Diagnosis of Enzyme Inhibition Using Excel Solver: A Combined Dry and Wet Laboratory Exercise

Abstract

In enzyme kinetic studies, linear transformations of the Michaelis–Menten equation, such as the Lineweaver–Burk double-reciprocal transformation, present some constraints. The linear transformation distorts the experimental error and the relationship between x and y axes; consequently, linear regression of transformed data is less accurate when compared with methodologies that use nonlinear regression. However, linear transformations are widely used. Explanations for this are the facility to determine model parameters by hand calculations, and until recently, the use of nonlinear regression was difficult as specialized software was not readily available to most scientists and students. Because utilization of personal computers is widespread, these constraints are no longer applicable. This work describes how to perform nonlinear regression with the Solver supplement of Microsoft Office Excel. It is easy to use and to view the results graphically. The F-test was applied to discriminate between models. These methodologies are important in any biochemistry syllabus and can be used to create an active-learning environment where students discriminate between different kinetic models and explore their own experimental results based on several hypotheses.

Repositioning of Thiourea-Containing Drugs as Tyrosinase Inhibitors

“…The simultaneous nonlinear fitting with the data from all concentrations of substrate and inhibitor generated the kinetic parameters in each model with minimized χ 2 value. The χ 2 is defined as the sum of the squared deviations between experimental and fitted values [ 52 ]. Comparison of χ 2 values between the four models by means of F -statistics enabled the selection of the most appropriate model [ 52 , 53 ].…”

A study towards drug discovery for the management of type 2 diabetes mellitus through inhibition of the carbohydrate-hydrolyzing enzymes α-amylase and α-glucosidase by chalcone derivatives

“…The chalcone analogues with methyl (30,36), methoxy (31,37), chloro (32), fluoro (33), and bromo (34) substituents were not able to inhibit α-glucosidase up to the highest tested concentration. As far as we know, these chalcone analogues with two double bonds between the A and B rings were evaluated here for the first time.…”

“…Among the chalcone analogues (29)(30)(31)(32)(33)(34)(35)(36)(37)(38)(39)(40)(41), the last derivative (41) with a 2′-hydroxy group at the A ring and a 2-nitro group at the B ring was almost nine times more active with an IC 50 value of 41 ± 1 μM when compared with the positive control acarbose (Fig. 3).…”

“…where V inic = initial velocity of formation of absorbing CNP in ΔAbsorbance per minute, V max = maximum achievable velocity when for the 0.1 U mL −1 of enzyme had used all the catalytic sites and is saturated by the substrate, S = CNPG3 concentration in mM, K m = Michaelis-Menten constant in mM, K ic = inhibitor dissociation constant of enzyme inhibitor expressed in µM −1 , K iu = inhibitor dissociation constant of enzyme-substrate-inhibitor complex expressed in µM −1 . Data fitting was performed in an Excel Microsoft Office™ spreadsheet using the Solver™ supplement according to Bezerra et al 35 and Dias et al 36 For each tested condition, the equation parameters were estimated by Solver™. The obtained values for the parameters of the simplest model (without inhibition) were used as initial guesses and for the other types of inhibition, competitive, non-competitive, uncompetitive, and finally the more complex model, mixed inhibition.…”

“…In the first set of compounds (1-28), one is unsubstituted (1), four are hydroxylated (2-5), ten are hydroxylated and methoxylated (6)(7)(8)(9)(10)(11)(12)(13)(14)(15), two are only methoxylated (16,17), two display methyl substituents (18,19), two display nitro groups (20,21) and seven possess chloro substituents (22)(23)(24)(25)(26)(27)(28). Among the chalcone analogues (29)(30)(31)(32)(33)(34)(35)(36)(37)(38)(39)(40)(41), one is unsubstituted (29), eleven present a single substitution, including methyl (30,36), methoxy (31,37), chloro (32), fluoro (33), bromo (34), nitro (35,38,39) and hydroxy (40). Compound (41) is di-substituted with a hydroxy group and a nitro group.…”