Aqueous chromatography utilizing hydrophobicity-modified anionic temperature-responsive hydrogel for stationary phases.

A new pH-/temperature-responsive poly(N-isopropylacrylamide-co-acrylic acid-co-N-tert-butylacrylamide) (poly(IPAAm-co-AAc-co-tBAAm)) hydrogel grafted on silica beads was evaluated as column matrix for a cation-exchange thermoresponsive chromatography. The stationary phase showed simultaneous changes in temperature-responsive surface charge density and hydrophobicity by incorporation of anionic AAc and hydrophobic tBAAm into IPAAm sequences. Thermoresponsive polymer property alterations were confirmed by temperature-responsive phase transition and shift in apparent pKa values. Catecholamine derivatives were retained on poly(IPAAm-co-AAc-co-tBAAm)-modified column at pH 7.0. Analyte retention was primarily due to the electrostatic interaction. It was noted that the temperature-induced phase transition of poly(IPAAm-co-AAc-co-tBAAm) hydrogel layer on the stationary phases was evidenced by the apparent inflection point in van't Hoff plots around 36 degrees C. This suggests that solute interactions should be changed below and above the stationary phase transition temperature, reducing electrostatic interaction above the transition temperature.