Development of capacitance and impedance measurement instrumentation system for real time enzyme kinetics

Abstract

The electrochemical methods are direct, precise, and quick measurement for the target analyte which overcomes the limitation of popular methods (HPLC, Mass spectrometry, Radiometric, fluorescence, MALDI-TOF, etc.) using enzyme systems for many applications. Recent developments in analytical instrumentation with electrical properties of impedance and capacitance at the sensitivity levels desired for measuring enzymatic reactions are now opening up new and versatile methods for real-time, endpoint, and kinetic analysis. Many enzymatic reactions are involved in the ion generation or depletion that leads to the change in capacitance or impedance. The charge- based enzymatic reactions would lead to a change in the charge status (either charged substrate or product) of the reaction during the course of a reaction. Real-time measurements are possible in capacitance and impedance measurement for the charge-based enzymatic reaction. In recent times, Electrochemical Impedance Spectroscopy (EIS) is an emerging technique for the development of enzyme biosensors. We have employed enzymes such as alkaline phosphatase, urease, apyrase, glucose oxidase, and esterase. These charge-based enzymatic reactions liberate the charged products of phosphate (PO43-), acetate (CH3COO-), and ammonia (NH4+) ions in the solution. These enzymes are well known for the charge-based enzymatic reactions and are commonly used for many applications in electrochemical measurement. The change in the ionic concentration during the course of the reaction has the electrical signal for the measurement. In this work, we demonstrated the liberated ions from the enzymatic reaction that causes the change of capacitance or impedance with respect to time in a fabricated prototype instrument. Capacitance measurement shows the enzyme newline