Unravelling the influence of choline chloride-based deep eutectic solvents on lysozyme: a comparative study of fructose and formic acid donors

Advances in medicine and pharmaceutics have led to an increasing need for efficient structure stabilization methods for therapeutic proteins. Lysozyme is an antimicrobial enzyme present in mucosal secretions and animal and plant tissues and merited by a wide range of applications. However, its inherent vulnerability to high temperatures and external stress limits these applications. To overcome these drawbacks, biocompatible solvents (deep eutectic solvents (DESs)) are used to stabilize the structure and activity of lysozyme. Consequently, a comprehensive assessment was undertaken to evaluate the effects of two DES solutions-choline chloride-fructose (ChCl/F) and choline chloride-formic acid (ChCl/FA)-on the conformation, thermal stability and enzymatic activity of lysozyme. This was done using UV-visible spectroscopy, Fourier transform infrared (FTIR) spectroscopy, steady-state fluorescence, dynamic light scattering (DLS), circular dichroism (CD) measurements, transmission electron microscopy (TEM) and activity assays, as a function of DES concentration. ChCl/FA is shown to preserve the structural and thermal stability, and enhance the enzymatic activity of lysozyme, while ChCl/F was shown to destabilize lysozyme. Moreover, high concentrations of both DES solutions weaken the activity of lysozyme. Overall, DESs can be described as potential biocompatible, sustainable media for preservating the stability and activity of lysozyme.