http://hdl.handle.net/1889/666 Wed, 19 Jun 2013 15:49:45 GMT 2013-06-19T15:49:45Z http://hdl.handle.net/1889/1906 Title: Composizione e dinamiche della microflora in differenti ecosistemi caseari Authors: Santarelli, Marcela Abstract: Il formaggio è un cibo biologicamente e biochimicamente dinamico contenente microorganismi sia intenzionalmente aggiunti come colture starter sia presenti come contaminanti avventizi di natura non-starter. La composizione della popolazione microbica si modifica sotto l’influenza di continui cambiamenti delle condizioni ambientali e delle interazioni tra i microorganismi durante la produzione e la stagionatura. Nel processo produttivo del formaggio, la selezione di parametri tecnologici può influenzare o anche indurre vari processi biochimici necessari per l’ottenimento di questo alimento. Le popolazioni microbiche presenti nel formaggio sono complesse e la loro crescita e attività rappresentano le fasi più importanti ma meno controllabili del processo di caseificazione. Lo scopo della presente tesi di dottorato è lo studio della diversità microbica e delle dinamiche delle colture starter, non-starter e dei microrganismi contaminanti coinvolti nei processi di produzione e stagionatura del formaggio. A tale scopo, diversi ecosistemi, dal latte crudo al formaggio, e la crosta sono stati studiati al fine di comprendere il ruolo specifico svolto dai vari microorganismi in ciascuna fase del processo di produzione e correlare la presenza di alcune specie microbiche con lo sviluppo di aromi e strutture desiderate del prodotto. Nello specifico, gli ecosistemi microbici di sieroinnesti naturali, formaggi duri italiani (Grana Padano e Parmigiano Reggiano), e della crosta naturale di un formaggio erborinato stagionato, sono stati studiati per tutto il processo di produzione e stagionatura mediante tecniche culture-dependent e culture-independent. Attraverso lo studio della microflora di sieroinnesti naturali e formaggi a partire dai primi giorni di produzione, è stato possibile descrivere l’attività fermentativa dei batteri lattici starter (SLAB) che determinano il processo di stagionatura del formaggio. L’acidità titolabile del sieroinnesto non è risultata essere in relazione né con la quantità di cellule (totali e coltivabili) né con il contributo specifico delle differenti specie. Elevate concentrazioni di acido lattico e aminoacidi liberi sono state trovate in formaggi con maggiori quantità della specie Lactobacillus helveticus e con alte densità di batteri termofili coltivabili. La presenza di galattosio residuo è risultata associata ad alti contenuti della specie Streptococcus thermophilus. Inoltre, la composizione in termini di biotipi dei sieroinnesti è apparsa molto più importante che la composizione in termini di specie nell’assicurare le buone performance dei sieroinnesti stessi. Una correlazione diretta tra la composizione delle specie SLAB e l’efficacia di acidificazione nei sieroinnesti non è stata trovata. Al contrario, nella matrice formaggio, l’attività fermentativa degli SLAB è sembrata essere specie-dipendente. Negli ecosistemi dei formaggi duri, sia gli SLAB che i LAB non-starter (NSLAB) sono apparsi contribuire all’acidificazione e alla maturazione. Tuttavia, gli SLAB, ed in particolare L. helveticus, sono risultati essere principalmente soggetti alla lisi che si verifica a 2 mesi di stagionatura. I NSLAB sono stati in grado di crescere dopo la salatura, diventando la maggioranza dei microorganismi durante la stagionatura. I NSLAB potrebbero provenire sia dal latte crudo che dal sieroinnesto naturale ed il loro contributo allo sviluppo delle caratteristiche del formaggio è ancora sconosciuto. Inoltre, la presenza di L. helveticus e Lactobacillus delbrueckii subsp. lactis in stato non coltivabile, fino a 13 mesi di stagionatura, suggerisce che queste specie potrebbero svolgere un ruolo diverso ma ancora sconosciuto nella stagionatura del formaggio. La microflora della superficie di un formaggio erborinato durante lo sviluppo naturale della crosta ha mostrato una diversità microbica composta da 14 generi di batteri (Enterococcus; Lactococcus; Leuconostoc; Macrococcus; Staphylococcus; Klebsiella; Brevibacterium; Corynebacterium; Brachybacterium;, Nocardiopsis;, Cobetia; Psychrobacter; Halomonas; Haererehalobacter), due generi di lieviti (Candida; Debaryomyces) e un genere di fungo filamentoso (Penicillium). Sono state osservate elevate e comparabili densità di batteri e lieviti vitali. E’ stata osservata inoltre, un’evoluzione batterica durante la formazione della crosta e nessun genere è rimasto costantemente presente per tutta la durata della stagionatura. Staphylococcus si è dimostrato il genere dominante durante le fasi precoci e successivamente è stato sostituito da Brevibacterium alla fine della stagionatura. Sfruttando esperimenti di interazione, sono stati osservati effetti di inibizione e stimolazione tra le specie; queste interazioni potrebbero spiegare come alcuni microorganismi contribuiscano alla formazione della comunità microbica. Candida catenulata e Debaryomyces hansenii hanno incrementato la crescita di Staphylococcus equorum mentre C. catenulata ha inibito la crescita di D. hansenii. Tuttavia, studi approfonditi dovranno essere svolti per valutare se le specie batteriche, di lievito e muffe sopracitate possano essere utili e svolgere un ruolo nello sviluppo dell’aroma e della struttura di altre varietà di formaggi che presentino simile crosta naturale. In conclusione, grazie ad approcci complementari culture-dependent e culture-independent, è stato possibile identificare quali microrganismi fossero principalmente coinvolti in ciascuna delle matrici casearie studiate, e definire l’importanza della loro presenza che, se in giusto equilibrio, può favorire l’ottenimento delle diverse caratteristiche peculiari di ciascun prodotto.; Cheese is a biologically and biochemically dynamic food containing microorganisms both deliberately added as starters and non-starter adventitious contaminants. The composition of microbial population changes under the influence of continuous shifts in environmental conditions and interactions occurring among microorganisms during manufacturing and ripening. In cheese manufacturing, the selection of technological parameters can influence and even induce several biochemical processes needed for this product. The microbiota present in cheese is complex and its growth and activity represent the most important, but the least controllable steps. The aim of the present thesis is the study of microbial diversity and dynamics of starter, non-starter and adventitious microorganisms involved in the cheese manufacturing and ripening processes. Thus, different ecosystems from raw milk to cheese and rind were investigated in order to comprehend the specific role played by microorganisms in each cheese making phase and to correlate the occurrence of certain microbial species with desired flavors and textures. In particular, the microbial ecosystems in natural whey starters, hard Italian cheeses (i.e. Grana Padano and Parmigiano Reggiano), and natural rind of mold-ripened cheese (i.e. blue cheese) were investigated throughout cheese manufacturing and ripening by culture-dependent and culture-independent techniques. By studying the microbiota in natural whey starters and cheese