Control of amylase and protease activities in a phytase preparation by ampholyte-free preparative isoelectric focusing for unrefined cereal-containing bread

Direct addition of a food additive-grade Aspergillus niger phytase preparation to 30% brown rice flour-added bread ingredients reduced the bread myo-inositol hexaphosphate (IP₆) content. The phytase preparation had protease and amylase activities. Addition of the crude phytase preparation induced bread crust collapse. Protease-free high- and low-amylase phytase fractions were prepared using preparative ampholyte-free isoelectric focusing (autofocusing). Addition of the protease-free low-amylase phytase fraction did not significantly affect loaf volume and did not collapse the crust. Addition of the protease-free high-amylase phytase fraction significantly increased loaf volume approximately 1.5× in the presence of amylase at 193–1029 U. However, addition of 1029 and 2058 U of amylase induced partial and whole bread crust collapse, respectively. Therefore, removal of protease and control of amylase activities in the phytase preparation are crucial in the preparation of brown rice flour-added bread with a low myo-inositol phosphate (IP₆) and good swelling property.