Tesi sul tema "Cheese"

The cheese making process at Anchor Products Hautapu was having difficulty reaching the now outdated New Zealand Dairy Board uniformity targets. Even though there is no longer a direct financial benefit in reaching these targets consistently, there is the benefit of being able to show the customer that the product which they are receiving is as consistent as possible. This project was carried out by systematically investigating each section of the cheese making process, looking for variations that were likely to affect the final product. Where variations were found methods for eliminating them or minimising their effect on the final product were developed. The largest source of variation was found to be caused by fluctuations in the curd depths on the belts of the Alf-O-Matic cheddaring machine. Overlapping the ends of consecutive cheese making tanks as well as the re-calculations of the pump out flow rates have been proposed to remedy this problem. Where the curd is drawn off from the end of the Alf-O-Matic cheddaring machine was also found to be causing variation in the product due to particle stratification affecting the salt levels of blocks that were being produced. Recommendations for methods to reduce the level of stratification have been suggested including using a capacitance probe to control the curd level. Further variations were occurring within the cheese making tanks with cutting and stirring speeds differing from tank to tank. The tanks were also being flushed with cold water causing moisture spikes in the product. Both of these problems have been eliminated by changes to the PLC program. Small improvements have been seen in the process with the changes that have already been carried out. Large improvements are expected if the rest of the recommendations are implemented. The largest improvements should be seen with the realisation of an overlapped pump out system.

Tableau d'honneur de la Faculté des études supérieures et postdoctorales, 2016-2017
Le fromage Mozzarella entre dans la composition de plusieurs mets populaires d’Amérique du Nord. L’aptitude de ce fromage à être râpé et ses propriétés caractéristiques de cuisson en font un ingrédient idéal. Ces qualités sont attribuées principalement aux propriétés physiques particulières de ce fromage sous certaines conditions de cisaillement et de température. Le but de ce projet était d’évaluer l’impact de différentes stratégies couramment mises en oeuvre dans l’industrie fromagère sur la composition, la microstructure et les propriétés physiques du fromage. Diverses stratégies ont été étudiées : les conditions de filage du caillé lors du procédé de « pasta filata », l’addition de protéines sériques dénaturées, le contrôle de la minéralisation et le vieillissement du fromage. Les résultats ont démontré que le contrôle de l’intensité mécanique et thermique fournie lors du filage permettait respectivement de réduire les pertes de solides et d’améliorer la répartition de la phase aqueuse dans la matrice fromagère. L’aptitude au râpage du fromage peut être optimisée en combinant l’utilisation de plusieurs stratégies dont la réduction du calcium colloïdal, un temps de vieillissement adéquat et un râpage à basse température. Par ailleurs, des changements aux facteurs mentionnés précédemment sont apportés lors de l’ajout de protéines sériques dénaturées, ces dernières ayant un impact sur la composition et la structure du fromage. Des modèles prédictifs de l’aptitude au râpage ont été développés en sélectionnant uniquement les descripteurs de composition et de texture pertinents. La perception sensorielle du fromage cuit sur pizza et les propriétés physiques du fromage fondu ont été considérablement influencées par l'évolution physico-chimique du fromage au cours du vieillissement. L’utilisation d’une nouvelle approche pour la caractérisation des propriétés rhéologiques du fromage fondu sous fortes contraintes a permis d’établir de bonnes relations avec les descripteurs sensoriels de texture. Ce travail a permis de valider l’hypothèse que l’utilisation d’une ou plusieurs stratégies simples et accessibles pouvait être mise de l’avant afin d’optimiser les propriétés physiques du fromage Mozzarella. Cela contribue à une meilleure compréhension des facteurs pouvant être contrôlés afin de développer des fromages avec des attributs spécifiques, lorsqu’utilisés comme ingrédient.
Mozzarella cheese is expected to perform various key attributes when used as a food ingredient. The shreddability and the melting properties of cheese during and after baking are mainly governed by the physical properties of cheese when subjected to external factors such as shear and temperature. Therefore, the goal of this project was to evaluate the impact of cheese-making strategies commonly used in the dairy processing industry on the cheese composition, microstructure, and physical properties. Various strategies were studied: pasta filata process conditions, addition of denatured whey protein (WP-D) to milk, control of curd mineralization, and cheese aging. Results showed that controlling the mechanical and thermal intensity during the pasta filata process can lead to reduced cheese solid losses and a better distribution of water within cheese microstructure, respectively. The ability of cheese to be shredded can be increased using a combination of multiple factors such as lowering colloidal calcium phosphate associated with casein, proper aging, and by reducing cheese temperature before shredding. However, an optimisation of the previous factors should be done if WP-D is added because of its impact on cheese composition and structure. Predictive models to assess cheese shreddability were built using only few relevant compositional and textural descriptors. Sensory perception of baked cheese texture and physical properties of melted cheese were dramatically influenced by the physico-chemical evolution of cheese during aging. Melted cheese texture was satisfactorily related to different sensory attributes using a novel approach to determine the rheological properties under the large stress experienced during mastication. This work validated the hypothesis that simple cheese-making strategies, alone or combined, can be used to optimize the cheese physical properties. This contributes to a better understanding of the factors that can be controlled to improve or develop cheese ingredient with specific attributes.

The effect of reducing the fat content of low-moisture part-skim Mozzarella cheese from 19% to less than 5% on melted cheese properties, i.e., apparent viscosity, cheese melt, and cook color, was investigated. Functional properties of melt and stretch and cook color were evaluated at d 1, 7, 14, and 28. A rapid microwave oven method underestimated the moisture content of the low fat cheeses by approximately 10%. Low fat cheese did not melt as well as did the low-moisture part-skim Mozzarella cheese although the moisture content of the low fat cheese (moisture content ranged from 62.5% to 63.6%) was greater than the moisture content of the part-skim control (52.1 %). Storage for 28 d only marginally increased the meltability of low fat cheese. Lower fat content increased cook color. The amount of intact a αs-CN decreased by at least 48% in all cheeses as a result of proteolysis during 28 d of storage. The relative proportion of bound, entrapped, and expressible water was determined for a reduced-fat (8% fat) and control (19% fat) Mozzarella cheese on d 1, 7, 14, and 21 of refrigerated storage. Changes in the state of water were related to changes in cheese microstructure of a commercial Mozzarella cheese and to changes in cheese meltability of the control cheese. The amount of expressible water was proportional to fat content. Throughout storage, fat/serum channels became smaller and the protein matrix expanded into the areas between fat globules. The meltability of both cheeses increased during storage. Both cheeses contained 0.71g bound water/ g protein. Expressible water decreased in both cheeses until by d 21 no water was expressible. Entrapped moisture increased from approximately 10% to 60% for the control cheese and from approximately 33% to 50% for the reduced-fat cheese. An objective test was developed for measuring stretch, a characteristic of melted cheese. Three nonfat and four low-moisture part-skim cheeses were evaluated using the new test and the results compared with conventional test methods. Two new melted cheese parameters were defined: melt strength, the maximum load (g) obtained during the test, and stretch quality, the average load (g) as the cheese fibers stretched and elongated. Melt strength correlated with apparent viscosity. Stretch quality was determined for selected nonfat and low-moisture part-skim cheeses. A three-pronged probe was used to pull cheese vertically from a melted cheese pool. Use of this elongation stretch test, along with more traditional melted cheese tests, provides more complete information about the functional properties of Mozzarella cheese.

The use of slower acid-producing starter bacteria for the production of lower fat Cheddar cheese has lead to milder flavor Cheddar cheeses that lack intense Cheddar notes. The metabolism of methionine leads to the production of methanethiol, which is one of the desirable Cheddar cheese flavor compounds. The influence of NaCl and reduced pH was determined for aminopeptidase, lipase/ esterase, and methanethiol-producing capability in selected lactic acid bacteria and brevibacteria in simulated cheese-like conditions. The activity of each enzyme decreased with NaCl addition and pH reduction to approximate a Cheddar cheese environment (5% NaCl and pH 5.2). The mechanism for methanethiol production by the starter and adjunct bacteria was also investigated. Different enzyme systems were found to be responsible for methanethiol production in starter lactococci, lactobacilli, and brevibacteria. In the lactococci, enzymes that acted primarily on cystathionine were responsible for methanethiol production from methionine. Lactobacilli also contained cystathionine-degrading enzymes, but these enzymes have properties different from the lactococcal enzymes. Brevibacterium linensBL2 lacked cystathionine-degrading enzymes, but was capable of the direct conversion of methionine to methanethiol. L-Methionine γ-lyase from B. linens BL2 was purified to homogeneity, and was found to catalyze the α, γ elimination of methionine resulting in the production of methanethiol, α-ketobutyrate, and ammonia. Characterization of the pure enzyme demonstrated that it is pyridoxal phosphate dependent, which is active at salt and pH conditions existing in ripening Cheddar cheese. The addition of either B. linens BL2 or L-methionine γ-lyase to aseptic cheese curd slurries increased methanethiol and total volatile sulfur compound production. In an attempt to increase methanethiol production and Cheddar cheese flavor in reduced-fat Cheddar cheese, B. linens BL2 was added as a starter adjunct to 60% reduced-fat cheese. Sensory evaluation of the cheese indicated that B. linens BL2 improved the flavor of 60% reduced-fat Cheddar cheese. This suggests that the addition of B. linens BL2 is an alternative to the addition of lactic acid bacteria to improve Cheddar cheese flavor via the metabolism of methionine.

Various techniques for accelerating mature flavour development in Cheddar cheese were compared. Control cheese ( c) was manufactured by using Streptococcus cremor is AM2, a starter used in normal commercial manufacture. A combination of S. cremoris AM2 and Streptococcus lactis C2 Lac- Prt- mutant was used in the manufacture of test cheeses (M). §._. lactis C2 mutant was grown in glucose broth at 30°c and pH 6 .O for 16 hours, followed by concentration and diafiltration to 1011cfu mL - 1 using microfiltration equipment. The control cheesemilk was inoculated to 6x107 streptococci pe mL with S. cremoris AM2 and the mutant vat cheesemilk to 2x109 per mL with a combination of ~ cremoris AM2 and ~ lactis C2 mutant. The starter population in cheese containing mutant starter was 100 times that in control cheese (1010 compared to 10a). Cheeses were also made with added bacterial neutral proteinase (Neutrase, N) and stored at a0c (a) and 15°c ( 15) for 32 weeks. This resulted in cheese being subjected to the following treatments: ca (control), C15, CNa, CN15, Ma·, M15, MNa, and MN15. Cheddaring times were slightly reduced and milling acidities slightly higher in the vat . containing mutant starter. However the composition of all cheese was satisfactory. Bacteriological counts, proteolysis, rheological properties and flavour development of these cheeses were monitored at regular intervals throughout maturation. The order of the effectiveness of the treatment in accelerating ripening was MN15, >M15,> CN15,> C15,> MNa,> Ma,> CNa,> ca. Cheeses from these treatments attained the characteristics of control cheese stored at a0 c (Ca) for 6 months after 1.4, 1.7, 2.0, 2.6, 2.8, 3.2, 4.3 and 6.0 months respectively. Cheese quality was not adversely affected except for bitterness in CN8 cheese and overmaturity in CN15 cheese late in the storage period. The possible mechanisms and relative merits of the various treatments are discussed with special reference to an "active role" theory of starter bacteria in flavour development.

