Inhibition of ATP synthase reverse activity restores energy homeostasis in mitochondrial pathologies.

The maintenance of cellular function relies on the close regulation of adenosine triphosphate (ATP) synthesis and hydrolysis. ATP hydrolysis by mitochondrial ATP Synthase (CV) is induced by loss of proton motive force and inhibited by the mitochondrial protein ATPase inhibitor (ATPIF1). The extent of CV hydrolytic activity and its impact on cellular energetics remains unknown due to the lack of selective hydrolysis inhibitors of CV. We find that CV hydrolytic activity takes place in coupled intact mitochondria and is increased by respiratory chain defects. We identified (+)-Epicatechin as a selective inhibitor of ATP hydrolysis that binds CV while preventing the binding of ATPIF1. In cells with Complex-III deficiency, we show that inhibition of CV hydrolytic activity by (+)-Epichatechin is sufficient to restore ATP content without restoring respiratory function. Inhibition of CV-ATP hydrolysis in a mouse model of Duchenne Muscular Dystrophy is sufficient to improve muscle force without any increase in mitochondrial content. We conclude that the impact of compromised mitochondrial respiration can be lessened using hydrolysis-selective inhibitors of CV.

Cocoa flavanols, Nrf2 activation, and oxidative stress in peripheral artery disease: mechanistic findings in muscle based on outcomes from a randomized trial.

The pathophysiology of muscle damage in peripheral artery disease (PAD) includes increased oxidant production and impaired antioxidant defenses. Epicatechin (EPI), a naturally occurring flavanol, has antioxidant properties that may mediate the beneficial effects of natural products such as cocoa. In a phase II randomized trial, a cocoa-flavanol-rich beverage significantly improved walking performance compared with a placebo in people with PAD. In the present work, the molecular mechanisms underlying the therapeutic effect of cocoa flavanols were investigated by analyzing baseline and follow-up muscle biopsies from participants. Increases in nuclear factor erythroid 2-related factor 2 (Nrf2) target antioxidants heme oxygenase-1 (HO-1) and NAD(P)H dehydrogenase [quinone] 1 (NQO1) in the cocoa group were significantly associated with reduced accumulation of central nuclei, a myopathy indicator, in type II muscle fibers (P = 0.017 and P = 0.023, respectively). Protein levels of the mitochondrial respiratory complex III subunit, cytochrome b-c1 complex subunit 2 (UQCRC2), were significantly higher in the cocoa group than in the placebo group (P = 0.032), and increases in UQCRC2 were significantly associated with increased levels of Nrf2 target antioxidants HO-1 and NQO1 (P = 0.001 and P = 0.035, respectively). Exposure of non-PAD human myotubes to ex vivo serum from patients with PAD reduced Nrf2 phosphorylation, an indicator of activation, increased hydrogen peroxide production and oxidative stress, and reduced mitochondrial respiration. Treatment of myotubes with EPI in the presence of serum from patients with PAD increased Nrf2 phosphorylation and protected against PAD serum-induced oxidative stress and mitochondrial dysfunction. Overall, these findings suggest that cocoa flavanols may enhance antioxidant capacity in PAD via Nrf2 activation.NEW & NOTEWORTHY The current study supports the hypothesis that in people with PAD, cocoa flavanols activate Nrf2, thereby increasing antioxidant protein levels, protecting against skeletal muscle damage, and increasing mitochondrial protein abundance. These results suggest that Nrf2 activation may be an important therapeutic target for improving walking performance in people with PAD.

Effects of epicatechin on cardiovascular function in middle-aged diet-induced obese rat models of metabolic syndrome.

