Preparation, characterization and application of natural deep eutectic solvents in the extraction of phenolics from pomegranate (Punica granatum L.) peel

Natural deep eutectic solvents (NADES) have been used as alternative means for the extraction of secondary plant metabolites due to advantages such as non-toxicity, ease of preparation and extraction of compounds with a wide range of polarity. In this study, it was sought to prepare and characterize NADES based on choline chloride (CC), a quaternary ammonium salt, with different hydrogen bond donor (HBD) compounds. The aim was to use NADES to extract phenolics from pomegranate (Punica granatum L.) peel, an agro-industrial residue with a high antioxidant potential, which can be reused in value-added applications (coatings, formulations, cosmetics). NADES were formulated at 60°C by mixing of CC with 20% (w/w) of water and different HBD (1:1 ratio): glucose (CC-GLUC), sucrose (CC-SUC), glycerol (CC-GLYC), lactic acid (CC-LAC) and citric acid (CC-CIT). NADES were characterized in terms of their pH and density, and in terms of their viscosity as a function of temperature, using an AR-1000 N rheometer. The extraction of phenolics from pomegranate peel took place in a 1:30 ratio (peel: NADES) at 45 °C, in an ultrasonic bath for 1 h. An ethanolic extract was prepared as a reference, following the same procedure. The extracts were centrifuged, filtered and stored under refrigeration. Their total phenolic content (TPC) was determined by the Folin-Ciocalteu method, using gallic acid as standard. The extracts were also characterized in terms of density and pH. Regarding the characterization of NADES, it was observed that the solvents containing organic acids (CC-LAC and CC-CIT) presented themselves as the most acidic, with pH's close to or less than 1.0. The solvents with higher density were also the most viscous, CC-GLUC>CC-SUC>CC-CIT>CC-LAC~CC-GLYC>ethanol. The viscosity of all NADES decreased according to the increase in temperature, a good result when it is aimed their application as extracting solvents, since the high viscosity is one of the limiting factors of their use. Extracts from CC-LAC and CC-CIT were the most acidic (pH around 2.3, while all the others had pH values between 4-5), and were also the extracts with the highest TPC: 4.14 and 3.84 mg gallic acid equivalent (GAE)/mL of extract, respectively. This is probably related to their greater acidity, since phenolic compounds are more easily extracted when the pH of the medium is lower than their pKa. These two NADES were the only capable of extracting more phenolics than ethanol (TPC of 3.55 mg GAE/mL of ethanolic extract). Extracts from CC-GLUC and CC-SUC presented the lowest TPC (2.51 and 0.81 mg GAE mL-1, respectively), which may be related to the high viscosity of these solvents. Therefore, this study was able to successfully develop natural deep eutectic solvents as a novel technology for phenolics extraction; it was observed a strong influence of pH and solvent viscosity on the extraction efficiency, and the NADES prepared with organic acids overcame ethanol in the amount of phenolics extracted. Finally, from the extracts obtained with NADES, it is expected to facilitate the reuse of an agro-industrial residue with high antioxidant capacity, the pomegranate peel.