Pc. Pandey et Hh. Weetall, AN EVANESCENT-WAVE SENSOR FOR THE DETECTION OF ORGANOPHOSPHORUS COMPOUNDS BASED ON THE INHIBITION OF CHOLINESTERASE, INDIAN JOURNAL OF CHEMICAL TECHNOLOGY, 2(5), 1995, pp. 261-265
A flow injection analysis (FIA) system coupled with an evanescent wave
(EW) sensor employing total internal reflection of fluorescence radia
tion (TIRP) for the detection of the organophosphorus compounds is rep
orted. The detection is based on the measurement of acetylcholinestera
se (AChE) inhibition in the presence of organophosphorus compounds. AC
hE was immobilized on controlled pore glass (CPG) particles and packed
into a teflon column which was then coupled to the evanescent wave se
nsor. A constant concentration of acetylthiocholine in Tris-HCl buffer
(10 mM, pH 7.5) was pumped through the enzyme column. The thiocholine
formed as a result of the AChE catalyzed reaction in the column was m
onitored by the injection of thylamino-3-(4'-maleimidylphenyl)-4-methy
lcoumarin (CPM) which forms a highly fluorescent compound with thiocho
line. The CPM-thiocholine complex was excited with the evanescent wave
and the resulting fluorescence was detected by a Himamastsu S-2387/66
R detector. Varying concentrations of paraoxon were injected through t
he enzyme column and the extent of enzyme inhibition was recorded by t
he injection of the optimum concentration of the CPM. The difference o
f the two responses in the presence and absence of the paraoxon determ
ines the extent of inhibition. The acetycholinesterase activity was th
en reactivated by the injection of pyridine 2-aldoxime methiodide (2-P
AM) which results in the recovery of the initial response.