Study of proteolysis in river buffalo mozzarella cheese using a proteomics approach

The guarantee of the origin and quality of raw material is essential for the protection and valorization of Campana buffalo mozzarella cheese. The risk of utilization of semifinished products and stored milk in substitution for fresh milk is increasing, due to the continuous desire to reduce production costs. A proteomics approach and electrophoresis survey of retail mozzarella cheeses indicated different rates of proteolysis in the production of dairy industries. The use of fresh milk and correct cheesemaking protocol yielded only gamma-caseins, which are derived from beta-casein by plasmin, and para-kappa-casein, which is derived from kappa-casein by chymosin. The detection of abnormal hydrolysis resulting in beta- and alpha(S1)-casein fragments, identified by mass spectrometry, indicates the use of stored milk or stored and pressed curd, or the reuse of unsold mozzarella cheese, to produce mozzarella. The formation of gamma-caseins and other fragments during a long storage of raw materials at room or refrigeration temperature was ascribed to plasmin (endogenous milk enzyme), whereas formation of alpha(S1)-casein fragments, mainly alpha(S1)-I-6P and alpha(S1)-I-7P-casein during the storage of curd was ascribed to the action of chymosin (exogenous enzyme) from rennet. Sodium dodecyl sulfate-PAGE and alkaline urea-PAGE permitted us to evaluate the freshness of the raw materials used in the manufacturing of buffalo mozzarella cheese and to reveal possible inappropriate preservation.