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B6-OP:DEVELOPMENT OF A BIOSENSOR FOR THE DETERMINATION OF PESTICIDES,
BASED ON THE INHIBITION OF ACETYLCHOLINESTERASE
G. Csiffáry (1), P. Nagy (1), N. Adányi (2), A. Kiss (1)
1Eszterházy Károly College, Egerfood Regional Knowledge Centre, Eger, Hungary
2Central Food Research Institute, Budapest, Hungary
(
csiffary.gabor@ektf.hu)
The main objective of our research was to develop a new biosensor device for the determination of pesticide compounds based on inhibition reactions of acetylcholinesterase enzyme. We developed an amperometric system using immobilized enzyme on a natural protein membrane (thin-layer enzyme cell) [1].
We modified the FIA system: by applying the enzyme cell in stopped flow mode, we were able to study enzyme inhibition more thoroughly. We used the electrochemically active acetyltiocholin (AcTCh) as a substrate. The proper polarization potential was found at 550 mV. We used 100 mM (pH 6.5) phosphate buffer. The optimal measuring temperature was 37-38ºC, the flow rate 0.66 ml/min. The residence time for the inhibition reaction was determined at 60 sec in the stopped flow mode. The linear measuring range of AcTCh was between 1 µM and 100 mM.
After the use of the enzyme cell containing immobilized acetylcholinesterase enzyme was optimized, the inhibition of organophosphate type dichlorvos (DDVP) and carbamate type carbofuran, methomyl and pirimicarb pesticides were measured amperometrically. The linear measuring ranges of the analytes were 1-100 nM; 1-100 nM; 200-1000 nM and 100-10000 nM respectively. The detection limits were: 1; 1; 200; 100 nM for dichlorvos, carbofuran, methomyl and pirimicarb, respectively.
The thin-layer enzyme cell proved to be very stable. It can be stored at 4°C for several months without any significant decrease of the signals. Stable and proper signal response was obtained after hundreds of injections.
The system was also applied for real sample measurements. We examined ground water samples with spiking method. Promising preliminary results were acquired during these experiments.
The developed measurement system might be efficiently utilized as a screening device for multiple toxic analytes in both real food and environmental samples.
References
- N. Adányi, M. Tóth-Markus, E.E. Szabó, M. Váradi, M.P. Sammartino, M. Tomassetti, L. Analytica Chimica Acta, 2004, 501, 219–225.
