(640c) Assesment of the Proteins Separation Efficiency in Polyacrylamide Hydrogels Prepared Using Templates | AIChE

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(640c) Assesment of the Proteins Separation Efficiency in Polyacrylamide Hydrogels Prepared Using Templates

Authors 

Carranza Oropeza, M. V. - Presenter, Tennessee Technological University
Sherrill, A. - Presenter,
Giudici, R. - Presenter, University of São Paulo
Arce, P. - Presenter, Tennessee Technological University
Sanders, R. - Presenter, Tennessee Tech University

Nanoporous polymer hydrogels offer a desirable combination of mechanical, optical, and transport characteristics that have placed them at the core of a variety of biomedical technologies including engineered tissue scaffolds, substrates for controlled release of pharmaceutical compounds, and sieving matrices for electrophoretic separation of DNA and proteins. All these applications are closely related to the role of the nanoporous morphology of the hydrogel matrix, especially the mean pore size and its distribution. Crosslinked polyacrylamide is a widely used gel matrix for protein and DNA electrophoresis because it offers a number of attractive properties including excellent separation resolution, optical/UV transparency, and electro-neutrality, amng others. Unfortunately, when properties of the biomolecules to be separated becomes similar to each other, challenges in the separation resolution become important. Modifications, beyond the usual crosslinking parameters are been explored in this investiagation by using templating agents, i. e. nanovoids, in order to modify the internal structure of the hydrogel so that improved separation can potentially be achieved.

In this work we study the efficiency of separation in polyacrylamide hydrogel prepared with 9% of acrylamide and 5%, 10% and 15% of template agent added. In this research,  SDS is used  in order to evaluate the efficiency of separation of specific proteins from  different sources, such as: Albumin Chicken Egg grade V (Ovalbumina, 44.3kDa, pI: 4.54), Alpha1-Acid Glycoprotein from human (40.8kDa, pI:2.7), both from Sigma Aldrich. Endostatine (20kDa, pI: ~8.9), TsnC (15kDa, pI: ~5) and YBBN (31kDa, pI: ~4.5);  these last ones from our collaborator Biotechnology Center (IPEN, Brazil). In addition to this  group,  we used besides those examples related to the standard proteins from GE Health Care ([1] Phosphorylase b, [2] Albumin, [3] Ovalbumin, [4] Carbonic anhydrase, [5] Trypsin inhibitor and [6] α-lactalbumin). The purpose in this research project is to asses the performance of the templated hydrogel in the separation of proteins with different pI (Point isoelectric), but with close molecular weight (ovalbumin and alpha1-acid), in an effort to test the role of the nanovoid in helping to improve the proteing separation efficiency. One particular aspect of interest is the  testing of the separation efficiency of the nanovoid modified hydrogel in   a mixture of proteins (ovalbumin and standard proteins). Other evaluation done in these type of nanomodified hydrogels is the comparison between the behavior of the proteins prepared in two different conditions: denatured and non denatured. The results regarding the efficiency of separation of the proteins samples tested are compared and discussed. Directions for future research are also included.

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