El objetivo general del presente TAF es el estudio del lanzamiento de un nuevo producto de fast food en el centro comercial Jockey Plaza, el mismo que se ofrece como un snack salado hecho de una masa crocante rellena con queso mozarella (ingrediente preponderante) y un ingrediente adicional Se plantean como objetivos específicos, analizar el entorno competitivo de empresas que comercializan productos similares o sustitutos con el fin de plantear estrategias de participación de mercado

The aim of this PhD thesis was the study of the genomic, biological and phenotypic background of bovine cheese-related traits. The primary goal of this PhD thesis was to unravel the genomic background of bovine milk technological and cheese-related traits to specific chromosomic regions (CHAPTERS 1 to 3). To achieve this, the cow’s ability to produce cheese was decomposed into 11 milk coagulation (MCP) and curd-firming properties (CFt), and 7 cheese yield and milk component recoveries into the curd (REC) traits. Besides, to tackle the problem of the large number of variables required to describe the cow’s ability to produce cheese, posing restrictions in the construction of selection indices, and thereby selection decisions, factor analysis (FA) was used (CHAPTERS 4 and 5). The MCP traits were: 3 traditional single point lacto-dynamographic properties (RCT: rennet coagulation time, min; k20: time to a curd firmness (CF) of 20 mm, min; a30: CF 30 min after rennet addition), 6 parameters modeling 360 CF data for each milk sample (CFP: potential asymptotic CF at infinite time, mm; kCF: curd firming instant rate constant, %×min-1; kSR: syneresis instant rate constant, %×min-1; RCTeq: RCT from modeling; CFmax: maximum CF, mm; tmax: time at CFmax, min), milk- protein (%) and pH. The 3 CY traits were the weight (wt) of fresh curd (%CYCURD), curd solids (%CYSOLIDS), and curd moisture (%CYWATER) as % of wt of milk processed. The 4 REC (RECFAT, RECPROTEIN, RECSOLIDS, and RECENERGY) were calculated as the % ratio between the nutrient in curd and the corresponding nutrient in processed milk. For FA 26 traits related to milk yield and quality (including milk protein fractions), MCP-CFt and CY-REC traits were analyzed. Single marker genome-wide association analyses (GWAS) complemented by gene-set enrichment and pathway-based analyses were conducted. In total, 1,152 Italian Brown Swiss cows reared in 85 herds were genotyped with the Illumina SNP50 Beadchip v.2. Single marker regression GWAS were fitted using the GenABEL R package (GRAMMAR-GC). The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway databases were queried for the enrichment analyses. In GWAS (CHAPTERS 1 and 2), sharp peaks were detected on Bos taurus autosome (BTA) 6, at 84 to 88 Mbp, with the highest peak detected at 87.4 Mbp in the region harboring the casein genes and more precisely of κ-CN (CSN3). Marker Hapmap52348-rs29024684 (~87.4 Mbp), closely located to the casein genes on BTA6, was strongly associated with RECFAT (P = 1.91×10-15) and CFP (P = 1.62×10-17). Evidence of quantitative trait loci at 82.6 and 88.4 Mbp on the same chromosome was found. On BTA11, marker ARS-BFGL-NGS-104610 (~104.3 Mbp) was highly associated with RECPROTEIN (P = 6.07×10-36). Apart from BTA6 and 11, SNP located in 15 more chromosomes (1, 2, 9, 12, 13, 14, 15, 16, 18, 19, 20, 23, 26, 27 and 28) were significantly associated to the MCP-CFt and CY-REC traits. The gene-set enrichment and pathway-based analysis (CHAPTER 3) revealed 21 GO and 17 KEGG categories significantly associated (false discovery rate controlled at 5%) with 7 of the traits (RCT, RCTeq, kCF, %CYSOLIDS, RECFAT, RECSOLIDS and RECENERGY), with some being in common between traits. The significantly enriched categories included calcium signaling pathway, salivary secretion, metabolic pathways, carbohydrate digestion and absorption, the tight junction and the phosphatidylinositol pathways, as well as pathways related to the bovine mammary gland health status, and contained a total of 150 genes located in all chromosomes but 9, 20, and 27. In FA (CHAPTERS 4 and 5), ten mutual orthogonal Fs were obtained using a varimax rotation. The 10 Fs explaining 74% of the original variability. Those Fs captured basic concepts of the “milk to cheese” process. More precisely, the first four Fs, sorted by variance explained, were able to capture the underlying structure of the CY percentage (F1%CY), the CF process with time (F2CFt), the milk and solids yield (F3Yield) and the presence of nitrogen (N) into the cheese (F4Cheese N). Moreover, 4 Fs (F5 αs1-β-CN, F7β-κ-CN, F8αs2-CN, F9αs1-CN-P) were related to the basic milk caseins (as1-CN, as2-CN, β-CN, κ-CN, and the phosphorylated form of as1-CN) and 1 factor was associated with the α-LA whey protein (F10α-LA). A factor describing the udder health status of a cow (F6Udder health), mainly loaded on lactose, other nitrogen compounds and SCS, was also obtained. In general, FA results were coherent to the given name of the factor. Stage of lactation had a significant effect for the majority of the Fs, followed by parity. Moreover, considerable genetic correlations existed among the Fs (CHAPTER 4). All Fs showed significant associations (P < 5 ×10-5) in GWAS, but F5Yield. High peaks on BTA6 (~87Mbp) and at the tail of BTA11 (~104Mbp) were mainly associated to F6β-κ-CN and F1Cheese N, respectively. In addition, 33 GO terms and 6 KEGG categories were mainly enriched for F8αs2-CN, but also for F1%CY, F4Cheese N, and F10α-LA. Biological pathways were mainly related to the broader categories of ion activity, neurons and the tight junction. Moreover, the considerably large number of enriched GO and KEGG terms for F8αs2-CN suggests that, perhaps, more focus should be given on αs2-CN (CHAPTER 5).
Lo scopo di questa tesi di dottorato è stato lo studio del background genomico, biologico e fenotipico di caratteri legati al processo di caseificazione nella specie bovina. L’obiettivo primario è stato quello di determinare il background genomico di caratteri tecnologici del latte bovino legati al processo di caseificazione (CAPITOLI da 1 a 3). Per raggiungere questo obiettivo, l’abilità della bovina di produrre formaggio è stata ripartita in 26 fenotipi: 11 caratteri di attitudine casearia e proprietà di coagulazione, comprendenti le tradizionali proprietà di coagulazione del latte (MCP) e nuovi parametri modellizzati di consistenza della cagliata (CFt), e 7 fenotipi di resa in formaggio (CY) e recupero dei nutrienti del latte nella cagliata (REC). Tuttavia, l'elevato numero di variabili necessarie per descrivere la produzione di formaggio bovino pone delle restrizioni nella costruzione di indici di selezione, e quindi nel prendere decisioni di selezione. Per superare il problema della elevata dimensionalità, è stata utilizzata un’analisi fattoriale (FA) per studiare la struttura latente dei 26 caratteri coinvolti nel processo di caseificazione (CAPITOLI 4 e 5). I caratteri MCP includevano le 3 proprietà lattodinamografiche tradizionali basate su singola misurazione dello strumento (RCT: tempo di coagulazione, in min; k20: tempo di rassodamento, in min; a30: consistenza del coagulo (CF) 30 min dopo l'aggiunta del caglio, in mm). I fenotipi CFt comprendevano un set di 6 parametri modellizzati sulla base di 360 dati di CF misurati per ciascun campione di latte (CFp: CF potenziale, in mm; kCF: tasso di rassodamento del coagulo, in % × min-1; kSR: tasso di sineresi, in % × min-1; RCTeq: RCT stimato dal modello; CFmax: massima CF, in mm; tmax: tempo necessario per raggiungere CFmax, in min), delle proteine del latte (%) e del pH. I 3 caratteri CY includevano resa a fresco (% CYCURD), resa in solidi (% CYSOLIDS), e acqua ritenuta nella cagliata (% CYWATER), espresse come percentuale del latte trasformato. Le 4 misure di REC (RECFAT, RECPROTEIN, RECSOLIDS, e RECENERGY) sono state calcolate come rapporto espresso in percentuale tra il valore di nutrienti nella cagliata e il corrispettivo nel latte. L’analisi FA ha considerato tutti i 26 caratteri oggetti di studio, comprendenti produzione e qualità del latte (incluse le frazioni proteiche del latte), parametri CFt e caratteri CY-REC. La metodologia adottata comprendeva analisi di associazione genome-wide (GWAS), accompagnata da analisi di arricchimento genetico e di tipo pathway-based. Le analisi genomiche hanno considerato un totale, 1.152 bovine di razza Bruna Italiana allevate in 85 allevamenti, genotipizzate attraverso il v.2 Illumina SNP50 Beadchip. Le analisi GWAS sono state condotte mediante analisi di regressione a singolo marcatore, fittate utilizzando il pacchetto GenABEL del software R (GRAMMAR-GC). I database Gene Ontology (GO) e Kyoto Encyclopedia of Genes and Genomes (KEGG) sono stati interrogati per le analisi di arricchimento. Nell’analisi GWAS (CAPITOLI 1 e 2) sono stati individuati picchi nitidi sull’autosoma 6 di Bos taurus (BTA) tra 84-88 Mbp, con il picco più alto rilevato a 87,4 Mbp nella regione ospitante i geni della caseina e più precisamente della κ-CN (CSN3). Il marcatore Hapmap52348-rs29024684 (~ 87,4 Mbp), localizzato in prossimità dei geni della caseina su BTA6, ha mostrato una forte associazione con RECFAT (P = 1.91 × 10-15) e CFP (P = 1.62 × 10-17). Sullo stesso cromosoma, è stata trovata evidenza di loci per i caratteri quantitativi a 82,6 e 88,4 Mbp. Su BTA11, il marcatore ARS-BFGL-NGS-104.610 (~ 104,3 Mbp) è risultato fortemente associato con RECPROTEIN (P = 6,07 × 10-36). Oltre a BTA6 e 11, altri SNP situati in altri 15 cromosomi (1, 2, 9, 12, 13, 14, 15, 16, 18, 19, 20, 23, 26, 27 e 28) sono risultati significativamente associati con MCP, CFt e con i caratteri CY-REC. L'analisi di arricchimento e pathway-based (CAPITOLO 3) ha rivelato 21 categorie GO e 17 categorie KEGG significativamente associate (tasso di errore controllato al 5%) con 7 tra i caratteri fenotipici considerati (RCT, RCTeq, kCF, %CYSOLIDS, RECFAT, RECSOLIDS e RECENERGY) e alcune categorie sono risultate in comune tra i caratteri. Le categorie significativamente arricchite includevano vie di segnalazione del calcio, di secrezione salivare, vie metaboliche, di digestione e assorbimento dei carboidrati, di giunzioni occludenti e del fosfatidilinositolo, così come vie legate allo stato di salute della ghiandola mammaria bovina, per un totale di 150 geni situati in tutti i cromosomi tranne 9, 20 e 27. Nella FA (CAPITOLI 4 e 5) sono stati ottenuti dieci Fs mutualmente ortogonali utilizzando una rotazione varimax. I 10 Fs spiegavano il 74% della variabilità originale. Tali Fs erano biologicamente riconducibili a elementi base del processo di trasformazione “dal latte al formaggio". Più precisamente, i primi 4 Fs, ordinati sulla base della varianza spiegata, sono stati in grado di definire la struttura latente della CY percentuale (F1% CY), del processo di CF nel tempo (F2CFt), del rendimento di latte e solidi (F3Yield) e della presenza di azoto (N) nel formaggio (F4Cheese N). Inoltre, 4 Fs (F5αs1-β-CN, F7β-κ-CN, F8αs2-CN, F9αs1-CN-P) erano associati alle caseine del latte (as1-CN, as2-CN, β-CN, κ-CN, e la forma fosforilata as1-CN) e 1 fattore alla proteina del siero α-LA (F10α-LA). É stato inoltre ottenuto un fattore in grado di descrivere lo stato di salute della mammella bovina (F6Udder health), basato principalmente sulla produzione di lattosio e di altri composti azotati e sulle cellule somatiche. In generale, i risultati nell’analisi FA sono risultati coerenti con l’attribuzione del significato biologico dato al fattore. La maggior parte degli Fs è risultata significativamente influenzata dallo stadio di lattazione, seguito dall’ordine di parto. Sono state inoltre riscontrate correlazioni genetiche rilevanti tra i fattori (CAPITOLO 4). Nell’analisi GWAS tutti gli Fs hanno mostrato associazioni significative (P <5 × 10-5), ad eccezione di F5Yield. I picchi elevati su BTA6 (~ 87Mbp) e sulla coda di BTA11 (~ 104Mbp) erano principalmente associati a F6β-κ-CN e F1Cheese N, rispettivamente. Inoltre, 33 termini GO e 6 categorie KEGG sono risultati arricchiti e associati con F1% CY, F4Cheese N, F8αs2-CN e F10α-LA. Le vie di segnalazione biologica descritte dai fattori erano principalmente correlate alle categorie più generali di attività ionica, neuroni e giunzioni occludenti. Poichè un numero considerevole di categorie arricchite GO e KEGG è risultato associato al fattore F8αs2-CN, maggiore attenzione dovrebbe essere posta sulla frazioneαs2-CN (CAPITOLO 5).

In this study, production of organic acids from cheese-whey was studied. Optimization of organic acids production was performed in semi-batch and batch reactors. Two sets of experiments were performed. First set of experiments were performed in semi-batch reactors for the optimization of organic loading rate (OLR) and hydraulic retention time (HRT). As a result of Set 1 experiments optimum OLR was found to be 15 g COD l-1. Second set of experiments were performed in batch reactors by using the optimum OLR found in Set 1 experiments. Set 2 experiments were conducted to study the effect of using different seed cultures and Basal Media (BM) on Volatile fatty avid (VFA) production. Main acidogenesis products were acetic acid (Hac), butyric acid (Buty) and propionic acid (HPr) with smaller quantities of i-butyric acid (i-Buty), valeric acid (Val) and caproic acid (Cap). It was seen that BM had a suppressive effect on ethanol (EtOH) production while it stimulated the VFA production. Higher VFA productions and variety of VFA types were observed in Test Reactors seeded with acidogenic culture (R3 and R6).

