Characterization of milk-clotting activity of brewer´s spent grain proteases

PEPE, ALFONSO; TITO, FLORENCIA ROCIO; DALEO, GUSTAVO RAÚL; TONÓN, CLAUDIA VIRGINIA; GUEVARA, MARIA GABRIELA

Paraná

Congreso; LIV Reunión anual Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2018

Study and characterization of the residues of agri-food industries has generated great interest in the scientific community since last thirty years, due to they can constitute an intermediate raw material for the manufacture of value-added products. Brewers? spent grain (BSG) is a rich source of protein that is currently only used to supplement livestock feed, or discarded. However, the proteins present, specifically enzymes, could be better used if they were isolated and used to carry out another process, reducing the environmental impact of direct discarding. The aim of this work was to analyze the milk- clotting activity (MCA) of BSG proteases. From BSG, the remaining proteins were extracted with Tris-HCl buffer pH 8 100 mM and precipitated with NH4SO4 at a final concentration of 80% w/v. Proteolytic activity was determined against azocasein. Activity was observed in the supernatant (SN). SN proteolytic activity was optimized, finding the optimum values at pH 7 and 40ºC. MCA of SN was determined by Arima method, using different SN: milk ratios. Incubations were at 37ºC overnight. The 1:1 ratio was found to be more suitable for obtaining the best clot. Caseinolytic activity of SN on α-, β- and κ-casein subunits was also evaluated to determine the degree of hydrolysis. Incubations were performed for 6 h at 37ºC and the percentage of hydrolysis was evaluated by SDS-PAGE. Positive proteolytic activity was observed on the κ-casein subunit, and little or no activity on α and β- subunits. These results suggest that BSG may be a possible source of enzymes that can be used for the production of spreadable cheeses.