Showing papers on "Xanthine published in 2015"
TL;DR: The main inhibition mechanism of ka Kempferol on XO activity may be due to the insertion of kaempferol into the active site of XO occupying the catalytic center of the enzyme to avoid the entrance of the substrate and inducing conformational changes ofXO.
Abstract: Xanthine oxidase (XO), a key enzyme in purine catabolism, is widely distributed in human tissues. It can catalyze xanthine to generate uric acid and cause hyperuricemia and gout. Inhibition kinetics assay showed that kaempferol inhibited XO activity reversibly in a competitive manner. Strong fluorescence quenching and conformational changes of XO were found due to the formation of a kaempferol-XO complex, which was driven mainly by hydrophobic forces. The molecular docking further revealed that kaempferol inserted into the hydrophobic cavity of XO to interact with some amino acid residues. The main inhibition mechanism of kaempferol on XO activity may be due to the insertion of kaempferol into the active site of XO occupying the catalytic center of the enzyme to avoid the entrance of the substrate and inducing conformational changes of XO. In addition, luteolin exhibited a stronger synergistic effect with kaempferol than did morin at the lower concentration.
199 citations
TL;DR: The results indicated that ginger rhizome fractions and its active constituents having promising antioxidant, anti-inflammatory, and anti-gout properties and might be used as potential natural drug against oxidative stress and inflammatory related diseases after successful in vivo study and clinical trials.
126 citations
TL;DR: A detailed review of methods of isolation, determination of xanthine oxidase activity, and the effect of plant extracts and their constituents on it is presented.
Abstract: Xanthine oxidase (XO) is an important enzyme catalyzing the hydroxylation of hypoxanthine to xanthine and xanthine to uric acid which is excreted by kidneys. Excessive production and/or inadequate excretion of uric acid results in hyperuricemia. This paper presents a detailed review of methods of isolation, determination of xanthine oxidase activity, and the effect of plant extracts and their constituents on it. Determining the content and activities of XO can be used for diagnostic purposes. Testing inhibition of XO is important for detection of potentially effective compounds or extracts that can be used to treat diseases that are caused by increased activity of XO. In vitro bioassays are used to examine test material for XO inhibition, as inhibitors of XO may be potentially useful for the treatment of gout or other XO induced diseases. Several authors reported on the XO inhibitory potential of traditionally used medicinal plants.
109 citations
TL;DR: A novel nanocomposite host matrix for enzyme immobilization of xanthine oxidase was developed by incorporating MWCNT in poly(GMA-co-VFc) copolymer film which played an important role in the biosensor performance and exhibited high good storage stability and satisfactory anti-interference ability.
80 citations
TL;DR: It appears that the steric effect of the ligand and the hydrophobicity of the complex both play a role in the chemosensitivity of these compounds, with greater steric bulk and greater hydrophilicity delivering higher cytotoxicity.
Abstract: A new library of silver(I)-N-heterocyclic carbene complexes prepared from the natural products caffeine, theophylline and theobromine is reported. The complexes have been fully characterised using a combination of NMR spectroscopy, mass spectrometry, elemental analysis and X-ray diffraction analysis. Furthermore, the hydrophobicity of the complexes has been measured. The silver(I)-N-heterocyclic carbenes have been evaluated for their antiproliferative properties against a range of cancer cell lines of different histological types, and compared to cisplatin. The data shows different profiles of response when compared to cisplatin in the same panel of cells, indicating a different mechanism of action. Furthermore, it appears that the steric effect of the ligand and the hydrophobicity of the complex both play a role in the chemosensitivity of these compounds, with greater steric bulk and greater hydrophilicity delivering higher cytotoxicity.
77 citations
TL;DR: This review focuses on studies of Xor inhibitors and their implications for understanding the chemical nature and reaction mechanism of the Moco active site of XOR.
