Purification and Characterization of Endogenous α-Amylase from Glutinous Rice Flour

Endogenous α-amylase activity is crucial for determining the end-use value of glutinous rice flour (GRF), and controlling it is a key goal in the milling process. Although the structure and properties of starch and protein in GRF have been extensively studied, there is little information on endogenous α-amylase in GRF. In this study, endogenous α-amylase isolated from GRF was purified and characterized. It was found to have a molecular weight of about 32 kDa, with the highest specific activity at 60 °C and a pH of 6.0. The enzyme is stable below 50 °C and in the pH range of 4.0-7.0. Its activity is Ca²⁺-independent but inhibited by Cu²⁺, Zn²⁺, Mg²⁺, Mn²⁺, and Ba²⁺. Its activity is also reduced by β-mercaptoethanol. The enzyme hydrolyzes amylopectin most efficiently. Circular dichroism spectroscopy showed that the enzyme contains 7.9% α-helix, 35.4% β-folding, 21.1% β-turning, and 35.9% random coils, with a T_m value of 63.68 °C. These results suggest that temperature control may be the best strategy for reducing amylase activity in dry-milled GRF, providing a new approach for the development of GRF dry-milling techniques.