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Biomarkers of effect from exposure to mixtures of organophosphate pesticides
Project Code: T10002
Health and Safety Lab (HSL)
The project’s overall objective was to investigate the possible development of new biomarkers of effect for organophosphate pesticide exposure. Such novel biomarkers should have better sensitivity than the historically used effect biomarker of looking for OP-induced depression in the activity of two blood enzymes, namely plasma butyrylcholinesterase (pChE) and erythrocyte acetylcholinesterase. Three main strands of investigation were encompassed within the overall objective.
- The possible development of an immunoassay for OP-modified pChE or AChE based on monoclonal antibodies preferentially binding the OP-modified protein in comparison to the native, active enzyme. OP-modified pChE or AChE being used as the antigen to raise antibodies.
- The possible development of an immunoassay that detects the active site or serine residue within the active site after organophosphorylation by OP pesticides. Here a peptide sequence around the serine within the active site of the enzyme and modified by OPs is used as immunogen to raise antibodies.
- The development of immunochemical or chromatographic assays for the measurement of organophosphorylated free serine residues. We hypothesise that such species may be formed from the turnover of pChE, AChE and importantly any other proteins that may undergo organophosphorylation at serine residue. Such abnormal amino-acids could not be reused in the amino-acid pool and therefore are likely to be excreted in urine.
This report details work funded by the Food Standards Agency (FSA) as project T10002 on novel biological effect markers of organophosphate pesticide exposures. The project builds upon HSL’s previous and on-going work in the development and use of biological markers of exposure and effect for pesticides, notably the anti-cholinergic organophosphate (OP) pesticides. HSL has undertaken for many years routine biological effect monitoring for OP pesticides, based on blood cholinesterase measurements, and also biological monitoring based on the urinary measurement of alkyl phosphates. These measurements have been applied by HSL for the routine monitoring of occupationally exposed workers, those with accidental OP exposure and in controlled human volunteer studies [1-9], and they have helped define the comparable levels of sensitivity of these monitoring methods [1, 3]. Previous attempts at improvement in terms of sensitivity to biological effect monitoring methods based on blood cholinesterase measurements have been based on using specific activity rather than simple measurement of activity changes [10-14]. This involves ratioing the enzyme activity of plasma butyrylcholinesterase (pChE) to its plasma concentration measured by immunoassay. This is to reduce the influence of intra-individual variation of pChE synthesis and degradation rates on the detection of enzyme inhibition through organophosphorylation of a key serine residue at the enzyme’s active site. However, this technique has not been shown to significantly improve the ability to detect low level OP exposure within occupational monitoring schemes. Rather it reduces the reliance on comparison against baseline/unexposed activity measurements in the individual in detecting exposure.
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