Effects of In Vitro Digestion of Polyphenols from Coffee on Binding Parameters to Human Topoisomerase II α.

Type II topoisomerase (TOPII) is an enzyme that influences the topology of DNA. DNA breaks generated by TOPII may result in mutagenic or cytotoxic changes in cancer cells. In this study, we characterized interactions of TOPIIα with coffee extracts and individual chlorogenic acids (CHAs) from the extracts by performing isothermal titration calorimetry (ITC) and molecular docking (MD) simulations. The study showed that the highest affinity to TOPIIα was found in green coffee (ΔG = -38.23 kJ/mol) and monochlorogenic acids fraction of coffee extracts (ΔG = -35.80 kJ/mol), resulting from the high content of polyphenols, such as CHAs, which can bind to the enzyme in the active site. Coffee extracts and their fractions maintained a high affinity for TOPIIα after simulated digestion in the presence of probiotic bacteria. It can be concluded that coffee may be a potential TOPIIα inhibitor considered as a functional food for cancer prevention.

The Interaction of Heptakis (2,6-di-O-Methyl)-ß-cyclodextrin with Mianserin Hydrochloride and Its Influence on the Drug Toxicity.

One tetracyclic antidepressant, mianserin hydrochloride (MIA), has quite significant side effects on a patients' health. Cyclodextrins, which are most commonly used to reduce the undesirable features of contained drugs within their hydrophobic interior, also have the potential to alter the toxic behavior of the drug. The present paper contains investigations and the characteristics of interaction mechanisms for MIA and the heptakis (2,6-di-O-methyl)-ß-cyclodextrin (DM-ß-CD) system, and evaluated the effects of the complexation on MIA cytotoxicity. In order to assess whether there was an interaction between MIA and DM-ß-CD molecules, isothermal titration calorimetry (ITC) have been chosen. Electrospray ionization mass spectrometry (ESI-MS) helped to establish the complex stoichiometry, and circular dichroism spectroscopy was used to describe the process of complex formation. In order to make a wider interpretative perspective, the molecular docking results have been performed. The viability of Chinese hamster cells were investigated in the presence of DM-ß-CD and its complexes with MIA in order to estimate the cytotoxicity of the drug and the conjugate with the chosen cyclodextrin. The viability of B14 cells treated with MIA+DM-ß-CD is lower (the toxicity is higher) than with MIA alone, and no protective effects have been observed for complexes of MIA with DM-ß-CD in any ratio.

Phenolics-Rich Extracts of Dietary Plants as Regulators of Fructose Uptake in Caco-2 Cells via GLUT5 Involvement.

The latest data link the chronic consumption of large amounts of fructose present in food with the generation of hypertension and disturbances in carbohydrate and lipid metabolism, which promote the development of obesity, non-alcoholic fatty liver disease, insulin resistance, and type 2 diabetes. This effect is possible after fructose is absorbed by the small intestine cells and, to a lesser extent, by hepatocytes. Fructose transport is dependent on proteins from the family of glucose transporters (GLUTs), among which GLUT5 selectively absorbs fructose from the intestine. In this study, we examined the effect of four phenolic-rich extracts obtained from A. graveolens, B. juncea, and M. chamomilla on fructose uptake by Caco-2 cells. Extracts from B. juncea and M. chamomilla most effectively reduced fluorescent fructose analogue (NBDF) accumulation in Caco-2, as well as downregulated GLUT5 protein levels. These preparations were able to decrease the mRNA level of genes encoding transcription factors regulating GLUT5 expression-thioredoxin-interacting protein (TXNIP) and carbohydrate-responsive element-binding protein (ChREBP). Active extracts contained large amounts of apigenin and flavonols. The molecular docking simulation suggested that some of identified phenolic constituents can play an important role in the inhibition of GLUT5-mediated fructose transport.

Bioavailability and metabolism of selected cocoa bioactive compounds: A comprehensive review.

