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Keywords = in vitro simulated gastro-duodenal digestion

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26 pages, 5877 KiB  
Article
Citrus Pomace as a Source of Plant Complexes to Be Used in the Nutraceutical Field of Intestinal Inflammation
by Mariarosaria Ingegneri, Maria Rita Braghini, Michela Piccione, Cristiano De Stefanis, Manuela Mandrone, Ilaria Chiocchio, Ferruccio Poli, Martina Imbesi, Anna Alisi, Antonella Smeriglio and Domenico Trombetta
Antioxidants 2024, 13(7), 869; https://doi.org/10.3390/antiox13070869 - 19 Jul 2024
Viewed by 969
Abstract
This study aims to recover the main by-product of Citrus fruits processing, the raw pomace, known also as pastazzo, to produce plant complexes to be used in the treatment of inflammatory bowel disease (IBD). Food-grade extracts from orange (OE) and lemon (LE) [...] Read more.
This study aims to recover the main by-product of Citrus fruits processing, the raw pomace, known also as pastazzo, to produce plant complexes to be used in the treatment of inflammatory bowel disease (IBD). Food-grade extracts from orange (OE) and lemon (LE) pomace were obtained by ultrasound-assisted maceration. After a preliminary phytochemical and biological screening by in vitro assays, primary and secondary metabolites were characterized by proton nuclear magnetic resonance (1H-NMR) and liquid chromatography coupled to diode array detection and electrospray ionization mass spectrometry (LC-DAD-ESI-MS) analyses. The intestinal bioaccessibility and antioxidant and anti-inflammatory properties were investigated by in vitro simulated gastro-intestinal digestion followed by treatments on a lipopolysaccharide (LPS)-stimulated human colorectal adenocarcinoma cell line (Caco-2). The tight junctions-associated structural proteins (ZO-1, Claudin-1, and Occludin), transepithelial electrical resistance (TEER), reactive oxygen species (ROS)-levels, expression of some key antioxidant (CAT, NRF2 and SOD2) and inflammatory (IL-1β, IL-6, TNF-α, IL-8) genes, and pNFkB p65 nuclear translocation, were evaluated. The OE and LE digesta, which did not show any significant difference in terms of phytochemical profile, showed significant effects in protecting against the LPS-induced intestinal barrier damage, oxidative stress and inflammatory response. In conclusion, both OE and LE emerged as potential candidates for further preclinical studies on in vivo IBD models. Full article
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<p><sup>1</sup>H NMR profiling of LE (top) and OE (bottom). Full spectra (<b>A</b>) and extended spectral regions from δ 5.95 to 7.30 (<b>B</b>), from δ 3.2 to 5.5 (<b>C</b>), and from δ 3.1 to 0.7 (<b>D</b>). 1 = sucrose, 2 = <span class="html-italic">α</span>-glucose, 3 = <span class="html-italic">β</span>-glucose, 4 = citric acid, 5 = tyrosine, 6 = asparagine, 7 = fructose, 8 = malic acid, 9 = GABA, 10 = aspartic acid, 11 = succinic acid, 12 = proline, 13 = alanine.</p>
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<p>Distribution percentage of phytochemical classes identified in orange and lemon raw pomace extracts (OE and LE, respectively).</p>
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<p>Representative LC-DAD chromatograms of orange raw pomace extract (OE, panel <b>A</b>) and lemon raw pomace extract (LE, panel <b>B</b>) pre- (black) and post-gastro-duodenal digestion (orange and green chromatogram, respectively) acquired at 292 nm.</p>
