Determination of Selected Heavy Metals Using Amperometric Horseradish Peroxidase (HRP) Inhibition Biosensor

Abstract

Fabrication of an amperometric biosensor for determination of heavy metals in tap water based on the inhibition of horseradish peroxidase activity is reported. The inhibition of the biosensor's response to 0.95 mM H₂O₂ at −200 mV (vs. Ag/AgCl) followed the order, Cd²⁺ < Cu²⁺ < Pb²⁺. The detection limits were 0.09 ppb, 0.03 ppb, and 0.10 ppb for Cd²⁺, Pb²⁺, and Cu²⁺, respectively. Kinetic parameters (apparent Michaelis-Menten constant) and I_max (maximum current) obtained in the absence ( = 1.16 mM, I_max = 0.51 µA) and the presence ( = 1.13 mM, I_max = 0.36) of 3.13 ppm Cd²⁺ showed that the enzyme inhibition process was reversible and non-competitive, and the corresponding K_I (inhibition constant) value was 0.026 mM. After two weeks, the biosensor retained >95% of its initial response and gave a reproducibility of 3.8% and precision of 2.1%.

Keywords:

• Amperometric biosensor
• Heavy metal
• Horseradish peroxidase
• Inhibition
• Polyaniline

Acknowledgments

This paper is part of a Special Issue of Analytical Letters focusing on papers presented at the 10th International Symposium on Kinetics in Analytical Chemistry (KAC-10).

Philiswa Nomngongo is grateful for NRF funding through both Dr. Anna Soares of University of Kwazulu Natal and Prof. Emmanuel Iwuoha of the University of Western Cape, NRF Grand-holders. She also acknowledges the Sensor Lab at University of Western Cape where part of the work for this study was carried out.

Notes

Concentrations were determined in ppb; ND: not detected, ±standard error (, where s is the sample standard deviation and n is the size of the sample).