Residues cholinesterase inhibition

Mice that were exposed dermally to residues of methyl parathion in emulsifiable concentrate on foliage, and were muzzled to prevent oral intake, developed inhibition of plasma cholinesterase and erythrocyte cholinesterase after two 10-hour exposures (Skinner and Kilgore 1982b). For the organophosphate pesticides tested in this study, cholinergic signs generally were seen in mice with cholinesterase inhibition >50% results for this end point were not broken down by pesticide. 

The inhibition of brain cholinesterase is a biomarker assay for organophosphorous (OP) and carbamate insecticides (Chapter 10, Section 10.2.4). OPs inhibit the enzyme by forming covalent bonds with a serine residue at the active center. Inhibition is, at best, slowly reversible. The degree of toxic effect depends upon the extent of cholinesterase inhibition caused by one or more OP and/or carbamate insecticides. In the case of OPs administered to vertebrates, a typical scenario is as follows sublethal symptoms begin to appear at 40-50% inhibition of cholinesterase, lethal toxicity above 70% inhibition. 

Presently available methods to diagnose and biomonitor exposure to anticholinesterases, e.g., nerve agents, rely mostly on measurement of residual enzyme activity of acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) in blood. More specific methods involve analysis of the intact poison or its degradation products in blood and/or urine. These approaches have serious drawbacks. Measurement of cholinesterase inhibition in blood does not identify the anticholinesterase and does not provide reliable evidence for exposure at inhibition levels less than 20 %. The intact poison and its degradation products can only be measured shortly after exposure. Moreover, the degradation products of pesticides may enter the body as such upon ingestion of food products containing these products. 

The WHO/FAO Joint Meeting of Experts on Pesticide Residues (JMPR) has given recommendations on interpretation of cholinesterase inhibition (FAO 1998, 1999), see Section 4.7.7.3.1. 

AChEj induction correlated with the body burden of total pesticides in 15 pooled flounder livers at the Benelux tunnel (site 2) and Noordwijk (site 6) locations. The highest concentrations of several OCPs (total 150 00 o,g/kg lipid weight) were found in flounder from the Rotterdam transect (De Boer et ah, 2001). The Rotterdam transect (sites 6 to 2) is characterised by acethyl-cholinesterase inhibition (mainly brain), which may reflect contamination by pesticides in this transect. The concentrations of total OCPs in SPM varied in the Rotterdam and Amsterdam transects from below the detection limit to 1.8 mg kg and 1.6 mg kg respectively. Rotterdam showed relatively higher levels of OCPs from upstream, caused by one of the major sources of OCP residues in the rivers Meuse and Rhine. The eontamination might be classified as historical, or industrial centres might still be emitting these eompounds (Voorspoels et al., 2004). 

The reactivator may form, either with the inhibitor or with its residue in inhibited cholinesterase, a stable secondary inhibitor of cholinesterase. 

The coupling of HPLC with a cholinesterase-inhibition AutoAnalyzer for the determination of organophosphate and carbamate insecticides has much potential for the routine screening of residues of these compounds [58]. 

Methods of Assessing Cholinesterase Inhibition. The increased use of cholinesterase-inhibiting insecticides has stimulated research in many areas of scientific endeavor. One such area has been concerned with the in vivo toxicological properties of cholinesterase inhibitors. However, the area of concern here is in the field of analytical chemistry where cholinesterases are used as a tool for the quantitative determination of unknown amounts of inhibitors (8). Such procedures are frequently used for the analysis of certain pesticide residues and can be categorized into the following types of methods ... 

WHO/FAO Guidelines for Cholinesterase-Inhibiting Pesticide Residues in Food... 

Birds killed by organophosphorus compounds in the wild consistently show 80-95% depression of brain-cholinesterase activity. Depression of brain-cholinesterase activity by >20% in birds has been used as a conservative criterion to indicate significant exposure to organophosphorus chemicals. Depression of brain-cholinesterase activity by >50% and confirmation of suspected organophosphorus chemical residues in tissues or ingesta are criteria for cause-effect diagnosis of death in birds exposed to cholinesterase-inhibiting chemicals. Death occurs in many avian species... 

This paper summarizes the development and application of both a philosophic and quantitative framework for unifying research approaches and findings in residue decay, exposure assessment, and cholinesterase response (Popcndorf Lefflngwell. Res. Rev. 82 125, 1982). Examples are provided for using this model to Interpret the potential cholinesterase response from a known foliar residue and to establish reentry intervals to prevent excessive cholinesterase Inhibition. The potential and limitations of extrapolating this approach to other settings is also discussed, as are the needs for future research to support a comprehensive approach to pesticide use, residues, and exposure controls. 

Organophosphate and carbamate pesticides are potent inhibitors of the enzyme cholinesterase. The inhibition of cholinesterase activity by the pesticide leads to the formation of stable covalent intermediates such as phosphoryl-enzyme complexes, which makes the hydrolysis of the substrate very slow. Both organophosphorus and carbamate pesticides can react with AChE in the same manner because the acetylation of the serine residue at the catalytic center is analogous to phosphorylation and carbamylation. Carbamated enzyme can restore its catalytic activity more rapidly than phosphorylated enzyme [17,42], Kok and Hasirci [43] reported that the total anti-cholinesterase activity of binary pesticide mixtures was lower than the sum of the individual inhibition values. 

Other serine hydrolases such as cholinesterases, carboxylesterases, lipases, and fl-lactamases of classes A, C, and D have a hydrolytic mechanism similar to that of serine peptidases [25-27], The catalytic mechanism also involves an acylation and a deacylation step at a serine residue in the active center (see Fig. 3.3). All serine hydrolases have in common that they are inhibited by covalent attachment of diisopropyl phosphorofluoridate (3.2) to the catalytic serine residue. The catalytic site of esterases and lipases has been less extensively investigated than that of serine peptidases, but much evidence has accumulated that they also contain a catalytic triad composed of serine, histidine, and aspartate or glutamate (Table 3.1). 

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