(650d) Competitive Protein Adsorption On Silica

Protein separations are critical in the pharmaceutical industry; unfortunately, the behavior of complex protein mixtures can be difficult to predict. To investigate this problem, the competitive adsorption of myoglobin (17.6 kDa, pI=7.2) and lysozyme (14.3 kDa, pI=11.35) on were used as test proteins for competitive adsorption on acid-washed 1000Å porous silica. Adsorption isotherms were obtained at room temperate using 0.02 M potassium phosphate buffer at pH 7; initial liquid-phase protein concentrations ranged from 0 – 570 μM for myoglobin and from 0 – 2800 μM for lysozyme.  These proteins were chosen due to their similar molecular weights and high isoelectric points. In the absence of myoglobin, lysozyme follows a Type II isotherm, while in the absence of lysozyme, myoglobin follows a Type I isotherm. In competitive adsorption experiments, lysozyme decreases myoglobin adsorption as expected. In the presence of myoglobin, lysozyme follows a Type I isotherm; at low concentrations, myoglobin decreases lysozyme adsorption. As additional myoglobin is added, lysozyme adsorption increases. The observed influence of myoglobin on lysozyme adsorption is not consistent with simple competition for surface area or functional groups. A thermodynamically-based mathematical model is used to explain the observed adsorption behavior.