PHOSPHOLIPID IMMOBILIZATION ON SOLID-SURFACES

Single chain ether phospholipids (PLs) containing omega-carboxyl groups in the alkyl chain were immobilized on silica propylamine (SPA) to form IAM chromatography packing material. The PL ligands are analogs of phosphatidylglycerol (PG), phosphatidylserine (PS), phosphatidylethanolamine (PE), and phosphatidic acid (PA). Ah of these PLs contain polar functional groups in the Lipid head group that require protection prior to PL immobilization and then deprotection after immobilization. The IAM surface was prepared in four steps: (i) the omega-carboxyl group was activated with carbonyldiimidazole, (ii) the activated PL-imidazolide ligand was bonded to SPA, (iii) the surface was end capped with a long chain anhydride and then end-capped with a short chain anhydride, and (iv) protecting groups were removed to form the IAM surface. The extent of deblocking the protecting groups was typically greater than or equal to 90%. This immobilization strategy generated a phospholipid surface that was stable when solvated with all organic solvents and aqueous buffers between pH 2 and 8. Both FT-IR spectroscopy and elemental analysis indicated that the bonding densities were 64-83 mg of PL/g of SPA, which corresponds to an area per molecule of 66-104 Angstrom(2). These bonding densities for the immobilized PLs are very close to the area per molecule of mobile phospholipids comprising liposome membranes. The similar areas per molecule of immobilized PLs and mobile phospholipid in liposomes indicate that the lipid environments are similar.