Search

Introduction: Cicer arietinum L. is the choice of health food for people with diabetes, hypertension, and hyperlipidemia. As an essential source of high‐nutrition legumes, it is also an important source of dietary isoflavones. Objectives: In order to improve the preparation efficiency of natural plants, a rapid biological activity screening and preparation of xanthine oxidase inhibitors from C. arietinum L. was established. Methods: Xanthine oxidase (XOD) inhibitors were rapidly screened using ultrafiltration liquid chromatography–mass spectrometry (UF‐LC–MS) based on receptor–ligand affinity. The change in XOD activity was evaluated by enzymatic reaction kinetics measurement. The potential bioactive compounds were verified through molecular docking. In addition, the biological activity of ligands screened was separated and purified by complex chromatography. The structures of the compounds were identified by nuclear magnetic resonance spectroscopy. Results: Three active ingredients, namely daidzin, daidzein, calycosin with XOD binding affinities were identified and isolated from the raw plant materials via semi‐preparative high‐performance liquid chromatography (HPLC), 0–60 min, 5–50% B and countercurrent chromatography (CCC) (ethyl acetate/acetic acid/water [5:0.8:10, v/v/v]). Conclusion: This study will help to elucidate the mechanisms of action of natural plants of interest at the molecular level and could also provide more opportunities for the discovery and development of new nutritional value from other natural resources. A novel new techniques for extraction, biological activity screening, and preparation of XOD inhibitors from Cicer arietinum L. have been established. XOD inhibitors were rapidly screened using UF‐LC‐MS. Enzymatic reaction kinetics experiments have been performed to explore its mechanism of action. Meanwhile, the potential anti‐gout effects of the obtained bioactive compounds were verified using molecular docking. Next, three active ingredients, namely, daidzin, daidzein, and calycosin with XOD binding affinities, were identified and isolated by a complex chromatography (semi‐preparative HPLC and CCC). [ABSTRACT FROM AUTHOR]

A new micro‐kinetic model of the enzyme‐catalyzed synthesis of fructo‐oligosaccharides (FOS) was developed. A commercial enzyme mixture Pectinex® Ultra SP‐L derived from Aspergillus aculeatus was used. A variety of initial enzyme concentrations (1–5 vol%) and sucrose concentrations (400–600 g/L) were experimentally investigated and included in kinetic modeling. Several variations of kinetic mechanisms and corresponding models have been examined. A hybrid genetic algorithm was used to predict the kinetic parameters simultaneously for all experimental data. The best fitting model has been adopted, and with an average error of 13.34%, it describes the experimental data very well. The influence of initial concentrations on the conversion of sucrose and production of FOS is being carefully investigated. It was shown that the initial sucrose concentration significantly affects the highest level of FOS concentration, but the enzyme concentration controls the time at which maximum is reached as well as the rate of FOS decomposition. [ABSTRACT FROM AUTHOR]

Inspired by the dissipative assembly in biological systems, transient hydrogels based on supramolecular interactions have been developed that are under thermodynamic nonequilibrium states. Host–guest interactions possess excellent properties including high selectivity and adjustable association constants, which are beneficial for controlling the properties and behaviors of transient colloidal materials. In this work, a host‐fueled transient supramolecular hydrogel system is reported. The hydrogels based on host–guest interactions are formed by addition of a chemical fuel, α‐cyclodextrin (α‐CD), to the aqueous solutions containing Pluronic F127 and α‐amylase. Meanwhile, as the host molecule of α‐CD consumes, the hydrogel networks start to collapse. The lifetime of the transient supramolecular hydrogels can be precisely controlled by adjusting the temperature and hydrogel composition, and repeated sol‐to‐gel‐to‐sol transitions can be realized by refueling the system with α‐CD. This study provides a new approach to regulate the nonequilibrium host–guest inclusion system by fueling it with host molecules. [ABSTRACT FROM AUTHOR]

