(200n) Increasing Capacity and Lifetime of Reverse Phase Resin

Development time and cost are two major impact factors for the biopharmaceutical industry in bringing a new molecule into the market. Chromatography resin is one of the more cost-intensive items used in drug substance manufacturing. One important consideration for chromatography resin in particular is its lifetime, since operating costs can be significantly reduced if the resin can maintain its resolution and capacity across multiple cycles of use. Column lifetime can vary significantly between different resins and especially between resins of different chromatographic modalities. This research focuses on reverse phase chromatography, which is a critical step in the purification of a wide variety of therapeutic proteins. A single, packed reverse phase chromatography column can be used for approximately 50 cycles without suffering from diminished performance. However, column lifetime can be adversely affected if feeds contain compounds that have the potential to foul the resin, either by precipitation or by strongly binding to the resin such that it will not elute during the method. This fouling was observed in one such program, which utilized an enzyme to improve specific productivity of an upstream reaction. This enzyme, in combination with residual sodium citrate in the feed, was found to cause column fouling of a Kromasil C8 column used for purification. The fouling resulted in dramatically reduced resolution in subsequent cycles, and therefore the effective lifetime of the column would end once this fouling occurred. The reverse phase columns could not be successfully regenerated using typically employed cleaning techniques, such as the use of sodium hydroxide. This research screens multiple potential regeneration solutions using both high throughput slurry plate screening as well as small scale column runs. A cleaning method with ethanol and acetonitrile in series was developed that successfully removed 98% fouling caused by citrate and enzyme. At this level of reduction, resolution on the column in subsequent cycles was similar to column resolution prior to fouling.