Mechanisms of theaflavins against gout and strategies for improving the bioavailability.

BACKGROUND: Gout is a crystal related arthropathy caused by monosodium urate deposition. At present, the identification of appropriate treatments and new drugs to reduce serum uric acid levels and gout risk is a major research area. PURPOSE: Theaflavins are naturally occurring compounds characterized by a benzodiazepine skeleton. The significant benefits of theaflavins have been well documented. A large number of studies have been carried out and excellent anti-gout results have been achieved in recent years. STUDY DESIGN: A comprehensive analysis of the mechanism of the anti-gout effect of theaflavins is presented through a literature review and network pharmacology prediction, and strategies for increasing the bioavailability of theaflavins are summarized. METHODS: In this review, the active components and pharmacological mechanisms of theaflavins in the treatment of gout were summarized, and the relationship between theaflavins and gout, the relevant components, and the potential mechanisms of anti-gout action were clarified by reviewing the literature on the anti-gout effects of theaflavins and network pharmacology. RESULTS: Theaflavins exert anti-gout effects by down regulating the gene and protein expression of glucose transporter 9 (GLUT9) and uric acid transporter 1 (URAT1), while upregulating the mRNA expression levels of organic anion transporter 1 (OAT1), organic cation transporter N1 (OCTN1), organic cation transporters 1/2 (Oct1/2), and organic anion transporter 2 (OAT2). Network pharmacology prediction indicate that theaflavins can regulate the AGE-RAGE and cancer signaling pathways through ATP-binding cassette subfamily B member 1 (ABCB1), recombinant mitogen activated protein kinase 14 (MAPK14), telomerase reverse tranase (TERT), signal transducer and activator of transcription 1 (STAT1), matrix metalloproteinase 2 (MMP2), B-cell lymphoma-2 (BCL2), and matrix metalloproteinase 14 (MMP14) targets for anti-gout effects. CONCLUSION: This review presents the mechanisms of anti-gout action of theaflavins and strategies for improving the bioavailability of theaflavins, as well as providing research strategies for anti-gout treatment measures and the development of novel anti-gout drugs.

Xanthine oxidase inhibitory activity and antihyperuricemic effect of Moringa oleifera Lam. leaf hydrolysate rich in phenolics and peptides.

ETHNOPHARMACOLOGICAL RELEVANCE: Moringa oleifera Lam. leaf (MOL), a rich source of protein and phenolics, was traditionally used to treat various diseases including headaches, fevers, sore throat and dyslipidemia. Recently, MOL was reported to possess antioxidant, anti-dyslipidemia and hepato-renal protective activities, indicating that MOL could become a potential agent to improve metabolic disorders associated with hyperuricemia. The antihyperuricemic effect of MOL hydrolysate (MOLH) with high contents of phenolics and peptides remains unknown. AIM OF THE STUDY: The aim of this study is to investigate xanthine oxidase (XO) inhibitory activity of MOLH, to clarify phenolic and peptide profiles of MOLH, and to evaluate possible mechanism underlying the antihyperuricemic effect of MOLH. MATERIALS AND METHODS: MOLH was prepared by enzymatic hydrolysis using commercial trypsin. XO inhibitory activity was determined by XO reaction-UPLC-MS coupling method. The chemical profiles of the phenolic and peptide fractions of MOLH were determined by UPLC-QTOF-MS/MS. The antihyperuricemic effect of MOLH was evaluated in a potassium oxonate-induced hyperuricemic rat model at doses of 200 and 500 mg/kg. Serum uric acid (UA), urea nitrogen, creatinine (CRE), triglyceride (TG), total cholesterol, high-density lipoprotein cholesterol and low-density lipoprotein cholesterol levels, serum XO activity, liver malondialdehyde (MDA) equivalent level, renal tumor necrosis factor-α and interleukin-1ß levels, and protein expression of renal urate-anion transporter 1, glucose transporter 9 and ATP-binding cassette transporter G2 were determined. RESULTS: The phenolic and peptide fractions played key roles in inhibiting XO activity and blocking uric acid production. Five flavonoids and sixteen polypeptides were identified in the phenolic and peptide fractions of MOLH, respectively. MOLH (200 and 500 mg/kg) could effectively reduce the serum UA level of hyperuricemic rats (p < 0.001) by regulation of serum XO activity (p < 0.05 at 200 mg/kg, p < 0.01 at 500 mg/kg) and renal urate transporters. Besides, MOLH could improve metabolic disorders associated with hyperuricemia by its multiple actions on liver MDA (p < 0.001), serum CRE (p < 0.05 at 500 mg/kg) and serum TG (p < 0.001). CONCLUSION: The results provided scientific evidence that MOLH rich in phenolics and peptides ameliorated hyperuricemia and metabolic disorders. This study validated the potential use of MOLH for regulation of hyperuricemia.