Abstract: Xanthine oxidoreductase (XOR), which is widely distributed from humans to bacteria, has a key role in purine catabolism, catalyzing two steps of sequential hydroxylation from hypoxanthine to xanthine and from xanthine to urate at its molybdenum cofactor (Moco). Human XOR is considered to be a target of drugs not only for therapy of hyperuricemia and gout, but also potentially for a wide variety of other diseases. In this review, we focus on studies of XOR inhibitors and their implications for understanding the chemical nature and reaction mechanism of the Moco active site of XOR. We also discuss further experimental or clinical studies that would be helpful to clarify remaining issues.
65 citations
TL;DR: Lineweaver-Burk plot analysis revealed that representative compound 4f acted as a mixed-type inhibitor for xanthine oxidase, rationalized by its molecular docking into the active site ofxanthine dehydrogenase.
61 citations
TL;DR: In this paper, a novel and sensitive amperometric xanthine biosensor was developed by preparing a nanocomposite film that was constructed by embedding reduced expanded graphene oxide (REGO) sheets decorated with iron oxide (Fe 3 O 4 ) nanoparticles into poly(glycidyl methacrylate-co-vinylferrocene) (P(GMA- co -VFc)) phase, and by covalent immobilization of Xanthine oxidase (XOD) on the surface of P(Gma-co)-
55 citations
TL;DR: Analysis of circular dichroism demonstrated that chrysin induced the conformational change of XO with increases in α-helix and β-sheet and reductions in β-turn and random coil structures, and the interaction was predominately driven by hydrogen bonds and van der Waals forces.
54 citations
TL;DR: This work characterized NO formation by rat and human XO/XD and AO, the first characterization of human enzymes, and the results support the employment of rat liver enzymes as suitable models of the human counterparts.
Abstract: Nitrite is presently considered a NO "storage form" that can be made available, through its one-electron reduction, to maintain NO formation under hypoxia/anoxia. The molybdoenzymes xanthine oxidase/dehydrogenase (XO/XD) and aldehyde oxidase (AO) are two of the most promising mammalian nitrite reductases, and in this work, we characterized NO formation by rat and human XO/XD and AO. This is the first characterization of human enzymes, and our results support the employment of rat liver enzymes as suitable models of the human counterparts. A comprehensive kinetic characterization of the effect of pH on XO and AO-catalyzed nitrite reduction showed that the enzyme's specificity constant for nitrite increase 8-fold, while the Km(NO2(-)) decrease 6-fold, when the pH decreases from 7.4 to 6.3. These results demonstrate that the ability of XO/AO to trigger NO formation would be greatly enhanced under the acidic conditions characteristic of ischemia. The dioxygen inhibition was quantified, and the Ki(O2) values found (24.3-48.8 μM) suggest that in vivo NO formation would be fine-tuned by dioxygen availability. The potential in vivo relative physiological relevance of XO/XD/AO-dependent pathways of NO formation was evaluated using HepG2 and HMEC cell lines subjected to hypoxia. NO formation by the cells was found to be pH-, nitrite-, and dioxygen-dependent, and the relative contribution of XO/XD plus AO was found to be as high as 50%. Collectively, our results supported the possibility that XO/XD and AO can contribute to NO generation under hypoxia inside a living human cell. Furthermore, the molecular mechanism of XO/AO-catalyzed nitrite reduction was revised.
53 citations
TL;DR: In this paper, a highly sensitive method for the determination of traces of xanthine based on the aggregation of citrate-stabilized gold nanoparticles (AuNPs) was described.
Abstract: We describe a highly sensitive method for the determination of traces of xanthine based on the aggregation of citrate-stabilized gold nanoparticles (AuNPs). It is found that, under optimal conditions of pH, the imide group of xanthine is adsorbed on the surface of the AuNPs, thereby displacing citrate ions. This leads to an aggregation of the AuNPs via hydrogen-bond interactions. As a result, the color of the solution changes from red to blue which can be seen with bare eyes and also can be measured by spectrophotometry. The ratio of the absorbances at 630 nm and 520 nm is linearly related to the concentration of xanthine in the 125 nM to 6.0 μM range (r = 0.9988), and the detection limit (3σ/slope) is 23 nM. The method is simple, feasible and fast.
