Monitoring antioxidants by coulometry: Quantitative assessment of the strikingly high antioxidant capacity of bergamot (Citrus bergamia R.) by-products.

A recently optimized rapid, cheap, and accurate coulometric method has been exploited to determine the antioxidant capacity of bergamot (Citrus bergamia Risso) by-products, including first (FPJ) and second press juices (SPJ), in comparison to analogous products from several citrus species. Extracts from the entire edible part (i.e., juice and pulp) and de-oiled peel of bergamot were also assayed. The Coulometrically Determined Antioxidant Capacity (CDAC) data, expressed as moles of electrons per mass of sample, were evaluated with other parameters such as total phenolic compounds, ascorbic acid, total carotenoids, and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical inhibition. The CDAC of bergamot FPJ (39 mmol e- kg-1) was comparable with other citrus juices (20-65 mmol e- kg-1 range), whereas the CDAC of bergamot SPJ (816 mmol e- kg-1) was strikingly higher than the counterparts from other citrus fruits. This value approached that of bergamot peel extracts (822 mmol e- kg-1). Bergamot peel and SPJ also exhibited the highest DPPH inhibition. The CDAC values were associated with the HPLC-determined content of flavonoids, namely neoeriocitrin, naringin, and neohesperidin, which were 4-10-fold more concentrated in bergamot SPJ and peel than in SPJ from other citrus species. These findings contribute to point at bergamot by-products as rich sources of antioxidant compounds on a quantitative basis, highlighting their enormous potential for pharmaceutical, nutraceutical and food applications.

Antiproliferative and antioxidant effect of polar hemp extracts (Cannabis sativa L., Fedora cv.) in human colorectal cell lines.

Total phenolic content and antioxidant activity of polar extracts of edible resources from Fedora hemp cultivar (Cannabis sativa L.), namely seed, flour and oil, were evaluated. The main components in the polar extracts were identified using HPLC-DAD and HPLC-ESI-MS/MS. As expected, the molecular profile of components from seeds and flour was strictly similar, dominated by N-trans-caffeoyltyramine. The profile of oil polar extracts contained hydroxycinnamic acid derivatives and cannabinoids at lower extent. While the extracts from hemp seed and flour did not interfere with growth of Caco-2 and HT-29 cell, the one from oil (150 µg/mL) significantly reduced cell viability after 24 h of treatment. This effect was associated with the activation of apoptotic cell death and was independent from the antioxidant capacity of the oil polar extract. Notably, HT-29 cells differentiated with sodium butyrate were not sensitive to the cytotoxic effect of the oil extract.

Tolerogenic Effect Elicited by Protein Fraction Derived From Different Formulas for Dietary Treatment of Cow's Milk Allergy in Human Cells.

Several formulas are available for the dietary treatment of cow's milk allergy (CMA). Clinical data suggest potentially different effect on immune tolerance elicited by these formulas. We aimed to comparatively evaluate the tolerogenic effect elicited by the protein fraction of different formulas available for the dietary treatment of CMA. Five formulas were compared: extensively hydrolyzed whey formula (EHWF), extensively hydrolyzed casein formula (EHCF), hydrolyzed rice formula (HRF), soy formula (SF), and amino acid-based formula (AAF). The formulas were reconstituted in water according to the manufacturer's instructions and subjected to an in vitro infant gut simulated digestion using a sequential gastric and duodenal static model. Protein fraction was then purified and used for the experiments on non-immune and immune components of tolerance network in human enterocytes and in peripheral mononuclear blood cells (PBMCs). We assessed epithelial layer permeability and tight junction proteins (occludin and zonula occludens-1, ZO-1), mucin 5AC, IL-33, and thymic stromal lymphopoietin (TSLP) in human enterocytes. In addition, Th1/Th2 cytokine response and Tregs activation were investigated in PBMCs from IgE-mediated CMA infants. EHCF-derived protein fraction positively modulated the expression of gut barrier components (mucin 5AC, occludin and ZO-1) in human enterocytes, while SF was able to stimulate the expression of occludin only. EHWF and HRF protein fractions elicited a significant increase in TSLP production, while IL-33 release was significantly increased by HRF and SF protein fractions in human enterocytes. Only EHCF-derived protein fraction elicited an increase of the tolerogenic cytokines production (IL-10, IFN-γ) and of activated CD4+FoxP3+ Treg number, through NFAT, AP1, and Nf-Kb1 pathway. The effect paralleled with an up-regulation of FoxP3 demethylation rate. Protein fraction from all the study formulas was unable to induce Th2 cytokines production. The results suggest a different regulatory action on tolerogenic mechanisms elicited by protein fraction from different formulas commonly used for CMA management. EHCF-derived protein fraction was able to elicit tolerogenic effect through at least in part an epigenetic modulation of FoxP3 gene. These results could explain the different clinical effects observed on immune tolerance acquisition in CMA patients and on allergy prevention in children at risk for atopy observed using EHCF.

