OBJECTIVE： To investigate the function change of intestinal CYP3A4 and P-glycoprotein （P-gp） under diabetic conditions and its mechanism. METHODS： Caco-2 cells were respectively added with 25， 50 and 100 μmol/L midazolam （CYP3A4 probe substrate） for incubation for 15， 30， 60， 120， 180 min； with 0.1， 0.2， 0.4 μg/ml rhodamine 123 （P-gp substrate） for incubation for 15， 30， 60， 90， 120 min to determine the substrate concentrations and incubation time. Caco-2 cells were added with insulin， glucose and fatty acid （palmitic acid and oleic acid） at different concentrations， and then the production of 1′ -OH-midazolam， the metabolite of midazolam， and the intake of rhodamine 123 were determined， in order to investigated the effect on the function of CYP3A4 and P-gp. RESULTS： The optimal substrate concentrations and incubation time were as follows as 50 μmol/L midazolam， 0.1 μg/ml rhodamine 123 and incubation for 2 h. With the increase in the concentration of insulin， glucose and palmitic acid， the production of 1′ -OH-midazolam and the activity of CYP3A4 reduced； the intake of rhodamine 123 increased， and the efflux transport function of P-gp decreased. Oleic acid had no significant effect on the production of 1′ -OH-midazolam or the intake of rhodamine 123. CONCLUSIONS： Under diabetes condition， the increase of insulin， glucose and palmitic acid may reduce the function of CYP3A4 and P-gp， while oleic acid has no effect on the function.