Cocoa beans and their co-products are a rich source of beneficial compounds for health promotion, including polyphenols and methylxanthines. Knowledge of bioavailability and in vivo bioactivity of these phytochemicals is crucial to understand their role and function in human health. Therefore, many studies concerning bioavailability and bioactivity of cocoa bioactive compound have been done in both in vivo animal models and in humans. This critical review comprehensively summarizes the existing knowledge about the bioavailability and the major metabolic pathways of selected cocoa bioactive compounds (i.e. monomeric flavan-3-ols, procyanidins, anthocyanins, flavonols, phenolic acids, N-phenylpropenoyl-L-amino acids, stilbenes, and methylxanthines). The compiled results indicated that many of these compounds undergo extensive metabolism prior to absorption. Different factors have been suggested to influence the bioavailability of polyphenols and methylxanthines among them the role of gut microbiota, structure of these compounds, food matrix and occurrence of other substances were the most often considered. Aforementioned factors decided about the site where these bioactive compounds are digested and absorbed from the alimentary tract, as well as the pathway by which they are metabolized. These factors also determine of the type of transport through the intestine barrier (passive, involving specific enzymes or mediated by specific transporters) and their metabolic path and profile.

Cicer arietinum L. Sprouts' Influence on Mineralization of Saos-2 and Migration of MCF-7 Cells.

In the present study, we investigated the biological activity of four extracts obtained from Cicer arietinum L. sprouts. The fermentation of the sprouts with Lactobacillus casei and their incubation with ß-glucosidase elevated the concentrations of isoflavonoids, especially coumestrol, formononetin and biochanin A. To study the biological activity of C. arietinum, the human osteosarcoma Saos-2 and human breast cancer MCF-7 cell lines were used. The extracts obtained from fermented sprouts exhibited the strongest ability to decrease intracellular oxidative stress in both types of cells. They augmented mineralization and alkaline phosphatase activity in Saos-2 cells, as well as diminished the secretion of interleukin-6 and tumor necrosis factor α. Simultaneously, the extracts, at the same doses, inhibited the migration of MCF-7 cells. On the other hand, elevated concentrations of C. arietinum induced apoptosis in estrogen-dependent MCF-7 cells, while lower doses stimulated cell proliferation. These results are important for carefully considering the use of fermented C. arietinum sprouts as a dietary supplement component for the prevention of osteoporosis.

Evaluation of Isoflavones as Bone Resorption Inhibitors upon Interactions with Receptor Activator of Nuclear Factor-κB Ligand (RANKL).

Receptor activator of nuclear factor-κB ligand (RANKL) is a cytokine responsible for bone resorption. It binds its receptor RANK, which activates osteoporosis. High levels of osteoprotegerin (OPG) competitively binding RANKL limit formation of ligand-receptor complexes and enable bone mass maintenance. The new approach to prevent osteoporosis is searching for therapeutics that can bind RANKL and support OPG function. The aim of the study was to verify the hypothesis that isoflavones can form complexes with RANKL limiting binding of the cytokine to its receptor. Interactions of five isoflavones with RANKL were investigated by isothermal titration calorimetry (ITC), by in silico docking simulation and on Saos-2 cells. Daidzein and biochanin A showed the highest affinity for RANKL. Among studied isoflavones coumestrol, formononetin and biochanin A showed the highest potential for Saos-2 mineralization and were able to regulate the expression of RANKL and OPG at the mRNA levels, as well as osteogenic differentiation markers: alkaline phosphatase (ALP), collagen type 1, and Runt-related transcription factor 2 (Runx2). Comparison of the osteogenic activities of isoflavones showed that the use of physicochemical techniques such as ITC or in silico docking are good tools for the initial selection of substances showing a specific bioactivity.

Influence of Diet Enriched with Cocoa Bean Extracts on Physiological Indices of Laboratory Rats.

