Amperometric microbial biosensor for direct determination of organophosphate pesticides using recombinant microorganism with surface expressed organophosphorus hydrolase

An amperometric microbial biosensor for the direct measurement of organopho sphate nerve agents is described. The sensor is based on a carbon paste ele ctrode containing genetically engineered cells expressing organophosphorus hydrolase (OPH) on the cell surface. OPH catalyzes the hydrolysis of organo phosphorus pesticides with p-nitrophenyl substituent such as paraoxon, para thion and methyl parathion to p-nitrophenol. The later is detected anodical ly at the carbon transducer with the oxidation current being proportional t o the nerve-agent concentration. The sensor sensitivity was optimized with respect to the buffer pH and loading of cells immobilized using paraoxon as substrate. The best sensitivity was obtained using a sensor constructed wi th 10 mg of wet cell weight per 100 mg of carbon paste and operating in pH 8.5 buffer. Using these conditions, the biosensor was used to measure as lo w as 0.2 muM paraoxon and 1 muM methyl parathion with very good sensitivity , excellent selectivity and reproducibility. The microbial biosensor had ex cellent storage stability, retaining 100% of its original activity when sto red at 4 degreesC for up to 45 days. (C) 2001 Elsevier Science B.V. All rig hts reserved.