Antiviral Properties of Flavonoids and Delivery Strategies.

This review summarizes the latest advancements in phytochemicals as functional antiviral agents. We focused on flavonoids, like apigenin, vitexin, quercetin, rutin and naringenin, which have shown a wide range of biological effects including antiviral activities. The molecular mechanisms of their antiviral effects mainly consist in the inhibition of viral neuraminidase, proteases and DNA/RNA polymerases, as well as in the modification of various viral proteins. Mixtures of different flavonoids or combination of flavonoids with antiviral synthetic drugs provide an enhancement of their antiviral effects. Recent strategies in drug delivery significantly contribute to overcoming the low bioavailability of flavonoids. Frequent viral infections worldwide have led to the need for new effective antiviral agents, which can be identified among the various phytochemicals. In this light, screening the antiviral activities of a cocktail of flavonoids would be advantageous in order to prevent viral infections and improve current antiviral therapies.

Morphological analysis of the seeds of three pseudocereals by using light microscopy and ESEM-EDS.

The seed morphology of three Pseudocereal Grains (PSCg), i.e. quinoa (Chenopodium quinoa Willd, Chenopodiaceae), buckwheat (Fagopyrum esculentum Moench, Polygonaceae) and amaranth (Amaranthus caudatus L., Amaranthaceae) was studied by light microscopy (LM) and Environmental Scanning Electron Microscopy coupled with Energy Dispersive Spectroscopy (ESEM-EDS). LM was used with visible light to evaluate either unstained sections or sections stained with Azan mixture and with fluorescent light. The aim of the study was to compare the architecture of the three seeds in order to connect their morphology with nutrient localization. The Azan staining allowed for the visualization of the seed coat, the embryo - with its shoot apical meristem - and the radicle cell layers, whereas the use of fluorescent microscopy identified the cells rich in phenolic compounds. Finally, the ESEM-EDS analysis revealed that the seed coat of the quinoa was thinner than that of amaranth or buckwheat. In all PSCg, starch granules appeared to be located in large polygonal cells, surrounded by a thin cell wall. Several globoids of proteins were observed in the embryo cells. In the radicle section, the vascular bundles of the procambium were evident, while Amaranth only showed a consistent layer of calcium crystals, located between the embryo and the perysperm. The morphological differences of the three PSCg were discussed in the context of their structural resistance to processing technologies which impact on nutritional value of derived foods.

C-Glycosyl Flavonoids from Beta vulgaris Cicla and Betalains from Beta vulgaris rubra: Antioxidant, Anticancer and Antiinflammatory Activities-A Review.

The green beet (Beta vulgaris var. cicla L.) and red beetroot (B. vulgaris var. rubra L.) contain phytochemicals that have beneficial effects on human health. Specifically, the green beet contains apigenin, vitexin, vitexin-2-O-xyloside and vitexin-2-O-rhamnoside, while the red beetroot is a source of betaxanthins and betacyanins. These phytochemicals show considerable antioxidant activity, as well as antiinflammatory and antiproliferative activities. Vitexin-2-O-xyloside, in combination with betaxanthins and betacyanins, exerts antiproliferative activity in breast, liver, colon and bladder cancer cell lines, through the induction of both intrinsic and extrinsic apoptotic pathways. A significant body of evidence also points to the role of these phytochemicals in the downregulation of the pro-survival genes, baculoviral inhibitor of apoptosis repeat-containing 5 and catenin beta-1, as well as the genes controlling angiogenesis, hypoxia inducible factor 1A and vascular endothelial growth factor A. The multi-target action of these phytochemicals enhances their anticancer activity. Vitexin-2-O-xyloside, betaxanthins and betacyanins can be used in combination with conventional anticancer drugs to reduce their toxicity and overcome the multidrug resistance of cancer cells. In this review, we describe the molecular mechanisms that enable these dietary phytochemicals to block the proliferation of tumor cells and inhibit their pro-survival pathways. Copyright © 2017 John Wiley & Sons, Ltd.

Comparison of reduced sugar high quality chocolates sweetened with stevioside and crude stevia 'green' extract.