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<p>OE and LE effects on Caco-2 cell viability. Cell viability evaluated by XTT assay and expressed as percentage of cell viability in Caco-2 cells untreated or treated with different concentrations of OE for 24 h (<b>A</b>) and 48 h (<b>B</b>); and in Caco-2 cells untreated or treated with different concentrations of LE for 24 h (<b>C</b>) and 48 h (<b>D</b>). Values are the mean ± SD of three independent experiments repeated at least in quintuplicate. Data were analyzed by 2-tailed Student’s <span class="html-italic">t</span> test. * <span class="html-italic">p</span> &lt; 0.05; ** <span class="html-italic">p</span> &lt; 0.01; ns: non-significant.</p>
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<p>Cell viability and proliferation in Caco-2 cells under different treatments. Cell viability evaluated by XTT assay and expressed as percentage of cell viability in Caco-2 cells untreated or treated with different concentrations of LPS for 24 h (<b>A</b>) and 48 h (<b>B</b>); and in Caco-2 cells untreated (Ctrl) or treated with LPS, LPS + 200 µg/mL OE and LPS + 200 µg/mL LE for 24 h (<b>C</b>) and 48 h (<b>D</b>). Cell proliferation monitored by using the Incucyte live cell imaging system was expressed as fold change of mean cell confluence in Caco-2 cells Ctrl, LPS, LPS + 200 µg/mL OE, and LPS + 200 µg/mL LE for 24 h (<b>E</b>) and 48 h (<b>F</b>). Values are the mean ± SD of three independent experiments repeated at least in quintuplicate. Data were analyzed by 2-tailed Student’s <span class="html-italic">t</span> test. * <span class="html-italic">p</span> &lt; 0.05; ** <span class="html-italic">p</span> &lt; 0.01; *** <span class="html-italic">p</span> &lt; 0.001; ns: non-significant.</p>
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<p>Intestinal permeability of Caco-2 cells under different treatments. (<b>A</b>) TEER values expressed as percentage of initial values of unit area resistance calculated by dividing resistance values by the effective membrane area in Caco-2 cells Ctrl, LPS, LPS + 200 µg/mL OE, and LPS + 200 µg/mL LE. Values are the mean ± SD of three independent experiments. Data were analyzed by 2-tailed Student’s <span class="html-italic">t</span> test. * <span class="html-italic">p</span> &lt; 0.05; ** <span class="html-italic">p</span> &lt; 0.01; *** <span class="html-italic">p</span> &lt; 0.001. (<b>B</b>) Representative immunofluorescence by confocal imaging of ZO-1, Claudin-1, and Occludin in Caco-2 cells Ctrl, LPS, LPS + 200 µg/mL OE, and LPS + 200 µg/mL LE. 40× magnification.</p>
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<p>Oxidative stress of Caco-2 cells under different treatments. Fold change of the relative mean fluorescence (<b>A</b>) and representative images (<b>B</b>) of CM-H2DCFDA (green) staining in Caco-2 cells Ctrl, LPS, LPS + 200 µg/mL OE, and LPS + 200 µg/mL LE. Hoechst nuclear staining (blue). 40× magnification. Relative mRNA expression of CAT (<b>C</b>), SOD2 (<b>D</b>), and NRE2L2 (<b>E</b>) genes measured by qPCR in Caco-2 cells Ctrl, LPS, LPS + 200 µg/mL OE, and LPS + 200 µg/mL LE. Values are the mean ± SD of three independent experiments. Data were analyzed by 2-tailed Student’s <span class="html-italic">t</span> test. * <span class="html-italic">p</span> &lt; 0.05; ** <span class="html-italic">p</span> &lt; 0.01; *** <span class="html-italic">p</span> &lt; 0.001; ns: non-significant.</p>