TL;DR: The inhibition may be attributed to the insertion of Gen into the active site of XO occupying the catalytic center of the enzyme to avoid entry of the substrate and inducing conformational changes ofXO (more compact), which was further unfavorable for forming the active cavity and further reduced the landing and oxidation of substrate.
Abstract: Genistein (Gen), widely distributed in soybean, is proved to be important in homeostasis in the human body. Herein, the inhibitory mechanism of Gen against xanthine oxidase (XO) was studied through multispectroscopic methods and molecular simulation. The inhibition kinetics showed that Gen competitively inhibited XO with an inhibition constant of (1.39 ± 0.11) μM by competing with xanthine for binding to the active site of XO. Fluorescence titration study suggested that the fluorescence quenching mechanism of XO was static, resulting from the formation of a Gen-XO complex at one fold site. The calculated thermodynamic parameters revealed that the interaction process was driven mainly by hydrophobic interactions and hydrogen bonds with affinity of (5.24 ± 0.02) × 10(4) Lmol(-1). Conformational analyses demonstrated that the microenvironment and the secondary structure of XO were changed upon binding of Gen. The molecular docking displayed that Gen bound to the active cavity of XO by interacting with the surrounding amino acid residues (Leu648, Phe649, Glu802, Ser876, Glu879, Arg880, Phe914, Phe1009, Thr1010 and Phe1013). Thus, the inhibition may be attributed to the insertion of Gen into the active site of XO occupying the catalytic center of the enzyme to avoid entry of the substrate and inducing conformational changes of XO (more compact), which was further unfavorable for forming the active cavity and further reduced the landing and oxidation of substrate. This study may offer novel insights into the inhibition mechanism of Gen on XO.
TL;DR: A simple three-step approach to be used for diagnosis, typing and confirmation of HX is suggested and this safe and non-invasive diagnostic algorithm is advocated instead of the traditional allopurinol loading test and intestinal or liver biopsy used in the past.
Abstract: Hereditary xanthinuria (HX) is a rare inherited disorder caused by a deficiency of xanthine dehydrogenase/oxidase (XDH/XO). Missing XDH/XO activity leads to undetectable levels of uric acid excessively replaced by xanthine in serum/urine. The allopurinol loading test has been traditionally used to differentiate between HX types I and II. Final confirmation of HX has been based on the biopsy finding of the absent XDH/XO activity in the small intestine or liver. We present the clinical, biochemical, ultrasound and molecular genetics findings in three new patients with HX and suggest a simple three-step approach to be used for diagnosis, typing and confirmation of HX. In the first step, the diagnosis of HX is determined by extremely low serum/urinary uric acid excessively replaced by xanthine. Second, HX is typed using urinary metabolomics. Finally, the results are confirmed by molecular genetics. We advocate for this safe and non-invasive diagnostic algorithm instead of the traditional allopurinol loading test and intestinal or liver biopsy used in the past.
TL;DR: A primary focus of this mini-review relates to how spectroscopy and computational methods have been used to develop an understanding of critical relationships between geometric structure, electronic structure, and catalytic function.
Abstract: We review the xanthine oxidase (XO) family of pyranopterin molybdenum enzymes with a specific emphasis on electronic structure contributions to reactivity. In addition to xanthine and aldehyde oxidoreductases, which catalyze the two-electron oxidation of aromatic heterocycles and aldehyde substrates, this mini-review highlights recent work on the closely related carbon monoxide dehydrogenase (CODH) that catalyzes the oxidation of CO using a unique Mo–Cu heterobimetallic active site. A primary focus of this mini-review relates to how spectroscopy and computational methods have been used to develop an understanding of critical relationships between geometric structure, electronic structure, and catalytic function.
TL;DR: It is demonstrated for the first time that XOD is activated in THP-1 human myeloid cells in response to pro-inflammatory and growth factor stimulation and plays an important role in maintaining mTOR-dependent translational control during the biological responses of human myELoid cells.