Lack of long‐time stability of dsDNA‐based supramolecular assemblies is an important issue that hinders their applications. In this work, 20 base pairs long dsDNA fragments [(dCdG)20‐65 %] composed of 65 % dCdG and 35 % dAdT nucleotides were tethered via a thiol to the surface of a gold electrode. The self‐assembled (dCdG)20‐65 % monolayer was immersed in solutions containing ectoine, a compatible solute. Electrochemical results showed that these monolayers were stable for one month. In situ IR spectroscopy indicated that ectoine interacts weakly with the phosphate‐ribose backbone, dehydrating the phosphate groups and stabilizing the A‐DNA conformation. This structural reorganization led to a reorientation of nucleic acid base pairs and a local disruption of the double‐helix structure. However, the conformation and orientation of the dsDNA fragment was stable in the −0.4

The present work deals with fabrication and characterization of the zinc oxide (ZnO) nanowire based novel two‐electrode capacitive biosensors on flexible Polyethylene terephthalate (PET) substrates for accurate estimation of glucose by analyzing the fundamental dielectric nature of the relevant sample. The morphology and crystalline quality of grown nanowires are analyzed by using field‐emission scanning electron microscope (FESEM) and X‐ray diffractometer (XRD), respectively. Current and capacitance values of the device have been studied for ten different glucose concentrations relevant to the physiological standards. The analytical performance of the devices in terms of enzyme activity, reliability and flexibility has also been evaluated. [ABSTRACT FROM AUTHOR]

Aroma is an important factor affecting mushroom character and quality. According to the different reaction pathway, the key aroma metabolites (sulfur and eight‐carbon volatiles) formation can be classified into enzymatic reactions and nonenzymatic reactions. Aroma volatiles are generated from precursors via the biocatalytic activities of various synthases during the growth stages of shiitake mushrooms. Understanding the specific relationships between the key aroma metabolites and their synthases is key to improving shiitake mushroom quality. At the same time, to reduce forest logging and burning of agricultural by‐products in farmland, agricultural by‐products have been applied to shiitake mushroom cultivation. Nevertheless, how to further improve the production of aroma volatiles in mushroom cultivated with agricultural waste is still a challenge. In order to understand the biosynthesis of volatiles via enzymatic reactions and screen the agricultural by‐products that can improve the production of aroma volatiles in mushroom cultivation, the mechanism of producing aroma volatiles needs to be further elucidated. In this study, the activities and gene expression levels of the key synthases involved in volatile metabolism, the contents of key aroma volatiles, and the correlations between related synthetase, volatiles, and cultivation substrate (CS) were investigated. Network models for visualizing the links between synthetase, volatiles, and CSs were built through partial least squares (PLS) regression analysis. The correlation coefficients among three related synthetase and enzymatic gene expression were high, and the combined effects of multiple synthetase promoted the production of volatiles. PLS analysis showed that the corncob and corn meal were more related to the production of volatiles and synthetase gene expression, and they can be added to the CSs as flavor promoting substances. The enrichment of key aroma volatiles in shiitake mushroom cultivated by the gradient of 20% corn meal combination CS was noticeable. [ABSTRACT FROM AUTHOR]

To ensure sustainable consumption and production patterns, alternative process design without plastics for chemical and biological reactions will benefit future generations. Reaction flasks used in chemical and biological laboratories have been made from glass, metals, and plastics so far. If containerless processing can be realized, researchers will have a next‐generation reaction process, which will be reactor and plastic‐free, and without risks of unforeseen issues induced by contact with reactions flasks, including contamination and alteration of the reactants. Here, polymerization, click chemistry, and enzymatic reactions can proceed effectively in a floating droplet at a node of standing wave generated by ultrasonic levitation. These results demonstrate that floating droplets levitated by acoustic waves can represent a revolutionary containerless reactor for performing various reactions in the fields of chemistry and biology. [ABSTRACT FROM AUTHOR]

The versatile design of stimuli‐responsive microparticles embedding valuable biomolecules has great potential in a variety of engineering fields, such as sensors, actuators, drug delivery, and catalysis. Here, results are reported on thermoresponsive core–gap–shell (TCGS) microcapsules made of poly(N‐isopropylacrylamide) (PNIPAm), which encapsulate hydrophilic payloads in a simple and stable manner. These are realized by a one‐step microfluidic approach using the phase separation of a supersaturated aqueous solution of NIPAm. Various designs of the microcapsules are achieved by individual control of the swelling or by incorporating pH‐responsive comonomers of the inner core and outer shell. The gap, i.e., the space between the inner core and outer shell, can be loaded with cargo‐like nanoparticles. The outer shell can serve as a stimuli‐responsive gateway for the transport of smaller molecules from the external solution. It is shown that the TCGS microcapsules are suitable as temperature controllable glucose sensors and hold promise in the design of controllable enzymatic reactions. The proposed platform provides an avenue for developing a new‐generation of microparticles for diverse and efficient engineering applications. [ABSTRACT FROM AUTHOR]

