(244f) Solution Effects On Protein Adsorption Mechanisms Onto SBA-15

Authors: 
Desch, R. J., University of Cincinnati
Guliants, V. V., University of Cincinnati


Because many drugs and drug targets are proteins, protein separation plays a critical role in the pharmaceutical industry. Solid silica based materials are extensively used as the adsorbent material for protein separations. However, commercial silica adsorbents have suboptimal efficiency and resolution. SBA-15, a highly ordered mesoporous silica material, has been developed with high surface area and controlled pore size and morphology. SBA-15 provides the potential of more efficient, higher resolution protein separations for the pharmaceutical industry. To test this potential, the adsorption of Burkholderia cepacia lipase, a well characterized commercially available protein, on SBA-15 was studied. The rates and mechanisms of protein adsorption onto silica are yet to be fully determined due to the complexity of the different system components. Protein adsorption onto a solid surface is determined by interactions among the proteins, the surrounding water and salt molecules, and the adsorbent. Changing the ionic strength or pH has been found to dramatically affect adsorption capacity, rate, and isotherm shape. The pH impacts electrostatic charges on the surface of both the protein and adsorbent. The ionic strength affects the amount of ions shielding each protein, thus impacting the protein solution interactions. The large effects of changing solution conditions on the thermodynamics and kinetics of protein adsorption imply alterations in the mechanism of adsorption.

To elucidate these solution effects on adsorbent capacity and isotherm shape, batch adsorption experiments were conducted at two pH values and two ionic strengths. Confocal laser scanning microscopy (CLSM) was used to visualize intraparticle concentration profiles of fluorescently tagged proteins (lipase, 33 kDa, and bovine serum albumin, 69 kDa) in the pores of a mesoporous silica adsorbent. Images were captured of individual adsorbent particles at staggered time points and statistically averaged to develop the protein concentration profiles. Quantitative thermodynamic data were obtained by measuring the heat of adsorption of the protein onto a packed bed using flow microcalorimetry. Correlation between calorimetric events and diffusion patterns in single particles allowed comprehensive determination of the mechanisms of diffusion. These data was then applied to the behavior of these proteins in high performance liquid chromatography (HPLC), providing a laboratory-scale model for industrial preparative chromatographic separations.