Abstract: Xanthine oxidase (XOD) is an enzyme which plays a central role in purine catabolism by converting hypoxanthine into xanthine and then further into uric acid. Here we report that XOD is activated in THP-1 human myeloid cells in response to pro-inflammatory and growth factor stimulation. This effect occurred following stimulation of THP-1 cells with ligands of plasma membrane associated TLRs 2 and 4, endosomal TLRs 7 and 8 as well as stem cell growth factor (SCF). Hypoxia-inducible factor 1 (HIF-1) and activator protein 1 (AP-1) transcription complexes were found to be responsible for XOD upregulation. Importantly, the mammalian target of rapamycin (mTOR), a major myeloid cell translation regulator, was also found to be essential for XOD activation. Specific inhibition of XOD by allopurinol and sodium tungstate led to an increase in intracellular AMP levels triggering downregulation of mTOR activation by phosphorylation of its T2446 residue. Taken together, our results demonstrate for the first time that XOD is not only activated by pro-inflammatory stimuli or SCF but also plays an important role in maintaining mTOR-dependent translational control during the biological responses of human myeloid cells.
TL;DR: In this paper, a nanocomposite modified Au electrode was characterized by Fourier transform infrared (FTIR), cyclic voltammetry (CV), scanning electron microscopy (SEM) and electrochemical impedance spectroscopy (EIS) before and after immobilization of XOD.
Abstract: Xanthine oxidase (XOD) extracted from bovine milk was immobilized covalently via N-ethyl-N'-(3-dimethylaminopropyl) carbodiimide (EDC) and N-hydroxy succinimide (NHS) chemistry onto cadmium oxide nanoparticles (CdO)/carboxylated multiwalled carbon nanotube (c-MWCNT) composite film electrodeposited on the surface of an Au electrode. The nanocomposite modified Au electrode was characterized by Fourier transform infrared (FTIR), cyclic voltammetry (CV), scanning electron microscopy (SEM) and electrochemical impedance spectroscopy (EIS) before and after immobilization of XOD. Under optimal operation conditions (25 degrees C, + 0.2 V vs. Ag/AgCl, sodium phosphate buffer, pH 7.5), the following characteristics are attributed to the biosensor: linearity of response up to xanthine concentrations of 120 mu M, detection limit of 0.05 mu M (S/N = 3) and a response time of at most 4 s. After being used 100 times over a period of 120 days, only 50% loss of the initial activity of the biosensor was evaluated when stored at 4 degrees C. The fabricated biosensor was successfully employed for the determination of xanthine in fish meat.
TL;DR: The potential application of xanthine derivative 6 and its chitosan formulation (CS-6) in the treatment of the diabetes mellitus syndrome is demonstrated.
TL;DR: Results suggest that hydrogen peroxide generated by xanthine/xanthine oxidase has the potential to suppress colorectal cancer cell proliferation.
Abstract: The twin character of reactive oxygen species is substantiated by a growing body of evidence that reactive oxygen species within cells act as inducers and accelerators of the oncogenic phenotype of cancer cells, while reactive oxygen species can also induce cancer cell death and can therefore function as anti-tumorigenic species. The aim of this study was to assess a possible influence of xanthine/xanthine oxidase on the proliferation of colorectal cancer cell line Caco-2. xanthine/xanthine oxidase (2.5 µM/0.25 mU/ml–25 µM/2.5 mU/ml) dose-dependently inhibited the proliferation of Caco-2 cells. Experiments utilizing reactive oxygen species scavengers (superoxide dismutase, catalase and mannitol) and exogenous hydrogen peroxide revealed a major role of hydrogen peroxide in the xanthine/xanthine oxidase effect. Investigations utilizing annexin V-fluorescein/PI assay using flow cytometry, and the lactate dehydrogenase extracellular release assay indicated that hydrogen peroxide induced necrosis, but not apoptosis, in Caco-2 cells. These results suggest that hydrogen peroxide generated by xanthine/xanthine oxidase has the potential to suppress colorectal cancer cell proliferation.
TL;DR: This work evaluated the effect of allopurinol, an inhibitor of NAD(P)H-generating xanthine dehydrogenase (XDH), on Clostridium beijerinckii grown in furfural-supplemented medium and found that it unexpectedly increased the rate of detoxification of Furfural by 1.4-fold.
