We investigated the carbon dioxide metabolism of Streptococcus thermophilus, evaluating the phenotype of a phosphoenolpyruvate carboxylase-negative mutant obtained by replacement of a functional ppc gene with a deleted and inactive version, ⌬ppc. The growth of the mutant was compared to that of the parent strain in a chemically defined medium and in milk, supplemented or not with L-aspartic acid, the final product of the metabolic pathway governed by phosphoenolpyruvate carboxylase. It was concluded that aspartate present in milk is not sufficient for the growth of S. thermophilus. As a consequence, phosphoenolpyruvate carboxylase activity was considered fundamental for the biosynthesis of L-aspartic acid in S. thermophilus metabolism. This enzymatic activity is therefore essential for growth of S. thermophilus in milk even if S. thermophilus was cultured in association with proteinase-positive Lactobacillus delbrueckii subsp. bulgaricus. It was furthermore observed that the supplementation of milk with aspartate significantly affected the level of urease activity. Further experiments, carried out with a p ureI -gusA recombinant strain, revealed that expression of the urease operon was sensitive to the aspartate concentration in milk and to the cell availability of glutamate, glutamine, and ammonium ions.

No abstract

Extending ripening of hard cheeses well beyond the traditional ripening period is becoming increasingly popular, although little is known about the actual evolution of their characteristics. The present work aimed at investigating selected traits of Parmigiano Reggiano cheese ripened for 12, 18, 24, 30, 40 and 50 months. Two cheeses per each ripening period were sampled. Although moisture constantly decreased and was close to 25% in 50-month cheeses, with a parallel increase in cheese hardness, several biochemical changes occurred involving the activity of both native and microbial enzymes. Capillary electrophoresis demonstrated degradation of αs1- and β-casein, indicating residual activity of both chymosin and plasmin. Similarly, continuous release of free amino acids supported the activity of peptidases deriving from lysed bacterial cells. Volatile flavor compounds, such as short-chain fatty acids and some derived ketones, alcohols and esters, evaluated by gas chromatography with solid-phase micro-extraction, accumulated as well. Cheese microstructure was characterized by free fat trapped in irregularly shaped areas within a protein network, with native fat globules being no longer visible. This study showed for the first time that numerous biochemical and structural variations still occur in a hard cheese at up to 50 months of aging, proving that the ripening extension deserves to be highlighted to the consumer and may justify a premium price.

Fat separation, gelation or sedimentation of UHT milk during shelf-storage represent instability phenomena causing the product rejection by consumers. Stability of UHT milk is of increasing concern because access to emerging markets currently implies for this product to be stable during shipping and prolonged storage, up to 12 months. The role of microfiltration prior to UHT process in avoiding or retarding the gelation or sediment formation was studied by comparing microfiltered UHT milk to conventional UHT milk. A second trial was set up to study the effects of double ultra-high pressure homogenization in delaying the cream rising and UHT milk homogenized once at lower pressure was taken as control. All milk samples were produced at industrial plant level. Milk packages were stored at 22 °C, opened monthly for visually inspecting the presence of cream layer, gel or sediment and then analysed. Microfiltration markedly delayed the formation of both gel particles and sediment, with respect to the control, and slowed down the proteolysis in terms of accumulation of peptides although no correlation was observed between the two phenomena. The double homogenization, also evaluated at ultra-structural level, narrowed the fat globule distribution and the second one (400 MPa), performed downstream to the sterilization step, disrupted the fat-protein aggregates produced in the first one (250 MPa). The adopted conditions avoided the appearance of the cream layer in the UHT milk up to 18 months. This study contributes important knowledge for developing strategies to delay instability phenomena in UHT milk destined to extremely long shelf storage.

No abstract

-The sensorial and physico-chemical characteristics described in the product specification for most PDO cheeses are inadequate to verify the compliance of cheeses on the market with the registered designation, particularly for grated products. During the past few years, much research has indicated the analytical parameters suitable for distinguishing Grana Padano (GP) from other similar hard cheeses. The characterization of grated GP is currently based on 3 analytical parameters, related to different aspects of cheese processing, which are: (i) the measurement of alkaline phosphatase (ALP) activity, a marker for possible heat treatment applied to milk, in the outermost layer of the cheese, just below the rind; (ii) the identification of specific peptides, that are identified only in the rind, due to the very slow progress of proteolysis in the rind during GP cheese ripening; and (iii) the free amino acid (FAA) composition. In the present study, we developed an extraction method, based on density gradient centrifugation of solubilized cheese, to separate the outermost layer of the cheeses from the rest in grated cheese, and we proposed a simplified criterion to evaluate the "typicalness" of the FAA pattern. The quality control scheme based on ALP activity, detection of specific peptides and FAA pattern was applied to more than 300 samples of marketed grated GP collected over three years, 10% of which were collected outside Italy, and ∼ 100 samples of grated generic ("Grana-type") hard cheeses. The results demonstrate that the simultaneous application of the three parameters allows one to distinguish grated GP from similar, non-PDO grated hard cheeses, and to recognize irregular GP cheeses.Grana Padano cheese / quality control / alkaline phosphatase / cheese peptides / free amino acidsArticle published by EDP Sciences 596 S. Cattaneo et al. Résumé -Fromage Grana Padano râpé : nouvelles indications pour le contrôle de qualité et pour reconnaître les imitations.Les caractéristiques sensorielles et physico-chimiques indiquées dans le cahier des charges de la plupart des fromages AOP sont peu utiles pour vérifier la conformité des produits mis sur le marché avec l'appellation, surtout pour les fromages râpés. Au cours des dernières années, plusieurs recherches ont mis en évidence les paramètres analytiques capables de distinguer le fromage Grana Padano (GP) d'autres fromages à pâte dure similaires. Actuellement, la caractérisation du GP râpé se base sur trois paramètres analytiques, liés à différents aspects de la production : (i) l'activité de la phosphatase alcaline (ALP), indicateur d'un possible traitement thermique du lait, mesurée dans la pâte du fromage, juste en-dessous de la croûte ; (ii) l'identification de peptides spécifiques de la croûte, du fait de la protéolyse très réduite dans cette partie durant l'affinage ; et (iii) la composition en acides aminés libres (AAL). Dans la présente étude, nous avons développé une méthode d'extraction basée sur la centrifugation en gradient de densité du fromage...

No abstract

21The late blowing defect still represents a problem for hard cheeses. Thus, the behaviour of the 22 cheese spoiling bacterium C. tyrobutyricum was studied throughout the cheesemaking and ripening and then were no longer cultivable. However, 2 x 10 2 spores appeared at the end of this stage, likely 28 triggered by the exponential growth phase, and were present until 6-month ripening. In cheese, C. 29tyrobutyricum UC7086 proved to convert free arginine to citrulline and then to ornithine, and to 30 produce γ-aminobutyric acid by glutamate transamination rather than by decarboxylation.