Therapeutic potential of plant-derived extracellular vesicles as nanocarriers for exogenous miRNAs.

Cell-to-cell communication strategies include extracellular vesicles (EVs) in plants and animals. The bioactive molecules in a diet rich in vegetables and fruits are associated with disease-preventive effects. Plant-derived EVs (PDEVs) are biogenetically and morphologically comparable to mammalian EVs and transport bioactive molecules, including miRNAs. However, the biological functions of PDEVs are not fully understood, and standard isolation protocols are lacking. Here, PDEVs were isolated from four foods with a combination of ultracentrifugation and size exclusion chromatography, and evaluated as vehicles for enhanced transport of synthetic miRNAs. In addition, the role of food-derived EVs as carriers of dietary (poly)phenols and other secondary metabolites was investigated. EVs from broccoli, pomegranate, apple, and orange were efficiently isolated and characterized. In all four sources, 4 miRNA families were present in tissues and EVs. miRNAs present in broccoli and fruit-derived EVs showed a reduced RNase degradation and were ferried inside exposed cells. EVs transfected with a combination of ath-miR159a, ath-miR162a-3p, ath-miR166b-3p, and ath-miR396b-5p showed toxic effects on human cells, as did natural broccoli EVs alone. PDEVs transport trace amounts of phytochemicals, including flavonoids, anthocyanidins, phenolic acids, or glucosinolates. Thus, PDEVs can act as nanocarriers for functional miRNAs that could be used in RNA-based therapy.

Antinociceptive and antiedema effects produced in rats by Brassica oleracea var. italica sprouts involving sulforaphane.

Natural products are recognized as potential analgesics since many of them are part of modern medicine to relieve pain without serious adverse effects. The aim of this study was to investigate the antinociceptive and anti-inflammatory activities of an aqueous extract of Brassica oleracea var. italica sprouts (AEBS) and one of its main reported bioactive metabolites sulforaphane (SFN). Antinociceptive activity of the AEBS (30, 100, and 300 mg/kg, i.p. or 1000 and 2000 mg/kg, p.o.) and SFN (0.1 mg/kg, i.p.) was evaluated in the plantar test in rats to reinforce its analgesic-like activity at central level using the reference drug tramadol (TR, 50 mg/kg, i.p.). The anti-inflammatory-like response was determined in the carrageenan-induced oedema at the same dosages for comparison with ketorolac (KET, 20 mg/kg, i.p.) or indomethacin (INDO, 20 mg/kg, p.o.). A histological analysis of the swollen paw was included to complement the anti-inflammatory response. Additionally, acute toxicity observed in clinical analgesics as the most common adverse effects, such as sedation and/or gastric damage, was also explored. As a result, central and peripheral action of the AEBS was confirmed using enteral and parenteral administration, in which significant reduction of the nociceptive and inflammatory responses resembled the effects of TR, KET, or INDO, respectively, involving the presence of SFN. No adverse or toxic effects were observed in the presence of the AEBS or SFN. In conclusion, this study supports that Brassica oleracea var. italica sprouts are a potential source of antinociceptive natural products such as SFN for therapy of pain alone and associated to an inflammation condition.

Winery By-Products as Sources of Bioactive Tryptophan, Serotonin, and Melatonin: Contributions to the Antioxidant Power.