The development of a nanographite (NG)‐based fluorescent biosensor for detecting microRNA (miRNA) is reported. Duplex‐specific nuclease (DSN)‐assisted signal amplification was key to its function. In the absence of a target, with the assistance of p‐stacking interactions, the NG adsorbed the double carboxyfluorescein (FAM)‐labelled probe (DFP) whose surface was perfectly complementary to miRNA, leading to quenching of FAM fluorescence. In the presence of a target, double‐stranded DNA/RNA hybrids were repelled by the NG and fluorescence was restored. Meanwhile, the considerable increase in signal strength and sensitivity suggests DSN‐mediated target recycling as an application. The detection limit of the proposed biosensor for miRNA was 10 pmol/L; there was a linear correlation when the miRNA concentration ranged from 50 pmol/L to 5 nmol/L. Additionally, the method could distinguish let‐7b from most let‐7 miRNA family members and was successfully used in a sample assay. This biosensor is a novel and highly sensitive tool for miRNA detection and has great potential for biochemical research, disease diagnosis, and therapy. [ABSTRACT FROM AUTHOR]

The new synthetic route to α‐acyloxy amides based on Passerini multicomponent reaction followed by transformation into the corresponding five‐, six‐, and seven‐membered 3‐hydroxy‐lactams has been proposed. The influence of the reaction conditions, which includes substrate structure on the reaction course that led to desired lactams, was studied. The use of various oxo components in the Passerini multicomponent reaction (P‐MCR) has been evaluated. The first example of a semicyclic O,O‐acetal employed as an aldehyde equivalent in the P‐MCR has been reported. Finally, the established protocol was successfully applied for the enantioselective synthesis of 3‐hydroxy‐lactams, which play a crucial role in the synthesis of the number of bioactive compounds. [ABSTRACT FROM AUTHOR]

During the last decade, we have revealed biosynthetic pathways responsible for the formation of important and chemically complex natural products isolated from various organisms through genetic manipulation. Detailed in vivo and in vitro characterizations enabled elucidation of unexpected mechanisms of secondary metabolite biosynthesis. This personal account focuses on our recent efforts in identifying the genes responsible for the biosynthesis of spirotryprostatin, aspoquinolone, Sch 210972, pyranonigrin, fumagillin and pseurotin. We exploit heterologous reconstitution of biosynthetic pathways of interest in our study. In particular, extensive involvement of oxidation reactions is discussed. Heterologous hosts employed here are Saccharomyces cerevisiae, Aspergillus nidulans and A. niger that can also be used to prepare biosynthetic intermediates and product analogs by engineering the biosynthetic pathways using the knowledge obtained by detailed characterizations of the enzymes. (998 char.) [ABSTRACT FROM AUTHOR]

BACKGROUND: Activation and stabilization of enzymes have received considerable attention. In recent years, chemical modification has re-emerged as a powerful complementary approach to site-directed mutagenesis and directed evolution because it is simple and effective. Chemical modification of enzymes by dicarboxylic acid anhydride enhanced their activities and stabilities. RESULTS: Lipase, α-chymotrypsin and subtilisin modified by citraconic anhydride were examined. Lipase and α-chymotrypsin were successfully modified by citraconic anhydride, while subtilisin showed poor reactivity with it. Optimum pH range for lipase and α-chymotrypsin, and the secondary structure of α-chymotrypsin was found to be unchanged by the modification process. Effect of the modification on the activity and thermostability of lipase was small. On the other hand, modification of lysine residues on the enzyme surface enhanced the catalytic activity and thermostability of α-chymotrypsin. Further modification of lysine residues caused the reduction of thermostability. Modification of more hydrophobic dicarboxylic acid anhydride resulted in an increase in thermostability. CONCLUSION: The modification of the lysine residues on the enzyme surface with citraconic anhydride shows a small effect on the catalytic reactivity and enhances the thermostability of the enzymes. [ABSTRACT FROM AUTHOR]

