Objective To investigate the influence of pH and commonly coadministered amlodipine on the transmembrane uptake transport of telmisartan. Methods Human colorectal cancer Caco-2 cells were cultured in vitro, and its monolayer integrity was confirmed by cell morphology, transepithelial electrical resistance (TEER) measurement and lucifer yellow permeability test. ATPase activity measurement and bidirection permeability experiment were performed to analyse whether telmisartan was a substrate of P-glycoprotein (P-gp). The influence of pH and amlodipine on the transmembrane uptake transport of telmisartan was assessed. Results The cultured Caco-2 cell monolayer exhibited tight junction, which was applicable for transport study. The ratio of ATPase activity of telmisartan to that of blank control was 1.73, and the efflux rate was 4.53 in the bidirection permeability experiment. With the decrease of pH from 8.0 to 5.0, the transmembrane uptake transport of telmisartan gradually increased in a pH-dependent manner. The transmembrane uptake was significantly enhanced at pH of 7.4 with coadministration of amlodipine. Conclusion The transmembrane transport of telmisartan may be mediated via P-gp, and its uptake transport is influenced by both pH and coadministration of amlodipine.