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<p>Inflammatory response of Caco-2 cells under different treatments. Relative mRNA expression of <span class="html-italic">IL-1β</span> (<b>A</b>), <span class="html-italic">IL-6</span> (<b>B</b>), <span class="html-italic">IL-8</span> (<b>C</b>), and <span class="html-italic">TNF-α</span> (<b>D</b>) genes measured by qPCR in Caco-2 cells Ctrl, LPS, LPS + 200 µg/mL OE, and LPS + 200 µg/mL LE. Representative immunofluorescence by confocal imaging (<b>E</b>) and QFIA (<b>F</b>) of pNFκB p65 (red) in Caco-2 cells Ctrl, LPS, LPS + 200 µg/mL OE, and LPS + 200 µg/mL LE. Hoechst nuclear staining (blue). 40× magnification. Values are the mean ± SD of three independent experiments. Data were analyzed by 2-tailed Student’s <span class="html-italic">t</span> test. * <span class="html-italic">p</span> &lt; 0.05; ** <span class="html-italic">p</span> &lt; 0.01; *** <span class="html-italic">p</span> &lt; 0.001; ns: non-significant.</p>
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19 pages, 4517 KiB  
Article
In Vitro Assessment of Cortisol Release Inhibition, Bioaccessibility and Bioavailability of a Chemically Characterized Scutellaria lateriflora L. Hydroethanolic Extract
by Daniele Giuseppe Buccato, Hammad Ullah, Lorenza Francesca De Lellis, Roberto Piccinocchi, Alessandra Baldi, Xiang Xiao, Carla Renata Arciola, Alessandro Di Minno and Maria Daglia
Molecules 2024, 29(3), 586; https://doi.org/10.3390/molecules29030586 - 25 Jan 2024
Viewed by 1761
Abstract
Excess cortisol release is associated with numerous health concerns, including psychiatric issues (i.e., anxiety, insomnia, and depression) and nonpsychiatric issues (i.e., osteoporosis). The aim of this study was to assess the in vitro inhibition of cortisol release, bioaccessibility, and bioavailability exerted by a [...] Read more.
Excess cortisol release is associated with numerous health concerns, including psychiatric issues (i.e., anxiety, insomnia, and depression) and nonpsychiatric issues (i.e., osteoporosis). The aim of this study was to assess the in vitro inhibition of cortisol release, bioaccessibility, and bioavailability exerted by a chemically characterized Scutellaria lateriflora L. extract (SLE). The treatment of H295R cells with SLE at increasing, noncytotoxic, concentrations (5–30 ng/mL) showed significant inhibition of cortisol release ranging from 58 to 91%. The in vitro simulated gastric, duodenal, and gastroduodenal digestions, induced statistically significant reductions (p < 0.0001) in the bioactive polyphenolic compounds that most represented SLE. Bioavailability studies on duodenal digested SLE, using Caco-2 cells grown on transwell inserts and a parallel artificial membrane permeability assay, indicated oroxylin A glucuronide and oroxylin A were the only bioactive compounds able to cross the Caco-2 cell membrane and the artificial lipid membrane, respectively. The results suggest possible applications of SLE as a food supplement ingredient against cortisol-mediated stress response and the use of gastroresistant oral dosage forms to partially prevent the degradation of SLE bioactive compounds. In vivo studies and clinical trials remain necessary to draw a conclusion on the efficacy and tolerability of this plant extract. Full article
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Figure 1
<p>PDA chromatograms of <span class="html-italic">S. lateriflora</span> extract not submitted to digestion, obtained by RP-UHPLC-PDA-ESI-MS/MS analysis. 1: apigenin derivative, 2: scutellarin, 3: baicalein 6-glucuronide, 4: naringenin 7-<span class="html-italic">O</span>-<span class="html-small-caps">d</span>-hexoside 6″ acetate, 5: oroxilyn A glucuronide, and 6: genistein.</p>
