Evaluation of the polyphenolic profile of native Ecuadorian stingless bee honeys (Tribe: Meliponini) and their antibiofilm activity on susceptible and multidrug-resistant pathogens: An exploratory analysis.

Biofilms are associated with infections that are resistant to conventional therapies, contributing to the antimicrobial resistance crisis. The need for alternative approaches against biofilms is well-known. Although natural products like stingless bee honeys (tribe: Meliponini) constitute an alternative treatment, much is still unknown. Our main goal was to evaluate the antibiofilm activity of stingless bee honey samples against multidrug-resistant (MDR) pathogens through biomass assays, fluorescence (cell count and viability), and scanning electron (structural composition) microscopy. We analyzed thirty-five honey samples at 15% (v/v) produced by ten different stingless bee species (Cephalotrigona sp., Melipona sp., M. cramptoni, M. fuscopilosa, M. grandis, M. indecisa, M. mimetica, M. nigrifacies, Scaptotrigona problanca, and Tetragonisca angustula) from five provinces of Ecuador (Tungurahua, Pastaza, El Oro, Los Ríos, and Loja) against 24-h biofilms of Staphylococcus aureus, Klebsiella pneumoniae, Candida albicans, and Candida tropicalis. The present honey set belonged to our previous study, where the samples were collected in 2018-2019 and their physicochemical parameters, chemical composition, mineral elements, and minimal inhibitory concentration (MIC) were screened. However, the polyphenolic profile and their antibiofilm activity on susceptible and multidrug-resistant pathogens were still unknown. According to polyphenolic profile of the honey samples, significant differences were observed according to their geographical origin in terms of the qualitative profiles. The five best honey samples (OR24.1, LR34, LO40, LO48, and LO53) belonging to S. problanca, Melipona sp., and M. indecisa were selected for further analysis due to their high biomass reduction values, identification of the stingless bee specimens, and previously reported physicochemical parameters. This subset of honey samples showed a range of 63-80% biofilm inhibition through biomass assays. Fluorescence microscopy (FM) analysis evidenced statistical log reduction in the cell count of honey-treated samples in all pathogens (P <0.05), except for S. aureus ATCC 25923. Concerning cell viability, C. tropicalis, K. pneumoniae ATCC 33495, and K. pneumoniae KPC significantly decreased (P <0.01) by 21.67, 25.69, and 45.62%, respectively. Finally, scanning electron microscopy (SEM) analysis demonstrated structural biofilm disruption through cell morphological parameters (such as area, size, and form). In relation to their polyphenolic profile, medioresinol was only found in the honey of Loja, while scopoletin, kaempferol, and quercetin were only identified in honey of Los Rios, and dihydrocaffeic and dihydroxyphenylacetic acids were only detected in honey of El Oro. All the five honey samples showed dihydrocoumaroylhexose, luteolin, and kaempferol rutinoside. To the authors' best knowledge, this is the first study to analyze stingless bees honey-treated biofilms of susceptible and/or MDR strains of S. aureus, K. pneumoniae, and Candida species.

Can the phenolic compounds of Manuka honey chemosensitize colon cancer stem cells? A deep insight into the effect on chemoresistance and self-renewal.

Manuka honey, which is rich in pinocembrin, quercetin, naringenin, salicylic, p-coumaric, ferulic, syringic and 3,4-dihydroxybenzoic acids, has been shown to have pleiotropic effects against colon cancer cells. In this study, potential chemosensitizing effects of Manuka honey against 5-Fluorouracil were investigated in colonspheres enriched with cancer stem cells (CSCs), which are responsible for chemoresistance. Results showed that 5-Fluorouracil increased when it was combined with Manuka honey by downregulating the gene expression of both ATP-binding cassette sub-family G member 2, an efflux pump and thymidylate synthase, the main target of 5-Fluorouracil which regulates the ex novo DNA synthesis. Manuka honey was associated with decreased self-renewal ability by CSCs, regulating expression of several genes in Wnt/ß-catenin, Hedgehog and Notch pathways. This preliminary study opens new areas of research into the effects of natural compounds in combination with pharmaceuticals and, potentially, increase efficacy or reduce adverse effects.

Influence of Altitudes and Development Stages on the Chemical Composition, Antioxidant, and Antimicrobial Capacity of the Wild Andean Blueberry (Vaccinium floribundum Kunth).