Taken as an engineering material, mozzarella cheese can be considered as a complex food system that has dynamic structure and complex flowproperties. Food scientists have been actively developing methods to characterize mozzarella cheese rheologically, but most of these methods are empirical in nature. In the past decades, there has been a paradigm shift towards the utilization of well-developed rheological methods which have been widely applied in the study of commercial synthetic polymers. In this work, the rheology of mozzarella cheese was studied using well-developed rheological techniques. Utilizing various rheometers, the linear and non-linear rheology of mozzarella cheese was examined. General practical properties of mozzarella cheese such as meltability, flowability and stretchability were extracted from these results. Capillary flow and rolling experiments were also performed to determine their suitability as innovative post-production processing techniques for mozzarella cheese. Finally, a comparative study on the effect of frozen storage on the rheology of three different brands of mozzarella cheese was performed. In general, it was found that mozzarella cheese can be classified as a pseudoplastic (shear thinning) semi-solid material possessing a yield stress at room temperature. Upon heating, the yield stress gradually diminishes and it can be considered as a viscoelastic fluid. The results obtained from the various rheometers indicate that the yield stress, duration of experiment, sample geometry and temperature greatly affect the consistency of the results. It was also shown that extrusion can be used as a processing technique for mozzarella cheese above a certain temperature where the cheese is in a melt state. Rolling was also found to be a potentially feasible processing method. Finally, in terms of the effect of frozen storage, in general, the dynamic moduli decrease with the period of storage due to the freezing of the proteins in the cheese.

Blue Stilton is a blue-veined cheese manufactured in a restricted area of the UK, using lactic starter cultures plus a secondary culture of Penicillium roquefotti. The aim of this study was to determine the change in microflora during ripening of the cheese and to investigate potential microbial interactions. Additionally, the volatile compounds present in mature samples of cheeses exhibiting few blue veins were compared with those in good quality cheeses, showing ample blue veining. Experiments on cheeses from a single dairy, monitored during the ripening process, showed that the total Lactobacillus count increased from levels of around 103 cfu g-1 on day one to around 107 cfu g-1 after 8 weeks of ripening. This is comparable to values found in other cheeses including both mould-ripened and non mould-ripened varieties. Yeast counts were generally higher than those found in other cheeses and also increased to levels in the region of 107 g-1. The total viable count (TVC) decreased from around 109 g-1 initially, reflecting the presence of the starter bacteria, to 107 g.1, suggesting a decline in the starter bacteria similar to that found in other cheeses. Mature cheeses always exhibited similar numbers of microorganisms although the species varied between cheeses. High quality, mature, cheeses were compared with sub-standard cheeses from the same production site. The predominant species of lactobacilli in good quality cheeses were Lb. plantarum and Lb. curvatus, whereas in poor quality cheeses Lb. brevis predominated. This corresponded to the results of gas chromatography-olfactometry, which indicated the presence of fruity off flavours in poor quality cheeses. Several strains of these species were isolated, as indicated by differing capabilities in utilisation of a range of carbon sources. Yeast species also varied between good and poor quality cheeses with Candida sphaerlca and C. catenulata predominating in good cheeses and C. famata, C. lipolytica and C. catenulata also occurring in both good and poor quality samples. Strain differences were observed by the biochemical profiles and two strains of C. famata demonstrated inhibitory effects against P. roqueforti when incubated under anaerobiosis. It was concluded that these strains may affect the development of blue veins in Stilton cheese when maturation conditions encourage their proliferation. Comparisons were made between samples of cheeses from several Stilton producers and the results suggested that although the levels of the groups of microorganisms tested were similar, the species of lactobacilli and yeasts present were different. This suggests that the indigenous microflora may have a significant impact on the flavour of cheeses from individual production sites. It was concluded that the microflora of Blue Stilton cheese may have a significant impact on the quality of the product both in terms of flavour and the development of the blue veins.

Mestrado em Biotecnologia - Biotecnologia Alimentar
Over the last years there has been an increasing interest to replace synthetic materials by biodegradable ones, due to the ecological problems. Edible and biodegradable films can be produced using polysaccharides, lipids, proteins and composites, and act as a package without damaging the environment. By choosing a suitable coating composition it is possible to preserve several desired properties of a certain food product. Important properties should be considered, such as mechanical, functional and barrier properties. The main goal of this study was to evaluate edible films and coatings from plant proteins (pea, soy), with incorporated natural antimicrobial and antioxidant agents, to potentially protect cheese from physico-chemical and microbial deterioration and to preserve the organoleptic characteristics, especially of sliced cheeses. The work performed focused mainly on the preparation and characterization of pea protein films, with added chitooligosaccharides (COs) (0.5%, 1% and 2%) and two types of essential oils at 1%, bay and thyme oils. Films with 0.5% of COs showed the highest values of Young’s modulus, tensile strength and elongation. Regarding the barrier properties, the film with 1% of COs showed the lower permeability value. Addition of small amounts of COs may be advantageous to improve the mechanical properties of the PPI films, besides the expected antimicrobial effects. An intermediate COs concentration (1%) could be advantageous to reduce the water vapor permeability, but it will also result in detrimental effects on the mechanical properties. Film’s hydrophobicity was also dependent on the amount of added COs and essential oils. For the films with COs, the presence of the essential oils increased the film’s hydrophobicity, an effect dependent on the type of added oil. The observed effects seem complex and they are probably dependent on the interactions among film components; certainly these aspects deserve further studies in order to improve and better understand the interactions/adhesion of the coating onto the cheese surface. The protein films by their own showed already some antioxidant activity, and the addition of COs or the essential oils results mainly on a higher rate of this effect (lower times to observe the antioxidant effects). Even so the films prepared with the bay oil revealed a higher antioxidant activity, which can be useful and complement the expected effects on the organoleptic properties of cheese samples treated with these films.
O interesse na substituição de materiais sintéticos por biodegradáveis tem vindo a aumentar devido aos problemas ecológicos. Filmes comestíveis e biodegradáveis podem ser produzidos utilizando polissacarídeos, lípidos, proteínas e compósitos e atuar como embalagens, sem danificar o meio ambiente. Ao escolher uma composição adequada para um revestimento é possível preservar várias propriedades desejáveis de um produto alimentar. Propriedades importantes como mecânicas, funcionais e de barreira devem ser consideradas. O principal objetivo deste estudo foi avaliar filmes e revestimentos comestíveis de proteínas vegetais (ervilha, soja), com agentes antimicrobianos e antioxidantes naturais incorporados, para proteger queijo de deterioração físico-química e microbiana e para preservar as características organolépticas, especialmente de queijos cortados/fatiados. O trabalho realizado foca-se principalmente na preparação e caracterização de filmes de proteína de ervilha, com a adição de oligoquitosanos (OQ) (0,5%, 1% e 2%) e dois tipos de óleos essenciais (1%), óleos de louro e tomilho. Filmes com 0,5% de OQs apresentaram valores mais elevados de módulo de Young, tensão de rutura e alongamento. Em relação às propriedades de barreira, o filme com 1% de OQs mostrou o valor de permeabilidade mais baixa. A adição de pequenas quantidades de OQs pode ser vantajosa para melhorar as propriedades mecânicas dos filmes de proteína de ervilha, além dos esperados efeitos antimicrobianos. Uma concentração OQs intermediária (1%) poderia ser vantajosa para reduzir a permeabilidade ao vapor de água, mas também resultaria em efeitos prejudiciais sobre as propriedades mecânicas. A hidrofobicidade dos filmes foi dependente da quantidade de OQs e óleos essenciais adicionados. Para os filmes com OQs, a presença dos óleos essenciais aumentou a hidrofobicidade dos filmes, um efeito dependente do tipo de óleo adicionado. Os efeitos observados parecem complexos e provavelmente dependem das interações entre os diferentes componentes do filme; Estes aspetos merecem mais estudos a fim de melhorar e compreender as interações / aderência do revestimento sobre a superfície do queijo. Os filmes de proteínas por si só mostraram alguma atividade antioxidante, e os resultados da adição de OQs ou óleos essenciais mostram uma taxa mais elevada deste efeito (diminuição do tempo de reação para observar os efeitos antioxidantes). Os filmes com óleo de louro revelaram uma maior atividade antioxidante, podendo ser útil e complementar aos efeitos esperados sobre as propriedades organolépticas de amostras de queijo revestidas.

Modern, large scale Cheddar cheese manufacture is dependent on reliable acid production by Lactococcus lactis subspecies cremoris and subspecies lactis starter cultures. Any inhibition of acid production may affect cheese quality, disrupt production schedules and reduce profitability. The presence of antibiotic residues in manufacturing milk resulting from the treatment of mastitis in lactating cattle is a potential source of starter culture inhibition. Therefore, a range of antibiotic concentrations was assessed for measurable inhibitory effects on acid production and compared to the minimum detectable concentrations by approved screening test procedures. Antibiotics were selected from formulations approved for use on lactating cattle for the treatment of mastitis. Novobiocin, lincomycin, oleandomycin and oxytetracyline HCl, all non-b-lactam antibiotics, inhibited acid production of one or more L. lactis strains at antibiotic concentrations below the detectable limit of standard screening procedures.
Depending on the antibiotic, either or both the Bacillus stearothermophilus (var. calidolactis) disk assay and/or the Delvo SP assay were ineffective at detecting the antibiotics at concentrations required to inhibit the starter strains. Consequently, antibiotic residues below the detectable limits of these testing procedures could cause significant starter culture inhibition, disrupting cheese making schedules. Another potential source of starter culture inhibition is related to raw milk quality and the practice of refrigerated storage prior to processing. Previous studies differed as to whether the growth of psychrotrophic organisms would have a detrimental impact on subsequent acid production by starter bacteria employed in cheese manufacture. In this study, no inhibition of acid production by a commercial L. lactis subsp. cremoris strain was evident when grown in milk that had undergone short term temperature abuse. Antimicrobial systems native to bovine milk may also have an adverse impact on starter culture performance. The present study assessed the inhibitory effect of an activated lactoperoxidase system (LPS) on a range of L. lactis cultures. All of the strains were significantly inhibited when grown on reconstituted skim milk in the presence of an active LPS. Inhibition of acid production by strains grown on glucose was also observed, leading to further investigations to describe the inhibitory process. A non-phosphoenolpyruvate phosphotransferase (PEP/PTS) dependent glucose transport system, first observed in 1980 in one L. lactis subsp. lactis strain, was hypothesised as a link in strain variations in LPS sensitivity. However, the LPS sensitive L. lactis subsp. cremoris strains tested did not take up glucose in a PEP depleted state, most likely due to their inability to utilise arginine as an ATP generating energy source. The questions remain unanswered whether cremoris strains possess this glucose transport mechanism and whether it could contribute to strain variations in LPS sensitivity.
In a subsequent investigation, galactose phosphotransferase system (PTS) deficient L. lactis strain ATCC 7962 demonstrated log phase growth inhibition when grown on galactose in the presence of the model LPS. Previously reported LPS mediated effects on the glycolytic enzyme hexokinase do not appear to explain this result. The present study confirmed strain variability in sensitivity to the model LPS among both Lactococcus lactis subspecies lactis and subspecies cremoris strains. Further, the observation that dithiothreitol significantly alleviated the inhibition of a highly sensitive cremoris strain, implicated the involvement of sulphydryl groups as the target of the transient inhibitory factors. Data collected excluded the possibility that portions of the metabolic pathways involved in fructose and galactose metabolism are sensitive to the LPS in cells possessing PEP/PTS capability. This study also identified potential directions of further work to elucidate the mechanism(s) of LPS inhibition.

In the blue cheese Stilton the starter mould Penicillium roqueforti grows and sporulates during the ripening period and is considered to be responsible for the unique blue cheese aroma. However, the sporulation of the mould, which results in the formation of blue veins, takes place in a fraction of the Stilton matrix which overall is very heterogeneous. Most blue cheeses develop a secondary microflora of yeasts which may affect their aroma. The aim of this study was to investigate the yeast flora of Stilton, the aroma profile of the cheese and the role of the yeasts in the aroma production. The approach in this work was to study individually the different sections of Stilton (the blue veins, the white core and the outer crust) as previous studies have demonstrated each section has a differing bacterial flora. In addition to the classical microbiology, a series of molecular techniques (Denaturing Gradient Gel Electrophoresis, Restriction Fragment Length Polymorphism and Terminal RFLP) were compared and applied for the screening of the local fungal communities in the cheese. The results showed that the two approaches were complementary. It was concluded that the structure of the fungal community was different for each section of the cheese. The aroma profiles of the three different sections of Stilton were studied using solvent extraction Gas Chromatography-Mass Spectrometry (GC-MS), a headspace GC-MS technique (SPME GC-MS) and direct headspace analysis (Atmospheric Pressure Chemical Ionisation [APCI]-MS). The different sections of Stilton presented different aroma profiles. Overall, the blue and the outer crust had similar profiles. These two sections contained higher amount of ketones while the white contained higher amounts of alcohols and aldehydes. Yeast isolates and the starter Penicillium roqueforti were cultivated alone and in combination in a cheese model and the aroma production was studied with SPME GC-MS analysis. The co-culture of the starter Penicillium roqueforti and individual yeast isolates resulted in aroma profiles different from those that were produced by the mould or the yeasts individually. The model of Penicillium roqueforti with Yarrowia lipolytica resulted in an aroma more similar to blue cheese than produced by the mould alone. Sensory analysis (Flash profile technique) was used in order to compare the aroma of this model with the aroma of blue cheeses and the perception of the combined culture was found to be similar to Stilton cheese, whereas that of the mould alone was not. Yeasts are a significant part of the microflora of Stilton and they are able to affect the aroma production. Selected isolates of Yarrowia lipolytica could be used in combination with Penicillium roqueforti for the production of blue cheese aroma e.g. as a starter culture.