Biopharmaceutical characterization of a novel sustained-release formulation of allopurinol with reduced nephrotoxicity.

The present study was aimed to develop a novel sustained-release formulation for allopurinol (ALP/SR) with the use of a pH-sensitive polymer, hydroxypropyl methylcellulose acetate succinate, to reduce nephrotoxicity. ALP/SR was evaluated in terms of crystallinity, the dissolution profile, pharmacokinetic behavior, and nephrotoxicity in a rat model of nephropathy. Under acidic conditions (pH1.2), sustained release behavior was seen for ALP/SR, although both crystalline ALP and ALP/SR exhibited rapid dissolution at neutral condition. After multiple oral administrations of ALP samples (10 mg-ALP/kg) for 4 days in a rat model of nephropathy, ALP/SR led to a low and sustained plasma concentration of ALP, as evidenced by half the maximum concentration of ALP and a 2.5-fold increase in the half-life of ALP compared with crystalline ALP, possibly due to suppressed dissolution behavior under acidic conditions. Repeated-dosing of ALP/SR resulted in significant reductions in plasma creatinine and blood urea nitrogen levels by 73% and 69%, respectively, in comparison with crystalline ALP, suggesting the low nephrotoxic risk of ALP/SR. From these findings, a strategic SR formulation approach might be an efficacious dosage option for ALP to avoid severe nephrotoxicity in patients with nephropathy.

RDP3, A Novel Antigout Peptide Derived from Water Extract of Rice.

Gout and hyperuricemia can seriously affect the quality of life; at present, however, existing medicines are unable to meet all clinical needs. In the current study, a novel peptide (i.e., rice-derived-peptide-3 (RDP3), AAAAMAGPK-NH2, 785.97 Da) in water extract obtained from shelled Oryza sativa fruits was identified. Testing revealed that RDP3 (minimum effective concentration 100 µg/kg) did not show both hemolytic and acute toxicity, and reduced uric acid levels in the serum of hyperuricemic mice by inhibiting xanthine oxidase activity and decreasing urate transporter 1 expression. RDP3 also alleviated renal injury in hyperuricemic mice by decreasing NLRP3 inflammasome expression. Furthermore, RDP3 alleviated formalin-induced paw pain and reduced monosodium urate crystal-induced paw swelling and inflammatory factors in mice. Thus, this newly identified peptide reduced uric acid levels and renal damage in hyperuricemic mice and showed anti-inflammatory and analgesic activities, indicating the potential of RDP3 as an antigout medicine candidate.

Formulation and in vitro Evaluation of Fast Dissolving Tablets of Febuxostat Using Co-Processed Excipients.

BACKGROUND: Febuxostat is a novel, orally-administered, powerful, non-purine, xanthine oxidase inhibitor used for treating gout and ceaseless tophaceous gout. The drug exhibits low bioavailability (about 49%) which is ascribed to its dissolution rate-limited absorption. OBJECTIVE: The current work is aimed to provide a novel strategy to improve the dissolution profile and thus, the bioavailability of Febuxostat. METHODS: Formulation of Fast Dissolving Tablets (FDT) is anticipated to provide immediate release of the drug, which in turn, will improve its dissolution profile to provide the initial surge in plasma concentration required in an acute gout attack. Incorporation of co-processed excipients in a tablet is known to improve the compressibility and disintegration characteristics of the tablets, which, in turn, result in enhanced in vitro drug release and improved bioavailability. A combination of crospovidone (it rapidly wicks saliva into the tablet to create the volume development and hydrostatic weight important to give quick disintegration) and microcrystalline cellulose (a highly compressible ingredient with good wicking and absorbing capacity) was, therefore, used as co-processed excipients. RESULTS: The tablets were prepared by direct compression technique with the application of a 32 randomized full factorial design. The prepared tablets were able to release more than 80% of the drug within 10 minutes of the start of dissolution testing and were able to show a better drug release profile in comparison to available marketed formulation. CONCLUSION: So, it can be concluded that the developed fast release formulation was found to exhibit convincing in vitro results and may prove a boon in the treatment of acute gout attack after establishing in vivo potential.

