Electrostatic control of protein–vesicle interactions in catanionic long-chain ionic liquid systems: insights from human serum albumin

Soft nanosystems (micelles, vesicles, reverse micelles, <i>etc.</i>) are highly significant in the biomedical field as a delivery system of biomolecules and drugs because of their easily modulable shape. In recent years, surface-active ionic liquid (SAIL)-based vesicles have gained popularity, with applications in gene therapy, micellar catalysis, and protein and drug delivery, among others. This article presents a comparative study on the interaction of cationic- and anionic-rich vesicles prepared using a SAIL (1-hexadecyl-3-methylimidazolium chloride, C₁₆MimCl) and aerosol OT (AOT) with a model protein (HSA) in aqueous buffer medium at pH 7.4. The interaction study used various physicochemical, spectroscopic, thermodynamic, and morphological measurements. The steady-state fluorescence spectra of the Trp214 residue of HSA were found to change slightly in cationic-rich vesicles, but nearly disappeared in anionic-rich vesicles. This suggests that the tertiary structure of HSA was preserved in cationic-rich vesicles, but broken down in anionic-rich vesicles. DLS experiments show that the volume of cationic-rich vesicles rapidly rises (197 ± 10 nm to 1091 ± 55 nm upon the addition of ∼4 µM HSA) without disrupting their vesicular structure, but anionic-rich vesicles split into smaller vesicles upon protein incorporation. The volume expansion of cationic-rich vesicles during protein adsorption is readily visible in TEM micrographs of the protein-vesicle assembly. Zeta potential measurements indicate that electrostatic attraction plays a major role in protein adsorption on cationic-rich vesicle surfaces, whereas electrostatic repulsion on anionic-rich vesicles leads to the disintegration of the vesicle structure during protein adsorption. According to the CD analysis, a little structural alteration (∼2%) of HSA occurs in anionic-rich vesicles, while no secondary structural change occurs in cationic-rich vesicles. Overall, by integrating multiple techniques, these results provide mechanistic insights into protein-vesicle interaction, revealing that, in cationic-rich vesicles, protein loading preserves both the vesicle and protein structure, making it suitable for use as an injectable drug delivery system (DDS); this is not the case with anionic-rich vesicles. This understanding of electrostatic effects offers a framework for the rational design of stable SAIL-based nanocarriers, highlighting their potential in protein stabilization and drug delivery.