Hesperitin-Copper(II) Complex Regulates the NLRP3 Pathway and Attenuates Hyperuricemia and Renal Inflammation.

BACKGROUND: Hyperuricaemia (HUA) is a disorder of purine metabolism in the body. We previously synthesized a hesperitin (Hsp)-Cu(II) complex and found that the complex possessed strong uric acid (UA)-reducing activity in vitro. In this study we further explored the complex's UA-lowering and nephroprotective effects in vivo. METHODS: A mouse with HUA was used to investigate the complex's hypouricemic and nephroprotective effects via biochemical analysis, RT-PCR, and Western blot. RESULTS: Hsp-Cu(II) complex markedly decreased the serum UA level and restored kidney tissue damage to normal in HUA mice. Meanwhile, the complex inhibited liver adenosine deaminase (ADA) and xanthine oxidase (XO) activities to reduce UA synthesis and modulated the protein expression of urate transporters to promote UA excretion. Hsp-Cu(II) treatment significantly suppressed oxidative stress and inflammatory in the kidney, reduced the contents of cytokines and inhibited the activation of the nucleotide-binding oligomerization domain (NOD)-like receptor thermal protein domain associated protein 3 (NLRP3) inflammatory pathway. CONCLUSIONS: Hsp-Cu(II) complex reduced serum UA and protected kidneys from renal inflammatory damage and oxidative stress by modulating the NLRP3 pathway. Hsp-Cu(II) complex may be a promising dietary supplement or nutraceutical for the therapy of hyperuricemia.

Improving the gel properties of salted egg white/cooked soybean protein isolate composite gels by ultrasound treatment: Study on the gelling properties and structure.

In this study, the effects of ultrasound treatment on the texture, physicochemical properties and protein structure of composite gels prepared by salted egg white (SEW) and cooked soybean protein isolate (CSPI) at different ratios were investigated. With the increased SEW addition, the ζ-potential absolute values, soluble protein content, surface hydrophobicity and swelling ratio of composite gels showed overall declining trends (P < 0.05), while the free sulfhydryl (SH) contents and hardness of exhibited overall increasing trends (P < 0.05). Microstructural results revealed that composite gels exhibited denser structure with the increased SEW addition. After ultrasound treatment, the particle size of composite protein solutions significantly decreased (P < 0.05), and the free SH contents of ultrasound-treated composite gels were lower than that of untreated composite gels. Moreover, ultrasound treatment enhanced the hardness of composite gels, and promoted the conversion of free water into non-flowable water. However, when ultrasonic power exceeded 150 W, the hardness of composite gels could not be further enhanced. FTIR results indicated that ultrasound treatment facilitated the composite protein aggregates to form a more stable gel structure. The improvement of ultrasound treatment on the properties of composite gels was mainly by promoting the dissociation of protein aggregates, and the dissociated protein particles further interacted to form denser aggregates through disulfide bond, thus facilitating the crosslinking and reaggregation of protein aggregates to form denser gel structure. Overall, ultrasound treatment is an effective approach to improve the properties of SEW-CSPI composite gels, which can improve the potential utilization of SEW and SPI in food processing.

Treatments of heating and ultrasound improve the inhibition of gallocatechin gallate on tyrosinase.

BACKGROUND: Gallocatechin gallate (GCG), a catechin of tea polyphenols, possesses inhibitory ability against tyrosinase, but few studies have reported how common processing methods affect it. In this research, the influence of heating and ultrasound treatments on the inhibition of GCG against tyrosinase was explored by ultraviolet-visible absorption, fluorescence spectroscopy, high-performance liquid chromatography and liquid chromatography-tandem mass spectrometry. RESULTS: Both heating and ultrasound treatments of GCG alone improved GCG's inhibitory ability against tyrosinase compared with the untreated, and a combination of heating and ultrasound treatment (100 °C, 20 min + 630 W, 20 min) further decreased the relative tyrosinase activity to 26.8%. The treated GCG exhibited a stronger fluorescence quenching effect on tyrosinase, but did not have any influence on the static quenching mechanism. Compared to the untreated GCG, the binding constants of treated GCG by heating, ultrasound and their combination with tyrosinase significantly increased, but the number of binding sites was still approximately one and the main driving force of the treated GCG was still hydrophobic interaction. After treatments of heating, ultrasound and their combination, the composition of GCG solutions was changed. CONCLUSION: The enhanced inhibition of treated GCG on tyrosinase may be due to partial conversion of GCG into epigallocatechin-3-gallate (EGCG) and gallic acid (GA), which may cooperate with GCG to better inhibit the enzyme activity. This study has provided some valuable information for the application of catechins against tyrosinase in food processing and cosmetic industry. © 2022 Society of Chemical Industry.

