Application of the Enzymatic Electrochemical Biosensors for Monitoring Non-Competitive Inhibition of Enzyme Activity by Heavy Metals.

Ashrafi Ashrafi,Adam Adam,Sýs Sýs,Farag Farag,Přibyl Přibyl,Richtera Richtera,Sedláčková Sedláčková

doi:10.3390/s19132939

Abstract

The inhibition effect of the selected heavy metals (Ag+, Cd2+, Cu2+, and Hg2+) on glucose oxidase (GOx) enzyme from Aspergillus niger (EC 1.1.3.4.) was studied using a new amperometric biosensor with an electrochemical transducer based on a glassy carbon electrode (GCE) covered with a thin layer of multi-wall carbon nanotubes (MWCNTs) incorporated with ruthenium(IV) oxide as a redox mediator. Direct adsorption of multi-wall carbon nanotubes (MWCNTs) and subsequent covering with Nafion® layer was used for immobilization of Gox. The analytical figures of merit of the developed glucose (Glc) biosensor are sufficient for determination of Glc in body fluids in clinical analysis. From all tested heavy metals, mercury(II) has the highest inhibition effect. However, it is necessary to remember that cadmium and silver ions also significantly inhibit the catalytic activity of Gox. Therefore, the development of Gox biosensors for selective indirect determination of each heavy metal still represents a challenge in the field of bioelectroanalysis. It can be concluded that amperometric biosensors, differing in the utilized enzyme, could find their application in the toxicity studies of various poisons.

Highlights

Enzymes are organic catalysts produced within the living organisms
Nafion® is better than chitosan for the covering of the electrode as it did not cause a significant decrease in the current compared to the glassy carbon electrode (GCE)/multi-wall carbon nanotubes (MWCNTs)
Glucose oxidase (GOx) obtained from Aspergillus niger ≥100,000 U·g−1 solid (EC 1.1.3.4), β-d-glucose, ruthenium oxide used as mediator, hydrogen peroxide (H2O2), multi-walled carbon nanotubes (MWCNTs), Nafion®, acetic acid (AA), chitosan (~50 kDa) and N,N-dimethylformamide (DMF), ethanol (99.98%), nitric acid (70%), ammonia solution (25%), and paraffin oil were purchased from Sigma-Aldrich

Summary

Introduction

Enzymes are organic catalysts produced within the living organisms. They speed up the biological reactions by lowering the activation energy. They can speed up the conversion of the substrate to the products in cellular metabolism up to 10 million times or more [1]. The conversion of the substrates by enzyme is highly specific. To initiate an enzyme-catalyzed reaction, the enzyme must bind to its substrate forming an enzyme–substrate complex [3]. Considering that the enzymes remain unchanged after the reactions, they catalyze and can be reused. They are effective in a very small amount [4].

Methods

Results

Conclusion