RESUMEN
The discovery of enzyme inhibitors from natural products is a crucial aspect in the development of therapeutic drugs. However, the complexity of natural products presents a challenge in developing simple and efficient methods for inhibitor screening. Herein, we have developed an integrated analytical model for screening xanthine oxidase (XOD) inhibitors that combines simplicity, accuracy, and efficiency. This model utilizes a colorimetric sensor and affinity chromatography technology with immobilized XOD. The colorimetric sensor procedure can quickly identify whether there are active components in complex samples. Subsequently, the active components in the samples identified by the colorimetric sensor procedure were further captured, separated, and identified through affinity chromatography. The integrated analytical model can significantly enhance the efficiency and accuracy of inhibitor screening. The proposed method was applied to screen for an activity inhibitor of XOD in five natural medicines. As a result, a potential active ingredient for XOD, polydatin, was successfully identified from Polygoni Cuspidati Rhizoma et Radix. This work is anticipated to offer new insights for the screening of enzyme inhibitors from natural medicines.
Asunto(s)
Técnicas Biosensibles , Cromatografía de Afinidad , Colorimetría , Inhibidores Enzimáticos , Xantina Oxidasa , Xantina Oxidasa/antagonistas & inhibidores , Xantina Oxidasa/química , Cromatografía de Afinidad/métodos , Colorimetría/métodos , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/análisis , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/aislamiento & purificación , Técnicas Biosensibles/métodos , Enzimas Inmovilizadas/química , Evaluación Preclínica de Medicamentos , HumanosRESUMEN
Establishing a rapid and accurate method for monitoring the freshness of aquatic products is of great importance. Hypoxanthine has been considered an essential indicator of aquatic products' freshness. Here, a novel smartphone colorimetric / inductively coupled plasma mass spectrometry (ICP-MS) / photothermal three-mode sensing strategy was established for monitoring hypoxanthine. Hypoxanthine can be catalyzed by xanthine oxidase to H2O2 and uric acid, which can simultaneously degrade MnO2 nanosheets (NSs) to Mn2+. After filter-assisted separation, the smartphone and ICP-MS were performed by monitoring the color of the membrane and the Mn2+ in the filtrate. Additionally, MnO2 NSs can facilitate the oxidation of dopamine to form polydopamine nanoparticles, which exhibit strong photothermal efficiency. The approach successfully monitored the deterioration of aquatic products under various storage conditions through portable thermometers and smartphones with low limits of detection (LODs), providing a potential application for in-situ evaluation of the freshness of aquatic products.
Asunto(s)
Técnicas Biosensibles , Hipoxantina , Óxidos , Hipoxantina/análisis , Técnicas Biosensibles/instrumentación , Técnicas Biosensibles/métodos , Óxidos/química , Animales , Compuestos de Manganeso/química , Almacenamiento de Alimentos , Contaminación de Alimentos/análisis , Alimentos Marinos/análisis , Límite de Detección , Colorimetría/métodos , Colorimetría/instrumentación , Espectrometría de Masas , Peróxido de Hidrógeno/química , Peróxido de Hidrógeno/análisis , Peces , Xantina Oxidasa/química , Xantina Oxidasa/metabolismo , Teléfono Inteligente , Indoles , PolímerosRESUMEN
The development of facile, low-cost reliable, and precise onsite assays for the bioactive component hypoxanthine (Hx) in meat products is significant for safeguarding food safety and public health. Herein, we proposed a smartphone-assissted aggregation-induced emission (AIE) fluorogen tetraphenylethene (TPE)-incorporated amorphous Fe-doped phosphotungstates (Fe-Phos@TPE) nanozyme-based ratiometric fluorescence-colorimetric dual-mode biosensor for achieving the onsite visual detection of Hx. When the Hx existed, xanthine oxidase (XOD) catalyzed Hx into H2O2 to be further catalyzed into â¢OH by the prominent peroxidase activity of Fe-Phos@TPE at pH = 6.5, resulting in the oxidization of nonfluorescent o-phenylenediamine (OPD, naked-eye colorless) to be yellow fluorescent emissive 2,3-diaminophenazine (DAP, naked-eye dark yellow) at 550 nm as well as the intrinsic blue fluorescence of Fe-Phos@TPE at 440 nm to be decreased via inner-filter effect (IFE) action, thereby realizing a multi-enzyme cascade catalytic reaction at near-neutral pH to overcome the traditional acidity dependence-induced time-consuming and low sensitivity troublesome.
