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A Fiber-Optic Biosensor Based on Monooxygenases and Sol–Gel Entrapped Fluoresceinamine for Trichloroethene and Tetrachloroethene

D. Ryoo, Y. Kim, K. Jeun, T. Wood and Y. Choi
Jeonju Univ., KR

TCE, PCE, optical fiber, biosensor

Tetrachloroethene (a.k.a. perchloroethene, PCE) and trichloroethene (TCE) are the most frequently detected groundwater contaminants which are suspected to cause cancer in humans. Monitoring of these carcinogens suffers from high costs and inaccuracy caused by cumbersome sampling procedure altering the analyte concentration through volume averaging. A couple of whole cell biosensors were tested for TCE, but no sensor has been developed for PCE. A biosensor system based on a fiber optic enzyme sensor was developed to detect TCE and PCE. Directed evolution called shuffling was performed for toluene-o-monooxygenase (TOM) and toluene/xylene-o-monooxygenase (ToMO) to obtain differential and improved reaction rates with respect to each chlorinated ethenes. Peroxide shunt was applied to remove the requirement for NADH, thus eliminating the need for complex NADH regeneration schemes for live cells on the biosensor tip and long-term application of enzyme biosensor would be possible. A three-layer sandwich structured sensing tip was constructed. The monooxygenases were immobilized on the outer layer, consisting of hydrophilic modified polyvinylidenefluoride membrane. The membrane was in contact with an intermediate sol–gel that incorporated fluoresceinamine (FLA), layered on an inner glass disk. The sensor operated in a static mode at room temperature and the intensity variation caused by hydrogen ion served as an analytical signal. Calibration curves were obtained for TCE and PCE, with concentration ranges 0.2–100 mg l?. The detection limits were 2 g l?1 for TCE and PCE. The method reproducibility was tested. The method was successfully applied to the detection and determination of these chlorinated ethenes in water samples, without sample preparation steps.

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