Dissertation 'consists of a selection of 71 publications produced over the period 1975-2001, which were drawn from the more complete list of 198 original research papers, review articles and books published over the same period' -- declaration.
Thesis (PhD Food Sc )--Stellenbosch University, 2001.
ENGLISH ABSTRACT: One of the curious facts about the food industry is that many of the processes in use today were being practised, in some form or other, by the Roman legions as they marched across Europe and beyond. Certainly they were familiar with the basic techniques of fermentation, and much current research into fermented foods is concerned with understanding the fundamental nature of these traditional processes, and how the individual stages in a particular fermentation can be better controlled. Recent developments in the dairy industry have tended to reflect this pattern and, over the years, my research group has done much to support the expanding markets for yoghurt and similar fermented milks. Our evaluation of the polysaccharide-producing characteristics of starter cultures, for example, encouraged yoghurt manufacturers to match physical properties to the perceived demands of consumers, and most culture suppliers followed this lead by labelling their products with precise designations as to their potential for imparting viscosity to a retail item. Similarly, my group was the first to record the unique physical properties of the concentrated yoghurt, labneh, C 230 g 1-1 total solids) that had been made for hundreds of years by draining whey from natural yoghurt hanging in a cloth or animal-skin bag. This detailed analysis of the product facilitated the application of ultra-filtration to natural yoghurt to generate a product with a quality that matched traditionallabneh and, today, factories in the Middle East, Greece and elsewhere are using modern membrane-filtration plants to satisfy a growing market demand. Our success in publicising the attractive properties of concentrated yoghurt encouraged me to devote time to yet another 'historical' concept, namely the apparent 'health benefits' derived by small communities in Eastern Europe from consuming kefir and koumiss. In the West, the flavour and texture of these latter products have never been accepted, but employing similar cultures to produce 'health-promoting' bio-yoghurts opened an entirely new avenue for research. As clinical evidence in support of the prophylactic and therapeutic properties of Lactobacillus acidophilus and a species of Bifidobacterium became available, so it became apparent that the therapeutic advantage that accompanies the regular ingestion of 'bio-yoghurts' depended on the survival of these microfloras over the stipulated shelf-lives of the retail vehicles. However, no laboratory medium was immediately available for the simultaneous enumeration of Lb. acidophilus and Bifidobacterium along with the yoghurt cultures, i.e. Streptococcus thermophilus and Lactobacillus delbrueckii sub-sp. bulgaricus. Designing such a medium became a priority for one of my students, and, even today, the procedures that he derived are being used by consumer groups that monitor the performance of the major dairy companies in England. If the improved quality of yoghurts and 'bio-yoghurts' had a major impact on consumer perceptions of fermented milks, the food sector in England gradually became aware of an even more dramatic change in consumer attitudes. Thus twenty years ago, cheese meant 'Cheddar' but, following a 'deluge' of television publicity about the attractions of 'exotic' catering, housewives began demanding mozzarella and mascarpone for lavish desserts, Feta to sprinkle over salads and Halloumi to grill or fry. In turn, exporting countries like Italy, Greece and Cyprus came under intense pressure to increase supplies of top quality products. Local manufacturers soon realised, however, that there was little information available concerning the scientific basis to the procedures employed to make some of these traditional cheeses, and my research group was selected by Funding Agencies in Greece and Cyprus to act as a focus for a series of studies of Feta and Halloumi cheese. The need to eliminate pathogens from the storage brines of Feta cheese without killing the yeasts and bacteria associated with maturation became an important consideration for exporters, and one of my students exploited a novel procedure employing furocoumarins and long-wave ultra-violet light to achieve the desired selective inactivation. At present, the economics of commercial application are somewhat dubious but, as soon as cheap, synthetic, non-toxic furocoumarins become more readily available, the system may well merit re-evaluation. We did confirm, however, that the metabolic activities of the yeasts and bacteria typically isolated from storage brines are essential for flavour development in Feta cheese, and that similar microfloras are instrumental in the development of the important charactistics of traditional Halloumi cheese. In particular, a new species of lactic acid bacterium, Lactobacillus cypricasei, was isolated from samples of the traditional ovine cheese, but whether or not the species has a unique role(s) in the maturation process remains an open question. Clearly there is still much to learn but, if the activities of my reseach group have added just a little to the scientific background essential for future studies of cheese and fermented milks, then their completion will have been worthwhile.
AFRIKAANSE OPSOMMING: Een van die merkwaardigste feite omtrent die voedselindustrie is dat baie van die prosesse wat vandag gebruik word, in een of ander vorm deur die Romeinse magte gebruik is toe hulle deur Europa marsjeer het. Basiese fermentasie tegnieke was aan hulle bekend, en heelwat huidige navorsing oor gefermenteerde voedsel is gemik daarop om die fundamentele natuur van hierdie tradisionele prosesse te verstaan en hoe die individuele stappe in 'n spesifieke fermentasie beter beheer kan word. Onlangse ontwikkelinge in die suiwelindustrie reflekteer hierdie patroon, en my navorsingsgroep het oor die jare heelwat gedoen om die groeiende markte vir joghurt en soortgelyke gefermenteerde melk te ondersteun. Ons evaluasie van die polisakkariedproduserende eienskappe van suursels het byvoorbeeld joghurtvervaardigers gehelp om fisieke eienskappe daar te stel wat verbruikers tevrede sal stel. Meeste verskaffers van kulture het hierdie voorbeeld gevolg deur hul produkte so te etiketteer dat duidelik gewys word watter potensiaal dit het om viskositeit aan die finale produk te verleen. Verder was my groep die eerste om die unieke fisiese eienskappe van die gekonsentreerde joghurt, labneh (230 g 1-1 totale vastestowwe) te bepaal, wat vir honderde jare gemaak is deur die wei van natuurlike joghurt te dreineer deur dit in "n materiaal- of diervelsak te hang. "n Gedetailleerde analise van hierdie tradisionele produk het bygedra tot die gebruik van ultrafiltrasie op natuurlike joghurt om "n produk te gee met dieselfde kwaliteit as tradisionele labneh. Vandag gebruik fabrieke in die Midde Ooste, Griekeland en elders moderne membraan-filtrasie aanlegte om in die groeiende vraag na die produk te voorsien. Ons sukses met die bekendmaking van die aantreklike eienskappe van gekonsentreerde joghurt het my aangespoor om tyd te spandeer aan nog 'n sogenaamde "historiese" konsep, naamlik die skynbare gesondheidsvoordele van klein gemeenskappe in Oos-Europa wat kefir en koumiss verbruik. In die Weste is die smaak en tekstuur van hierdie produkte nooit werklik aanvaar nie, maar om soortgelyke kulture te gebruik om "gesondheidsbevorderende" bio-joghurt te produseer, het 'n hele nuwe navorsingsveld daargestel. Soos kliniese bewyse van die terapeutiese en voorkomende voordele van Lactobacillus acidophilus en 'n spesie van Bifidobacterium bekend gemaak is, het dit duidelik geword dat die terapeutiese voordele wat saamgaan met die gereelde inname van "bio-joghurts", afhang van die oorlewing van hierdie mikroflora oor die gestipuleerde rakleeftyd van die kommersiële produkte. Geen laboratorium medium was egter onmiddellik beskikbaar vir die gelyktydige telling van Lb. actdophilus en Bifldobacterium tesame met die joghurt kulture Streptococcus thermophilus en Lb. delbrueckii sub-sp. bulgaricus. Die ontwikkeling van so 'n medium het een van my studente se prioriteit geword, en selfs vandag word die prosedures wat deur hom ontwikkel is, gebruik deur verbruikersgroepe wat die optrede van groot suiwelmaatskappye in Engeland monitor. lndien die verbeterde kwaliteit van joghurts en bio-joghurts 'n groot impak gehad het op verbruikers se persepsie van gefermenteerde melk oor Wes-Europa heen, het die voedselsektor in Engeland bewus geraak van selfs 'n meer dramatiese verandering in verbruikers se houding. Twintig jaar terug het kaas "Cheddar" beteken, maar na 'n stortvloed televisie advertensies oor die aanloklikheid van eksotiese geregte, het daar by huisvroue 'n vraag ontstaan na Mozzarella en Mascarpone vir nageregte, Feta oor slaai en Halloumi om te bak of te braai. Italië, Griekeland en Siprus wat hierdie produkte uitgevoer het, het onder kwaai druk gekom om groter hoeveelhede, top-kwaliteit produkte te lewer. Plaaslike vervaardigers het gou agtergekom dat min inligting beskikbaar was oor die wetenskaplike basis van die prosedures wat gebruik word om hierdie tradisionele kase te maak en my navorsingsgroep is deur befondsingsagentskappe in Griekeland en Siprus genader om studies te doen oor sekere aspekte van die vervaardiging van Feta en Halloumi kaas. Dit het vir beide in- en uitvoerders belangrik geword om die patogene te elimineer uit die soutoplossing waarin Fetakaas gestoor word, sonder om die giste en bakterieë wat rypwording aanhelp, te dood. Een van my studente het 'n innoverende prosedure ontwikkel wat furocoumarins en lang-golf-ultra-violet lig gebruik om selektiewe inaktivering te kry. Op die oomblik is daar effense onsekerheid oor die ekonomiese implikasies van die kommersiële toepassing, maar sodra goedkoop, sintetiese, nie-toksiese furocoumarins geredelik beskikbaar word, moet die sisteem weer geëvalueer word. Ons het egter bevestig dat die metaboliese aktiwiteite van die giste en bakterieë in die stooroplossing noodsaaklik is vir geurontwikkeling in Feta kaas en dat soortgelyke mikrofloras instrumenteel is in die ontwikkeling van die belangrike karaktereienskappe van tradisionele Halloumi kaas. 'n Unieke melksuurbakterium, Lactobacillus cypricasei, is uit monsters tradisionele skaap Halloumi geïsoleer, maar of hierdie spesie 'n unieke rol speel in die verouderingsproses is nog 'n ope vraag. Duidelik is daar nog baie om te leer, maar indien my navorsingsgroep se aktiwiteite slegs 'n klein bydrae gemaak het tot die wetenskaplike agtergrond wat essensieel is vir toekomstige navorsing, was die voltooiing daarvan die moeite werd.

International Telemetering Conference Proceedings / October 21, 2002 / Town & Country Hotel and Conference Center, San Diego, California
For the last 30 years Magnetic Tape Systems have been the primary means of recording data from airborne instrumentation systems. Increasing data rates and harsh environmental requirements have often exceeded the ability of tape-based systems to keep pace with platform technology. This paper examines operational and data reduction benefits when employing the IRIG 106 Chapter 10 Solid State Recorder Standard introduced by the Range Commanders Council (RCC) Telemetry Group (TG). The Standard and this paper address media formatting, data formatting for a variety of different data types, data downloading, and data security, along with serial command and control and discrete command and control of the recorder. This paper also addresses software data processing and raw data reconstruction of Chapter 10 data.

The characteristics (chemical, microbiological and organoleptic) of commercial fresh and mature Halloumi cheeses were examined. In addition, the effect of the feeding regime of lactating animals on the flavour of fresh Halloumi cheeses was also assessed. Three different brands of Halloumi cheese (industrial, Traditional 1 and Traditional 2), based on milk origin and location of the dairies, were selected for subsequent analysis. The industrial cheese was made with bovine milk as the major ingredient (an insignificant mixture of ovine/caprine milk might be included), while the traditional cheeses were primarily manufactured with ovine milk (a small amount of caprine milk might be included). The traditional cheeses also differed in the location of the dairies, in that Traditional 1 cheese was manufactured in the province of Paphos while Traditional 2 cheese was made in Nicosia province. Differences were revealed both between the types of cheese and age, i.e. fresh versus mature. The characteristics that differed included the microbiological load of fresh samples, the type and quantity of flavour compounds, and the rate of proteolysis. Some of the volatile compounds present in Traditional 1 cheese probably originated from plants present in the grazing plains (thyme and burnet), while the mint added during manufacture contributed, almost exclusively, to the plant volatile compounds that were present in Traditional 2 cheese. The sensory analysis identified significant differences between the three brands. Moreover, the panels (young and older) assessed the cheeses in a different way, reflecting a probable effect of panel age on sensory testing. During the microbiological analysis of the mature traditional Halloumi cheeses a new Lactobacillus species was recovered, which was named Lactobacillus cypricasei. In order to improve the organoleptic characteristics of bovine Halloumi cheese, two 'natural starter cultures' were isolated from raw milks and incorporated into bovine milk for the manufacture of cheese in the laboratory. One selected starter culture, Lactobacillus rhamnosus, was used to ripen the bovine milk prior to Halloumi cheese manufacture, but its activity failed to bring about the anticipated results. The degradation of protein and fat was monitored throughout the maturation of the cheese, and only minor differences were observed between 'experimental' (starter culture) and 'control' (no starter culture) Halloumi cheeses. The sensory analysis of fresh and mature Halloumi cheeses reflected the results of the instrumental analysis, for the panel did not detect any significant differences between 'experimental' and 'control' cheeses. In the light of the high frequency of occurrence of Enterococcus faecium in the fresh samples of traditional commercial samples and ovine milks, it was decided to assess the survival of the micro-organism under normal cheesemaking conditions. Hence, Halloumi cheese with a starter culture of E. faecium was produced in the laboratory. The results showed that some cells of E. faecium survived the harsh cooking treatment (90°CI 1 h) of the cheese blocks during Halloumi cheese manufacture. The experiments were not completed due to the controversy about using Enterococcus spp. in cheesemaking. Overall, the analysis of commercial and laboratory cheeses has given an insight into the special characteristics of Halloumi cheese, and provided with a better understanding of how flavour and texture develops with cheese maturation.