Cocoa bean is a rich source of polyphenols, mainly flavonoids which have a wide range of biological properties. The aim of the study was to determine the physiological indices of laboratory rats as a response to diets containing water extracts of raw or roasted cocoa beans of Forastero variety, as well as purified monomeric flavan-3-ols fraction isolated from them. The influence of these extracts on selected parameters was studied during 4 weeks feeding. The samples of rats feces were collected throughout the experiment and after its completion, biological samples (intestines content, blood, and organs) were retrieved individually from each rat and subjected to analyses. The observed changes in the gastrointestinal tract functioning indices and metabolism indicators, determined throughout the study and after its completion, confirm to some extent the biological activity of polyphenol extracts of cocoa beans. The differences in the results obtained for the analyzed parameters of the gastrointestinal tract revealed that the cocoa bean extracts differently affected the physicochemical properties of rats' intestines. The results indicate the beneficial effects of the applied nutrition treatment on the activity of cecal enzymes and the content of volatile fatty acids in the gut.

Binding of Red Clover Isoflavones to Actin as A Potential Mechanism of Anti-Metastatic Activity Restricting the Migration of Cancer Cells.

Actin functions are crucial for the ability of the cell to execute dynamic cytoskeleton reorganization and movement. Nutraceuticals that form complexes with actin and reduce its polymerization can be used in cancer therapy to prevent cell migration and metastasis of tumors. The aim of this study was to evaluate the ability of isoflavones to form complexes with actin. Docking simulation and isothermal titration calorimetry were used for this purpose. The formation of complexes by hydrogen bonds, hydrophobic and π-π interactions was demonstrated. Interactions occurred at the ATP binding site, which may limit the rotation of the actin molecule observed during polymerization and also at the site responsible for contacts during polymerization, reducing the ability of the molecule to form filaments. The greatest therapeutic potential was demonstrated by isoflavones occurring in red clover sprouts, i.e., biochanin A and formononetin, being methoxy derivatives of genistein and daidzein.

The effects of baking conditions on acrylamide content in shortcrust cookies with added freeze-dried aqueous rosemary extract.

The aim of the present study was to determine the effects of baking air humidity and dough supplementation with freeze-dried aqueous rosemary extract on acrylamide content in shortcrust cookies, as well as on their antioxidant properties and phenolic composition. Shortcrust cookies were baked at 170 °C in dry or humid (90%) air using 0.1, 0.2, or 0.5% of rosemary extract, and were compared to control samples without the extract. Acrylamide concentration in the obtained products ranged from 22.49 to 28.38 µg kg-1. Furthermore, cookies baked in humid air had less acrylamide (by 6% on average) than those baked in dry air, irrespective of extract content. On the other hand, a significant relationship was found between humidity conditions and total phenolic content in the final products. Shortcrust cookies containing 0.5% of rosemary extract and baked in humid air revealed the best antioxidant properties as their total polyphenol content was approx. Three times higher than that in control samples. Furthermore, shortcrust cookies with 0.5% of rosemary extract showed superior DPPH radical scavenging capacity.

Phenolic Profile and Antioxidant Potential of Leaves from Selected Cotoneaster Medik. Species.