The current study aimed to investigate the cardiovascular effects of epicatechin, a flavonoid found in green tea and cocoa, in attenuating complications associated with metabolic syndrome in diet-induced obese rats. Male Wistar-Kyoto (WKY) rats aged 16 weeks were fed either standard rat chow or given a high-fat-high-carbohydrate (HFHC) diet for 20 weeks. Epicatechin treatment (5 mg/kg/d) was administered to a subset of WKY rats commencing at week 8 of the 20 week HFHC feeding period. Body weights, food, water and energy intakes, blood pressure, heart rate and glucose tolerance were measured throughout the treatment period. Oxidative stress and inflammatory markers, lipid levels, cardiac collagen deposition, cardiac electrical function, aortic and mesenteric vessel reactivity were examined after the treatment. Twenty weeks of HFHC feeding in WKY rats resulted in the development of metabolic syndrome indicated by the presence of abdominal obesity, dyslipidaemia, glucose intolerance and increased blood pressure. Epicatechin treatment was found to enhance the oxidative stress status in HFHC groups through an increase in serum nitric oxide levels and a decrease in 8-isoprostane concentrations. Furthermore, WKY-HFHC rats displayed a decrease in IL-6 levels. The lipid profiles in HFHC groups showed improvement, with a decrease in LDL-cholesterol and TAG and an increase in HDL-cholesterol levels observed in WKY-HFHC rats. However, epicatechin was not effective in preventing weight gain, glucose intolerance or hypertension in HFHC fed rats. Overall, the results of this study suggest that epicatechin has the potential to improve the underlying mechanisms associated with metabolic syndrome in obese rats.

A novel optimized eco-friendly simple spectrofluorimetric method for the determination of total catechins in green tea extract: Application to commercial tablet.

Green tea extract (GTE) contains antioxidants that are present in green tea. The active constituents of green tea extract are catechins. This study demonstrates a spectrofluorimetric method for measuring GTE's catechin concentration based on its native fluorescence. To design a quick, sensitive, and ecological spectrofluorimetric approach, all features were investigated and adjusted. This method relies on determining the GTE ethanolic solution's native fluorescence at 312 nm after excitation at 227 nm. The calibration graph displayed a linear regression for values between 0.05 and 1.0 µg mL-1. The detection and quantification limits of the proposed technique were 0.008 and 0.026 µg mL-1, respectively. Two pure catechins present in GTE, (-)-epicatechin and (-)-epigallocatechin gallate, were examined by the proposed method. The analytical estimation of GTE in the pharmaceutical tablet was achieved effectively using this approach. An adequate degree of agreement was found when the findings were compared to those obtained by the comparative technique. Therefore, the novel strategy may be used in the GTE quality control study with minimal risks to people or the environment. The quantum yields of catechins were estimated. The validated technique was accepted by the International Council of Harmonization criteria.

Separation and Detection of Catechins and Epicatechins in Shanxi Aged Vinegar Using Solid-Phase Extraction and Hydrophobic Deep Eutectic Solvents Combined with HPLC.

This research presents a new, eco-friendly, and swift method combining solid-phase extraction and hydrophobic deep eutectic solvents (DES) with high-performance liquid chromatography (SPE-DES-HPLC) for extracting and quantifying catechin and epicatechin in Shanxi aged vinegar (SAV). The parameters, such as the elution solvent type, the XAD-2 macroporous resin dosage, the DES ratio, the DES volume, the adsorption time, and the desorption time, were optimized via a one-way experiment. A central composite design using the Box-Behnken methodology was employed to investigate the effects of various factors, including 17 experimental runs and the construction of three-dimensional response surface plots to identify the optimal conditions. The results show that the optimal conditions were an HDES (tetraethylammonium chloride and octanoic acid) ratio of 1:3, an XAD-2 macroporous resin dosage of 188 mg, and an adsorption time of 11 min. Under these optimal conditions, the coefficients of determination of the method were greater than or equal to 0.9917, the precision was less than 5%, and the recoveries ranged from 98.8% to 118.8%. The environmentally friendly nature of the analytical process and sample preparation was assessed via the Analytical Eco-Scale and AGREE, demonstrating that this method is a practical and eco-friendly alternative to conventional determination techniques. In summary, this innovative approach offers a solid foundation for the assessment of flavanol compounds present in SAV samples.

