Validation and comparison of analytical methods for the determination of uric acid in pulses and cereals by salting out assisted extraction by Rapid resolution liquid chromatography.

An analytical method was developed and validated for the quantitative determination of uric acid in cereals and pulses based on salting-out assisted extraction and subsequent analysis by Rapid Resolution Liquid Chromatography (RRLC). Uric acid is a degradation product of purines, which is an indicator of insect infestation and the state of stored grains and pulses. This study aims to compare and validate a high-performance liquid chromatography method with Diode-array detection (HPLC-DAD) and fluorescence detection (HPLC-FLD) for the estimation of uric acid. Protein precipitation with ammonium sulfate and acetonitrile was used for sample cleanup and pre-treatment. The addition of inorganic ions results in preferential solvation and precipitates proteins. The separation was performed on a Zorbax SB C18 column (150 × 4.6 mm, 5 µm) with an isocratic elution using water-acetonitrile containing 10 mM sodium dihydrogen phosphate (95:5, v/v), at a flow rate of 0.5 mL/min. The relative coefficient (r2) for the calibration curve was more than 0.995 over the concentration range of 25-200 mg/kg. This method's precision at concentrations of 25-150 mg/kg was within 7.25%, and the accuracy was 85.1%-92.7%. The method was validated in terms of the LOD, LOQ, repeatability, reproducibility, linearity, uncertainty, specificity & system suitability. The limit of detection and limit of quantification was 16.60 mg/kg and 50.34 mg/kg, respectively.

Molecular Imprinting Technology for Determination of Uric Acid.

The review focuses on the overview of electrochemical sensors based on molecularly imprinted polymers (MIPs) for the determination of uric acid. The importance of robust and precise determination of uric acid is highlighted, a short description of the principles of molecular imprinting technology is presented, and advantages over the others affinity-based analytical methods are discussed. The review is mainly concerned with the electro-analytical methods like cyclic voltammetry, electrochemical impedance spectroscopy, amperometry, etc. Moreover, there are some scattered notes to the other electrochemistry-related analytical methods, which are capable of providing additional information and to solve some challenges that are not achievable using standard electrochemical methods. The significance of these overviewed methods is highlighted. The overview of the research that is employing MIPs imprinted with uric acid is mainly targeted to address these topics: (i) type of polymers, which are used to design uric acid imprint structures; (ii) types of working electrodes and/or other parts of signal transducing systems applied for the registration of analytical signal; (iii) the description of the uric acid extraction procedures applied for the design of final MIP-structure; (iv) advantages and disadvantages of electrochemical methods and other signal transducing methods used for the registration of the analytical signal; (vi) overview of types of interfering molecules, which were analyzed to evaluate the selectivity; (vi) comparison of analytical characteristics such as linear range, limits of detection and quantification, reusability, reproducibility, repeatability, and stability. Some insights in future development of uric acid sensors are discussed in this review.

Urate in fingernail represents the deposition of urate burden in gout patients.

Urate in the fingernails of gout patients and healthy volunteers was successfully detected by high-performance liquid chromatography (HPLC) with ultraviolet (UV) in our previous research. This study aimed to further investigate whether nail urate could be a proxy for the burden of monosodium urate (MSU) crystals deposits in gout. To this end, we conducted a study in two parts. Firstly, we successfully detected urate in the nail by HPLC-UV and evaluated nail urate concentrations in control subjects and patients with gout. As expected, we found that levels of nail urate were significantly higher in patients with gout than in healthy controls, and the nail urate level was significantly correlated with the volume of MSU crystals deposits measured by dual-energy CT (DECT). Secondly, we found that nail urate can reflect changes in urate levels in the body during urate lowering therapy through a 3-month follow-up study. Our results provide the possibility of quantification of urate in human fingernails as a non-invasive alternative for assessing MSU crystals deposits in gout.

Theacrine and strictinin, two major ingredients for the anti-influenza activity of Yunnan Kucha tea.