starting from the first days of manufacture, it was possible to describe the fermentative activity of starter lactic acid bacteria (SLAB) determining the ripening progress of the cheese. Whey starter titratable acidity did not seem to be related to the cell amount (total and cultivable cells) nor to the different species contribution. High concentrations of lactic acid and free aminoacids were found in cheeses with higher levels of Lactobacillus helveticus species and cultivable thermophilic bacterial densities. The presence of residual galactose was associated with higher contents of Streptococcus thermophilus species. In addition, the biotype composition of whey starters seemed to be far more important than the species composition in ensuring their good performances. A direct correlation between the composition of SLAB species and the acidifying efficacy in whey starters was not found. Contrarily, in the cheese matrix, the SLAB fermentative activity seemed to be species-dependent. In hard cheese ecosystems, both SLAB and non-starter LAB (NSLAB) seemed to contribute to acidification and ripening. However, SLAB, and in particular L. helveticus, resulted to be the species mainly subjected to the lysis occurring at 2 months of ripening. NSLAB were able to grow after brining, and became more relevant during ripening. NSLAB could arise both from the raw milk and the natural whey starter but their contribution to the development of cheese characteristics is still unknown. Furthermore, the presence of L. helveticus and Lactobacillus delbrueckii subsp. lactis in a non-cultivable state, up to 13 months of ripening, suggests that these species could play a different but still unknown role in cheese ripening. The microbiota on the surface of a blue cheese during the natural rind development showed microbial diversity comprising fourteen genera of bacteria (Enterococcus; Lactococcus; Leuconostoc; Macrococcus; Staphylococcus; Klebsiella; Brevibacterium; Corynebacterium; Brachybacterium;, Nocardiopsis;, Cobetia; Psychrobacter; Halomonas; Haererehalobacter), two yeasts genera (Candida; Debaryomyces) and one filamentous fungal genus (Penicillium). High and comparable densities of viable bacteria and yeasts were observed. Bacterial succession was observed during rind formation and no genus remained constant throughout ripening. The Staphylococcus genus dominated the early stages and then Brevibacterium the later stages. By using interaction experiments, inhibition and stimulation were observed among several species; these interactions could explain how some microorganisms contribute to community formation. Candida catenulata and Debaryomyces hansenii enhanced the growth of Staphylococcus equorum while C. catenulata inhibited D. hansenii growth. However, thorough studies need to be performed in order to evaluate whether the above mentioned bacteria, yeasts and molds can be beneficial and play a role in flavor and texture development of other cheese varieties with similar natural rinds. Overall, thanks to culture-dependent and culture-independent complementary approaches it was possible to identify which microorganisms were mainly involved in each dairy matrix and to address the importance of their presence that, if balanced, can help obtaining the distinctive features of each product. Sat, 31 Dec 2011 23:00:00 GMT http://hdl.handle.net/1889/1906 2011-12-31T23:00:00Z http://hdl.handle.net/1889/1905 Title: Batteri lattici non starter durante la stagionatura del formaggio: sopravvivenza, crescita e produzione di molecole potenzialmente coinvolte nella formazione dell'aroma Authors: Sgarbi, Elisa Abstract: I batteri lattici (LAB) costituiscono un gruppo eterogeneo di batteri tradizionalmente utilizzati nella produzione di formaggi. Essi possono svolgere diversi ruoli durante la caseificazione, gli starter LAB (SLAB) sono i protagonisti del processo di fermentazione, mentre i non starter LAB (NSLAB) esplicano il loro ruolo durante la maturazione del formaggio. Il lavoro di ricerca di questo dottorato si è focalizzato sullo studio del contributo dei NSLAB durante la stagionatura del formaggio. In particolare è stato preso in esame uno dei più famosi formaggi Italiani a lunga stagionatura, il Parmigiano Reggiano. Durante la maturazione del formaggio gli SLAB muoiono e lisano di conseguenza; i NSLAB, invece, iniziano a crescere e si sviluppano durante la stagionatura. Durante il periodo di maturazione del Parmigiano Reggiano i NSLAB crescono in mancanza di zuccheri del latte, perché il lattosio è già stato consumato dagli SLAB durante le fasi iniziali della produzione del formaggio. La capacità dei NSLAB di sopravvivere e crescere in queste condizioni potrebbe essere dovuta alla loro abilità di utilizzare fonti energetiche alternative, come i prodotti derivanti dalla lisi cellulare dei SLAB. Il primo studio riportato in questa tesi di dottorato, ha confermato la capacità dei NSLAB di sopravvivere e crescere utilizzando come fonte di energia solo i prodotti di lisi cellulare degli SLAB. Gli esperimenti condotti per sostenere questa ipotesi hanno dimostrato che i prodotti derivanti dalla lisi di L. helveticus (SLAB) possono fornire nutrienti sufficienti a consentire la crescita in vitro di due specie di NSLAB, L. casei e L. rhamnosus. Inoltre, l’osservazione di cambiamenti nella composizione del prodotto di lisi degli SLAB dopo la crescita dei NSLAB, suggerisce che alcuni composti possano essere consumati o prodotti dal metabolismo della microflora non starter durante la crescita. Ulteriore scopo di questa tesi di dottorato è stato indagare se il metabolismo dei NSLAB, che crescono utilizzando le fonti di nutrimento disponibili nel formaggio stagionato, potesse contribuire alla produzione di molecole volatili coinvolte nella formazione del flavour del formaggio. I risultati ottenuti hanno portato a supporre che molte molecole importanti, che contribuiscono al flavour, derivino da vie metaboliche seguite dai NSLAB, come il metabolismo del lattato/citrato, il catabolismo degli aminoacidi e degli acidi grassi liberi e dall’uso dei pochi zuccheri contenuti nel formaggio stagionato (come N-acetilglucosamina derivante dalla parete cellulare degli SLAB). La presenza di questi composti volatili ha confermato che, durante la stagionatura, la microflora NSLAB contribuisce a sviluppare l’aroma del formaggio. Infine, è stata valutata la capacità dei LAB di sintetizzare nuove molecole, coinvolte nell’aroma caratteristico del formaggio, γ-glutamil-amminoacidi e lactoyl-amminoacidi. E’ stato osservato che, L. helveticus, scelto come rappresentante della microflora starter, e L. rhamnosus, rappresentante dei NSLAB, sono in grado di produrre sia γ-glutamil-fenilalanina e lactoyl-fenilalanina grazie probabilmente ad una particolare un'attività enzimatica batterica. Mentre il primo composto è stato prodotto solo dalle cellule dopo lisi, la lactoyl-fenilalanina è stata prodotta dalle cellule dei LAB sia durante la crescita che dopo lisi, in quantità diverse a seconda della specie, della condizione cellulare e del tempo di incubazione. Vista la bassa resa delle reazioni, la maggiore quantità di queste molecole, che si trova nel formaggio, potrebbe essere una conseguenza del loro accumulo durante il lungo periodo di maturazione che caratterizza i formaggi a lunga stagionatura. Le capacità dei