Abstract: Chromatographic packing materials containing poly(N-isopropylacrylamide-co-2-acr-ylamido-2-methyl propane sulfonic acid) and poly(N-isopropylacrylamide-co-N-tert-butylacryla-mide) were prepared via conventional radical polymerization. Characterization was performed with thermogravimetric analysis (TGA) and elemental analysis to determine the grafting ratio. Mixtures of Val, Leu, Phe, Trp, Art and Ala derivatized with phenylisothiocyanate were separated using prepared stationary phases by HPLC. The results indicated that separation can be accomplished only by controlling column temperatures, which proves that chromatographic retention of analytes alters with the change of column temperature, and the materials are of temperature-responsive property.