Peroxynitrite is essential for aerenchyma formation in rice roots under waterlogging conditions.

MAIN CONCLUSION: In this study, we report that peroxynitrite is necessary for ethylene-mediated aerenchyma formation in rice roots under waterlogging conditions. Plants under waterlogging stress face anoxygenic conditions which reduce their metabolism and induce several adaptations. The formation of aerenchyma is of paramount importance for the survival of plants under waterlogging conditions. Though some studies have shown the involvement of ethylene in aerenchyma formation under waterlogging conditions, the implication of peroxynitrite (ONOO-) in such a developmental process remains elusive. Here, we report an increase in aerenchyma formation in rice roots exposed to waterlogging conditions under which the number of aerenchyma cells and their size was further enhanced in response to exogenous ethephon (a donor of ethylene) or SNP (a donor of nitric oxide) treatment. Application of epicatechin (a peroxynitrite scavenger) to waterlogged plants inhibited the aerenchyma formation, signifying that ONOO- might have a role in aerenchyma formation. Interestingly, epicatechin and ethephon co-treated waterlogged plants were unable to form aerenchyma, indicating the necessity of ONOO- in ethylene-mediated aerenchyma formation under waterlogging conditions. Taken together, our results highlight the role of ONOO- in ethylene-mediated aerenchyma formation in rice and could be used in the future to develop waterlogging stress-tolerant varieties of rice.

Anti-adhesive activity of some secondary metabolites against Staphylococcus aureus on 3D printing medical materials.

Recent improvements in 3D printing technology have increased the usage of 3D printed materials in several areas. An exciting and emerging area of applying these next-generation manufacturing strategies is the development of devices for biomedical applications. The main aim of this work was to investigate the effect of tannic acid, gallic acid, and epicatechin gallate on the physicochemical characteristics of acrylonitrile butadiene-styrene (ABS) and Nylon 3D printing materials using the contact angle method. The adhesion of Staphylococcus aureus on untreated and treated materials was evaluated by scanning electron microscopy (SEM) analysis and the images were treated by MATLAB software. The results of the contact angle measurements showed a significant change in the physicochemical properties of both surfaces, indicated an increase in the electron donor character of 3D printing materials following treatment. Thus, the ABS surfaces treated with tannic acid, gallic acid, and epicatechin gallate have become more electron donating. Furthermore, our results proved the ability of S. aureus to adhere on all materials with a percentage of 77.86% for ABS and 91.62% for nylon. The SEM has shown that all actives molecules were sufficient to obtain better inhibition of bacterial adhesion, which tannic acid has shown a total inhibition of S. aureus on ABS. From these results, our treatment presents a high potential for utilization as an active coating to prevent bacterial attachment and the eventual biofilm development in medical field.

Mur ligase F as a new target for the flavonoids quercitrin, myricetin, and (-)-epicatechin.

MurC, D, E, and F are ATP-dependent ligases involved in the stepwise assembly of the tetrapeptide stem of forming peptidoglycan. As highly conserved targets found exclusively in bacterial cells, they are of significant interest for antibacterial drug discovery. In this study, we employed a computer-aided molecular design approach to identify potential inhibitors of MurF. A biochemical inhibition assay was conducted, screening twenty-four flavonoids and related compounds against MurC-F, resulting in the identification of quercitrin, myricetin, and (-)-epicatechin as MurF inhibitors with IC50 values of 143 µM, 139 µM, and 92 µM, respectively. Notably, (-)-epicatechin demonstrated mixed type inhibition with ATP and uncompetitive inhibition with D-Ala-D-Ala dipeptide and UM3DAP substrates. Furthermore, in silico analysis using Sitemap and subsequent docking analysis using Glide revealed two plausible binding sites for (-)-epicatechin. The study also investigated the crucial structural features required for activity, with a particular focus on the substitution pattern and hydroxyl group positions, which were found to be important for the activity. The study highlights the significance of computational approaches in targeting essential enzymes involved in bacterial peptidoglycan synthesis.

