Amperometric Determination of Serum Cholesterol with Pencil Graphite Rod
Nidhi Chauhan, Jagriti Narang, Chandra S. Pundir
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DOI: 10.4236/ajac.2010.12006   PDF    HTML     6,602 Downloads   13,088 Views   Citations

Abstract

A cholesterol oxidase from Streptomycin sp. was immobilized onto pencil graphite rod and employed for amperometric determination of serum cholesterol. The method has the advantage over earlier amperometric methods that it requires low potential to generate electrons from H2O2, which does not allow ionization of serum substances. The optimum working conditions of amperometric determination were pH 6.8, 25?C and 30 s. The current measured was in proportion to cholesterol concentration ranging from 1.29×10-3 to 10.33×10-3 M. Minimum detection limit of the method was 0.09 ×10-3 M. Mean analytical recovery of added cholesterol (100 mg/dl and 200 mg/dl) in serum was 85.0% & 90.0% respectively. Within batch and between batch coefficients of variations were 1.59% & 4.15% respectively. A good correlation (r = 0.99) was obtained between serum cholesterol values by standard enzymic colorimetric method and the present method. No interference by metabolites was observed in the method. The enzyme electrode was reused 200 times over a period of 25 days, when stored at 4?C

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N. Chauhan, J. Narang and C. Pundir, "Amperometric Determination of Serum Cholesterol with Pencil Graphite Rod," American Journal of Analytical Chemistry, Vol. 1 No. 2, 2010, pp. 41-46. doi: 10.4236/ajac.2010.12006.

Conflicts of Interest

The authors declare no conflicts of interest.