Abstract: In addition to glucans, xylans, and arabinans, lignocellulosic biomass hydrolysates contain significant levels of nonsugar components that are toxic to the microbes that are typically used to convert biomass to biofuels and chemicals. To enhance the tolerance of acetone–butanol–ethanol (ABE)-generating Clostridium beijerinckii NCIMB 8052 to these lignocellulose-derived microbial inhibitory compounds (LDMICs; e.g., furfural), we have been examining different metabolic perturbation strategies to increase the cellular reductant pools and thereby facilitate detoxification of LDMICs. As part of these efforts, we evaluated the effect of allopurinol, an inhibitor of NAD(P)H-generating xanthine dehydrogenase (XDH), on C. beijerinckii grown in furfural-supplemented medium and found that it unexpectedly increased the rate of detoxification of furfural by 1.4-fold and promoted growth, butanol, and ABE production by 1.2-, 2.5-, and 2-fold, respectively. Since NAD(P)H/NAD(P)+ levels in C. beijerinckii were largely unchanged upon allopurinol treatment, we postulated and validated a possible basis in DNA repair to account for the solventogenic gains with allopurinol. Following the observation that supplementation of allopurinol in the C. beijerinckii growth media mitigates the toxic effects of nalidixic acid, a DNA-damaging antibiotic, we found that allopurinol elicited 2.4- and 6.7-fold increase in the messenger RNA (mRNA) levels of xanthine and hypoxanthine phosphoribosyltransferases, key purine-salvage enzymes. Consistent with this finding, addition of inosine (a precursor of hypoxanthine) and xanthine led to 1.4- and 1.7-fold increase in butanol production in furfural-challenged cultures of C. beijerinckii. Taken together, our results provide a purine salvage-based rationale for the unanticipated effect of allopurinol in improving furfural tolerance of the ABE-fermenting C. beijerinckii.
TL;DR: The results suggest that insulin resistance induced by acute caffeine administration is mediated by A1 and A2B adenosine receptors, and both Glut4 and NO seem to be downstream effectors involved in insulin resistanceinduced by acute coffee.
TL;DR: In this article, the formation of π-stacked complexes between water-soluble porphyrins was studied, and it was shown that xanthine and its derivatives can quench the fluorescence of the porphrin according to the number of methyl groups in the molecule of quencher.
TL;DR: In this article, a series of pyrazole-based 1,3,4-oxadiazole derivatives were rationally designed and synthesized in good yields by following a convenient route.
TL;DR: A mutation in the human molybdenum cofactor sulfurase (MCSU) gene was detected and the pathogenicity of the human MCSU gene was strongly confirmed, indicating that hypouricemia is caused by genetic defects of the molybdoflavoprotein enzymes.
Abstract: Background: Hypouricemia is caused by various diseases and disorders, such as hepatic failure, Fanconi renotubular syndrome, nutritional deficiencies and genetic defects. Genetic defects of the molybdoflavoprotein enzymes induce hypouricemia and xanthinuria. Here, we identified a patient whose plasma and urine uric acid levels were both extremely low and aimed to identify the pathogenic gene and verify its mechanism. Methods: Using next-generation sequencing (NGS), we detected a mutation in the human molybdenum cofactor sulfurase (MCSU) gene that may cause hypouricemia. We cultured L02 cells, knocked down MCSU with RNAi, and then detected the uric acid and MCSU concentrations, xanthine oxidase (XOD) and xanthine dehydrogenase (XDH) activity levels, and xanthine/hypoxanthine concentrations in cell lysates and culture supernatants. Results: The NGS results showed that the patient had a mutation in the human MCSU gene. The in vitro study showed that RNAi of MCSU caused the uric acid, human MCSU concentrations, the XOD and XDH activity levels among cellular proteins and culture supernatants to be extremely low relative to those of the control. However, the xanthine/hypoxanthine concentrations were much higher than those of the control. Conclusions: We strongly confirmed the pathogenicity of the human MCSU gene.
TL;DR: The data suggest that quercetin and hydroxytyrosol protects cardiomyocytes against X/XO-induced oxidative toxicity by diminishing intracellular ROS and the regulation of stress-sensitive protein kinase cascades and transcription factors.
