Urease Enzyme Research Articles

Swarms of Enzymatic Nanobots for Efficient Gene Delivery.

This study investigates the synthesis and optimization of nanobots (NBs) loaded with pDNA using the layer-by-layer (LBL) method and explores the impact of their collective motion on the transfection efficiency. NBs consist of biocompatible and biodegradable poly(lactic-co-glycolic acid) (PLGA) nanoparticles and are powered by the urease enzyme, enabling autonomous movement and collective swarming behavior. In vitro experiments were conducted to validate the delivery efficiency of fluorescently labeled NBs, using two-dimensional (2D) and three-dimensional (3D) cell models: murine urothelial carcinoma cell line (MB49) and spheroids from human urothelial bladder cancer cells (RT4). Swarms of pDNA-loaded NBs showed enhancements of 2.2- to 2.6-fold in delivery efficiency and 6.8- to 8.1-fold in material delivered compared to inhibited particles (inhibited enzyme) and the absence of fuel in a 2D cell culture. Additionally, efficient intracellular delivery of pDNA was demonstrated in both cell models by quantifying and visualizing the expression of eGFP. Swarms of NBs exhibited a >5-fold enhancement in transfection efficiency compared to the absence of fuel in a 2D culture, even surpassing the Lipofectamine 3000 commercial transfection agent (cationic lipid-mediated transfection). Swarms also demonstrated up to a 3.2-fold enhancement in the amount of material delivered in 3D spheroids compared to the absence of fuel. The successful transfection of 2D and 3D cell cultures using swarms of LBL PLGA NBs holds great potential for nucleic acid delivery in the context of bladder treatments.

Theoretical Study on the Disparate Spatial Arrangement of 3’,4’,5,7-tetrahydroxy-3-flavene Towards Jack Bean Urease Enzyme Active Site

Urease enzyme plays crucial role in the hydrolysis of urea. Excessive hydrolysis of urea can have significant impacts on the environment. In recent years, there has been growing interest in identifying natural compounds that can inhibit urease activity. Flavonoids, phytochemical compounds present in plants, have shown promising potential as urease inhibitors. Studies have revealed that certain flavonoids, such as 3’,4’,5,7-tetrahydroxy-3-flavene, abbreviated as H4FLA for convenient, exhibit potent inhibition of urease, surpassing the efficacy of well-known synthetic inhibitors. In the present study, we report quantum mechanical calculations that mainly investigate the interaction of H4FLA towards urease at disparate spatial arrangement. It was found that the most favourable position between H4FLA and active site of urease has interaction energy of -3.80 eV. Topology analysis revealed that there is no typical covalent bond found between atoms involved in the interaction. Only weak interactions were detected. The hydroxyl groups with highest (negative) local potential throughout the structure, contribute mainly to the formation of non-covalent interaction with the nickel centers, indicating their potential involvement in the inhibitory activity of flavonoids against urease.

Novel Copper (II) Complexes with Fluorine-Containing Reduced Schiff Base Ligands Showing Marked Cytotoxicity in the HepG2 Cancer Cell Line.

Several novel copper (II) complexes of reduced Schiff bases containing fluoride substituents were prepared and structurally characterized by single-crystal X-ray diffraction. The complexes exhibited diverse structures, with the central atom in distorted tetrahedral geometry. The biological effects of the products were evaluated, specifically their cytotoxicity, antimicrobial, and antiurease activities, as well as affinity for albumin (BSA) and DNA (ct-DNA). The complexes showed marked cytotoxic activities in the HepG2 hepatocellular carcinoma cell line, considerably higher than the standard cisplatin. The cytotoxicity depended significantly on the substitution pattern. The best activity was observed in the complex with a trifluoromethyl group in position 4 of the benzene ring-the dichloro[(±)-trans-N,N'-bis-(4-trifluoromethylbenzyl)-cyclohexane-1,2-diamine]copper (II) complex, whose activity (IC50 28.7 μM) was higher than that of the free ligand and markedly better than the activity of the standard cisplatin (IC50 336.8 μM). The same complex also showed the highest antimicrobial effect in vitro. The affinity of the complexes towards bovine serum albumin (BSA) and calf thymus DNA (ct-DNA) was established as well, indicating only marginal differences between the complexes. In addition, all complexes were shown to be excellent inhibitors of the enzyme urease, with the IC50 values in the lower micromolar region.

