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Twenty oenotannins from different botanical origins were studied in model wine solution (1 g/L, 12% ethanol, pH 3.5). An original device was created for measuring Oxidation-Reduction potential ( ORp ) of the solutions at 20 °C in strict anoxic condition by the electrochemical method of the platinum electrode zero-current potential. Reactivity against proteins and antioxidant properties were related to the chemical structure and, consequently, to the botanical origin of the oenotannins. The highest turbidity after BSA addition (ΔNTU > 1000) values were measured for the gallic hydrolysable tannins. The ORp versus standard hydrogen electrode ranged from 420 to 260 mV. The ellagitannins had the highest antioxidant power ( AP %), followed by condensed tannins and gallotannins, highlighting a correlation with the phenolic profile. Based on these findings, two formulations were prepared as a blend of some of the tested oenotannins, with the ability to increase (MIX1) and decrease (MIX2) the ORp of the model wine.

The effects of the phenolic compounds of extra virgin olive oil (EVOO) on AGS cells have never been studied so far, which is the aim of this study. The profiles of the main phenolic components in EVOOs, mainly secoiridoid compounds derived from the transformation of oleuropein during the olive milling process, were evaluated and compared. Oils of different origins were evaluated aiming at verifying whether chemical differences in the phenolic composition of the dry extracts played a role in the metabolism and in maintaining the cellular redox state of AGS cells. The following key enzymes of some metabolic pathways were studied: lactate dehydrogenase, enolase, pyruvate kinase, glucose 6-phosphate dehydrogenase, citrate synthase, 3-Hydroxyacyl-CoA dehydrogenase and hexokinase. As confirmed through PCA analysis, pretreatments with the dry extracts of EVOOs at different concentrations appeared to be able to counteract the enzymatic activity alterations due to oxidative stress induced by H 2 O 2 1 mM and 2 mM. The studied phytocomplexes showed the ability to protect AGS cells from oxidative damage and the secoiridoid derivatives from both oleuropein and ligstroside contributed to the observed effects. The results suggested that EVOOs with medium to high concentrations of phenols can exert this protection.

Millet is the sixth-highest yielding grain in the world and a staple crop for millions of people. Fermentation was applied in this study to improve the nutritional properties of pearl millet. Three microorganism combinations were tested: Saccharomyces boulardii (FPM1), Saccharomyces cerevisiae plus Campanilactobacillus paralimentarius (FPM2) and Hanseniaspora uvarum plus Fructilactobacillus sanfranciscensis (FPM3). All the fermentation processes led to an increase in minerals. An increase was observed for calcium: 254 ppm in FPM1, 282 ppm in FPM2 and 156 ppm in the unfermented sample. Iron increased in FPM2 and FPM3 (approx. 100 ppm) with respect the unfermented sample (71 ppm). FPM2 and FPM3 resulted in richer total phenols (up to 2.74 mg/g) compared to the unfermented sample (2.24 mg/g). Depending on the microorganisms, it was possible to obtain different oligopeptides with a mass cut off ≤10 kDalton that was not detected in the unfermented sample. FPM2 showed the highest resistant starch content (9.83 g/100 g) and a prebiotic activity on Bifidobacterium breve B632, showing a significant growth at 48 h and 72 h compared to glucose ( p < 0.05). Millet fermented with Saccharomyces cerevisiae plus Campanilactobacillus paralimentarius can be proposed as a new food with improved nutritional properties to increase the quality of the diet of people who already use millet as a staple food.

For improving the management of the production chain of PGI Mantua pears (which comprises many varieties, including Abate Fetel), applying the cardinal principles of circular economy and sustainability, the fruits with diseases or defects were recovered for producing dried rounds of pears from the Abate Fetel cultivar, a new product with high nutritional value that extends the remaining life. This process led to the production of secondary and residual by-products, which are mainly composed of the highest and lowest part of the fruits, comprising seeds, pulps, peels and petioles. Hence, this study was focused on the valorization of these secondary by-products of Abate Fetel pears through the production of pear extracts using traditional and "green" extraction methods that involve the use of supercritical CO 2 fluid extraction. The produced extracts, together with a reference solvent-derived extract, were analyzed by HPLC-ESI-MS, and in parallel, their direct and cellular antioxidant activity were assessed. Evidence has indicated that all the tested extracts reduced the H 2 O 2 -induced reactive oxygen species (ROS), lipid peroxidation and nitric oxide (NO) levels, respectively, in human intestinal Caco-2 cells. Hence, this study clearly suggests that extracts obtained from Mantuan PGI pear by-products may be used as valuable sources of bioactive upcycled phytocomplex for the development of dietary supplements and/or functional foods.

