Phenolic compounds include a large variety of analytes relevant in health care and pollution monitoring. Tyrosinase is an enzyme which has broad substrate specificity toward a broad range of phenols. It converts electrochemically inactive monophenols into active quinones, and most tyrosinase biosensors are based on the electrochemical reduction of quinone produced by the enzymatic reaction. Immobilization of the enzyme while retaining its specific biological function, is a key step in the construction of enzyme-based biosensors because it ensures an intimate contact between the enzyme and the underlying transducer, can improve enzyme stability, and largely determines the ultimate sensor performance.

Here, we present our recent data for measurement of tyrosinase kinetics using electrochemical methods and compare the results with those obtained from UV-Vis spectrophotometry.  Further, we show the procedure for immobilization of tyrosinase in a thin layer cell. The immobilization method is based on introducing sulfhydryls group into the enzyme through a bifunctional linker which then can bond to the gold electrode surface via thiol bonds. The electrode is placed in a thin layer cell of volume 3×10^-10 m³. Different parameters including immobilization time, incubation time, and phenol concentration are studied. Our study shows that the enzyme remains active after immobilization and that activity can be measured even with  small amounts of immobilized enzyme.

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