Abstract: OBJECTIVE To Study on the nanofiltration separation mechanism of catalpol of Rehmannia glutinosa in ethanol-water solution. METHODS Based on solution-diffusion effect， charge effect and molecular sieving in nanofiltration separation， the influence of ethanol concentration on membrane flux， rejection and swelling coefficient， the mass transfer coefficient of catalpol was fitted with series of ethanol concentration to analyze the nanofiltration separation mechanism. RESULTS The experiment showed that there was a linear relationship between operation pressure and membrane flux. Meanwhile， the membrane flux gradually decayed and the absolute value of swelling coefficient increased with the increase of ethanol concentration. The rejection of catalpol increased from 50.14% to 91.16%， and the mass transfer coefficient decreased from 19.27×10^-6 m/s to 9.72×10^-6 m/s with the swelling of pore diameter in ethanol-water solution. CONCLUSION The molecular interaction force among catalpol， membrane material and solution are changed by ethanol. The charge rejection effect between nanofiltration membrane and catalpol is strengthened with the swelling of pore diameter. The nanofiltration separation mechanism of catalpol is preliminarily clarified in ethanol-water solution， which provides theoretical and technical support for concentrating heat-sensitive traditional Chinese medicine ingredients from the organic solvent-water solution at normal temperature.