NSLAB e le loro attività durante la maturazione del formaggio si sono rivelate molto interessanti e potrebbero trovare una applicazione tecnologica nella produzione di formaggi o altri prodotti fermentati stagionati. In particolare, lo studio e la conseguente conoscenza delle abilità specie-specifiche dei NSLAB che portano alla formazione di composti aromatici, potrebbero essere di grande interesse per i produttori di formaggi in tutto il mondo.; Lactic Acid Bacteria (LAB) constitute a heterogeneous group of bacteria that are traditionally used in cheese manufacturing. They may play different roles in cheese-making where starter LAB (SLAB) participate in the fermentation process, whereas non starter LAB (NSLAB) are implicated in cheese ripening. This PhD thesis focused on NSLAB role during cheese ripening, in particular in a famous long ripened cheese, Parmigiano Reggiano. During ripening the death and lysis of SLAB (the dominant microflora at the culmination of curd acidification) and the growth of NSLAB, occur. In Parmigiano Reggiano cheese NSLAB grow in lacking sugars conditions, because the lactose has already been exhausted by SLAB during the initial step of cheese making. The ability of NSLAB to survive in these conditions could be due to their particular capabilities to use alternative energy sources, such as the SLAB cell lysis products. The first study reported in this PhD thesis, confirmed the NSLAB ability to survive and grow using as nutrient source only SLAB cell lysis products. In fact, the experiments conducted to sustain this hypothesis confirmed that L. helveticus (SLAB) lysate products provide sufficient nutrients to allow the growth of two NSLAB species, L. casei and L. rhamnosus, in vitro. Furthermore the changes in the SLAB lysis product composition suggest that some compounds can be either consumed or produced by bacterial metabolism during growth. Secondly this PhD research aimed to investigate if the NSLAB metabolism in these lacking sugar conditions, could contribute to the production of volatile molecules involved in cheese flavour formation. The “volatilomes” of L. rhamnosus and L. casei, grown using SLAB cell lysis products or a cheese model medium, were studied. The results allowed to suppose that some important molecules, involved in flavour formation, can be derived from NSLAB metabolic pathways, such as lactate/citrate metabolism, amino acids catabolism, free fatty acids metabolism and from the use of the few sugars contained in ripened cheese (i.e. N-acetyl glucosamine from SLAB cell wall). The presence of these volatile compounds confirmed that, during aging, the NSLAB contribute to develop flavour of ripened cheese. Finally, the ability of LAB to synthesize specific new molecules, γ-glutamil-amino acids and lactoyl-amino acids, potentially involved in flavour was evaluated. It was observed that, L. helveticus, representative of SLAB, and L. rhamnosus, representative on NSLAB, were able to produce both γ-glutamyl-phenylalanine and lactoyl-phenylalanine by enzymatic activity. While the first molecule was produced only by lysed cells, the latter was produced either by growing or lysed cells in different amount depending on the species, the cell condition and time of incubation. Having regard to the low yield of the reactions, the greater amount of these compounds, found in cheese, is a consequence of the long time of ageing and of the accumulation of the amino acidic precursors. The observed abilities of NSLAB and their activities during cheese ripening could be harnessed to several technological extents. In particular, the description of species specific features involved in flavour formation could be of great interest for the worldwide cheese producers. Sat, 31 Dec 2011 23:00:00 GMT http://hdl.handle.net/1889/1905 2011-12-31T23:00:00Z http://hdl.handle.net/1889/1902 Title: Probing quality and safety of tomato products by genomic and proteomic tools Authors: Bencivenni, Mariangela Abstract: La presente tesi di dottorato riguarda l’utilizzo di tecniche avanzate di genomica e proteomica per la valutazione della qualità e della sicurezza di una delle più importati e rinomate colture tradizionali italiane: il pomodoro. Nella prima sezione, approcci innovativi di proteomica sono stati applicati al problema delle allergie al pomodoro. Tecniche immunologiche classiche, basate sull’elettroforesi su gel per la separazione delle proteine, affiancate da sistemi di spettrometria di massa avanzati, sono state applicate ad un set di 12 ecotipi di pomodoro, consentendo l’identificazione e la caratterizzazione dei principali allergeni. Inoltre, sono state individuate e caratterizzate diverse isoforme della proteina allergenica del pomodoro Lipid Transfer Protein (LTP), sia negli estratti di buccia che di semi di pomodori comuni, mediante tecniche di spettrometria di massa ad alta risoluzione accoppiate a strategie di purificazione classiche ed innovative. Nella seconda sezione, tecniche di genomica avanzata sono state applicate alla questione dell’identificazione varietale in pomodoro. In particolare, è stata effettuata la discriminazione di sette varietà di pomodoro mediante l’utilizzo di sonde specifiche, chiamate acidi peptido nucleici (PNAs), che riconoscono SNP caratteristiche del pomodoro. Dopo che le prestazioni di PNA diversamente modificati (2D- and 5L- chiral-box), in termini di affinità di legame e di riconoscimento di mismatch, sono state testate e confrontati con quelle del PNA chirale, sia in soluzione che su superficie, il miglior modello di PNA è stato utilizzato per il design di un microarray basato sui PNA e per lo sviluppo di un sistema PNA-mediated PCR clamping per il riconoscimento di SNP specifiche del pomodoro.; The present Ph.D. thesis deals with the use of advanced genomic and proteomic techniques for the assessment of quality and safety of one of the most important traditional and worldwide renowned Italian crop products: tomato. In the first section, innovative proteomic approaches have been applied to the issue of tomato allergy. Classical immunological techniques, based on gel electrophoresis for the separation of proteins, along with advanced mass spectrometric systems, have been applied to a set of 12 tomato ecotypes, allowing the detection and the characterization of the main allergens. Moreover, several novel isoforms of allergenic tomato Lipid Transfer Protein (LTP) have been detected and characterized, both in peel and in seed fractions of common tomatoes, by means of high-resolution mass spectrometric techniques, coupled with classic and innovative purification strategies. In the second section, advanced genomic techniques have been applied to the problem of tomato variety identification. In particular, the discrimination of seven tomato varieties has been achieved by means of specific probes, namely Peptide Nucleic Acids (PNAs), recognizing characteristic tomato SNPs. After that the performances, in terms of binding affinity and mismatch recognition, of different modified PNAs (2D- and 5L- chiral-box) have been tested and compared to fully ‘achiral’ PNA, both in solution and on solid surfaces, the best PNA model was exploited in the design of a PNA-based microarray and in a PNA-mediated PCR clamping system for the recognition of tomato specific SNPs. Sat, 31 Dec 2011 23:00:00 GMT