Polymethoxyflavones Research Articles

Effects of spray-drying temperature on the physicochemical properties and polymethoxyflavone loading efficiency of citrus oil microcapsules

A polymethoxyflavone (PMF)-loaded citrus oil emulsion was prepared as microcapsules via spray-drying. The effects of spray-drying temperature ranges from 110 °C to 190 °C on the physical properties of the citrus oil microcapsule powder, encapsulation efficiency of PMF and the alteration of citrus oil aroma were investigated in this study. The microcapsule powder formed at 150 °C spray-drying temperature possessed low moisture content (1.7%, w.b.), high powder yield (51.4%), good particle integrity and the corresponding reconstituted emulsion had a small droplet size. The encapsulation efficiency of PMF in microcapsule powder decreased as the spray-drying temperature increased from 110 °C to 190 °C (75.8%→45.2%). Electronic nose and GC-MS measurements revealed that the spray-dried powder was close to the aroma of the original citrus oil, although partial volatile components were lost when spray-drying. The results from this study are expected to facilitate the development of PMF-loaded citrus oil microcapsule products for functional foods.

Polymethoxyflavones as food factors for the management of inflammatory diseases

Recent researches have expanded the concept that inflammation is a critical component of human diseases. The possible mechanisms by which inflammation contribute to various conditions include increase of reactive species and inflammatory cytokines and growth factors leading to alteration of biomolecules, induction of genetic change, cellular dysfunction and enhancement of transformation or proliferation. Thus, targeting the inflammation is potential strategy for prevention of chronic diseases. Numerous epidemiological and laboratory studies suggest that polymethoxyflavones (PMFs) exhibit a broad spectrum of biological activities. Recently, we isolated and identified hydroxylated PMFs from citrus peels and have investigated their biological activities, including anti-inflammation and cancer chemopreventive property. We suggest that hydroxylated PMFs possess greater biological activity potency for chemoprevention on targeting inflammation.

Seasonal Effect of Ultraviolet Irradiation on Polymethoxyflavone and Hesperidin Content in Ponkan and Tachibana Flavedo

Ponkan (Citrus reticulate) and Tachibana (Citrus tachibana) contain large amounts of polymethoxyflavones (PMFs). To produce highly functional food with high potential, it is important to develop technologies to increase their PMF content. Nobiletin content of the flavedo changed in harvested ‘Ohta Ponkan’ fruits after ultraviolet (UV)-C irradiation and PMF content increased for only the first 2 days following irradiation. The effect of UV-C or UV-B irradiation on PMF content at each harvest time (UV-C, July to December; UV-B, July to September) was observed in Ponkan and Tachibana. We found that UV-C had the greatest impact earlier in the season. The effects of UV-B irradiation were similar to UV-C results. Therefore, it is beneficial to harvest early and extract PMFs from fruit 24 hours after UV-C irradiation.

Multi-Component Comparative Pharmacokinetics in Rats After Oral Administration of Fructus aurantii Extract, Naringin, Neohesperidin, and Naringin-Neohesperidin.

Citrus × aurantium L., Chinese name: Fructus Aurantii (FA) has been largely used as Qi-invigorating herb in China for centuries. The main components (meranzin hydrate, naringin, neohesperidin, meranzin, nobiletin) have good physiological activity with relatively high abundance in FA. Few multi-component comparative pharmacokinetics are simultaneously accessible for the flavone glycosides, polymethoxy flavones, and coumarins in FA. In this work, a reliable and rapid ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was established and validated to determine the five ingredients in the SD rat plasma, and further applied to the pharmacokinetic studies after oral administration of monomer, drugs in compatibility, and FA extract. After hydrolysis with β-glucuronidase and sulfatase, the concentration of naringin and neohesperidin in rat plasma were expressed respectively by the total concentration of naringenin and hesperitin which was determined by UPLC-MS/MS. Double-peak phenomenon was observed for naringin and neohesperidin, which may be due to the enterohepatic circulation or multiple site absorption of the two flavone glycosides. Meranzin hydrate and meranzin (coumarins) were absorbed rapidly (Tmax, about 1.0 h) but eliminated slowly (t1/2z exceeds 6.5 h). Nobiletin, a typical polymethoxy flavone, was also rapidly absorbed according to Tmax and AUC(0-t). DAS 3.1 software suggests the pharmacokinetic profiles of the five components in rats be depicted as a two-compartment pharmacokinetic model. There were significant differences in pharmacokinetic parameters for naringenin and hesperetin between the compatibility, FA extract group vs monomer group: ① remarkable increases in the values of AUC(0–∞), AUC(0–t) and Cmax; ② obvious decrease of CLZ/F; and ③ longer tmax and t1/2z. The results suggest that compatibility can promote mutual absorption and affect the elimination behaviors.