Anti-Inflammatory and Anti-Hyperuricemic Functions of Two Synthetic Hybrid Drugs with Dual Biological Active Sites.

The present study aimed to test the anti-inflammatory and xanthine oxidase inhibitory activities of two synthesized molecules and compare them to routinely prescribed nonsteroidal anti-inflammatory drugs (NSAIDs), such as diclofenac and the serum urate-lowering drug, allopurinol. The anti-inflammatory effects of the designed compounds (A and B) were evaluated in carrageenan (CAR)-induced paw edema in mice. The levels of nitric oxide and myeloperoxidase activity were measured in paw skin using biochemical methods. Additionally, prostaglandin E2 (PGE2), C-reactive protein (CRP), cyclooxygenase-2 (Cox-2), tumor necrosis factor-α (TNFα), interleukin (IL)-1ß, IL-2 and IL-10, and monocyte chemoattractant protein-1 (MCP1) were assessed by enzyme-linked immunosorbent assay (ELISA). The expression of inflammation-related genes was confirmed by real-time qPCR. The expression of inducible nitric oxide synthase (iNOS) and nuclear factor-kappa B (NF-κB) were estimated using immunohistochemistry, and xanthine oxidase inhibitory activity was evaluated using an in vitro assay. The results revealed that compounds A and B decreased inflammation, as was observed by a reduction in the elevation of all the tested markers. In addition, the tested compounds markedly decreased paw swelling, mobilization of inflammatory cells, iNOS-, and NF-κB-immunoreactive cells in a mouse model of paw edema. Interestingly, both compounds were potent xanthine oxidase inhibitors as well as Cox inhibitors with higher activity in favor of compound B providing potential dual acting series of anti-hyperuricemic and anti-inflammatory therapeutic agents.

Novel Carbon Dots Derived from Puerariae lobatae Radix and Their Anti-Gout Effects.

Gout is a disease with a high incidence and causing great harm, and the current treatment drugs are not satisfactory. In this study, novel water-soluble carbon dots (CDs) with anti-gout effect, named Puerariae lobatae Radix CDs (PLR-CDs), are reported. PLR-CDs were synthesized with an improved pyrolysis method at 300 °C, and their characterization was performed with multifaceted approaches, such as transmission electron microscopy (TEM) and ultraviolet-visible (UV-vis) and Fourier-transform infrared (FTIR) spectroscopy. In addition, the biocompatibility of PLR-CDs was studied using the cell counting kit (CCK)-8 in LO2 cells and RAW264.7 cells, and the anti-gout activity of PLR-CDs was examined on animal models of hyperuricemia and gouty arthritis. The characterization of PLR-CDs indicated that they were nearly spherical, with diameters ranging from 3.0 to 10.0 nm, and the lattice spacing was 0.283 nm. The toxicity experiment revealed that PLR-CDs were non-poisonous for LO2 cells and RAW264.7 cells at concentrations below 250 µg/mL. The results of pharmacodynamic experiments showed that PLR-CDs could lower the blood uric acid level in model rats by inhibiting the activity of xanthine oxidase and reduce the degree of swelling and pathological damage of gouty arthritis. Thus, PLR-CDs with anti-gout biological activity and good biocompatibility have the prospect of clinical application for the treatment of gout.

Preparation, optimisation, and in vitro-in vivo evaluation of febuxostat ternary solid dispersion.

