Derivation of Kinetic Parameter Dependent Model for the Quantification of the Concentration and Molar Mass of an Enzyme in Aqueous Solution: A Case Study on Aspergillus oryzea α-amylase

Aim: The objective of the research was to verify a simple mathematical model for the determination of mass concentration and the molar mass of an enzyme in any solution using standard solution of Aspergillus oryzea alpha amylase and the action of A. oryzea alpha amylase on heat treated soluble potato starch.

Study Design: Experimental.

Place and Duration of Study: Chemistry & Biochemistry Department, Research Division of Ude International Concepts limited (RC: 862217). The research lasted for about 4 months between May and Sep, 2015, on a non – continuous basis.

Methodology: Bernfeld method of enzyme assay was used to generate data on catalytic activity of A. oryzea alpha amylase. One stock solution of the enzyme was prepared by dissolving 0.01 g of the enzyme in 100 ml of tris – HCl buffer, at pH 5 and temperature 20°C and at pH 6.9 and temperature 37°C. In each case the stock solutions were diluted to the desired concentrations.

Results: The calculated mass concentration (Emp[E]) and the calculated molar mass (EmpMPROT) of the enzyme were 107.99±7.32 mg/l and 54.58±4.87 kDa respectively at pH, 6.9 and temperature, 37°C (all mean values were from 12 calculations); The Emp[E] and EmpMPROT values were 101.47±8.06 mg/l and 51.68±5.98 kDa respectively, at pH 5 and temperature 20°C (all mean values were from 11 calculations).

Conclusion: The model can be used to determine the mass concentration of the enzyme in its crude extract and as a corollary, its molar mass can also be determined given accurate concentration and the strict adoption of the protocol described.