Abstract: The increased activity of xanthine/xanthine oxidase (X/XO) has been suggested as a risk factor for heart disease and herbal polyphenols exhibits cardioprotection in vitro and in vivo. To understand the cardioprotective action mechanisms of polyphenol quercetin and hydroxytyrosol, the expression levels of stress-responsive proteins were studied in X/XO-induced toxicity model of H9c2 cardiomyocyocytes. Pretreatment with each polypenol (0.1-10 μg/ml; 24 h) enhanced viability (p < 0.01; MTT test) and inhibited reactive oxygen species (ROS) generation (p < 0.001; H2DCFDA assay) against 12 h exposure to a free radical generating system, X (0.5 mM) and XO (5 mU/ml). Western blotting experiments showed that X/XO increases the phosphorylation of downstream substrate of p38, MAPK-activated protein kinase 2 (MAPKAPK-2), p44/42-MAPK (Erk1/2) and cleaved caspase-3 (p < 0.001, vs. Control), however inhibits the levels of phosphorylated c-Jun and Hsp27 (p < 0.01, vs. Control). Pretreatment with quercetin or hydroxytyrosol attenuated the phosphorylation of MAPKAPK-2 and cleaved caspase-3 in X/XO-exposed cells (p < 0.01, vs. X/XO). Hydroxytyrosol enhanced the reduction of phosphorylation of a transcriptional target c-Jun and led to overphosphorylation in protective proteins, p44/42-MAPK and Hsp27 in X/XO-exposed cells (p < 0.01, vs. X/XO). Our data suggest that quercetin and hydroxytyrosol protects cardiomyocytes against X/XO-induced oxidative toxicity by diminishing intracellular ROS and the regulation of stress-sensitive protein kinase cascades and transcription factors.
TL;DR: Data obtained here indicate that transmembrane cycling of GSH may be involved in E. coli protection against extracellular ROS and may promote rapid growth recovery.
TL;DR: In this paper, various chromatographic separations of the ethyl acetate-soluble fraction of Aster glehni (AGEF) led to the isolation of five new caffeoylglucoside derivatives.
Abstract: In the search for xanthine oxidase (XOD) inhibitors as anti-gout agents from natural products, various chromatographic separations of the ethyl acetate-soluble fraction of Aster glehni (AGEF) led to the isolation of five new caffeoylglucoside derivatives, namely 6′-O-caffeoyl-(6S,9R)-roseoside (1), 6′-O-caffeoylampelopsisionoside (2), 6′-O-caffeoylsonchuinoside C (3), 6′-O-caffeoyldihydrosyringin (4), and (2E)-2-methyl-but-2-ene-1,4-diol-6′-O-caffeoyl-1-O-β-glucopyranoside (glehnoside, 5), together with 13 known compounds. The absolute stereochemistry of the 6′-O-caffeoylsonchuinoside C (3) was established with the help of spectroscopic analyses, enzymatic hydrolysis, and Mosher's method, as well as in comparison with literature data. All isolated substances were determined for their inhibitory activities on uric acid production by the xanthine/XOD system. Among them, 4,5-O-dicaffeoylquinic acid methyl ester (12) showed the most potent inhibitory activity with an IC50 value of 2.6 ± 0.1 μM, which was comparable to that of allopurinol used as a positive control. Furthermore, hypouricemic effects of AGEF were assessed by measuring serum uric acid levels 3 h after potassium oxonate treatment (250 mg/kg, i.p.) to induce hyperuricemia in rats. When preadministered orally once a day at doses of 50, 100, and 300 mg/kg for 7 days, AGEF reduced the potassium oxonate-induced elevated serum uric acid level by 15.4, 39.8, and 32.3%, respectively. The results suggest that AGEF has the potential to be a new source of agents for the prevention and/or treatment of hyperuricemia and gout.
TL;DR: In this article, a novel electrochemical detecting system has been constructed by introducing xanthine oxidase, which can catalyze H or X to uric acid during the detection process.
