A new type magnetic immobilized metal affinity chromatography (IMAC) material for phosphoproteomic analyses

In this thesis a new magnetic immobilized metal (ion) affinity choromatography(IMAC) material was synthesized for use in phosphoproteomic analysis. Magnetic nanoparticles as support material, 4–hydroxy 2,6–dicarboxylate (chelidamic acid) as chelator and lanthanide(III) ions with high affinity for phosphate ions as metal ions were selected in the developed magnetic IMAC material.The cubic magnetic Fe3O4 nanoparticles were synthesized by the co–precipitation method and their size was determined as 8 ± 2 nm. To protect the magnetic nanoparticles from unwanted reactions, their surfaces are coated in 2 different ways. In the first method, core–shell nanoparticles were obtained by coating magnetic nanoparticles with silica. The size and morphology of silica–coated magnetic nanoparticles were characterized by TEM and XRD measurements, the particles were found to be 120 ± 10 nm in size, in the form of spherical particles surrounded by a cubic magnetic Fe3O4 core surrounded by an amorphous silica shell. In the second method, the surface of Fe3O4 nanoparticles was directly covered with 3–aminopropyltriethoxysilane (APTES) and particles of 9 ± 2 nm size were obtained. The surface of silane and silica coated magnetic nanoparticles was modified with chelidamic acid and it was proved by FTIR and TGA measurements that they were bound to the particle surfaces by 2.5% and 5% respectively. Two different types of magnetic IMAK material were synthesized by binding lanthanide(III) (Nd3+, Dy3+ and Er3+) ions to the surface of the obtained chelidamic acid modified magnetic nanoparticles.The performance of developed magnetic IMAC materials for phosphoproteomic analyses was determined by phosphate adsorption studies carried out at 25oC. Phosphate adsorption studies have shown that magnetic IMAC material prepared by silanizing the surface has higher phosphate adsorption capacity than silica coated magnetic IMAC materials.