Cholesterol-lowering activity of adzuki bean (Vigna angularis) polyphenols.

BACKGROUND: Adzuki beans (ABs; Vigna angularis) were reported to show potential for prevention of cholesterol absorption and lowering of the blood cholesterol level. However, the main active compounds and some cellular effects remain unknown. In this study, we evaluated the potential cholesterol-lowering effects of (+)-catechin 7-O-ß-D-glucopyranoside (C7G) and (+)-epicatechin 7-O-ß-D-glucopyranoside (E7G), identified as abundant polyphenols in ABs. METHODS AND RESULTS: To investigate the cholesterol-lowering activity in vitro, cholesterol micelles, bile acids, and Caco-2 cells as an intestinal model were used in the study. C7G and E7G each inhibited micellar solubility in a dose-dependent manner, and their inhibitory activity was as strong as that of (+)-catechin (IC50 values: C7G, 0.23 ± 0.03 mg/ml; E7G, 0.22 ± 0.02 mg/ml; (+)-catechin, 0.26 ± 0.11 mg/ml). The AB polyphenols showed binding activity toward bile acids and changed them into an insoluble form. When Caco-2 cells were treated with C7G or E7G, the amount of incorporated cholesterol was significantly decreased compared with vehicle-treated control cells, and no cytotoxicity was observed under the experimental conditions used. Meanwhile, quantitative real-time PCR revealed that the mRNA level of the cholesterol transporter NPC1L1 remained unchanged in the treated cells. CONCLUSIONS: Taken together, the present findings suggest that C7G and E7G are the main active compounds in ABs, and have the ability to inhibit micellar solubility, bind to bile acids, and suppress cholesterol absorption. The present study supports the health benefits of ABs as a medicinal food and the application of AB polyphenols as medicinal supplements to suppress cholesterol elevation.

Optimization parameters for the production of dimer of epicatechin from an endophytic fungus Curvularia australiensis FC2AP using response surface methodology (RSM).

The search for natural therapeutic agents has intensified due to their potential to treat various diseases. Bioactive secondary metabolites from endophytes offer high therapeutic profiles and can be mass-produced after optimizing medium parameters and purification. This investigation aimed to maximize crude pigmented secondary metabolite (CPSM) production from Curvularia australiensis FC2AP by optimizing fermentation conditions statistically. The endophytic fungus produced a maximum yield of 8.81 UL/g from biomass using Sabouraud's Dextrose Broth. After screening essential factors, the Plackett-Burman design was used for factorial optimization, and the Box Behnken design was employed to investigate three significant factors. The final CPSM yield was 12.3 UL/g, approximately 4-fold higher than the preliminary growth medium. Chromatographic purification using a gradient solvent system resulted in six fractions, with the fourth fraction demonstrating the highest bioactivity profile. Structural characterization confirmed this fraction to be a dimer of epicatechin, which has anti-cancer properties, as confirmed through in vivo studies on Sprague Dawley rats. This is the first report of a epicatechin dimer produced from C. australiensis.

Quantification of Risperidone Contained in Precipitates Produced by Tea Catechins Using Nuclear Magnetic Resonance.

A mixture of risperidone and eight major tea catechins: (-)-epicatechin-3-O-gallate (ECg), (-)-epigallocatechin-3-O-gallate (EGCg), (-)-catechin-3-O-gallate (Cg), (-)-gallocatechin-3-O-gallate (GCg), (-)-epicatechin (EC), (-)-epigallocatechin (EGC), (+)-catechin (CA), or (+)-gallocatechin (GC), in tartaric acid buffer (pH 3.0) afforded a precipitate. Amounts of risperidone and these catechins in the precipitate were measured by quantitative 1H-NMR (qNMR). About half or more of risperidone used was precipitated by gallated catechins ECg, EGCg, Cg, and GCg; on the other hand, it was precipitated little by non-gallated catechins EC, EGC, CA, and GC. Furthermore, risperidone was precipitated more by 2,3-trans gallated catechins Cg and GCg than 2,3-cis gallated catechins ECg and EGCg. Regarding the amount of tea catechins in the precipitate obtained by a mixture of risperidone and Catechin Mixture, the amounts of 2,3-cis gallated catechins EGCg and ECg were much larger than those of the other green tea catechins GCg, EC, EGC, CA, and GC. It was considered that risperidone was mainly precipitated by EGCg and ECg in Catechin Mixture. Therefore, it can be concluded that when patients take risperidone with catechin-rich beverages, the efficacy of risperidone reduces, mainly because the 2,3-cis gallated catechins EGCg and ECg form complexes with risperidone and precipitate.