Agri-food industry wastes and by-products include highly valuable components that can upgraded, providing low-cost bioactives or used as an alternative protein source. In this context, by-products from olive production and olive oil extraction process, i.e., seeds, can be fostered. In particular, this work was aimed at extracting and characterizing proteins for Olea europaea L. seeds and at producing two protein hydrolysates using alcalase and papain, respectively. Peptidomic analysis were performed, allowing to determine both medium- and short-sized peptides and to identify their potential biological activities. Moreover, an extensive characterization of the antioxidant properties of Olea europaea L. seed hydrolysates was carried out both in vitro by 2,2-diphenyl-1-picrylhydrazyl (DPPH), by ferric reducing antioxidant power (FRAP), and by 2,2'-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) assays, respectively, and at cellular level by measuring the ability of these hydrolysates to significant reduce the H 2 O 2 -induced reactive oxygen species (ROS) and lipid peroxidation levels in human intestinal Caco-2 cells. The results of the both hydrolysates showed significant antioxidant properties by reducing the free radical scavenging activities up to 65.0 ± 0.1% for the sample hydrolyzed with alcalase and up to 75.7 ± 0.4% for the papain hydrolysates tested at 5 mg/mL, respectively. Moreover, similar values were obtained by the ABTS assays, whereas the FRAP increased up to 13,025.0 ± 241.5% for the alcalase hydrolysates and up to 12,462.5 ± 311.9% for the papain hydrolysates, both tested at 1 mg/mL. According to the in vitro results, both papain and alcalase hydrolysates restore the cellular ROS levels up 130.4 ± 4.24% and 128.5 ± 3.60%, respectively, at 0.1 mg/mL and reduce the lipid peroxidation levels up to 109.2 ± 7.95% and 73.0 ± 7.64%, respectively, at 1.0 mg/mL. In addition, results underlined that the same hydrolysates reduced the activity of dipeptidyl peptidase-IV (DPP-IV) in vitro and at cellular levels up to 42.9 ± 6.5% and 38.7 ± 7.2% at 5.0 mg/mL for alcalase and papain hydrolysates, respectively. Interestingly, they stimulate the release and stability of glucagon-like peptide 1 (GLP-1) hormone through an increase of its levels up to 660.7 ± 21.9 pM and 613.4 ± 39.1 pM for alcalase and papain hydrolysates, respectively. Based on these results, olive seed hydrolysates may represent new ingredients with antioxidant and anti-diabetic properties for the development of nutraceuticals and functional foods for the prevention of metabolic syndrome onset.

Phenolic and triterpenoid compounds of the olive tree are recognized as having a key role in health promotion, thanks to their multiple protective actions in humans. To expand the source of these bioactive compounds, the phenolic and triterpenoid profiles of leaf, branch, destoned fruit, destoned pomace, shell, seed, and extra virgin olive oil from the Frantoio , Leccino , and Moraiolo olive cultivars were simultaneously characterized by HPLC-DAD-MS. Overall, 43 molecules were quantitated and expressed on the obtained dry extracts. Oleuropein was mainly concentrated in branches (82.72 g/kg), fruits (55.79 g/kg), leaves (36.71 g/kg), and shells (1.26 g/kg), verbascoside (4.88 g/kg) in pomace, and nüzhenide 11-methyl oleoside (90.91 g/kg) in seeds. Among triterpenoids, which were absent in shells, the highest amount of oleanolic acid was found in olive leaves (11.88 g/kg). HCT-116 colorectal cells were chosen to assess the cytotoxicity of the dry extract, using the phytocomplex from Frantoio, which was the richest in phenols and triterpenoids. The IC 50 was also determined for 13 pure molecules (phenols and terpenoids) detected in the extracts. The greatest inhibition on the cell's proliferation was induced by the branch dry extract (IC 50 88.25 μg/mL) and by ursolic acid (IC 50 24 μM). A dose-dependent relationship was observed for the tested extracts.