ETHNOPHARMACOLOGICAL RELEVANCE: Kucha tea plant (Camellia assamica var. kucha Chang et Wang) is regarded as a mutant variety of wild Pu'er tea plant found in few mountain areas of Yunnan, China. Its fresh young leaves and shoots are picked by the indigenous aborigines in these local areas to prepare an herbal tea for the treatment of common cold empirically. MATERIALS AND METHODS: Two extra compounds of relative abundance were detected in Kucha tea in comparison with Pu'er tea, and their chemical structures were identified as chlorogenic acid and theacrine. These two compounds as well as two major compounds, strictinin and caffeine, in Kucha tea were evaluated for their cytotoxicity and inhibitory effects on human influenza virus A/Puerto Rico/8/34 by analyzing viral protein expression and progeny production. RESULTS: No or low cytotoxicity was detected for the four Kucha compounds when their concentrations were below 100 µM. Expression of viral NS1 protein was significantly inhibited by chlorogenic acid, theacrine or strictinin, but not caffeine at a concentration of 100 µM. The relative inhibitory potency was detected as chlorogenic acid < theacrine < strictinin, and both theacrine and strictinin displayed significant inhibition at a concentration of 50 µM. According to a plaque assay, viral progeny production was significantly reduced by theacrine or strictinin, but not by chlorogenic acid or caffeine under the same concentration of 100 µM. CONCLUSION: It is suggested that theacrine and strictinin are two major ingredients responsible for the anti-influenza activity of Yunnan Kucha tea traditionally used for the treatment of common cold.

Construction of Kevlar nanofiber/graphene oxide composite beads as safe, self-anticoagulant, and highly efficient hemoperfusion adsorbents.

Recently emerged hemoperfusion absorbents, e.g. ion-exchange resin, activated carbon, and other porous materials, provide numerous novel possibilities to cure chronic liver failure (CLF) and renal failure (CRF). However, the limited adsorption performance and unsatisfactory blood compatibility significantly impede the development of the absorbents. Hence, designing safe and self-anticoagulant hemoperfusion absorbents with robust toxin clearance remains a considerable challenge. Here, brand new Kevlar-based composite gel beads for hemoperfusion are prepared by interface assembly based on π-π interaction. First, Kevlar nanofiber-graphene oxide (K-GO) beads are produced by liquid-liquid phase separation. Then, sodium p-styrenesulfonate (SS) is adsorbed onto the K-GO interface by π-π interaction and initiated to achieve the composite gel (K-GO/PSS) beads with an interfacial crosslinked structure. Such composite gel beads possess superior mechanical strength and self-anticoagulation capability, owing to the dual-network structure and heparin-mimicking gel structure, respectively. Furthermore, the K-GO/PSS beads show robust adsorption capacities for different kinds of toxins due to their strong charge and π-π interactions. A simulated hemoperfusion experiment in vitro demonstrates that the concentrations of the toxins in the blood can be restored to normal values within 30 minutes. In general, we envision that such composite gel beads will provide new strategies for future clinical CLF and CRF treatments.

Bimetallic nanoparticles decorated hollow nanoporous carbon framework as nanozyme biosensor for highly sensitive electrochemical sensing of uric acid.

An ultrasensitive electrochemical biosensor was developed to identify the low levels of uric acid (UA) in human serum. The gold/cobalt (Au/Co) bimetallic nanoparticles (NPs) decorated hollow nanoporous carbon framework (Au/Co@HNCF) was synthesized as a nanozyme by pyrolysis of the Au (III)-etching zeolitic imidazolate framework-67 (ZIF-67). The external Au NPs combined with internal Co NPs on the hollow carbon framework exhibited enhanced activity for UA oxidation, thereby generating superior signals. Accordingly, the Au/Co@HNCF biosensor presented ranking performances with a low detection limit of 0.023 µM (S/N = 3), an ultrahigh sensitivity of 48.4 µA µM-1 cm-2, and an extensive response in the linear region of 0.1-25 µM and the logarithmic region of 25-2500 µM. Owing to the ordered nanoporous framework and carbon interfacial features, the Au/Co@HNCF biosensor displayed adequate selectivity for UA sensing over a series of biomolecules. In addition, the Au/Co@HNCF biosensor was employed to quantify UA in human serum samples. The test results were basically consistent with those of a commercial apparatus, and thus demonstrated that the proposed Au/Co@HNCF biosensor was reliable for UA determination in clinical research.