Epicatechin Prevents Cryocapacitation of Bovine Spermatozoa through Antioxidant Activity and Stabilization of Transmembrane Ion Channels.

Epicatechin (EPC) is a flavonoid belonging to the family of catechins; it has been described as a powerful scavenger of a wide spectrum of reactive oxygen species (ROS) and a modulator of ex vivo sperm vitality. In this study, we assessed the potential protective abilities of EPC on cryopreserved bovine spermatozoa. We focused on conventional quality parameters, as well as the oxidative profile of spermatozoa alongside capacitation patterns, and expression profiles of proteins involved in the process of capacitation. Semen samples were cryopreserved in the presence of 25, 50 or 100 µmol/L EPC and compared to native semen (negative control) as well as ejaculates frozen in the absence of EPC (positive control). A dose-dependent improvement of conventional sperm quality parameters was observed following EPC administration, particularly in case of the sperm motility, membrane, acrosome and DNA integrity in comparison to the positive control. Experimental groups exposed to all EPC doses presented with a significantly lower proportion of capacitated spermatozoa as opposed to the positive control. While no significant effects of EPC were observed in cases of superoxide production, a significant decrease in the levels of hydrogen peroxide and hydroxyl radical were recorded particularly in the experimental groups supplemented with 50 and 100 µmol/L EPC. Western blot analysis revealed that supplementation of particularly 100 µmol/L EPC to the semen extender prevented the loss of the cation channel of sperm (CatSper) isoforms 1 and 2, sodium bicarbonate cotransporter (NBC) and protein kinase A (PKA), which play important roles in the process of sperm capacitation. In summary, we may hypothesize that EPC is particularly effective in the stabilization of the sperm membrane during the freeze-thaw process through its ability to quench ROS involved in damage to the membrane lipids and to prevent the loss of membrane channels crucial to initiate the process of sperm capacitation. These attributes of EPC provide an additional layer of protection to spermatozoa exposed to low temperatures, which may be translated into a higher post-thaw structural integrity and functional activity of male gametes.

In Vivo Reversal of P-Glycoprotein-Mediated Drug Resistance in a Breast Cancer Xenograft and in Leukemia Models Using a Novel, Potent, and Nontoxic Epicatechin EC31.

The modulation of P-glycoprotein (P-gp, ABCB1) can reverse multidrug resistance (MDR) and potentiate the efficacy of anticancer drugs. Tea polyphenols, such as epigallocatechin gallate (EGCG), have low P-gp-modulating activity, with an EC50 over 10 µM. In this study, we optimized a series of tea polyphenol derivatives and demonstrated that epicatechin EC31 was a potent and nontoxic P-gp inhibitor. Its EC50 for reversing paclitaxel, doxorubicin, and vincristine resistance in three P-gp-overexpressing cell lines ranged from 37 to 249 nM. Mechanistic studies revealed that EC31 restored intracellular drug accumulation by inhibiting P-gp-mediated drug efflux. It did not downregulate the plasma membrane P-gp level nor inhibit P-gp ATPase. It was not a transport substrate of P-gp. A pharmacokinetic study revealed that the intraperitoneal administration of 30 mg/kg of EC31 could achieve a plasma concentration above its in vitro EC50 (94 nM) for more than 18 h. It did not affect the pharmacokinetic profile of coadministered paclitaxel. In the xenograft model of the P-gp-overexpressing LCC6MDR cell line, EC31 reversed P-gp-mediated paclitaxel resistance and inhibited tumor growth by 27.4 to 36.1% (p < 0.001). Moreover, it also increased the intratumor paclitaxel level in the LCC6MDR xenograft by 6 fold (p < 0.001). In both murine leukemia P388ADR and human leukemia K562/P-gp mice models, the cotreatment of EC31 and doxorubicin significantly prolonged the survival of the mice (p < 0.001 and p < 0.01) as compared to the doxorubicin alone group, respectively. Our results suggested that EC31 was a promising candidate for further investigation on combination therapy for treating P-gp-overexpressing cancers.