The research aimed to prepare febuxostat (FEB) solid dispersion through solvent evaporation. Optimised solid dispersion composed of FEB, polyvinylpyrrolidone (PVP K30) and poloxamer at a ratio of 1:3:3 was characterised. Powder X-ray diffraction (XRD) and differential scanning calorimetry (DSC) indicated FEB was transformed from crystalline into the amorphous state in solid dispersion and scanning electron microscopy (SEM) revealed the morphology. Fourier transform infrared spectroscopy (FT-IR) suggested the interactions formed between FEB and polymers. A remarkable increase was observed of the optimised formulation in saturation solubility, dissolution studies (96.17 ± 0.79% in pH 6.0), and bioavailability (Cmax 18.25 ± 2.44 vs. 7.72 ± 0.48 µg/mL and AUC0-∞ 53.62 ± 7.63 vs. 34.76 ± 2.45 µg·h/mL). Besides, the FEB solid dispersion showed great stability after 90 days storage. Thus, the present study supports the rationality of PVP K30 and poloxamer188 as co-carriers for the preparation of FEB solid dispersion.

S-allyl cysteine as potent anti-gout drug: Insight into the xanthine oxidase inhibition and anti-inflammatory activity.

S-allyl cysteine (SAC) is known for its various beneficial effects such as neuroprotection and immunomodulation. The beneficial effect of SAC against gout has not been explored. The present study aims to describe the two roles of SAC: (1) inhibitory effect against xanthine oxidase (XO) enzyme activity; and (2) anti-inflammatory property against MSU crystal-induced gouty inflammation in rat. The inhibitory effect of SAC against bovine XO enzyme activity was determined in vitro. In silico analysis was carried out to determine the intermolecular interaction between SAC and bovine XO. MSU crystal was injected in the right paw of the rat to induce gouty inflammation. SAC (40 mg/kg body weight) and colchicine (positive control; 1 mg/kg body weight) was given for 3 days. At the end of the treatment, the oxidative stress, antioxidant parameters and mitochondrial function were determined in the ankle joint tissue. The concentration of inflammatory cytokines such as TNF-α and IL-1ß was measured in the serum using ELISA. SAC inhibited (IC50 value, 33 µg/ml) XO enzyme activity in a competitive mode with corresponding Ki value of 4 µg/ml. In silico analysis predicted the interaction of SAC with the amino acids such as Arg880, Phe798, Phe914 and Phe1009 of XO enzyme. The root mean square deviation, root mean square fluctuation and free energy calculation values confirmed the stable SAC-XO interaction. The inhibition of SAC on XO enzyme activity in in vivo was further confirmed by silkworm model. SAC through reducing oxidative stress, enhancing antioxidants, protecting mitochondrial function has shown anti-inflammatory effect against MSU crystal-induced gout which was observed as reduced level of inflammatory markers in the serum. The medicinal potential of SAC as a preventive agent through its XO inhibitory property as well as curative agent through its anti-inflammatory property against gout has been understood from the present study.

Anti-Nociceptive Effect and Standardization from Mixture of Agrimonia pilosa Ledeb and Salvia miltiorrhiza Bunge Extracts.

Agrimonia pilosa Ledeb has been previously reported to produce an anti-nociceptive effect in ICR mice in both tail-flick and hot-plate tests. Studies have shown that Salvia miltiorrhiza Bunge, also renowned in traditional Chinese medicine, is an effective anti-inflammatory treatment. Among the extraction solvents investigated, a 50% ethanol (EtOH) extract of A. pilosa produced the highest anti-nociceptive effect in monosodium uric acid-induced gout pain models and the greatest yield. The 80% EtOH extract of S. miltiorrhiza moderately inhibited lipopolysaccharide-induced nitric oxide release from RAW 264.7 murine macrophages and exhibited outstanding yield. The mixture of optimized A. pilosa and S. miltiorrhiza extracts were evaluated for enhanced anti-nociceptive effects in gout arthritis treatment. To control extract quality, four marker compounds were determined using an HPLC-DAD method. A 1:1 mixture of A. pilosa 50 and S. miltiorrhiza 80% EtOH extracts of produced better results than when the extracts were administered individually.

Xanthine Oxidase Inhibitory Activity and Thermostability of Cinnamaldehyde-Chemotype Leaf Oil of Cinnamomum osmophloeum Microencapsulated with ß-Cyclodextrin.