Identification of Xanthomicrol as a Major Metabolite of 5-demethyltangeretin in Mouse Gastrointestinal Tract and Its Inhibitory Effects on Colon Cancer Cells

Abstract Objectives Biotransformation may greatly affect the biological properties of dietary compounds. The metabolites generated in the body by the biotransformation may have different bioactivities in comparison with the ingested parent compounds. Polymethoxyflavones (PMFs) are natural bioactive flavonoids almost exclusively present in citrus fruits. 5-demethyltangeretin (5DT) is a unique PMF that has been demonstrated with potent chemopreventive effects on various human cancer cells. Methods In this study, we determined the metabolic fate of 5DT in mice and the bioactivities of its major metabolite. We found that 5DT underwent extensive biotransformation in 5DT-fed mice, and demethylated and hydroxylated metabolites were produced via phase I metabolism, while glucuronide and sulfate conjugates were generated via phase II metabolism. In particular, 4′ position on the B-ring of 5DT was the primary site for demethylation (formation of xanthomicrol, XAN), 3′ position on the B-ring was the primary site for hydroxylation, and 5 position on the A-ring was the primary site for methylation. Importantly, it is observed that the colonic level of XAN was much higher than that of 5DT. Results Therefore, next, we investigated the chemopreventive effects of XAN on HCT116 human colon cancer cells. It was found that XAN significantly inhibited the growth of HCT116 cells by inducing cell cycle arrest and cellular apoptosis and modulated multiple key signaling pathways associated with colon cancer in an anti-carcinogenic direction. For example, XAN treatment upregulated p53, and p21 and downregulated cyclin D and CDK4 level, compared to untreated control cells. Conclusions In conclusion, this study identified major metabolites of 5DT in the gastrointestinal tract, and demonstrated the suppressive effects of its primary colonic metabolite —XAN— on colon cancer in vitro. Funding Sources NIH, USDA-NIFA.

Protective effects of polymethoxyflavone-rich cold-pressed orange peel oil against ultraviolet B-induced photoaging on mouse skin

Abstract The protective effects of the topical application of orange peel cold-pressed oil (CPO), a rich polymethoxyflavones (PMFs) source, on ultraviolet B (UVB)-induced skin injury were studied. Depilated KM mice were irradiated by UVB at one minimal erythemal dose every day and the treated groups were smeared with CPO before radiation. After two weeks we found that CPO helped to maintain the dermal structure and enhanced serum levels and skin tissue expression of superoxide dismutase, catalase and glutathione peroxidase. In serum CPO lowered interleukin-6/-1β and tumor necrosis factor α levels; in skin it decreased the levels of type III collagen and H2O2 but increased hydroxyproline and type I collagen. Moreover, the skin expression of matrix metalloproteinases 2/9 was downregulated, whereas the expression of tissue inhibitor of metalloproteinases 1/2 was upregulated. The above effects followed a dose-dependent manner. Liquid chromatography analysis of irradiated skin tissue showed partial decomposition of PMFs into 5-hydroxy-PMFs.

Flavones as tyrosinase inhibitors: kinetic studies in vitro and in silico.