The chemical composition and biological capacities of berries depend on environmental parameters, maturity, and location. The Andean blueberry (Vaccinium floribundum Kunth), also known as mortiño, presents a unique combination of several phytochemicals, which play a synergistic role in its characterization as a functional food. We aimed to expose the possible variations that exist in the profile of the phenolic compounds as well as the antioxidant and antimicrobial capacity of the wild Andean blueberry with respect to three ripeness stages and two different altitudes. We found that polyphenols are the predominant compounds in the berry during the early ripeness stage and are the main bioactive compounds that give rise to the antioxidant capacity and inhibition effect on the growth of gram-positive and gram-negative bacteria. Moreover, the accumulation of ascorbic acid, free amino acids, and anthocyanins increases as the ripening process progresses, and they were the main bioactive compounds in the ripe berry. The latter compounds influence the production of the typical bluish or reddish coloration of ripe blueberries. In addition, it was determined that environmental conditions at high altitudes could have a positive influence in all cases. Overall, our data provide evidence regarding the high functional value of the wild Andean blueberry.

The reciprocal interaction between polyphenols and other dietary compounds: Impact on bioavailability, antioxidant capacity and other physico-chemical and nutritional parameters.

Polyphenols are plant secondary metabolites, whose biological activity has been widely demonstrated. However, the research in this field is a bit reductive, as very frequently the effect of individual compound is investigated in different experimental models, neglecting more complex, but common, relationships that are established in the diet. This review summarizes the data that highlighted the interaction between polyphenols and other food components, especially macro- (lipids, proteins, carbohydrates and fibers) and micronutrients (minerals, vitamins and organic pigments), paying particular attention on their bioavailability, antioxidant capacity and chemical, physical, organoleptic and nutritional characteristics. The topic of food interaction has yet to be extensively studied because a greater knowledge of the food chemistry behind these interactions and the variables that modify their effects, could offer innovations and improvements in various fields ranging from organoleptic, nutritional to health and economic field.

Three Amazonian palms as underestimated and little-known sources of nutrients, bioactive compounds and edible insects.

Mauritia flexuosa, Bactris gasipaes, and Oenocarpus bataua are among the main palms in the Amazon used for food and medicinal purposes. The food most commonly derived from these are fruits, oil, and the larvae of the insect Rhynchophorus palmarum reared in their trunks. Palm fruits are used for oil extraction as they are rich in saturated fatty acids, fiber, pro-vitamin A, carotenoids, tocopherols, macro and microelements, and polyphenols. Furthermore, the larvae of R. palmarum are rich in lipids, vitamin E, and proteins. This review analyzes the chemical composition of the fruit and oil of these palm species, as well as the R. palmarum larvae that breed in them. Our aim is to present information that is not widely known in order to demonstrate the potential of these palms as sources of plant-based and animal food with high nutritional and functional values.

Manuka honey in combination with 5-Fluorouracil decreases physical parameters of colonspheres enriched with cancer stem-like cells and reduces their resistance to apoptosis.

The aim of the present work was to evaluate the in vitro effect of Manuka honey and its combination with 5-Fu, the most common drug used in the treatment of colon cancer, on the morphological and physical parameters of colonspheres enriched with cancer stem-like cells deriving from HCT-116 colon adenocarcinoma cell line and on the apoptosis rate. Manuka honey, alone and more in combination with 5-Fu, reduced the weight, the diameter and mass density of the spheroids and induced apoptosis through the downregulation of many apoptosis inhibitors, including IAPs (Livin, Survivin, XIAP), IGFs (IGF-I, IGF-II and IGF-IR) and HSPs (HSP-27, HSP-60 and HSP-70). These results led to a reduction in the survival ability of cancer stem-like cells, as well as to a chemosensitizing effect of honey towards 5-Fu, considering that apoptosis resistance is one of the main causes of cancer stem-like cells chemoresistance.

Strawberry tree honey in combination with 5-fluorouracil enhances chemosensitivity in human colon adenocarcinoma cells.

