Surfactant-Mediated Protein Chromatography

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This project is in the general area of analytical and surface chemistry and in the subfield of separation science. The objectives of this research are (a) to solubilize and separate, by hydrophobic interaction chromatography, protein mixtures without denaturation, and (b) to explore and exploit changes in chromatographic selectivity due to mild surfactants. Mobile chromatographic phases containing n-octyl glucoside, CHAPS, or Triton X-100 will be examined over a range of concentrations below and above the respective critical micelle concentrations. The dependence of protein retention on surfactant concentration and ionic strength will be determined by isocratic chromatography for an ensemble of proteins. Isotherms for the binding of surfactants to stationary phases will be determined. The effects of surfactants on chromatographic efficiency, resolving power, and linear loading capacity will also be determined. Protein recoveries (native and total) will be ascertained and indications of on-column denaturation will be tested by measurements of specific activity, circular dichroism, or intrinsic fluorescence. The correlation of solvent strength with interfacial tension measurements will be explored and convenient means will be devised for the separation of surfactant from protein following chromatography. The results of these studies will be of broad and significant potential impact in protein chemistry and biotechnology.