Tyrosinase is a multifunctional copper-containing oxidase enzyme that catalyses the first steps in the formation of melanin pigments. Identification of tyrosinase inhibitors is of value for applications in cosmetics, medicine and agriculture. To develop an analytical method that allows identification of drug-like natural products that can be further developed as tyrosinase inhibitors. Results of in vitro and in silico studies will be compared in order to gain a deeper insight into the mechanism of action of enzyme inhibition. Using an in vitro assay we tested tyrosinase inhibitor effects of five structurally related flavones, i.e. luteolin (1), eupafolin (2), genkwanin (3), nobiletin (4), and chrysosplenetin (5). The strongest inhibitors were further investigated in silico, using enzyme docking simulations. All compounds tested showed modest tyrosinase inhibitory effect compared to the positive control, kojic acid. The polymethoxy flavones 4 and 5 exhibited the strongest tyrosinase inhibitory effect with the half maximal inhibitory concentration (IC50 ) values of 131.92 ± 1.75 μM and 99.87 ± 2.38 μM respectively. According to kinetic analysis 2, 4 and 5 were competitive inhibitors, whereas 1 and 3 were non-competitive inhibitors of tyrosinase. Docking studies indicated that methoxy groups on 4 and 5 caused steric hindrance which prevented alternative binding modes in the tyrosinase; the methoxy groups on the B-ring of these flavones faced the catalytic site in the enzyme. The docking simulations nicely complemented the in vitro kinetic studies, opening the way for the development of predictive models for use in drug design.

Eupatorin Suppressed Tumor Progression and Enhanced Immunity in a 4T1 Murine Breast Cancer Model.

Eupatorin is a polymethoxy flavone extracted from Orthosiphon stamineus and was reported to exhibit cytotoxic effects on several cancer cell lines. However, its effect as an anti–breast cancer agent in vivo has yet to be determined. This study aims to elucidate the potential of eupatorin as an anti–breast cancer agent in vivo using 4T1 challenged BALB/c mice model. In this article, BALB/c mice (20-22 g) challenged with 4T1 cells were treated with 5 mg/kg or 20 mg/kg eupatorin, while the untreated and healthy mice were fed with olive oil (vehicle) via oral gavage. After 28 days of experiment, the mice were sacrificed and blood was collected for serum cytokine assay, while tumors were harvested to extract RNA and protein for gene expression assay and hematoxylin-eosin staining. Organs such as spleen and lung were harvested for immune suppression and clonogenic assay, respectively. Eupatorin (20 mg/kg) was effective in delaying the tumor development and reducing metastasis to the lung compared with the untreated mice. Eupatorin (20 mg/kg) also enhanced the immunity as the population of NK1.1+ and CD8+ in the splenocytes and the serum interferon-γ were increased. Concurrently, eupatorin treatment also has downregulated the expression of pro-inflammatory and metastatic related genes (IL-1β. MMP9, TNF-α, and NF-κB). Thus, this study demonstrated that eupatorin at the highest dosage of 20 mg/kg body weight was effective in delaying the 4T1-induced breast tumor growth in the animal model.

Improved bioaccessibility of polymethoxyflavones loaded into high internal phase emulsions stabilized by biopolymeric complexes: A dynamic digestion study via TNO's gastrointestinal model

In this work, the bioaccessibility of polymethoxyflavones (PMFs) loaded in high internal phase emulsions (HIPE, ϕoil = 0.82) stabilized by whey protein isolate (WPI)-low methoxy pectin (LMP) complexes was evaluated using in vitro lipolysis and dynamic in vitro intestinal digestion studies. PMFs loaded HIPE was prepared by using aqueous dispersion of pre-formed biopolymeric complexes (WPI-LMP, 2:1 ratio) as the external phase and medium chain triglycerides oil (containing PMFs extracted from citrus peel) as the dispersed phase. The in vitro lipolysis study revealed that PMFs in HIPE became bioaccessible much higher than PMFs in medium chain triacylglycerols oil (MCT oil). In addition, by simulating the entire human gastrointestinal (GI) tract, the GI model TIM-1 demonstrated a 5- and 2-fold increase in the total bioaccessibility for two major PMFs encapsulated in HIPE, i.e. tangeretin (TAN) and nobiletin (NOB), respectively, as opposed to PMFs in MCT oil. Together these results from the digestion study showed that the incorporation of a high amount of PMFs into the viscoelastic matrix of HIPE could represent an innovative and effective way to design an oral delivery system. Such a system could be used to control and to improve the delivery of lipophilic bioactive compounds within the different compartments of the digestive tract, especially the human upper GI tract.