Colorectal cancer remains a challenging health burden worldwide. This study aimed to assess the potentiality of Strawberry tree honey (STH), a polyphenol-enriched food, to increase the effectiveness of 5-Fluorouracil (5-FU) in adenocarcinoma (HCT-116) and metastatic (LoVo) colon cancer cell lines. The combined treatment reduced cell viability and caused oxidative stress, by increasing oxidative biomarkers and decreasing antioxidant defence, in a more potent way compared to 5-FU alone. The expression of endoplasmic reticulum (ATF-6, XBP-1) and MAPK (p-p38 MAPK, p-ERK1/2) markers were also elevated after the combined treatment, enhancing the cell cycle arrest through the modulation of regulatory genes (i.e., cyclins and CDKs). Apoptotic gene (i.e., caspases) expressions were also increased after the combined treatment, while those of proliferation (i.e., EGFR), cell migration, invasion (i.e., matrix metallopeptidase) and epithelial-mesenchymal transition (N-cadherin, ß-catenin) were suppressed. Finally, the combined treatment led cell metabolism towards a quiescent stage, by reducing mitochondrial respiration and glycolysis. In conclusion, this work represents an initial step to highlight the possibility to use STH in combination with 5-FU in the treatment of colon cancer, even if further in vitro an in vivo studies are strongly needed to confirm the possible chemo-sensitizing effects of STH.

The Molecular Basis of Different Approaches for the Study of Cancer Stem Cells and the Advantages and Disadvantages of a Three-Dimensional Culture.

Cancer stem cells (CSCs) are a rare tumor subpopulation with high differentiation, proliferative and tumorigenic potential compared to the remaining tumor population. CSCs were first discovered by Bonnet and Dick in 1997 in acute myeloid leukemia. The identification and isolation of these cells in this pioneering study were carried out through the flow cytometry, exploiting the presence of specific cell surface molecular markers (CD34+/CD38-). In the following years, different strategies and projects have been developed for the study of CSCs, which are basically divided into surface markers assays and functional assays; some of these techniques also allow working with a cellular model that better mimics the tumor architecture. The purpose of this mini review is to summarize and briefly describe all the current methods used for the identification, isolation and enrichment of CSCs, describing, where possible, the molecular basis, the advantages and disadvantages of each technique with a particular focus on those that offer a three-dimensional culture.

Nutritional Value and Preventive Role of Nigella sativa L. and Its Main Component Thymoquinone in Cancer: An Evidenced-Based Review of Preclinical and Clinical Studies.

In recent times, scientific attention has been paid to different foods and their bioactive components for the ability to inhibit the onset and progress of different types of cancer. Nigella sativa extract, powder and seed oil and its main components, thymoquinone and α-hederin, have showed potent anticancer and chemosensitizing effects against various types of cancer, such as liver, colon, breast, renal, cervical, lung, ovarian, pancreatic, prostate and skin tumors, through the modulation of various molecular signaling pathways. Herein, the purpose of this review was to highlight the anticancer activity of Nigella sativa and it constitutes, focusing on different in vitro, in vivo and clinical studies and projects, in order to underline their antiproliferative, proapoptotic, cytotoxic and antimetastatic effects. Particular attention has been also given to the synergistic effect of Nigella sativa and it constitutes with chemotherapeutic drugs, and to the synthesized analogs of thymoquinone that seem to enhance the chemo-sensitizing potential. This review could be a useful step towards new research on N. sativa and cancer, to include this plant in the dietary treatments in support to conventional therapies, for the best achievement of therapeutic goals.

Bioactive compounds, phenolic profile, antioxidant capacity and effectiveness against lipid peroxidation of cell membranes of Mauritia flexuosa L. fruit extracts from three biomes in the Ecuadorian Amazon.

The bioactive compounds, total antioxidant capacity and protective effect against lipid oxidative damage of red blood cell (RBC) membranes of Mauritia flexuosa L. extracts from three altitude levels (low, middle and high zones) were analyzed. The identification of the main polyphenols in the fruit was performed using High Performance Liquid Chromatography/Electrospray Ionization Tandem Mass Spectrometry analysis, while High-Performance Liquid Chromatography - Diode Array Detector was applied for the vitamin C and carotenoid analysis. The ability of the extracts (methanolic, free polyphenols and bound polyphenols) to protect RBC membranes against oxidative destruction was assessed utilizing the thiobarbituric acid reactive substance assay (TBARS) assay. Among the 14 phenolic compounds identified, quercetin glycosides were the most predominant ones. The fruit from the middle altitude zone showed higher vitamin C and lutein contents than those from the low and high areas, whilst higher values of ß-Carotene were obtained in M. flexuosa L. from the low zone. The contents of flavonoids and total polyphenols were lower in fruit from the low zone, whilst the fruit from the middle zone had the highest values. Similar results were observed in the results of the antioxidant capacity assays. However, a significant difference was not found in the protective effect of the extracts from the three altitude zones against lipid peroxidation in RBC membranes.

