Zhong Zhong1, Sean Pieper2, Manasi Katragadda2, Thomas K. Wood3, Kevin L. Lear2, David S. Dandy1, and Kenneth F. Reardon1. (1) Department of Chemical and Biological Engineering, Colorado State University, 1370 Campus Delivery, Fort Collins, CO 80523, (2) Department of Electrical and Computer Engineering, Colorado State University, 1370 Campus Delivery, Fort Collins, CO 80523, (3) Artie McFerrin Department of Chemical Engineering, Texas A&M University, 200 Jack E. Brown Building, MS 3122, College Station, TX 77843-3122
Groundwater monitoring plays an important role in safeguarding our water supply, especially drinking water resources. Toluene and trichloroethylene (TCE) are two common groundwater contaminants, as well as critical contaminants in the National Primary Drinking Water Regulations. Traditional monitoring for groundwater contaminants usually involves field sampling, transport, and laboratory measurements (usually by GC with an FID or MS detector, or by HPLC). These measurements are expensive and slow and require removal of the sample from the site, which adds to the cost and alters the analyte concentration through volatile losses and nonlinear volume averaging during the sampling process. Sensors, and especially biosensors, could avoid these disadvantages by providing real-time in situ measurement with highly specific and sensitive detection. A biosensor usually consists of a biological component that detects the analyte through an enzymatic reaction or biological binding, a transducer that translates the biochemical signal into an optical or electrical signal, and a detector that recognizes the signal and allows it to be correlated to the analyte concentration.
In this project, oxygenase-based fiber-optic biosensors are developed to measure toluene and TCE respectively. These biosensors are made by using oxygenase-expressing bacteria as the biocomponent and oxygen optodes as the transducer. The optical signal change (OSC) has been found to correlate to the analyte concentration. Employing different oxygenases as the biocomponent yielded biosensors with different responses to toluene and TCE. For example, biosensors constructed with ttoluene ortho-monooxygenase (TOM) exhibit an OSC/ΔCtoluene = 148 counts•L/mg with limits of detection of 300 μg/L, while biosensors made with TOM-Green (a selected mutation of TOM derived from directed evolution) were found to have an OSC/ΔCTCE = 694 counts•L/μg with limits of detection of 0.2 μg/L. The calibrations of these biosensors were verified by GC/MS. The average response time for these biosensor is around 1 hour. The biosensors could retain more than 75% of their original activity for at least a week and could be regenerated by exposure to formate or glucose.