Action mechanisms of two key xanthine oxidase inhibitors in tea polyphenols and their combined effect with allopurinol.

BACKGROUND: Tea polyphenols have been reported to have the effect of lowering uric acid. However, there are few studies on the inhibitory effects and molecular mechanisms of specific catechins on the urate-metabolizing enzyme xanthine oxidase (XO). In this research, multiple spectroscopic methods and computer simulations were used to determine the inhibitory ability and mechanisms of epigallocatechin gallate (EGCG) and gallocatechin gallate (GCG) on XO. RESULTS: Herein, EGCG and GCG reversibly inhibited XO activity in a mixed manner, with IC50 values of 40.50 ± 0.32 and 33.60 ± 0.53 µmol L-1 , and also decreased the superoxide anion radical (O2 - ) of the catalytic system by reducing the XO molecule and inhibiting the formation of uric acid. The combination of EGCG or GCG with allopurinol showed synergistic inhibition on XO. The binding of EGCG or GCG to XO with moderate affinity formed a stable complex by hydrogen bonds and van der Waals forces. The presence of EGCG and GCG made the structure of XO more stable and compact. The two inhibitors bound to the vicinity of flavin adenine dinucleotide (FAD) in XO, hindering the entry of substrate; thus the activity of XO was suppressed. CONCLUSION: Both EGCG and GCG are excellent natural XO inhibitors, and inhibited the activity of XO by occupying the channel of the substrate to enter the active center and interfering with the dual substrate reaction catalyzed by XO. These findings provide a scientific basis for the application of catechins in dietary supplements and medicines with lowering uric acid effects. © 2022 Society of Chemical Industry.

Improvement of gel properties and digestibility of the water-soluble polymer of tea polyphenol-egg white under thermal treatment.

In this study, the improvement of gel properties and digestibility of the water-soluble polymer of tea polyphenol (TP)-egg white protein (TEP) under heat induction (HTEP), was studied. Results indicated that the particle size and turbidity of TEP increased with TP concentration, and the absolute value of ζ-potential decreased. After heat induction, the surface hydrophobicity of HTEP decreased with TP concentration, and the degree of protein aggregation increased. Microstructure and T2 showed that the gel structure became compact and stable, and HTEP had a strong water-binding ability. The ionic and disulfide bonds were the main chemical bonds in HTEP. The hardness and disulfide bond increased, but the digestion of HTEP increased initially and then decreased (caused by the change of gel structure). Infrared spectroscopy indicated the mutual conversion of intermolecular and intramolecular ß-sheets. In short, TP could modify egg white gel through forming stable disulfide bonds and dense gel network structures.

Effects of stewing with tea polyphenol on the gel properties, microstructure, and secondary structure of boiled egg white.

This study aimed to investigatethe mechanism of stewing with tea polyphenols (TP) on the properties of boiled egg white gel (BEWG). The results indicated that, during the stewing process, soluble protein and hardness showed an overall increasing trend, while surface hydrophobicity showed a decreasing trend with blue-shift. The free sulfhydryl group showed that TP could promote the formation of disulfide bonds, and the position of immobilized water at T2 showed a decreasing trend. Environmental scanning electron microscopy and SDS-PAGE showed that the protein gel aggregation degree increased. Moreover, Fourier transform infrared spectrometry showed that protein polarity increased and that α-helices, ß-turn, intramolecular ß-sheets, as well as intermolecular antiparallel ß-sheets showed an increasing trend. Generally, TP strengthened protein aggregation by promoting the formation of disulfide and hydrogen bonds, thus enhancing the gel strength of BEWG. Moreover, the secondary structure of proteins became more stable under the action of TP, and the higher the concentration of TP, the greater the effect on BEWG. PRACTICAL APPLICATION: TP, an ideal, cheap, and safe natural food additive, can be applied to the processing of egg products because the addition of TP can significantly improve the gel strength of egg white.

Inhibitory effect of epicatechin gallate on protein glycation.

Protein glycation can produce the advanced glycation end products (AGEs), which has a potential health risk due to its relevance with diabetic complications. Searching for potent inhibitors to suppress hyperglycemia-induced protein glycation is of great significance. Here, we probed the anti-glycation potential of epicatechin gallate (ECG), an important ingredient of green tea, by spectroscopy and chromatography supplemented by molecular docking. It was found that ECG suppressed the carbonylation and the formation of amyloid cross-ß structures of bovine serum albumin (BSA) as well as the generation of AGEs in BSA-fructose model by 65.6%, 73.7% and 87.0%, resepctively. A combination of ECG with epigallocatechin gallate (EGCG) at molar ratios from 0.5:1 to 2:1 exhibited the interactive effects on AGEs formation from moderate antagonism to synergy. Moreover, ECG at 44.2 µg/mL showed a trapping efficiency of 81.6% for the important precursor methylglyoxal (MGO) of AGEs within 24 h, and the formed ECG-mono-MGO and ECG-di-MGO adducts were verified by LC-QqQ-MS analysis. The inhibitory effect of ECG on the protein glycation may be due to its MGO trapping and the specific binding with certain amino acid residues of BSA. These findings may provide the theoretical basis for the development of ECG as an inhibitor of protein glycation.

