novel tyrosinase-based biosensor!

Phenolic derivatives determination in medical, food and environmental samples is very important because of the high toxicity of these compounds. Currently used phenol detection methods are neither inexpensive nor rapid screening techniques. Tyrosinase-based biosensors have been proven to be promising tools for this purpose because of advantages such as selectivity, low cost, fast and cheap screening of samples. Many materials, such as different polymers, alumina sol-gels, magnetic nanoparticles-chitosan nanocomposite, have been employed to immobilize tyrosinase in biosensors. These compounds were used in electrochemical biosensors and the enzyme was immobilized on the electrode surface. Moreover orderly protein films on a solid support can be prepared using a layer-by-layer assembly, a simple process based on the sequential adsorption of polyanions and polycations guided by electrostatic interactions. Using this technique, we obtained a mushroom tyrosinase/poly(dimethyldiallylammonium chloride) multilayer on a quartz support. The catecholase activity of the immobilized enzyme was detected using L-DOPA as substrate. Dopachrome formation was revealed at 475 nm. The kinetic parameters (Km and Vmax) of the tyrosinase were determined and compared with the behaviour of the enzyme in solution. Our results demonstrate for the first time that “layer by layer” deposition preserves the enzymatic properties of tyrosinase. For this reason we propose this layering method as a useful potential tool for the production of a tyrosinase-based biosensor.