Selective Sorbents - Silica-immobilized affinity ligands and molecularly imprinted polymers

Abstract: The development of selective sorbents for the separation of various molecules, preferentially biomolecules, is described in this work. Affinity ligands were immobilized on small silica particles and used in high-performance liquid affinity chromatography (HPLAC). With this technique, it is possible to perform fast, selective and efficient bioseparations. Methods were developed for the preparation of silica-immobilized phenyl boronic acid and its use in general ligand affinity chromatography of, e.g., carbohydrates, catechols and nucleotides. Cibacron Blue is another general ligand, with selectivity for many biomolecules, such as enzymes and binding proteins. It was used for the HPLAC separation of proteins such as kinases from yeast extracts. Automatic detection was performed using an on-line enzyme assay system. Molecular imprinting was employed for the preparation of various selective sorbents. To improve the performance, a composite technology was developed in which small rigid particles of silica or TRIM (trimethylolpropane trimethacrylate) were employed for the anchoring of molecularly imprinted polymers. Proteins were for the first time used as templates in molecular imprinting. The boronate interaction with sialic acid moieties of the glycoprotein transferrin was employed for molecular imprinting in aqueous environments using polymerizable organic silanes on the surface of silica. Bovine pancreatic ribonuclease A, with exposed surface histidines was imprinted using monomers capable of forming metal chelates with the protein. Finally, the advantages of the various technologies described in this work will be discussed, as well as current trends and future prospects.