Asunto(s)
Técnicas Biosensibles , Hipoxantina , Productos de la Carne , Técnicas Biosensibles/instrumentación , Hipoxantina/análisis , Hipoxantina/química , Productos de la Carne/análisis , Xantina Oxidasa/química , Xantina Oxidasa/metabolismo , Contaminación de Alimentos/análisis , Animales , Colorantes Fluorescentes/química , Fluorescencia , Teléfono Inteligente , Colorimetría/métodosRESUMEN
Hypoxanthine (Hx), produced by adenosine triphosphate (ATP) metabolism, is a valuable indicator that determines the quality and degradation status of meat products and is also an important biochemical marker to certain diseases such as gout. The rapid emergence of paper-based enzyme biosensors has already revolutionized its on-site determination. But it is still limited by the complex patterning and fabrication, unstable enzyme and uneven coloration. This work aims to develop an eco-friendly method to construct engineered paper microfluidic, which seeks to produce reaction and non-reaction zones without any patterning procedure. Chito-oligosaccharide (COS), derived from shrimp shells, was used to modify nitrocellulose membranes and immobilize xanthine oxidase (XOD) and chromogenic agent of nitro blue tetrazolium chloride (NBT). After modification, micro fluids could converge into the modification area and Hx could be detected by XOD-catalyzed conversion. Due to the positively charged cationic basic properties of COS, the enzyme storage stability and the color homogeneity could be greatly strengthened through the electrostatic attraction between COS and XOD and formazan product. The detection limit (LOD) is 2.30 µM; the linear range is 0.05-0.35 mM; the complete test time can be as short as 5 min. The COS-based biosensor shows high specificity and can be used directly for Hx in complex samples such as fish and shrimp samples, and different broths. This biosensor is eco-friendly, nontechnical, economical and therefore a compelling platform for on-site or home-based detection of food freshness.
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Técnicas Biosensibles , Colodión , Hipoxantina , Oligosacáridos , Xantina Oxidasa , Animales , Oligosacáridos/química , Oligosacáridos/análisis , Técnicas Biosensibles/métodos , Hipoxantina/análisis , Hipoxantina/química , Colodión/química , Xantina Oxidasa/química , Xantina Oxidasa/metabolismo , Peces , Quitina/química , Enzimas Inmovilizadas/química , Enzimas Inmovilizadas/metabolismo , Tecnología Química Verde/métodos , Propiedades de Superficie , Límite de DetecciónRESUMEN
Gout, a painful condition marked by elevated uric acid levels often linked to the diet's high purine and alcohol content, finds a potential treatment target in xanthine oxidase (XO), a crucial enzyme for uric acid production. This study explores the therapeutic properties of alkaloids extracted from sunflower (Helianthus annuus L.) receptacles against gout. By leveraging computational chemistry and introducing a novel R-based clustering algorithm, "TriDimensional Hierarchical Fingerprint Clustering with Tanimoto Representative Selection (3DHFC-TRS)," we assessed 231 alkaloid molecules from sunflower receptacles. Our clustering analysis pinpointed six alkaloids with significant gout-targeting potential, particularly emphasizing the fifth cluster's XO inhibition capabilities. Through molecular docking and the BatchDTA prediction model, we identified three top compounds-2-naphthylalanine, medroxalol, and fenspiride-with the highest XO affinity. Further molecular dynamics simulations assessed their enzyme active site interactions and binding free energies, employing MM-PBSA calculations. This investigation not only highlights the discovery of promising compounds within sunflower receptacle alkaloids via LC-MS but also introduces medroxalol as a novel gout treatment candidate, showcasing the synergy of computational techniques and LC-MS in drug discovery.
