Abstract
A polymer-based amperometric enzyme electrode (Pt/PoPD/ChOx) was prepared for the amperometric detection of free cholesterol. Firstly, poly(o-phenylenediamine) (PoPD) polymer film was prepared in acetonitrile-water medium containing o-phenylenediamine (oPD) monomer and (±)-10-camphorsulfonic acid (HCSA) on Pt electrode by the use of cyclic voltammetry technique. Cholesterol oxidase (ChOx) enzyme was immobilized onto Pt/PoPD electrode surface. The determination of cholesterol was performed via monitoring of the oxidation current of enzymatically produced H2O2 at +0.7 V vs. Ag/AgCl. Optimum buffer concentration, pH and working temperature were found as 0.05 mol L-1, 7.5 and 40 oC, respectively. The working range of enzyme electrode to cholesterol was 9.8 × 10-3-11 µmol L-1 and response time 150 s. The effects of possible interferences present in serum samples on response of enzyme electrode were examined. The determination of total cholesterol in serum samples was performed by using proposed Pt/PoPD/ChOx enzyme electrode and results were in good agreement with those obtained by spectrophotometric method.
Highlights
Poly(o-phenylenediamine) films were used for elimination interference and determination of cholesterol in human serum could not be performed by using prepared biosensor.[12,51]
The response time that is defined as the time it takes for the electrode to reach 95% of the steady-state current of the Pt/PoPD/Cholesterol oxidase (ChOx) enzyme electrode was determined at two different cholesterol concentrations
The concentration of total cholesterol of three serum samples were analyzed with the proposed Pt/PoPD/ChOx by standard addition method and results were compared with hospital method (Table 2)
Summary
Pt/PoPD electrode was dipped into 250 μL cholesterol oxidase (ChOx) (2.4 U mL-1) solution containing 5 μL glutaraldehyde (25%) for 12 h at +4 °C for immobilization of enzyme molecules. Current-cholesterol concentration plots of the Pt/PoPD/ChOx enzyme electrode were obtained in four different phosphate buffer concentrations (0.05‐0.2 mol L−1) at a pH value of 7.5.
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