A biochemical approach to study wheat products digestibility and wheat allergens degradation

This thesis is focused on wheat digestibility, particularly referring to proteins, starch and fibre nutrients, which can plays a crucial role in several fields as in food allergic reactions, glycemic index assessment and in the fibre degradation. Wheat is one of the most important crops in the world, used to make a vast range of food products, such as bread, pasta and cakes. Allergic reactions to food, including wheat products, are becoming more common in Western countries affecting up to 2% of the adult population and 6% of children (Sotkovsky et al., 2008). The stability of proteins to the gastro-intestinal digestion process after heat treatment may contribute to their allergenic potential. It is thought that resistance to digestion plays an important role in the development of sensitisation, and it is postulated that resistance to gastric conditions (low pH and pepsinolysis) may have a significant impact on the solubility and allergenic potential of food proteins. Indeed, both intact proteins and digested polypeptides have the potential to elicit an allergic reaction. In order to better understand how chemical and biochemical changes induced by food processing (such as cooking) affect digestibility of wheat flour proteins, one dimensional (1D) and two-dimensional electrophoresis (2-DE) digestion maps of heated and unheated flour have been produced. Experiments were carried out using an in vitro static system with an additional ‘chewing’ step including human salivary amylase, in order to study oral degradation of wheat flour proteins. Afterwards, an in vitro dynamic model of digestion has been performed to gain a more reliable comprehension on protein degradation. Besides, as known, starch is the main constituent in cereal food and it influences the rate and the extent of digestion and, consequently, its nutritional and health properties. Foods having a low glycemic index have been suggested from FAO/WHO organisations (1998) since it promotes the slow release of glucose in the body. This work involved also the study of three types of pasta which were subjected to in vitro and in vivo digestions in order to assess their glycemic index. Finally, another crucial nutrient component in wheat is represented by arabinoxylans (AX) which are hemicellulose belonging to the fibre group. This fibre have been described as resistant to gastrointestinal digestion, since it is usually degraded in the colon tract, by macrobiota. However in vitro simulated digestions of arabinoxylans, extracted from wheat dough, were performed, and the effect on arabinose substitution level was investigated. Results mainly showed a reduced protein degradation in heated flour, suggesting that heat treatment can modify protein susceptibility to digestion. Results concerning the glycemic index study gave evidence that the wheat genotype can affect the rate of starch hydrolysis due to some nutrients, as proteins, which may vary among wheat varieties. Finally, concerning the arabinoxylans digestions, interesting findings showed that enzymes and surfactants and either the acid gastric pH, were able to affect AX region IR spectra.Il lavoro sviluppato in questa tesi riguarda la digeribilità dei nutritivi del frumento, con particolare riferimento a proteine, amido e fibra. La digestione dei nutritivi gioca un ruolo chiave nelle reazioni allergiche agli alimenti, nella valutazione dell’indice glicemico e infine nella degradazione della fibra. Il frumento è una tra le più importanti produzioni cerealicole mondiali e viene utilizzato per la produzione di svariati prodotti: pane, pasta, dolci. Le reazioni allergiche agli alimenti, incluse quelle indotte dal frumento, stanno diventando sempre piu’ diffuse nei paesi occidentali, colpendo circa il 2% degli adulti e il 6% dei bambini (Sotkovsky et al., 2008). Tra i fattori che contribuiscono al potenziale allergenico di un alimento si deve considerare la stabilità delle proteine in seguito al processo digestivo e di cottura. Infatti è noto che la resistenza degli allergeni al trattamento di digestione è tra i parametri coinvolti nello sviluppo della sensitizzazione individuale, e le condizioni ambientali gastriche possono influenzare la solubilità di potenziali allergeni. È altresi’ noto che proteine o polipeptidi non digeriti hanno la potenzialità di scaturire una reazione allergica. In questo lavoro è stato studiato l’effetto del trattamento termico e della digestione, in farine di frumento cotta e non, attraverso l’analisi di profili proteici 1D e di mappe bidimensionali 2D. Questi esperimenti sono stati effettuati utilizzando un sistema di digestione statico simulando la fase orale, gastrica e intestinale. Inoltre, successivamente, è stato messo a pun to un sistema di digestione dinamico, per lo studio della degradazione delle proteine in farina cotta. Inoltre, è noto che l’amido è il principale componente dei prodotti cerealicoli e ne influenza le proprietà salutistiche nutrizionali. Alimenti con basso indice glicemico sono stati suggeriti dalle organizzazioni FAO/WHO. Questo lavoro ha coinvolto l’utilizzo di tre varietà di pasta, ove, esperimenti di digeribilità dell’amido in vitro e in vivo, hanno permesso di fornire una valutazione del loro indice glicemico. Infine, componenti della fibra in farine di frumento sono stati studiati attraverso la messa a punto di un sistema di digestione in vitro, per valutare l’effetto dell’ambiente gastro-intestinale sugli arabinoxilani estratti da impasti di farina. I risultati mostrano come il trattamento termico della farina di frumento modifichi la degradazione degli allergeni durante la digestione gastro-intestinale in vitro. Inoltre, lo studio delle tre varietà di pasta dimostra differenze significative nei valori di indice glicemico, imputabili al genotipo. Infine dalla digestione degli arabinoxilani, i risultati riportano sorprendentemente un effetto della digestione gastrica e intestinale, sulla parziale midificazione di queste fibre