Nobiletin and Derivatives: Functional Compounds from Citrus Fruit Peel for Colon Cancer Chemoprevention.

The search for effective methods of cancer treatment and prevention has been a continuous effort since the disease was discovered. Recently, there has been increasing interest in exploring plants and fruits for molecules that may have potential as either adjuvants or as chemopreventive agents against cancer. One of the promising compounds under extensive research is nobiletin (NOB), a polymethoxyflavone (PMF) extracted exclusively from citrus peel. Not only does nobiletin itself exhibit anti-cancer properties, but its derivatives are also promising chemopreventive agents; examples of derivatives with anti-cancer activity include 3′-demethylnobiletin (3′-DMN), 4′-demethylnobiletin (4′-DMN), 3′,4′-didemethylnobiletin (3′,4′-DMN) and 5-demethylnobiletin (5-DMN). In vitro studies have demonstrated differential efficacies and mechanisms of NOB and its derivatives in inhibiting and killing of colon cancer cells. The chemopreventive potential of NOB has also been well demonstrated in several in vivo colon carcinogenesis animal models. NOB and its derivatives target multiple pathways in cancer progression and inhibit several of the hallmark features of colorectal cancer (CRC) pathophysiology, including arresting the cell cycle, inhibiting cell proliferation, inducing apoptosis, preventing tumour formation, reducing inflammatory effects and limiting angiogenesis. However, these substances have low oral bioavailability that limits their clinical utility, hence there have been numerous efforts exploring better drug delivery strategies for NOB and these are part of this review. We also reviewed data related to patents involving NOB to illustrate the extensiveness of each research area and its direction of commercialisation. Furthermore, this review also provides suggested directions for future research to advance NOB as the next promising candidate in CRC chemoprevention.

Enrichment of polymethoxyflavones from Citrus reticulata ‘Chachi’ peels and their hypolipidemic effect

In the present study, a simple and efficient method based on orthogonal experimental design and macroporous resin chromatography was established for the first time for enrichment of polymethoxyflavones (PMFs) from the peels of Citrus reticulata ‘Chachi’ (CRC). The optimum conditions of extraction and enrichment process were as follows: use of a liquid to solid ratio of 1 to 14, two-hour extraction time repeated twice with 70% ethanol; use of HPD-450 macroporous resin, wash solvent of purified water and 25% aqueous ethanol, and 70% aqueous ethanol as desorption solvent. The purity of PMFs in the resulting extract reached 62.26%. Our data indicate that the PMFs purified could potently alleviate high-fat diet-induced hyperlipidaemia. The PMFs were nontoxic as determined by acute toxicity test. The method established was suitable for large-scale separation of PMFs from CRC peels and the PMFs might be developed as an anti-hyperlipidaemia agent or dietary supplement.

Supercritical fluid CO2 extraction of three polymethoxyflavones from Citri reticulatae pericarpium and subsequent preparative separation by continuous high-speed counter-current chromatography

Polymethoxyflavones (PMFs) are widely found in Citri Reticulatae Pericarpium (CRP) and have been investigated with a broad spectrum of biological activities as well as health promoting properties. However, separation of the PMFs from a complex sample, especially preparative separation of these PMFs with high purity, remains challenging. In the present study, an efficient method based on supercritical fluid extraction (SFE) and continuous high-speed counter-current chromatography (HSCCC) has been developed for extracting and preparative purification PMFs from CRP. Various experimental conditions were investigated to optimize the SFE and HSCCC processes. Under these optimized conditions, crude extract of CRP (extract I) was obtained with a maximum contents of nobiletin, 3,5,6,7,8,3′,4′-heptamethoxyflavone and tangeretin. Further extraction of crude extract I was carried out to obtain crude extract II, which was further isolated and purified by HSCCC. It was worth mentioned that continuous injection HSCCC process were realized without lost of separation efficiency, which allowed for multiple purification cycles and therefore saved a lot of labor and time. Furthermore, high-performance liquid chromatography (HPLC) was employed to analyze the fractions separated by HSCCC, which revealed that the purities of the three PMFs were all above 98%. The structures of the three PMFs were identified by LC–MS and 1H NMR spectroscopy.