An inhibition potential on some metabolic enzymes (urease and xanthine oxidase), essential oil contents and antioxidant effect of Sideritis lanata L.

An inhibition potential on some metabolic enzymes (urease and xanthine oxidase), essential oil contents and antioxidant effect of Sideritis lanata L.

Synthesis, In Vitro Enzymatic Inhibition, and Molecular Modeling of Novel Piperazine‐Based Bis‐Schiff Base Derivatives as Promising Anti‐urease Agents

AbstractThe current study was aimed to synthesize piperazine‐based bis‐Schiff base derivatives (1–15) and were also screened in vitro for their inhibition against urease enzyme under the positive control of thiourea drug (IC50 value of 36.40±2.35 μM). Among the tested analogs, the maximum potency was shown by analog 2 with the lowest IC50 value of 2.10±1.10 μM, whereas the minimum activity was demonstrated by the scaffold 5 having an IC50 value of 49.60±6.10 μM. The experimental results of urease activity prompted us to investigate and propose a possible mechanism of how actives analogs of piperazine derivatives will interacts with the catalytic sites of targeted urease enzymes. For this purpose, molecular docking with Auto Dock Vina gave us an insight into the binding interactions of the active compounds to different amino acid residues of the targeted urease enzyme.

ESSENTIAL OIL CONTENTS AND BIOLOGICAL ACTIVITIES OF THYMUS CANOVIRIDIS JALAS AND THYMUS SIPYLEUS BOISS.

Objective: Members of the Lamiaceae family are considered to be major sources of bioactive therapeutic agents. Many of them are important medicinal and aromatic plants used in traditional and modern medicine and in the food, cosmetic and pharmaceutical industries. The aim of this study was to investigate in detail the biological activities and chemical composition of the essential oils of Thymus canoviridis Jalas and Thymus sipyleus Boiss. species belonging to the genus Thymus, one of the most important genera of the Lamiaceae family. Material and Method: The essential oil content of the species was determined by GC-MS. Antioxidant activities of the essential oils were determined using lipid peroxidation, DPPH free radical, ABTS cation radical and CUPRAC methods. In addition, cytotoxic activities against breast cancer (MCF-7) and colon cancer (HT-29) cell lines and anticholinesterase (against AChE and BChE enzymes), urease, tyrosinase, elastase, collagenase and angiotensin converting enzyme inhibition activities were determined. Result and Discussion: When the essential oil composition of T. sipyleus was analyzed, the major compounds were 1,8-cineole (eucalyptol) (18.16%), camphor (15.08%) and endo-borneol (11.63%), while T. canoviridis was found to be rich in carvacrol (72.88%). T. canoviridis showed high antioxidant activity in lipid peroxidation (IC50: 45.72±0.12 μg/ml), ABTS (IC50: 6.12±0.03 μg/ml) and CUPRAC (IC50: 5.31±0.01 μg/ml) methods. The selectivities of T. canoviridis and T. sipyleus species against MCF-7 cell line were 4.39 and 6.81, respectively. In the enzyme inhibition studies, both Thymus species showed moderate inhibition activity against BChE enzyme (Inhibition%: 57.88±1.14, 39.21±0.89, respectively). In addition, T. sipyleus showed moderate inhibition of elastase enzyme (Inhibition%: 25.33±0.79). When the results are evaluated in general, it can be said that T. canoviridis essential oil with its rich carvacrol content and high cytotoxic and antioxidant activity can be preferred as a safer and natural option instead of synthetic preservatives in food, pharmaceutical and cosmetic industries to extend shelf life and ensure food safety.

