muramidase and 1-2-distearoyllecithin

Distearoylphosphatidylcholine (DSPC)/cholesterol/distearoylphosphatidylethanolamine (DSPE)-polyethylene glycol 5000 [PEG(5000)] lipid disks, mimicking biological membranes, were used as pseudostationary phase in partial filling electrokinetic capillary chromatography (EKC) to study interactions between pharmaceuticals and lipid disks. Capillaries were coated either noncovalently with a poly(1-vinylpyrrolidone)-based copolymer or covalently with polyacrylamide to mask the negative charges of the fused-silica capillary wall and to minimize interactions between positively charged pharmaceuticals and capillary wall. Although the noncovalent copolymer coating method was faster, better stability of the covalent polyacrylamide coating at physiological pH 7.4 made it more reliable in partial filling EKC studies. Migration times of pharmaceuticals were proportional to the amount of lipids in the pseudostationary phase, and partition coefficients were successfully determined. Because the capillary coatings almost totally suppressed the electroosmotic flow, it was not practical to use the EKC-based method for partition studies involving large molecules with low mobilities. Hence, the applicability of the biomembrane mimicking lipid disks for interactions studies with large molecules was verified by the quartz crystal microbalance technique. Biotinylated lipid disks were then immobilized on streptavidin-coated sensor chip surface, and interactions with a high-molecular-mass molecule, lysozyme, were studied. Cryo-transmission electron microscopy and asymmetrical flow field-flow fractionation were used to clarify the sizes of lipid disks used.

Topics: Animals; Bacterial Proteins; Chickens; Cholesterol; Chromatography, Micellar Electrokinetic Capillary; Lipids; Membranes, Artificial; Muramidase; Phosphatidylcholines; Phosphatidylethanolamines; Polyethylene Glycols; Protein Interaction Mapping; Quartz Crystal Microbalance Techniques; Streptavidin; Streptomyces