Role of tangeritin against cognitive impairments in transgenic Drosophila model of Parkinson’s disease

Parkinson’s disease (PD) is the second most common neurodegenerative disorder after Alzheimer’s disease. As there is no permanent cure for the disease, the use of herbal compounds with antioxidant potential will be an effective approach for controlling the progression of disease. In this context the effect of tangeritin (a polymethoxy flavone concentrated in the peels of citrus fruits) was studied at final doses of 5, 10 and 20 μM on PD model flies. The doses were established in diet and the PD flies were allowed to feed on it for 24 days. The effect was studied on cognitive impairments. Immunostaining of brain sections for tyrosine hydroxylase was also performed. The docking studies were also carried out to give a plausible binding site of tangeritin on alpha synuclein molecule. The results of the study showed that tangeritin is effective in improving the cognitive impairments.

Simultaneous separation of six pure polymethoxyflavones from sweet orange peel extract by high performance counter current chromatography

Successful isolation of polymethoxyflavones (PMFs) from citrus peels has led to numerous evaluations of PMFs in a broad spectrum of biological activities, such as inhibition of chronic inflammation, cancer prevention and anti-atherogenic properties. Recent reports associated with the health promoting properties of PMFs in citrus fruits have dramatically increased. However, the limiting factor in animal and human study of PMFs is still the supply of pure PMFs, such as tangeretin, nobiletin, sinensetin and 3,5,6,7,3′,4′-hexamethoxyflavone. Herein, we introduce the newly developed efficient separation method using high-performance counter-current chromatography (HPCCC) in isolating multiple pure single PMFs simultaneously in one cycle process. With the smallest preparation loop on the semi-preparative HPCCC instrument, the optimized solvent system of hexanes/ethyl acetate/methanol/water resulted in the isolation of pure sinensetin, tangeretin, nobiletin, 3,5,6,7,3′,4′-hexamethoxyflavone, 5,6,7,4′-tetramethoxyflavone and 3,5,6,7,8,3′,4′-heptamethoxyflavone directly from crude sweet orange peel extract in one cycle of separation process by HPCCC in the mode of reverse phase. The purity of each of the six isolated PMFs is greater than 96.6% analyzed by high-performance liquid chromatography and proton nuclear magnetic resonance. Scale-up and high purity of individual PMFs can be separated by using a large separation loop in preparative HPCCC model. The renovated HPCCC methodology can be practically used in natural product isolation and consequent biological property evaluation.

Three Polymethoxyflavones Purified from Ougan (Citrus reticulata Cv. Suavissima) Inhibited LPS-Induced NO Elevation in the Neuroglia BV-2 Cell Line via the JAK2/STAT3 Pathway.

In order to establish an efficient method for separation of polymethoxyflavones (PMFs) and explore the anti-inflammatory mechanism of PMF monomers, a citrus variety rich in PMFs, Ougan (Citrus reticulata cv. Suavissima), was selected, and three monomers, including nobiletin, tangeretin, and 5-demethylnobiletin, were purified by ultrasonic-assisted extraction, solid phase extraction, and high-speed countercurrent chromatography separation. UPLC-MS was used to identify the three monomers. UPLC determined purities of 99.87% to nobiletin, 99.76% to tangeretin, and 98.75% to 5-demethylnobiletin with the standard curve method. A lipopolysaccharide (LPS)-induced NO releasing model was performed in the mouse microglia BV-2 cell line. Results illustrated that PMF monomers inhibited the NO release and the inflammation-related cytokines, including IL-1β, IL-6, and TNFα elevation. QRT-PCR revealed that PMFs alleviated LPS-induced upregulation of iNOS, IL-6, JAK2, TNFα, IL-1β, and NF-κB and LPS-induced downregulation of IκBα, while they did not affect TLR1, TLR2, TLR4, and TLR6. STAT3 expression was repressed by tangeretin and 5-demethylnobiletin, but not by nobiletin. Western blot assay also showed a suppression of expression and phosphorylation of JAK2 by all three PMF monomers, while STAT3 phosphorylation was restrained by tangeretin and 5-demethylnobiletin. The mechanism was primarily verified by the JAK2 inhibitor Ruxolitinib and the STAT3 inhibitor Stattic.