http://hdl.handle.net/1889/1902 2011-12-31T23:00:00Z http://hdl.handle.net/1889/1901 Title: Free and Hidden Fumonisins in Corn (Zea Mays L.): Occurrence and Masking Mechanism Authors: Falavigna, Claudia Abstract: Nel presente lavoro di tesi sono stati studiati diversi aspetti riguardanti le fumonisine, metaboliti secondari tossici prodotti da funghi del genere Fusarium, focalizzando l’attenzione sulle forme nascoste. In particolare, il meccanismo di mascheramento è stato ampiamente studiato attraverso modelli in vitro, allo scopo di capire qual è la natura delle interazioni che intercorrono tra queste micotossine e le macromolecole del mais. Successivamente, è stato effettuato uno studio allo scopo di individuare quali sono i principali fattori in grado di influenzare positivamente l’infezione fungina, la sintesi e il mascheramento delle fumonisine sulla pianta in fase di maturazione, con particolare attenzione riguardo al ruolo esercitato dal genotipo di mais, dalla sua composizione macromolecolare e dalle interazioni pianta-patogeno nel fenomeno di mascheramento. Infine, è stato sviluppato un metodo LC-ESI-MS/MS per l’analisi multiresiduale di diversi analoghi delle fumonisine appartenenti alle serie minori A e C, allo scopo di monitorarli in brodi di coltura di Fusarium sottoposti a condizioni di crescita diverse, ottenendo nuove informazioni riguardanti il pattern di produzione di tali composti.; The work is focused on the study of specific aspects of fumonisins, toxic metabolites produced by a number of Fusarium species in corn, considering especially hidden forms. In particular, masking mechanism has been largely investigated through in vitro experiments, in order to establish which type of interactions occur between such mycotoxins and maize macromolecules. Subsequently, the main factors able to promote fungal infection and fumonisins production and masking in growing maize have been researched, focusing the attention on the role played by the maize genotype and the macromolecular composition as well as by the plant-pathogen interaction in masking phenomenon. Finally, a multiresidual LC-ESI-MS/MS method has been developed to allow the monitoring of several fumonisin analogues produced by Fusarium species under different growing condition, collecting several information concerning the biosynthetic pattern of such compounds. Sat, 31 Dec 2011 23:00:00 GMT http://hdl.handle.net/1889/1901 2011-12-31T23:00:00Z http://hdl.handle.net/1889/1899 Title: Flavonoids and related compounds: new data on dietary sources, metabolism and availability in humans Authors: Calani, Luca Abstract: The present doctoral thesis deals with dietary polyphenols from several viewpoints. After a brief introduction to the topic, the work starts with an investigation into the polyphenolic composition of hazelnut skin. Hazelnuts are typically consumed whole (raw - with skin, or roasted - without skin) or used as ingredient in a variety of processed foods, especially in bakery and confectionery products. Hazelnut skin, hazelnut hard shell and hazelnut green leafy cover, as well as hazelnut tree leaf, are by-products of roasting, cracking, shelling/hulling, and harvesting processes, respectively and are now investigated for their composition in the attempt to add economic value to waste from the hazelnut industry. In this first work, skins from different hazelnut samples were characterised for total polyphenol content, total antioxidant capacity (TAC) and for their content in specific polyphenolic compounds. The second investigation deals with a source of peculiar polyphenols: the milk thistle. This study evaluated the absorption and metabolism of milk thistle flavonolignans (silychristin, silydianin, silybin and isosilybin isomers - all together known as silymarin) in humans. Fourteen volunteers consumed an extract of milk thistle and urine was collected up to 48 hours after consumption. Thirty-one metabolites were identified in urine by means of HPLC-MS/MS, monoglucuronides being the most common excreted form, followed by sulphate-glucuronides and diglucuronides, respectively. The third study aimed to investigate green tea flavan-3-ol catabolism, plasma pharmacokinetic and urinary excretion up to 24h from tea ingestion in humans, demonstrating that green tea catechins are more bioavailable than previously observed when colonic ring fission metabolites are taken into consideration. Regular consumption of green tea containing flavan-3-ols allows a non marginal exposition of the human body to their catabolites, somehow justifying the numerous beneficial actions described as linked to green tea intake. The data obtained in this last study along with those provided by other researchers, allowed to review the most recent findings on green tea flavan-3-ol bioavailability in humans in the fourth work presented in this thesis. The work also reviewed the bioavailability of coffee chlorogenic acids. The fifth study represents an update on the bioavailability of green tea flavan-3-ols. Novel information was obtained by monitoring urinary excretion of flavan-3-ol metabolites up to 48h from tea consumption by human volunteers. The estimated bioavailability was close to the 62% of the total ingested flavan-3-ols and the main reasons of this increment were the longer urinary collection period, allowing an almost complete excretion of flavan-3-ol metabolites, and the identification and quantification of novel molecules derived by microbial modification of flavan-3-ols. In conclusion, this doctoral thesis updates the knowledge about polyphenols by providing new information about their sources, their human and microbial metabolism and their bioavailability. All these aspects are paramount when the use of polyphenols as functional ingredients is suggested, as the defined identification and quantification of these molecules in food items, as well as their absorption and modifications occurring within the human body, are fundamental steps to unravel their putative beneficial effects toward human health. Sat, 31 Dec 2011 23:00:00 GMT http://hdl.handle.net/1889/1899 2011-12-31T23:00:00Z http://hdl.handle.net/1889/1675 Title: Cottura a vapore in forno: effetto di diverse umidità relative sulle caratteristiche fisiche, lo stato dell’acqua e la qualità sensoriale di carne di tacchino. Authors: Mora, Beatrice Abstract: La cottura a vapore in forno di carne di tacchino è una tecnica ampiamente applicata nei processi industriali, nella ristorazione collettiva e nella cucina domestica. Nei forni a vapore la cottura a convezione forzata dell'aria viene accoppiata ad un’iniezione di vapore nella camera di cottura. Nonostante la grande applicazione del vapore in cottura, l’effetto del vapore su resa di cottura, profili riscaldamento, consistenza, colore, e proprietà sensoriali della carne cotta non è ancora stato spiegato chiaramente. I dati pubblicati circa l'uso del forno a vapore a convezione sono ancora piuttosto limitati e contraddittori, e riguardano prevalentemente la cottura a vapore in condizioni di saturazione confrontata con cotture in assenza di vapore. Solo poche informazioni sono disponibili sull'uso di quantità di vapore inferiori alla condizione di saturazione. In questo lavoro e stato valutato con un approccio sistematico e multi-analitico l’effetto di diversi livelli di vapore (umidità relative