ANALYSIS OF PEPTIDES AND PROTEINS BY TEMPERATURE-RESPONSIVE CHROMATOGRAPHIC SYSTEM USING N-ISOPROPYLACRYLAMIDE POLYMER-MODIFIED COLUMNS

A new method of HPLC using packing materials modified with a temperatu re responsive polymer, poly(N-isopropylacrylamide) (PIPAAm), was devel oped. Homogeneous PIPAAm polymer and its copolymer with butyl methacry late (BMA) were synthesized and grafted to aminopropyl silica by activ ated ester-amine coupling and they were used as packing materials. The surface properties and functions of the stationary phases are control led by external temperature. Isocratic elution by aqueous mobile phase alone is the basis for separation of peptides and protein. The separa tion of the mixture of three peptides, insulin chain A and B and beta- endorphin fragment 1-27 was achieved by changing the column temperatur e with 0.9% NaCl aqueous solution as the sole eluent. Retention of pep tides and proteins was controlled both by column temperature and by Na Cl concentration in the aqueous mobile phases in this chromatographic system. (C) 1997 Elsevier Science B.V.