Prevention of Obesity and Hyperlipidemia by Heptamethoxyflavone in High-fat Diet-induced Rats.

Polymethoxyflavones (PMFs) have been shown to prevent obesity, ameliorate type 2 diabetes, and regulate lipid metabolism in vitro and in vivo. However, little is known about the contribution of 3,5,6,7,8,3',4'-heptamethoxyflavone (HMF) to prevent obesity and regulate lipid metabolism in vivo. We aimed to investigate the potential efficacy of HMF on preventing obesity and hyperlipidemia in rats fed a high-fat diet (HFD) and its underlying mechanisms. Male Sprague-Dawley rats were fed a normal diet or an HFD with or without HMF (0.02%, 0.04% and 0.08%, w/w) for 6 weeks. The supplementation of HMF not only significantly decreased body weight gain (HFD, 336.50 ± 18.84 g; LHMF, 309.43 ± 20.74 g; MHMF, 296.83 ± 13.88 g; HHMF, 265.71 ± 19.09 g; respectively, p < 0.05) and adipose tissues weight ( p < 0.05), but also markedly lowered serum levels of total cholesterol, triacylglycerol, and low-density lipoprotein cholesterol ( p < 0.05) in the sixth week in a dose-dependent manner compared with the HFD group. HMF also significantly alleviated hepatic steatosis in the liver (liver weight g/100 g body weight of HFD, 4.86 ± 0.11%; LHMF, 4.02 ± 0.33%; MHMF, 4.05 ± 0.31%; HHMF, 3.72 ± 0.34%; respectively, p < 0.05). Furthermore, transcriptome analysis and real-time quantitative RT-PCR demonstrated that HMF supplementation markedly downregulated hepatic genes related to adipogenesis transcription and inflammatory responses, and significantly upregulated genes related to fatty acid oxidation and energy expenditure. These results indicated that HMF could effectively prevent obesity and hyperlipidemia by regulation of the expression of lipid metabolism-related and inflammatory response-related genes.

Nobiletin induces growth inhibition and apoptosis in human nasopharyngeal carcinoma C666-1 cells through regulating PARP-2/SIRT1/AMPK signaling pathway.

Background/AimNobiletin, a major polymethoxyflavones (PMFs) from citri reticulatae pericarpium (CRP), can inhibit several forms of cancer proliferation. However, the effects of nobiletin on nasopharyngeal carcinoma (NPC) C666‐1 cells remain largely unknown.Materials and MethodsCell counting kit 8 (CCK8) assay was used to measure cell vitality. Flow cytometry was performed to measure the apoptosis rate. Quantitative real‐time polymerase chain reaction (qRT‐PCR) and Western blot analysis were applied to determine the expression of mRNA and protein, respectively.ResultsWe showed that the proliferation rate of C666‐1 cells was inhibited and the apoptosis rate was raised after treating with nobiletin. Moreover, nobiletin inhibited the expression of poly(ADP‐ribose)polymerase‐2 (PARP‐2), and the tumor suppression effect of nobiletin on C666‐1 is associated with PARP‐2‐dependent pathway.ConclusionWe demonstrated for the first time that nobiletin inhibited the growth of C666‐1 cells, which may be relative to its regulation on PARP‐2/SIRT1/AMPK signaling pathway. Our result implied that nobiletin may serve as a strategy to treat nasopharyngeal carcinoma.

Polymethoxyflavones: Chemistry and Molecular Mechanisms for Cancer Prevention and Treatment

Polymethoxyflavones (PMFs) are one group of the flavonoid compounds, with tangeretin (Tan) and nobiletin (Nob) being the most abundant PMFs in citrus peel. Numerous biological activities of PMFs have been intensively studied, including anti-inflammatory and anticancer activities. Because of their methoxy groups, PMFs are more lipophilic than hydroxyl flavones, which may affect their biological activities. In addition, researchers found that hydroxylated PMFs (HPMFs) are one of the major metabolites of PMFs in animal urine and feces. Although PMF and HPMFs do show anticancer activity against different types of cancers, but their low hydrophilicity is still a crucial factor that may affect their biological effectiveness. Therefore, from the pharmaceutical aspect, chemical modifications of PMFs have been carried out to obtain acetylated PMFs (Ac-PMFs) for enhancing their biological effects. From the past centuries to the present, cancer is still a critical disease that needs to be solved. Carcinogenesis can be simply divided into three stages: initiation, promotion, and progression. These three stages involve different biological events, such as DNA mutation, cell proliferation, cell growth, and metastasis. In this paper, we aim to illustrate the biological effects of different PMFs, HPMFs, PMF derivatives, and metabolites against different types of cancer and related molecular mechanisms.