The amino acid tryptophan and its derived molecules serotonin and melatonin are involved in a wide range of physiological functions that contribute significantly to human health, namely antioxidant, immune-active, and neurological properties. Grapes and wine are a source of these compounds, but their presence in wine by-products remains underexplored. Therefore, the aim of this work was the identification and quantification of tryptophan, serotonin, and melatonin in winery by-products (grape stems, grape pomace, and wine lees) by ultra-high performance liquid chromatography coupled to electrospray ionization and mass spectrometer with triple-quadrupole technology (UHPLC-ESI-QqQ-MS/MS), as well as the evaluation of the extracts obtained (by applying specific extraction conditions for each of them) for their antioxidant and reducing capacity (by three different and complementary methods: FRAP, ABTS•+, and ORAC). Furthermore, correlation analyses were developed to establish the contribution of the different analytes to the total antioxidant activity. The main results obtained pointed out grape stems as the by-product with the highest tryptophan content (96.28 mg/kg dw) and antioxidant capacity (142.86, 166.72, and 363.24 mmol TE/kg dw, FRAP, ABTS•+, and ORAC, respectively), while serotonin and melatonin were the predominant derivatives in grape pomace (0.086 and 0.902 µg/kg dw, respectively). The antioxidant capacity of the standards was also analysed at the concentrations found in the matrices studied. A significant correlation was found between the concentration of the pure tryptophan standard and the antioxidant capacity (ABTS•+, r2 = 0.891 at p < 0.001 (***); FRAP, r2 = 0.885 at p < 0.01 (**); and ORAC, r2 = 0.854 at p < 0.01 (**)). According to these results, winery by-products can be highlighted as valuable materials to be used as novel ingredients containing tryptophan, serotonin, and melatonin, while tryptophan was identified as the most relevant contributor (out of phenolic compounds) to the antioxidant capacity exhibited by wine by-products.

Microwave Treatment vs. Conventional Pasteurization: The Effect on Phytochemical and Microbiological Quality for Citrus-Maqui Beverages.

This study uncovered the impacts of microwave (MW) treatments compared to conventional pasteurization (TP) on the quality of functional citrus-maqui beverages, with added sucrose or stevia. The influence of these thermal treatments on the microbiological burden and phytochemical composition was determined by processing under two MW power levels (600 W and 800 W) and TP at 85 °C for 15 s for 60 days at room temperature (20 °C). The results indicated that, beyond the microbiological quality achieved in the juices treated by both MW and TP technology, there were no differences among the treatments regarding the stability of vitamin C, anthocyanin, and flavanone concentrations. However, anthocyanins were more stable in those beverages with sucrose added, rendering a better red color. Besides, all treatments ensured microbiological stability throughout the entire storage time. In conclusion, MW treatment could be considered as an alternative to TP, which ensures microbial safety, protecting functional compounds associated with health effects.

Potential Activity of Abrantes Pollen Extract: Biochemical and Cellular Model Studies.

The aim of this study was to determine the grain composition and (poly)phenolic profile of pollen from Abrantes (Portugal), as well as its antioxidative and antidiabetic properties, and abilities to protect human erythrocytes against induced hemoglobin oxidation, lipid peroxidation, and hemolysis. The phytochemical profile of the Abrantes' bee pollen revealed twenty phenolic compounds, identified by high-performance liquid chromatography with electrospray ionization mass spectrometry coupled with photodiode array detection. Among them, quercetin derivatives were the most abundant. Concerning the biological potential, the pollen extract showed notable capacity for 2,2-diphenyl-1-picrylhydrazyl, nitric oxide, and superoxide radicals, as well as for inhibition of α-glucosidase action, and protection of human erythrocytes against oxidative damage. Non-cytotoxic effects regarding the NHDF normal cell line, human adenocarcinoma Caco-2, and human liver HepG2 cells were observed. The results obtained contributed to further research on modes of action related to oxidative damage and metabolic health problems, to generate deeper knowledge of potential health-promoting effects to develop novel pharmaceutical drugs, nutraceuticals, and dietary supplements.

Valorisation of Prunus avium L. By-Products: Phenolic Composition and Effect on Caco-2 Cells Viability.

Prunus avium L. by-products, such as stems, leaves, and flowers, are used in folk medicine to prevent and treat some diseases. However, their phenolic composition and in vitro bioactivities against tumor cells are poorly known. In this work, we compared the phenolic profile and the biological potential of aqueous infusions and hydroethanolic extracts of P. avium leaves, stems, and flowers from Saco cultivar, collected from the Fundão region (Portugal). Among the fifty-two phenolic compounds tentatively identified by HPLC-DAD-ESI/MSn, the hydroxycinnamic acids were the most abundant. Both extracts of stems revealed a higher activity against DPPH•. Meanwhile, hydroethanolic extracts from stems and flowers and aqueous infusions of flowers were the most effective in inhibiting the growth of the human epithelial colorectal adenocarcinoma (Caco-2) cells at concentrations above 200 µg/mL. More detailed knowledge about the phenolic composition and health-promoting properties of Portuguese P. avium by-products allows for increasing the biological and commercial value of these bio-wastes, which may have a positive impact on food and pharmaceutical industries, as on the valorization of the local economy.