Dissertation presented in partial fulfillment of the requirements for the degree of Master in Biotechnology
Polyhydroxyalkanoates (PHAs) are polyesters produced by a variety of microorganisms. Due to the similarity of chemical and physical properties with the conventional plastics, and full biodegradability, PHAs constitute one of the best alternatives for synthetic polymers replacement. However, the production costs of these biopolymers are very high when compared to synthetic polymers production. One way to reduce the production costs is the utilization of low cost raw materials, such as industrial wastes and by-products as carbon source. An example of raw material is cheese whey, a by-product form cheese industry rich in lactose (4-5%). In this work, cheese whey was supplied to Escherichia coli strains harbouring the PHB synthesis genes from Cupriavidus necator for the production of poly(3hydroxybutyrate) (PHB). During this study, diverse reactor operating strategies were tested: feeding controlled by pH under oxygen limitation, feeding without oxygen limitation and continuous feeding. The best results were achieved in a fed-batch system with feeding controlled by pH and oxygen limitation, where 44.93% of PHB content,33.76 g/L of PHB concentration, 78.65 g/L of active biomass concentration and a volumetric productivity of 0.57 gPHB/L.h, were obtained.

A new method for manufacture of natural cheese was developed using 5X ultrafiltered whole milk retentate. The retentate was prefermented to pH 5.0-5.2 before curd formation to simplify the cheese making process. It was demonstrated that the process can be made commercially feasible. Ultrafiltration and diafiltration of whole milk were done so that the desired level of residual lactose was left in the retentate. Retentate was inoculated with lactic starter culture and incubated (prefermented) until all lactose was converted to lactic acid. The final pH stabilized at about 5.0-5.2; the desired pH of the cheese curd. Incubation at 25°C allowed the retentate to remain liquid during prefermentation and easily pumped through pipes. After prefermentation, retentate was passed through a mechanical curd former where rennet was injected and mixed. The retentate-rennet mixture coagulated as it traveled upward through a cylinder. The curd emerged from the curd former and was cut into cubes by a grid of knives. The curd cubes were treated for removal of moisture by cooking in liquid and evaporation under vacuum. After moisture removal, curd was drained. salted and pressed. Cheddaring and milling were unnecessary. Experiments were performed to determine proper methods for preparing retentate. Diafiltration level was significantly related to lactose concentration in retentate (p=.OOO 1) and final pH of fermented retentate (p=.OOO1). Acidified permeate and acidified, deionized water were evaluated as cooking liquids. Cheese made from curd cooked in permeate had acid defects. but curd cooked in water produced cheese with a pH similar to conventionally produced Cheddar cheese. Moisture content of all finished cheese was excessive for Cheddar cheese. Body and texture of cheese made from curd cooked in water was close to conventionally produced Cheddar cheese. Diafiltration may be used to control final pH of fermented retentate. Prefermentation of retentate prior to cheese making will simplify equipment and shorten manufacturing time. Commercial application of the process is discussed.

Variables affecting the physical properties of Mozzarella cheese were investigated. The effects of various milk-clotting enzymes were examined. The type of milk coagulating enzyme used played a significant role in determining physical properties of direct acid Mozzarella cheese. Cook color was not affected by enzyme type, but melt and stretch were significantly affected. Proteolytic nature of starter cultures was reviewed and recommendation s were give n. Cheese made with proteinase-deficient strains had more stretch after holding for 14 and 28 d than cheese made with non-deficient strains. Cheese made with pairs or single strains of L. helveticus had the same melt, more stretch, and less cook color than cheeses made with paired strains of Lactobacillus delbrueckii ssp. bulgaricus and Streptococcus salivarius ssp. thermophilus. Frozen storage, thawing, and shredding of Mozzarella cheese were described and suggestions given for optimizing shelf life. Shredding, freeze temperature, thaw temperature , and time of storage had no effect on cook color. Frozen, shredded cheese stretched more and melted less than non-shredded frozen cheese. Reduced fat, high moisture Mozzarella cheese was made and found acceptable when compared to low moisture part-skim Mozzarella cheese. Reduced fat cheeses decreased in stretch and increased in melt throughout storage. Differences in stretch, melt, and cook color were not significant from one casein-to-fat ratio to another. Reduced fat, high moisture Mozzarella cheese was made with partial or total replacement of L. helveticus with L. casei ssp. casei and was found to compare well with low moisture part-skim Mozzarella cheese. Cheese made with L. casei ssp. casei cultures, paired with either S. salivarius ssp. thermophilus and L. helveticus or just S. salivarius ssp. thermophilus, had the least stretch and the greatest melt.

The lactic acid fermentation of cheese whey permeate by Lactobacillus helveticus was studied. Precipitate formation during autoclaving of whey permeate was examined. Precipitation was found to be pH and temperature dependent. Qualitative analysis suggested that the precipitate was a calcium-phosphate complex. Solubilisation was achieved both by acidification and use of the sequestering agent EDTA. Optimisation of L. helveticus growth in whey permeate was carried out using factorial design, as opposed to a traditional univariate approach. Using this technique, the variation of specific growth rate with pH, temperature and stiirer speed was assessed. Cell growth and lactic acid formation in whey permeate containing various supplements, were investigated. Yeast extract was the most effective nitrogen/growth factor supplement. Maximum lactic acid production was achieved in permeate containing yeast extract (0.75% w/v), Tween 80 (0.1% v/v) and sodium acetate (0.05% w/v). Optimisation of lactic acid production in supplemented whey permeate was performed using factorial design. Optimum conditions for both acid formation and cell growth were pH 5.9, temperature 42°C and stirrer speed 200 rpm. Fourier transform infrared spectroscopy was applied to the on line and off line quantitative analysis of lactose and lactic acid during the fermentation process. This technique enabled substrate and product levels to be assessed quickly and simply, with no sample pre-treatment. Continuous culture of L. helveticus in MRS medium and supplemented whey permeate was carried out. Substrate conversion and lactic acid productivity decreased with increasing dilution rate. Maximum productivity corresponded to a dilution rate of 0.3 h" 1, whereas minimum residual substrate occured at a dilution rate of 0.1 h' 1 . Translation of the fermentation process from bench scale (11) to pilot scale (161) appeared to be successful. Completion times, productivity and lactose utilisation compared favourably with bench scale results.

Defects such as mold and bacterial stains can appear on cheese. Manually detecting defects is a time-consuming, cost-ineffective, and ergonomically unsatisfactory process for a dairy because the quality technicians must inspect each cheese before packaging. Instead, dairies would prefer an automatic detection system, but it is unclear whether reliable options are available. One potential approach is hyperspectral image analysis, which can interpret and classify chemical information from a sample. We collected hyperspectral images from a dairy using a short-wave infrared (SWIR) camera and compared three prediction models: a PLS-discriminant analysis with the software Breeze from the analysis company Prediktera and two classifiers based on a convolutional neural network and a support vector machine, both coded in Python. We found mold and lactobacilli stains using all methods, but dirt was more challenging to detect. Also, the methods had issues with false positives of lactobacilli stains. To improve accuracy, we recommend collecting more data, especially samples with lactobacilli stains and dirt, and using more non-defect cheese for validation. To find smaller defects, we propose that future work should test a visible and near-infrared (VNIR) camera with higher resolution. Though PLS-discriminant analysis did not achieve the highest accuracy, it was not far off and had the most time-effective predictions. Since Breeze already integrates PLS-discriminant analysis, it should remain in focus, but for higher accuracy Prediktera should continue to explore other methods such as neural networks.

Natamycin is an antimycotic compound that is widely used in the cheese industry to increase the shelf life of cheeses, especially shredded cheeses, by inhibiting the growth of molds. Natamycin is applied to the surface of cheese as an aqueous suspension or as a powder. However, natamycin is not readily water soluble making it harder to distribute evenly over shredded cheese Natamycin is degraded by ultraviolet (UV) light at wavelengths of 350 nm and below. Typical packaging applications do not provide adequate UV protection causing natamycin to degrade. This work was undertaken to determine the efficacy of UV absorber film to prevent UV light degradation of natamycin on the surface of shredded cheese. Current accepted methods to determine concentration of natamycin were evaluated for appropriateness in natamycin degradation studies. The use of cyclodextrins to increase water solubility was tested to see if a uniform distribution of natamycin over the shredded cheese could be done effectively. Furthermore, a known application of mold was performed to see how well natamycin and each of its applications could prevent visible mold growth from occurring. The International Dairy Federation recognizes two methods to quantify natamycin on shredded cheese: high performance liquid chromatography (HPLC) and spectrophotometry. Concentrations of natamycin in aqueous suspensions were determined using both methods. Results show that spectrophotometry is flawed when quantifying the amount of active natamycin because the method gives erroneously high results. The amount of active natamycin is not accurately quantified using spectrophotometric techniques because it cannot separate the active form from the inactive form of natamycin. Polymer packages containing a UV absorber (11.4% light transmission at 350 nm) allow significantly less UV-associated degradation of natamycin than those packages that lacked a UV protectant (90.0% light transmission at 350 nm) (p<0.05). Incorporating a UV absorber into a package helps protect natamycin and its various complexes from UV light degradation, which can increase the shelf life of shredded cheese. However, even with a UV absorber, natamycin is still able to degrade. Natamycin was complexed with different cyclodextrins to help better solubilize natamycin â β-cyclodextrin, hydroxy-propyl β-cyclodextrin and γ-cyclodextrin. Using cyclodextrins to apply natamycin more uniformly onto shredded cheese did not significantly increase the consistency of distribution (p<0.05). Variability was uniform throughout all treatments with the exception of HPBCD complex. After 27 days, all of the UV packages treated with each of the cyclodextrin treatments containing shredded cheese began to show visible mold growth. Those packages stored in total darkness remained mold free through the duration of the experiment ending on day 62. When untreated with natamycin and an initial concentration of 101-102 spores/gram of Penicillium roqueforti, shredded cheese remained free from visible mold growth for 24 days in total darkness at 4°C. Samples treated with one of the natamycin treatments were able to remain mold free for at least 9 more days, showing visible signs of mold growth at day 33. There was no statistical difference between the treatments of dry natamycin, aqueous suspension natamycin, β-cyclodextrin-natamycin complex, and γ-cyclodextrin-natamycin complex (p<0.05). However, there was a difference with the use of hydroxy-propyl β-cyclodextrin-natamycin complex. Hydroxy-propyl β-cyclodextrin-natamycin complex allowed the shredded cheese to last for 41 days, 17 days longer than the control sample.
Master of Science

Mestrado em Biotecnologia
O queijo Serra da Estrela é um queijo tradicional Português produzido a partir de leite de ovelha crú, sal e cardo (Cynara cardunculus, L.). Como outros queijos, particularmente os feitos a partir de leite crú, o Queijo Serra tem na sua composição vários microrganismos. Este trabalho teve como objetivo estudar o efeito da Alta-Pressão (AP) no Queijo Serra da Estrela - Denominação de Origem Protegida (DOP), após processamento e durante o armazenamento (100 dias a 5 °C). Queijos com 45 dias de maturação foram tratados com 400, 500 e 600 MPa durante 10, 5 e 3 minutos, respetivamente, a 4 °C. Amostras não pressurizadas foram utilizadas como controlo. Os resultados revelaram que os microrganismos benéficos para a maturação, bactérias lácticas, foram as menos afetadas pela AP, sendo reduzidas no máximo em ~ 0,50 log UFC/g (amostras tratadas a 600 MPa foram as mais afetadas). Os microrganismos totais aeróbios mesófilos foram reduzidos em ~ 1,0 log UFC / g (amostras tratadas a 400 MPa foram as menos afetadas). No entanto, a contagem de Enterobacteriaceae mostrou ≥ 3,5 log reduções, valores que se mantiveram constantes durante o armazenamento; bolores e leveduras apresentaram reduções ≥ 3,6 logo após o processamento. As amostras inoculadas com L. innocua a 8,6 log UFC/g apresentaram reduções ≥ 4,8 log após tratamento a AP, mas passados 14 dias revelaram níveis abaixo do limite de deteção. Foram observadas pequenas alterações nos parâmetros físico-químicos (pH e acidez titulável) entre o controlo e queijo tratado durante o armazenamento. Durante esse período, a oxidação lipídica foi mais intensa no queijo não processado. Os resultados obtidos permitem concluir que AP apresenta um bom potencial para tornar o Queijo Serra livre de microrganismos potencialmente patogénicos, sem alterações significativas nas características do queijo (azoto solúvel em água, atividade e conteúdo de água).
Serra da Estrela Cheese is a traditional Portuguese cheese manufactured from raw milk, salt and cardoon (Cynara cardunculus, L.). As other cheeses, particularly those made from raw milk, Serra Cheese has in its composition several microorganisms. This work aimed to study the effect of High Pressure Processing (HPP) on Serra da Estrela Cheese – Denominação de Origem Protegida (DOP) after pressure processing and during storage (100 days at 5 °C). Cheeses with 45 days of ripening were treated at 400, 500 and 600 MPa pressures during 10, 5 and 3 minutes, respectively, at 4 °C. Non-processed samples were used as controls. The results revealed that microorganisms beneficial to cheese maturation (lactic acid bacteria; LAB) were the least affected by HPP, being reduced at maximum by ~ 0.50 Log CFU/g (samples treated at 600 MPa were the most affected). Total aerobic mesophilic microorganisms were reduced by ~ 1.0 Log CFU/g (samples treated at 400 MPa were least affected). However, Enterobacteriaceae counts showed ≥ 3.5 log cycle reductions, remaining unchanged during the storage; yeasts and moulds counts exhibited ≥ 3.6 log cycle reductions after process. Samples inoculated with L. innocua at 8.6 Log CFU/g presented ≥ 4.8 log cycle reductions after HPP, but after 14 days revealed levels below the detection limit. Small changes in physicochemical parameters (pH values and titratable acidity) were observed between control and treated cheese during storage. Throughout this period, lipid oxidation was more intense in non-processed cheese. The results obtained allow concluding that HPP has good potential to render Serra Cheese free of potential pathogenic microorganisms, with no significant changes in cheese characteristics (water soluble nitrogen, water activity and content).

