Bridged Ethylene Polyethylene Oxide Surfaces to Improve Packing Materials for Widepore Size Exclusion Chromatography

Here, we describe the preparation of bridged ethylene polyethylene oxide (BE-PEO) surface-modified silica packing materials for size exclusion chromatography. BE-PEO surface-modified silica was hydrolyzed and subsequent ¹H nuclear magnetic resonance analysis of hydrolysis products confirmed the successful formation of BE-PEO bonded surface. Silica particles exhibiting 3 µm diameters and 1000 Å nominal pore diameters were selected as a base material for this work out of the critical need to improve analytical capabilities for the testing of cell and gene therapy drug products. Accelerated high pH aging study revealed significant enhancement in column stability. Multi-angle light scattering noise measurements showed inordinately lower baseline noise. Moreover, we evaluated the chromatographic performance of BE-PEO silica-packed columns through separations of a protein test mixture, DNA ladder, monoclonal antibody-based therapeutics, and adeno-associated viruses. BE-PEO silica columns demonstrated high resolution, high recovery separations that were confirmed to be reproducible and capable of extended column lifetimes and exhibited low ionic and hydrophobic secondary interactions. In summary, BE-PEO silica particles have yielded a new level of performance, improved base stability, and inherently lower baseline noise. These novel widepore particles will facilitate more sensitive size-based detection and characterization of large biologics in the form of advanced gene therapy products.