Effects of pH and temperature on mAP activity. AP activity was measured by a spectrophotometric method using p-nitrophenyl phosphate (pNPP). (A) Optimum pH. Enzymatic activity was assayed at 37°C in the presence of 0.5 mM pNPP as a substrate using 50 mM Tris–HCl buffer (pH 7.0–8.5) or 50 mM DEA buffer (pH 8.0 to 10.5). (B) Optimum temperature. Enzymatic activity was assayed at 20–80°C in the presence of 0.5 mM pNPP as a substrate using 50 mM DEA buffer (pH 9.0). (C) Thermal stability. Residual activity of mAP (●) was determined at 37°C after 15 min incubation at temperature ranges from 20°C to 100°C. Bacterial AP (▲) was also assayed under the same conditions. (D) Kinetic studies of mAP. Enzymatic activity was assayed under standard condition with 0.5 mM CaCl2 using a range of pNPP.