Fabrication of an amperometric urea biosensor based on chitosan modified pencil graphite electrode

An amperometric biosensor was fabricated by using urease enzyme, graphene oxide (GO), gold nanoparticles (AuNPs) and chitosan (CHIT) composite. Pencil graphite electrode (PGE) was used for the electrodeposition of nanomaterials and the detection of urea. Various techniques such as transmission electron microscopy (TEM), scanning electron microscopy (SEM), fourier-transform infrared (FTIR) spectra, UV–visible spectroscopy, zeta-potential and cyclic voltammetry (CV) were used for characterization. The biosensor showed an optimum response at pH 8.0 and 40 °C temperature. The maximum current of the proposed biosensor was due to electron generated at 0.79 V against Ag/AgCl electrode. Fabricated biosensor showed a wide linear range (0.1–1.0 mg/mL) and low limit of detection (0.9956 mg/mL) with satisfactory selectivity, sensitivity (0.00555 μA/mg/mL/cm2), and stability. It was successfully employed for detecting urea concentration in sera samples of healthy individuals and patients with kidney disorders.

Pyridylpiperazine-based carbodithioates as urease inhibitors: synthesis and biological evaluation.

The urease enzyme is recognized as a valuable therapeutic agent for treating the virulent Helicobacter pylori bacterium because of its pivotal role in aiding the colonization and growth of the bacterium within the gastric mucosa. In order to control the harmful consequences of bacterial infections, urease inhibition presents itself as a promising and effective approach. The current research aimed to synthesize pyridylpiperazine-based carbodithioate derivatives 5a-5n and 7a-7n that could serve as potential drug candidates for preventing bacterial infections through urease inhibition. The synthesized carbodithioate derivatives 5a-5n and 7a-7n were explored to assess their ability to inhibit the urease enzyme after their structural explication by gas chromatography-mass spectrometry (GC-MS). In the in vitro evaluation with thiourea as a standard drug, it was observed that all the synthesized compounds exhibited significant inhibitory activity compared to the reference drug. Among the compounds tested, 5j (bearing an o-tolyl moiety) emerged as the most effective inhibitor, displaying strong urease inhibition with an IC50 value of 5.16 ± 2.68μM. This IC50 value is notably lower than that of thiourea (23 ± 0.03μM), indicating the significantly most potent potential of inhibition. In molecular docking of 5j within the active site of urease, numerous noteworthy interactions were identified.

Effects of urease and cementing solution concentrations on micro-scale enzymatic mineralisation characteristics

Enzymatically induced carbonate precipitation holds significant promise in geotechnical engineering. The concentrations of urease enzyme and cementing solution play vital roles in precipitation behaviours under distinct injection strategies. In situ microscale experiments, conducted in S-shaped microchips, were aimed at exploring precipitation distribution characteristics and micro-clogging effects due to biological precipitation. In the one-phase continuous injection methods, elevated urease concentrations correlated with increased amorphous calcium carbonate formation and more heterogeneous distributions. Conversely, lower cementing solution concentrations produced larger, fewer crystals and more uniform distributions. Higher urease improved efficiency but increased cost, while lower cementing solution concentrations displayed efficient performance. Permeability reduction exhibited a power-law relationship with the precipitation area fraction. In the two-phase staged injection method, elevated urease concentrations fostered more regularly shaped crystals. Higher calcium ion concentrations could inhibit ureolysis rate, reducing precipitates. Higher urease concentrations proved beneficial in improving efficiencies. Permeability exhibited a marginal decrease initially, followed by a significant decline as the precipitation area fraction increased. The results offer microscale insights into concentration effects, aiding the understanding of upscaling mechanisms and numerical simulations.

Evaluation of The Performance of Enzyme-Induced Calcite Precipitations (EICP) Solutions Based on pH and Electrical Conductivity

