 P-MRS allowed the detection of NAD+ signals on every subject in 16 min. Spectral fitting provided an NAD+ concentration of 107 ± 28 μM for 1 H-MRS and 367 ± 78 μM and 312 ± 65 μM for 31 P-MRS when uridine diphosphate glucose (UDPG) was excluded and included, respectively, as an overlapping signal. NAD+ detection by 1 H-MRS is a simple method that comes at the price of reduced NMR visibility. NAD+ detection by 31 P-MRS has near-complete NMR visibility, but it is complicated by spectral overlap with NADH and UDPG. Overall, the 1 H- and 31 P-MR methods both provide exciting opportunities to study NAD+ metabolism on human brain in vivo. © 2016 International Society for Magnetic Resonance in Medicine. Magn Reson Med 78:828-835, 2017. © 2016 International Society for Magnetic Resonance in Medicine.

Synergistic effect of pyrroloquinoline quinone and graphene nano-interface for facile fabrication of sensitive NADH biosensor

A self-assembly composite of graphene-pyrroloquinoline quinone (PQQ) was fabricated and modified on glassy carbon electrode (GCE) for sensitive detection of nicotinamide adenine dinucleotide (NADH). Chitosan (CTS) was applied to disperse graphene to form a stable robust film on GCE. A synergistic effect between PQQ and graphene was observed during the electrocatalytic oxidation of NADH, with about 260mV reduction in the oxidation potential and 2.5-fold increase in the oxidation current compared with those on the bare GCE. The electrochemical sensors based on the modified electrodes allowed the detection of NADH with a good linear dependence from 0.32 to 220µM with a high sensitivity of 0.421µAµM−1cm−2 and a low detection limit of 0.16µM (S/N=3). It could also eliminate the interference of electroactive substances like ascorbic acid (AA), uric acid, and dopamine and its derivatives. The outstanding performances of graphene-PQQ/CTS composite capable of improving the electrical conductivity and accelerating the electron transport suggested its promising applications for design of different graphene based composites used in electrochemical sensing and energy fields.

Amperometric Quantification of NADH Based on Graphene/Methylene Blue Nanocomposite Thin Films on Au(111)

Graphene (Gr)/methylene blue (MB) composites were successfully prepared and amperometric detection of nicotinamide adenine dinucleotide (NADH) was studied on the Gr/MB composite thin films modified Au(111) electrode. Gr sheets were fabricated by using two‐step electrochemical approaches: intercalation of sodium dodecyl sulfate (SDS) into graphite working electrode by electrooxidation and exfoliation of SDS‐intercalated graphite electrode by electrochemical reduction to obtain Gr/SDS suspension. In order to prepare Gr/MB composite structure, first SDS was removed from Gr/SDS suspension and then MB was added into Gr suspension. Gr/MB dispersion was ultrasonicated, vacuum‐filtered through a membrane, and then re‐dispersed into water. Gr/MB nanocomposite was supported on Au(111) substrates by drop‐casting of this composite dispersion. Characterization of Gr/MB composites was carried out by using scanning electron microscopy (SEM), electron dispersive spectroscopy (EDS), UV–Visible absorption spectroscopy, Fourier transform infrared spectroscopy (FTIR), and cyclic voltammetry techniques. Gr‐MB/Au(111) electrode, having an excellent electrocatalytic activity, was used for amperometric determination of NADH. The oxidation current density was linearly proportional to NADH concentration in the range between 1.0 to 264 μM. The sensitivity of the sensor was calculated as 0.316 µA µM−1 cm−2 with a detection limit of 0.3 µM at a signal‐to‐noise ratio of 3.0. POLYM. COMPOS., 38:E118–E127, 2017. © 2016 Society of Plastics Engineers

A label-free fluorescence strategy for selective detection of nicotinamide adenine dinucleotide based on a dumbbell-like probe with low background noise.

In this work we developed a novel label-free fluorescence sensing approach for the detection of nicotinamide adenine dinucleotide (NAD(+)) based on a dumbbell-like DNA probe designed for both ligation reaction and digestion reaction with low background noise. SYBR Green I (SG I), a double-helix dye, was chosen as the readout fluorescence signal. In the absence of NAD(+), the ligation reaction did not occur, but the probe was digested to mononucleotides after the addition of exonuclease I (Exo I) and exonuclease I (Exo III), resulting in a weak fluorescence intensity due to the weak interaction between SG I and mononucleotides. In the presence of NAD(+), the DNA probe was ligated by Escherichia coli DNA ligase, blocking the digestion by Exo I and Exo III. As a result, SG I was intercalated into the stem part of the DNA dumbbell probe and fluorescence enhancement was achieved. This method was simple in design, fast to operate, with good sensitivity and selectivity which could discriminate NAD(+) from its analogs.

Label-free detection of nicotinamide adenine dinucleotide based on ligation-triggered exonuclease III-assisted signal amplification

Schematic illustration of the Exo III-assisted amplification strategy for NAD+detection.

Ligation-triggered fluorescent silver nanoclusters system for the detection of nicotinamide adenine dinucleotide

Herein, we demonstrate a novel silver nanocluster-based fluorescent system for the detection of nicotinamide adenine dinucleotide (NAD(+)), an important biological small molecule involved in a wide range of biological processes. A single-stranded dumbbell DNA probe was designed and used for the assay, which contained a nick in the stem, a poly-cytosine nucleotide loop close to 5' end as the template for the formation of highly fluorescent silver nanoclusters (Ag NCs) and another loop close to 3' end. Only in the presence of NAD(+), the probe was linked at 5' and 3' ends by Escherichia coli DNA ligase, which blocked the DNA polymerase-based extension reaction, ensuring the formation of fluorescent Ag NCs. This technique provided a logarithmic linear relationship in the range of 1pM-500nM with a detection limit of as low as 1 pM NAD(+), and exhibited high selectivity against its analogues, and was then successfully used for the detection of NAD(+) level in four kinds of cell homogenates. In addition, this new approach was conducted in an isothermal and homogeneous condition without the need of any thermal cycling, washing, and separation steps, making it very simple. Overall, this label-free protocol offers a promising alternative for the detection of NAD(+), taking advantage of specificity, sensitivity, cost-efficiency, and simplicity.