Phenolic-Based Discrimination between Non-Symptomatic and Symptomatic Leaves of Aesculus hippocastanum Infested by Cameraria ohridella and Erysiphe flexuosa.

The herbivore Cameraria ohridella (kingdom Animalia) and the pathogen Erysiphe flexuosa (kingdom Fungi) are considered pests and biotic stressors of Aesculus hippocastanum (chestnut trees). The impact of both pests on the accumulation of secondary metabolites in chestnut leaves was investigated. Specifically, the interactive effect of both pests on metabolite accumulation and their potential role in enhancing the resistance of chestnut trees to biological stress was the focus of this study. Aesculus hippocastanum leaves with varying degrees of Cameraria ohridella infestation and Erysiphe flexuosa infection were used in this research. Leaf samples were collected during the plant vegetative growth phase and evaluated for pest infection and secondary metabolite content. Eight main polyphenols were identified in the leaves: (1) neochlorogenic acid, (2) (-)-epicatechin, (3) procyanidin trimer A-type, (4) procyanidin tetramer A-type, (5) quercetin-3-O-arabinoside, (6) quercetin-3-O-rhamnoside, (7) kaempferol-3-O-arabinoside, and (8) kaempferol-3-O-rhamnoside. It was found that the accumulation of metabolites, primarily those derived from epicatechin and quercetin, during the initial vegetation phase (up to 11.05 or 09.05), strongly depended on the later degree of pest infection. The differences observed in the metabolite dynamics in the chestnut leaves, depending on the extent of infection, indicate the development of a metabolic response mechanism in chestnut trees to biological stress.

Study of Influence of Extraction Method on the Recovery Bioactive Compounds from Peel Avocado.

The avocado peel is a waste material from consumption avocado (Persea americana Mill.) with big biotechnology potential. The purpose of the present work was to study the influence of six extraction methods, maceration (M), maceration plus ß-cyclodextrin (MßC), solid-state fermentation (SSF), sonication with water or ethanol, wet grinding (WG), wet grinding plus maceration (WGM), on the recovery of bioactive compounds from the avocado peel such as total phenols, epicatechin and chlorogenic acid. The results showed that the extraction method has a significant effect on the content of total phenols, the WGM method obtaining the highest value of total phenols (2143.1 mg GAE/100 g dry weight). Moreover, the results indicated that the extraction method had a significant effect on chlorogenic acid and epicatechin recovery, the WGM method obtaining the highest amount of epicatechin and chlorogenic acid, 181.7 and 244.3 mg/100 g dry matter, respectively. Additionally, the characterization of WGM extract was realized by UPLC-ESI-MS/MS and GC-MS. Thus, the WGM method allowed for obtaining good yields of recovery of phenolic compounds using an accessible technology and a more environment-friendly solvent.

(-)-Epicatechin Inhibits Metastatic-Associated Proliferation, Migration, and Invasion of Murine Breast Cancer Cells In Vitro.