Abstract EICP is an innovative, sustainable technology that utilizes specific chemicals, particularly urea and calcium chloride, in conjunction with the urease enzyme to induce the precipitation of calcium carbonate. The abovementioned technology is widely acknowledged as contemporary and environmentally responsible, aligning with established sustainability principles and green construction criteria. The quantity of precipitated calcium carbonate is contingent upon the composition of the materials employed in the precipitation process and their respective proportions. The investigation focused on determining the best percentage of calcium carbonate precipitation by conducting experiments on different concentrations of urea, calcium chloride, and the urease enzyme and examining the specific characteristics of the chemical solution that facilitate the precipitation process. The augmentation of urea and calcium chloride to a certain proportion resulted in a corresponding augmentation in the quantity of precipitated substances such as calcium carbonate. In the course of furthering the increase process, it was observed that the value of the precipitated materials remained constant and did not exhibit any further augmentation for some mixing ratios. According to the results of current study, the best ratio to precipitated calcium carbonate can be attributed to using an optimal ratio of 1 mol/L of urea and 0.5 mol/L of calcium chloride. Due to the notable efficacy of urease, the concentration increment of urease did not exhibit significant variation beyond 1 g/l. However, the alteration in the rate of reaction velocity will influence the outcome of EICP process. The experimental results and integration of several methodologies demonstrated a positive correlation between temperature elevation and reaction rate augmentation, resulting in a rapid reaction termination.

A novel urease-assisted ratiometric fluorescence sensing platform based on pH-modulated copper-quenched near-infrared carbon dots and methyl red-quenched red carbon dots for selective urea monitoring.

A novel and sensitive fluorescence ratiometric method is developed for urea detection based on the pH-sensitive response of two fluorescent carbon dot (CD) systems: R-CDs/methyl red (MR) and NIR-CDs/Cu2+. The sensing mechanism involves breaking down urea using the enzyme urease, releasing ammonia and increasing pH. At higher pH, the fluorescence of NIR-CDs is quenched due to the enhanced interaction with Cu2+, while the fluorescence of R-CDs is restored as the acidic MR converts to its basic form, removing the inner filter effect. The ratiometric signal (F608/F750) of the R-CDs/MR and NIR-CDs/Cu2+ intensities changed in response to the pH induced by urea hydrolysis, enabling selective and sensitive urea detection. Detailed spectroscopic and morphological investigations confirmed the fluorescence probe design and elucidated the sensing mechanism. The method exhibited excellent sensitivity (0.00028 mMLOD) and linearity range (0.001 - 8.0 mM) for urea detection, with successful application in milk samples for monitoring adulteration, demonstrating negligible interference and high recovery levels (96.5% to 101.0%). This ratiometric fluorescence approach offers a robust strategy for selective urea sensing in complicated matrices.

IMPACT OF FOREST CONVERSION TO PASTURE ON SOIL ENZYMATIC ACTIVITY IN THE NORTHERN AMAZON

The conversion of forests to pastures in the Amazon results in deforestation and the loss of environmental services. This practice affects biogeochemical cycles and impairs soil enzyme activity, which is essential for maintaining soil quality. This study aimed to investigate the impact of converting part of the Amazon forest into pastures, focusing on soil enzyme activity. The study was conducted at Fazenda Canto Verde, Roraima, comparing native forest areas and pastures of Brachiaria brizantha and Brachiaria humidicola, established on Haplic acrisol, without fertilization or tillage, under a management regime of 30 days of grazing and 60 days of rest. Sampling involved 12 mini trenches per hectare at two depths, with analysis of enzyme activity post-incubation. Higher activity of carbon cycle enzymes (Cellulase, Invertase, β-Glucosidase) was observed in the forest compared to pastures, especially with B. humidicola. Nitrogen cycle enzymes (Urease, BAA-Protease) were more active in the forest, while B. humidicola showed the highest Casein-Protease activity. In the phosphorus and sulfur cycles, the forest led in Phosphomonoesterase and Phosphodiesterase, while B. humidicola excelled in Arylsulfatase. This study demonstrates that replacing forest with pastures significantly alters soil functionality, impacting biogeochemical cycles and their ecological functions.