The antioxidant efficiency of 70% aqueous methanolic extracts from the leaves of twelve selected Cotoneaster Medik. species was evaluated using four complementary in vitro tests based on SET- (single electron transfer) and HAT-type (hydrogen atom transfer) mechanisms (DPPH, FRAP, O2(•-) and H2O2 scavenging assays). The samples exhibited the dose-dependent responses in all assays with activity parameters of EC50 = 18.5-34.5 µg/mL for DPPH; 0.9-3.8 mmol Fe(2+)/g for FRAP; SC50 = 27.7-74.8 µg/mL for O2(•-); and SC50 = 29.0-91.3 µg/mL for H2O2. Significant linear correlations (|r| = 0.76-0.97, p < 0.01) between activity parameters and total contents of phenolics (5.2%-15.4% GAE) and proanthocyanidins (2.1%-15.0% CYE), with weak or no effects for chlorogenic acid isomers (0.69%-2.93%) and total flavonoids (0.28%-1.40%) suggested that among the listed polyphenols, proanthocyanidins are the most important determinants of the tested activity. UHPLC-PDA-ESI-QTOF-MS analyses led to detection of 34 polyphenols, of which 10 B-type procyanidins, 5 caffeoylquinic acids and 14 flavonoids were identified. After cluster analysis of the data matrix, the leaves of Cotoneaster zabelii, C. splendens, C. bullatus, C. divaricatus, C. hjelmqvistii and C. lucidus were selected as the most promising sources of natural antioxidants, exhibiting the highest phenolic levels and antioxidant capacities, and therefore the greatest potential for pharmaceutical applications.

Chickpeas­composition, nutritional value, health benefits, application to bread and snacks: a review.

Chickpea is grain legumes grown mainly in areas with temperate and semiarid climate. It is characterized by a high content of protein, fat, vitamins, fiber, and a lower content of carbohydrates than flour of wheat. Chickpeas may contain antinutritional compounds that can impair utilization of the nutrients by people. Heat treatment is an effective method to increase the amount of protein available for intestinal digestibility. Adding chickpeas to a foodstuff can increase their nutritional value and reduce the acrylamide content. Acrylamide is an antinutritional substance present in foods, such as bread, snacks, and chips. Chickpea flour and protein may be new way to a reduce the content of acrylamide in products of this type. The addition of chickpea flour affects the sensory and textural properties.

The Influence of Maceration and Flavoring on the Composition and Health-Promoting Properties of Roasted Coffee.

Over the years, many methods of refining green beans have been developed, including maceration aimed at enriching the coffee aroma and improving the overall quality. This study aimed to evaluate the influence of different methods of maceration (fruit and wine) and the addition of food flavors to coffee beans on antioxidant activity, caffeine, phenolic and organic acid content, as well as health-promoting properties. This research showed that the use of the maceration in melon and apple fruit pulp (100 g of fruit pulp per 100 g of green coffee, incubated for 24 h, coffee roasting at 230 °C, control trial roasted coffee) ensured the highest polyphenol (hydroxycinnamic acids and their esters-chlorogenic acids) content (in melon pulp-13.56 g/100 g d.b. (dry bean); in apple pulp-13.22 g/100 g d.b., p < 0.05 (one-way ANOVA)) and antioxidant activity. Melon (92.11%, IC50 = 3.80 mg/mL extract) and apple (84.55%, IC50 = 4.14 mg/mL) showed the highest α-amylase (enzyme concentration 10 µmol/mL) inhibition activity (0.5 mg/mL for both fruits). The addition of food flavors reduced the total content of chlorogenic acids to the range of 4.64 to 6.48 g/100 g d.b. and increased the content of acrylamide and 5-HMF, which positively correlated with a low antioxidant potential compared to the macerated samples and the control. Studies have shown that coffee macerated in the pulp of melon and apple fruit, due to its great potential to inhibit α-amylase in vivo, may have a preventive effect on type II diabetes. This study complements the current knowledge on the potential health-promoting properties of coffee flavored using different methods; further research should include more advanced models for testing these health-promoting properties. Statistical analysis was based on the determination of the average values of six measurements and their standard deviation, as well as on the one-way ANOVA (analysis of variation) and the Pearson correlation coefficient, using Statistic 10.0 software. The significance was defined at p ≤ 0.05.

Evaluation of the effect of roasting and digestion on biological activity of compounds of coffee extracts - in vitro assessment of the bioavailability, cytoprotective properties and modulation of inflammatory response.

