Interaction of soybean hull peroxidase with syntethic polielectrolytes: an alternative method for enzyme purification using simple and escalable methods

CAMISCIA, PAOLA; NATALIA DE FAZIO; BOSIO, BÁRBARA; PICÓ, GUILLERMO A.; WOITOVICH VALETTI, NADIA

Saltillo

Congreso; 9th International Forum on Industrial Bioprocessing; 2021

Peroxidases are ubiquitous enzymes, with a wide variety of functions and applications. Currently, the main source of peroxidase is the roots of A. Rusticana. An alternative source of peroxidases could be soybean hulls (SBH). While processing soybeans, one of the first steps is the separation of its “hull”. This residue represent 2% of the grains weight and have little applications. SBH contain 12% soluble proteins, 10-12% pectin, 40% cellulose and 1-3% lignin [1]. Among the soluble proteins exists the soybean peroxidase (SBP), which has proven to be a good substitute of the existing commercial enzyme [2]. The main objective of the present work was to develop a new method for the purification of SBP using precipitation with polyelectrolytes: Eudragit L and Eudragit S. The working material consisted of a protein extract obtained by leaching the SBH using 50 mM phosphate buffer, pH 6.00 as the extraction phase [3]. The precipitation pH for both polymers was in the range of 4.00 and 5.00. When the pH increased, the precipitation lowered considerably falling to zero when pH values were above 5.00. Titration curves of the extract at different pH were performed with the polymers. In both cases, similar curves were obtained where the amount of precipitated SBP increases until reaching an equilibrium point. The amount of polymer required to reach maximum precipitation was in the order of 1.10-5 % w/v for Eudragit S and 2.5.10-5% w/v for Eudragit L. It was also observed that none of the polymers affects the activity of the protein. The data obtained were applied for the purification of the protein with successful results. When Eudragit S was used, the enzyme was purified 6.2 times with a yield of 75%, working at pH 5.00. While with Eudragit L a purification factor of 2.6 was obtained with a yield of 73.7% when working at pH 5.50. The developed protocol is presented as an inexpensive, scalable and non-polluting alternative for the purification of SBP from an agro-industrial waste.