Asunto(s)
Etanolaminas , Gota , Helianthus , Helianthus/metabolismo , Ácido Úrico/metabolismo , Ácido Úrico/uso terapéutico , Simulación del Acoplamiento Molecular , Inhibidores Enzimáticos/farmacología , Gota/tratamiento farmacológico , Xantina Oxidasa/química , Xantina Oxidasa/metabolismoRESUMEN
Background: Unsymmetrical thioureas 1-20 were synthesized and then characterized by various spectroscopy techniques such as UV, IR, fast atom bombardment (FAB)-MS, high-resolution FAB-MS, 1H-NMR and 13C-NMR. Methods: Synthetic compounds 1-20 were tested for their ability for antioxidant, lipoxygenase and xanthine oxidase activities. Results: Compounds 1, 2, 9, 12 and 15 exhibited strong antioxidant potential, whereas compounds 1-3, 9, 12, 15 and 19 showed good to moderate lipoxygenase activity. Ten compounds demonstrated moderate xanthine oxidase inhibition. Conclusion: Compound 15 displayed the highest potency among the series, exhibiting good antioxidant, lipoxygenase and xanthine oxidase activities. Theoretical calculations using density functional theory and molecular docking studies supported the experimental findings, indicating the potential of the synthesized compounds as potent antioxidants, lipoxygenases and xanthine oxidase agents.
Asunto(s)
Antioxidantes , Lipooxigenasa , Antioxidantes/química , Simulación del Acoplamiento Molecular , Xantina Oxidasa/química , Xantina Oxidasa/metabolismo , Inhibidores Enzimáticos/química , Tiourea/farmacología , Tiourea/química , Relación Estructura-ActividadRESUMEN
Background: Cinnamic acid and its derivatives have gained significant attention in recent medicinal research due to their broad spectrum of pharmacological properties. However, the effects of these compounds on xanthine oxidase (XO) have not been systematically investigated, and the inhibitory mechanism remains unclear.
Objectives: The objective of this study was to screen 18 compounds and identify the XO inhibitor with the strongest inhibitory effect. Furthermore, we aimed to study the inhibitory mechanism of the identified compound.
Methods: The effects of the inhibitors on XO were evaluated using kinetic analysis, docking simulations, and in vivo study. Among the compounds tested, 4-NA was discovered as the first XO inhibitor and exhibited the most potent inhibitory effects, with an IC50 value of 23.02 ± 0.12 µmol/L. The presence of the nitro group in 4-NA was found to be essential for enhancing XO inhibition. The kinetic study revealed that 4-NA inhibited XO in a reversible and noncompetitive manner. Moreover, fluorescence spectra analysis demonstrated that 4-NA could spontaneously form complexes with XO, referred to as 4-NA-XO complexes, with the negative values of â³H and ΔS.
Results: This suggests that hydrogen bonds and van der Waals forces play crucial roles in the binding process. Molecular docking studies further supported the kinetic analysis and provided insight into the optimal binding conformation, indicating that 4-NA is located at the bottom outside the catalytic center through the formation of three hydrogen bonds. Furthermore, animal studies confirmed that the inhibitory effects of 4-NA on XO resulted in a significant reduction of serum uric acid level in hyperuricemia mice.
Conclusion: This work elucidates the mechanism of 4-NA inhibiting XO, paving the way for the development of new XO inhibitors.
.Asunto(s)
Hiperuricemia , Xantina Oxidasa , Ratones , Animales , Simulación del Acoplamiento Molecular , Xantina Oxidasa/química , Xantina Oxidasa/metabolismo , Ácido Úrico , Cinética , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/químicaRESUMEN
Xanthine oxidase (XO), a rate-limiting enzyme in uric acid production, is the pivotal therapeutic target for gout and hyperuricemia. In this study, 57 peptides from α-lactalbumin and ß-lactoglobulin were obtained via virtual enzymatic hydrolysis, and 10 XO inhibitory peptides were virtually screened using molecular docking. Then toxicity, allergenicity, solubility, and isoelectric point of the obtained 10 novel peptides were evaluated by in silico tools. The XO activity of these synthetic peptides was tested using an in vitro assay by high-performance liquid chromatography. Their inhibitory mechanism was further explored by molecular docking. The results showed that 4 peptides GL, PM, AL, and AM exhibited higher inhibitory activity, and their half maximal inhibitory concentration in vitro was 10.20 ± 0.89, 23.82 ± 0.94, 34.49 ± 0.89, and 40.45 ± 0.92 mM, respectively. The peptides fitted well with XO through hydrogen bond, hydrophobic interaction, and van der Waals forces, and amino acid residues Glu802, Leu873, Arg880, and Pro1076 played an important role in this process. Overall, this study indicated 4 novel peptides GL, PM, AL, and AM from whey protein exhibited XO inhibitory activity, and they might be useful and safe XO inhibitors for hyperuricemia prevention and treatment.
