Preparation of pH-responsive stationary phase for reversed-phase liquid chromatography and hydrophilic interaction chromatography

Novel pH-responsive polymer-grafted silica was successfully synthesized through the radical “grafting from” polymerization on azo initiator-immobilized silica. The immobilization of azo initiator onto the silica surface was achieved by the reaction of surface amino groups with 4,4′-azobis(4-cyanovaleric acid chloride). The polymer-grafted silica was prepared by stirring suspension of the azo initiator-immobilized silica in anhydrous dioxane containing acrylic acid (AAc) and butyl acrylate (BA). The resulting polymer-grafted silica was demonstrated to be pH responsive to hydrophobic/hydrophilic property by reversed-phase liquid chromatography (RPLC) and hydrophilic interaction chromatography (HILIC). In RPLC mode, the retention of aromatic compounds decreased with the increase in the pH of mobile phase. However, the opposite result was obtained in HILIC mode; the retention of soybean isoflavones was stronger with the mobile phase at higher pH. Finally, the separations of sulfonamides and soybean isoflavones were carried out in RPLC mode and the separation of some nucleotides was achieved in HILIC mode.