Synthesis of 1,2,3-triazole functionalized derivatives of metronidazole as potent inhibitors of urease: Synthesis via click chemistry, in vitro urease inhibition, kinetics and molecular docking studies

Current study is based on multistep synthesis of sixteen new 1,2,3-triazole functionalized derivatives of metronidazole drug 3–18 by using a Cu(I)-catalysed 1,3-dipolar azide-alkyne cycloaddition (CuAAC) reaction via a click chemistry approach. All the compounds were purified by using conventional silica gel column chromatography with isocratic eluent. Their structures were confirmed by using various spectroscopic techniques including UV, IR, LR and HR MS, 1H-, 13CNMR , and 2D-NMR spectroscopy. The library of 1,2,3-triazole derivatives of metronidazole 3–18 was evaluated for urease inhibitory activity, and cytotoxicity against normal cells. Compound 8 (IC50= 6.8 ± 0.8 µM) was found to be the most potent urease inhibitor, while acetohydroxamic acid (IC50 = 20.3 ± 0.4 µM) was used as experimental standard. Compounds 5 (IC50 = 7.9 ± 0.9 µM), 6 (IC50 = 9.5 ± 0.2 µM), 7 (IC50 = 8.5 ± 1.0 µM), 10 (IC50 = 9.3 ± 1.1 µM), and 18 (IC50 = 7.2 ± 0.6 µM) were also found to be potent inhibitors of urease. Parent drug metronidazole also showed a potent urease enzyme inhibitory activity with IC50= 10.54 ± 0.8 µM. All compounds were found to be non-cytotoxic against the human fibroblast (B.J.) cell line. Molecular docking and kinetic studies were performed for compounds 5–8, 10, and 18 which showed that these compounds are competitive inhibitors of urease enzyme inhibition. Urease inhibition is a therapeutic approach to help in the treatment of peptic ulcers, caused by ureolytic bacteria. The initial findings presented herein therefore, deserve further studies.

Dual-enhanced enzyme cascade hybrid hydrogel for the construction of optical biosensor

The biomimetic enzyme cascade system plays a key role in biosensing as a sophisticated signal transduction and amplification strategy. However, constructing a regulated enzyme cascade sensing system remains challenging due to the mismatch of multiple enzyme activities and poor stability. Herein, we design an efficient dual-enhanced enzyme cascade hybrid system (UFD-DEC) containing DNA-controlled nanozymes (Fe-cdDNA) and enzyme (urease) via combining the electrostatic contact effect with the hydrogel-directed confinement effect. Precise modulation of Fe-cdDNA nanozyme by DNA offers a means to control its catalytic efficiency. This regulated UFD-DEC system accelerates the reaction rate and provides remarkable stability compared with the free enzyme system. Benefiting from the plasticity properties of hydrogels, a “lab-in-a-tube” platform was constructed by encapsulating UFD-DEC in a microcentrifuge tube. Such a UFD-DEC-based hydrogel tube exhibits sufficient adaptability to profile urea when used in conjunction with a smartphone-assisted image processing algorithm, which on-site delivers urea information with a detection limit of 0.12 mmol L−1. This customizable and inexpensive miniaturized biosensor platform for monitoring urea may facilitate point-of-care testing applications.

Effects of the rice-mushroom rotation pattern on soil properties and microbial community succession in paddy fields.

Currently, straw biodegradation and soil improvement in rice-mushroom rotation systems have attracted much attention. However, there is still a lack of studies on the effects of rice-mushroom rotation on yield, soil properties and microbial succession. In this study, no treatment (CK), green manure return (GM) and rice straw return (RS) were used as controls to fully evaluate the effect of Stropharia rugosoannulata cultivation substrate return (SRS) on soil properties and microorganisms. The results indicated that rice yield, soil nutrient (organic matter, organic carbon, total nitrogen, available nitrogen and available potassium) and soil enzyme (urease, saccharase, lignin peroxidase and laccase) activities had positive responses to the rice-mushroom rotation. At the interannual level, microbial diversity varied significantly among treatments, with the rice-mushroom rotation significantly increasing the relative alpha diversity index of soil bacteria and enriching beneficial microbial communities such as Rhizobium, Bacillus and Trichoderma for rice growth. Soil nutrients and enzymatic activities were significantly correlated with microbial communities during rice-mushroom rotation. The fungal-bacterial co-occurrence networks were modular, and Latescibacterota, Chloroflexi, Gemmatimonadota and Patescibacteria were closely related to the accumulation of nutrients in the soil. The structural equation model (SEM) showed that fungal diversity responded more to changes in soil nutrients than did bacterial diversity. Overall, the rice-mushroom rotation model improved soil nutrients and rice yields, enriched beneficial microorganisms and maintained microbial diversity. This study provides new insights into the use of S. rugosoannulata cultivation substrates in the sustainable development of agroecosystems.