Pasteurized skimmilk at 4°C was acidified to pH 5.8 with 85.5% phosphoric acid (136g H3Po4;100 kg skimmilk), then warmed to 54°C and ultrafiltered to a protein concentration 9.1 ± 0.2%. The retentate was heated to 76.5°C for 16 s then cooled to 2°C. Phosphoric acid (85.5%) was added at a rate of 3.41g per kg retentate. The acidified retentate was slowly warmed to 29.5 °C (3°C/5 min) when the pH was checked. The pH at this point was no lower than 5.4. Heating was continued until a temperature of 32.2°C was reached. Glucono delta lactone was added to the retentate (17.6 g/kg retentate) and left undisturbed for approximately 80 min. The curd was cut at pH 4.7 with 0.64 cm curd knives and allowed 10 min for syneresis. Permeate obtained from the same lot of milk was acidified to pH 4.8 (66 g H3Po4;100 kg permeate), then added to the curd at 32.2°C (three parts permeate to four parts retentate) and used as a cooking vehicle. The curd was cooked to 59°C in 90 min. The curd was held at 59°C for 10 min, drained and washed once with ice water. Cream dressing containing 12.5% fat and 3% salt was used at the rate of two parts curd to one part dressing. Control cottage cheese was produced by a direct acid method from the same skimmilk used to produce ultrafiltered curd. Use of ultrafiltered skimmilk retentate for cottage cheese making resulted in 2.24% more curd (corrected to 20% solids) and 2.24% more curd per kg original milk protein than the control. However, satisfactory firmness in UF curd required slightly more than 20% solids in the final product. Sensory evaluations indicated that creamed cottage cheese was not significantly different (p

