Organophosphates cholinesterase inhibition detection

Tests for specific classes of chemicals include the measurement of transmitter stimulated adenyl cyclase and Na/K-ATPase for chemicals that affect receptor function or cholinesterase inhibition for organophosphates or carbamates. Electrophysiological techniques may detect chemicals such as DDT or pyrethroids, which affect impulse transmission. 

Several detection methods are based on the biological activities of certain compounds. Cholinesterase inhibiting pesticides (e.g., organophosphates, carbamates) are detected sensitively by treating the layer with the enzyme and a suitable substrate, which react to produce a colored product over the entire layer except where colorless pesticide zones are located due to their inhibition of the enzyme-substrate reaction. 

Organophosphates, thiocarbamates, carbamates, carbamoyloximes, dithiocarbamates, and ureas were included among 100 pesticides and metabolites detected on TLC plates by their cholinesterase inhibiting properties. After developing the plates in ether, xylene, di-n-butyl ether, n-butyl acetate or methyl-isobutyl ketone, the plates were exposed to bromine vapor. The compounds were oxidized to their oxo derivatives, which exhibited more effective cholinesterase inhibitory activity. Bovine liver suspension served as enzyme source and the substrates 2-naphthyl acetate and Fast Blue B salt as the chromogenic agents (163a). 

Hydraulic fluids themselves cannot be measured in blood, urine, or feces, but certain chemicals in them can be measured. Aliphatic hydrocarbons, which are major components of mineral oil hydraulic fluids and polyalphaolefin hydraulic fluids, can be detected in the feces. Certain components of organophosphate ester hydraulic fluids leave the body in urine. Some of these fluids inhibit the enzyme cholinesterase. Cholinesterase activity in blood can be measured. Because many other chemicals also inhibit cholinesterase activity in blood, this test is not specific for organophosphate ester hydraulic fluids. This test is not available at most doctor s offices, but can be arranged at any hospital laboratory. See Chapters 2 and 6 for more information. 

Disulfoton and its breakdown products can be measured in the blood, urine, feces, liver, kidney, or body fat of exposed people. In cases of occupational or accidental exposure to disulfoton, the breakdown products are often measured in the urine. The breakdown products are relatively specific for disulfoton and a few other similar organophosphate pesticides and can be detected in urine for up to one week after people were last exposed. Because disulfoton inhibits cholinesterase in blood and in blood cells, inhibition of this enzyme activity may also suggest exposure to disulfoton. Cholinesterase activity in blood and in blood cells may remain inhibited for as long as 1-2 weeks after the last exposure. Because other organophosphate pesticides also inhibit cholinesterase activity in blood and blood cells, this test is not specific for disulfoton. The measurement of cholinesterase in blood and blood cells and the amount of disulfoton breakdown products in the urine cannot always predict how much disulfoton you were exposed to. Your doctor can send samples of your blood or urine to special laboratories that perform these tests. Chapters 2 and 6 provide more information about medical tests. 

Method. A diagram of the apparatus is shown in Fig.4.29. Any suitable liquid chromatograph may be used. The AutoAnalyzer is modified such that the liquid sampler is fitted to the end of the chromatographic column. The proportioning pump is by-passed. The set-up of the AutoAnalyzer is the same as that for normal measurements of cholinesterase. The application of this technique to the determination of CGA 18809 in plum-leaf extract is shown in Fig.4.30. A comparison is made with UV analysis of the same extract. The limit of detection for CGA 18809 is c . 20 ng at a 3 1 signal to noise ratio. The relative inhibitions of several organophosphates and carbamates are compared in Table 4.9. Diazoxon may be detected in low picogram quantities. 

The detection of organophosphate and other pesticides based on the inhibition of the enzyme acetylcholinesterase by these compounds has received considerable attention primarily due to high specificity and sensitivity [1,7-16]. Cholinesterases, such as acetylcholinesterase,... 

This method of pesticide detection is based on the inhibition of cholinesterase activity. This is a non-specific measurement and as such cannot determine which pesticide the enzyme electrode had been exposed to. This protocol has described a method for extraction and detection of diazinon, but other organophosphate pesticides can be substituted (e.g. chlorfenvinphos) bearing in mind that the enzyme has different inhibition constants for other pesticides and quantities may need to be altered. 

Methods have been published that allow the classification of two types of esterases, the carboxylic ester hydrolases (CEHs) and the phosphoric triester hydrolases (PTEHs) (Anspaugh and Roe, 2004). The CEHs contain the B-esterases, which are inhibited by organophosphates. B-esierases include many other esterases, such as CarbE, acetylcholinesterase (AChE), cholinesterases (ChE), aryleslerases, sterol esterases, insect juvenile hormone esterases, aixl others. The determination of A-esterases uses a protocol for the detection of PTEHs. The PTEH assay allows for the identification of two subclasses of esterases, the A-esterase (known as aiyldialkylphos-phatase) and ditsopropyl fluorophosphatase. Both these enzymes metabolize OP compounds. 

Selective behavior monitors specific toxicity and employs enzyme inhibition tests and reporter genes tests. Examples, are use of cholinesterase for organophosphates and urease for heavy metal ions detection and also transgenic microorganism for specific analyte assessment. 

The cholinesterase electrode was constructed on this basis for the total determination of organophosphates and carbamates [32]. This electrode is also sensitive to other toxic compounds with very low detection limits (paraquat (3 ppm) trichlorophenol (20 ppm) methylazinphos (150 ppb) lindane (15 ppm) and mercury (4 ppm)) [135]. This type of biosensor uses enzymatic inhibition and can be incorporated in an automated system for the quality control of water, and, notably, waste water [291]. 

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