A Simple Method to Tune Up Screen‐Printed Carbon Electrodes Applicable to the Design of Disposable Electrochemical Sensors

AbstractWe report an easy method to tune up screen‐printed carbon electrodes (SPCEs) for application in fabricating disposable electrochemical sensors. Simply by ultrasonic polishing a bare SPCE in a γ‐Al2O3 slurry, the surface roughness was drastically smoothed coupled with a large increase in hydrophilicity. The as‐generated micromorphology on the surface of the SPCE was found to be ideal for the immobilization of catechol to minimize the overpotential in the sensitive detection of nicotinamide adenine dinucleotide (NADH) and hydrazine. Physical characterization by both XPS and AFM studies specify that the adsorption behavior is related to the carbon surface functionalities and the trapping of γ‐Al2O3 on the polished‐SPCE.

Highly sensitive detection of NADH using electrochemiluminescent nanocomposites

Electrodes, modified with [Ru(bpy)2PVP10]n(ClO4)2 metallopolymer:gold nanoparticle composites, are used for the high sensitivity detection of nicotinamide adenine dinucleotide, NADH, using electrochemiluminescence, ECL; bpy is 2,2′-bipyridyl and PVP is poly (4-vinylpyridine). Incorporation of 12.5nm gold nanoparticles protected with 4-dimethylaminopyridine, DMAP-AuNPs, causes the overall ECL efficiency to decrease with increasing nanoparticle loading. However, the absolute emission intensity and signal-to-noise ratio are both higher for the nanocomposite than for the parent metallopolymer. The ECL intensity depends linearly on [NADH] from 10fM to 10μM with a limit of detection, LOD, of 5fM. The wide dynamic range and low limit of detection make these nanocomposite films attractive for biosensor development.

A label-free fluorescence DNA probe based on ligation reaction with quadruplex formation for highly sensitive and selective detection of nicotinamide adenine dinucleotide

A simple label-free fluorescent sensing scheme for sensitive and selective detection of nicotinamide adenine dinucleotide (NAD(+)) has been developed based on DNA ligation reaction with ligand-responsive quadruplex formation. This approach can detect 0.5 nM NAD(+) with high selectivity against other NAD(+) analogs.

Ultrasensitive and selective detection of nicotinamide adenine dinucleotide by target-triggered ligation–rolling circle amplification

An ultrasensitive fluorescence assay for nicotinamide adenine dinucleotide (NAD(+)) was developed by target-triggered ligation-rolling circle amplification (L-RCA). This novel approach can detect as low as 1 pM NAD(+), much lower than those of previously reported biosensors, and exhibits high discrimination ability even against 200 times excess of NAD(+) analogs.

Molecular Beacon Based Bioassay for Highly Sensitive and Selective Detection of Nicotinamide Adenine Dinucleotide and the Activity of Alanine Aminotransferase

We have developed a new approach to detect nicotinamide adenine dinucleotide (NAD(+)) with high specificity and sensitivity using molecular beacons (MBs) and employed it in the investigation of NAD(+) related biological processes, such as calorie restriction and alanine aminotransferase (ALT) activation. The E. coli DNA ligase would catalyze the ligation of two short oligonucleotides that complement with an MB only in the presence of NAD(+), resulting in the opening of the MB and the restoration of fluorescent signal. Thanks to the high sensitivity of the MB probe and the fidelity of E. coli DNA ligase toward its substrates, this approach can detect 0.3 nM NAD(+) with high selectivity against other NAD(+) analogs. This novel assay can also provide a convenient and robust way to analyze NAD(+) in biological samples such as cell lysate. As NAD(+) plays an essential role in many biochemical processes, this method can be used to investigate NAD(+) related life processes. For instance, the effect of calorie restriction on the intracellular NAD(+) level in MCF7 cells has been studied using this new assay. Moreover, this approach was also successfully used to analyze the activity of ALT. Therefore, this novel NAD(+) assay holds wide applicability as an analytical tool in biochemical and biomedical research.

Electrochemical detection of nicotinamide adenine dinucleotide based on molecular beacon-like DNA and E. coli DNA ligase

An electrochemical method for nicotinamide adenine dinucleotide (NAD +) detection with high sensitivity and selectivity has been developed by using molecular beacon (MB)-like DNA and Escherichia coli DNA ligase. In this method, MB-like DNA labeled with 5′-SH and 3′-biotin was self-assembled onto a gold electrode in its duplex form by means of facile gold–thiol chemistry, which resulted in blockage of electronic transmission. It was eT OFF state. In the presence of NAD +, E. coli DNA ligase was activated, and the two nucleotide fragments which were complementary to the loop of the MB-like DNA could be ligated by the NAD +-dependent E. coli DNA ligase. Hybridization of the ligated DNA with the MB-like DNA induced a large conformational change in this surface-confined DNA structure, which in turn pushed the biotin away from the electrode surface and made the electrons exchange freely with the electrode. Then the generated electrochemical signals can be measured by differential pulse voltammetry (DPV). Under optimized conditions, a linear response to logarithmic concentration of NAD + range from 3 nM to 5 μM and a detection limit of 1.8 nM were obtained. Furthermore, the proposed strategy had sufficient selectivity to discriminate NAD + from its analogues.