Asunto(s)
Supresores de la Gota , Hiperuricemia , Animales , Supresores de la Gota/farmacología , Supresores de la Gota/uso terapéutico , Hiperuricemia/tratamiento farmacológico , Hiperuricemia/veterinaria , Xantina Oxidasa/química , Xantina Oxidasa/metabolismo , Proteína de Suero de Leche , Simulación del Acoplamiento Molecular , Inhibidores Enzimáticos/química , Péptidos/farmacologíaRESUMEN
Five new polyacetylene derivatives (1-5), cyclocodonlandiynosides A-E, and eight known analogues (6-13) were isolated and identified from the fruits of Cyclocodon lancifolius. Their structures were established via spectroscopic and chemical methods, including NMR, HRESIMS, enzymatic hydrolysis, Mo2(OAc)4-induced circular dichroism and sugar derivatization. Compound 1 contains a nitrogenous fragment, which was rarely found in C14 polyacetylenes. Compounds 3 and 4 are polyacetylene glucosides possessing novel aglycones. All the isolated polyacetylenes (except 12) were screened for their xanthine oxidase (XO) inhibitory activity. All the tested compounds, at the concentration of 62.5 µg/mL, showed XO inhibiting effects. Among them, 13 and 3 showed the most potent XO inhibitory activity with IC50 values of 87.65 and 96.32 µM, compared to the positive control allopurinol with an IC50 value of 19.25 µM.
Asunto(s)
Frutas , Xantina Oxidasa , Polímero Poliacetilénico , Xantina Oxidasa/química , Estructura Molecular , Extractos Vegetales/química , Poliinos/química , Poliinos/farmacología , Inhibidores Enzimáticos/farmacologíaRESUMEN
Recently, the incidence of hyperuricemia increased with patient rejuvenation, searching for new xanthine oxidase (XOD) inhibitors from natural products becomes important. In our previous work, a flavonoid extract of saffron floral bio-residues (SFB) was found to alleviate hyperuricemia via inhibiting XOD. In this study, an integrated approach combining two-dimensional liquid chromatography, surface plasmon resonance (SPR) and isothermal titration calorimetry (ITC) was developed to online screen and character the potential XOD inhibitors from SFB. The two-dimensional liquid chromatography consisted of affinity chromatography and reverse phase chromatography (2D-AR), in which an XOD column, an inactive XOD column, and a control column were used in the first dimensional liquid chromatography to avoid phenomena of "false positive" and "missing screen of compounds with weak affinity to XOD" that often occur in the screening process, and a C18 column was used in the second dimensional liquid chromatography to separate the mixed XOD binders. Four flavonoid glycosides, i.e., quercetin-3-O-sophoroside (QS), kaempferol-3-O-sophoroside (KS), kaempferol-3-O-rutinoside (KR), and kaempferol-3-O-glucoside (KG), were thus successfully screened and identified from SFB extract by the 2D-AR method. The affinity of QS, KS, KR, KG, kaempferol (aglycone of KS, KR and KG), and quercetin (aglycone of QS) binding to XOD was investigated using SPR method, with KD ranged from 4.8 µM to 47.6 µM. The inhibitor constant (KI) of KS, KR, KG, quercetin and kaempferol were 4.92 mM, 1.11 mM, 0.294 mM, 4.93 µM and 3.27 µM, respectively, determined using ITC method. Finally, the anti-XOD activities of KS, the most abundant flavonoid in SFB extract, and kaempferol in hyperuricemia mice were verified, which suggested that the multi-hyphenated approach established herein can be applied for screen and character the XOD inhibitors in natural products.
