Production and Evaluation of Novel Exogenous Phytase Enzymes Using Several Testing Variables

Broiler chickens reared in the United States are mainly fed corn-soybean based diets. Most of the phosphorus in these diets is bound to a molecule called phytate; this source of phosphorus is unavailable to the bird because they lack adequate amount of endogenous phytase enzyme. Since the 1990\u27s exogenous phytase enzymes have been incorporated in broiler diets to liberate the phytate-bound phosphorus. These enzymes must be able to resist thermal denaturation associated with pelleting and maintain high efficacy in the bird. Most of the literature focuses on single aspects of phytase efficacy. More specifically, experiments evaluate phytase based on either in vitro retention or in vivo measures, but rarely are both considered simultaneously. Therefore, six experiments were conducted at West Virginia University to elucidate variable effects in order to demonstrate how to properly assess the efficacy of microbial expressed phytase variants. The authors believe that several testing experiments are essential to properly assess phytase and sequence is important. The following variables were considered: 1) in vivo assays using lead candidates in mash feed; 2) optimization of enzyme production based on a specific expression host; 3) in vivo dosing assay using the best enzyme/host combination; 4) in vivo assay using incrementally increased pelleting temperatures. In vitro activity was determined for each experiment and in vitro retention using increasing pelleting temperatures was determined for experiments 1 and 2. All experiments utilized Cobb 500 broiler chickens and included a standard curve, generated by feeding incrementally increased non-phytate phosphorus (nPP) via inorganic phosphorus. Linear and quadratic regressions of the standard curve were tested to determine nPP sparing effects. Four experimental phytases were determined from in vitro activity; two of the four enzymes were chosen after the in vivo assay using mash feed. These two enzymes were expressed in yeast and bacterial hosts. The best enzyme/host combination was chosen for in vivo testing at various doses. Finally the enzyme/host combination at an appropriate dose was pelleted at incremental temperatures for in vivo testing. The best enzyme expressed in the yeast host at 750 FTU/kg was found to perform similar to a positive control diet. However, conditioning temperature during pelleting decreased performance compared to mash feed despite similar in vitro activity