Comparative analysis of protein composition and digestibility of Ceratonia siliqua L. and Prosopis spp. seed germ flour.

By virtue of exclusive nutrient composition and nutritional properties, seed germ flours from both European carob (Ceratonia siliqua L.) and South American algarrobo (Prosopis alba and Prosopis nigra) or vinal (Prosopis ruscifolia) have potential as a high nutritional value and health-promoting ingredient for food formulations. In order to define their compositional and functional properties, we investigated the germ protein content of carob compared to the P. alba, P. nigra and P. ruscifolia counterparts, applying proteomics and complementary methods. The mono- and two-dimensional electrophoretic profiles of Prosopis spp. were very similar among one another, while C. siliqua exhibited significant differences. Attenuated total reflectance-Fourier transform infrared (ATR-FTIR) spectroscopy highlighted a dominant ß-sheet structural conformation for C. siliqua, suggesting that carob germ flour might more suited than Prosopis germ flour for baking and food technological applications. In contrast, Prosopis spp. contained a more adequate nutritional value than C. siliqua, in terms of essential amino acid complement. Both carob and algarrobo germ flour samples were highly digestible, as demonstrated by in vitro simulated gastrointestinal digestion, releasing high amounts of free amino acids and only minor proportions of low molecular weight peptides.

Comparative Study of Chemical, Biochemical Characteristic and ATR-FTIR Analysis of Seeds, Oil and Flour of the Edible Fedora Cultivar Hemp (Cannabis sativa L.).

A series of chemical and biochemical parameters of edible hemp resources (seeds, oil, and flour) from the monoecious EU registered hemp genotype Fedora, was determined, including fatty acid profile, phytosterol composition, total phenolics, antioxidant activity, macro- and micro-elements. The fatty acid ω-3/ω-6 approached the nutritionally optimal 3/1 ratio. ß-sitosterol and other phytosterols sterols dominated the unsaponifiable fraction. Hemp seeds, flour, and oil contained 767 ± 41, 744 ± 29, and 21 ± 5 mg GAE kg-1 total polyphenols, respectively. The antioxidant potential of Fedora flour and seeds, evaluated through the DPPH (2,2-Diphenyl-1-picrylhydrazyl) assay, was higher than that of oil. K and Mg were the most abundant macro-elements, particularly in flour, while the concentration of trace elements was Fe > Cu > Ni > Mn. The presence of an array of bioactive compound candidate Fedora products as health-promoting food matrices. The ATR-FTIR spectra of hemp-derived products indicated the proximate composition of macro-nutrients.

Oxidative Stability of Pomegranate (Punica granatum L.) Seed Oil to Simulated Gastric Conditions and Thermal Stress.

The fatty acid composition of pomegranate (Punica granatum L.) seed oil (PSO) is dominated by punicic acid, a conjugated linolenic acid (18:3ω-5). As a free fatty acid, punicic acid is rapidly oxidized in air and extensively isomerizes upon acid-catalyzed methylation at 90 °C. In contrast, triacylglycerol-bound punicic acid in PSO was unchanged by simulated gastric conditions and was degraded by 5-7% by severe heating (up to 170 °C for 4 h), as herein assessed by gas chromatography, attenuated total reflectance-Fourier transform infrared spectroscopy, 1H and 13C NMR, and high-resolution electrospray ionization mass spectrometry. Total polar compounds of PSO were slightly affected by thermal stress, accounting for 5.71, 6.35, and 9.53% (w/w) in the unheated, heated at mild temperature (50 °C, 2 h), and heated at frying temperature (170 °C, 4 h) PSO, respectively. These findings support from a structural standpoint the potential use of PSO as a health-promoting edible oil.