Asunto(s)
Crocus , Hiperuricemia , Humanos , Animales , Ratones , Quempferoles/farmacología , Xantina Oxidasa/química , Quercetina/farmacología , Inhibidores Enzimáticos/química , Flavonoides/farmacologíaRESUMEN
Three novel antioxidant candidates based on phenolic polyketide, monoacetylphloroglucinol (MAPG), a natural antibiotic compound produced by plant growth-promoting rhizobacteria (PGPR), Pseudomonas fluorescens F113 have been proposed. Initially, a green and highly efficient route to the synthesis of MAPG and its two analogues from phloroglucinol (PG) has been developed. Afterward, their rational mechanism of antioxidant activity has been investigated based on thermodynamic descriptors involved in the double (2H+/2e-) radical trapping processes. These calculations have been performed using the systematic density functional theory (DFT) method at the B3LYP/Def2-SVP level of theory in the gas phase and aqueous solution. Our findings reveal that the double formal hydrogen atom transfer (df-HAT) mechanism is preferred in the gas phase, while the double sequential proton loss electron transfer (dSPLET) mechanism is preferred in aqueous solution for all MAPGs. The 6-OH group represents the most favorable site for trapping radical species for all MAPGs, which is supported by the pKa values obtained from DFT calculations. The role of acyl substituents on the PG ring has been comprehensively discussed. The presence of acyl substituents has a strong influence on the thermodynamic parameters of the phenolic O-H bond in PG. These results are supported by frontier molecular orbitals (FMOs) analysis, where the addition of acyl substituents increases the chemical reactivity of MAPGs significantly. Based on molecular docking and molecular dynamic simulations (MDs), MAPGs are also predicted to be promising candidates for xanthine oxidase (XO) inhibition.HighlightsThe antioxidant activity of the three synthesized monoacetylphloroglucinols (MAPGs) has been investigated using the density functional theory (DFT) method.Acyl substituents increase the chemical reactivity and antioxidant activity of MAPGs.Double formal hydrogen atom transfer (df-HAT) is the preferred mechanism in the gas phase.Double sequential proton loss electron transfer (dSPLET) seems to be more favored in aqueous solution.MAPGs are expected to be promising xanthine oxidase (XO) inhibitors.
Asunto(s)
Antioxidantes , Xantina Oxidasa , Antioxidantes/química , Xantina Oxidasa/química , Protones , Simulación del Acoplamiento Molecular , Hidrógeno , Fenoles , Agua , Radicales Libres , TermodinámicaRESUMEN
Oral intake of 1,4-naphthoquinones could be a potential risk factor for hyperuricemia and gout via activation of xanthine oxidase (XO). Herein, 1,4-naphthoquinones derived from food and food-borne pollutants were selected to investigate the structure and activity relationship (SAR) and the relative mechanism for activating XO in liver S9 fractions from humans (HLS9) and rats (RLS9). The SAR analysis showed that introduction of electron-donating substituents on the benzene ring or electron-withdrawing substituents on the quinone ring improved the XO-activating effect of 1,4-naphthoquinones. Different activation potential and kinetics behaviors were observed for activating XO by 1,4-naphthoquinones in HLS9/RLS9. Molecular docking simulation and density functional theory calculations showed a good correlation between -LogEC50 and docking free energy or HOMO-LUMO energy gap. The risk of exposure to the 1,4-naphthoquinones was evaluated and discussed. Our findings are helpful to guide diet management in clinic and avoid adverse events attributable to exposure to food-derived 1,4-naphthoquinones.
Asunto(s)
Inhibidores Enzimáticos , Naftoquinonas , Humanos , Ratas , Animales , Inhibidores Enzimáticos/química , Simulación del Acoplamiento Molecular , Xantina Oxidasa/química , Medición de Riesgo , DietaRESUMEN
BACKGROUND: Flos Sophorae Immaturus (FSI) is rich in polyphenols and a potential uric acid-lowering food. However, the processing of FSI is greatly restricted due to the heat sensitivity and low solubility of polyphenols. In this study, hydrothermal treatment - an effective strategy - was applied to FSI processing. The variation of xanthine oxidase (XO) inhibitory effect and polyphenol composition of FSI during hydrothermal treatment were recorded. RESULTS: The XO inhibition rate of FSI increased from 32.42% to 89.00% after hydrothermal treatment at 220 °C for 30 min, as well as total polyphenols (from 0.66 to 1.11 mg mL-1 ) and flavonoids (from 1.21 to 1.58 mg mL-1 ). However, high thermal temperature (>160 °C) and extended thermal time (>90 min) caused the degradation of polyphenols. Rutin, kaempferol-3-O-rutinoside and narcissoside rapidly degraded and converted to quercetin, kaempferol and isorhamnetin when the temperature exceeded 160 °C. The maximum yields of quercetin, kaempferol and isorhamnetin were at 220 °C for 30 min, 90 min and 90 min, respectively. Meanwhile, the conversion kinetics conformed to the first-order model. Interestingly, these newly formed polyphenols possessed better XO inhibitory effects than their derivatives with 3-O-rutinoside. CONCLUSION: Polyphenol conversion during hydrothermal treatment was the main reason for enhancing XO inhibitory activity. Therefore, hydrothermal treatment is an appropriate method for improving the XO inhibitory effect of FSI. © 2022 Society of Chemical Industry.
