Analytical aspects of enzyme reversible inhibition

A simple graphical method for the determination of reversible inhibition type, inhibition constant ( K i ) and estimation of fifty percent of inhibition I 50 of an enzyme reaction is described. The method consists of plotting experimental data as “degree of inhibition” versus the inhibitor concentration at two or more concentrations of substrate. Diagnosis of inhibition type is based on determination of I 50 and the observation of the shift of the inhibition curves. Relationship between I 50 and inhibition constant K i was discussed. A simplified hyperbolae equation of degree of inhibition showing kinetic orders of 1 and zero at low and high concentrations of inhibitors respectively is proposed. The relative error of inhibitor concentration increased drastically when degree of inhibition reached values of 90%. Examples of published inhibition reports as well as an experimental example of amperometric biosensor based on tyrosinase inhibition by benzoic acid were in agreement with the proposed theoretical approach.