Natural deep eutectic systems as cryoprotective agents for cryopreservation process of cells: Characterization of thermophysical and cytotoxicity properties.

Natural deep eutectics systems (NADES) are composed mostly of natural primary metabolites such as sugars, sugar alcohols, organic acids, amino acids, and amines. Additionally, water can also be incorporated in their structure. These components have been identified in animals living in extreme temperature amplitudes environments.
These eutectic solvents are the result of the creation of intermolecular hydrogen bonds between two and/or three compounds, in correct molar ratio. More recently, studies have demonstrated that stability and other properties, such as viscosity, conductivity and toxicity can be improved with the introduction of water as a tertiary component. This water is strongly retained in the NADES structure and cannot be evaporated.
In this work, we report new NADES for possible application in cryopreservation, composed by sugars (e.g., trehalose, sucrose, and fructose), alcohols (e.g., glycerol), sugar alcohols (e.g., mannitol), amino acids (e.g., proline and histidine) and water. We report also experimental data for these NADES, namely physical properties and cytotoxicity. We studied the rheological properties such as viscosity in a temperature range from -15 C to 40 C. Different water contents (10% to 80%) were tested and we observed that the viscosity is greatly decreased by the presence of water. The evaluation of the thermal behaviour of the systems, between -90 C and 40 C, showed that NADES has a strong effect on the crystallization of water. At high NADES contents no crystallization event were observed. The cytotoxicity of the NADES studied, presented different cellular responses and
increasing water concentration increases cell viability, which may be related to the decrease of the viscosity of the solutions.
These results demonstrate that tuning the physical properties of NADES will lead to different biological responses. In this sense, it is essential to understand how the physical and biological properties are related and to perform the characterization of the systems.