Asunto(s)
Quempferoles , Quercetina , Polifenoles , Xantina Oxidasa/química , RutinaRESUMEN
Gout is an inflammatory joint disease caused by urate crystal deposition, which is associated with hyperuricemia. Gout will take place when the uric acid accumulates. Xanthine oxidase (XO) is a crucial enzyme in the formation of uric acid. Inhibiting XO is one of the means to ameliorate gout. Luteoloside is a kind of natural flavonoid, which has an excellent prospect for relieving gout. But there are few reports on the interaction mechanism between luteoloside and XO currently. In this study, the interaction mechanism between luteoloside and XO was explored using spectroscopy and molecular docking. The fluorescence spectroscopy results indicated that luteoloside could make the intrinsic fluorescence of XO quenched, and the binding constant between luteoloside and XO was (1.85 ± 0.22) × 103 L mol-1 at 298 K. The synchronous fluorescence spectroscopy results showed that the absorption peaks of Tyr and Trp shifted blue, and the hydrophobicity of the microenvironment increased. Moreover, CD spectra showed that α-helix of XO decreased, ß-sheet and ß-turn increased after adding luteoloside. The results of molecular docking analysis showed that XO could combine with luteoloside through hydrogen bonds and hydrophobic force. The results indicated that luteoloside could remarkably interact with XO. Insights into the interaction mechanism provide a necessary basis for the search for low-toxic natural products as targets of XO. HIGHLIGHTS: Luteoloside and xanthine oxidase was a strong binding mode and had only one binding site. Luteoloside could cause α-helix reduced, ß-sheet and ß-turn increased, and change the secondary structure of XO. The binding between luteoloside and xanthine oxidase was a spontaneous process. The main binding force was hydrophobic force between luteoloside and xanthine oxidase.
Asunto(s)
Gota , Xantina Oxidasa , Humanos , Xantina Oxidasa/química , Xantina Oxidasa/metabolismo , Simulación del Acoplamiento Molecular , Ácido Úrico , Inhibidores Enzimáticos/farmacología , Espectrometría de FluorescenciaRESUMEN
Some seed-derived antioxidant peptides are known to regulate cellular modulators of ROS production, including those proposed to be promising targets of anticancer therapy. Nevertheless, research in this direction is relatively slow owing to the inevitable time-consuming nature of wet-lab experimentations. To help expedite such explorations, we performed structure-based virtual screening on seed-derived antioxidant peptides in the literature for anticancer potential. The ability of the peptides to interact with myeloperoxidase, xanthine oxidase, Keap1, and p47phox was examined. We generated a virtual library of 677 peptides based on a database and literature search. Screening for anticancer potential, non-toxicity, non-allergenicity, non-hemolyticity narrowed down the collection to five candidates. Molecular docking found LYSPH as the most promising in targeting myeloperoxidase, xanthine oxidase, and Keap1, whereas PSYLNTPLL was the best candidate to bind stably to key residues in p47phox. Stability of the four peptide-target complexes was supported by molecular dynamics simulation. LYSPH and PSYLNTPLL were predicted to have cell- and blood-brain barrier penetrating potential, although intolerant to gastrointestinal digestion. Computational alanine scanning found tyrosine residues in both peptides as crucial to stable binding to the targets. Overall, LYSPH and PSYLNTPLL are two potential anticancer peptides that deserve deeper exploration in future.
Asunto(s)
Antineoplásicos/metabolismo , Antioxidantes/metabolismo , Quimioinformática/métodos , Descubrimiento de Drogas/métodos , Péptidos/metabolismo , Extractos Vegetales/metabolismo , Semillas/química , Antineoplásicos/química , Antioxidantes/química , Dominio Catalítico , Estabilidad de Medicamentos , Humanos , Proteína 1 Asociada A ECH Tipo Kelch/química , Proteína 1 Asociada A ECH Tipo Kelch/metabolismo , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Péptidos/química , Peroxidasa/química , Peroxidasa/metabolismo , Extractos Vegetales/química , Unión Proteica , Xantina Oxidasa/química , Xantina Oxidasa/metabolismoRESUMEN
Photoelectrochemical (PEC) enzymatic biosensors integrate the excellent selectivity of enzymes and high sensitivity of PEC bioanalysis, but the drawbacks such as high cost, poor stability, and tedious immobilization of natural enzymes on photoelectrodes severely suppress their applications. AgCu@CuO aerogel-based photoelectrode materials with both remarkable enzyme-like activities and outstanding photoelectric properties were innovatively designed and synthesized to evaluate the activity of xanthine oxidase with a wide linear detection range and a low limit of detection.