Kaempferia parviflora Ethanol Extract, a Peroxisome Proliferator-Activated Receptor γ Ligand-binding Agonist, Improves Glucose Tolerance and Suppresses Fat Accumulation in Diabetic NSY Mice.

This study assessed the effect of Kaempferia parviflora, also known as black ginger (BG), and its ethanol extract (BGE) on peroxisome proliferator-activated receptor (PPAR) γ agonistic activity, glucose tolerance, fat accumulation, and lipids-induced hypertriglyceridemia in mice. PPARγ ligand-binding capacity in vitro and polymethoxy flavone contents were highly observed in organic solvent extracts. In an animal experiment A, male diabetic Nagoya-Shibata-Yasuda mice were divided into five dietary groups and fed each diet for 8 weeks: AIN-93G diet (low-fat [LF] diet), high-fat (HF) diet, HF diet supplemented with 1% BG, HF diet supplemented with 0.19% BGE, and HF diet supplemented with pioglitazone (PPARγ agonist, 3 mg/kg/day) as a PPARγ agonistic positive control. As determined from glucose and insulin tolerance tests, plasma glucose levels were improved in the BG and BGE groups. The BGE extract suppressed fat accumulation in adipose tissues, liver, and muscles without changing the plasma adiponectin level. In an animal experiment B, in order to investigate the effect of BG and BGE on lipid-induced hypertriglyceridemia, male ddY mice were divided into three test groups: control, BG-administered group (500 mg/kg), and BGE-administered group (100 mg/kg). The plasma triacylglycerol level was not different among the groups during the lipids administration test. These results conclude that the BGE extract containing several kinds of polymethoxy flavones showed PPARγ ligand-binding capacity in vitro and prevented obesity and insulin resistance independent of adiponectin secretion in mice. PRACTICAL APPLICATION: Kaempferia parviflora, also known as black ginger (BG), is often used as a folk medicine and a functional food material to prevent metabolic syndrome mainly in Asian regions. Here, we have clarified that ethanol extract from BG (BGE) contains several kinds of polymethoxy flavones to show PPARγ ligand-binding capacity and is an active extract for the improvement of obesity and insulin resistance. The BGE is expected to be applied for functional food materials in health food markets. Also, polymethoxy flavones to show PPARγ ligand-binding capacity can be generally applied as a physiological active compound of functional food supplements.

Characteristic of interaction mechanism between β-lactoglobulin and nobiletin: A multi-spectroscopic, thermodynamics methods and docking study

Nobiletin (Nob) is a major component among the most reported polymethoxyflavones (PMFs), which possesses multiple efficacious healthcare activities. Owing to its high melting point and poor water solubility, the oral bioavailability of Nob needs to be improved via loading Nob on carriers. To take full advantage of Nob, the interaction mechanism between Nob and vehicles should be clarified. Herein, β-lactoglobulin (β-LG) was selected as the vehicle and further investigated the binding mechanism between Nob and β-LG. The binding stoichiometry of complex was found to be 1:1 by analysis of intrinsic fluorescence experiment. The results also confirmed by isothermal titration calorimetry (ITC) measurement that the binding behavior between β-LG and Nob was a spontaneously endothermic process driving mainly by hydrophobic interaction. Moreover, competitive binding and molecular docking method indicated the Nob was primary bound to internal calyx of β-LG at neutral pH. UV spectrophotometry revealed that the solubility of Nob was enhanced to 3 times by forming complex. Furthermore, Nob could alter secondary structure of β-LG by a transition from α-helix to β-sheet and lead to small increase on surface hydrophobicity of β-LG. This work will provide some valuable information on clarifying the interaction between protein and PMFs, which contributing to improve the poor bioavailability of PMFs.