Antinociceptive effects of maqui-berry (Aristotelia chilensis (Mol.) Stuntz).

Maqui-berry is characterised by presenting a high concentration of (poly)phenols, accounting anthocyanins (cyanidin and delphinidin) for over 85% of the total. These coloured flavonoids have demonstrated potential neurological activity, but the evidence of their antinociceptive properties is scarce. In order to cover this gap, different doses (suitable for human administration) of a maqui-berry powder (1.6% anthocyanin), using enteral and parenteral routes of administration, were compared at central and peripheral levels using a nociceptive pain model (formalin test) in mice. Gastric damage analysis as possible adverse effects of analgesic and anti-inflammatory drugs was also explored. Dose-antinociceptive response was confirmed using both routes of administration and in both neurogenic and inflammatory phases of the formalin test, without gastric damage. In conclusion, these preliminary data provide evidence of pharmacological properties of maqui-berry to alleviate nociceptive pain.

Maqui berry (Aristotelia chilensis) extract improves memory and decreases oxidative stress in male rat brain exposed to ozone.

Introduction: Prolonged ozone exposure can produce a state of oxidative stress, which in turn causes alterations in the dynamics of the brain and affects memory and learning. Moreover, different investigations have shown that high flavonoid content berries show a great antioxidant activity. The relationship between the protective effect of the maqui berry extract and its antioxidant properties in the brain has not been studied in depth. Objectives: The present study evaluated whether the protection exerted by the aqueous extract of maqui berry in brain regions associated with cognitive performance is due to its antioxidant capacity. Methods: Sprague Dawley rats were exposed to 0.25 ppm ozone and administered with maqui berry extracts. At the end of the treatments, spatial learning and short- and long-term memory were evaluated, as well as oxidative stress markers. Results: The administration of 50 and 100 mg/kg of the aqueous extract of maqui berry was effective in preventing the cognitive deficit caused by chronic exposure to ozone. The antioxidant effect of the administration of maqui berry was analyzed in the prefrontal cortex, hippocampus, and amygdala. Oxidative stress markers levels decreased and the enzymatic activity of superoxide dismutase diminished in animals exposed to ozone treated with the 50 mg/kg dose of maqui berry. Discussion: These results show a relationship between protection at the cognitive level and a decrease in oxidative stress markers, which suggests that the prevention of cognitive damage is due to the antioxidant activity of the maqui berry.

Chicory (Cichorium intybus L.) as a food ingredient - Nutritional composition, bioactivity, safety, and health claims: A review.

Chicory (Cichorium intybus L.) is a perennial herb from the Cichorium genus, Asteraceae family, and is worldwide cultivated. So far, chicory has been used mainly in animal feed, but also in several cases in the food industry: as salad, for teas and tea blends, for coffee supplementation, and as a source for the inulin production. Nowadays there is an increasing interest in chicory utilization for food production and supplementation. Some compounds present in chicory, such as polyphenols, inulin, oligofructose and sesquiterpene lactones may be considered as potential carriers of food functionality. This review describes nutritional, mineral and bioactive composition of the chicory plant and summarized the main biological activities associated with the presence of bioactive compounds in the different plant parts. Finally, the review explores possibilities of uses of chicory and its implementation in food products, with intention to design new functional foods.

Stevia vs. Sucrose: Influence on the Phytochemical Content of a Citrus-Maqui Beverage-A Shelf Life Study.

