Shankar Balasubramanian1, Valber Pedrosa1, Sheetal Paliwal1, Virginia A. Davis2, James Wild3, and Aleksandr Simonian1. (1) Materials Research and Education Center, Auburn University, Wilmore, Auburn, AL 36849, (2) Department of Chemical Engineering, Auburn University, Auburn, AL 36849, (3) Biochemistry and Biophysics, Texas A & M University, 103 Biochemistry/Biophysics Building, Texas A & M University, 2128 TAMU, College Station, TX 77843
A simple approach is described for design and development of highly sensitive and stable electrochemical biosensor for direct detection of organophosphate pesticides. Organophosphorus hydrolase (OPH) functionalized single and multi-walled carbon nanotube (CNT) conjugates provided the platform for direct amperometric detection of Paraoxon, a model organophosphate. The catalytic hydrolysis of paraoxon produces equimoles of p-nitrophenol whose oxidation was monitored amperometrically in real time under flow-injection (FIA) mode. Buffer pH, flow rate and method of OPH immobilization were found to influence the sensitivity of paraoxon detection. The dynamic concentration range was 0.5 to 8.5 mmol L-1 with a detection limit of 2.3 mmol L-1 (S/N = 3). Our studies show that immobilized enzyme retains a significant degree of enzymatic activity, provided high sensitivity and showed remarkable stability with only 8% decrease in signal over 10-day period. These results suggest that covalent immobilization of OPH on CNT can be used for specific immobilization of OPH with advantages of low price, simplicity and biocompatibility.