Coffee is the most consumed beverage in the world. Consumption of phenolic compounds present in coffee protects the body against oxidative stress generation, inflammatory response, and cancer development. The aim of the study was evaluation of biological activity of coffee extracts (obtained from green, as well as light and dark roasted Robusta and Arabica beans) and isolated fractions on human colon adenocarcinoma Caco-2 cells, which are used as a cellular model of intestinal barrier in bioavailability studies. Additionally, impact of coffee phenolics on oxidative stress level and anti-inflammatory activity has been studied with RAW 264.7 macrophages used in immunomodulatory research. It was demonstrated that the coffee constituents protection against oxidative stress, lipotoxicity and secretion of proinflammatory mediators is correlated with the presence of mono- and dichlorogenic acids and roasting process. It was demonstrated that coffee phytochemicals can decrease cells proliferation and bind to topoisomerase IIα being a dietary tool in cancer prevention.

Assessment of the Interaction of Acetylcholinesterase Binding with Bioactive Compounds from Coffee and Coffee Fractions Digested In Vitro in the Gastrointestinal Tract.

The aim of the study was to evaluate the degree of acetylcholinesterase (AChE) inhibition by green and light- and dark-roasted coffee extracts and their fractions after digestion in a simulated gastrointestinal tract. The analysis was carried out using isothermal titration calorimetry, molecular docking, and dynamics simulations. The results showed that 3-O-caffeoylquinic acid binds strongly to AChE through hydrogen interactions with the amino acids ARG289A, HIS440A, and PHE288A and hydrophobic interactions with TYR121A in the active site of the enzyme. The Robusta green coffee extract (ΔG = -35.87 kJ/mol) and dichlorogenic acid fraction (ΔG = -19-29 kJ/mol) showed the highest affinity. Dichlorogenic acids (3,4-O-dicaffeoylquinic acid, 4,5-O-dicaffeoylquinic acid, and 3,4-O-dicaffeoylquinic acid) have high affinity for AChE as single compounds (ΔG(ITC) = -48.99-55.36 kJ/mol, ΔG(LF/AD) = -43.38-45.38 kJ/mol). The concentration necessary to reduce AChE activity by 50% amounted to 0.22 µmol/µmol chlorogenic acids to the enzyme.

Effect of Inhibiting Butyrylcholinesterase Activity Using Fractionated Coffee Extracts Digested In Vitro in Gastrointestinal Tract: Docking Simulation and Calorimetric and Studies.

Butyrylcholinesterase (BChE) is a major enzyme from the alpha-glycoprotein family that catalyzes the hydrolysis of neurotransmitter acetylcholine (ACh), lowering the concentration of ACh in the nervous system, which could cause aggravation of Alzheimer's disease (AD). In select pathological conditions, it is beneficial to reduce the activity of this enzyme. The aim of this study was to evaluate the degree of BChE inhibition by coffee extracts fractionated into mono- and diesters of caffeic acid/caffeine, digested in vitro in the gastrointestinal tract. The bioactive compounds from coffee showed high affinity for BchE, -30.23--15.28 kJ/mol, and was the highest for the caffeine fraction from the green Arabica extract. The isolated fractions were highly effective in inhibiting BChE activity at all in vitro digestion phases. It has been shown that the fractionation of coffee extracts could be potentially used to obtain high prophylactic or even therapeutic effectiveness against AD.

Reply to Zarguan et al. Comment on "Budryn et al. Hydroxybenzoic Acids as Acetylcholinesterase Inhibitors: Calorimetric and Docking Simulation Studies. Nutrients 2022, 14, 2476".

We thank Prof. Zarguan's group for their comment [...].

Evaluation of the thermal stability of bioactive compounds in coffee beans and their fractions modified in the roasting process.