Role of compost in assisted phytostabilization via three naturally occurring species on mine-contaminated soil and health risk alleviation for livestock

Phytostabilization emerges as an efficient and enduring remediation technique for heavy metal-contaminated mine soils using plants acclimatized to such circumstances. In this research, the synergistic effects of Marrubium cuneatum, Verbascum speciosum, and Stipa arabica species, alongside municipal solid waste compost (MSWC) amendment at 0, 1, 3, and 5 % rates, were assessed for a six-month pot experiment aimed at remediating naturally polluted soil containing lead (Pb) and cadmium (Cd) and mitigating the health risks associated with these metal exposures for grazing ruminants. Applying compost improved the aboveground biomass of M. cuneatum and V. speciosum and S. arabica by 13, 19, and 18 %, respectively. Also, soil dehydrogenase and urease enzyme activities were enhanced up to 131 and 34 %. Upon MSWC application, all three species uptake Cd and Pb at a significantly lower rate, except in the V. speciosum shoot at 3 and 5 % doses, and bioaccumulation factors were markedly diminished. Compost notably augmented the antioxidant system, leading to a reduction in malondialdehyde content of V. speciosum, M. cuneatum, and S. arabica by 35, 29, and 23 %, respectively, and an elevation in chlorophyll level. Superoxide dismutase, catalase, glutathione peroxidase, and ascorbate peroxidase activities either increased or displayed no significant differences. Risk assessment revealed that MSWC substantially decreased the daily intake of Pb and Cd and their accumulation in animal organs, thereby eliminating limits on meat consumption. Findings affirm the combined efficacy of MSWC and the study species in stabilizing contaminated Pb and Cd soils, diminishing risks to animal feed security and human health.

Isatin-based ibuprofen and mefenamic acid Schiff base derivatives as dual inhibitors against urease and α–glucosidase: In vitro, in silico and cytotoxicity studies

α-Glucosidase and urease inhibitors have emerged as crucial for developing therapeutic drugs targeting diabetes and gastrointestinal disorders. This study reports on new series of ibuprofen and mefenamic acid Schiff base derivatives incorporating isatin as dual inhibitors of α-glucosidase and urease enzymes. These synthesized derivatives (7a-r) were structurally characterized by 1H NMR, 13C NMR and HRMS (EI). Biological evaluation (IC50) identified several derivatives i.e.,7a (urease = 17.37 ± 1.37 µM, α-glucosidase = 44.1 ± 1.15 µM), 7j, (urease = 16.61 ± 1.37 µM, α-glucosidase = 81.2 ± 1.33 µM), 7o, (urease = 18.63 ± 1.27 µM, α-glucosidase = 70.3 ± 1.14 µM), 7r (urease = 11.36 ± 1.32 µM, α-glucosidase = 39.3 ± 1.17 µM), as dual inhibitors of urease (thiourea 21.37 ± 1.76 µM) and α–glucosidase (acarbose 375.82 ± 1.76 µM) enzymes. These bioactive derivatives were explored for cell viability studies against mononuclear cells revealing a good cytocompatibility. In silico molecular docking studies were also conducted to predict the binding mode of new derivatives with target enzymes that were found consistent with the results of in vitro research.

A Multienzyme Logic H+ and Na+ Biotransducer.

