Amperometric Cholesterol Determination Using HRP Incorporated Carbon Paste Electrode

Vikas Hooda*and Anjum Gahlaut

Centre for Biotechnology, Maharshi Dayanand University, Rohtak-124001, Haryana, India Corresponding Author E-mail : advance.biotech@gmail.com

DOI : http://dx.doi.org/10.13005/bbra/2809

ABSTRACT: Elevated level of serum cholesterol has become an utmost risk factor for several diseases. A method has been described for the construction of an HRP incorporated carbon paste based amperometric biosensor for cholesterol detection mounted on cellulose acetate (CA) membrane bounded with enzymes cholesterol esterase and cholesterol oxidase. Electrochemical measurements were implemented with Ag / AgCl as the reference electrode and silver wire as the auxiliary electrode under a capacity of −50mV. The optimum pH of the biosensor was pH 7.0. The incubation temperature and reaction time displayed in a wide temp range from 35 °C to 50 °C was 25 seconds. The linear relationship shown by the sensor was in range from 2.5 to 60 mg dL^-1 of the substrate, cholesteryl acetate and amount of current (mA). The correlation coefficient, r was 0.9916 for n=20, being compared to standard colorimetric methods. The mean analytical recovery observed was 99.24±0.81 & 99.12±0.84 after addition of 100mg dL^-1 and 200mg dL^-1 cholesteryl acetate in serum samples respectively. Coefficient of variation (CV) calculated were <2% & <4 % within and between batch respectively. The present working electrode retained 50% of its initial activity when used continuously for 180 times over 35 days, stored in sodium phosphate buffer, (0.1 M, pH 7.0) at 4^oC. There were no practical interferences observed in the present method.

KEYWORDS: Cholesterol Esterase; Cholesterol Oxidase; Carbon Paste Electrode; Cellulose Acetate; Horseradish Peroxidase

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