In-situ reduction of Ag+ on black phosphorene and its NH2-MWCNT nanohybrid with high stability and dispersibility as nanozyme sensor for three ATP metabolites.

The environmental stability, water-processibility and life-span of black phosphorene (BP) severely limit the application of its electronic devices in aqueous system containing oxygen. We reported the controllable preparation of in-situ reduction and deposition of silver nanoparticles on the BP surface and its amino-functionalized multi-walled carbon nanotubes (NH2-MWCNT) nanocomposite. With the addition of both NH2-MWCNT and Ag+, the BP-based nanocomposite was prepared by ultrasonic-assisted liquid-phase exfoliation and was dispersed in carboxymethyl cellulose sodium (CMC) aqueous solution. The morphology, microstructure, and electrochemical properties of the nanohybrid were characterized. NH2-MWCNT-BP-AgNPs showed high environmental stability, good water-processibility, satisfactory life-spans, superior electrocatalytic capacity with enzyme-like kinetic characteristics. The nanohybrid was applied as electrochemical sensors for single/simultaneous analysis of uric acid (UA), xanthine (XT) and hypoxanthine (HX). Excellent voltammetric responses for simultaneous determination in linear ranges of 0.1-800 µM with a limit of detection (LOD) of 0.052 µM for UA, 0.5-680 µM with a LOD of 0.021 µM for XT, and 0.7-320 µM with a LOD of 0.025 µM for HX under optimal conditions. Besides, the developed nanozyme sensor was employed for simultaneous voltammetric analysis of UA, XT and HX in real samples with acceptable recoveries. This work will provide theoretical guidance and experimental support for the preparation and application of two-dimensional materials, nanozymes and sensing devices.

Simultaneous Voltammetric Determination of Acetaminophen, Ascorbic Acid and Uric Acid by Use of Integrated Array of Screen-Printed Electrodes and Chemometric Tools.

In the present work, ternary mixtures of Acetaminophen, Ascorbic acid and Uric acid were resolved using the Electronic tongue (ET) principle and Cyclic voltammetry (CV) technique. The screen-printed integrated electrode array having differentiated response for the three oxidizable compounds was formed by Graphite, Prussian blue (PB), Cobalt (II) phthalocyanine (CoPc) and Copper oxide (II) (CuO) ink-modified carbon electrodes. A set of samples, ranging from 0 to 500 µmol·L-1, was prepared, using a tilted (33) factorial design in order to build the quantitative response model. Subsequently, the model performance was evaluated with an external subset of samples defined randomly along the experimental domain. Partial Least Squares Regression (PLS) was employed to construct the quantitative model. Finally, the model successfully predicted the concentration of the three compounds with a normalized root mean square error (NRMSE) of 1.00 and 0.99 for the training and test subsets, respectively, and R2 ≥ 0.762 for the obtained vs. expected comparison graphs. In this way, a screen-printed integrated electrode platform can be successfully used for voltammetric ET applications.

An electrochemical biosensor based on novel butylamine capped CZTS nanoparticles immobilized by uricase for uric acid detection.

Quaternary chalcopyrite, i.e., Cu2ZnSnS4 (CZTS) nanoparticles films have been proposed as a novel matrix system for enzyme-based electrochemical biosensors providing a non-toxic, low-cost alternative for the fabrication of bioelectrodes. The easy tuneability of the band gap of CZTS by varying the cation ratio and size of nanoparticles facilitate to impart desirable electrical properties in the material. Butylamine capped spherical CZTS nanoparticles of size 15-16 nm and band gap 2.65 eV have been synthesized by colloidal hot injection technique. The films of CZTS onto ITO substrates are deposited using dip coating technique, and uricase enzyme have been immobilized onto CZTS films using EDC-NHS binding chemistry. Electrochemical analyses of this bioelectrode revealed that the uricase/CZTS/ITO/glass electrode exhibits good linearity over a wide range of 0-700 µM uric acid concentration with a limit of detection (LOD) of 0.066 µM. The low value of 0.13 × 10-4 M of Michaelis-Menten constant (Km) indicate the enhanced affinity of immobilized enzyme (uricase) towards uric acid. Thus, the present report confirms the promising application of the p-type CZTS thin film matrix for the realization of an electrochemical biosensor.

