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ORGANOPHOSPHORUS HYDROLASE-BASED ASSAY FOR ORGANOPHOSPHATE PESTICIDES
Citation:
Rogers, K R., Y. Wang, A. Mulchandani, P. Mulchandani, AND W. Chen. ORGANOPHOSPHORUS HYDROLASE-BASED ASSAY FOR ORGANOPHOSPHATE PESTICIDES. BIOTECHNOLOGY PROGRESS 15(3):517-521, (1999).
Impact/Purpose:
The overall objective of this task is to develop scientifically sound sampling and bioanalytical approaches for screening and monitoring of hazardous wastes. These techniques are expected to provide the Agency with improved screening and field portable methods to characterize, reduce, and control risk to human health and the environment. Specific objectives will include development and characterization of the following concepts:
SPMDs for passive accumulation of TICs
Bioassays for toxic and genotoxic compounds
MIPs for volatile and semivolatile toxic organics
Rapid screening assays using the previously listed components.
Description:
We report a rapid and versatile Organophosphorus hydrolase (OPH)-based method for measurement of organophosphates. This assay is based on a substrate-dependent change in pH at the local vicinity of the enzyme. The pH change is monitored using fluorescein isothiocyanate (FITC), which is covalently Immobilized to the enzyme. This method employs the use of poly(methyl methacrylate) beads to which the FITC-labeled enzyme is adsorbed. Analytes were then measured using a microbead fluorescence analyzer. The dynamic concentration range for the assay extends from 25 to 400 uM for paraoxon with a detection limit of 8 uM. Organophosphorus insecticides measured using this technique included ethylparathion, methylparathion, dursban, fensulfothion, crotoxyphos, diazinon, mevinphos, dichlorvos, and coumaphos. This technique was used to measure coumaphos in biodegradation samples of cattle dip wastes and showed a high correlation (r2 = 0.998) to an HPLC method.
The U.S. Environmental Protection Agency (EPA), through its Office of Research and Development (ORD), funded the work involved in preparing this article. It has been subject to the Agency's peer review and has been approved for publication. Mention of trade names or commercial products does not constitute endorsement or recommendation by the U.S. EPA. This work was supported, in part, by EPA Grant R823663-01-0 (to A.M. through UCR).