Gma-the Dema Overall Column Preparation And Application

Porous monolithic columns have been prepared by the direct free radical copolymerization of glycidyl methacrylate and ethylene dimethacrylate within the confines of chromatographic column in the presence of porogens. The porosity and flow characteristics of monoliths intended for separation of proteins have been adjusted during their preparation by changing key variables such as temperature, composition of the pore-forming solvent mixture, and content of crosslinking monomer. The epoxide groups of these monoliths were modified to different extents by reaction with diethylamine to afford l-N,N-diethylamino-2-hydroxypropyl functionalities useful for ion-exchange chromatography. Following characterization of the monoliths, the columns were tested in the chromatographic separation of a homologous series of proteins. Very good separations of the proteins were achieved. The binding capacity was determined experimentally under dynamic conditions using frontal analysis using Bovine Serum Albumin (BSA). Reproducibility was checked and protein concentration, temperature and the flow rate were varied. Preliminary results confirm the flow independence of the dynamic binding capacity in the whole range of applied flow rates.In the second part of this paper, a systematical method for the analysis and purification of a small peptide (O8 peptide) , which is synthesised through chemical method, was build and optimized in terms of stationary phase, particle size, mobile phase composition and gradient program. Furthermore, this method was adjusted and scaled up to preparative scale according to the relationship between experimental condition and separation result.