References

[1] S. Hirany, D. Li and I. Jialal, “A More Valid Measurement of Low Density Lipoprotein Cholesterol in Diabetic Patients”, American Journal of Medicine, Vol. 102, No. 1, 1997, pp. 48-53.
[2] C. C. Allain, L.S. Poon, C. S. G. Chan, W. Richmond and P.C. Fu, “Enzymatic Determination of Serum Total Cholesterol,” Clinical Chemistry, Vol. 20, No. 4, 1974, pp. 470-475.
[3] H. Osman and Kwee Yap, “Comparative Sensivity of Cholesterol Analysis Using GC, HPLC and Spectrophotometric Methods,” Malaysian Journal of Food Science, Vol. 10, No. 2, 2006, pp. 205-210.
[4] D. L. Witte, D. A. Barrett and D. A. Wycoff, “Evaluation of an Enzymatic Procedure for Determination of Serum Cholesterol with the Abbott ABA-100,” Clinical Chemistry, Vol. 20, No. 10, 1974, pp. 1282-1286.
[5] I. Karubeet, K. Hera, H. Matsuoka and S. Suzuki, “Amperometric Determination of Total Cholesterol in Serum with Use of Immobilized Cholesteriol Esterase and Cholesterol Oxidase,” Analytical Chimica Acta, Vol. 139, No. 10, 1982, pp. 127-132.
[6] B. Shahnaz, S. Tada, T. Kajikawa, T. Ishida and K. Kawanishi, “Automated Fluimetric Determination of Cellular Cholesterol,” Annals of Clinical Biochemistry, Vol. 35, No. 6, 1998, p. 665.
[7] M. J. Tabata and T. Murachi, “Automated Analysis of Total Cholesterol in Serum Using Coimmobilized Cholesterol Ester Hydrolase and Cholesterol Oxidase,” Journal of Applied Biochemistry, Vol. 3, 1981, pp. 84-92.
[8] Suman and C. S. Pundir, “Co-immobilization of Cholesterol Esterase, Cholesterol Oxidase & Peroxidase onto Alkylamine Glass Beads for Measurement of Total Cholesterol in Serum,” Current Applied Physics, Vol. 3, No. 2-3, 2003, pp. 129-133.
[9] V. Hooda, A. Gahlautb, H. Kumar and C. S. Pundir, “Biosensor Based on Enzyme Coupled PVC Reaction Cell for Electrochemical Measurement of Serum Total Cholesterol,” Sensor and Actuator B, Vol. 136, No. 1, 2009, pp. 235-241.
[10] L. Braco, J. A. Daros and M. de la. Guardia, “Enzymatic Flow Injection Analysis in Nonaqueous Media,” Analytical Chemistry, Vol. 64, No. 2-3, 1992, pp. 129-133.
[11] B. Sean, P. Narine, D. Singh and A. Guiseppi-Elie, “Amperometric with in Determination of Cholesterol in Serum Using A Biosensor of Cholesterol Oxidase Contained Polypyrol-Hydrogel Membrane,” Analytical Chimica Acta, Vol. 448, No. 1-2, 2001, pp. 27-36.
[12] P. Nuria, Gloria Ruiz, A. Julio Reviejo and Jose M. Pingarron, “Graphite Teflon Composite Bienzyme Electrodes for the Determination of Cholesterol in Reversed Micelles: Application to Food Samples,” Analytical Chemistry, Vol. 5, No. 6, 2001, pp. 1190-1195.
[13] L. Jianping, T. Peng and Y. Peng, “Cholesterol Biosensor Based on Entrapment of Cholesterol Oxidase in a Silicic Sol-Gel Matrix at a Prussian Blue Modified,” Electroanalysis, Vol. 15, No. 12, 2003, pp. 1031-1033.
[14] H. Endo, M. Masashi, T. Mio, R. Huifeng, H. Tetushito, U. Naoto and M. Kohji, “Enzyme Sensor System for Determination of Total Cholesterol in Fish Plasma,” Fisheries Science, Vol. 69, No. 6, 2003, pp. 1194-1199.
[15] S. Singh, A. Chaubey and B. D. Mahlotra, “Amperometric Cholesterol Biosensor Based on Immobilized Cholesterol Esterase and Cholesterol Oxidase on Conducting Polypyrol Films,” Analytical Chimica Acta, Vol. 502, No. 2, pp. 229-234, 2004.
[16] M. J. Song, D. H. Yun, J. H. Jin, N. K. Min and S. Hong, “Comparison of effective working electrode area on planar and porous silicon substrates for cholesterol biosensor,“ Japanese Journal of Applied Physics, Vol. 45, No. 9A, 2006, pp. 7197-7202.
[17] ?. B. Cem, ?. Haluk, Celebi, S. Serda and Y. Atilla, “Amperometric Enzyme Electrode for Free Cholesterol Determination Prepared with Cholesterol Oxidase Immobilized in Poly(Vinylferrocenium) Film,” Enzyme Microbial Technology, Vol. 40, No. 2, 2007, pp. 262-265.
[18] W. Gao, J. Song and N. Wu, “Voltametric Behaviour and Square-Wave Voltametric Determination of Trepibutone at a Pencil Graphite Electrode,” Journal of Electroanalytical Chemistry, Vol. 576, No. 1, 2005, pp. 1-7.
[19] M. D. Vestergaard, K. Kerman and E. Tamiya, “An Electrochemical Approach for Detecting Copper-Chelating Properties of Flavonoids Using Disposable Pencil Graphite Electrodes: Possible Implications in Copper-Mediated Illnesses,” Analytical Chimica Acta, Vol. 538, No. 1-2, 2005, pp. 273-281.
[20] A. M. Bond, J. Peter, Mahon, J. G. Schiewe and V. Vicente-Beckett, “An Inexpensive and Renewable Pencil Electrode for Use in Field Based Stripping Voltammetry,” Analytical Chimica Acta, Vol. 345, No. 1-2, 1997, pp. 67- 74.
[21] B Erable, N. Duteanu, S. M. Kumar, Y. Feng, M. M. Ghangrekar and K. Scott, “Nitric Acid Activation of Graphite Granules to Increase the Performance of the Non-Catalyzed Oxygen Reduction Reaction (ORR) for MFC Applications,” Electrochemistry Communication, Vol. 11, No. 7, 2009, pp. 1547-1549.
[22] J. C. Vidal, J. Espuelas and J. R. Castillo, “Amperometric Cholesterol Biosensor Based on In Situ Reconstituted Cholesterol Oxidase on an Immobilized Monolayer of Flavin Adenine Dinucleotide Cofactor,” Analytical Biochemistry, Vol. 333, No. 1, 2004, pp. 88-98.
[23] A. Kumar, R. R. Pandey and B. Brantley, “Tetraethylorthosilicate Film Modified with Protein to Fabricate Cholesterol Biosensor,” Talanta, Vol. 69, No. 3, 2006, pp. 700-705.
[24] A. Noma and K. Nakayama, “Polarographic Method for Rapid Micro Determination of Cholesterol with Cholesterol Esterase and Cholesterol Oxidase,” Clinical Chemistry, Vol. 22, No. 3, 1976, pp. 336-40.
[25] T. Yao, M. Sato, Y. Kobayashi and T. Wasa, “Amperometric Assays of Total and Free Cholesterol in Serum by the Combined Use of Immobilized Cholesterol Esterase and Cholesterol Oxidase Reactors and Peroxidase Electrode in Flow Injection System,” Analytical Biochemistry, Vol. 149, 1985, pp. 387-391.
[26] H. S. Huang, S. S. Kuan and G. G. Guiblault, “Amperometric Determination of Total Cholesterol in Serum, with Use of Immobilized Cholesterol Ester Hydrolase and Cholesterol Oxidase,” Clinical Chemistry, Vol. 23, No. 1, 1977, pp. 671-676.
[27] R. J. Fernandez, M. M. D. L. De Castro and M. Valcarcel, “Determination of Total Cholesterol in Serum by Flow Injection Analysis with Immobilized Enzyme,” Clinical Chimica Acta, Vol. 31, No. 2, 1987, pp. 97-104.
[28] C. Heuck, I. Erbe and D. Mathias, “Cholesterol Determination in Serum after Fractionation of Lipoproteins by Immunoprecipitation,” Clinical Chemistry, Vol. 31, No. 1-2, 1985, pp. 252-256.
[29] S. Singh, P. R. Solanki and B. D. Malhotra, “Covalent Immobilization of Cholesterol Esterase and Cholesterol Oxidase on Polyaniline Films for Application to Cholesterol Biosensor,” Analytica Chimica Acta, Vol. 568, No. 1-2, 2006, pp. 126-132.

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