Urease-catalyzed Hydrolysis Of Urea Research Articles

Highly Sensitive Fiber-Optic SPR Urea Sensor Based on ZIF-8/Urease

A highly-sensitive fiber-optic urea sensor was designed and experimentally demonstrated based on surface plasmon resonance (SPR) effect and metal-organic zeolite framework (ZIF-8)/urease modification. The SPR effect was excited by plating gold film on the thin core fiber in multimode-thin core-multimode fiber structure, which behaved high refractive index sensitivity. The ZIF-8/urease composite film was grown on the gold film by rapid aqueous phase synthesis method, in which the ZIF-8 could immobilize more urease and made urease and urea fully contact. Since the urease catalyzed urea hydrolysis would change the refractive index, the measurement of urea concentration was realized with high sensitivity and good selectivity. Experimental results showed that the urea measurement sensitivity of the proposed sensor could reach 6 nm/mM within the concentration range of 1-7 mM, which has great application potential in the field of human health monitoring.

Single-layer HNb3O8 with strong and nearby Lewis and Brønsted acid sites boosts amide bond hydrolysis for urease mimicking.

Urea pollution is a growing environmental concern, and its removal via catalytic hydrolysis is challenging due to the resonance-stabilized amide bonds. In nature, this reaction is catalyzed by ureases in many soil bacteria. However, the remedy of this problem with natural enzymes is not feasible as they are easily denatured and require high costs for both preparation and storage. Given this, the development of nanomaterials bearing enzyme-like activity (nanozymes) with advantages such as low production cost, simple storage, and pH/thermal stability has attracted much attention over the past decade. As inspired by the mechanism of urease-catalyzed urea hydrolysis, the co-presence of Lewis acid (LA) and Brønsted acid (BA) sites is imperative to proceed with this reaction. Herein, layered HNb3O8 samples with intrinsic BA sites were adopted for investigation. The layer reduction of this material to few-/single layers can expose Nb sites with various LA strengths depending on the degree of NbO6 distortion. Among the catalysts examined, single-layer HNb3O8 bearing strong LA and BA sites displays the best hydrolytic activity towards acetamide and urea. This sample with high thermal stability was found to outperform urease at temperatures higher than 50 °C. The acidity-activity correlation established in this study is believed to guide the future design of industrial catalysts to remediate urea pollution.

Low-temperature in situ CO2 enhanced oil recovery

The new generation in situ CO2 enhanced oil recovery (ICE), which delivers CO2 by injection of a CO2 generating agent solution, was proven to offer robust recovery performances and potential cost and workflow benefits. However, the developed ICE systems are either formulated with a complex fluid recipe or a simplified version of a fluid system limited by high-temperature requirements. This work focused on removing the requirement of high reservoir temperature in our previous work while maintaining the simplicity of the single fluid injection system. Urease is studied as the catalyst for urea hydrolysis and evaluated experimentally. The denaturation behavior of urease and the reaction kinetics of the catalyzed urea hydrolysis process are tested at different urea concentrations, urease concentrations, and temperatures. Four sets of one-dimensional sand pack flowthrough experiments are conducted to verify the tertiary recovery potential of the newly developed low-temperature ICE system with different lithology. The extent of wettability alteration and the recovery mechanism for different lithologies was determined through core sample imbibition experiments and direct contact angle measurements. From the experimental results, urease-catalyzed urea hydrolysis is proven to be effective in tertiary oil recovery applications below the 50 °C reservoir temperature range with urea conversion ratio up to 68.7 %. The selected low-temperature ICE system (10 wt% urea solution with 31 U/g urease) had superior tertiary recoveries(Etr) in the flowthrough test for sandstone(Etr = 31.3 %) and carbonate(Etr = 27.5 %) than the corresponding high-temperature cases. Imbibition tests of this improved formulation with various porous media show distinct wettability reversal trends towards a more water-wet state post imbibition. This trend is associated with the produced NH3 and CO2 due to urea hydrolysis. Both oil-aged sandstone and carbonate imbibition tests show noticeable water wetness improvement. The observations of this work significantly expand the operational envelopes of the current ICE system.

