Sugar-based Cryoprotectants Stabilize Liposomal Vesicles by Exhibiting a Cholesterol-like Effect

Abstract

Liposomal vesicles tend to fuse and aggregate during lyophilization. To avoid these events, cryoprotectants are added to the dispersion before lyophilization. Herein, we have compared the effect of three commonly used cryoprotectants (mannitol, MTL; trehalose, THL; and β-cyclodextrin, β-CD) upon structural characteristics of liposomes. The formulation was prepared using ethanol injection method, and cryoprotectants were tested at three dose levels (2, 6, and 10 mM). We have elucidated their effect on soy lecithin (SL) liposomes formulated with and without cholesterol (CHL). Characterizations were performed using scattering, thermal, and spectroscopic techniques. CHL molecules interacted hydrophobically with the SL bilayer. In spite of triggering a noticeable increase in the hydrodynamic diameter (about 30 nm), CHL promoted the stabilization of vesicles. Hydrogen bonding interactions were verified by the shift in −OH stretching over 3300−3500 cm −1 . This manifested in an increased phase transition temperature (T m ) of SL liposomes. T m increased further upon incorporation of cryoprotectants, particularly with β-CD. Enthalpic changes were indicative of an affinity interaction between phospholipids and cryoprotectants, regardless of the presence of CHL. β-CD showed concentration-dependent changes in the energetics of this interaction. The affinity of cryoprotectant-liposome interaction has been ranked as β-CD ≫ THL > MNT.

Alkanols Regulate the Fluidity of Phospholipid Bilayer in Accordance to Their Concentration and Polarity

“…Liposomal vesicles were prepared by the ethanol injection method . A required amount of phospholipid was dissolved in ethanol (2 mL) and it was injected dropwise (1 mL/min) into alcoholic solutions (10 and 20 mM) under mild magnetic stirring.…”

“…Such structures, vesicles in particular, are encapsulated with hydrophilic and hydrophobic payloads. In addition, phospholipid vesicles have gained wide popularity in fundamental research due to their resemblance to cell membranes. − Membrane properties can be tuned by combining phospholipids with small molecular organic substances including alcohols. , For instance, short-chain alcohols ( n -alkanols) are incorporated in liposomal vesicles as permeation enhancers. , With their polar hydroxyl group and nonpolar hydrocarbon chain, alcohols exhibit excellent affinity for the bilayers. It has been shown that alkanols can modulate the bilayer properties, and affect their overall organization in fluid as well as gel phase. , The extent of alcohol-induced disorder in interacyl chain spacing of vesicles differs in between fluid and gel phases .…”