The separation speed is very important to the development of two dimensional liquid chromatography. The viscosity of the mobile phase decreases and the analyte diffusivity increases at higher temperature. An ion exchange chromatography column was used as the first dimension mode and reversed phase chromatography as the second dimension mode, which were connected by the interface consisting of a ten-port valve and two trap columns. A high temperature two dimensional weak anion exchange (WAX)/ reversed phase (RP) liquid chromatography system was developed by the second dimensional column operated at 80 ℃ and higher flow rate up to 3 mL/min. A tryptic digest of 4 proteins was separated on this system. 33 fractions from the first dimension were introduced to the second dimension by the interface. 187 peaks were separated by the high temperature two dimensional liquid chromatography system.

Key words: high temperature, two-dimensional liquid chromatography, ion exchange/reversed phase liquid chromatography, peptide