Nutraceutical Compounds Targeting Inflammasomes in Human Diseases.

The macromolecular complex known as "inflammasome" is defined as an intracellular multi-protein complex composed of a sensor receptor (PRR), an adaptor protein and an effector enzyme (caspase-1), which oligomerize when they sense danger, such as how the NLR family, AIM-2 and RIG-1 receptors protect the body against danger via cytokine secretion. Within the NLR members, NLRP3 is the most widely known and studied inflammasome and has been linked to many diseases. Nowadays, people's interest in their lifestyles and nutritional habits is increasing, mainly due to the large number of diseases that seem to be related to both. The term "nutraceutical" has recently emerged as a hybrid term between "nutrition" and "pharmacological" and it refers to a wide range of bioactive compounds contained in food with relevant effects on human health. The relationship between these compounds and diseases based on inflammatory processes has been widely exposed and the compounds stand out as an alternative to the pathological consequences that inflammatory processes may have, beyond their defense and repair action. Against this backdrop, here we review the results of studies using several nutraceutical compounds in common diseases associated with the inflammation and activation of the NLRP3 inflammasomes complex. In general, it was found that there is a wide range of nutraceuticals with effects through different molecular pathways that affect the activation of the inflammasome complex, with positive effects mainly in cardiovascular, neurological diseases, cancer and type 2 diabetes.

Protective effect of the medicinal herb infusion "horchata" against oxidative damage in cigarette smokers: An ex vivo study.

Cigarette smoking has been associated with an increase in oxidative stress (OS) and is considered a predisposing factor to chronic noncommunicable diseases, whilst dietary antioxidants has been proposed as an alternative to cope with this oxidative stress. In this study, 20 smokers and 20 non-smokers were studied with the aim of determining their antioxidant status, as well as the ability of an infusion of 23 medicinal plants, to counteract the damage caused by OS. The plasma, red blood cells (RBCs) and polymorphonuclear cells (PBMCs) of both groups were incubated or not with the horchata infusion extract and then the OS markers, genotoxicity, nanostructure of RBCs membrane and genes related to oxidative responses and cellular functionality were evaluated. Up to 33 different compounds, mainly quercetin glycosides, were identified in the extract. A significant deterioration in the antioxidant status in smokers compared to non-smokers was found. The horchata infusion extract improved the nanostructure of RBCs and DNA damage, as well as the activity of the endogenous antioxidant enzymes and markers of oxidative damage to lipid, and proteins in plasma, RBCs and PBMCs in both groups, whilst no significant changes were found in the expression of different genes related to OS response.

Phytochemical Analysis and Biological Investigation of Nepeta juncea Benth. Different Extracts.

This study was carried out to screen the amount and the classes of secondary metabolites and to evaluate the antioxidant, cytotoxic, antifungal, and antibacterial activities of the methanolic, ethanolic, and water extracts of the roots, leaves, and flowers of Nepeta juncea Benth. The results show that the highest total phenol (69.54 ± 0.31 mg gallic acid equivalents (GAE)/g dry weight), total flavonoid (41.37 ± 0.17 mg quercetin equivalents (QE)/g dry weight), anthocyanin (6.52 ± 0.21 mg cyanidin/100 g dry weight), and tannin (47.36 ± 0.33 mg catechin/g dry weight) concentrations were recorded in the methanolic extract of the leaves of N. juncea. The gas chromatography-mass spectrometry (GC-MS) analysis of the extracts showed that 1,8-cineole, 4aα-7α-7aα-nepetalactone, ß-pinene, terpinen-4-ol, and α-terpineol were the major compounds, respectively. The best 2, 2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging and ferric-reducing antioxidant, cytotoxic, antifungal, and antibacterial activities were observed for the methanolic extract of the leaves. For the two latter activities, the best activity was revealed on Staphylococcus aureus, Bacillus cereus, and Candida albicans. The minimum inhibitory concentration (MIC) values for the antimicrobial of the methanolic extract from the leaves were in the range of 25-100 µg/mL, whereas the minimum bactericidal concentration (MBC) values were in the range of 50-200 µg/mL. The results reported herein show that, for the first time in the literature, N. juncea is a remarkable source of antioxidant, antifungal, and antibacterial compounds.