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<p>Caco-2 cell viability after treatment with the duodenal-digested <span class="html-italic">S. lateriflora</span> extract at a concentration range of 60–7.5 mg/mL and a control group (CTR). Viability values are calculated as averages from three independent assays, and the standard deviation is provided. The CTR represents cells not treated with <span class="html-italic">S. lateriflora</span> extract, set at 100% viability. Statistical significance is denoted as follows: * <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.01, **** <span class="html-italic">p</span> &lt; 0.0001, and ns: not significant.</p>
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<p>Morphology of Caco-2 cell monolayers. Fluorescent microscope image showing the top view of a Caco-2 monolayer, where the cells’ borders can be distinguished by immunocytochemical staining of the tight junction protein ZO-1 (red) and DAPI counterstaining for nuclei identification (blue).</p>
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<p>RP-UHPLC-PDA-ESI-MS/MS analysis of the solution taken from the basolater compartment, related to the absorption experiment on Caco-2 cells grown on a transwell insert treated with the duodenal digested <span class="html-italic">S. lateriflora</span> extract. (<b>A</b>,<b>B</b>) Mass chromatograms; (<b>C</b>,<b>D</b>) MS and MS/MS spectra of oroxylin A.</p>
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<p>RP-UHPLC-PDA-ESI-MS/MS analysis of the solution taken from the basolater compartment, relating to the PAMPA experiment on the artificial lipid membrane treated with duodenal digested <span class="html-italic">S. lateriflora</span> extract. (<b>A</b>,<b>B</b>) Mass chromatograms; (<b>C</b>,<b>D</b>) MS and MS/MS spectra of oroxylin A.</p>
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<p>H295R Cell viability after treatment with <span class="html-italic">S. lateriflora</span> extract in the concentration range of 60–7.5 mg/mL. CTR: cells not treated with <span class="html-italic">S. lateriflora</span> extract. Data were expressed as mean ± SD from three biological replicates. ** <span class="html-italic">p</span> &lt; 0.01, *** <span class="html-italic">p</span> &lt; 0.001, **** <span class="html-italic">p</span> &lt; 0.0001, and ns: not significant.</p>
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<p>Concentration of cortisol (ng/mL) released from H295R cells in the basal state (CRT), following the treatment with forskolin (CRT + forskolin), and following the treatment of cells with increasing concentrations (5–30 mg/mL) of <span class="html-italic">S. lateriflora</span> extract. Data were expressed as mean ± SD from three biological replicates. **** <span class="html-italic">p</span> &lt; 0.0001.</p>
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25 pages, 2182 KiB  
Article
In Vitro Digestibility and Bioaccessibility of Nutrients and Non-Nutrients Composing Extruded Brewers’ Spent Grain
by Maria Belen Gutierrez-Barrutia, Sonia Cozzano, Patricia Arcia and Maria Dolores del Castillo
Nutrients 2022, 14(17), 3480; https://doi.org/10.3390/nu14173480 - 24 Aug 2022
Cited by 7 | Viewed by 3449
Abstract
This study aimed to evaluate the effect of the extrusion process on the bioaccessibility of brewers’ spent grain (BSG) nutrients (carbohydrates and proteins) and non-nutrients (bioactive compounds). BSG and extruded BSG (EBSG) were digested in vitro simulating human oral-gastro-intestinal digestion and colonic fermentation. [...] Read more.
