Amperometric determination of high-density lipoprotein cholesterol using polyethylene glycol-modified enzymes and a...

A peroxidase-entrapped and ferrocene-embedded carbon paste (POD-Fc-CP) electrode allows a highly sensitive detection of H2O2 at levels as low as 10 nmol/L with practically no interference by coexisting substances, turbidity or coloration of samples. The electrode was applied to the amperometric determination of high-density lipoprotein (HDL)-cholesterol in a very small volume (1-2 microL) using polyethylene glycol (PEG)-modified cholesterol esterase and cholesterol oxidase without prior precipitation or separation of HDL. PEG-modified enzymes exhibit a selective activity toward HDL-cholesterol in the presence of dextran sulphate and MgCl2 to generate H2O2. The HDL-cholesterol concentrations of human serum samples determined by this method showed a good correlation with those determined by an ordinary spectrophotometric method using PEG-modified enzymes and peroxidase or by a conventional precipitation method.