Anti-inflammatory effect of the medicinal herbal mixture infusion, Horchata, from southern Ecuador against LPS-induced cytotoxic damage in RAW 264.7 macrophages.

The phytochemical composition and the antioxidant and anti-inflammatory activities of a mixture of 23 plants, named Horchata, traditionally consumed in Ecuador, have been evaluated. The study was carried out using the hydroalcoholic extract (HHext) and infusion (IHext) of the horchata plant mixture. It was verified that thermal treatment affected the contents of vitamin C and carotenoids, but hardly those of polyphenols, which would be the main bioactive compounds in the infusion, the common form of preparation of horchata for consumption. Among phenolic compounds, caffeoylquinic acids, flavones and flavonols (mostly quercetin glycosides) were prominent. Both HHext and IHext extracts managed to protect RAW 264.7 macrophages against LPS-induced cytotoxic damage, increasing the levels of endogenous antioxidant enzymes and modulating the production of pro-inflammatory and anti-inflammatory cytokines. Greater protective effects were obtained for HHext compared to IHext, which was in agreement with its higher content of phenolic compounds favoured by a more efficient extraction in the hydroalcoholic medium. Nonetheless, the infusion still maintained a significant antioxidant and anti-inflammatory activity, which would support the protective effects on health traditionally attributed to its consumption by the population.

Chemical Composition and Antioxidant Activity of the Main Fruits, Tubers and Legumes Traditionally Consumed in the Andean Regions of Ecuador as a Source of Health-Promoting Compounds.

Thirteen fruits, eight legumes and three tubers consumed in the Andean regions of Ecuador were studied to determine their bioactive compounds, organic acids, sugars content, total antioxidant capacity, as well as to determine which among them showed the greatest contribution in terms of antioxidant activity and which compounds contributed to it. Among fruits, taxo fruits (Passiflora mollissima (Kunth) L.H. Bailey) presented the highest values of total phenolic, carotene content, and total antioxidant capacity. The ají ratón (Capsicum chinense Jacq) showed the highest content of vitamin C. Taxo showed the highest content of ß carotene, whilst lycopene was identified only in guayaba fruits (Psidium guajava L.) and ají ratón was the principal source of lutein. In legumes, chocho perla (Lupinus mutabilis Sweet) showed the highest values for both total phenolic and flavonoid content, whilst frejol negro (Phaseolus vulgaris L.) and frejol canario (Vigna unguiculata (L.) Walp) showed the highest values for FRAP and DPPH assay, respectively. Between tubers, the jícama (Smallanthus sonchifolius (Poepp.) H. Rob.) had the majors values in terms of total phenolic, flavonoid content, and total antioxidant capacity. In terms of total antioxidant capacity, taxo fruits have the highest contribution in terms of total antioxidant capacity, whilst the dendogram shown the occurrence of five distinct groups in which taxo was located in the first largest group. Our data contributing towards gaining better knowledge about the Andean Ecuadorian diet and the composition of Andean food in order to estimate dietary intakes of health-promoting components.

Guava (Psidium guajava L. cv. Red Suprema) Crude Extract Protect Human Dermal Fibroblasts against Cytotoxic Damage Mediated by Oxidative Stress.

We analyzed guava fruits (Psidium guajava L. cv. Red Suprema) from Cuba to determine their chemical composition, total antioxidant capacity, as well as their protective effect against oxidative damage using an in vitro model of human dermal fibroblasts. The guava fruit is a natural source of bioactive compounds, such as polyphenols, vitamin C, folates and beta carotenes with proven health benefits. Human dermal fibroblasts were pre-incubated with different concentrations of guava crude extract and then subjected to oxidative stress using the AAPH stressor. The number of apoptotic and dead cells, as well as the markers of oxidative damage such as lipid and protein oxidation significantly decreased when cells were pre-incubated with guava crude extract and then exposed to the stressor. The activity of antioxidant enzymes also improved when cells were pre-incubated with guava crude extract in comparison to cells subjected to stress without prior pre-incubation with the guava extract. The results obtained in this study highlight the health benefits of guava regarding oxidative stress, proving it to be an important source of bioactive compounds associated with important biological properties.

Overexpression of the Anthocyanidin Synthase Gene in Strawberry Enhances Antioxidant Capacity and Cytotoxic Effects on Human Hepatic Cancer Cells.