dell’aria) sulla cottura di carne di tacchino. Le prove di cottura sono state effettuate con un dispositivo sperimentale di cottura a vapore espressamente concepito per modulare l'umidità relativa dell'aria. La valutazione della qualità della carne di tacchino cotto è stata effettuata studiando le caratteristiche fisiche (consistenza, colore, contrazione di volume), lo stato dell’ acqua e le proprietà sensoriali. La descrizione dello stato dell’ acqua con diversi parametri, quali, il contenuto di umidità, la capacità di trattenere l’acqua (WHC) e la mobilità molecolare (1H NMR), è risultato uno strumento molto utile per una migliore comprensione delle caratteristiche fisiche e delle proprietà organolettiche della carne cotta, che sono strettamente legate alla diversa denaturazione delle proteine indotta dal processo di cottura. I risultati indicano che diversi livelli di vapore (umidità relative) determinano diverse prestazioni in cottura e caratteristiche qualitative della carne cotta. L’applicazione del vapore porta a tempi di cottura più brevi rispetto alla cottura in assenza di vapore quando la temperatura della superficie della carne risulta sufficientemente superiore alla temperatura del centro del prodotto fissata come termine della cottura. Cotture con bassa quantità di vapore e tempi di cottura brevi rispetto a quelli ottenuti in assenza di vapore, hanno determinato una maggiore resa di cottura e una maggiore tenerezza percepita della carne rispetto alla cottura con elevate quantità di vapore (prossime alla condizione di saturazione). Sono stati definiti i valori minimi di umidità relativa a basse temperature di cottura (80-150 ° C) in grado di determinare effetti positivi sulla qualità della carne e un elevato potenziale risparmio di acqua. A basse temperature di cottura l'impiego di ridotte quantità di vapore potrebbe essere considerato una valida alternativa alla cottura in condizione di saturazione, in modo particolare per applicazioni industriali, grazie alla maggiore qualità della carne di tacchino e al potenziale risparmio di acqua.; Steam oven cooking of turkey meat is widely used in industrial processing, foodservice and domestic cooking. Steam cooking is achieved by steam injection in the oven chamber coupled with the air forced convection. Despite the large application of steam in cooking, its effect on cooking yield, heating profiles, texture, colour and sensory properties of cooked meat has not yet been clearly explained. Published data about the use of steam-convection oven are still rather limited and contradictory, and mainly deal with steam cooking in the saturation condition compared to dry oven cooking. Only little information is available on the use of steam quantities lower than the saturation condition. In this work the effect of different steam levels (air relative humidities) on turkey meat cooking with a systematic and multi-analytical approach was evaluated. The cooking trials were performed with an experimental steam cooking device expressly designed to modulate the air relative humidity. Quality evaluation of cooked turkey meat was carried out investigating physical characteristics (texture, colour, shrinkage), water status and sensorial properties. The description of the water status with different parameters such as moisture content, water holding capacity and 1H NMR mobility resulted a very valuable tool for a better understanding of physical characteristics and sensorial properties of cooked meat, that were closely related to the different denaturation of protein induced by the cooking process. Results indicated that different steam levels (relative humidities) led to different meat cooking performances and cooked meat quality. The application of steam in cooking allows for shorter cooking times than dry oven cooking when the temperature of the surface of meat results sufficiently higher than the temperature of the centre. Low steam cooking, characterized by shorter cooking times than those with no steam, resulted in higher cooking yield and greater perceived tenderness of meat than high steam cooking (close to the saturation condition). The minimum relative humidity values allowing positive effects on meat quality and higher potential water saving at low cooking temperatures (80-150°C), were defined. The use of low steam quantities at low cooking temperatures could be considered as a valid alternative to saturation cooking, particularly indicated for industrial applications, due to the increased quality of turkey meat and the potential water saving. Fri, 31 Dec 2010 23:00:00 GMT http://hdl.handle.net/1889/1675 2010-12-31T23:00:00Z http://hdl.handle.net/1889/1674 Title: Highly proteolized novel and traditional foods Authors: Lambertini, Francesca Abstract: The present Ph.D. Thesis concerns the molecular characterization of highly proteolyzed foods, with particular attention to the influence on their biological and functional properties exerted by oligopeptides and amino acids. Different kinds of foods containing low molecular weight nitrogen compounds have been considered: traditional food products derived from a natural proteolytic processes, , and in particular Parmigiano-Reggiano cheese and Culatello sausages, and novel foods obtained by a proteolytic digestion through an innovative biocatalytic processes, considering in particular proteic hydrolyzates obtained from the recovery of poultry industry leftovers. The nitrogen fractions of the proteinaceous hydrolyzates extracted from the different food matrices, both traditional and produced according to novel technologies, have been extensively studied by means of mass spectrometry coupled with liquid chromatography. This technique has been demonstrated to be a valid approach to monitor the composition of complex peptide mixtures, because of its high sensibility and selectivity. In particular a new LC/MS/MS methods to quantify free and total amino acids without any derivatization step has been developed and validated, and improved LC/MS methodologies have been set up also for the analysis of other nitrogen components Cheese is a natural product with a potential high biological value. Proteolysis occurring during ageing time leads to the release of a large number of peptides from caseins, which have been demonstrated to be biologically active by in vitro studies. The composition of the oligopeptide fraction of different Parmigiano-Reggiano cheeses at different ageing times has been extensively studied in a very detailed way by means of LC/MS methodologies. Antioxidant and antihypertensive properties of these peptide fractions have been studied. Moreover, since data on the effective bioavailability of these peptides are still quite scarce, the work of the present PhD. thesis also addressed the study of their bioaccessibility through simulated gastrointestinal digestion. Low molecular weight nitrogen compounds in Culatello sausages, one of the most famous Italian dry pork meat products, have also been studied. For the first time data on the amount of characteristic’s meat compounds, such as carnosine, anserine, creatine and creatinine, in cured meat are reported and their amount was found to be strictly related to the technological production process. Finally, free and total amino acid analysis of the biosynthetic proteinaceous hydrolyzates derived from poultry