Breast cancer, due to its high incidence and mortality, is a public health problem worldwide. Current chemotherapy uses non-specific cytotoxic drugs, which inhibit tumor growth but cause significant adverse effects. (-)-Epicatechin (EC) is part of a large family of biomolecules called flavonoids. It is widely distributed in the plant kingdom; it can be found in green tea, grapes, and cocoa. Several studies in animals and humans have shown that EC induces beneficial effects in the skeletal muscle and the cardiovascular system, reducing risk factors such as arterial hypertension, endothelial dysfunction, damage to skeletal muscle structure, and mitochondrial malfunction by promoting mitochondrial biogenesis, with no adverse effects reported. Recently, we reported that EC had an antitumor effect in a murine triple-negative mammary gland tumor model, decreasing tumoral size and volume and increasing survival by 44%. This work aimed to characterize the effects of flavanol EC on proliferation, migration, and metastasis markers of triple-negative murine breast (4T1) cancer cells in culture. We found proliferation diminished and Bax/Bcl2 ratio increased. When the migration of culture cells was evaluated, we observed a significant reduction in migration. Also, the relative expression of the genes associated with metastasis, Cdh1, Mtss1, Pten, Bmrs, Fat1, and Smad4, was increased. In conclusion, these results contribute to understanding molecular mechanisms activated by EC that can inhibit metastatic-associated proliferation, migration, and invasion of murine breast cancer cells.

Induction, Flavonoids Contents, and Bioactivities Analysis of Hairy Roots and True Roots of Tetrastigma hemsleyanum Diels et Gilg.

The flavonoids in Tetrastigma hemsleyanum Diels et Gilg (T. hemsleyanum) have high medicinal value. However, because of slow growth and harsh ecological environments, T. hemsleyanum is currently an endangered species. In light of this, we present a detailed hairy root induction procedure as a promising alternative to true roots with medicinal value. The percentage of explants induced by Agrobacterium rhizogenes (A. rhizogenes) to produce hairy roots out of the total number of explants infected (induction rate 1) was 95.83 ± 7.22%, and the proportion of hairy roots that contained Rol B fragments among all the hairy roots with or without Rol B fragments (positive rate) was 96.57 ± 1.72%. The transformation was further confirmed by the expression of the GUS protein. A high-productive hairy root line was screened for the comparative profiling of six flavonoids with true roots using high-performance liquid chromatography (HPLC). The contents of (+)-catechin, (-)-epicatechin, neochlorogenic acid, luteolin-6-C-glucoside, and orientin were 692.63 ± 127.24, 163.34 ± 31.86, 45.95 ± 3.46, 209.68 ± 6.03, and 56.82 ± 4.75 µg/g dry weight (DW) of 30-day-old hairy roots, respectively, which were higher than those of 3-year-old true roots. Hairy roots have stronger antioxidant activity than true roots. Overall, the hairy roots of T. hemsleyanum could serve as promising alternative sources for the production of flavonoids with medicinal uses.

Preventing Adipogenesis and Preserving Mitochondria and GLUT-4 Functions by Extracts and Isolated Compounds of Australian Acacia saligna.

Acacia saligna's secondary metabolites show promise in treating type 2 diabetes mellitus and its related conditions. We previously discovered that methanolic extracts, isolated flavonoids, and cyclitols effectively preserve mitochondria in 3T3-L1 adipocytes. In this current work, quantification of lipid droplet levels with Oil Red O assay showed a noticeable decrease in lipogenesis in 3T3-L1 cells. Methanolic leaf and bark extracts and isolated compounds, (-)-epicatechin 6 and myricitrin 8, reduced cellular lipid levels by 21.15% to 25.28%, respectively. mRNA levels of key regulators of mitochondrial biogenesis, such as adiponectin, PGC-1α, and mtTFA, were increased. Methanolic flower extract (FL-MeOH) and its chemical components, naringenin 1 and D-(+)-pinitol 5a, increased these gene levels from 10% to 29% at the higher dose. Our study found that FL-MeOH slightly reduced pro-inflammatory cytokines TNF-α and IL-6, attributed to two phytochemicals, naringenin-7-O-α-L-arabinofuranoside 2 and D-(+)-pinitol 5a. Western blot analysis also showed that adipocytes treated with MeOH extracts had higher GLUT-4 expression levels than untreated adipocytes. Overall, A. saligna extracts and their isolated compounds demonstrated anti-lipogenesis activity during 3T3-L1 cell differentiation, modulation of transcriptional levels of adiponectin, PGC-1α, and mtTFA, reducing TNF-α and IL-6 mRNA levels, promoting mitochondrial biogenesis, and enhancing GLUT-4 expression.

