Novel chromatographic separation materials for proteomics and pharmaceutics

In the field of proteome research new techniques for high throughput protein identification are of great interest. Currently, enzymatic cleavage followed by mass spectrometry (MS)-based identification of proteins is state-of-the-art. Enzymatic cleavage of proteins is usually performed by the serine protease trypsin either in-gel or in-solution. In order to prevent autodigestion of trypsin leading to tryptic fragments, which might complicate the interpretation of mass spectra most protocols employ a low concentration of protease in combination with long incubation times. Immobilization of the digestion enzyme is a useful tool to overcome these limitations. By attaching the enzyme via a covalent bond to a stationary phase autodigestion is eliminated. When monolithic materials are used as stationary phase, digestion of the protein is accelerated because of the excellent mass transfer within monolithic materials resulting in digestion times in the range of minutes. Furthermore, the preparation of a monolith in a fused silica capillary (FSC) enables the use of the monolithic enzyme reactor in HPLC experiments with off-line or on-line coupling to various MS devices.