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Main purpose of the work is assembling, testing and optimizing new disposable amperometric biosensors to analyze substances in different application fields as agribusiness, clinical chemistry and environment protection. Many kinds of modified electrodes have been prepared and tested to build portable devices to analyze quickly many analytes, in a simple and cost-effective manner. Bare electrodes of the screen-printed type, with silver as reference, have been used for modification. The glassy carbon electrodes with multi-walled carbon nanotubes or graphene or gold nanoparticles depositions were modified with generation IV ionic liquids. Choline as cation and amino acids, such as glycine, serine, phenylalanine and histidine, as anions have been employed for these ionic liquids. The presence of nanostructured materials on the electrode brings an increased contact surface between analytes and receptor and, consequently, an amplification of the amperometric signal and a better sensibility. Moreover the use of new ionic liquids of generation IV, biologically friendly and water soluble, improves the electronic transfer, facilitating and strengthening the redox reaction nearby the electrode. By immobilizing the proper enzymes onto the modified electrode surface, different compounds of analytical interest can be determined by means of sensitive, properly designed amperometric biosensors. Analytes such as antioxidant components in extra-virgin olive oils, alcohols in beverages and glucose in food matrices have been tested, using a suitable enzyme: microbial lipase, alcohol dehydrogenase and glucose oxidase, respectively.