Hierarchical bi-continuous Pt decorated nanoporous Au-Sn alloy on carbon fiber paper for ascorbic acid, dopamine and uric acid simultaneous sensing.

In this work, Pt nanoparticles modified nanoporous AuSn(Pt@NP-AuSn) alloy on Ni buffered flexible carbon fiber paper (CFP) is fabricated by a simple replacement reaction in which NP-AuSn is fabricated by controllable dealloy of electrodeposited Au-Sn alloy films. The as prepared Pt@NP-AuSn/Ni/CFP possesses hierarchical pore structure, high specific surface area and excellent catalytic activity. Due to the bi-functions of both the large surface area of nanoporous metal and macroporous of carbon fiber paper facilitating mass transfer, the Pt@NP-AuSn/Ni/CFP shows high sensitivity of detecting ascorbic acid (AA), dopamine (DA) and uric acid (UA), with sensitivities of 0.14 µA µM-1 cm-2, 15.23 µA µM-1 cm-2, 0.28 µA µM-1 cm-2 under the concentration ranging from 200 to 2000 µM, 1-10 µM, and 25-800 µM for AA, DA and UA, respectively. Further, the Pt@NP-AuSn/Ni/CFP possesses long-term stability of sensing AA, DA and UA and presents great anti-interference towards a variety of common compounds in body fluid. All of these results manifest the Pt@NP-AuSn/Ni/CFP can be a promising candidate for the application of the electrochemical sensor for simultaneous detection of AA, DA and UA.

Simultaneous detection of ATP metabolites in human plasma and urine based on palladium nanoparticle and poly(bromocresol green) composite sensor.

A sensitive voltammetric sensor based on palladium nanoparticles (PdNPs) and poly-bromocresol green (pBG) composite layer immobilized on amide functionalized single-walled carbon nanotubes (AmSWCNTs) modified pyrolytic graphite (PdNPs:pBG/AmSWCNTs/PG) has been prepared for the simultaneous determination of adenosine triphosphate (ATP) catabolites, inosine (INO), hypoxanthine (HX), xanthine (XT), and uric acid (UA). The modified PdNPs:pBG/AmSWCNTs/PG was characterized by electrochemical experiments and surface analysis, which exhibited exceptional electrocatalytic effects towards the oxidation of INO, HX, XT, and UA with a significant enhanced peak current and well resolved peaks separation for all the analytes. The linear calibration curves were obtained in the concentration range of 0.001-175 µM, 0.001-200 µM, 0.001-150 µM, and 0.001-200 µM and limits of detection were found as 0.95 nM, 1.04 nM, 1.07 nM, and 0.43 nM corresponding to INO, HX, XT, and UA, respectively. The common metabolites present in the biological fluids did not interfere in the determination. The applicability of the proposed sensor was successfully demonstrated by determining INO, HX, XT, and UA in the human plasma and urine and the obtained results were validated by using HPLC.

Universal method for direct bioconjugation of electrode surfaces by fast enzymatic polymerization.

We report HRP-catalyzed polymerization of Tannic acid (TA) and application of the poly (Tannic acid) (p(TA)) as a versatile platform for covalent immobilization of biomolecules on various electrode surfaces based on electrochemical oxidation of the p(TA) and subsequent oxidative coupling reactions with the biomolecules. We also used this method for capturing cancer cells through a linker molecule, folic acid (FA). Furthermore, we have demonstrated that enhanced electrocatalytic activity of the p(TA)-modified surface could be used for simultaneous electrochemical determination of biologically important electroactive molecules such as ascorbic acid (AA), dopamine (DA), and uric acid (UA). This HRP-catalyzed polymerization of TA and p(TA)-mediated surface modification method can provide a simple and new framework to construct multifunctional platforms for covalent attachment of biomolecules and development of sensitive electrochemical sensing devices.

Identification of urate hydroperoxide in neutrophils: A novel pro-oxidant generated in inflammatory conditions.