NQS-Doped PDMS Solid Sensor: From Water Matrix to Urine Enzymatic Application.

The development of in situ analytical devices has gained outstanding scientific interest. A solid sensing membrane composed of 1,2-naphthoquinone-4-sulfonate (NQS) derivatizing reagent embedded into a polymeric polydimethylsiloxane (PDMS) composite was proposed for in situ ammonium (NH4+) and urea (NH2CONH2) analysis in water and urine samples, respectively. Satisfactory strategies were also applied for urease-catalyzed hydrolysis of urea, either in solution or glass-supported urease immobilization. Using diffuse reflectance measurements combined with digital image processing of color intensity (RGB coordinates), qualitative and quantitative analyte detection was assessed after the colorimetric reaction took place inside the sensing membrane. A suitable linear relationship was found between the sensor response and analyte concentration, and the results were validated by a thymol-PDMS-based sensor based on the Berthelot reaction. The suggested sensing device offers advantages such as rapidity, versatility, portability, and employment of non-toxic reagents that facilitate in situ analysis in an energy-efficient manner.

A visual detection of human immunodeficiency virus gene using ratiometric method enabled by phenol red and target-induced catalytic hairpin assembly

Relying on the specific coordination of Ag+ and mismatched cytosine-cytosine (C–C), the high-efficiency inhibition of urease by Ag+ ion, and the rapid and sensitive response of phenol red to pH, a sensitive ratiometric sensor has been designed for visual detection of human immunodeficiency virus gene (HIV DNA). This sensor utilizes the HIV DNA to initiate catalytic hairpin assembly (CHA) process, releasing Ag+ to inhibit subsequent urease-catalyzed urea hydrolysis and prevent the pH of the solution from rising. The CHA process and the absorbance ratio of phenol red at different wavelengths (A559/A432) amplify the signal, allowing the sensor to detect HIV DNA from 10 to 130 nM in a sensitive and highly selective manner with a low detection limit of 7.8 nM. In addition, this sensor can visually distinguish different concentrations of HIV DNA within a certain range and possesses a good recovery in 1% of serum samples, which will provide new ideas for biosensor design, dipstick test, blood test, and other clinical disease prevention.

A pH sensitive fluorescent carbon dots for urea and urease detection

In this manuscript, a pH-sensitive carbon dots were prepared by directly carbonizing m-phenylenediamine aqueous solution in high pressure autoclave. The prepared carbon dots were gradually protonated with the increase of hydrogen ions concentrations. The fluorescence of prepared carbon dots gradually decreased with the pH value increasing from 3.0 to 9.4, and there existed a well linear relationship between fluorescence intensity at 520 nm and pH value in the range from 6.2 to 8.6. Therefore, the prepared carbon dots showed the potential to be a fluorescent probe for pH detection. Based on the pH-responsive fluorescent carbon dots and urease-catalyzed hydrolysis of urea, we further proposed an assay system for the determination of urea and urease respectively in the range from 0.05 to 3.0 mmol·L−1 and 2.5 to 80 mg·L−1.

A fast and simple headspace gas chromatographic technique for quantitatively analyzing urea in human urine

A fast and convenient headspace gas chromatographic (HS-GC) approach was described for the estimation of urea in human urine. The HS-GC could detect the generated carbon dioxide derived from the urease-catalyzed hydrolysis of urea. It was found that the hydrolysis of urea catalyzed by urease was completed within 40 min at 35 °C. The results proved the great accuracy (relative errors ≤ 8.48%) and precision (RSD ≤ 2.66%) of the HS-GC approach. Moreover, the recoveries ranged from 97.9% to 101.5%. The new approach is rapid and automated, which provides a new way to routinely analyze urea in urine for the control of metabolic disease.