The consumption of sugar-sweetened beverages has been related with the risk of cardiovascular diseases and other pathophysiological situations, such as obesity or diabetes mellitus. Given the increasing awareness on this fact, food industries are developing new products to reduce the amount of added sugar in development of food products development. Accordingly, in the present work, new functional beverages, constituting a dietary source of bioactive phenolics and supplemented with stevia or sucrose, were designed in order to study the influence of the sweetener during processing and shelf-life. This study is of critical for the informed selection of the sweetener based on its effect on the final phytochemical profile of beverages, especially taking into consideration that there are no previous studies on Stevia rebaudiana. Physicochemical features and phytochemical composition, as well as stability of the different beverages concerning these parameters, were evaluated for 90 days during storage under different conditions (refrigeration (4 °C) and room temperature (25 °C) under light or darkness conditions). Physicochemical parameters (pH, titratable acidity, total soluble solids, and color) did not display statistically significant differences between beverages. Storage temperature was the greatest determinant affecting the stability of all the analyzed bioactive compounds (vitamin C, anthocyanins, and flavanones). The main difference between sweeteners was observed in flavanones, which exhibited a higher loss during storage under day light conditions when stevia was added instead of sucrose. In addition, the juices' colors were rather stable, keeping a reddish coloration and natural appearance throughout the shelf life. Hence, stevia could be considered as an alternative sweetener by the beverage industry.

Differential Catabolism of an Anthocyanin-Rich Elderberry Extract by Three Gut Microbiota Bacterial Species.

Elderberries are good sources of anthocyanins, which are poorly absorbed in the upper gastrointestinal tract but extensively transformed into phenolic metabolites at the colonic level. Because different gut microbiota strains have different metabolism, the catabolism of anthocyanins may lead to interindividual differences in metabolite production. In this work, an anthocyanin-rich elderberry extract was incubated with three single gut microbial strains (Enterobacter cancerogenous, Bifidobacterium dentium, and Dorea longicatena) up to 4 days, to assess differences in their phenolic metabolism. All of the strains degraded the elderberry anthocyanins, but the metabolic pathways followed were different. Although some metabolites were common for all of the strains, a wide disparity was observed in the kind and amount of several phenolic metabolites produced by each species. These in vitro preliminary results may be of help in the interpretation of the bioavailability of anthocyanins and give a clue to understand interindividual variability in metabolite production.

Sorting out the Value of Cruciferous Sprouts as Sources of Bioactive Compounds for Nutrition and Health.

Edible sprouts with germinating seeds of a few days of age are naturally rich in nutrients and other bioactive compounds. Among them, the cruciferous (Brassicaceae) sprouts stand out due to their high contents of glucosinolates (GLSs) and phenolic compounds. In order to obtain sprouts enriched in these phytochemicals, elicitation is being increasing used as a sustainable practice. Besides, the evidence regarding the bioavailability and the biological activity of these compounds after their dietary intake has also attracted growing interest in recent years, supporting the intake of the natural food instead of enriched ingredients or extracts. Also, there is a growing interest regarding their uses, consumption, and applications for health and wellbeing, in different industrial sectors. In this context, the present review aims to compile and update the available knowledge on the fundamental aspects of production, enrichment in composition, and the benefits upon consumption of diverse edible cruciferous sprouts, which are sources of phenolic compounds and glucosinolates, as well as the evidence on their biological actions in diverse pathophysiological situations and the molecular pathways involved.

Waking Up from Four Decades' Long Dream of Valorizing Agro-Food Byproducts: Toward Practical Applications of the Gained Knowledge.

The late 1970s was the onset of literature about the first research outputs on alternatives to create added-value to agro-food byproducts focused on the reduction of the dependency on raw materials and, simultaneously, helping to reduce the environmental impacts of the agricultural activities. This trend, increased over the years and during the past decade, has been boosted by the growing concern of the socio-economic impact of wastes from agro-food activities, however, with little success of the proposed applications. Throughout four decades an array of studies have emerged, aimed to gain evidence on the relevance of innovation in the agro-food industry, as well as to overcome this situation. To our knowledge, only a few cases, summarized in the present perspective, represent the main alternatives currently available for the valorization of agro-food byproducts, with indications of some constraints that need to be addressed, in the coming years, to obtain a real profit from these products.