TL;DR: Observations suggest the involvement of ligand-mediated oligomerization and conformational changes in the process of activation of PfHGXPRT, a unique mechanism of activation where the enzyme switches from a low activity (unactivated) to a high activity (activated) state upon pre-incubation with substrate/products.
Abstract: P. falciparum (Pf) hypoxanthine guanine xanthine phosphoribosyltransferase (HGXPRT) exhibits a unique mechanism of activation where the enzyme switches from a low activity (unactivated) to a high activity (activated) state upon pre-incubation with substrate/products. Xanthine phosphoribosylation by unactivated PfHGXPRT exhibits a lag phase, the duration of which reduces with an increase in concentration of the enzyme or substrate, PRPP·Mg2+. Activated PfHGXPRT does not display the lag phase and exhibits a ten-fold drop in the Km value for PRPP·Mg2+. These observations suggest the involvement of ligand-mediated oligomerization and conformational changes in the process of activation. The dipeptide Leu–Lys in the PPi binding site of human and T. gondii HG(X)PRT that facilitates PRPP·Mg2+ binding by isomerization from trans to cis conformation is conserved in PfHGXPRT. Free energy calculations using the well-tempered metadynamics technique show the ligand-free enzyme to be more stable when this dipeptide is in the trans conformation than in the cis conformation. The high rotational energy barrier observed for the conformational change from experimental and computational studies permits delineation of the activation mechanism.
TL;DR: The first on-flow characterization of tight binders of xanthine oxidase is described, including 4CBALO, an allopurinol ruthenium derivative, which exhibited 100% inhibition at 200 μM compared to 86% inhibition from allopURinol at the same concentration.
Abstract: Xanthine oxidase (XO) is an enzyme in the purine salvage pathway that catalyzes the oxidation of hypoxanthine to xanthine with subsequent production of uric acid from the xanthine oxidation, and it has been considered an important target of newly developed inhibitors. Based on the advantages of using immobilized capillary enzyme reactors (ICERs) in a 2D LC system as a tool for screening new enzymatic ligands, this work validated an XO-ICER using allopurinol as a positive control. Despite the complex interaction between XO and allopurinol due its tight binding nature, it was possible to recognize the inhibitory kinetics parameters through Morrison's equation. The tight binding nature of inhibition was established by varying the IC50 values according to the substrate concentration. The kinetic inhibitory profile of allopurinol was used to validate the XO-ICER. Then, the XO-ICER was used to screen specific ruthenium derivatives. The selected compound, 4CBALO, an allopurinol ruthenium derivative, exhibited 100% inhibition at 200 μM compared to 86% inhibition from allopurinol at the same concentration. The inhibitory effect on the immobilized XO was reversible after the elution of the compound, with immediate recovery of the ICER activity. Additionally, 4CBALO behaved as a selective and competitive tight binder of xanthine oxidase with a true Ki value of 0.29 μM, which was obtained from the Morrison equation. This report describes the first on-flow characterization of tight binders of xanthine oxidase.
TL;DR: In this article, the xanthine, despite the destabilization of hemoglobin, guarantees the stability of phosphorylation-dephosphorylation balance, the caspase silence, and the integrity of cytoplasmic domain of band 3 protein promoting several positive metabolic conditions, resulting in an increase of cellular reducing power.
Abstract: Caffeine, as many molecules of the vegetable world, acting on a wide range of molecular targets, is able to exert its beneficial effects on the metabolic function in the human body. Several caffeine effects derive from its action as an antagonist of adenosine receptors, many others from its antioxidant properties. Particularly, in the erythrocytes the xanthine, despite the destabilization of hemoglobin, guarantees the stability of phosphorylation–dephosphorylation balance, the “caspase silence,” and the integrity of cytoplasmic domain of band 3 protein promoting several positive metabolic conditions, resulting in an increase of the cellular reducing power. All these findings associated with the strong scavenger activity of caffeine toward hydroxyl radical already evident at low doses of the alkaloid could be connected with the positive effects of xanthine with respect to the risk of neurodegenerative diseases, conditions now mostly associated with oxidative stress.