The anti-oxidative activity of plant-derived extracts is well-known and confers health-promoting effects on functional foods and food supplements. Aim of this work is to evaluate the capability of two different assays to predict the real biological antioxidant efficiency. At this purpose, extracts from five different plant-derived matrices and commercial purified phytochemicals were analyzed for their anti-oxidative properties by using well-standardized in vitro chemical method (TEAC) and an ex vivo biological assay. The biological assay, a cellular membrane system obtained from erythrocytes of healthy volunteers, is based on the capability of phytochemicals treatment to prevent membrane lipid peroxidation under oxidative stress by UV-B radiation. Plant extracts naturally rich in phenols with different structure and purified phytochemicals showed different in vitro and ex vivo antioxidant capacities. A high correlation between phenolic contents of the plant-derived extracts and their ability to prevent oxidative injuries in a biological system was found, thus underlying the relevance of this class of metabolites in preventing oxidative stress. On the other hand, a low correlation between the antioxidant capacities was shown between in vitro and ex vivo antioxidant assay. Moreover, data presented in this work show how food complex matrices are more effective in preventing oxidative damages at biological level than pure phytochemicals, even if for these latter, the antioxidant activity was generally higher than that observed for food complex matrices.

Two extra virgin olive oil (EVOO) phenolic extracts (BUO and OMN) modulate DPP-IV activity. The in vitro DPP-IV activity assay was performed at the concentrations of 1, 10, 100, 500, and 1000 μg/mL, showing a dose-dependent inhibition by 6.8 ± 1.9, 17.4 ± 6.1, 37.9 ± 2.4, 57.8 ± 2.9, and 81 ± 1.4% for BUO and by 5.4 ± 1.7, 8.9 ± 0.4, 28.4 ± 7.2, 52 ± 1.3, and 77.5 ± 3.5% for OMN. Moreover, both BUO and OMN reduced the DPP-IV activity expressed by Caco-2 cells by 2.9 ± 0.7, 44.4 ± 0.7, 61.2 ± 1.8, and 85 ± 4.2% and by 3 ± 1.9, 35 ± 9.4, 60 ± 7.2, and 82 ± 2.8%, respectively, at the same doses. The concentration of the most abundant and representative secoiridoids within both extracts was analyzed by nuclear magnetic resonance ( 1 H-NMR). Oleuropein, oleacein, oleocanthal, hydroxytyrosol, and tyrosol, tested alone, reduced the DPP-IV activity, with IC 50 of 472.3 ± 21.7, 187 ± 11.4, 354.5 ± 12.7, 741.6 ± 35.7, and 1112 ± 55.6 µM, respectively. Finally, in silico molecular docking simulations permitted the study of the binding mode of these compounds.

Extra virgin olive oil (EVOO) is well known for containing relevant amounts of healthy phenolic compounds. The European Food Safety Authority (EFSA) allowed a health claim for labelling olive oils containing a minimum amount of hydroxytyrosol (OHTyr) and its derivatives, including tyrosol (Tyr). Therefore, harmonized and standardized analytical protocols are required in support of an effective application of the health claim. Acid hydrolysis performed after extraction and before chromatographic analysis has been shown to be a feasible approach. Nevertheless, other fast, green, and easy methods could be useful for on-site screening and monitoring applications. In the present research, a natural deep eutectic solvent (NADES) composed of lactic acid and glucose was used to perform a liquid/liquid extraction on EVOO samples, followed by UV-spectrophotometric analysis. The spectral features of the extracts were related with the content of total OHTyr and Tyr, determined by the acid hydrolysis method. The second derivative of spectra allowed focusing on three single wavelengths (i.e., 299 nm, 290 nm, and 282 nm) significantly related with total OHTyr, total Tyr, and their sum, respectively. In particular, the sum of OHTyr and Tyr could be determined with a root mean square error of prediction of 29.5 mg kg -1 , while the limits of quantitation and detection were respectively 11.8 and 4.9 mg kg -1 . The proposed method, therefore, represents an easy screening tool, with the use of a green, food-derived solvent, and could be considered as an attempt to pave the way for food grade analytical chemistry.