leftovers have shown that this potential novel foods posses an high nutritional value, due to the high content of bioavailable essential amino acids. These proteic hydrolyzates showed interesting antioxidant and antihypertensive properties. Amino acidic composition, and overall oligopeptide fractions, have been found to be strictly related to the proteolytic conditions used for food production, thus bioactive compound amounts may be regulated on the basis of the desired features. For example, samples with the highest hydrolysis degree may be used as functional food ingredients in virtue of their high antioxidant capacity. In conclusion, the present thesis demonstrated that proteolyzed foods can be considered high quality food and food ingredients, either if traditional food obtained by natural proteolytic processes or novel foods obtained through innovative technologies. In the latter case, it has been demonstrated that leftover of the food industry can be transformed in new food and feed not only having a high nutritional value, but also possessing biofunctional properties. Thu, 31 Mar 2011 22:00:00 GMT http://hdl.handle.net/1889/1674 2011-03-31T22:00:00Z http://hdl.handle.net/1889/1673 Title: Strumenti per l'innovazione di processo nel settore dei formaggi a pasta cotta: image analysis delle dimensioni dei granuli di cagliata e predizione delle cinetiche di scambio termico per raffreddamento convettivo della forma tramite analisi FEM Authors: Iezzi, Romina Abstract: Il settore lattiero caseario ricopre il 12 % dell’intero fatturato dell’industria alimentare italiana e vale quasi il 10 % dell’export alimentare italiano. Grana Padano e Parmigiano Reggiano hanno alti prezzi di vendita rispetto ad altri formaggi considerati ormai “commodities” del mercato alimentare; ciò dimostra le difficoltà che tali prodotti devono superare per operare nel mercato e, quindi, evidenzia la necessità di acquisire strumenti di conoscenza che possano tradursi in strumenti operativi utili per ridurre i costi di produzione, senza pagare un prezzo eccessivo in termini di qualità e nel rispetto dei dettami previsti dal regolamento CE 510/2006. Fra i diversi aspetti che girano attorno al binomio costi produttivi-qualità, due possono assumere un ruolo importante per tutto il comparto caseario: lo studio delle dimensioni dei granuli di cagliata, come condizione per ottimizzare le operazioni di taglio del coagulo, e la previsione delle cinetiche di raffreddamento della cagliata in fascera in funzione di dimensioni, forma e condizioni esterne. Per il primo obiettivo è stata impiegata l’image analysis bidimensionale per determinare le dimensioni dei granuli di cagliata, dopo le fasi di rottura e di cottura, provenienti da 3 caseifici di Parmigiano Reggiano e Grana Padano per un totale di 93 campioni. L'analisi ha permesso di evidenziare che l'area proiettata media dei granuli di cagliata risultanti dalla spinatura è pari a 2,80±4,10 mm2, e che la cottura riduce di circa il 38,6 % tale area, portandola a 1,72±2,12 mm2. L'analisi statistica descrittiva ha evidenziato una distribuzione non simmetrica delle dimensioni attorno al valore medio e valori importanti di differenza tra le dimensioni maggiori e minori dei granuli, differenze queste anche molto variabili a seconda dei caseifici. Inoltre lo studio della distribuzione delle dimensioni dei granuli (subito dopo rottura e cottura) in classi di frequenza progressivamente crescente da 0,1 a 24 mm2 ha evidenziato, come prevedibile, la maggiore numerosità dei granuli di ridotte dimensioni, ma anche come questi granuli ricoprano un valore ridotto dell'intera area proiettata. Considerando infine che il numero di granuli non dovrebbe cambiare in misura importante in seguito all'agitazione durante la cottura, la riduzione del valore di area proiettata potrebbe essere un indice della quantità di siero espulso. È stata, infatti, osservata una concordanza nel valore medio tra il rapporto di contenuto di solidi totali TS di CCG e FCG ed i valori di riduzione di area. L'approccio tramite analisi d'immagine ha dimostrato la fattibilità della misura e potrà consentire di razionalizzare il processo di taglio in modo da ridurre la variabilità delle dimensioni dei granuli. Per il secondo obiettivo l’approccio predittivo, anziché quello empirico di conoscenza delle temperature di raffreddamento, ha portato allo sviluppo di un modello matematico che, tramite l'analisi numerica, ha permesso di simulare gli effetti dello scambio termico (raffreddamento convettivo naturale) di forme cilindriche di formaggio in funzione delle variabili di interesse. Per aumentare la precisione della predizione, il modello ha incluso le funzioni temperatura-dipendenti (k e cp) dell'alimento, le caratteristiche termo-fisiche degli stampi di contenimento ed i parametri dell'aria circostante. Il confronto tra i risultati delle simulazioni ed i dati sperimentali è stato eseguito mediante il calcolo del valore di RMSE. I valori di RMSE ottenuti per i campioni di formaggio (valore medio massimo di 1,23 °C), caratterizzati da temperature iniziali, dimensioni e composizioni differenti, hanno consentito di validare l'attendibilità del modello proposto, quando le assunzioni che lo caratterizzano vengono rispettate. Nello stesso tempo hanno permesso anche di valutare, tramite la validazione delle curve predette riferite ai campioni di cagliata, quale potrebbe invece essere l'ordine di grandezza dell'errore nel caso in cui le assunzioni esposte non siano completamente rispettate. La validazione del modello ha, altre sì, permesso di valutare l'impatto che la scelta dei materiali costituenti lo stampo può avere sulla velocità di raffreddamento della cagliata. L'accuratezza del modello, soprattutto quando sono rispettati i principali assunti, ne potrà permettere l'applicazione in tutti quei settori in cui i fenomeni temperatura-dipendenti influenzano il raggiungimento e/o il mantenimento dei requisiti di qualità, riducendo la necessità di ripetere sperimentazioni onerose. L'integrazione di questo modello con i modelli di previsione della crescita microbica potrà rappresentare un passo avanti negli studi di previsione della crescita microbica stessa in matrici alimentari reali.; The dairy sector covers 12 % of the entire turnover of the Italian food industry and it is worth 10 % of the Italian food export. Selling prices of Grana Padano and Parmigiano Reggiano are higher than those of the cheeses that are considered “commodities” in the food market; this fact explains the difficulties that these products need to overcome in order to operate in the market and, therefore, it proves the necessity to acquire a new knowledge allowing for a reduction in the production costs, both safeguarding cheese quality and the dictates of CE 510/2006 Regulation. Among the various aspects regarding the binomial production cost-quality, two of them can assume an important role for the dairy products: the study of the dimensions of cheese curd grains, as a condition to optimize the cut operations of the coagulum, and the prediction of heat exchange during cheese curd cooling in the mould according to its dimensions, shape and external conditions. The size of curd grains after cutting and cooking were measured by means of bi-dimensional image analysis; 93 samples from 3 factories of Parmigiano Reggiano