This study aimed to evaluate the effect of the extrusion process on the bioaccessibility of brewers’ spent grain (BSG) nutrients (carbohydrates and proteins) and non-nutrients (bioactive compounds). BSG and extruded BSG (EBSG) were digested in vitro simulating human oral-gastro-intestinal digestion and colonic fermentation. The duodenal bioaccessibility of glucose, amino acids and phenolic compounds was analyzed. The fermentability of the dietary fiber was assessed by analysis of short-chain fatty acids. Additionally, assessment of the bioaccessibility of phenolic compounds after colonic fermentation was undertaken. The antioxidant, anti-inflammatory and antidiabetic properties of the bioaccessible compounds were studied. Extrusion caused no change in the digestibility of gluten and glucose bioaccessibility (p > 0.05). Moreover, the bioaccessibility of amino acids and phenolic compounds significantly increased (p < 0.05) due to extrusion. However, higher short-chain fatty acid content was formed in colonic fermentation of BSG (p < 0.05) compared to EBSG. The latter inhibited intracellular ROS formation in IEC-6 cells and showed anti-inflammatory properties in RAW264.7 cells. With respect to antidiabetic properties, glucose absorption was lower, and the inhibition of carbohydrases higher (p < 0.05), in the presence of EBSG compared to BSG. The effects of EBSG and BSG digests on glucose transporters were not significantly different (p > 0.05). In conclusion, extrusion positively affected the nutritional value and health-promoting properties of BSG. Full article
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<p>Intracellular ROS formation under physiological conditions in IEC-6 for untreated cells (Control (+)), cells treated with DBSG, cells treated with DEBSG and an antioxidant standard (FA). Percentage of ROS formation was calculated with respect to the Control (+). Different letters within columns show significant differences (<span class="html-italic">p</span> &lt; 0.05). DBSG: brewers’ spent grain intestinal digest; DEBSG: extruded brewers’ spent grain intestinal digest; FA: ferulic acid.</p>
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<p>NO formation (µg/mL) in RAW 264.7 induced by LPS (1 µg/mL). Different letters within columns show significant differences (<span class="html-italic">p</span> &lt; 0.05). DBSG: brewers’ spent grain intestinal digest; DEBSG: extruded brewers’ spent grain intestinal digest; FA: ferulic acid.</p>
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<p>Brewers’ spent grain intestinal digests (DBSG) and extruded brewers’ spent grain intestinal digests (DEBSG) glucose absorption in IEC-6 cells over time.</p>
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<p>Glucose absorption in IEC-6 cells over time against reference inhibitors of glucose transporters (phloridzin and phloretin), ferulic acid, brewers’ spent grain intestinal digest (DBSG) and extruded brewers’ spent grain intestinal digest (DEBSG) under sodium-dependent conditions.</p>
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<p>Glucose absorption in IEC-6 cells over time against reference inhibitors of glucose transporters (phloridzin and phloretin), ferulic acid, brewers’ spent grain intestinal digest (DBSG) and extruded brewers’ spent grain intestinal digest (DEBSG) under sodium-free conditions.</p>
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13 pages, 1302 KiB  
Article
In Vitro Hypoglycemic and Anti-Inflammatory Potential and Toxicity of Powders from Pulp and by-Products of Ziziphus mistol from Argentina
by María Eugenia Orqueda, Sebastian Torres, Iris Catiana Zampini and María Inés Isla
Foods 2022, 11(14), 2125; https://doi.org/10.3390/foods11142125 - 18 Jul 2022
Cited by 3 | Viewed by 1808
Abstract
Background: The Ziziphus mistol fruit (vulgar name mistol) is used in northwestern Argentina in traditional food and beverage preparations and popular medicines for liver and respiratory disorders. Aims: The aim of this research was to evaluate the hypoglycemic and anti-inflammatory activity in pulp [...] Read more.
Background: The Ziziphus mistol fruit (vulgar name mistol) is used in northwestern Argentina in traditional food and beverage preparations and popular medicines for liver and respiratory disorders. Aims: The aim of this research was to evaluate the hypoglycemic and anti-inflammatory activity in pulp powders and sub-products (skin and seeds) of mistol fruit, along with their toxicity. Methods: Powders from mistol seeds, pulp, and skin were obtained. Antioxidant capacity and inhibitory activity against key enzymes involved in metabolic syndrome were determined by in vitro assays. Results: The mistol powders obtained from the different fruit parts reduced glucose bioaccessibility. Before and after simulated gastroduodenal digestion, the polyphenol-enriched extracts (PEE) obtained from mistol powders increased glucose uptake by yeast cells and inhibited the pivotal enzymes of the inflammatory pathway (cyclooxygenase-2, lipooxygenase-1, and phospholipase A2). The analyzed mistol powders did not show acute toxicity or genotoxicity in model organisms and cell cultures. Conclusions: These results evince the potentiality of both the pulp from Z. mistol fruits and residual biomass (seeds and skin) to obtain biofunctional powders to use as supplements for metabolic disorders associated with chronic diseases. Full article