Coffee is used as flavor or health-promoting additive in thermally processed food. In this study, ground coffee and freeze-dried coffee extracts were evaluated in terms of their thermal stabilities, and for the first time heat resistance of fractions (mono-, dichlorogenic acids and caffeine) with different roasting levels was evaluated. It observed that the degradation of green coffee bean ingredients began at 150 °C, and for the re-heated light and dark roasted, in the range of 171-188 °C. The lyophilized extracts were more stable and their degradation began around 160 °C. However, with the re-treatment (cooking, baking, frying) of the coffee extract fractions, the degradation of the monochlorogenic acids commenced at 114 °C, while for dichlorogenics at 108 °C and caffeine at 146 °C. Monochlorogenic acids in Robusta coffee showed high antioxidant activity (55-70%) and the highest content of fiber (13-17%). Coffee could be used to fortify food.

Evaluation of Activity of Sesquiterpene Lactones and Chicory Extracts as Acetylcholinesterase Inhibitors Assayed in Calorimetric and Docking Simulation Studies.

The aim of the study was to explain the effects of sesquiterpene lactones (SLs) from chicory (Cichorium intybus L.) root extracts as inhibitors of acetylcholinesterase (AChE) at the molecular level and to determine the inhibition of AChE activity by specific SLs (lactucin and lactucopicrin) and different chicory extracts. The obtained SLs-rich extracts were purified by the countercurrent partition chromatography (CPC) technique. AChE inhibitors were analyzed using two models: isothermal titration calorimetry (ITC) and docking simulation. The results of ITC analysis of the enzyme and the ligands' complexation showed strong interactions of SLs as well as extracts from chicory with AChE. In a test of enzyme activity inhibition after introducing acetylcholine into the model system with SL, a stronger ability to inhibit the hydrolysis of the neurotransmitter was observed for lactucopicrin, which is one of the dominant SLs in chicory. The inhibition of enzyme activity was more efficient in the case of extracts, containing different enzyme ligands, exhibiting complementary patterns of binding the AChE active site. The study showed the high potential of using chicory to decrease the symptoms of Alzheimer's disease.

Hydroxybenzoic Acids as Acetylcholinesterase Inhibitors: Calorimetric and Docking Simulation Studies.

One of the symptoms of Alzheimer's disease (AD) is low acetylcholine level due to high acetylcholinesterase (AChE) activity. For this reason, AChE inhibitors are used in the treatment of AD, the prolonged use of which may cause a cholinergic crisis. There is a need to search for safe natural AChE inhibitors. The study analyzed 16 hydroxybenzoic acids using calorimetry and docking simulation as AChE inhibitors. All tested compounds were shown to inhibit the hydrolysis of ACh. The best properties were shown by methyl syringinate, which acted as competitive inhibitor at a catalytic site. The tested compounds also interacted with the anionic or peripheral binding site known to block ß-amyloid plaques formation. The activity of the tested hydroxybenzoic acids IC50 ranged from 5.50 to 34.19 µmol/µmol of AChE, and the binding constant Ka from 20.53 to 253.16 L/mol, which proves their reversible, non-toxic effect, and activity at physiological concentrations.

Bioaccessibility of coffee bean hydroxycinnamic acids during in vitro digestion influenced by the degree of roasting and activity of intestinal probiotic bacteria, and their activity in Caco-2 and HT29 cells.

In coffee beans, especially roasted, a significant part of hydroxycinnamic acid (HCAs) and their esters chlorogenic acids (CHAs) is attached to melanoidins through both covalent and non-covalent bonds. Bound and, to a greater extent, unbound HCAs, including those released from the polymerized material during digestion, can be pivotal in preventing of many chronic civilization diseases. The aim of the study was to determine the amount of free CHAs and those released from coffee extracts during in vitro digestion in various sections of the gastrointestinal tract, in the presence and absence of probiotic bacteria. The concentration of free CHAs was the lowest in the stomach and achieved the highest levels in the large intestine. Probiotic bacteria caused significant release of CHAs, and in the colon their concentration was the highest. The studies with Caco-2 and HT-29 cell lines showed that digested coffee extracts had cytoprotective potential against tert-BOOH induced oxidative stress.