Digeribilit\ue0\ua0 in vitro dell'amido e risposta glicemica in vivo di pasta ottenuta da antiche specie di frumento

La qualit\ue0 nutrizionale degli alimenti riveste oggigiorno un\u2019importanza strategica sia per i produttori che per i consumatori. I comitati di consultazione per la nutrizione umana (FAO/WHO) raccomandano il consumo di alimenti a basso indice glicemico, o lenta digeribilit\ue0 dell\u2019amido, che hanno un effetto sulla riduzione della glicemia e della risposta insulinemica ed esplicano altri ben noti effetti salutistici nei confronti di diverse malattie metaboliche (diabete tipo II, sindrome metabolica). Per quanto riguarda la pasta, questi indici qualitativi possono dipendere sia dalla materia prima (origine, variet\ue0, grado di macinazione delle semole) sia dai trattamenti tecnologici che essa subisce durante la sua trasformazione (impastamento, estrusione, essiccazione, cottura). Recentemente, vi \ue8 stato un crescente interesse per i 'grani antichi' che comprendono specie diverse appartenenti al genere Triticum (Triticum monococcum, o farro piccolo, T. dicoccum, o farro medio, T. spelta, o farro grande, T. polonicum, o Kamut\uae) che secondo alcuni autori (Bonafaccia et al. 2000, Marques et al., 2007), potrebbero vantare elevate caratteristiche nutrizionali e di digeribilit\ue0 rispetto ai frumenti moderni. In questo studio \ue8 stata valutata l\u2019influenza del genotipo sulla digeribilit\ue0 dell\u2019amido in vitro, e in vivo, di pasta alimentare ottenuta da T. dicoccum (farro medio) e da T. polonicum (Kamut\uae) rispetto a quella ottenuta da frumento moderno (T. durum). I risultati ottenuti dalle prove di digeribilit\ue0 dell\u2019amido in vitro e dalla misurazione dell\u2019indice glicemico in vivo mostrano differenze significative tra la pasta ottenuta da Triticum durum e quella ottenuta da T. polonicum

Pasta ottenuta da antiche specie di frumento: studio preliminare della digeribilit\ue0 dell'amido

FAO and WHO organisations recommend to consume foods with a low glycemic index, that are able to reduce the glycemic and insulinemic response. Moreover these foods are involved in the prevention of metabolic deseases. In these last years the interest for \u201cancient grains\u201d has increased a lot, so, this study has involved pasta made from 3 Triticum grains (Triticum dicoccum, T. polonicum, and T. durum). Results show statistical significant differences in glycemic index between pasta of T. durum and T. polonicum

Assessment of the fining efficiency of zeins extracted from commercial corn gluten and sensory analysis of the treated wine

