Wide Pore Monolithic Silica of Various Functionalization: Protein A, C18, C8 and C4, in High Performance Liquid Chromatography for Large Molecule Separations

Egidijus Machtejevas, Applied Solutions SMI, Merck KGaA, Darmstadt, Germany
Tom Kupfer, Applied Solutions Smi, Merck Kgaa, Darmstadt, Germany
Benjamin Peters, Applied Solutions Smi, Merck Kgaa, Darmstadt, Germany

In the last years the pharmaceutical market has changed dramatically from small molecules to protein-based drugs. The development of biological entities is strong increasing because biotechnological processes now enable a production with reasonable costs. That implies a demand of suitable analytical methods for process monitoring and quality control of biomolecules with therapeutic purposes. Especially the HPLC is the mostly used analysis method. Most important for the HPLC analysis are the properties of the column. For bigger peptides, proteins or antibodies a new type of columns is needed which provide a good permeability, better mass transfer and a better selectivity.

In contrast to conventional packed-particle columns, wide pore (300 A) monolithic silica columns are made of a single continuous-bed rod of high purity porous silica that is then bonded with C18, C8, CN, Protein A. Monolithic columns remove backpressure as the primary consideration in method development and give back the flexibility of choices in flow rates for much higher throughput, column lengths for superior resolution, and solvent choices for optimum selectivities that smaller and smaller sized packed-particle columns have slowly taken away over the decades. Because they have no individual particles to shift or break, column performance is very consistent over much longer lifetimes, making them ideal for relatively ”dirty/matrix rich” sample analysis. Their high permeability also makes them very forgiving of shortcuts and timesaving in sample preparation as well as easier to aggressively flush out to reequilibrate.

This presentation will guide you through the world of wide pore monolithic silica materials. Benefits will be demonstrated with many application examples including pharmaceutical and bioanalysis separations (proteins, antibodies, etc.), calibration curves, recovery calculations