Uric acid is the final product of purine metabolism in humans and is considered to be quantitatively the main antioxidant in plasma. In vitro studies showed that the oxidation of uric acid by peroxidases, in presence of superoxide, generates urate free radical and urate hydroperoxide. Urate hydroperoxide is a strong oxidant and might be a relevant intermediate in inflammatory conditions. However, the identification of urate hydroperoxide in cells and biological samples has been a challenge due to its high reactivity. By using mass spectrometry, we undoubtedly demonstrated the formation of urate hydroperoxide and its corresponding alcohol, hydroxyisourate during the respiratory burst in peripheral blood neutrophils and in human leukemic cells differentiated in neutrophils (dHL-60). The respiratory burst was induced by phorbol myristate acetate (PMA) and greatly increased oxygen consumption and superoxide production. Both oxygen consumption and superoxide production were further augmented by incubation with uric acid. Conversely, uric acid significantly decreased the levels of HOCl, probably because of the competition with chloride by the catalysis of myeloperoxidase. In spite of the decrease in HOCl, the overall oxidative status, measured by GSH/GSSG ratio, was augmented in the presence of uric acid. In summary, the present results support the formation of urate hydroperoxide, a novel oxidant in neutrophils oxidative burst. Urate hydroperoxide is a strong oxidant and alters the redox balance toward a pro-oxidative environment. The production of urate hydroperoxide in inflammatory conditions could explain, at least in part, the harmful effect associated to uric acid.

Graphene oxide reinforced core-shell structured Ag@Cu2O with tunable hierarchical morphologies and their morphology-dependent electrocatalytic properties for bio-sensing applications.

In this study, a facile solution approach was developed for the synthesis of a series of core-shell structured Ag@Cu2O nanocrystals of various shapes including triangles, spheres, and cubes with well-defined stable heterojunctions. The electrooxidation of dopamine (DA), uric acid (UA), guanine (G), and adenine (A) using these hybrids revealed morphology-dependent sensing properties, with activities and accumulation ability following the order, triangular Ag@Cu2O > spherical Ag@Cu2O > cubic Ag@Cu2O. Further, we constructed a novel graphene oxide (GO) nanosheet-reinforced triangular Ag@Cu2O ternary hetero-nanostructure. Such a hybrid with a three-dimensional interconnected hierarchical architecture is suitable for catalysis, since it not only leads to improved interfacial electron transfer, but also readily exposes the highly catalytic Ag@Cu2O to the reactants. Therefore, more enhanced electrochemical activities were observed for the oxidation of DA, UA, G, and A. This study provides an efficient way to synthesize morphology-controlled Ag@Cu2O heterogeneous catalysts for the fabrication of potential biosensors, and also opens up attractive avenues in the design of multifunctional ternary noble metal-semiconductor-carbon hybrids.

Portable electrochemical sensor based on 4-aminobenzoic acid-functionalized herringbone carbon nanotubes for the determination of ascorbic acid and uric acid in human fluids.

A new portable electrochemical sensor based on 4-aminobenzoic acid-modified herringbone carbon nanotubes (hCNTs-4ABA/Au-IDA) has been developed for the simultaneous determination of ascorbic acid (AA) and uric acid (UA) in physiological fluids. AA and UA were quantified by chronoamperometry at 0.1 and 0.32 V, respectively, in phosphate buffer solution (PBS 0.25 M, pH 7.0). Significant results were obtained for the separate quantification of AA and UA, with a limit of detection (LOD) of 0.65 µM for both analytes, and sensitivities of (9.0 ±â€¯0.4) A g-1 mM-1 and (8.8 ±â€¯0.3) A g-1 mM-1 for AA and UA, respectively. Repeatability was studied at 50 µM for AA and UA, providing relative standard deviations (RSD) lower than 9%. Additions of glucose, dopamine and epinephrine did not interfere with the AA and UA determination. Furthermore, UA did not interfere with AA determination at 0.1 V, although AA additions increased the current recorded at 0.32 V. The method has been successfully applied to human urine, perspiration and serum samples, without significant matrix effects, which allows for the use of an external calibration and the analysis of all the matrices investigated.

One-step selective screening of bioactive molecules in living cells using sulfur-doped microporous carbon.

