Relation between degree of hydrolysis and hydrolyzed beta-lactoglobulin determined by electrophoretic measurements

BALLERINI, G. A.; BÁEZ, G. D.; MORO, A.; BUSTI, P. A.; DELORENZI, N. J.

Rosario

Congreso; XII. Congreso, XXX Reunión anual de la Sociedad de Biología Rosario 2010; 2010

Enzymatic hydrolysis of bovine whey proteins, whose main component is beta-lactoglobulin (b-LG), is used in order to obtain food additives with increased emulsifying and foaming properties. At pH 8.0, b-LG presents a conformational transition with the subsequent exposition of sites sensitive to protease attacks. A stock 25 mg mL-1 b-LG solution was prepared in 50 mM phosphate buffer at pH 8.0. This sample was then submitted to trypsin hydrolysis (1/167 enzyme/substrate ratio) at 37 ºC. Aliquots were withdrawn at 0, 5, 15, 30, 60, 120, 180 and 240 minutes of hydrolysis time. The reaction was stopped by heating (5 minutes at 85 ºC). Electrophoresis in polyacrylamide gels (SDS-PAGE) was carried out using a stacking gel of 10 % and a resolution gel of 15 %. The digitalized image of the respective stained gels was analyzed using specific software developed by our research group. Remaining b-LG was determined and expressed in percentage related to non-hydrolyzed sample (0 minutes). Hydrolyzed b-LG was obtained by difference. Trimethylbencenosulfonic acid method (TNBS) was applied to determine the degree of hydrolysis (DH) of the different samples. The results obtained showed a linear relation between DH (%) and hydrolyzed b-LG (%). The statistical analysis showed the following parameters: 0.127 (p