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Técnicas Biosensibles/métodos , Técnicas Electroquímicas/métodos , Geles/química , Xantina Oxidasa/análisis , Técnicas Biosensibles/instrumentación , Catálisis , Cobre/química , Técnicas Electroquímicas/instrumentación , Electrodos , Pruebas de Enzimas/instrumentación , Pruebas de Enzimas/métodos , Peróxido de Hidrógeno/química , Hipoxantina/química , Límite de Detección , Naftoles/química , Oxidación-Reducción , Procesos Fotoquímicos , Plata/química , Xantina Oxidasa/químicaRESUMEN
Xanthine oxidase (XO) plays a vital role in inducing hyperuricemia and increasing the level of superoxide free radicals in blood, and is proved as an important target for gout. Chrysoeriol (CHE) is a natural flavone with potent XO inhibitory activity (IC50 = 2.487 ± 0.213 µM), however, the mechanism of interaction is still unclear. Therefore, a comprehensive analysis of the interaction between CHE and XO was accomplished by enzyme kinetics, isothermal titration calorimetry (ITC), multi-spectroscopic methods, molecular simulation and ADMET. The results showed that CHE acted as a rapid reversible and competitive-type XO inhibitor and its binding to XO was driven by hydrogen bonding and hydrophobic interaction. Moreover, CHE exhibited a strong fluorescence quenching effect through a static quenching procedure and induced conformational changes of XO. Its binding pattern with XO was revealed by docking study and the binding affinity to XO was enhanced by the interactions with key amino acid residues in the active pocket of XO. Further, CHE showed good stability and pharmacokinetic behavior properties in molecule dynamic simulation and ADMET prediction. Overall, this study shed some light on the mechanism of interaction between CHE and XO, also provided some valuable information concerning the future therapeutic application of CHE as natural XO inhibitor.
Asunto(s)
Biología Computacional , Flavonas/química , Flavonas/metabolismo , Xantina Oxidasa/química , Xantina Oxidasa/metabolismo , Alopurinol/farmacología , Animales , Calorimetría , Bovinos , Dicroismo Circular , Inhibidores Enzimáticos/farmacología , Febuxostat/química , Febuxostat/farmacología , Colorantes Fluorescentes/metabolismo , Hemólisis/efectos de los fármacos , Cinética , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Estructura Secundaria de Proteína , Conejos , Espectrometría de Fluorescencia , Espectrofotometría Ultravioleta , Factores de Tiempo , Xantina Oxidasa/antagonistas & inhibidoresRESUMEN
Xanthine oxidase (XO) is an important target for the effective treatment of hyperuricemia-associated diseases. A series of novel 2-substituted 6-oxo-1,6-dihydropyrimidine-5-carboxylic acids (ODCs) as XO inhibitors (XOIs) with remarkable activities have been reported recently. To better understand the key pharmacological characteristics of these XOIs and explore more hit compounds, in the present study, the three-dimensional quantitative structure-activity relationship (3D-QSAR), molecular docking, pharmacophore modeling, and molecular dynamics (MD) studies were performed on 46 ODCs. The constructed 3D-QSAR models exhibited reliable predictability with satisfactory validation parameters, including q2 = 0.897, R2 = 0.983, rpred2 = 0.948 in a CoMFA model, and q2 = 0.922, R2 = 0.990, rpred2 = 0.840 in a CoMSIA model. Docking and MD simulations further gave insights into the binding modes of these ODCs with the XO protein. The results indicated that key residues Glu802, Arg880, Asn768, Thr1010, Phe914, and Phe1009 could interact with ODCs by hydrogen bonds, π-π stackings, or hydrophobic interactions, which might be significant for the activity of these XOIs. Four potential hits were virtually screened out using the constructed pharmacophore model in combination with molecular dockings and ADME predictions. The four hits were also found to be relatively stable in the binding pocket by MD simulations. The results in this study might provide effective information for the design and development of novel XOIs.