Pathogenesis of Proteus mirabilis Infection.

Proteus mirabilis, a Gram-negative rod-shaped bacterium most noted for its swarming motility and urease activity, frequently causes catheter-associated urinary tract infections (CAUTIs) that are often polymicrobial. These infections may be accompanied by urolithiasis, the development of bladder or kidney stones due to alkalinization of urine from urease-catalyzed urea hydrolysis. Adherence of the bacterium to epithelial and catheter surfaces is mediated by 17 different fimbriae, most notably MR/P fimbriae. Repressors of motility are often encoded by these fimbrial operons. Motility is mediated by flagella encoded on a single contiguous 54-kb chromosomal sequence. On agar plates, P. mirabilis undergoes a morphological conversion to a filamentous swarmer cell expressing hundreds of flagella. When swarms from different strains meet, a line of demarcation, a "Dienes line," develops due to the killing action of each strain's type VI secretion system. During infection, histological damage is caused by cytotoxins including hemolysin and a variety of proteases, some autotransported. The pathogenesis of infection, including assessment of individual genes or global screens for virulence or fitness factors has been assessed in murine models of ascending urinary tract infections or CAUTIs using both single-species and polymicrobial models. Global gene expression studies performed in culture and in the murine model have revealed the unique metabolism of this bacterium. Vaccines, using MR/P fimbria and its adhesin, MrpH, have been shown to be efficacious in the murine model. A comprehensive review of factors associated with urinary tract infection is presented, encompassing both historical perspectives and current advances.

Temporal Control of Gelation and Polymerization Fronts Driven by an Autocatalytic Enzyme Reaction.

Chemical systems that remain kinetically dormant until activated have numerous applications in materials science. Herein we present a method for the control of gelation that exploits an inbuilt switch: the increase in pH after an induction period in the urease‐catalyzed hydrolysis of urea was used to trigger the base‐catalyzed Michael addition of a water‐soluble trithiol to a polyethylene glycol diacrylate. The time to gelation (minutes to hours) was either preset through the initial concentrations or the reaction was initiated locally by a base, thus resulting in polymerization fronts that converted the mixture from a liquid into a gel (ca. 0.1 mm min−1). The rate of hydrolytic degradation of the hydrogel depended on the initial concentrations, thus resulting in a gel lifetime of hours to months. In this way, temporal programming of gelation was possible under mild conditions by using the output of an autocatalytic enzyme reaction to drive both the polymerization and subsequent degradation of a hydrogel.

PH-Sensitive gold nanoclusters: preparation and analytical applications for urea, urease, and urease inhibitor detection

Herein, we reported for the first time a facile synthetic process of gold nanoclusters (AuNCs) by using N-acetyl-L-cysteine both as a reducing agent and as a protection ligand. Based on the pH stimuli-responsive properties of the as-prepared AuNCs, we constructed a pH-sensing platform for the detection of urea, urease, and urease inhibitors.

Simple and reliable urea assay based on a signal accumulation type of ion-sensitive field-effect transistor

A simple urea assay was developed using a signal accumulation type of ion-sensitive field-effect transistor (SA-ISFET). Decreases in proton concentration resulting from urease-catalyzed hydrolysis of urea are detected by SA-ISFET as a change in potential. The method exhibits high sensitivity, linearity, and reproducibility when potential signals are accumulated 10-fold.

PH Wave-Front Propagation in the Urea-Urease Reaction

The urease-catalyzed hydrolysis of urea displays feedback that results in a switch from acid (pH ∼3) to base (pH ∼9) after a controllable period of time (from 10 to >5000 s). Here we show that the spatially distributed reaction can support pH wave fronts propagating with a speed of the order of 0.1−1 mm min−1. The experimental results were reproduced qualitatively in reaction-diffusion simulations including a Michaelis-Menten expression for the urease reaction with a bell-shaped rate-pH dependence. However, this model fails to predict that at lower enzyme concentrations, the unstirred reaction does not always support fronts when the well-stirred reaction still rapidly switches to high pH.