BACKGROUND: The demand for zero and reduced-sugar food products containing cocoa is expanding continuously. The present study was designed to evaluate the feasibility of producing high-quality chocolate sweetened with a crude extract of Stevia rebaudiana (Bertoni) prepared by a green microwave-assisted water-steam extraction procedure. Seven approximately isosweet chocolate formulations were developed, mixing cocoa paste, sucrose, commercial stevioside, crude green extract and maltitol in different proportions. All samples were analyzed for the determination of polyphenol and flavonoid content, antioxidant activity, and sensory acceptability. RESULTS: The use of a crude stevia extract allowed low-sugar, high-quality chocolates to be obtained that were also acceptable by consumers and had a significant increased antioxidant activity. Moreover, consumers' segmentation revealed a cluster of consumers showing the same overall liking for the sample with 50% sucrose replaced by the stevia crude extract as that obtained with the commercial stevioside and the control sample (without sucrose replacement). CONCLUSION: The results provide information that can contribute to promoting the development of sweet food products, with advantages in terms of an improved nutritional value (reduced sugar content and increased antioxidant activity) and a reduced impact of the production process on the environment. © 2016 Society of Chemical Industry.

Phytochemicals as Innovative Therapeutic Tools against Cancer Stem Cells.

The theory that several carcinogenetic processes are initiated and sustained by cancer stem cells (CSCs) has been validated, and specific methods to identify the CSCs in the entire population of cancer cells have also proven to be effective. This review aims to provide an overview of recently acquired scientific knowledge regarding phytochemicals and herbal extracts, which have been shown to be able to target and kill CSCs. Many genes and proteins that sustain the CSCs' self-renewal capacity and drug resistance have been described and applications of phytochemicals able to interfere with these signaling systems have been shown to be operatively efficient both in vitro and in vivo. Identification of specific surface antigens, mammosphere formation assays, serial colony-forming unit assays, xenograft transplantation and label-retention assays coupled with Aldehyde dehydrogenase 1 (ALDH1) activity evaluation are the most frequently used techniques for measuring phytochemical efficiency in killing CSCs. Moreover, it has been demonstrated that EGCG, curcumin, piperine, sulforaphane, ß-carotene, genistein and the whole extract of some plants are able to kill CSCs. Most of these phytochemicals act by interfering with the canonical Wnt (ß-catenin/T cell factor-lymphoid enhancer factor (TCF-LEF)) pathway implicated in the pathogenesis of several cancers. Therefore, the use of phytochemicals may be a true therapeutic strategy for eradicating cancer through the elimination of CSCs.

Enhancement of flavonoid ability to cross the blood-brain barrier of rats by co-administration with α-tocopherol.

Vitamin E and polyphenols could exhibit a therapeutic role in the treatment of oxidative stress-induced neurodegenerative diseases. Therefore, their ability to cross the blood-brain barrier (BBB) represents an important issue to be explored by different diet combinations. In this study, we have evaluated the ability of α-tocopherol to support epigallocatechin-3-gallate (EGCG), quercetin and rutin to cross the BBB, following oral administration. Eighteen rats were fed a standard diet (C), a diet supplemented with α-tocopherol (A), with a mixture of EGCG, quercetin and rutin (P); or with a mixture of α-tocopherol and the three flavonoids (AP). Flavonoids and their conjugated derivatives were assayed in brain and plasma by HPLC-MS, whereas α-tocopherol was detected by RP-HPLC. The oxidative damage, due to the potential pro-oxidant activity of flavonoids, was evaluated by the presence of 8-hydroxy-2'-deoxyguanosine (8-OHdG) in hippocampal Cornus Ammonis, one of the most vulnerable sites in the brain. Our results indicate that α-tocopherol is able to promote quercetin transport across the BBB. The mixture of rutin and quercetin seems to favour the accumulation of quercetin and/or its conjugated derivatives in the brain. In contrast, α-tocopherol does not affect EGCG transport across the BBB. The densitometric analysis of 8-OHdG immunoreactivity does not reveal any difference of oxidative damage among the experimental groups. Our results suggest that α-tocopherol may promote quercetin transport across the BBB, leading to a significant increase of α-tocopherol and quercetin concentration in the brain.

Caecal absorption of vitexin-2-O-xyloside and its aglycone apigenin, in the rat.