Asunto(s)
Ácidos Carboxílicos/química , Inhibidores Enzimáticos/química , Simulación del Acoplamiento Molecular/métodos , Simulación de Dinámica Molecular , Xantina Oxidasa/antagonistas & inhibidores , Xantina Oxidasa/química , Diseño de Fármacos , Inhibidores Enzimáticos/uso terapéutico , Humanos , Enlace de Hidrógeno , Interacciones Hidrofóbicas e Hidrofílicas , Hiperuricemia/tratamiento farmacológico , Estructura Molecular , Unión Proteica , Relación Estructura-Actividad CuantitativaRESUMEN
BACKGROUND: Abnormally elevated xanthine oxidase (XO) activity has been verified to cause various pathological processes, such as gout, oxidative stress injury and metabolic syndrome. Thus, XO activators may exhibit above potential toxicological properties. Plumbagin (PLB) is an important active compound in traditional Chinese medicine (TCM), while its obvious toxic effects have been reported, including diarrhea, skin rashes and hepatic toxicity. However, the potential toxicity associated with enhancement of XO activity has not been fully illuminated so far. METHODS: The present study investigated the effect of PLB on XO activity by culturing mouse liver S9 (MLS9), human liver S9 (HLS9), XO monoenzyme system with PLB and xanthine. Then, the molecular docking and biolayer interferometry analysis were adopted to study the binding properties between PLB and XO. Finally, the in vivo acceleration effect also investigated by injected intraperitoneally PLB to KM mice for 3 days. RESULTS: PLB could obviously accelerate xanthine oxidation in the above three incubation systems. Both the Vmax values and intrinsic clearance values (CLint, Vmax/Km) of XO in the three incubation systems increased along with elevated PLB concentration. In addition, the molecular docking study and label-free biolayer interferometry assay displayed that PLB was well bound to XO. In addition, the in vivo results showed that PLB (2 and 10 mg/kg) significantly increased serum uric acid levels and enhanced serum XO activity in mice. CONCLUSION: In summary, this study outlines a potential source of toxicity for PLB due to the powerful enhancement of XO activity, which may provide the crucial reminding for the PLB-containing preparation development and clinical application.
Asunto(s)
Naftoquinonas/farmacología , Xantina Oxidasa/metabolismo , Animales , Femenino , Humanos , Hígado/enzimología , Masculino , Ratones , Simulación del Acoplamiento Molecular , Naftoquinonas/química , Oxidación-Reducción , Xantina/química , Xantina/metabolismo , Xantina Oxidasa/químicaRESUMEN
In this study, a polydopamine (PDA)-modified hollow fiber-immobilized xanthine oxidase (XOD) was prepared for screening potential XOD inhibitors from flavonoids. Several parameters for the preparation of PDA-modified hollow fiber-immobilized XOD, including the dopamine concentration, modification time, XOD concentration and immobilization time, were optimized. The results show that the optimal conditions for immobilized XOD activity were a dopamine concentration of 2.0 mg/mL in 10.0 mM Tris-HCl buffer (pH 8.5), a modification time of 3.0 h, an XOD concentration of 1000 µg/mL in 10.0 mM phosphate buffer (pH 7.5) and an immobilization time of 3.0 h. Subsequently, the enzymatic reaction conditions such as the pH value and temperature were investigated, and the enzyme kinetics and inhibition parameters were determined. The results indicate that the optimal pH value (7.5) and temperature (37 °C) of the PDA-modified hollow fiber-immobilized XOD were consistent with the free enzyme. Moreover, the PDA-modified hollow fiber-immobilized XOD could still maintain above 50% of its initial immobilized enzyme activity after seven consecutive cycles. The Michaelis-Menten constant (Km) and the half-maximal inhibitory concentration (IC50) of allopurinol on the immobilized XOD were determined as 0.25 mM and 23.2 µM, respectively. Furthermore, the PDA-modified hollow fiber-immobilized XOD was successfully applied to evaluate the inhibitory activity of eight flavonoids. Quercetin, apigenin, puerarin and epigallocatechin showed a good inhibition effect, and their percentages of inhibition were (79.86 ± 3.50)%, (80.98 ± 0.64)%, (61.15 ± 6.26)% and (54.92 ± 0.41)%, respectively. Finally, molecular docking analysis further verified that these four active compounds could bind to the amino acid residues in the XOD active site. In summary, the PDA-modified hollow fiber-immobilized XOD is an efficient method for the primary screening of XOD inhibitors from natural products.