A novel chromatographic system utilizing temperature-responsive polymer carrying amino acid derivative as the HPLC stationary phase is proposed. In this study, we prepared the functional copolymers composed of N-isopropylacrylamide (NIPAAm) and N-acryloyl amino acid derivative (proline, phenylalanine, and tryptophan), which have temperature-responsiveness and molecular recognition. These copolymers underwent a reversible phase transition from a hydrophilic to a hydrophobic micro-structure in aqueous solution in vicinity of their lower critical solution temperature (LCST).  The temperature-responsive chromatographic system is based on a novel concept that the property of stationary phase surface is altered by external temperature.

In this chromatographic system, we could control the retention times of target analytes by changing the column temperature, such as PTH-amino acids, pharmaceutical agents and bioactive peptides.  On proline-derivative based polymer modified stationary phase, the retention of PTH-aromatic amino acids were dramatically increased derived from the CH- interaction.  In addition, phenylalanine-derivative based polymer was shown the expected - interaction to the PTH-tryptophan.  In the research, we developed novel separation system available the dual characteristic of temperature-responsibility and molecular recognition.

Additionally, temperature-responsive chromatography could achieve the separation of proteins under mild elution conditions, such as a physiological condition. Thus, temperature-responsive chromatography which can change the properties for HPLC by changing only the column temperature have advantages in maintaining the biological activity of proteins and in reduced pollution from an organic solvent.