Design of Temperature-Responsive Surfaces and Its Application for Aqueous Chromatography

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This project aimed to develop a novel aqueous chromatography system in which the polarity of stationary phases is controlled by external temperature changes to separate and purify solutes without changing their structure and functions. This system is quite different from reversed phase chromatography in which the polarity of stationary phases is altered by the addition of organic solvents in water with various mixing ration. In the proposed system, the surface of the aminated silica stationary phase was modified with temperature-responsive poly (N-isopropylacrylamide) (PIPAAm) with a carboxyl end. The successful modification of the surfaces with polymers were confirmed by means of titration of residual surface functional groups, X-ray photoelectron spectroscopy, and atmic force microscope. These surfaces showed temperature-responsive wettability changes, being hydrophilic at lower temperature below polymer transition temperature, and become hydrophobic above the transition temperature. … More The elution profiles of five steroids were then investigated by changing temperature using high-performance liquid chromatography column packed with PIPAAm-grafted silica using aqueous mobile phase. Retention times of hydrophobic steroids increased with increasing temperature without any change in the eluent composition. Retarded elution time of hydrophobic steroids resulted from the stronger interaction of these compounds with the PIPAAm-grafted surfaces in which the surface property becomes hydrophobic with temperature increase. As a consequence, gradient elution-like effect is achieved with a single mobile phase by controlling external temperature. We further investigated the possibility of this temperature-responsive chromatography for polypeptide separation. Polypeptides with similar number of amino acid residues were successfully separated at higher temperature where hydrophobic interaction of solutes with stationary phases are primarily influenced the separation efficacy. In conclusion, elution of target substances is controlled only by a small change in column temperature without any further modification of the aqueous mobile phase in the proposed chromatography. Less