The aims of the present work are to evaluate zeins as potential clarification agent for red wine and to assess the impact of zeins treatment on the wine \u201cbouquet\u201d through sensory analysis. Zeins have been extracted from commercial corn gluten which is a protein-rich by-product deriving from maize starch industry. Three red wines (cv. Cabernet Sauvignon, Merlot and Valpolicella) have been used for the fining experiments in order to test zein proteins (Z) in comparison with a commercial nonhydrolysed animal gelatins (G1). The new method of zeins extraction gave a good yield of extraction (about 72%). These proteins showed an excellent fining action if compared to wine treatment with G1. In fact, zeins treatment could decrease wine turbidity and remove phenolic compounds, such as anthocyanins and proanthocyanidins, and, at the same time, it did not affect red wine colour. Finally, the sensory analysis indicated that no significant differences were detected between the same wines clarified with Z or with G1. In conclusion, the results show that corn gluten can be used in place of potentially allergenic animal proteins to obtain a low cost fining agent that can be introduced during the wine clarification step

Assessment of the fining efficiency of zeins extracted from commercial corn gluten and sensory analysis of the treated wine

The aims of the present work are to evaluate zeins as potential clarification agent for red wine and to assess the impact of zeins treatment on the wine \u201cbouquet\u201d through sensory analysis.Zeins have been extracted from commercial corn gluten which is a protein-rich by-product deriving from maize starch industry. Three red wines (cv. Cabernet Sauvignon, Merlot and Valpolicella) have been used for the fining experiments in order to test zein proteins (Z) in comparison with a commercial non-hydrolysed animal gelatins (G1). The new method of zeins extraction gave a good yield of extraction (about 72%). These proteins showed an excellent fining action if compared to wine treatment with G1. In fact, zeins treatment could decrease wine turbidity and remove phenolic compounds, such as anthocyanins and proanthocyanidins, and, at the same time, it did not affect red wine colour. Finally, the sensory analysis indicated that no significant differences were detected between the same wines clarified with Z or with G1.In conclusion, the results show that corn gluten can be used in place of potentially allergenic animal proteins to obtain a low cost fining agent that can be introduced during the wine clarification step

Helicobacteraceae in Bulk Tank Milk of Dairy Herds from Northern Italy

Helicobacter pylori is responsible for gastritis and gastric adenocarcinoma in humans, but the routes of transmission of this bacterium have not been clearly defined. Few studies led to supposing that H. pylori could be transmitted through raw milk, and no one investigated the presence of other Helicobacteraceae in milk. In the current work, the presence of Helicobacteraceae was investigated in the bulk tank milk of dairy cattle herds located in northern Italy both by direct plating onto H. pylori selective medium and by screening PCR for Helicobacteraceae, followed by specific PCRs for H. pylori, Wolinella spp., and “Candidatus Helicobacter bovis.” Three out of 163 bulk milk samples tested positive for Helicobacteraceae, but not for the subsequent PCRs. H. pylori was not isolated in any case. However, given similar growth conditions, Arcobacter butzleri, A. cryaerophilus, and A. skirrowii were recovered. In conclusion, the prevalence of Helicobacteraceae in raw milk was negligible (1.8%), and H. pylori was not identified in any of the positive samples, suggesting that, at least in the farming conditions of the investigated area, bovine milk does not represent a potential source of infection

Effect of dough mixing on wheat endosperm cell walls

Dough-derived cell wall fragments isolated by ultracentrifugation were largely derived from the starchy endosperm, with some fragments deriving from the aleurone and outer layers, as indicated by fluorescence microscopy. Dough mixing had little effect on the structure and composition of cell wall fragments compared to thin grain sections, as determined by Fourier transform infrared (FTIR) and 1H nuclear magnetic resonance (NMR) spectroscopy. These analyses confirmed that the fragments largely comprised water-unextractable arabinoxylan and β-glucan. FTIR microspectroscopy of dough-derived cell wall fragments prepared from five bread wheat cultivars showed that two largely comprised highly substituted arabinoxylan (cv. Manital and San Pastore), one comprised a mixture of low, medium, and highly substituted arabinoxylan (cv. Hereward), and the remaining two comprised a greater proportion of low substituted arabinoxylan (cv. Claire and Yumai 34). Yumai 34 yielded a greater mass of cell wall material, and its cell walls comprised a high proportion of medium substituted arabinoxylan. Such methods will allow for the impact of bakery ingredients and processing on endosperm cells, including the addition of xylanases, to be investigated in the future to ensure any potential health benefits arising from wheat breeding are realized in the food that reaches the consumer