The in vivo bioavailability of the flavone-C-glycosides has been little studied compared to their O-glycoside analogues, which are both more common in nature and considered more easily hydrolyzed than C-glycosides, by both enterocytes and gut microbiota. In this study, we used vitexin-2-O-xyloside (VOX), an apigenin-8-C-glucoside-2-O-xyloside, purified from seeds of Swiss chard (Beta vulgaris cicla), to investigate VOX absorption into portal blood compared to its aglycone, apigenin. We used a rat model in which we ligated the ileo- and colo-caecal junctions, then administered apigenin or VOX directly into the caecum. Blood samples were drawn from the portal vein at timed intervals over 40 min. The kinetic profile of appearance in portal blood of the compounds and their metabolites was evaluated by HPLC-ESI-MS. Apigenin was found in portal blood both as the aglycone and as an apigenin-glucuronide derivative. The VOX was found unchanged and as a reduced monoglycoside, which underwent glucuronidation. By collecting the bile, we confirmed that the liver received unchanged VOX, which was returned to the gut by enterohepatic recirculation for reabsorption from the ileum. The amount of apigenin and VOX remaining in the caecum accounted for ∼15% and ∼26%, respectively. These data show for the first time that the C-glycoside VOX is absorbed unchanged and undergoes enterohepatic recirculation in addition to hydrolysis to the monoglycoside, reduction and conjugation to form a bioavailable glucuronide.

Nutritional and functional potential of Beta vulgaris cicla and rubra.

Swiss chard (Beta vulgaris cicla, BVc) and beetroot (Beta vulgaris rubra, BVr) are vegetables of the Chenopodiaceae family, widely consumed in traditional western cooking. These vegetables represent a highly renewable and cheap source of nutrients. They can be cultivated in soils with scarce organic material and little light and water. BVc and BVr have a long history of use in folk medicine. Modern pharmacology shows that BVc extracts possess antihypertensive and hypoglycaemic activity as well as excellent antioxidant activity. BVc contains apigenin flavonoids, namely vitexin, vitexin-2-O-rhamnoside and vitexin-2-O-xyloside, which show antiproliferative activity on cancer cell lines. BVr contains secondary metabolites, called betalains, which are used as natural dyes in food industry and show anticancer activity. In this light, BVc and BVr can be considered functional foods. Moreover, the promising results of their phytochemicals in health protection suggest the opportunity to take advantage of the large availability of this crop for purification of chemopreventive molecules to be used in functional foods and nutraceutical products.

Comparison of bioactive phytochemical content and release of isothiocyanates in selected brassica sprouts.

The consumption of brassica sprouts as raw vegetables provides a fair amount of glucosinolates (GLs) and active plant myrosinase, which enables the breakdown of GLs into health-promoting isothiocyanates (ITCs). This study reports the determination of the main constituents related to human health found in edible sprouts of two Brassica oleracea varieties, broccoli and Tuscan black kale, and two Raphanus sativus varieties, Daikon and Sango. Radish sprouts exhibited the highest ability to produce ITCs, with Daikon showing the greatest level of conversion of GLs into bioactive ITCs (96.5%), followed by Sango (90.0%). Tuscan black kale gave a value of 68.5%, whereas broccoli displayed the lowest with 18.7%. ITCs were not the exclusive GL breakdown products in the two B. oleracea varieties, since nitriles were also produced, thus accounting for the lower conversion observed. Measuring the release of plant ITCs is a valuable tool in predicting the potential level of exposure to these bioactive compounds after the consumption of raw brassica sprouts.

Total extract of Beta vulgaris var. cicla seeds versus its purified phenolic components: antioxidant activities and antiproliferative effects against colon cancer cells.

INTRODUCTION: Beta vulgaris var. cicla (BV) leaves contain chemopreventive compounds that have been investigated for new drug discovery. These compounds belong to the family of the apigenin-glycosides. Since the leaves are seasonal products containing high percentages of water, they are easily degradable during storage in fresh conditions. To be stored they require a drying process, consuming time and a large amount of energy. The extraction of apigenin-glycosides may also be conveniently performed from BV seeds, which represent a stable and year-long available biomass. OBJECTIVES: The present report was undertaken to find a strategy of purification of bioactive flavonoids from BV seeds and test their ability to inhibit proliferation both on human colon cancer (RKO) cells and normal human fibroblasts (HF). MATERIALS AND METHODS: The ethyl-acetate extract of BV seeds was fractionated on a Sephadex LH 20 column. A fraction of this extract, labeled as P4, exploited a marked antiproliferative activity on RKO cells. The components of P4 were purified on an RP18 column chromatography and identified by HPLC-ESI-MS as 2,4,5-trihydroxybenzaldehyde, 2,5-dihydroxybenzaldehyde, vanillic acid, xylosylvitexin, glucopyranosyl-glucopyrasyl-rhamnetin and glucopyranosyl-xylosyl-rhamnetin. All of them were tested for cytostatic and cytotoxic activity on RKO and HF cells. RESULTS: Xylosylvitexin exhibited the strongest antiproliferative activity on RKO cells, together with an enhancement of the apoptosis, an increase of cells in the G1 phase and a reduction of cells in the S phase; on the contrary, the proliferation of HF was significantly stimulated. CONCLUSION: Xylosylvitexin is the main and more efficient chemopreventive compound in BV seeds, but the natural cocktail of molecules, represented by P4 fraction, showed a better compromise between the antiproliferative activity on RKO cells and the enhancement of HF proliferation.