Food fortification through the increase and/or modulation of bioactive compounds has become a major goal for preventing several diseases, including cancer. Here, strawberry lines of cv. Calypso transformed with a construct containing an anthocyanidin synthase (ANS) gene were produced to study the effects on anthocyanin biosynthesis, metabolism, and transcriptome. Three strawberry ANS transgenic lines (ANS L5, ANS L15, and ANS L18) were analyzed for phytochemical composition and total antioxidant capacity (TAC), and their fruit extracts were assessed for cytotoxic effects on hepatocellular carcinoma. ANS L18 fruits had the highest levels of total phenolics and flavonoids, while those of ANS L15 had the highest anthocyanin concentration; TAC positively correlated with total polyphenol content. Fruit transcriptome was also specifically affected in the polyphenol biosynthesis and in other related metabolic pathways. Fruit extracts of all lines exerted cytotoxic effects in a dose/time-dependent manner, increasing cellular apoptosis and free radical levels and impairing mitochondrial functionality.

Dietary polyphenols: Structures, bioavailability and protective effects against atherosclerosis.

Epidemiological studies have demonstrated that nutritional habits, like those based on high consumption of fruits and vegetables, have been associated with a longer life expectancy and a significant decrease in the incidence and prevalence of several chronic diseases with inflammatory basis, such as cardiovascular diseases (CVD). This beneficial activity has been related to the content of several bioactive compounds in fruit and vegetables, such as polyphenols. The cardioprotective effects of polyphenols have been linked mainly to its antioxidant properties; however, recent findings attribute its anti-atherosclerotic potential to modulate simultaneous signaling and mechanistic pathways. Emerging data suggest that polyphenols can regulate cellular lipid metabolism; vascular and endothelial function; haemostasis; as well as platelet function; which represent primary conditions for atherosclerotic plaque formation and development. This review presents the results of a selection of experimental studies and clinical trials regarding the atheroprotective effects of the most common dietary polyphenols.

The protective effect of acerola (Malpighia emarginata) against oxidative damage in human dermal fibroblasts through the improvement of antioxidant enzyme activity and mitochondrial functionality.

Acerola fruits (Malpighia emarginata DC.) from the central region of Cuba were analyzed to determine their chemical composition and protective capacity against oxidative damage using an in vitro human dermal fibroblast (HDFa) model. The chemical composition analyses showed a high content of vitamin C, total polyphenols, ß-carotene and folates in the acerola fruit. From the HPLC-DAD/ESI-MSn analyses, two anthocyanins (cyanidin 3-O-rhamnoside and pelargonidin 3-O-rhamnoside), three hydroxycinnamoyl derivatives (caffeoyl hexoside, dihydrocaffeoylquinic acid and coumaroyl hexoside) and fifteen flavonols (mostly glycosylated forms of quercetin and kaempferol) were detected. HDFa were pre-incubated with an acerola crude extract (ACExt) and subsequently subjected to oxidative stress induced by AAPH. Apoptosis, intracellular ROS and the biomarkers of lipid and protein oxidation significantly increased after inducing stress, while the activities of the antioxidant enzyme catalase and superoxide dismutase and mitochondrial functionality were markedly affected. However, ACExt was able to protect against oxidative damage through decreasing apoptosis, intracellular ROS levels and lipid and protein damage, besides improving antioxidant enzyme activities and mitochondrial functionality. The obtained results support acerola fruits as relevant sources of functional compounds with promising effects on human health.

Strawberry consumption improves aging-associated impairments, mitochondrial biogenesis and functionality through the AMP-activated protein kinase signaling cascade.

Dietary polyphenols have been recently proposed as activators of the AMP-activated protein kinase (AMPK) signaling pathway and this fact might explain the relationship between the consumption of polyphenol-rich foods and the slowdown of the progression of aging. In the present work, the effects of strawberry consumption were evaluated on biomarkers of oxidative damage and on aging-associated reductions in mitochondrial function and biogenesis for 8weeks in old rats. Strawberry supplementation increased antioxidant enzyme activities, mitochondrial biomass and functionality, and decreased intracellular ROS levels and biomarkers of protein, lipid and DNA damage (P<0.05). Furthermore, a significant (P<0.05) increase in the expression of the AMPK cascade genes, involved in mitochondrial biogenesis and antioxidant defences, was also detected after strawberry intake. These in vivo results were then verified in vitro on HepG2 cells, confirming the involvement of AMPK in the beneficial effects exerted by strawberry against aging progression.