A proteomic approach for the characterization of typical meat products: definition of molecular markers of industrial and artisanal Naples-type salami
GUALTIERI, LIBERATA (2011) A proteomic approach for the characterization of typical meat products: definition of molecular markers of industrial and artisanal Naples-type salami. [Tesi di dottorato] (Inedito)
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Naples-type salami is dry fermented sausages with medium-size grana made of coarsely minced lean pig, mixed with fat, salt, pepper and various spices, stuffed into natural or artificial casings and ripened for 30-60 days, native to the Campania, today produced throughout Italy. The organoleptic and sensory properties of this product are due to the degradation events (proteolysis, lipolysis) occurring during the maturation: the lipid fraction undergoes hydrolytic and oxidative changes, involving liberation of free fatty acids (FFA), volatile compounds and precursor of odorous molecules. Endogenous enzymes, such as calpains and cathepsins, are primarily responsible for the initial degradation of the sarcoplasmic and myofibrillar proteins, then the most commonly found Lactobacillus species in dry fermented meats are able to hydrolyse myofibrillar and sarcoplasmic muscle proteins in vitro. The peptides produced can influence final taste of the salami and moreover are object of the activity of endogenous and exogenous amino peptidases releasing amino acids which represent precursors of aromatic compounds. The aim of this work has been to characterize the metabolites formed during ripening of Naples-type salami, in particular have been evaluated structural and aromatic differences between industrial and artisanal products. Analyses were carried out on two Naples-type salami experimental productions, an industrial and an artisanal, that differ in amount of pepper and for the use of microbiological starter in industrial salami and on four salami samples procured on the market. At first a proteomic analysis has been carried out on sarcoplasmic fraction of Naples-type salami by RP-HPLC technique coupled to MALDI-TOF-MS and ESI-MS, later on a metabolomic analysis of lipid and aromatic components using HS-SPME-GC-MS techniques. The proteolysis produced a different final proteic pattern probably due to the action of different microflora a microbial starter in industrial salami and autochthon microflora in artisanal one; the proteins less susceptible to proteolysis were carbonic anhydrase 3, glyceraldehyde-3-phosphate dehydrogenase , β-enolase 3 and triosephosphate isomerase; in both salami samples were observed the total proteolysis of creatine kinase, phosphofructokinase and phosphoglicerate kinase, instead myoglobin, fructose bisphosphate aldolase A and phosphoglicerate mutase disappeared only in industrial salami and troponine C 2, pyruvate kinase and glucose-6-P-isomerase in artisanal one. The fresh meat sample presented a few peptides because in an early stage of the tenderization process (48-72 h), however they derived mainly from the proteolysis of phosphofructokinase, piruvate kinase and β-enolase 3. In the sample of industrial salami after 30 days of ripening the proteolysis resulted very pronounced with production of a high number of novel peptides that derived mainly from glyceraldehyde-3-phosphate dehydrogenase, pyruvate kinase and fructose bisphosphate aldolase A. Moreover some peptides common to industrial samples after 7, 21 and 30 days of ripening, such as the peptide 284-304 of calpastatin and the peptide 87-107 of creatine kinase, have been identified and some other common to artisanal salami in order to use them as molecular markers as indicators of manufacturing and technological processes. Final proteic pattern of industrial salami resulted different from artisanal one because essentially endogenous muscle enzymes, such as calpains and cathepsins, act during ripening of artisanal salami together with enzymes of inside microflora. These differences can be put in evidence by creation of data bank could be achieved to quickly identify and safeguard typical products. Furthermore the characterisation of the peptides formed during ripening could be used to evaluate the degree of proteolysis in a given meat sample as well as to correlate the rheological and sensory characteristics with formation of typical compounds. The aromatic profile of industrial salami resulted more complex than artisanal one for the use of starter cultures. Pepper compounds were quantitatively the largest group of volatiles identified in the aromatic profile of industrial salami at 30 days of ripening. The absence of microbial starter in artisanal salami determine a minor concentration of compounds such as ethyl esters and methyl ketones related to activity of S. xylosus and carnosus species. Among volatile and semivolatile odorous molecules identified in salami samples, were present short chain fatty acids such as octanoic acid, decanoic, dodecanoic. These molecules are present in greater amount in the industrial salami to indicate a more advanced lipolytic process in this last one respect to artisanal salami. The characterization of sarcoplasmic proteins and peptides as well as the analysis of aromas could be used for identification of molecular markers of quality and typicality in order to obtain the Protected Designation of Origin (P.D.O.) mark, to differentiate an artisanal salami from an industrial one and to allow traceability of the products.
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