High-performance adsorption chromatography of proteins on deformed non-porous agarose beads coated with insoluble metal compounds : II. Coating: aluminium and zirconium (hydr)oxide with stoichiometrically bound phosphate

Abstract Two novel supports for adsorption chromatography of biomolecules, based on deformed non-porous cross-linked agarose beads coated with aluminium or zirconium (hydr)oxide, were prepared. Some fundamental chromatographic properties of these bed materials are discussed, including the adsorption mechanism and high-resolution separations of mixtures of proteins, nucleotides and coenzymes. Both aluminium and zirconium (hydr)oxide agarose irreversibly and stoichiometrically bind phosphate ions, thus some of the chromatographic properties resemble those of calcium phosphate and hydroxyapatite. Fractionation of a β-glucosidase preparation into two components possessing enzymatic activity was achieved on the aluminium (hydr)oxide phosphate agarose, whereas chromatography on zirconium (hydr)oxide phosphate agarose removed several inactive contaminants but gave only one peak associated with enzyme activity. This finding indicates that the adsorption mechanisms of the two adsorbents are not identical, although other experiments show that there are several analogies. For instance, model proteins chromatographed on these two supports were eluted in the same order, although the relative retention times differed, i.e. , the chromatograms did not have the same appearance. Compressed beds of cross-linked non-porous agarose beads, including those coated with aluminium and zirconium (hydr)oxide, have the unique feature that the resolution is roughly independent of flow-rate and that there is no need for small beads of narrow size distribution.