Naslov
55. Limitation of the Ellman Method: Cholinesterase activity measurement in the presence of oximes
(Limitation of the Ellman Method: Cholinesterase activity measurement in the presence of oximes)

Autori
Šinko, Goran ; Čalić, Maja ; Bosak, Anita ; Kovarik, Zrinka

Vrsta, podvrsta i kategorija rada
Radovi u zbornicima skupova, cjeloviti rad (in extenso), znanstveni

Izvornik
Proceedings of the Seventh International Chemical and Biological Medical Treatment Symposium / Price, Richard ; Brodbeck, Urs ; Price, Barbara ; Lehnherr, Irma - Kaneohe (HI) : Applied Science and Analysis, 2008, 1-6

Skup
The Seventh International Chemical and Biological Medical Treatment Symposium

Mjesto i datum
Spiez, Švicarska, 13.04.2008. - 18.04.2008

Vrsta sudjelovanja
Poster

Vrsta recenzije
Međunarodna recenzija

Ključne riječi
Ellman method; oxime; oximolysis; acetylthiocholine; cholinesterase

Sažetak
The Ellman method for assaying thiols is widely used for cholinesterase activity measurement. The enzyme activity is measured indirectly through increase in the absorption of 5-thio-2-nitrobenzoic acid (TNB) ion, a product of the reaction between thiol reagent DTNB (5, 5’ -dithiobis(2-nitrobenzoic acid)) and thiocholine. Thiocholine is the product of acetylthiocholine (ATCh) hydrolyses by cholinesterase. Oximes, reactivators of inhibited cholinesterase by organophosphorus compounds, are nucleophiles that also react with ATCh, producing thiocholine and indirectly TNB ion. The aim of this study was to characterize ATCh oximolysis. Therefore we measured reaction between oximes (K027 or HI-6) and ATCh with DTNB, using the Ellman method at different pH, and taking into account thiocholine concentration. To confirm oximate ion involvement in the nucleophilic attack we also determined the reaction rate between the oximes and ATCh, without DTNB, at different pH by measuring the decrease in oximate ion absorption over time. Oximate ion of K027 reacted 14 times faster with ATCh (306 M-1 min-1) than oximate ion of HI-6 (22 M-1 min-1). However the rate constants obtained with the Ellman method were 84 M-1 min-1 for K027 and 22 M-1 min-1 for HI-6. The rate obtained with K027 is actually rate of the Ellman reaction itself. This suggests that the Ellman’ s reaction can not be used uncritically to evaluate oxime reaction with choline esters, in particular when oximolysis is faster than the Ellman reaction itself at a given pH.

Izvorni jezik
Engleski

Znanstvena područja
Kemija

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