Characterization of silica particles prepared via urease-catalyzed urea hydrolysis and activity of urease in sol–gel silica matrix

Urease templated precipitation of silica synthesized by sol–gel chemistry produces a composite material allowing high urease activity. This study investigates the structural properties of the composite material that allow for the retention of the urease hydrolysis activity. Scanning (SEM) and transmission (TEM) electron microscopy reveal that the composite has a mesoporous structure composed of closely packed spherical structures ∼20–50nm in diameter. Brunauer–Emmett–Teller (BET) analysis revealed that the surface area and pore volume of the composite prepared under the conditions of 50mM urea and 25°C is relatively high (324m2/g and 1.0cm3/g). These values are equivalent to those of usual mesoporous silica materials synthesized from the self-assembly of triblock copolymers as organic templates. In addition, after encapsulating in a sol–gel silica matrix, urease retained high activity (∼90% of the activity compared with native urease). Our results suggest a new method for synthesizing mesoporous silica materials with highly tunable pore sizes and shapes under mild conditions.

Response surface design for the optimization of enzymatic detection of mercury in aqueous solution using immobilized urease from vegetable waste

Soluble and alginate immobilized urease was utilized for detection and quantitation of mercury in aqueous samples. Urease from the seeds of pumpkin, being a vegetable waste, was extracted and purified to apparent homogeneity (sp. activity 353U/mg protein; A280/A260=1.12) by heat treatment at 48±0.1°C and gel filtration through Sephadex G-200. Homogeneous enzyme preparation was immobilized in 3.5% alginate leading to 86% immobilization, no leaching of enzyme was found over a period of 15 days at 4°C. Urease catalyzed urea hydrolysis by soluble and immobilized enzyme revealed a clear dependence on the concentration of Hg2+. Inhibition caused by Hg2+ was non-competitive (Ki=1.2×10−1μM for soluble and 1.46×10−1μM for alginate immobilized urease.). Time-dependent inhibition both in presence and in absence of Hg2+ ion revealed a biphasic inhibition in activity. For optimization of this process response surface methodology (RSM) was utilized where two-level-two-full factorial (22) central composite design (CCD) has been employed. The regression equation and analysis of variance (ANOVA) were obtained using MINITAB® 15 software. Predicted values thus obtained were closed to experimental value indicating suitability of the model. 3D response surface plot, iso-response contour plot and process optimization curve were helpful to predict the results by performing only limited set of experiments.

Factorial design for the optimization of enzymatic detection of cadmium in aqueous solution using immobilized urease from vegetable waste

Free as well as alginate immobilized urease was utilized for detection and quantitation of cadmium (Cd2+) in aqueous samples. Urease from the seeds of pumpkin (Cucumis melo), being a vegetable waste, was extracted and purified to apparent homogeneity (Sp. Activity 353U/mg protein; A280/A260=1.12) by heat treatment at 48±0.1°C and gel filtration through Sephadex G-200. The homogeneous enzyme preparation was immobilized in 3.5% alginate leading to 86% immobilization and no leaching of the enzyme was found over a period of 15 days at 4°C. Urease catalyzed urea hydrolysis by both soluble and immobilized enzyme revealed a clear dependence on the concentration of Cd2+. The inhibition caused by Cd2+ was non-competitive (Ki=1.41×10−5M). The time dependent inhibition both in the presence and in absence of Cd2+ ion revealed a biphasic inhibition in the activity. A Response Surface Methodology (RSM) for the parametric optimization of this process was performed using two-level-two-full factorial (22), central composite design (CCD). The regression coefficient, regression equation and analysis of variance (ANOVA) was obtained using MINITAB® 15 software. The predicted values thus obtained were closed to the experimental value indicating suitability of the model. In addition to this 3D response surface plot and isoresponse contour plot were helpful to predict the results by performing only limited set of experiments.