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<p>Mistol distribution map in South America.</p>
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<p>Glucose adsorption capacity (determined by measuring the mM of glucose in dialysate) of digested and undigested mistol powders. Values are presented as mean ± SEM and scrutinized by 1 or 2-way ANOVA, followed by a Tukey or LSD test using R studio software. Different letters on the bars indicate significant differences between the three parts of the fruit, evaluated according to Tukey’s test (<span class="html-italic">p</span> ≤ 0.05). No significant differences (<span class="html-italic">p</span> ≤ 0.05) were observed between digested and undigested powders.</p>
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<p>Effect of different amounts (0.25 g, ▲; 0.5 g, ●; and 1.0 g, ■) of mistol powders on glucose diffusion: (<b>A</b>) seed, (<b>B</b>) pulp, and (<b>B</b>,<b>C</b>) skin powders. Results are expressed as the mean of three independent tests with SEM. The symbol * at a given time indicates significant differences (<span class="html-italic">p</span> ≤ 0.05) between the glucose content in the dialysate between the three amounts of mistol powder, according to Tukey’s test.</p>
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<p>Effect of mistol extracts (PEE) on glucose consumption by <span class="html-italic">S. cerevisiae</span>: Skin (■), pulp (●), and seed (▲) PEE. Results are expressed as the mean of three independent tests with SEM. The symbol * at a given extract concentration indicates significant differences (<span class="html-italic">p</span> ≤ 0.05) between percentage of improvement in glucose intake by yeast cells between the three parts of mistol fruits, according to Tukey’s test.</p>
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19 pages, 854 KiB  
Article
Prototype Gluten-Free Breads from Processed Durum Wheat: Use of Monovarietal Flours and Implications for Gluten Detoxification Strategies
by Rosa Pilolli, Maria De Angelis, Antonella Lamonaca, Elisabetta De Angelis, Carlo Giuseppe Rizzello, Sonya Siragusa, Agata Gadaleta, Gianfranco Mamone and Linda Monaci
Nutrients 2020, 12(12), 3824; https://doi.org/10.3390/nu12123824 - 14 Dec 2020
Cited by 4 | Viewed by 2379
Abstract
In this investigation, we reported the production of prototype breads from the processed flours of three specific Triticum turgidum wheat genotypes that were selected in our previous investigation for their potential low toxic/immunogenic activity for celiac disease (CD) patients. The flours were subjected [...] Read more.
In this investigation, we reported the production of prototype breads from the processed flours of three specific Triticum turgidum wheat genotypes that were selected in our previous investigation for their potential low toxic/immunogenic activity for celiac disease (CD) patients. The flours were subjected to sourdough fermentation with a mixture of selected Lactobacillus strains, and in presence of fungal endoproteases. The breads were characterized by R5 competitive enzyme linked immunosorbent assay in order to quantify the residual gluten, and the differential efficacy in gluten degradation was assessed. In particular, two of them were classified as gluten-free (<20 ppm) and very low-gluten content (<100 ppm) breads, respectively, whereas the third monovarietal prototype retained a gluten content that was well above the safety threshold prescribed for direct consumption by CD patients. In order to investigate such a genotype-dependent efficiency of the detoxification method applied, an advanced proteomic characterization by high-resolution tandem mass spectrometry was performed. Notably, to the best of our knowledge, this is the first proteomic investigation which benefitted, for protein identification, from the full sequencing of the Triticum turgidum ssp. durum genome. The differences of the proteins’ primary structures affecting their susceptibility to hydrolysis were investigated. As a confirmation of the previous immunoassay-based results, two out of the three breads made with the processed flours presented an exhaustive degradation of the epitopic sequences that are relevant for CD immune stimulatory activity. The list of the detected epitopes was analyzed and critically discussed in light of their susceptibility to the detoxification strategy applied. Finally, in-vitro experiments of human gastroduodenal digestion were carried out in order to assess, in-silico, the toxicity risk of the prototype breads under investigation for direct consumption by CD patients. This approach allowed us to confirm the total degradation of the epitopic sequences upon gastro-duodenal digestion. Full article
