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HIGHLY SENSITIVE ASSAY FOR ANTICHOLINESTERASE COMPOUNDS USING 96 WELL PLATE FORMAT
Citation:
Rogers, K R. AND N N. Mishra. HIGHLY SENSITIVE ASSAY FOR ANTICHOLINESTERASE COMPOUNDS USING 96 WELL PLATE FORMAT. Presented at 220th ACS National Meeting, Washington, DC, August 20-24, 2000.
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:
One of the approaches for reducing uncertainties in the assessment of human exposure is to better characterize concentrations of hazardous compounds that may be present in our immediate environment. A significant limitation to this approach, however, is that sampling and laboratory analysis of contaminated environmental and biological samples, can be slow and expensive; thus, limiting the number of samples that can be analyzed within time and budget constraints. Faster, simpler, and more cost-effective field screening methods can increase the amount of information available concerning the location, source and concentration of pollutants present in the environment. Among the compounds of interest to EPA for human exposure assessment are pesticides. More specifically, insecticides from the organophosphate and carbamate classes are widely used in industrial and residential settings. These compounds often show high acute toxicity due to their inhibition of the enzyme acetylcholinesterase (AChE).
Although there have been a variety of chromatographic methods reported for detection of these compounds, these techniques are typically expensive and time-consuming. In addition, there have been a number of bioanalytical and biosensor methods reported based on inhibition of AChE. Although these methods show considerable promise, they are, for the most part, not well suited for screening large numbers of environmental samples. There are also several other issues that must be addressed with respect to the application of an AChE inhibition assay to environmental screening. More specifically, a number of commonly used organophosphate insecticides require metabolic activation to inhibit AChE. In addition, organophosphorus and carbamate insecticides (and their metabolites) vary widely in their ability to inhibit AChE (i.e., their relative response in these inhibition assays). We report a screening assay for organophosphorus and carbamate classes of insecticides that is convenient, rapid, simple and sensitive. Incorporated into the assay protocol is an oxidation step to convert parent insecticides to their oxon forms. In addition to IC50 values, relative inhibition values (compared to paraoxon) are also reported.
The U.S. Environmental Protection Agency (EPA), through its Office of Research and Development (ORD), funded this research and approved this abstract as a basis for an oral presentation. Mention of trade names or commercial products does not constitute endorsement or recommendation of these products by the EPA. The actual presentation has not been peer reviewed by EPA. NNM is currently a National Research Council Fellow.