Enzymatic Hydrolysis Of Pichia Pastoris Cells

Yeast extract contains abundant nutritients and are widely used in microbial fermentation. However, enzymatic process for preparing yeast extract has not been systematically studied. Based on the yeast cell components, a novel process for yeast extract preparations was developed by using spent yeast cell of Pichia pastoris from enzymes production industry. The composition and contents of amino acids of the yeast extract were determined and its nutritional value evaluated. The main conclusion is as follows: Effective enzymatic hydrolysis of Pichia pastoris has been obtained by combined use of enzymes ?-Mannase, ?-glucanase, pectinase, neutral protease, DNase and RNase.The effects of temperature, pH, yeast cell concentration and time on the hydrolysis efficiency of the enzyme mixture-?-Mannase, ?-glucanase and pectinase were studied by orthogonal experiment. The optimum conditions are as follows: temperature 50?, pH5.5,yeast cell 10% (w/v) and digestion time 12 h.The enzymatic properties were determined for prepared neutral proteinase, DNase and RNase. For neutral proteinase, the optima pH and temperature were 7.0 and 50?,respectively.. Mn2+, Mg2+, Ca2+ enhanced its activity. The optima pH and temperature of DNase were 7.0 and 30?, respectively. Fe2+ Cu2+, Mn2+, Mg2+ enhanced its activity. The optima pH and temperature of RNase were 9.0 and 37?, 0.5 mmol/L Co2+, EDTA enhanced its activity. These enzymes are suitable for hydrolysis of Pichia cells.The optimized enzymatic process for preparing yeast extracts was established as follows: 1) The amounts of enzymes used included ?-Mannase of 508.5 U, ?-glucanase(508.5 U), pectinase (56.5 U), neutral protease (56.5 U), DNase (18.8 U) and RNase (300.0 U ); 2) Spent Pichia cells were re-suspended to a final concentration of 10% (w/v); 3)(3-Mannase, ?-glucanase and pectinase were then added simultaneously to hydrolyze yeast cells at 50? for 12 h; 4) The hydrolysis was then allowed to proceed with addition of neutral protease at 50? for 7 h; 5) The reaction mixture was then treated with DNase and RNase for another 4 h; 6) After all the steps of enzyme treatments, the hydrolysate was centrifuged, and yeast extract was obtained from the supernatant by vacuum freeze-drying or spray drying, with a dissolution rate of 66.2%, a dry matter yield of 65.7%. The resultant yeast extract contains 17 amino acids with a content of 3.64 % and amino nitrogen of 5.94%. Yeast extract applied to microbial cultivation showed that it outperformed the Oxoid yeast extract.