Higher precision in initial rates may be achievable: A test of a pseudo-statistical method

Abstract

There has been a concerted effort at establishing the best method for the measurement of initial rates for various purposes, including the calculation of kinetic parameters, the maximum velocity (Vmax), and the Michaelis-Menten constant (KM). The objectives of this research are: 1) to derive equations without KM for the determination of the Vmax in particular and vice versa; 2) to determine the KM and Vmax with other equations other than the Michaelian equation; and 3) to subject the calculated and extrapolated kinetic parameters to pseudo-statistical remediation where necessary as a test of their viability and usefulness. The study was experimental and theoretical. It is supported by the Bernfeld method of enzyme assay. By graphical means, the Vmax and KM values for galactosidase respectively range between 163 and 185 mM/min and between 2.07 and 2.77 mg/L; the range by calculations is 177 and 214 mM/min and 2.45 and 3.311 mg/L, subject to pseudo-statistical remediation. Overall, the ranges of Vmax and KM values for alpha-amylase from both the graphical method and calculation are, respectively, 1.095 to 1.018 mM/min and 18.15 to 20.554 g/L. The equations for the determination of the KM and Vmax, which are respectively invariant with respect to each other, were rederived. The initial rates must not be characterised by a mixture of conditions that validate different quasi-steady-state assumptions if the true KM and Vmax are of interest. The new pseudo-statistical method for the remediation of error in all measurements, if necessary, is viable, useful, and robust.