Abstract: The paper proposes an identification model of axial dispersion coefficient, phase e-quilibrium constant and lumped mass transfer coefficient in the process of large scale liquid chromatography separation. The parameters are determined by using pulse response experiment technique and liquid chromatography technique. The results show that (1) axial dispersion coefficient increases as the fluid velocity and packing density of fixed bed increase; (2) eddy dispersion is dominant in axial dispersion; (3) the effect of phase equilibrium constants on the chromatographic separation are more sensitive than lumped mass transfer coefficients; (4) phase equilibrium constants decrease as the temperature rises, lumped mass transfer coefficients increase as temperature rises and fluid velocity increases and (5) theoretical elution curves are in good agreement with experiment elution curves.