New insights into the binding mechanism between osthole and ß-lactoglobulin: Spectroscopic, chemometrics and docking studies.

Osthole, a type of coumarin derivative, owns many biological functions, but the poor water solubility and low bioavailability limit its usage in food and pharmaceutical fields. ß-Lactoglobulin (BLG) is a major protein in bovine milk whey which is usually used as a carrier of poorly soluble bioactive components. Researching the interaction between osthole and BLG can help us in understanding their binding mechanism and design an osthole delivery system using BLG as carrier protein to further promote the application of osthole in functional food and drug. Hence, this study was devoted to explore the binding properties of osthole with BLG and the effect on the protein structure by employing multispectroscopic approaches, chemometrics and molecular simulation studies. The results of the concentration profiles and pure spectra of the components resolved from highly overlapping spectral signals by the multivariate curve resolution-alternating least squares (MCR - ALS) algorithm verified the formation of an osthole-BLG complex. Moreover, osthole was found to strong quench the intrinsic fluorescence of BLG and the quenching mechanism was determined to be a static procedure. The binding constant of osthole with BLG at 298 K was (4.06 ±â€¯0.03) × 104 L mol-1. Hydrophobic forces and hydrogen bonds played major roles in the binding process. The analysis of synchronous fluorescence, circular dichroism and Fourier transform infrared spectra found that the presence of osthole caused BLG structure compact and the change in the polarity and hydrophobicity of the microenvironment around Tyr residues of the protein. Modeling docking predicted that osthole bound to the hydrophobic cavity of BLG through stable hydrogen bonds primarily with the amino acid residues Lys75 and Thr76.

Mechanistic insights into the inhibition of quercetin on xanthine oxidase.

Quercetin, one of the most abundant flavonoid in the daily diet, was found to reversibly inhibit the generation of uric acid and superoxide radicals (O2-)catalyzed by xanthine oxidase (XOD) in a mixed-type manner with IC50 values of (2.74±0.04)×10-6 and (2.90±0.03)×10-6molL-1, respectively, and the inhibition of quercetin on O2- generation may be ascribed to the reduced form of XOD by a ping-pong mechanism. XOD had one high affinity binding site for quercetin with a binding constant of 4.28×104Lmol-1 at 298K, and the binding process was predominately driven by van der Waals forces and hydrogen bonds on account of the negative enthalpy and entropy changes. Moreover, molecular docking confirmed that the binding site for quercetin located in the isoalloxazine ring of the flavin adenine dinucleotide (FAD) domain of XOD, then the diffusion of O2- out of the FAD site was blocked in favor of another electron transferred from FADH2 to O2- to form hydrogen peroxide (H2O2). This study may clarify the role of quercetin on inhibiting XOD catalysis and provide a potential nutritional supplement for preventing gout and peroxidative damage.

Inhibitory mechanism of morin on α-glucosidase and its anti-glycation properties.

It is important to investigate the inhibition of α-glucosidase due to its correlation with type 2 diabetes. Morin was found to exert significant inhibition activity on α-glucosidase in a reversible mixed-type manner with an IC50 value of (4.48 ± 0.04) µM. Analyses of fluorescence and circular dichroism spectra indicated that the formation of the morin-α-glucosidase complex was driven mainly by hydrophobic forces and hydrogen bonding, and caused the conformational changes of α-glucosidase. The phase diagrams of fluorescence showed that the conformational change process was monophasic without intermediates. Molecular docking indicated that morin mainly interacted with amino acid residues located close to the active site of α-glucosidase, which may move to cover the active pocket to reduce the binding of the substrate and then inhibit the catalytic activity. Morin was also found to exhibit inhibition in the generation of advanced glycation end products which was related to the long term complications of diabetes.

The inhibitory kinetics and mechanism of dietary vitamins D3 and B2 on xanthine oxidase.