A metal-free electrode using heteroatom-doped microporous carbon was fabricated for the ultrasensitive monitoring of mono-bioactive molecules and the selective signaling of dopamine (DA) secreted by living cells. The constructed electrode based on sulfur-doped microporous carbon (S-MC) shows a high surface area, a spherical construction, numerous carbon chain defects, and microporous structures, which are the key factors of the interactive signaling transducer, fast response, and active interfacial surfaces. The intrinsic features of S-MC with different %S-doping (S-MC-1, and S-MC-2) through the sp2-carbon chain create abundant catalytic active sites, facilitate molecular diffusion through the microporous structure, promote strong binding with the targeted molecules, and induce interactions at electrolyte-electrode interfaces. The S-MC-1 provides selective signaling in a tertiary mixture of DA, ascorbic acid (AA), and uric acid (UA) with a high sensitivity and a wide linear range of 0.01-5, 10-4000, and 1-2000 µM, respectively. The detection limits were set at 3 nM, 1.26 µM, and 0.23 µM for DA, AA, and UA respectively. The S-MC-1 demonstrated a selective screening of DA released from PC12 cells under a K+ ion- stimulator with high sensitivity and promoted high biocompatibility, low cytotoxicity, high stability, and reliable reproducibility (%RSD ranged from 1 to 2.7). Our findings indicated that the S-MC-1 can be utilized as an in-vitro model for simultaneously monitoring extracellular-DA secreted from living cells and sensing mono-bioactive molecules in biological samples.

Progress in utilisation of graphene for electrochemical biosensors.

This review discusses recent graphene (GR) electrochemical biosensor for accurate detection of biomolecules, including glucose, hydrogen peroxide, dopamine, ascorbic acid, uric acid, nicotinamide adenine dinucleotide, DNA, metals and immunosensor through effective immobilization of enzymes, including glucose oxidase, horseradish peroxidase, and haemoglobin. GR-based biosensors exhibited remarkable performance with high sensitivities, wide linear detection ranges, low detection limits, and long-term stabilities. Future challenges for the field include miniaturising biosensors and simplifying mass production are discussed.

Development of luminol-N-hydroxyphthalimide chemiluminescence system for highly selective and sensitive detection of superoxide dismutase, uric acid and Co2.

N-hydroxyphthalimide (NHPI), a well known reagent in organic synthesis and biochemical applications, has been developed as a stable and efficient chemiluminescence coreactant for the first time. It reacts with luminol much faster than N-hydroxysuccinimide, eliminating the need of a prereaction coil used in N-hydroxysuccinimide system. Without using prereaction coil, the chemiluminescence peak intensities of luminol-NHPI system are about 102 and 26 times greater than that of luminol-N-hydroxysuccinimide system and classical luminol-hydrogen peroxide system, respectively. The luminol-NHPI system achieves the highly sensitive detection of luminol (LOD = 70pM) and NHPI (LOD = 910nM). Based on their excellent quenching efficiencies, superoxide dismutase and uric acid are sensitively detected with LODs of 3ng/mL and 10pM, respectively. Co2+ is also detected a LOD of 30pM by its remarkable enhancing effect. Noteworthily, our method is at least 4 orders of magnitude more sensitive than previously reported uric acid detection methods, and can detect uric acid in human urine and Co2+ in tap and lake water real samples with excellent recoveries in the range of 96.35-102.70%. This luminol-NHPI system can be an important candidate for biochemical, clinical and environmental analysis.

Embroidered electrochemical sensors on gauze for rapid quantification of wound biomarkers.

Electrochemical sensors are an attractive platform for analytical measurements due to their high sensitivity, portability and fast response time. These attributes also make electrochemical sensors well suited for wearable applications which require excellent flexibility and durability. Towards this end, we have developed a robust electrochemical sensor on gauze via a unique embroidery fabrication process for quantitative measurements of wound biomarkers. For proof of principle, this biosensor was used to detect uric acid, a biomarker for wound severity and healing, in simulated wound fluid which exhibits high specificity, good linearly from 0 to 800µM, and excellent reproducibility. Continuous sensing of uric acid was also performed using this biosensor which reveals that it can generate consistent and accurate measurements for up to 7h. Experiments to evaluate the robustness of the embroidered gauze sensor demonstrate that it offers excellent resilience against mechanical stress and deformation, making it a promising wearable platform for assessing and monitoring wound status in situ.