Kenneth J. Fountain

In the following application, the impact of a 125 Å pore sub-2 µm packing material on the separation and resolution of small proteins and peptides will be demonstrated. We will also show the impact of both physical and chemical properties of SEC packing materials on SEC calibration curves used for molecular weight estimation.

This application presents the impact that mobile phase composition (pH and ionic strength) can have on this separation. Additionally, we will discuss some of the primary considerations necessary for the successful transfer of a developed SE-UPLC method.

Presented in this application are the advantages that may be realized using a 125Å pore size, sub-2-µm ethylene-bridged hybrid (BEH) silica packing material and Waters UltraPerformance Liquid Chromatography (UPLC) instrumentation for this traditional analysis. Among these advantages are faster run times, higher sensitivity, and higher resolving separations of insulin and covalent insulin HMW, while at the same time greatly reducing acetonitrile containing waste-stream volumes.