The xanthine oxidase inhibitory activity and thermostability of Cinnamomum osmophloeum leaf oil microencapsulated with ß-cyclodextrin were evaluated in this study. The yield of leaf oil microcapsules was 86.3% using the optimal reaction conditions at the leaf oil to ß-cyclodextrin ratio of 15:85 and ethanol to water ratio ranging from 1:3 to 1:5. Based on the FTIR analysis, the characteristic absorption bands of major constituent, trans-cinnamaldehyde, were confirmed in the spectra of leaf oil microcapsules. According to the dry-heat aging test, ß-cyclodextrin was thermostable under the high temperature conditions, and it was beneficial to reduce the emission of C. osmophloeum leaf oil. Leaf oil microcapsules exhibited high xanthine oxidase inhibitory activity, with an IC50 value of 83.3 µg/mL. It is concluded that the lifetime of C. osmophloeum leaf oil can be effectively improved by microencapsulation, and leaf oil microcapsules possess superior xanthine oxidase inhibitory activity.

Potential cow milk xanthine oxidase inhibitory and antioxidant activity of selected phenolic acid derivatives.

Xanthine oxidase (XO) found in all mammals and excess activity leads to urolithiasis. The cow milk XO was purified to 305-fold with a specific activity of 8.76 EU/mg and overall yield of 87% by using DEAE-Sepharose chromatography. The phenolics showed potent XO inhibitory effect with Ki , P1 (0.412), P2 (0.632), P3 (0.585), P4 (0.886), P5 (1.633), P6 (0.503), P7 (2.882), P8 (3.761), P9 (4.487), and P10 (5.841) µM. The phenolics P9 and P10 exhibited uncompetitive inhibition; the phenolics P1, P2, P3, P4, and P6 showed competitive inhibition, and other phenolic acids showed noncompetitive inhibition. The studied phenolic compounds showed potent antioxidant activity and expressed as EC50 , ranged from, DPPH (4.2-25.8 µg mL-1 ), ABTS (10.2-42.5 mmol TE 100 g-1 ), and FRAP (6.3-36.8 mol Fe (II) 100 g-1 ). The results obtained from this study might be utilized for design of XO inhibitors and as antigout agent.

Synthesis and evaluation of sulfonamide derivatives as potent Human Uric Acid Transporter 1 (hURAT1) inhibitors.

This letter presents synthesis and structure-activity relationship study of sulfonamide derivatives as inhibitors of Human Uric Acid Transporter 1 (hURAT1). Among all tested sulfonamide derivatives, compounds 9b, 16i and 19b exhibited excellent inhibition activity with IC50 value of 10, 2, and 83nM, respectively. In addition, compounds 9b and 19b demonstrated moderate PK profile in rats.

Phase behavior, intermolecular interaction, and solid state characterization of amorphous solid dispersion of Febuxostat.

The aim of this work was to prepare and characterize the amorphous molecular dispersion of Febuxostat (FXT) using PVP K30, HPMC-AS, Soluplus®, and PVP VA64. The solid dispersions were prepared by solvent evaporation technique. Their physical properties were studied by differential scanning calorimetry, powder X-ray diffraction, Fourier transformation infrared spectroscopy, and compared to that of same physical mixtures. The success of physicochemical stability of the dispersions is often revealed as glass transition temperature (Tg) versus composition (w) dependencies. The shape of the Tg versus composition was mathematically modeled using the Gordon-Taylor equation, Couchman-Karasz equation, Brekner-Schneider-Cantow equation, and a three-parameter BCKV equation. In this work, different types of Tg patterns obtained for FXT-polymer binary mixtures are analyzed in terms of the above equations and relations between their prime fitting parameters are presented. The theoretical values and modeled parameters were compared using various results obtained by thermal analysis. The influence of important physicochemical phenomena and properties of the mixtures on the shape of the Tg versus composition patterns are also illustrated. The interaction between drug and polymers and the model parameters were analyzed, aiming to assess the state of mixing and intermolecular interactions.

Charge transfer interaction of organic p-acceptors with the anti-hyperuricemic drug allopurinol: Insights from IR, Raman, 1H NMR and 13C NMR spectroscopies.

The topic of charge-transfer (CT) complexation of vital drugs has attracted considerable attention in recent years owing to their significant physical and chemical properties. In this study, CT complexes derived from the reaction of the anti-hyperuricemic drug allopurinol (Allop) with organic p-acceptors [(picric acid (PA), dichlorodicyanobenzoquinone (DDQ) and chloranil (CHL)] were prepared, isolated and characterized by a range of physicochemical methods, such as IR, Raman, 1H NMR and 13C NMR spectroscopy. The stoichiometry of the complexes was verified by elemental analysis. The results show that all complexes that were formed were based on a 1:1 stoichiometric ratio. This study suggests that the complexation of Allop with either the DDQ or CHL acceptor leads to a direct p®p* transition, whereas the molecules of Allop and PA are linked by intermolecular hydrogen- bonding interactions.