Dietary guidelines to promote health are usually based on the patterns' prediction on disease risk of foods and nutrients. Overactivity of xanthine oxidase (XO) is the underlying cause of gout. Herein, the inhibitory kinetics and mechanism of dietary vitamins D3 and B2 on XO were investigated by multispectroscopic methods and a molecular modeling technique. The results showed that vitamin D3 competitively inhibited XO with an inhibition constant of 26.93 ± 0.42 µM by inserting into the active cavity of XO interacting with the surrounding amino acid residues through hydrogen bond and van der Waals forces. Vitamin D3 bound to XO thereby induced the structural compactness of XO which in turn hindered the binding of substrate xanthine to cause the inhibition on XO. Vitamin B2 exhibited a mixed-type inhibition by binding to the vicinity of the active cavity with an inhibition constant of 37.76 ± 0.87 µM through hydrophobic interactions and a feeble hydrogen bond, and it induced the unfolding of the XO structure and an increase of the flexible loops (ß-turns and random coils) which might move to cover the active pocket and reduce the binding of the substrate xanthine, and then lead to a lower catalytic activity of the enzyme. In addition, vitamins D3 and B2 showed a synergistic effect on inhibiting the activity of XO in a certain range of concentration. These findings may provide new insights into the inhibitory mechanism of vitamins D3 and B2 on XO and functional research of the vitamins in the supplementary treatment of gout.

Organochlorine pesticides and pyrethroids in Chinese tea by screening and confirmatory detection using GC-NCI-MS and GC-MS/MS.

One hundred and one tea samples including green tea, dark tea, scented tea, black tea, and oolong tea were screened and confirmed for the contamination of 31 organochlorine pesticides (OCPs) and 19 pyrethroids (PYs) by gas chromatography-negative chemical ionization-mass spectrometry (GC-NCI-MS) and gas chromatography-tandem mass spectrometry (GC-MS/MS). 50 pesticides, 3 deuterium-labeled PYs, and 24 (13)C-labeled OCPs were separated well with the limits of detection (LODs) ranging from 0.02 to 4.5 µg/kg for GC-NCI-MS, and the positive samples were verified by GC-MS/MS with LODs of 0.1-5.0 µg/kg. High detection rates for some PYs, such as 63.4% for bifenthrin (not detected (ND)-3.848 mg/kg), 55.4% for λ-cyhalothrin (ND-3.244 mg/kg), 46.5% for cypermethrin (ND-0.499 mg/kg), and 24.8% for fenvalerate (ND-0.217 mg/kg), were found in the 101 tea samples. Endosulfan, DDTs, HCHs, and heptachlor, the persistent OCPs, were frequently detected with rates of 63.4% (ND-1.802 mg/kg), 56.4% (ND-0.411 mg/kg), 24.8% (ND-0.377 mg/kg), and 15.8% (ND-0.100 mg/kg), respectively.

Effect of luteolin on xanthine oxidase: inhibition kinetics and interaction mechanism merging with docking simulation.

Xanthine oxidase (XO) catalyses hypoxanthine and xanthine to uric acid in human metabolism. Overproduction of uric acid will lead to hyperuricemia and finally cause gout and other diseases. Luteolin is one of the major components of celery and green peppers, its inhibitory activity on XO and their interaction mechanism were evaluated by multispectroscopic methods, coupled with molecular simulation. It was found that luteolin reversibly inhibited XO in a competitive manner with inhibition constant (Ki) value of (2.38±0.05)×10(-6) mol l(-1). Luteolin could bind to XO at a single binding site and the binding was driven mainly by hydrophobic interactions. Analysis of synchronous fluorescence and circular dichroism spectra demonstrated that the microenvironment and secondary structure of XO were altered upon interaction with luteolin. The molecular docking results revealed luteolin actually interacted with the primary amino acid residues located within the active site pocket of XO.

Interaction of alpinetin with bovine serum albumin: Probing of the mechanism and binding site by spectroscopic methods.

The binding interaction between alpinetin and bovine serum albumin (BSA) in physiological buffer solution (pH 7.4) was investigated by fluorescence, UV-vis spectroscopy and Fourier transform infrared (FT-IR) spectroscopy. It was proved from fluorescence spectra that the fluorescence quenching of BSA by alpinetin was probably a result of the formation of BSA-alpinetin complexes, and the binding constant (K(a)) were determined according to the modified Stern-Volmer equation. The thermodynamic parameters, enthalpy change (DeltaH) and entropy change (DeltaS), were calculated to be 22.10kJmol(-1) and 166.04Jmol(-1)K(-1), respectively, which indicated that the interaction between alpinetin and BSA was driven mainly by hydrophobic interaction. Moreover, the competitive experiments of site markers suggested that the binding site of alpinetin to BSA was located in the region of subdomain IIA (sudlow site I). The binding distance (r) between the donor (BSA) and the acceptor (alpinetin) was 3.32nm based on the Förster theory of non-radioactive energy transfer. In addition, the results of synchronous fluorescence and FT-IR spectra demonstrated that the microenvironment and the secondary structure of BSA were changed in the presence of alpinetin.