Objective To investigate the effects of high-dose vitamin C on proliferation and glucose uptake in lung cancer A549 cells. Method The IC50 value of A549 cells treated with vitamin C was analyzed by CCK8 method. Different concentrations of vitamin C the final concentration was 0-8 mmol / L were applied to A549 cells for 24 hours and glucose concentrations were set at 5 mmol / L and 25 mmol / L respectively. Cell proliferation was detected by CCK8 method. A549 cells were interfered with 8 mmol / L vitamin C and intracellular vitamin C concentration was detected by FRASC Ascorbic acid kit Ⅱ. Human bronchial epithelioid cells HBE cell were used as a control to analyze the time change of intracellular vitamin C concentration for 4 h. A549 cells were then divided into four groups and interfered with 5 mmol / L glucose 5 mmol / L glucose + 8 mmol / L vitamin C 25 mmol / L glucose or 25 mmol / L glucose + 8 mmol / L vitamin C. Intracellular vitamin C concentration was detected and intracellular glucose intake was detected by using fluorescence-labeled 2-deoxyglucose as a probe. The expression levels of glucose transporters 1 and 3 GLUT1 and GLUT3 were detected by qPCR and Western Blot. Result The IC50 concentration of A549 cells treated with vitamin C for 24 h was 5. 192 mmol/ L. The inhibitory effect of vitamin C on A549 cell proliferation was enhanced with the increase of concentration P< 0. 05 and the cell viability of A549 cells after 5 mmol / L glucose intervention was lower than 25 mmol / L glucose when the concentration of vitamin C was 4 or 8 mmol / L P<0. 05 . Vitamin C concentration in A549 cells increased with time and was significantly higher than that in HBE cells at 0. 5 1 2 and 4 h P<0. 05 . After 8 mmol / L vitamin C intervention for 24 h the intracellular glucose intake of A549 cells was decreased P<0. 05 and after 25 mmol / L glucose intervention the intracellular vitamin C concentration was lower than 5 mmol/ L glucose P<0. 05 . When glucose concentration was 5 mmol / L the expression levels of RNA and protein of GLUT1 and GLUT3 were decreased after 8 mmol / L vitamin C intervention P< 0. 05 while the RNA level of GLUT3 and the protein level of GLUT1 were decreased when glucose concentration was 25 mmol / L P< 0. 05 . Conclusion At low glucose concentration high-dose vitamin C down-regulated the expression of GLUT1 and GLUT3 decreased glucose uptake in A549 cells and inhibited the proliferation of A549 cells.