Aqueous Extract of Pomegranate Alone or in Combination with Citalopram Produces Antidepressant-Like Effects in an Animal Model of Menopause: Participation of Estrogen Receptors.

It has been reported that the aqueous extract of pomegranate (AE-PG) has polyphenols with estrogenic-like activities. The present work determines if AE-PG alone or in combination with the selective serotonin reuptake inhibitor, citalopram, has antidepressant-like effects. It was also analyzed the participation of estrogen receptors (ER). AE-PG (0.1, 1.0, 10, or 100 mg/kg) was evaluated in ovariectomized female Wistar rats subjected to the forced swimming test. The effects induced by AE-PG were compared with those of citalopram (2.5, 5.0, 10, and 20.0 mg/kg) and 17ß-estradiol (E2; 2.5 5.0, and 10 µg/rat). Likewise, the combination of suboptimal doses of AE-PG (0.1 mg/kg) plus citalopram (2.5 mg/kg) was evaluated. To determine if ER participates in the antidepressant-like action of pomegranate, the estrogen antagonist tamoxifen (15 mg/kg) was administered with AE-PG (1 mg/kg). AE-PG produced antidepressant-like actions with a similar behavioral profile induced by citalopram and E2. Suboptimal doses of citalopram plus AE-PG produced antidepressant-like effects. Tamoxifen was able to block AE-PG's antidepressant-like actions. These results confirm the participation of ER in AE-PG's antidepressant-like effects. Furthermore, the additive effects observed with the combined treatment of AE-PG plus citalopram could be advantageous in the treatment of depressive disorders, such as menopause.

Broccoli sprouts in analgesia - preclinical in vivo studies.

BACKGROUND: Broccoli is a rich source of health-promoting glucosinolates, phenolic compounds, minerals and vitamins, which might have potential to alleviate pain. AIM: To explore the antinociceptive effects of a broccoli sprout aqueous extract (BSE) in experimental models of pain and an opioid mechanism. MATERIALS AND METHODS: the BSE was administered to mice and rats that were subjected to the writhing and formalin tests, respectively. Gastric damage or sedative-like response, as adverse effects observed in anti-inflammatory non-steroidal and opioid analgesic drugs, respectively, were also explored. RESULTS: Antinociception, but not sedative or gastric injury response, was observed in a significant and dose-dependent manner with the BSE (50-500 mg kg-1, i.p. and 500-2000 mg kg-1, p.o.) in comparison to the control group; these effects resembled those observed with the analgesic tramadol (30 mg kg-1, i.p.) in writhing and formalin assessments. Blockage of opioid receptors by naloxone (1 mg kg-1, i.p.) produced partial inhibition of the antinociceptive effect of the BSE in both assays. CONCLUSION: This study gives evidence of the potential activity of broccoli sprouts in pain therapy.

Metabolic Activity of Radish Sprouts Derived Isothiocyanates in Drosophila melanogaster.

We used Drosophila melanogaster as a model system to study the absorption, metabolism and potential health benefits of plant bioactives derived from radish sprouts (Raphanus sativus cv. Rambo), a Brassicaceae species rich in glucosinolates and other phytochemicals. Flies were subjected to a diet supplemented with lyophilized radish sprouts (10.6 g/L) for 10 days, containing high amounts of glucoraphenin and glucoraphasatin, which can be hydrolyzed by myrosinase to the isothiocyanates sulforaphene and raphasatin, respectively. We demonstrate that Drosophila melanogaster takes up and metabolizes isothiocyanates from radish sprouts through the detection of the metabolite sulforaphane-cysteine in fly homogenates. Moreover, we report a decrease in the glucose content of flies, an upregulation of spargel expression, the Drosophila homolog of the mammalian PPARγ-coactivator 1 α, as well as the inhibition of α-amylase and α-glucosidase in vitro. Overall, we show that the consumption of radish sprouts affects energy metabolism in Drosophila melanogaster which is reflected by lower glucose levels and an increased expression of spargel, a central player in mitochondrial biogenesis. These processes are often affected in chronic diseases associated with aging, including type II diabetes mellitus.