(This article belongs to the Section Nutrition and Public Health)
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<p>Protein quantification of the model breads by Bradford colorimetric assay carried out on the total protein extracts.</p>
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<p>Overview of the peptides identified in the low molecular weight fraction (LMW, &lt;3 kDa) by the HR-MS/MS analysis. Panel (<b>a</b>): peptide count according to the sequence length. Panel (<b>b</b>): peptide count according to the belonging protein (please consider that several peptides were shared among different accessions).</p>
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17 pages, 1210 KiB  
Article
A Comprehensive Peptidomic Approach to Characterize the Protein Profile of Selected Durum Wheat Genotypes: Implication for Coeliac Disease and Wheat Allergy
by Rosa Pilolli, Agata Gadaleta, Luigia Di Stasio, Antonella Lamonaca, Elisabetta De Angelis, Domenica Nigro, Maria De Angelis, Gianfranco Mamone and Linda Monaci
Nutrients 2019, 11(10), 2321; https://doi.org/10.3390/nu11102321 - 1 Oct 2019
Cited by 10 | Viewed by 3634
Abstract
The wheat varietal selection undertaken by breeders in recent decades has been tailored mainly to improve technological and productivity-related traits; however, the latter has resulted in a considerable impoverishment of the genetic diversity of wheat-based products available on the market. This pitfall has [...] Read more.
The wheat varietal selection undertaken by breeders in recent decades has been tailored mainly to improve technological and productivity-related traits; however, the latter has resulted in a considerable impoverishment of the genetic diversity of wheat-based products available on the market. This pitfall has encouraged researchers to revalue the natural diversity of cultivated and non-cultivated wheat genotypes in light of their different toxic/immunogenic potential for celiac disease and wheat-allergic patients. In the present investigation, an advanced proteomic approach was designed for the global characterization of the protein profile of selected tetraploid wheat genotypes (Triticum turgidum). The approach combined proteins/peptides sequence information retrieved by specific enzymatic digestions (single and dual proteolytic enzymes) with protein digestibility information disclosed by means of in-vitro simulated human gastroduodenal digestion experiments. In both cases, the peptide pools were characterized by discovery analysis with liquid chromatography high-resolution tandem mass spectrometry, and specific amino acid sequences were identified via commercial software. The peptide list was screened for in silico toxicity/immunogenicity risk assessment, with the aid of various open-source bioinformatics tools for epitopes matching. Given the global information provided by the designed proteomic approach, the in silico risk assessment not only tackled toxicity implication for celiac disease patients, but also scouted for immunogenic sequences relevant for wheat allergic patients, achieving a comprehensive characterization of the protein profile of the selected genotypes. These latter were assessed to encrypt a variable number of toxic/immunogenic epitopes for celiac disease and wheat allergy, and as such they could represent convenient bases for breeding practices and for the development of new detoxification strategies. Full article
(This article belongs to the Special Issue Contributions of Diet and Gastrointestinal Digestion to Food Allergy)
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<p>Description of the analytical workflow applied for the comprehensive peptidomic approach to characterize the protein profile of wheat genotypes.</p>
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<p>Venn diagrams highlighting the distribution of identified proteins (panels (<b>a</b>) and (<b>c</b>)) and peptides (panels (<b>b</b>) and (<b>d</b>)) among the three different protein fractions subjected to specific digestion with chymotrypsin.</p>
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<p>Workflow of the two-round identification protocol applied to maximize the number of GD resistant peptides identified.</p>
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<p>Distribution among main proteins of the detected CD (<b>a</b>,<b>b</b>) and WA (<b>c</b>,<b>d</b>) epitopes in Osborne fractions and GD digests.</p>
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<p>Distribution among the main proteins of the hazard peptides detected in GD digest containing intact epitopes relevant for CD (<b>a</b>) and WA (<b>b</b>).</p>
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