Chromatographic Characterization of Phosphonate Analog EDTA-Modified Zirconia Support for Biochromatographic Applications

Abstract

Zirconium dioxide (zirconia) has a great affinity for inorganic and organic phosphate. Previous work from this laboratory demonstrated the utility of phosphate-modified microparticulate zirconia as a support for protein separations. We have extended this investigation to include the study of ethylenediamine-N,N'-tetramethylphosphonic acid (EDTPA), a phosphonate analog of EDTA, as a surface modifier for zirconia. Our work explores the use of EDTPA-modified zirconia (PEZ) for its potential use as a high-performance inorganic cation-exchange support for the separation of proteins. The phosphate groups in EDTPA very effectively block the sites responsible for strong interactions of hard Lewis bases with zirconia's surface. Modification of zirconia with EDTPA provides a "biocompatible" stationary phase, resulting in high mass recoveries of proteins. We compare PEZ with inorganic phosphate-modified zirconia to show increased efficiency, as well as unique selectivities for chromatography of proteins on the chelator-modified surface. Finally, the selectivity, efficiency, and separation mechanism are reported. The studies show that PEZ is a useful high-performance ion-exchange support for the separation of cationic proteins and for modulating the sites responsible for the high affinity of zirconia toward certain classes of anions.