Amperometric cholesterol biosensor based on the direct electrochemistry of cholesterol oxidase and catalase on a graphene/ionic liquid-modified glassy carbon electrode

Cholesterol oxidase (ChOx) and catalase (CAT) were co-immobilized on a graphene/ionic liquid-modified glassy carbon electrode (GR-IL/GCE) to develop a highly sensitive amperometric cholesterol biosensor. The H2O2 generated during the enzymatic reaction of ChOx with cholesterol could be reduced electrocatalytically by immobilized CAT to obtain a sensitive amperometric response to cholesterol. The direct electron transfer between enzymes and electrode surface was investigated by cyclic voltammetry. Both enzymes showed well-defined redox peaks with quasi-reversible behaviors. An excellent sensitivity of 4.163 mA mM(-1) cm(-2), a response time less than 6 s, and a linear range of 0.25-215 mu M (R-2 > 0.99) have been observed for cholesterol determination using the proposed biosensor. The apparent Michaelis-Menten constant (K-M(app)) was calculated to be 2.32 mM. The bienzymatic cholesterol biosensor showed good reproducibility (RSDs < 5%) with minimal interference from the coexisting electroactive compounds such as ascorbic acid and uric acid. The CAT/ChOx/GR-IL/GCE showed excellent analytical performance for the determination of free cholesterol in human serum samples. (C) 2013 Elsevier B.V. All rights reserved.