In the framework of research aimed at promoting the nutraceutical properties of the phenolic extract (BUO) obtained from an extra virgin olive oil of the Frantoio cultivar cultivated in Tuscany (Italy), with a high total phenols content, this study provides a comprehensive characterization of its antioxidant properties, both in vitro by Trolox equivalent antioxidant capacity, oxygen radical absorbance capacity, ferric reducing antioxidant power, and 2,2-diphenyl-1-picrylhydrazyl assays, and at the cellular level in human hepatic HepG2 and human intestinal Caco-2 cells. Notably, in both cell systems, after H 2 O 2 induced oxidative stress, the BUO extract reduced reactive oxygen species, lipid peroxidation, and NO overproduction via modulation of inducible nitric oxide synthase protein levels. In parallel, the intestinal transport of the different phenolic components of the BUO phytocomplex was assayed on differentiated Caco-2 cells, a well-established model of mature enterocytes. The novelty of our study lies in having investigated the antioxidant effects of a complex pool of phenolic compounds in an extra virgin olive oil (EVOO) extract, using either in vitro assays or liver and intestinal cell models, rather than the effects of single phenols, such as hydroxytyrosol or oleuropein. Finally, the selective trans-epithelial transport of some oleuropein derivatives was observed for the first time in differentiated Caco-2 cells.

This study was aimed at demonstrating the substantial equivalence of two extra virgin olive oil samples extracted from the same batch of Coratina olives with (OMU) or without (OMN) using ultrasound technology, by performing chemical, biochemical, and cellular investigations. The volatile organic compounds compositions and phenolic profiles were very similar, showing that, while increasing the extraction yields, the innovative process does not change these features. The antioxidant and hypocholesterolemic activities of the extra virgin olive oil (EVOO) phenol extracts were also preserved, since OMU and OMN had equivalent abilities to scavenge the 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) radicals in vitro and to protect HepG2 cells from oxidative stress induced by H 2 O 2 , reducing intracellular reactive oxygen species (ROS) and lipid peroxidation levels. In addition, by inhibiting 3-hydroxy-3-methylglutarylcoenzyme a reductase, both samples modulated the low-density lipoprotein receptor (LDLR) pathway leading to increased LDLR protein levels and activity.

Pomegranate is receiving renewed commercial and scientific interest, therefore a deeper knowledge of the chemical composition of the fruits of less studied varieties is required. In this work, juices from arils and decoctions from mesocarp plus exocarp were prepared from fifteen varieties. Samples were submitted to High Performance Liquid Chromatography-Diode Array Detector-Mass Spectrometry, spectrophotometric and colorimetric CIEL*a*b* analyses. Antioxidant, antiradical and metal chelating properties, inhibitory activity against tyrosinase and α-amylase enzymes were also evaluated. All varieties presented the same main phenols; anthocyanins and ellagitannins were widely variable among varieties, with the richest anthocyanin content in the juices from the Wonderful and Soft Seed Maule varieties (approx. 660 mg/L) and the highest ellagitannin content in the peel of the Black variety (approx. 133 mg/g dry matter). A good correlation was shown between the colour hue and the delphinidin/cyanidin ratio in juices ( R 2 = 0.885). Total polysaccharide yield ranged from 3% to 12% of the peels' dry weight, with the highest content in the Black variety. Decoctions (24.44-118.50 mg KAE/g) showed better in vitro antioxidant properties and higher inhibitory capacity against tyrosinase than juices (not active-16.56 mg KAE/g); the inhibitory capacity against α-amylase was similar and quite potent for juices and decoctions. Knowledge about the chemical composition of different pomegranate varieties will allow for a more aware use of the different parts of the fruit., Competing Interests: The authors declare no conflict of interest.

Millet is underutilized in Europe, despite its advantages compared to other common cereals. In Asia and Africa, millet is mainly eaten in fermented form; its consumption has beneficial properties on human health. Three millet batches were compared in terms of free and bound phenols by High Performance Liquid Chromatography-Diode Array Detector-Mass Spectrometry (HPLC-DAD-MS). The richest one in terms of bound phenols was selected for testing via a basic (0.1 M NaOH) and an acidic (1.2 M H 2 SO 4 ) hydrolysis, in which 149.3 and 193.6 mg/100 g of phenols were recovered, respectively. The ability of fermentation, with yeast and Lactobacilli , to increase the content of phenolic compounds was verified. Five withdrawalswere performed to verify the influence of fermentation time on the total phenolic content. The greatest phenolic content was observed after 72 h. Fermentation increased the cinnamic acids and flavonoids contents by approximately 30%. Vitexin and vitexin 2″- O -rhamnoside contents were significantly higher in the fermented millet; these compounds partially inhibit the protein tyrosine phosphatase enzyme, which is overexpressed in type-2 diabetes. A molecular dynamic simulation showed the two flavonoids to be allosteric inhibitors. The phenolic extract from fermented millet demonstrated a higher level of antioxidant protection on human erythrocytes by ex vivo cellular antioxidant activity in red blood cells. In this context, functional foods based on fermented millet could represent a new trend in European markets.