In recent years, consumers have become increasingly interested in the quality aspects of food. Food quality, in turn, is strongly related to the sensory characteristics such as the flavor. Several scientific studies have shown that the Volatile Organic Compounds (VOCs) released by the food are related to the flavor and can be considered as assistive markers in the production chain. Today, the analysis of VOCs requires fast, non-invasive, and solvent free devices. It has been shown that the VOCs can be extracted, identified, and measured with a Gas Chromatography-Mass Spectrometry (GC-MS) without any pre-concentration or pre-treatment of the food. The main objective of this PhD thesis was to investigate the presence of volatile compounds in dairy products. More precisely, this study aimed in i) qualifying and quantifying VOCs in dairy products, ii) examining their formation and iii) integrating knowledge on VOCs by tracking their release during the whole production process from the raw materials till the final dairy product. In addition, statistical analysis was applied to link VOCs with the genetic characterization of animals, dairy system and individual cow-factors (e.g. stage of lactation, order of parity and milk yield). The identification and quantification of VOCs were performed using fast and non-invasive analytical approaches (Solid Phase Micro Extraction/Gas Chromatography-Mass Spectrometry SPME/GC-MS and Proton Transfer Reaction-Time of Flight-Mass Spectrometry PTR-ToF-MS) that can monitor the evolution of VOCs. To achieve the overall goal, the research was partitioned in four interrelated subparts as described below. The aim of the first chapter was to study the VOCs presence in the headspace of cheese. To this purpose, 150 cheeses ripened for two months were used. The cheeses were obtained through an individual model cheese-making approach using milk from individual Brown Swiss cows. Animals reared in 30 herds belonging to different dairy systems, from traditional (typical of the mountainous area) to modern ones. The study identified 55 VOCs classified in the chemical families of free fatty acids, esters, alcohols, aldehydes, ketones, lactones, terpenes, and pyrazines. We found that dairy system and individual cow characteristics (lactation stage, order of parity and daily milk yield) influenced the volatile compounds. In order, to test the instrument reproducibility and the model cheese-making procedure; data of GC analysis, order of injection of the sample into instrument, and vat were included in the statistical model. In many cases, these analytical factors did not affect the amount of VOCs released by cheese. In the second chapter, the potential of a new spectrometric technique (PTR-ToF-MS) was investigated to study cheese quality traits on a large scale. The PTR-ToF-MS allows direct injection of the sample headspace without extraction or pre-concentration steps, has a shorter analysis time (only a few seconds per sample) and greater sensitivity that permit to monitor on-line the evolution of volatile compounds. The resulting spectral information can provide a very detailed description of samples, which is useful for characterizing food quality and typicality. In particular, we analyzed the volatile fingerprint of 1,075 model cheeses produced using individual milk of Brown Swiss cows reared in 72 herds of different dairy systems. The output of PTR (spectrum) was characterized by more than 600 spectrometric peaks (variables). After removing interfering ions and background noise a set of 240 peaks was selected. Further, based on the results of the first contribution and literature, 61 peaks were identified. These peaks represent the major part of the cheese flavor. To summarize the amount of information, a multivariate analysis (PCA) was applied associating principal components (PC) with the 240 spectrometric peaks. Following, we tried to characterize the PCs through the correlations between PCs and the spectrometric peaks. The effects of dairy system, herd within dairy system, individual cows characteristics (lactation stage, order of parity and milk yield), and vat used for the cheese-making on the PCs and on the 240 peaks were analyzed. Dairy system was correlated with PC and 57 spectrometric peaks, especially when the herds were using Total Mixer Ration (TMR) as feeding technique, including or not maize silage in the diets. Regarding the individual animal characteristics, the most significant effect was the stage of lactation (139 peaks), followed by milk yield and parity, with 31 and 21 peaks, respectively. Finally, the vat used for the cheese-making was not found to be significant, confirming the good reproducibility of the model cheese-making procedure used to study cheese quality aspects. In the third chapter, the effect of cows’ genetics to the VOCs of ripened cheeses was assessed. Principal components and the 240 spectrometric peaks (as described above in the second contribution) were used fitting an animal model in a Bayesian framework. On average, heritability (h2) of 7% for PCs was found, which is similar to h2 of somatic cell count and much lower than the h2 of milk fat content and daily milk yield. It is interesting to note that only a small proportion of peaks showed very low h2 (<7%). The major part of them showed values similar to those found for PCs, while forty peaks presented heritability similar to that of milk yield and other milk quality traits. The variability attributed to the herd was different for the various PC. Results suggest a potential of improvement for several cheese VOCs through genetic selection in dairy cow breeding programs. The aim of the fourth chapter was to study the effect of summer transhumance on the quality traits of dairy products. Due to the extended work, this contribution was further splitted into two parts. In the first part, the evolution of milk and cheese quality characteristics were studied, while in the second part the evolution of VOC content of dairy products was analyzed. For the first part, chemical characteristics and technological properties of 11 dairy products obtained during summer transhumance of cows to Alpine pastures (Malga) were analyzed. Dairy products obtained throughout this period are known to give origin to high-value, healthier products, and extra tasty,. Bulk milk from 148 dairy cows reared day and night on Alpine pasture (1,860 m a.s.l.) was used. We performed 7 experimental cheese-making according to traditional mountain techniques, one every two weeks, using milk produced during the summer transhumance (from June to September). For each cheese-making we collected: milk from the evening milking (day before the cheese-making), the same milk the following morning (after natural creaming), the cream separated, the whole milk from the morning milking, the milk in vat obtained mixing the creamed evening milk with the whole morning milk, the fresh curd, the whey, the ricotta obtained from whey, and the residual scotta. Moreover, the curd was used to produce typical “Malga” cheese that was ripened for 6 and 12 months. The chemical characteristics were measured with infrared technology. Results highlighted variation in milk yield, milk chemical composition, cheese yield and curd recoveries and/or loss of nutrients in the traditional cheese-making. In particular, a reduction of milk yield, fat, protein and lactose contents of milk during summer transhumance was observed. Nevertheless, the return to lowland farming systems of the cows at the end of grazing season, positively affected milk yield and milk chemical composition. The average of cheese yield was 14.2%, while recoveries of fat, protein, total solids and energy were 85.1%, 77.8%, 49.4% and 58.1%, respectively. These results were in accordance to those found in the literature. For the second part of this chapter, the VOCs content of sample headspace was measured through SPME/GC-MS. Forth nine VOCs belonging to the chemical families of alcohols, aldehydes, free fatty acids, ketones, esters, lactones, terpenes, phenolic, and sulphur compounds were detected. In addition, the evolution of VOCs and their chemical family across the cheese- and ricotta-making processing as well as during the cheese ripening period was tracked. The comparison between VOCs concentration of 4 types of milk (whole evening, creaming milk, whole morning, milk in vat) showed that the creaming process significantly affected about half of all the volatile organic compounds analyzed, followed by the effects of milking (evening milking vs. morning milking) and the mixing (creamed milk mixed with whole morning milk). In general, the cream, in contrast to curd and ricotta, showed higher content of free fatty acids, sulphurs and terpenes compounds. Moreover, in ricotta a higher VOC concentration was observed compared to the curd, probably due to the high temperature required during the ricotta process. The effect of the progressive nutrient depletion of milk was investigated by contrasts between VOC concentration of milk in the vat, whey, and scotta. Although milk contains a greater amount of nutrients, whey and scotta have shown a higher concentration of VOCs with the exceptions of esters, sulphurs, terpenes and phenolic compounds. Finally, the effect of ripening was tested by comparing the quantity of VOCs of curd and of aged cheeses (6 and 12 months). The release of volatile compounds increased with increasing ripening period in relation with the enzymatic and microbiological activity of cheese. In summary, the spectrometric techniques (SPME/GC-MS and PTR-ToF-MS) used in this work demonstrated to be very efficient to characterize the volatile organic compounds of dairy products. The dairy system, and cow related factors affected the volatile fingerprint of ripened cheeses. Particularly, concerning the individual animal source of variation, lactation stage was the most important effect followed by the cow’s parity and the milk yield. On the basis of phenotypes used in this work, the traits collected offered the potential for a genetic analysis to be carried out. The genetic analysis demonstrated the existence of an exploitable genetic variability of the volatile profile of cheese that might be useful for an (in)direct selection of dairy cows for cheese quality traits in breeding programs. Nevertheless, further research is needed in this area. In the era of genomics for e.g., it might be interesting to associate genomic regions to specific VOCs. This information might be useful for genomic breeding programs. The evolution of volatile compounds across the production chain depends on specific technological aspects, such as the process of natural creaming, the temperature of coagulation, and the ripening period. The monitoring of volatile fingerprint permits to obtain dairy products with specific organoleptic characteristics useful to differentiate them on the market and to improve the supply chain efficiency on the basis of quality aspects.
Negli ultimi anni, il consumatore è diventato sempre più sensibile agli aspetti qualitativi degli alimenti, i quali sono fortemente influenzati dalle caratteristiche sensoriali come l’aroma. Diversi lavori scientifici hanno dimostrato che i composti volatili (VOCs) rilasciati dall’alimento sono correlati con il suo aroma e possono essere considerati come traccianti delle filiere alimentari. Oggi, l’analisi dei VOCs richiede strumenti rapidi, sensibili, non invasivi e che abbiano bisogno dell’impiego di pochi solventi durante la preparazione del campione. E’ stato dimostrato che i VOCs possono essere estratti, misurati e identificati con la Gas Cromatografia di Massa (GC-MS) senza pre-concentrazioni o pre-trattamenti dell’alimento da analizzare. Gli obiettivi principali della tesi di dottorato erano di studiare la presenza di composti volatili nei prodotti lattiero-caseari. Più precisamente, questo studio aveva come obiettivi di i) qualificare e quantificare i VOCs nei prodotti lattiero-caseari, ii) esaminare la loro formazione e iii) integrare le conoscenze acquisite su questi composti attraverso tutta la filiera di produzione dalla materia prima fino al prodotto finito. Inoltre, analisi statistiche sono state utilizzate per collegare i VOCs con la caratterizzazione genetica degli animali, il sistema di allevamento e le caratteristiche individuali delle vacche (es. stadio di lattazione, ordine di parto e produzione giornaliera di latte). L’identificazione e la quantificazione dei VOCs sono state fatte utilizzando tecniche analitiche precise, veloci e non invasive (Solid Phase Micro Extraction/Gas Chromatography-Mass Spectrometry SPME/GC-MS and Proton Transfer Reaction-Time of Flight-Mass Spectrometry PTR-ToF-MS). Per rispondere agli obiettivi generali della tesi, l’attività di ricerca è stata divisa in cinque parti connesse tra di loro. L’obiettivo nel primo capitolo era di studiare i composti volatili presenti nello spazio di testa di campioni di formaggio. Per questo scopo, sono stati analizzati 150 formaggi stagionati per due mesi. I formaggi sono stati prodotti utilizzando una metodica di caseificazione individuale usando latte individuale di vacche di razza Bruna. Gli animali sono stati allevati in 30 aziende appartenenti a diversi sistemi di allevamento, da tradizionale (tipico della realtà montana) a moderno. In questo studio sono stati identificati 55 VOCs per ogni formaggio, classificati in diverse famiglie chimiche: acidi grassi, esteri, alcoli, aldeidi, chetoni, lattoni, terpeni e pirazine. Dai risultati emerge che il sistema di allevamento e le caratteristiche individuali delle vacche (stadio di lattazione, ordine di parto e produzione giornaliera di latte) influenzano i composti volatili. Inoltre, per testare la riproducibilità dello strumento e della metodica di caseificazione; la data di analisi cromatografica, l’ordine d’iniezione del campione nello strumento (GC), e la caldaia di caseificazione erano inclusi nel modello statistico. In molti casi, questi fattori analitici/strumentali non influenzano la quantità di VOCs rilasciata dai formaggi. Nel secondo capitolo, il potenziale di una nuova tecnica analitica (PTR-ToF-MS) è stato approfondito per studiare, su larga scala, le caratteristiche qualitative del formaggio. Il PTR-ToF-MS dal punto di vista analitico, permette un’iniezione diretta del campione senza estrazione o pre-concentrazione, ha un breve tempo di analisi (solo pochi secondi per campione) e grande sensibilità consentendo di monitorare in tempo reale l’evoluzione dei composti volatili. L’analisi produce uno spettro molto dettagliato che può essere utile per la caratterizzazione delle qualità e della tipicità dell’alimento. In particolare, è stata analizzata l’impronta aromatica di 1,075 formaggi prodotti utilizzando latte individuale di vacche di razza Bruna allevate in 72 aziende appartenenti a diversi sistemi di allevamento. L’impronta aromatica (spettro) era caratterizzata da più di 600 picchi (variabili) per ogni formaggio. Gli spettri sono stati analizzati e dopo la rimozione degli ioni interferenti e del rumore di fondo è stato selezionato un data set costituito da 240 picchi per ogni formaggio. In seguito, basandosi sui risultati del primo contributo e sulla letteratura sono stati identificati i picchi più importanti (61) in termini quantitativi e qualitativi. Per sintetizzare la quantità di informazioni ovvero estrarre delle componenti principali (PC) è stata fatta un’analisi multivariata (PCA) a partire dai 240 picchi spettrometrici. In seguito, le PCs sono state caratterizzate sulla base delle loro correlazioni con i 240 picchi spettrometrici. Sono stati analizzati gli effetti del sistema di allevamento, dell’azienda entro sistema di allevamento, le caratteristiche individuali delle vacche (stadio di lattazione, ordine di parto e produzione di latte), e caldaia di caseificazione sulle PCs e sui 240 picchi. Dai risultati emerge che il sistema di allevamento è correlato con le PC e 57 picchi, specialmente quando le aziende come tecnica di alimentazione utilizzano il carro miscelatore (TMR) con e senza insilati nella dieta. Considerando le caratteristiche individuali delle vacche, l’effetto più significativo è lo stadio di lattazione (139 picchi), seguito dalla produzione di latte e dall’ordine di parto, con 31 e 21 picchi, rispettivamente. Infine, la caldaia di caseificazione è un effetto spesso non significativo, confermando la buona riproducibilità della micro-caseificazione utilizzata anche per lo studio di aspetti qualitativi del formaggio. Nel terzo capitolo è stato studiato l’effetto della genetica dell’animale sui composti volatili dei formaggi. A tale scopo, sono state analizzate le componenti principali (estratte come discusso sopra nel secondo contributo) e i 240 picchi spettrometrici (PTR-ToF-MS) utilizzando un modello animale con un approccio Baesiano. Dai risultati emerge in media un’ereditabilità (h2) del 7 % per le componenti principali, la quale è simile all’h2 trovata per le cellule somatiche e leggermente più bassa di quella del contenuto di grasso nel latte e della produzione giornaliera di latte stimate in precedenza sugli stessi animali. E’ interessante osservare che solo una piccola quantità di picchi ha una bassa h2 (<7%). La maggior parte di essi presenta valori simili a quelli trovati per le PCs, mentre 40 picchi presentano ereditabilità simile a quella trovata per la produzione giornaliera di latte e ad altre caratteristiche qualitative del latte. La variabilità attribuita all'azienda è risultata diversa per le PCs. Questi risultati dimostrano che esiste un’interessante variabilità genetica di alcuni VOCs che potrebbe essere potenzialmente utilizzata nei programmi di miglioramento genetico. L’obiettivo nel quarto capitolo era di studiare l’effetto della transumanza sulle caratteristiche qualitative di prodotti lattiero-caseari. Vista la grande mole di dati, questo contributo è stato diviso in due parti tra loro connesse. Nella prima parte è stata studiata l’evoluzione della qualità del latte e del formaggio, mentre nella seconda parte è stata analizzata l’evoluzione dei composti volatili dei prodotti lattiero-caseari nel processo di caseificazione. Nella prima parte, sono state analizzate le proprietà fisiche, chimiche e tecnologiche di 11 prodotti lattiero-caseari raccolti durante la transumanza al pascolo Alpino (Malga) di vacche da latte. E’ risaputo che i prodotti ottenuti durante il periodo di alpeggio possono avere un valore aggiunto dovuto alle elevate proprietà nutrizionali, salutistiche e aromatiche. Per approfondire le conoscenze finora acquisite, è stata fatta questa prova in cui è stato utilizzato il latte di massa prodotto da 148 vacche allevate giorno e notte al pascolo (1,860 m s.l.m.). Durante l’esperimento, sono state fatte 7 caseificazioni seguendo tecniche tradizionali, una ogni 2 settimane, utilizzando il latte prodotto durante la transumanza (da giugno a settembre). Sono stati raccolti per ogni caseificazione: il latte della mungitura della sera (giorno prima della caseificazione), lo stesso latte il mattino successivo (dopo il processo di scrematura naturale), la panna di affioramento, il latte della mungitura del mattino, il latte in caldaia ottenuta dalla miscela tra il latte scremato della mungitura della sera con il latte della mungitura del mattino, la cagliata, il siero, la ricotta ottenuta dal siero e il residuo della lavorazione ossia la scotta. Inoltre, la cagliata è stata usata per produrre formaggi di “Malga” che sono stati stagionati per 6 e 12 mesi. Le caratteristiche chimico-fisiche sono state misurate con una tecnologia a infrarosso. I risultati dimostrano una variazione della produzione giornaliera e composizione chimica del latte, resa in formaggio e recupero/o perdita di nutrienti nel processo di caseificazione tradizionale. In particolare, si è osservata una riduzione della produzione giornaliera di latte, grasso, proteine e lattosio del latte durante la transumanza estiva. Tuttavia, si è anche osservato un effetto positivo sulla produzione e la composizione chimica del latte del ritorno delle vacche nelle aziende di fondo valle alla fine della stagione dell’alpeggio. La resa media di formaggio in questo lavoro è risultata del 14.2%, mentre i recuperi di grasso, proteine, solidi totali ed energia sono del 85.1%, 77.8%, 49.4% e 58.1%, rispettivamente. Questi risultati sono in linea con quelli trovato in letteratura. Nella seconda parte di questo contributo, è stato misurato il contenuto di composti volatili nello spazio di testa dei campioni con la tecnica SPME/GC-MS. Dopo l’analisi, sono stati identificati 49 VOCs appartenenti alle famiglie chimiche degli alcoli, aldeidi, acidi grassi, chetoni, esteri, lattoni, terpeni e composti solforati e fenolici. Inoltre, è stata studiata l’evoluzione dei VOCs e delle loro famiglie chimiche attraverso i processi di caseificazione, di produzione della ricotta e di stagionatura del formaggio. Il confronto tra la concentrazione dei VOCs dei 4 tipi di latte (intero e scremato della sera, intero del mattino, caldaia) ha dimostrato che il processo di scrematura influenza la concentrazione di metà dei composti volatili analizzati, seguito dall’effetto della mungitura (intero della sera vs. intero del mattino) e dall’effetto del mescolamento (latte scremato della sera mescolato in parti uguali con il latte del mattino). In generale, la panna, rispetto a cagliata e ricotta, ha un maggiore contenuto di acidi grassi, terpeni e composti solforati. Inoltre, la ricotta rispetto alla cagliata ha un’elevata concentrazione di VOC, probabilmente dovuta alla maggiore temperatura utilizzata durante il processo di produzione. L’effetto del progressivo depauperamento di nutriente del latte è stato studiato attraverso il confronto tra latte in caldaia, siero e scotta. Sebbene il latte abbia un maggiore contenuto di nutrienti, il siero e la scotta hanno una maggiore concentrazione di VOC ad eccezione delle famiglie chimiche degli esteri, terpeni, composti solforati e fenolici. Infine, l’effetto della maturazione è stato valutato attraverso il confronto tra le quantità di VOC della cagliata e dei formaggi stagionati (6 e 12 mesi). Il rilascio dei composti volatili incrementa con l’aumento del periodo di maturazione probabilmente dovuto a una maggiore attività enzimatica e microbiologica nel formaggio. In conclusione, le tecniche analitiche di spettrometria di massa utilizzate in questo lavoro (SPME/GC-MS e PTR-ToF-MS) hanno permesso di caratterizzare i composti volatili dei prodotti lattiero-caseari in maniera efficiente. Il sistema di allevamento, le caratteristiche individuali delle vacche hanno influenzato l’impronta aromatica di formaggi individuali stagionati. In particolare, riguardo alle caratteristiche individuali degli animali il principale effetto era lo stadio di lattazione seguito da ordine di parto e produzione giornaliera di latte. Sulla base dei fenotipi raccolti in questo lavoro è stato possibile effettuare un’analisi genetica, la quale ha dimostrato l’esistenza di un’interessante variabilità genetica connessa ai composti volatili del formaggio che potrebbe essere utile per una selezione (in)diretta delle vacche da latte sulla base di aspetti qualitativi in programmi di miglioramento genetico. Tuttavia sono necessarie altre ricerche in quest’area per esempio, nell’era della genomica, sarebbe interessante associare qualche regione specifica del genoma ai composti volatili. L’evoluzione dei composti volatile attraverso la filiera di produzione dipende da specifici aspetti tecnologici, come l’affioramento della panna, la temperatura di coagulazione e il periodo di stagionatura. Il monitoraggio dell’impronta aromatica permette di ottenere prodotti lattiero-caseari con delle specifiche caratteristiche organolettiche utili a differenziare il prodotto sul mercato e a migliorare l’efficienza dell’intera filiera produttiva sulla base di aspetti qualitativi.