A small-angle X-ray scattering study of aggregation and gelation of colloidal silica

Abstract Aggregation and gelation in colloidal silica dispersions have been studied by time-resolved, small-angle X-ray scattering. Two different ways of raising the ionic strength, leading to destabilization of the dispersions, have been examined. In situ destabilization by urease-catalyzed hydrolysis of urea has been used as well as an approach with simple mixing of sols with salt solutions. For concentrated dispersions different structures result from the two destabilization methods. Structures differ also in the gelled state, with essentially no fractal scattering from the directly mixed samples whereas fractal scattering on intermediate length scales is observed for the in situ destabilized samples. In contrast, for more dilute particle concentrations, similar structures are generated in both cases and at gelation structures are in close agreement. However, gel times differ drastically in some cases for the two methods, as do the conditions for gelation. These differences are tentatively rationalized by some degree of irreversible aggregation occurring in the mixing stage that, given sufficient time, leads to gelation.

Nanoassembly of biocompatible microcapsules for urease encapsulation and their use as biomimetic reactors

Biocompatible polypeptide capsules with high enzyme loading and activity prepared by templating mesoporous silica spheres were used as biomimetic reactors for performing CaCO3 synthesis exclusively inside the capsule interior via urease-catalyzed urea hydrolysis.

Rapid coagulation of silicon carbide slurry via direct coagulation casting

Direct coagulation casting via urease-catalyzed hydrolysis of urea in SiC suspension was studied at room temperature. The results demonstrate that high-solids-loaded SiC slurries with low viscosity can be prepared at pH 9.5–10.5. The urease-catalyzed decomposition of urea can be used to shift the pH and to increase the ionic strength of the SiC slurry. The pH value decreased with the increase of coagulation time. The optimal coagulation temperature is 50 °C for 62 vol.% SiC slurry. The strength of wet green body after 1 h of coagulation at room temperature is high enough for demolding and handling without damage in the wet green stage.

Inhibition of chitosan-immobilized urease by slow-binding inhibitors: Ni 2+, F − and acetohydroxamic acid

The inhibitions by Ni 2+ and F − ions and by acetohydroxamic acid of jack bean urease covalently immobilized on chitosan membrane was studied (pH 7.0, 25°C) and compared with those of the native enzyme. The reaction progress curves of the immobilized urease-catalyzed hydrolysis of urea were recorded in the absence and presence of the inhibitors. They revealed that the inhibitions are of the competitive slow-binding type similar to those of native urease. The immobilization weakened the inhibitory effect of the inhibitors on urease as measured by the inhibition constants K i ∗ . The increase in their values: 17.9-fold for Ni 2+, 26.5-fold for F − and 1.7-fold for acetohydroxamic acid, was accounted for by environmental effects generated by heterogeneity of the urease–chitosan system: (1) mass transfer limitations imposed on substrate and reaction product in the external solution, and (2) the increase in local pH on the membrane produced by both the enzymatic reaction and the electric charge of the support. By relating the K M /K i ∗ ratio to the electrostatic potential of chitosan it was found that while the reduced Ni 2+ inhibition is mainly brought about by the potential, inhibition by acetohydroxamic acid is independent of the potential, and the acid inhibits urease in its non-ionic form. The reduction in F − inhibition was ascribed to the increased pH in the local environment of the immobilized enzyme.

Inhibition of jack bean urease by thiols.: Calorimetric studies

The influence of three thiol compounds: β-mercaptoethylamine thioglycolic acid and thioacetic acid on the urease-catalyzed hydrolysis of urea was studied. The reaction was carried out in phosphate buffer pH 7.0 at 25°C. The progress of the reaction was observed by means of a calorimetric method using an isoperibol calorimeter. The total reaction progress curves, Δ T vs. t, were interpreted with the help of the integration Jennings–Niemann method and the inhibition constants were calculated.