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A Mid-Infrared Flow-Through Sensor for Label-Free Monitoring of Enzyme Inhibition

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Abstract:

Label-free monitoring of acetylcholinesterase (AChE) activity was achieved with a mid-infrared flow-through sensor. The flow-through sensor comprised agarose beads, carrying covalently immobilized AChE, which were placed in a temperature-controlled (37 °C) CaF2 flow cell with an optical path of 60 μm. The sensor was incorporated into a computer-controlled sequential injection (SI) system for automated liquid handling. Different mixtures of enzyme substrate acetylcholine (ACh) and inhibitor (tacrine) were prepared and fed into the flow-through sensor. The flow was stopped as soon as the prepared mixtures reached the sensor. Enzymatic hydrolysis of ACh by AChE was directly monitored as it took place in the flow-through sensor. The inhibition effect of tacrine was calculated from the reaction-induced spectral changes, revealing an important decrease in the activity of AChE, approaching zero when the inhibitor concentration is high enough. The developed mid-infrared flow-through sensor is flexible and can be used to study the inhibitor activity of different target molecules as well as different enzymes.

Keywords: ACETYLCHOLINESTERASE; ENZYME; FLOW-THROUGH SENSOR; FOURIER TRANSFORM INFRARED SPECTROSCOPY; FT-IR SPECTROSCOPY; INHIBITION KINETICS; SEQUENTIAL INJECTION ANALYSIS; SIA

Document Type: Research Article

DOI: http://dx.doi.org/10.1366/000370208786822214

Affiliations: Institute of Analytical Chemistry, Vienna University of Technology, Getreidemarkt 9/164 AC, A-1060 Vienna, Austria

Publication date: December 1, 2008

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