Dairy sector is growing fast, contributing to important share of global and national economic sector and bringing nutrient components into human diets. However, dairy sector is one of the main contributors to environmental impacts arising from food sector. Cheese sector is strategic to Italian dairy sector and economy, exporting high quality and Protected Designation of Origin (PDO) cheese to international countries. There is urgent need to increase the sustainability of dairy sector, considering the whole dairy chain perspective. The aim of this Ph.D. Project was to assess the environmental impacts associated to the Italian dairy cheese chain. The Project has been conducted in Veneto Region (northeast Italy). Life Cycle Assessment (LCA) methodology have been applied to assess the impacts producing raw milk at farm gate (Manuscript I), Asiago PDO cheese from cradle to dairy plant gate (Manuscript II) and Mozzarella from cradle to grave (Manuscript III). In the Project, the primary data have been collected through direct interview to dairy farmers and visits to each dairy farm, and direct interview and visits to Asiago and Mozzarella cheese plants. Primary data collection regarded 34 dairy farms and an Asiago and Mozzarella cheese dairy plants. Ecoinvent® v3 and Agrifoodprint® v1 databases have been use for secondary data, while data from literature and national inventories have been use to model the post plant phases (distribution, retail, consumption and disposal) in the Manuscript III. Simapro© 8 was the modelling software. The impacts estimated affect human health, ecosystem and resources use. Manuscript I and Manuscript II represent the LCA of Asiago PDO cheese production. Indeed, the 34 dairy farms analyzed are located in the adjacent area of the Asiago plant, creating a unique narrow chain for producing Asiago PDO cheese which is manufactured by the PDO guidelines. In the Manuscript I, the functional unit was one kg of milk, and it was the first LCA on milk production in Veneto Region. The production of purchased feed and on-farm feed (which require land, water, chemical fertilizers and manure, machinery use) and animal emissions (enteric methane, and nitrogen emissions from manure management) were main hotspots for overall impact categories, such as climate change, acidification, eutrophication, water and land use, and energy usage. Minor contribution to final impacts originated from electricity, fuels and detergents use, and waste produced during farm activities (such milking, cleaning). Different allocation approaches were tested beside the biological default allocation, and all approaches modified the final results per kg of milk. The results were similar to those reported in literature. In the Manuscript II, the LCA model represents the production chain to produce one kg of Asiago PDO cheese, ready to sell, at dairy plant gate. The raw milk production represented the main contributor to all impact categories, except for ozone depletion where the cheese-making process was the first driver. Excluding farm phase from the assessment the manufacturing operations resulted the hotspots for overall impacts, except for eutrophication and water depletion mainly caused by wastewater treatment, and land occupation which was occurred due to primary and secondary paper packaging. The main contributor inside the cheese plant were electricity and natural gas usage, and process water, moreover transport of raw milk from farm to cheese plant impacted toxicity and photochemical oxidant formation. Economic allocation was applied and compared to milk solids content allocation, which reduced the final emissions per kg of Asiago than economic allocation. In fact, the milk solids allocation assigned more impact to the co-products (whey and other cheese) than economic allocation. Uncertainty analysis and sensitivity analysis of aging period were included into the study. In the Manuscript III the LCA methodology has been applied to an high industrialized mozzarella plant, the Italian third largest mozzarella plant. The LCA was performed in a cradle to grave perspective, including the post manufacturing phases, as distribution, retail, consumption and end of life phases. The plant used Italian and foreign milk, and distribute the mozzarella to Italy and international countries. The functional unit was one kg of mozzarella consumed. Result confirmed that raw milk production was the main contributor to overall impacts categories, except for ozone depletion where refrigerant used for cooling along the post farm chain were the main hotspot. Manufacturing and packaging were the second most important contributors to final impacts, followed by disposal of wastewater, while minor impacts were associated to distribution, retail and consumption; although relevant contribution was transport of milk and mozzarella, considering the international origin and delivery of the products. Electricity and natural gas usage, together with cardboard packaging for delivery drove the impacts during mozzarella-making process. While the impact arising from post plant phase were mainly determine by energy usage. The normalized results showed ecotoxicity, acidification, eutrophication and climate change as the main impact category contributing to the European impact and these categories are the first scope to apply strategies for reduction. A sensitivity analysis was performed to test different allocation approaches and to analyzed how the final results are influenced by allocation method; finally, a sensitivity analysis was performed determining the difference of impacts among the tradition high moisture mozzarella and the low moisture mozzarella. This analysis highlighted, excluding the difference derived from farm phase, transport is the main cause of larger impact for low moisture mozzarella, because foreign raw milk is generally used for this type of production, and the cooking in oven in the consumption phase, because low moisture mozzarella is largely used as pizza topping, in fact a cooking in an electric oven was assumed.
Il settore lattiero-caseario è in crescita, contribuendo in maniera importante al settore economico mondiale, inoltre è un settore che apporta fondamentali nutrienti nella dieta umana. Tuttavia, il medesimo settore è uno dei principali comparti che determinano l’impatto ambientale associato al settore alimentare. Il settore della produzione di formaggi è un ambito strategico del comparto lattiero caseario italiano e per l’economia italiana, che vanta esportazione di formaggi di alta qualità e formaggi DOP verso numerosi paesi internazionali. C’è un urgente bisogno di aumentare la sostenibilità del settore lattiero caseario, considerando questa necessità in una prospettiva “dalla culla alla tomba”, ossia dalla produzione delle materie prime per la produzione di prodotti lattiero caseari, al loro consumo, ed infine allo smaltimento dei rifiuti associati al loro ciclo di vita. Lo scopo generale del progetto di dottorato di ricerca è stato quello di valutare gli impatti ambientali derivanti dal settore lattiero caseario italiano con particolare riferimento alla Regione Veneto. L’analisi del ciclo di vita (Life Cycle Assessment, LCA) è stata la metodologia utilizzata per valutare gli impatti ambientali associati alla produzione di latte “dalla culla al cancello della azienda” (Manoscritto I), del formaggio Asiago DOP in una prospettiva “dalla culla alla cancello del caseificio” (Manoscritto II) e della mozzarella in una prospettiva “dalla culla alla tomba” (Manoscritto III). I dati primari del progetto sono stati raccolti tramite interviste agli allevatori delle 34 aziende agricole coinvolte nel progetto, e ai responsabili del caseificio produttore di Asiago e dello stabilimento di produzione della mozzarella. Ecoinvent® v3 and Agrifoodprint® v1 database sono stati usati come fonte di dati secondari usati nel progetto. Dati da letteratura scientifica e report nazionali sono stati utilizzati per modellare le fasi post-stabilimento (distribuzione, consumo e smaltimento) nel Manoscritto III. Simapro© 8 è stato il software utilizzato per stimare gli impatti ambientali nei tre Manoscritti. Gli impatti stimati hanno riguardato la sfera della salute umana, l’ecosistema e utilizzo di risorse. Manoscritto I e Manoscritto II presentano lo studio LCA per il formaggio Asiago DOP. Infatti le 34 stalle da latte analizzate sono localizzare nel territorio regionale, creando una unica filiera produttiva, come richiesto dal disciplinare di produzione del formaggio Asiago DOP. Nel Manoscritto I, l’unità funzionale è stata un kg di latte prodotto. In particolare, il Manoscritto I rappresenta il primo studio LCA relativo alla produzione di latte bovino nella Regione del Veneto. La produzione di mangimi extra-aziendali e la produzione dei mangimi aziendali (i quali richiedono terra, acqua, fertilizzanti e reflui zootecnici, e macchinari) e le emissioni riconducibili ai bovini (metano enterico, e emissioni di ossido di diazoto dai reflui) sono stati i principali driver per la totalità degli impatti stimati, tra cui cambiamento climatici, acidificazione, eutrofizzazione, utilizzo di acqua, suolo ed energia. Un minore contributo agli impatti stimati è associato all’utilizzo di elettricità, carburanti e detergenti, e ai rifiuti prodotti durante le attività svolte in stalla (es. mungitura e pulizie). Sono stati testati differenti metodi allocativi alternativi al metodo base, il quale era l’allocazione biologica tra latte e peso vivo prodotto in stalla. Diversi metodi allocativi hanno portato a diverse emissioni per unità funzionale come riscontrato in letteratura. Il Manoscritto II ha presentato LCA della filiera produttiva del formaggio Asiago DOP, e come unità funzionale aveva un kg di Asiago al cancello del caseificio, dopo la fase di stagionatura. La produzione di latte bovino in stalla è il driver principale per gli impatti stimati ad eccezione del depauperamento dello strato di ozono del quale il processo di caseificazione era il principale driver. Escludendo la fase di produzione del latte bovino, l’analisi LCA ha dimostrato come le fasi di caseificazione siano quelle maggiormente impattanti, ad esclusione di eutrofizzazione e utilizzo di acqua per le quali il maggiore responsabile è rappresentato dai processi di trattamento delle acque reflue ottenute durante i processi di caseificazione, ed infine l’occupazione di suolo, il quale è stato primariamente determinato dalla produzione del packaging cartaceo primario e secondario. I principali driver durante le fasi di caseificazione del formaggio sono risultati l’utilizzo di elettricità e gas naturale, e l’acqua utilizzata durante la caseificazione (principalmente in fasi di pulizia), inoltre il trasporto del latte dalla stalla al caseificio ha contributo agli impatti di tossicità e formazione di ossidi fotochimici. E’ stata comparata l’ allocazione economica e l’allocazione basata sul contenuto di solidi del latte presenti nel formaggio Asiago e nei coprodotti (altri formaggi prodotti in caseificio e siero liquido), ed è stato evidenziato che l’allocazione basata sul contenuto di solidi del latte ha determinato delle minori emissioni per kg di formaggio Asiago rispetto alla allocazione economica. Infatti, l’allocazione basata sul contenuto di solidi del latte assegna una maggior percentuale di impatti ai due coprodotti presenti. Infine è stata svolta l’analisi di incertezza dei risultati ottenuti, e analisi di sensitività basata sulla durata del processo di stagionatura. Il Manoscritto III a differenza del Manoscritto II ha applicato la metodologia LCA in un stabilimento altamente industrializzato per la produzione di mozzarella (terzo produttore di mozzarella a livello italiano). LCA è stato realizzato in una prospettiva “dalla culla alla tomba”, considerando le fasi post-stabilimento, quali distribuzione presso la GDO e i piccoli rivenditori, fase di consumo e il fine vita. Lo stabilimento produttivo usava sia latte bovino italiano che estero, e la mozzarella prodotta veniva distribuita sia sul mercato nazionale che internazionale. L’unità funzionale dello studio era un kg di mozzarella consumata. Il risultato dello studio ha confermato come la produzione di latte in stalla rappresenti il principale driver per la maggioranza degli impatti stimati, con eccezione del depauperamento dello strato di ozono, per il quale i gas refrigeranti utilizzati per la refrigerazione e lo stoccaggio dei prodotti nelle fasi di produzione e in quelle post-stabilimento produttivo erano i principali drivers. Le fasi di produzione in stabilimento e il packaging sono risultate essere secondi principali drivers per gli impatti finali, seguiti da trattamento e smaltimento delle acque reflue, mentre i minori impatti sono stati identificati per le fasi di distribuzione e consumo della mozzarella. I risultati ottenuti sono stati normalizzati e hanno mostrato nella eco-tossicità, acidificazione, eutrofizzazione e cambiamenti climatici le principali categorie di impatto; per queste categorie di impatto dovrebbero essere focalizzate primariamente le strategie per la riduzione delle emissioni. L’analisi di sensitività condotta nello studio ha evidenziato come il metodo allocativo influenzi i risultati finali. Inoltre un’analisi di sensitività è stata eseguita per differenziare le due principali tipologie di mozzarella nel mercato italiano ed anche internazionale: bocconcino di mozzarella (alto contenuto di umidità) e mozzarella da pizza (basso contenuto di umidità). La comparazione ha evidenziato che, una volta non considerata la differenza derivante dalla fase di stalla la quale contiene anche la maggior variabilità, il trasporto del latte in stabilimento rappresenta la causa principale del maggiore impatto per la mozzarella da pizza, infatti a livello italiano, come nello stabilimento analizzato, la mozzarella da pizza è prodotta utilizzando primariamente latte di origine estera, richiedendo una maggiore distanza di trasporto, con conseguente aumento della emissioni. Infine, un’altra rilevante parte degli impatti è stata associata alla cagliata acquistata da stabilimenti italiani e esteri ed utilizzata per la produzione di mozzarella da pizza. Infatti, tale semilavorato prima di essere trasformato in mozzarella, è stato a sua volta prodotta in stabilimenti appositivi, richiedendo vari input quali energia, materiali e packaging, ed infine trasportata nello stabilimento di produzione della mozzarella.

Gorgonzola is a blue-veined, mould-ripened cheese, made from pasteurized cow’s milk inoculated with starter cultures (Streptococcus thermophilus and Lactobacillus delbrueckii), along with Saccharomyces cerevisiae and Penicillium roqueforti, the main responsible of the aroma and flavour of the cheese at the end of ripening. For this reason, monitoring the fungal growth and unravel the microbial dynamics during the ripening process is crucial. Traditional methods based on colony forming unit (CFU) along with alternatives (ergosterol, dry weight, spores counting) are not suitable for hyphal filaments quantification in the cheese matrix. Here we monitored and compared the P. roqueforti development between two Gorgonzola productions from two different years and the same plant, by means of a qPCR assay, mass spectrometry-based chemical analysis and an assay with a glutamate-sensitive luminescent biosensor for the evaluation of proteolysis. qPCR assay was based on a species-specific primer set targeted on ari1 gene. This assay allowed the monitoring of P. roqueforti development during cheese ripening. The lipolysis and proteolysis cheese profiles obtained by mass-spectrometry and luminometric analysis were in good agreement with the qPCR data, revealing that the most relevant blooming of the mould occurs after 20 days of ripening, and that the most recent production had a lower amount of fungal mycelium produced and a lower enzymatic activity, thus affecting the final product quality. The metabolic analysis on sugars consumption revealed the importance of galactose degradation on the final product quality. The analytical methods here described are useful for a rapid evaluation of the maturation process and the quality of blue-cheeses.

Thesis (Ph. D.)--Ohio State University, 2005.
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