Abstract
In this study, boroxine derivative (K2[B3O3F4OH]) was tested as an inhibitor of horseradish peroxidase (HRP) by spectrophotometric and electrochemical methods. The activity of horseradish peroxidase was first studied under steady-state kinetic conditions by a spectrophotometric method which required the use of guaiacol as a second substrate to measure guaiacol peroxidation. The results of this method have shown that, by changing the concentration of guaiacol as the literature suggests, a different type of inhibition is observed than when changing the concentration of hydrogen peroxide as the substrate. This suggests that guaiacol interferes with the reaction in some way. The electrochemical method involves direct electron transfer of HRP immobilized in Nafion nanocomposite films on a glassy carbon (GC) electrode, creating a sensor with an electro-catalytic response to the reduction of hydrogen peroxide. The electrochemical method simplifies kinetic assays by removing the requirement of reducing substrates.
Highlights
The boroxine derivative, dipotassium-trioxohydroxytetrafluorotriborate K2 [B3 O3 F4 OH], has lately been intensively studied as a potential enzyme inhibitor
The goal of our research was to compare two methods and to show that, when using the spectrophotometric method, electron donors such as guaiacol could interfere with the inhibitor K2 [B3 O3 F4 OH] and cause problems when determining the type of inhibition
This study showed that the enzyme horseradish peroxidase follows the Michaelis–Menten kinetic model, in the absence and in the presence of the inhibitor K2 [B3 O3 F4 OH]
Summary
The boroxine derivative, dipotassium-trioxohydroxytetrafluorotriborate K2 [B3 O3 F4 OH], has lately been intensively studied as a potential enzyme inhibitor. Horseradish peroxidase is an important heme-containing enzyme and is widely used for analytical purposes, biosensors and enzymatic bioreactors [4,5,6]. The detection of HRP activity is widely used in labelling systems and a large number of procedures for monitoring its reaction have been developed. To monitor HRP activity, substances often referred to as peroxidase substrates need to be added to the reaction mixture. These substrates are electron donors and, when oxidized by HRP with hydrogen peroxide present as the oxidizing agent, a characteristic change that is detectable by the spectrophotometric method takes place [7,8]
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