• Biosensors
• Precision Enology
• Chemical functionalizations
• Polyphenols

Polyphenols are secondary metabolites synthetized during plants growth in response to stress. The recurrent chemical structure of polyphenol macromolecules contains aromatic rings with one or more hydroxyl substituents.
This broad class of chemical compounds exhibits several interesting biological properties, such as anti-inflammatory and anti-bacterial actions, as well as multiple positive effects on human health. Furthermore, polyphenols, and their sub-family of tannins, show a crucial role in the oenological field, determining the organoleptic characteristics of wines.
The analysis of polyphenols content in grapes, musts and wine id of great importance in the entire winemaking process, being crucial in identifying the moment of grape harvest and monitoring the fermentation process.
Thus, the analysis of polyphenols content in different phases of the process is crucial from an industrial point of view.
To date, UV-Vis or Near Infrared analysis are helpful instrumental methods used for the quantification of polyphenols. Such detections are costly solutions for a constant monitoring and requires a specialized laboratory.
The construction of cheaper and portable sensors would be desirable to reduce costs and speed up measurements.
In this work, I have collaborated with the NeuroSens group at the Istituto Nanoscienze of the National Research Council, located at the Laboratorio NEST, in the project SAWINE – Laboratorio on-chip basato su onde acustiche di superficie per enologia di precisione, funded by Fondazione Cassa di Risparmio di Lucca. The aim of the research project is the development of an effective and cheap strategy for the in-field analysis of polyphenols in grapes, musts and wines. To do this, we have developed a Lab-on-Chip (LoC) technology combining surface chemical modifications and acoustic waves devices, with the aim to develop a novel Surface Acoustic Waves (SAW)-based portable sensor for oenological applications.
My contribute to this project was to perform the preliminary analysis of chemical functionalizations with a Quartz Crystal Microbalance with Dissipation monitoring (QCM-D), testing the efficacy of functionalization.
Sensing of QCM-D relies on the detection of molecular adhesion on a Au-quartz sensor, and allows the study of biomolecular binding events at the solid-liquid interface.
Two different types of functionalization of QCM crystals were evaluated:
• - Functionalization with proteins.
• - Functionalization with peptides.
Explored proteins were Bovine Serum Albumin (BSA) and Gelatine type A (Gel-A); we fine-tuned a strategy to covalently bind the proteins to the surface of the QCM crystals. These proteins contain different sites that can interact with polyphenols, and, among such sites, proline residuals exhibit a crucial role.
In the functionalization with peptides, we studied the behaviour of two customized peptides derived from Istatine-5 (Ist-5) and Murine salivary Protein-5 (MP-5). Such peptides were selected on the basis of their chemical properties: Ist-5 aminoacidic sequence allows interactions with aromatic residuals of condensed tannins, while MP-5 has great affinity for hydrolysable tannins.
We exploited the cited chemical functionalities starting from the analysis of aqueous pure gallic tannin solutions and then we proposed the same strategies in the analysis of whole wines with known polyphenols concentrations.
Results indicate that the chemical procedure developed for the functionalization of the Au-coated sensors has a notable yield and good repeatability. The immobilized amounts of proteins over the surface ranges from 56,58 to 7061,01 ng/cm2, while for peptides ranges from 17.24 to 131.94 ng/cm2. We were able to analyze both aqueous solutions and complex wines with the same analytical setup. We were able to analyze solutions with increasing concentrations, from 0,02 g/L to 3.73 g/L of polyphenols, with limited saturation effects of the sensor functionalization. Moreover, we have demonstrated that polyphenols can be detected with the proposed technology, and tannin can be discriminated from polyphenols. The amounts of immobilized molecules over the sensor surface are found to be suitable for a sensitive detection, resulting from 0.05 to 0.847 ng of polyphenols over ng of immobilized molecules. Our results are interesting, notably because this work represents the first attempt to combine surface functionalization and acoustic sensors with the analysis of polyphenols from liquid solutions.
Results of the present work are currently exploited at the Laboratorio NEST for the development and the refining of an in-house high-frequency LoC SAW-based device, in fulfilment of the SAWINE project’s aim.