Improvement in botanical standardization of commercial freeze-dried herbal extracts by using the combination of antioxidant capacity and constituent marker concentrations.

Botanical extracts are standardized to > or = 1 marker compounds (MCs). This standardization provides a certain level of quality control, but not complete quality assurance. Thus, industries are looking for other satisfactory systems to improve standardization. This study focuses on the standardization of herbal medicines by combining 2 parameters: the concentration of the MC and antioxidant capacity. Antioxidant capacity was determined with the oxygen radical absorbance capacity (ORAC) method and the concentrations of the MCs, by high-performance liquid chromatography. Total phenols were also determined by the Folin-Ciocolteau method. The ORAC values, expressed as micromol Trolox equivalents/100 g (ORAC %), of 12 commercial herbal extracts were related to the ORAC values of the respective pure MCs at the concentrations at which the MCs occur in products (ORAC-MC %). The ORAC % values of 11 extracts were higher than those of the respective MCs and the ratios ORAC-MC %/ORAC % ranged from 0.007 to 0.7, whereas in the case of Olea europaea leaves, the same ratio was 1.36. The ORAC parameters and their ratios, as well as the linear relationship between ORAC-MC % and ORAC %, are described and discussed as tools for improving the standardization of herbal products and detecting modifications due to herb processing and storage.

Polyphenols and antioxidant capacity of vegetables under fresh and frozen conditions.

The phenolic and oxygen radical absorbance capacity (ORAC) values have been measured in six fresh and frozen vegetables (beet green, spinach, broccoli, carrot, onion, and celery) from the same cultivar by analyzing the whole juice (WJ) and the acetonic extract of the squeezed pulp. To exploit the effect of the acid environment on the stability and recovery of the phenolics, perchloric acid (PCA) was added directly to WJ and to the pulp before the extraction with acetone. In both fresh and frozen vegetables, PCA markedly increased the recovery of phenolics extracted from the pulp, but PCA had no effect on the WJ. Four of six frozen vegetables showed lower phenolic and ORAC values than the fresh vegetables, whereas in the other two cases, values were significantly higher compared to fresh samples. Among the fresh vegetables, beet green showed the highest ORAC and phenolic values; however, when measured in two different cultivars of beet green, the ORAC value showed as much as 4.5-fold variation, whereas total phenolics and flavonoids showed 1.2- and 3.5-fold variations, respectively. The results show that total phenolics and ORAC, compared in fresh and frozen vegetables, represent an index of the mildness of blanching in the industry of frozen vegetables and provide a measure of the gap in antioxidants in the diet of people who consume frozen instead of fresh vegetables. The plant genotype is an important source of variability in the ORAC value, which can be conveniently used to increase the intake of antioxidants from vegetables.

Tocopherols enhance neurogenesis in dentate gyrus of adult rats.

In the dentate gyrus of the mammalian hippocampus, neurogenesis carries on throughout postnatal life. The aim of this work was to identify an exogenous control factor of adult neurogenesis. Neurogenesis in the adult dentate gyrus was previously found to be enhanced in vitamin E-deficient rats. The effects of alpha- or beta-tocopherol supplementation on neurogenesis in the adult dentate gyrus were investigated by 5-bromo-2'-deoxyuridine labeling. Tocopherol was found to increase the survival of newborn cells and the total number of granule cells in the adult rat dentate gyrus. Newborn cells were phenotypically characterized by expression of the immature neuron marker TOAD-64 (turned on after division-64). Therefore tocopherol in high doses possibly increases the number of granule cells in the dentate gyrus by saving newborn cells from death.