390775 Efficiency of Separation of Macromolecules Using Electrochromatography Technique

Tuesday, November 18, 2014
Galleria Exhibit Hall (Hilton Atlanta)
Maria Veronica Carranza Oropeza1, Reinaldo Giudici2, J. Robby Sanders1 and Pedro E. Arce1, (1)Chemical Engineering, Tennessee Technological University, Cookeville, TN, (2)Chemical Engineering, Polytechnic School, University of São Paulo,, São Paulo, Brazil

Recent years have seen an increasing interest in use the electrochromatography technique for separation of macromolecules. Electrochromatography is a chemical separation technique in analytical chemistry, biochemistry and molecular biology used to resolve and separate mostly large biomolecules such as proteins. This technique is a combination of size exclusion chromatography (gel filtration chromatography) and gel electrophoresis. These separation mechanisms operate essentially in superposition along the length of a gel filtration column to which an axial electric field gradient has been added. The molecules are separated by size due to the gel filtration mechanism and by electrophorectic mobility due to the gel electrophoresis mechanism. Additionally there are secondary chromatographic solute retention mechanisms.

In this work we have developed a modified electrochromatography system in order to able to provide useful estimates of protein retention time and velocity in a column packed with Sephadex gel and subjected to an electric field. Uniformly sized polymer particles and with different shapes were utilized as the shape template to compare their suitability as a uniformly sized packing material for small-scale high-performance liquid chromatography (HPLC) or capillary electrochromatography (CEC).

A correlation of electrophoretic mobility of peptide and proteins with respect to their charge, molecular mass, and asymmetry enables the selection of solute target molecules for electrochromatographic separations. The results regarding the efficiency of separation of protein mixtures, and the characterization of the different columns packed are compared and discussed in order to understand the influence of the particles (used as template) in the separation process. Directions for future research are also included.

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