Sodium ions and protons regulate various fundamental processes at the cell and tissue levels across all biological kingdoms. It is therefore pivotal for bioelectronic devices, such as biosensors and biotransducers, to control the transport of these ions through biological membranes. Our study explores the regulation of proton and sodium concentrations by integrating an Na+-type ATP synthase, a glucose dehydrogenase (GDH), and a urease into a multienzyme logic system. This system is designed to operate using various chemical control input signals, while the output current corresponds to the local change in proton or sodium concentrations. Therein, a H+ and Na+ biotransducer was integrated to fulfill the roles of signal transducers for the monitoring and simultaneous control of Na+ and H+ levels, respectively. To increase the proton concentration at the output, we utilized GDH driven by the inputs of glucose and nicotinamide adenine dinucleotide (NAD+), while recorded the signal change from the biotransducer, together acting as an AND enzyme logic gate. On the contrary, we introduced urease enzyme which hydrolyzed urea to control the decrease in proton concentration, serving as a NOT gate and reset. By integrating these two enzyme logic gates we formed a simple multienzyme logic system for the control of proton concentrations. Furthermore, we also demonstrate a more complex, Na+-type ATP synthase-urease multienzyme logic system, controlled by the two different inputs of ADP and urea. By monitoring the voltage of the peak current as the output signal, this logic system acts as an AND enzyme logic gate. This study explores how multienzyme logic systems can modulate biologically important ion concentrations, opening the door toward advanced biological on-demand control of a variety of bioelectronic enzyme-based devices, such as biosensors and biotransducers.

Energetic features of H-bonded and antiparallel π-stacked supramolecular assemblies in pyrazolones: Experimental, computational and biological analyses

The work presented herein outlines a straightforward synthetic route to produce a series of pyrazolone compounds, which are significant in both coordination chemistry and pharmaceutical research. These heterocyclic compounds were thoroughly characterized using FTIR, 1H and 13C NMR spectroscopy, and their molecular structures were conclusively determined through X-ray crystallography. The resulting supramolecular architectures feature a variety of noncovalent interactions, including conventional NH…O and CH…O hydrogen bonds, CH…halogen contacts, CH…N interactions, and offset π-π stacking. Density Functional Theory (DFT) calculations, alongside Molecular Electrostatic Potential (MEP) surface analysis and Quantum Theory of Atoms in Molecules (QTAIM)/Non-Covalent Interaction (NCI) plot methodologies, were employed to probe these interactions further. This analysis highlighted the significance of both CH…O and NH…O hydrogen bonds and revealed the critical role of antiparallel π-stacking interactions, particularly within chlorophenyl and benzonitrile substituted pyrazolones, underscoring their stabilizing effects on the molecular structure. Biological assessment against urease enzyme unveiled compound 3a as the lead inhibitor with an IC50 value of 0.71 ± 0.09 μM and 31-folds strong inhibition than thiourea, thus making pyrazolone heterocycles as potential drug candidates.

Exploiting the pH-responsive behavior of zinc-dithizone complex for fluorometric urea sensing utilizing red-emission carbon dots

This study develops a novel fluorometric method for the sensitive and selective determination of urea, based on unique system comprising nitrogen doped red-emissive carbon dots (NRECDs), zinc-dithizone complex, and the urease enzyme. The underlying principle of this method relies on the pH increase resulting from the enzymatic breakdown of urea by urease. Initially, the fluorescence of the NRECDs is quenched by the red-colored zinc-dithizone complex. However, upon the addition of urea, the subsequent release of ammonia and the consequent rise in pH lead to the dissociation of the zinc-dithizone complex, causing a color change from red to yellow. This spectral shift eliminates the quenching effect, resulting in the restoration of the CDs’ fluorescence. The prepared NRECDs were comprehensively characterized using various spectroscopic techniques, including fluorometry, X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), UV–visible spectroscopy, and transmission electron microscopy (TEM) imaging. The proposed fluorometric method exhibits excellent sensitivity (Limit of detection = 0.0012 mM) and linearity (R2 = 0.9951) in the determination of urea. Notably, this approach addresses the selectivity limitations of previous pH-sensitive CDs-based methods, which relied solely on the intrinsic response of CDs, lacking specificity in either quenching or fluorescence enhancement. Furthermore, the developed method demonstrates remarkable selectivity, as evidenced by negligible interference from various potentially interfering substances, ensuring reliable and accurate urea quantification. When applied to human serum samples, the method showcased excellent recovery with low relative standard deviations, highlighting its practical applicability in biomedical and clinical applications.