Stability of the Ellagitannin Fraction and Antioxidant Capacity of Varietal Pomegranate Juices.

This work aimed to assess the effect of combining two pomegranate (Punica granatum L.) cultivars at different rates on the ellagitannin content, antioxidant capacity, and total phenolic content of varietal pomegranate juices. Widely distinct juices made from Mollar de Elche and Wonderful cultivars were used for the elaboration of blended juices. They were stored for 70 days at both room and refrigeration temperatures. This study revealed a significant cultivar effect on the stability of main pomegranate ellagitannins (punicalagins, punicalins, punicalagin-like compound, and ellagic acid derivatives) and on the antioxidant capacity measured by the ABTS+ and DPPH* in vitro assays. Blended juices enhanced and/or retained the initial ellagitannin content and antioxidant capacity of pure juices during storage. Thus, blending varietal juices could be suggested as a promising alternative to the development of fresh juices with a high, stable phytochemical load.

Maqui berry vs Sloe berry--liquor-based beverage for new development.

"Pacharin" is an aniseed liquor-based beverage made with sloe berry (Prunus spinosa L.) that has been produced in northern Spain. On the other hand, maqui berry (Aristotelia chilensis) is a common edible berry from Chile, and currently under study because of its multiple beneficial effects on health. The aim of this work was to design a new aniseed liquor-based beverage with maqui berry, as an industrial alternative to a traditional alcoholic product with bioactive berries. The characterization of its composition, compared with the traditional "Pacharin", and its evolution during maceration (6 and 12 months) showed that the new maqui liquor had significantly-higher anthocyanin retention over time. More studies on the organoleptic properties and bioactivity are underway.

Antinociceptive and anti-inflammatory activities of a pomegranate (Punica granatum L.) extract rich in ellagitannins.

Pomegranate (Punica granatum L.) has been used for centuries for the treatment of inflammatory diseases. However, there is a lack of comprehensive information focused on the properties of a certain pomegranate (poly)phenolic profile to cure pain and gastric injury induced by anti-inflammatory drugs. This study investigated the systemic effects of different doses of a HPLC-characterized pomegranate extract on the formalin-induced nociceptive behavior in mice. The effect of the extract against gastric injury caused by non-steroidal anti-inflammatory drugs and ethanol was also assessed. Pomegranate reduced nociception in both phases of the formalin test, suggesting central and peripheral activities to inhibit nociception. Indomethacin-induced gastric injury was not produced in the presence of pomegranate, which also protected against ethanol-induced gastric lesions. The present results reinforce the benefits of pomegranate (poly)phenolics in the treatment of pain as well as their anti-inflammatory properties.

The intake of broccoli sprouts modulates the inflammatory and vascular prostanoids but not the oxidative stress-related isoprostanes in healthy humans.

Current evidence supports the positive association between the consumption of plant foods and health. In this work, we assessed the effect of consuming a half-serving (30 g) or one serving (60 g) of broccoli sprouts on the urinary concentrations of biomarkers of oxidative stress (isoprostanes) and inflammation (prostaglandins and thromboxanes). Twenty-four volunteers participated in the project. A quantitative determination of sulforaphane and its mercapturic derivatives, eicosanoids, and total vitamin C in urine was performed. The intake of broccoli sprouts produced an increase in the urinary concentrations of sulforaphane metabolites and vitamin C. Among the 13 eicosanoids analyzed, tetranor-PGEM and 11ß-PGF2α as well as 11-dehydro-TXB2 showed a significant decrease in their urinary concentrations after the ingestion of broccoli sprouts. Therefore, the consumption of broccoli sprouts modulated the excretion of biomarkers linked to inflammation and vascular reactions without exerting a significant influence on the oxidation of phospholipids in vivo.