Identification of antihyperuricemic peptides in the proteolytic digest of shark cartilage water extract using in vivo activity-guided fractionation.

A peptide that exerts antihyperuricemic activity after oral administration was identified from a microbial protease (alcalase) digest of the water extract of shark cartilage by in vivo activity-guided fractionation, using oxonate-induced hyperuricemic rats. Water extract of shark cartilage was first fractionated by preparative ampholine-free isoelectric focusing, followed by preparative reversed-phase liquid chromatography. The antihyperuricemic activity of the alcalse digests of the obtained fractions was evaluated using an animal model. Alcalase digests of the basic and hydrophobic fractions exerted antihyperuricemic activity. A total of 18 peptides were identified in the alcalase digest of the final active fraction. These peptides were chemically synthesized and evaluated for antihyperuricemic activity. Tyr-Leu-Asp-Asn-Tyr and Ser-Pro-Pro-Tyr-Trp-Pro-Tyr lowered the serum uric acid level via intravenous injection at 5 mg/kg of body weight. Furthermore, orally administered Tyr-Leu-Asp-Asn-Tyr showed antihyperuricemic activity. Therefore, these peptides are at least partially responsible for the antihyperuricemic activity of the alcalase digest of shark cartilage.

Pharmacokinetics and comparative bioavailability of allopurinol formulations in healthy subjects.

Allopurinol is the most commonly used urate-lowering therapy in gout. This study was undertaken to evaluate the pharmacokinetics and relative bioavailability of two brands of allopurinol tablets. The in vivo study was established according to a single-center, randomized, single-dose, laboratory-blinded, Two Way, Cross-Over Study with a washout period of 1 week. Under fasting conditions, 24 healthy male volunteers were randomly allocated to receive a single oral dose (200 mg) of either test and reference formulations. Plasma samples were obtained over a 6-hour interval and analyzed for allopurinol by reversed phase liquid chromatography with ultraviolet detection. The 90% confidence intervals for the ratio of log transformed values of Cmax , AUC0-t , and AUCt-∞ of the two treatments were within the acceptable range (0.8-1.25) for bioequivalence. From PK perspectives, the two allopurinol formulations were considered bioequivalent, based on the rate and extent of absorption. No adverse events occurred or were reported after a single 200-mg allopurinol and both formulations were well tolerated.

Lack of efficacy during the switch from brand to generic allopurinol.

We report for the first time the lack of therapeutic effects after the switch from a brand formulation of allopurinol to a generic one. A 56-year-old man, with a 5 years history of well-treated gout arthropathy with allopurinol (Zyloric(®) 300 mg/die), developed acute gout arthropathy after the switch from the brand formulation of allopurinol to a generic one. Clinical evaluation and laboratory findings confirmed the diagnosis of acute gout arthropathy. Generic formulation of the drug was dismissed and Zyloric(®) was administered with an improvement of both clinical symptoms and laboratory findings. In conclusion, even if generic formulations are considered to have the same effects in comparison to the brand one, more data are necessaries in order to well define their effectiveness and rationale use.

Colchicine revisited.

Purified from a Mediterranean plant nearly two centuries ago, colchicine has been discovered to inhibit many steps in the inflammatory process. The drug has good oral bioavailability and some enterohepatic recirculation, requiring dose adjustments for kidney disease and avoidance in liver disease. Toxicities are primarily gastrointestinal, hepatic, and hematologic. Colchicine is approved by the U.S. Federal Drug Administration for the treatment and prophylaxis of gout flares but has also been tried with varying success in the treatment of familial Mediterranean fever, primary biliary cirrhosis, psoriasis, Behçet's disease, aphthous stomatitis, linear IgA dermatosis, relapsing polychondritis, Sweet's syndrome, scleroderma, amyloidosis, leukocytoclastic vasculitis, epidermolysis bullosa, and dermatomyositis.