The present study investigated the health effect of two types of diacylglycerol (DG) produced from (i) palm kernel (PK) oil of medium-chain saturated fatty acids and (ii) soy-canola oil (SC) blend of long-chain unsaturated fatty acids in C57BL/6N mice. As compared to diet containing 30% PK triacylglycerol (TG), 16 wk feeding trial on C57BL/6N mice with a diet consisting of 30% PKDG and 30% SCDG significantly ( p<0.05) reduced the fat accumulation in epididymal and retroperitoneal regions. Serum glucose, cholesterol, leptin and insulin levels were significantly ( p<0.05) suppressed in PKDG- and SCDG-fed mice. In terms of gene expression, PKDG diet induced expression of acyl-CoA synthase long chain (ACSL) and acyl-CoA synthase medium chain mRNA in the small intestine of the mice while SCDG fed mice upregulated ACSL in liver and small intestine. This suggests that the difference in fatty acid composition of DG may potentially induce β-oxidation in different organs in mice. Besides, expression of apolipoprotein B mRNA were reduced in mice fed with PKDG and SCDG, indicating the ability of PKDG and SCDG to reduce LDL levels. In conclusion, structural differences between DG and TG markedly influenced the metabolism of lipids in the body while fatty acid composition has only showed a minor effect. Practical applications: Medium chain type of DG oil, if proven to have similar effects to long chain type DG in this study, can be incorporated into food for health purposes and can replace conventional cooking oil as healthy functional cooking oil. [ABSTRACT FROM AUTHOR]

We propose the fabrication of nanostructured glassy carbon (GC) electrodes modified with horseradish peroxidase (HRP) for the detection of paracetamol. This was accomplished by inducing the nanostructuration of GC via the adsorption of polystyrene nanospheres (900 nm diameter) followed by electropolymerization of N-(10-azidodecyl)pyrrole. The nanospheres were then removed and nanostructured polypyrrole-GC was submitted to click reaction in presence of ethynyl-biotin that was further coupled to HRP-avidin. The electrode was then used to sense the electrochemical reduction of the enzymatically generated electroactive oxidized species of acetaminophen (NAPQI) in the presence of hydrogen peroxide. The nanostructured electrode with HRP exhibits a fast response towards NAPQI reduction and improved performances in terms of sensitivity and limit of detection compared to non structured electrode. [ABSTRACT FROM AUTHOR]

Time-temperature indicators (TTIs) are devices with a time-temperature-dependent change to reflect food quality status and temperature history. There is increasing interest in the application of TTIs to monitor food cold chain logistics for food safety and quality. In this paper, a new enzymatic TTI based on enzyme reaction, diffusion and enzyme demobilization technology is developed. The mathematical model of TTI is drawn up, which shows the relationships between the changes of time and temperature and the diffusion length of the indicator's colour changed. To assess the reliability of TTI as the quality indicating device for perishable foods in the process of storage and transportation, fresh milk is studied as an example. Through the research of dynamic characteristics of milk and TTIs, five different TTIs that matched with the activation energy of the prototype milk were designed and tested in several constant temperature profiles and a variable temperature profile. By comparing the end response value of the prototype TTI with the effect of forecasting the shelf life of milk, it was considered that the enzyme reaction-diffusion TTI has good stability and reliability at dynamic storage conditions and could be used to monitor the shelf life of perishable food during the cold chain transportation. Copyright © 2012 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Enzymatic hydrogelation is a totally different process to the heating-cooling gelation process, in which the precursors of the gelators can be involved during the formation of self-assembled structures. Using thixotropic hydrogels formed by a super gelator as our studied system, we demonstrated that the enzyme concentration/conversion rate of enzymatic reaction had a strong influence on the morphology of resulting self-assembled nanostructures and the property of resulting hydrogels. The principle demonstrated in this study not only helps to understand and elucidate the phenomenon of self-assembly triggered by enzymes in biological systems, but also offers a unique methodology to control the morphology of self-assembled structures for specific applications such as controlled drug release. [ABSTRACT FROM AUTHOR]