Coffee Silverskin Phytocompounds as a Novel Anti-Aging Functional Food: A Pharmacoinformatic Approach Combined with In Vitro Study.

Coffee became a beverage that was in demand in the world and consequently produced millions of tons of coffee byproducts namely coffee silverskin (CS). Unutilized CS will be waste and cause environmental pollution such as greenhouse gas emissions, landfill waste, and groundwater contamination. This is a research concern at this time, although many studies have been conducted to find newer applications of CS, exploration of its benefits in the health sector is still limited. Therefore, exploring the benefits of CS to prevent or delay aging will be very interesting to develop in functional food industry technology. Therefore, this study aims to report profiling metabolites or phytochemicals, biological activities in terms of antioxidant activity, and potential anti-aging of CS via molecular docking simulation and in vitro modulation of the mTOR/AMPK/SIRT1 pathway. Something new has been obtained from this work, the profile of phytocompounds, and biological activities both in molecular docking simulation and in vitro studies. Some of the compounds observed in Robusta CS extract (rCSE) such as Epicatechin, Kaempferol, and Quercitrin, and Arabica CS extract (aCSE) such as (+)-Catechin dan Naringin have promising potential as inhibitors of iNOS, mTOR, and HIF-1α via molecular docking simulation. Interestingly, the in vitro biological activity assay of antioxidant and anti-aging activity, rCSE showed the same promising potential as the results of a molecular docking simulation. More interestingly, AMPK/SIRT1/mTOR expressions are well modulated by rCSE compared to aCSE significantly (p < 0.05). This makes the rCSE have promising biological activity as a candidate for functional food development and/or treatment agent in combating free radicals that cause the aging process. In vivo studies and human trials are certainly needed to see the further efficacy of the rCSE in the future.

Antihyperglycemic Properties of Extracts and Isolated Compounds from Australian Acacia saligna on 3T3-L1 Adipocytes.

Our early work indicated that methanolic extracts from the flowers, leaves, bark, and isolated compounds of Acacia saligna exhibited significant antioxidant activities in vitro. The overproduction of reactive oxygen species (ROS) in the mitochondria (mt-ROS) interfered with glucose uptake, metabolism, and its AMPK-dependent pathway, contributing to hyperglycemia and diabetes. This study aimed to screen the ability of these extracts and isolated compounds to attenuate the production of ROS and maintain mitochondrial function via the restoration of mitochondrial membrane potential (MMP) in 3T3-L1 adipocytes. Downstream effects were investigated via an immunoblot analysis of the AMPK signalling pathway and glucose uptake assays. All methanolic extracts effectively reduced cellular ROS and mt-ROS levels, restored the MMP, activated AMPK-α, and enhanced cellular glucose uptake. At 10 µM, (-)-epicatechin-6 (from methanolic leaf and bark extracts) markedly reduced ROS and mt-ROS levels by almost 30% and 50%, respectively, with an MMP potential ratio 2.2-fold higher compared to the vehicle control. (-)-Epicatechin 6 increased the phosphorylation of AMPK-α by 43%, with an 88% higher glucose uptake than the control. Other isolated compounds include naringenin 1, naringenin-7-O-α-L-arabinopyranoside 2, isosalipurposide 3, D-(+)-pinitol 5a, and (-)-pinitol 5b, which also performed relatively well across all assays. Australian A. saligna active extracts and compounds can reduce ROS oxidative stress, improve mitochondrial function, and enhance glucose uptake through AMPK-α activation in adipocytes, supporting its potential antidiabetic application.