Potential anti-inflammatory effects of the hydrophilic fraction of pomegranate (Punica granatum L.) seed oil on breast cancer cell lines.

In this work, we characterized conjugated linolenic acids (e.g., punicic acid) as the major components of the hydrophilic fraction (80% aqueous methanol extract) from pomegranate (Punica granatum L.) seed oil (PSO) and evaluated their anti-inflammatory potential on some human colon (HT29 and HCT116), liver (HepG2 and Huh7), breast (MCF-7 and MDA-MB-231) and prostate (DU145) cancer lines. Our results demonstrated that punicic acid and its congeners induce a significant decrease of cell viability for two breast cell lines with a related increase of the cell cycle G0/G1 phase respect to untreated cells. Moreover, the evaluation of a great panel of cytokines expressed by MCF-7 and MDA-MB-231 cells showed that the levels of VEGF and nine pro-inflammatory cytokines (IL-2, IL-6, IL-12, IL-17, IP-10, MIP-1α, MIP-1ß, MCP-1 and TNF-α) decreased in a dose dependent way with increasing amounts of the hydrophilic extracts of PSO, supporting the evidence of an anti-inflammatory effect. Taken together, the data herein suggest a potential synergistic cytotoxic, anti-inflammatory and anti-oxidant role of the polar compounds from PSO.

Peptides from water buffalo cheese whey induced senescence cell death via ceramide secretion in human colon adenocarcinoma cell line.

SCOPE: Milk proteins are a source of bioactive peptides. Recent studies have indicated that protein-derived peptides released in buffalo cheese acid whey exert a cytomodulatory effect in human epithelial colon cancer (CaCo2) cells. The aim of the present study was to explain the molecular mechanism involved in the response of CaCo2 cells to oxidative stress in the presence of peptide fractions of buffalo cheese whey, purified and characterized by mass spectrometry. METHODS AND RESULTS: We demonstrated that treatment of CaCo2 treated with H2O2 (H-CaCo2) cells with a partially purified peptide sub-fraction (f3) from buffalo cheese acid whey induced a reduction of mitochondrial superoxide anion with subsequent decrease in heat shock protein 70 and 90 expression. Moreover, we observed a 5-fold decrease in cyclin A expression and cell cycle arrest in G1/G0 phases. These responses were associated with increased activity of alkaline phosphatase and beta-galactosidase, markers of differentiation and senescence respectively. CONCLUSIONS: The structural characterization of the active peptide fraction and the elucidation of the effects induced by its treatment on H-CaCo2 cells in vitro demonstrated an activity of this peptide sub-fraction in the modulation of cell cycle, thus suggesting potential application for the development of nutraceuticals as well as health-promoting functional foods.

Maldi-tof mass spectrometry profiling of polar and nonpolar fractions in heated vegetable oils.

Triacylglycerol oxidation of thermally stressed (6 h at 180 degrees C, simulating deep-frying conditions) edible vegetable oil (sunflower and olive) was studied using matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS). Chromatographic separation of the nonpolar and polar components from the heated oil performed on silica gel prior to MS analysis significantly enhanced the detection of oxidized components. The spectra contained signals that were assigned to triacylglycerols (TAG), diacylglycerols (DAG), triacylglycerol oxidative dimers, oxidized TAG, and TAG fragments arising from the homolytic beta-scission of linoleyl, peroxy, and alkoxy radicals. Enrichment of the polar compounds prevented mass spectrometric ion suppression, thus allowing the detection of minor species originating from thermal oxidation. In addition, this allowed the monitoring of polar compounds in vegetable oils undergoing mild thermal treatment. As such, chromatographic separation coupled with MALDI-TOF MS analysis provided a rapid, sensitive, and specific tool to assess the thermal oxidation of vegetable oils.