1-Octen-3-ol and 10-oxo- trans-8-decenoic acid are metabolites of the breakdown of linoleic acid (LA) by mushroom enzymes. These compounds can be produced in a bioreactor using a crude mushroom homogenate and the exogenous addition of LA and oxygen. The factors' duration of blending, mushroom–buffer ratio, effect of a surfactant, whole against partially clarified reaction broths, purity of LA, and utilization of stumps instead of whole mushrooms were studied for their effect on reaction yield using a 1-L bioreactor. The results showed the feasibility of using the more inexpensive 60%-pure LA instead of the 99%-pure LA even when a yield loss was involved. Waste stumps could be used instead of whole mushrooms with a yield decline of 26%. [ABSTRACT FROM AUTHOR]

The hydrogen-bond network in various stages of the enzymatic reaction catalyzed by HIV-1 protease was studied through quantum-classical molecular dynamics simulations. The approximate valence bond method was applied to the active site atoms participating directly in the rearrangement of chemical bonds. The rest of the protein with explicit solvent was treated with a classical molecular mechanics model. Two possible mechanisms were studied, general-acid/general-base (GA/GB) with Asp 25 protonated at the inner oxygen, and a direct nucleophilic attack by Asp 25. Strong hydrogen bonds leading to spontaneous proton transfers were observed in both reaction paths. A single-well hydrogen bond was formed between the peptide nitrogen and outer oxygen of Asp 125. The proton was diffusely distributed with an average central position and transferred back and forth on a picosecond scale. In both mechanisms, this interaction helped change the peptide-bond hybridization, increased the partial charge on peptidyl carbon, and in the GA/GB mechanism, helped deprotonate the water molecule. The inner oxygens of the aspartic dyad formed a low-barrier, but asymmetric hydrogen bond; the proton was not positioned midway and made a slightly elongated covalent bond, transferring from one to the other aspartate. In the GA/GB mechanism both aspartates may help deprotonate the water molecule. We observed the breakage of the peptide bond and found that the protonation of the peptidyl amine group was essential for the peptide-bond cleavage. In studies of the direct nucleophilic mechanism, the peptide carbon of the substrate and oxygen of Asp 25 approached as close as 2.3 Å. [ABSTRACT FROM AUTHOR]

Our results demonstrate that in monolayer films, the addition of compatible solutes, such as ectoine, to the electrolyte solution may have a protective effect on dsDNA stability over time. Keywords: DNA; self-assembled monolayer; ectoine; in situ spectroelectrochemistry; enzymatic reaction EN DNA self-assembled monolayer ectoine in situ spectroelectrochemistry enzymatic reaction 3780 3780 1 10/29/21 20211013 NES 211013 B The Cover Feature b shows double strand DNA monolayers on the gold electrode surface stored for a long time in aqueous solutions in the absence and presence of ectoine, compatible solute. [Extracted from the article]

Photoelectrochemical (PEC) bioassay system based upon cathodic measurement of enzymatic product hydrogen peroxide (H2O2) is highly attractive for accurate analyte detection as it can naturally avoid interference signals arising from readily oxidizable species in biological solutions. However, its practical utilization is restricted due to the similar reduction potential of H2O2 and oxygen, whereas solution oxygen levels can fluctuate significantly. To this end, this restriction is addressed via developing a novel PEC bioassay system containing a bio‐cathode with a solid–liquid–air triphase interface constructed by assembling an oxidase layer on the surface of superhydrophobic single‐crystal TiO2 nanowire arrays. The triphase bio‐cathode enables oxygen rapidly diffusing from the air phase to the bioassay reaction zone, thereby providing a constant interfacial oxygen concentration. The cathodic measurement enables PEC bioassay system exhibiting remarkably high detection selectivity, and compared with a normal diphase solid–liquid PEC bioassay system a 100‐fold extended linear detection range. The rapid electron transport in the single‐crystal TiO2 nanowire arrays endows the bioassay system an 845‐fold enhanced sensitivity, and three orders of magnitude lower minimum detection limit compared with TiO2 nanoparticle–based ones. The detection principle is general and thus applicable for quantification of other bio‐species. A high performance PEC bioassay system based on cathodic reaction at a bio‐cathode possessing a solid–liquid–air triphase joint interface is designed and constructed by assembling an oxidase layer on a superhydrophobic single‐crystal TiO2 nanowire arrays. The triphase PEC bioassay simultaneously exhibits high detection selectivity, wide linear detection up limit, high sensitivity, and low minimum detection limit. [ABSTRACT FROM AUTHOR]