Cholinium-Based Good's Buffers Ionic Liquids as Remarkable Stabilizers and Recyclable Preservation Media for Recombinant Small RNAs

RNA is a biopolymer of high relevance in the biopharmaceuticals field and in fundamental and applied research; however, preservation of RNA stability is still a remarkable challenge. Herein, we demonstrate the enhanced potential of aqueous solutions of self-buffering cholinium-based Good's buffers ionic liquids (GB-ILs), at 20 and 50% (w/w), as alternative preservation media of recombinant small RNAs. The thermal stability of RNA is highly enhanced by GB-ILs, with an increase of 14 degrees C in the biopolymer melting temperature-the highest increase observed to date with ILs. Most GB-ILs investigated improve the stability of RNA at least up to 30 days, both at 25 degrees C and at 4 degrees C, without requiring the typical samples freezing. Molecular dynamics simulations were applied to better understand the molecular-level mechanisms responsible for the observed RNA improved stability. The number of IL cations surrounding the RNA chain is similar, yet with differences found for the IL anions, which are responsible for the overall charge of the biopolymer first solvation sphere. No cytotoxicity of the studied solutions containing RNA and ILs at 20% (w/w) was observed onto two distinct human cell lines, reinforcing their potential to act as preservation media when foreseeing biopharmaceutical applications. Finally, RNA was successfully recovered from the ILs aqueous solutions, without changes in its structural integrity, and the ILs were successfully recycled and reused.