Toluidine Blue for the Determination of Binding of Anionic Polysaccharides to Lipid Raft Domains by Absorption Spectroscopy

Abstract

The complexes of negatively charged polysaccharides with lipid vesicles have been shown to have applications in medicine, bioremediation, water purification, and construction of nano-biosensors. This article presents research on the formation of these complexes based on the interactions between three types of liposomes, DOPC liposomes (which contain a lipid bilayer in the liquid-disordered (Ld) state), RAFT liposomes (which contain liquid-ordered (Lo) lipid raft domains surrounded by lipids in the Ld state) and SPH–CHL liposomes (which contain a lipid bilayer in the Lo state), and two selected anionic polysaccharides, polysialic acid (PSA) and polygalacturonic acid (PGA). The analysis was conducted using a toluidine blue (TB) probe and the absorption spectroscopy technique. In contrast to DOPC and SPH–CHL liposomes, binding of negatively charged PSA or PGA chains to RAFT liposomes induced a TB absorption maximum shift from 630 nm to 560 nm. The obtained results indicate that toluidine blue can be applied for monitoring the formation of these nano-complexes, and that the boundaries between Ld/Lo domains within membranes in RAFT liposomes can significantly enhance the binding affinity of negatively charged polysaccharides to the lipid bilayer surface. The observed metachromatic shift in TB absorption suggests that negatively charged PSA and PGA chains interact with the Ld/Lo boundaries within RAFT liposome membranes.

Keywords

• absorption spectroscopy
• liposomes
• polysialic acid
• polygalacturonic acid
• lipid rafts
• toluidine blue