Cholinesterase methods monitoring

Altstein, M., Segev, G., Aharonson, N., Ben-Aziz, O., Turniansky, A. and Avnir, D. (1998) Sol-gel-entrapped cholinesterases A microtiter plate method for monitoring anticholinesterase compounds. Journal of Agricultural and Food Chemistry, 46, 3318-3324. 

In AChE-based biosensors acetylthiocholine is commonly used as a substrate. The thiocholine produced during the catalytic reaction can be monitored using spectromet-ric, amperometric [44] (Fig. 2.2) or potentiometric methods. The enzyme activity is indirectly proportional to the pesticide concentration. La Rosa et al. [45] used 4-ami-nophenyl acetate as the enzyme substrate for a cholinesterase sensor for pesticide determination. This system allowed the determination of esterase activities via oxidation of the enzymatic product 4-aminophenol rather than the typical thiocholine. Sulfonylureas are reversible inhibitors of acetolactate synthase (ALS). By taking advantage of this inhibition mechanism ALS has been entrapped in photo cured polymer of polyvinyl alcohol bearing styrylpyridinium groups (PVA-SbQ) to prepare an amperometric biosensor for... 

M. Altstein, G. Segev, N. Aharonson, O. Ben-Aziz, A. Turniansky, and D. Avnir, Sol-gel-entrapped cholinesterases a microtiter plate method for monitoring anti-cholinesterase compounds. J. Agric. Food Chem. 46, 3318-3324 (1998). 

The Department has developed methods for monitoring the exposure of workers exposed to organophosphate and carbamate pesticides. These methods utilize the determination of plasma and red blood cell cholinesterase activities and urinary alkyl phosphates. Studies are reported vrti ich show that these methods have proven useful in evaluating the safety effectiveness of closed-transfer systems and in determining reentry times for field workers. 

For the purposes of occupational medicine, the determination of cholinesterases in the blood of workers intoxicated with OP is obhgatory. However, the normal values varied within the laboratories depending on the method of determination. Systematic monitoring of workers with nerve agents was performed at our department from 1962 to 1963 however, the results presented contain determinations during 40 years (1964-2004). Because of the large number of results, the results are limited to RBC AChE only although plasma BuChE aetivity was also determined. 

In order to adapt the system to on-line monitoring, in wastewater control, for example, the occurrence of pesticide in a flow buffer was investigated. It was found possible to differentiate between reversible and irreversible inhibition and to quantify a reversible inhibitor. Since it was possible with the calorimetric method to use the natural substrate acetylcholine to assay cholinesterase, instead of the commoly used thiocholines, this methodology might be useful in medical research as well. 

The activity of cholinesterases is usually determined by Ellman s method (19). This assay can also be carried out in microtitre plates. Thirty microliters of the ChE dilution and 30 pL of DNTB are added to 210pL of40mM Britton-Robinson-I buffer pH 8.0. The reaction is started by addition of 30 pL of the appropriate acylthiocholine substrate. A microtiter plate reader is used to monitor the development of the absorption at 595 nm. The corresponding slope is used to calculate the esteratic activity. 

Hackathom, D. R., W. J. Brinkman, T. R. Hathaway, T. D. Talbot, and L. R. Thompson. 1983. Validation of whole blood method for cholinesterase monitoring. American Industrial Hygiene Association Journal 44 547-551. 

Altstein M, Segev G, Aharonson N et al. (1998) Gel-entrapped cholinesterases a microtiter plate method for monitoring anti-choHnesterase compounds. J Agric Food Chem 46(8) 3318-3324... 

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