and Grana Padano were collected. Image analysis showed that the mean projected area of curd grains was 2.80±4.10 mm2 after cutting, while this area was 1.72±2.12 mm2 with a reduction of 38.6% at the end of cooking. The statistic analysis evidenced a not symmetrical distribution of the dimensions around the mean value and a high difference between the greater and smaller size of grains. Moreover the study of the distribution of the size of grains (immediately after cutting and cooking) in frequency classes, progressively increasing from 0.1 to 24 mm2, showed that the number of grains of small dimension was higher, but also that these grains covered a reduced part of the total projected area. Considering that the grain number did not change during stirring and cooking, the reduction of the projected area could be considered an index of the amount of expelled whey. The mean value of the ratio between the total solids of cooked grains and cutting grains and the mean reduction of area were also in agreement. Nevertheless the image analysis demonstrated the feasibility of this approach which may allow to improve the efficiency of milk coagulum cut, and to reduce the variability of cheese curd grains. A mathematical model was developed and used, through the numerical analysis, in order to simulate the effects of the heat exchange (free convection cooling) of cylindrical cheese. The model included cheese temperature-functions (k and cp), the thermo-physical characteristics of the mould and the parameters of the surrounding air. The comparison between the result of the simulations and the experimental data was performed by calculating the RMSE values. The values of RMSE obtained for the cheese samples (the maximum mean value was 1.23 °C), which were characterized by different initial temperatures, dimensions and compositions, confirmed the proposed model when the model assumptions were respected. At the same time, RMSE values allowed to estimate, through the curd samples, the possible error of the prediction if the assumptions were not fully respected. The model validation also allowed to estimate the impact of the mould materials on the cheese cooling rate. The accuracy of the proposed model makes it functional to be applied to all the fields where the temperature phenomena influence the achievement and/or the maintenance of quality requirement, reducing the need of onerous experiments. The heat exchange model can be integrated with a microbial growth prediction model, representing a step forward in the studies of microbial growth prediction in real food matrixes. Fri, 31 Dec 2010 23:00:00 GMT http://hdl.handle.net/1889/1673 2010-12-31T23:00:00Z http://hdl.handle.net/1889/1671 Title: Role of Lactobacillus rhamnosus in Parmigiano Reggiano cheese ripening: a genotypic and post-genomic study Authors: Bove, Claudio Giorgio Abstract: Lactic Acid Bacteria (LAB) constitute an heterogeneous group of bacteria that are traditionally used to produce fermented foods. They may play different roles in cheese-making where starter LAB (SLAB) participate in the fermentation process, whereas non starter LAB (NSLAB) are implicated in the maturation of cheese. Although the role of NSLAB in ripening has not yet been clarified, different authors have suggested their importance in the cheese ageing. In Parmigiano Reggiano (PR) cheese, NSLAB, autochthonic of raw milk and arising from the environment, are the protagonists of the different biochemical processes during the production and ripening stages. Recent studies in microbiological ecology of PR cheese allowed a deeper insight of the microbial composition during the manufacturing and ripening stages of the same cheese-making process. Further, the study of samples representative of the subsequent stages of the same cheese-making by culture-independent methods led to determine the microbial succession during 24 months of PR ripening. It was demonstrated that SLAB are dominant until the 2nd month of ripening. Differently, after cheese brining, the species NSLAB, especially Lb. rhamnosus, are able to grow and increase in number, while SLAB cells undergo to autolysis. Lb. rhamnosus was shown to be the dominant species present after a lack of essential nutrients, such as sugars. Therefore, this species seems to well adapt to the absence of lactose in cheese, confirming an optimal adaptability to unfavorable growth conditions. Presumably, this characteristic was due to the ability of Lb. rhamnosus to use nitrogen fraction as an alternative energy source. So far, only few studies are available about NSLAB, and in particular about Lb. rhamnosus, in PR and the exact role of these bacteria has not been investigated in greater depth. In particular, the biotypes of the dominant species during the maturation stage of cheese processing have never been studied. Indeed, it can only be hypothesized that the technological pressure determines, at different stages, the potential development of the biotype that may have a specific biochemical role leading to certain flavors and sensorial traits of the PR cheese. For this reason, Lb. rhamnosus, the dominant species among NSLAB in PR, has been chosen as subject for the present study. This Ph.D thesis aims to better understand the role played by this species in cheese-making and ripening process, and to hypothesize the strategies used to adapt its energy metabolism to the environmental conditions met in PR cheese. To reach this issue, three different steps has been pursued. In the first step, genotypic relatedness at the intraspecies level for 66 Lb. rhamnosus strains isolated during the same cycle of PR cheesemaking, was determined. Biodiversity has been estimated by means of two DNA fingerprinting techniques, i.e., randomly amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) and repetitive extragenic palindromic-polymerase chain reaction REP-PCR analysis. It was found an intraspecies heterogeneity in Lb. rhamnosus strains which is certainly correlated to their abilities to adapt to specific environmental and technological conditions. In this regard, the detection of biotypes that mark specific steps in cheese ripening, or of those that can develop differently throughout the ripening process, suggests that they may have also specific roles closely linked to their peculiar metabolical properties. In the second step of the research the metabolic data (fermentative profiles, synthesis of organic acids and FAA) and 2-DE analyses followed by the identification of a large number of proteins were combined to highlight Lb. rhamnosus physiological mechanisms of adaptation that are responsible for survival in cheese during ripening. Cultivation was carried out under cheese-like conditions (Cheese broth) and on pasteurized and micro-filtrated milk and MRS broth As shown by fermentative profiles analysis, the major part of the Lb. rhamnosus strains had the capacity to use numerous chemical compounds as energy sources. Growth and acidification differed depending on the culture media. Compared to growth on MRS or pasteurized and micro-filtrated milk, all strains cultivated in cheese-like medium (cheese broth, CB) showed a decrease of the synthesis of D,L-lactic acid and synthesized higher levels of acetic acid. Except for one strain, the others caused an increase of the concentration of free amino acids during cultivation on CB. A proteomic approach was applied to elucidate the (i) intra-specific diversity of Lb. rhamnosus and (ii) the metabolic mechanisms involved in growth in CB which mimicked the cheese during ripening. The proteomic maps of five strains showing different metabolic traits were comparatively determined after growth on MRS and CB. The result showed that synthesis of ca. 60 - 70% of the cytosolic proteins of Lb. rhamnosus strains was not affected by the culture media. The remaining part (ca. 30-40%) of the total proteome seemed to vary depending on strains and growth media. A total of 93 protein spots which showed an increased level of synthesis during growth in MRS (46 spots) or CB (47 spot) were identified by MALDI-time of flight mass spectrometry (MALDI-TOF-MS/MS) and nano-electrospray ionization-ion trap mass spectrometry (ESI-MS/MS). Compared to cells grown in MRS, Lb. rhamnosus strains cultivated under cheese-like conditions modified the synthesis of proteins related to protein biosynthesis, nucleotide and carbohydrate metabolisms, glycolysis pathway, proteolytic activity, cell wall and exopolysaccharide biosyntheses, cellular regulation, amino acid and citrate metabolisms, oxidation/reduction processes and stress response. It was confirmed that Lb. rhamnosus strains have the capacity to activate different metabolic pathways depending on the culture media. In the last step an innovative cDNA-AFLP protocol was developed to investigate the changes in gene expression profile during the growth of one Lb. rhamnosus strain isolated in 20 month ripened PR and therefore able to growth and survive in this hostile environment. Changes in gene expression was evaluated in CB versus a rich medium (MRS). This study represents one of the few concerning bacterial transcriptomic analysis through cDNA-AFLP approaches. This technique allowed to generate unique transcript tags from reverse-transcribed messenger RNA using restriction enzymes and selective PCR amplification. Results evidenced that Lb. rhamnosus is able to modify the expression of a large part of genes when cultivated in cheese-like conditions (CB) compared to growth under in vitro optimal conditions (MRS). However, these genes still have to be recognized thus it is not possible at the moment to correlate them with proteins found with the proteomic study. For this reason further analysis will have to be focused on gel extraction followed by the identification of expressed genes. This will allow to deepen the knowledge of the basal metabolism through the identification of constitutive genes, as well as to analyze the reactions of the cells to the dairy environment by the identification of differently expressed genes in the two cultured media. Moreover the expected results could lead to the discovery of Lb. rhamnosus genes putatively involved both in the physiological mechanisms of adaptation to cheese-like substrates, that are fundamental for survival during ripening, and in the definition of organoleptic characteristics of the PR cheese. Fri, 31 Dec 2010 23:00:00 GMT http://hdl.handle.net/1889/1671 2010-12-31T23:00:00Z http://hdl.handle.net/1889/1665 Title: Plant foods and cardiovascular health: looking for mechanisms of action Authors: Jechiu, Lucia Abstract: Given the importance of vegetable foods in the prevention of CVDs and in the maintenance of vascular health, this PhD researcher project was aimed to investigate the role of specific vegetable components, such as betaine, choline and polyphenol metabolites on different mechanisms putatively related to CVD health. Betaine and choline are two methyl donors that convert homocysteine to methionine while polyphenols diminish the oxidative stress. Specifically, the work has been focused on the evaluation of the role of betaine and choline on different inflammatory markers and on the role of phenolic metabolites on the modulation of NADPH oxidase activity, the key enzyme of oxidative stress through its role in reactive oxygen species production. In particular, the research work addressed two questions: 1) whether betaine and choline intake from the diet are related to inflammation, and 2) what is the effect of metabolites derived from berries and coffee polyphenols on the reduction of superoxide anion production in a cell line suitable as an in vitro model for atheroma. The two research lines hereby described resulted into two research papers. Survey on betaine and choline content in food products consumed in Italy, dietary intake and relation with traditional and emerging CVD risk factors. We have determined the content of betaine and free choline in products commonly consumed in Italy in order to estimate the daily intake in the Italian population and to assess the association with markers of inflammation and components of the metabolic syndrome in a cross-sectional study of a healthy population living in Northern Italy. Betaine and free choline content was measured by high-pressure liquid chromatography – tandem mass spectrometry (HPLC MS/MS) in 28 common cereal products mainly consumed in Italy such as pasta, bread and other bakery products and in 23 vegetables such as spinach, beet and others. Under the context of the cross-sectional survey that enrolled 469 healthy subjects living in the Parma area, data on clinical biochemistry and functional anthropometry were gathered and the correlation of betaine and choline with inflammatory marker was studied. Information on diet was collected via 3-day dietary questionnaires. The outcomes of this survey revealed a high content of betaine and free choline in wholemeal foods; betaine was also found at high concentrations in beetroot and spinach. In population of this study, betaine intake showed a positive correlation with component of metabolic syndrome and inflammatory biomarker C-reactive protein, an independent risk factor of cardiovascular event, while dietary choline showed no association. Superoxide anion production in cultured human promonocyte cells THP-1 due to polyphenol metabolites. In the in vitro study we have investigated the effect of different colonic metabolites derived from berries and coffee polyphenols on the modulation of NADPH oxidase-dependent superoxide anion production in monocytes THP-1. The molecules were tested at the low concentrations actually found in the bloodstream after absorption on a cell line implicated in atheroma after PMA stimulation. The bioluminescence technique was employed for the assessment of O2.- production. The results obtained in our in vitro study demonstrated that polyphenol catabolites of colonic origin are not capable to reduce the oxidative stress through modulation of NADPH system. Taken altogether, these data seem to indicate that the protection exerted by dietary sources of these classes of molecules, such as whole grain cereals, fruits and vegetables, is not directly related to the compounds assessed in this study, or at least, is not specifically related to the mechanisms investigated. Further studies are warranted in order to elucidate the role of betaine and choline, as well as the role of phenolic metabolites, in the beneficial effects of whole grain cereals, fruits and vegetables on risk of cardiovascular disease and atherosclerosis. Fri, 31 Dec 2010 23:00:00 GMT http://hdl.handle.net/1889/1665 2010-12-31T23:00:00Z