To explore whether ethyl acetate extract of Liujunzi Decoction（EAELD）can regulate the energy metabolism of TE-1 of esophageal cancer cells via the microRNA-1（miR-1）/hypoxia inducible factor-1α（HIF-1α）axis.

Stable transfection blocking miR-1 expression cell lines（miR-1 inhibitor，IN）and blank controls（miR-1 NC，NC）have been established in the preliminary experiments. The half maximal inhibitory concentration（IC₅₀）and 30% inhibitory concentration（IC₃₀）of EAELD on NC were obtained using MTT assay，and were used as the Liujunzi Decoction high and low concentrations to compare their effects on IN and NC，respectively. The effect of EAELD on miR-1 and HIF-1αmRNA in TE-1 cells was measured by quantitative real-time PCR（RT-qPCR）. Dual-luciferase gene reporter assay was used to validate the targeting relationship of miR-1 and HIF-1α. Then the experiments were divided into NC，Liujunzi Decoction low concentration，Liujunzi Decoction high concentration，IN＋Liujunzi Decoction low concentration and IN＋Liujunzi Decoction high concentration groups. Seahorse energy metabolism analysis system detected the mitochondrial pressure and glycolysis rate，and the protein expression level was detected by Western Blot.

MTT results showed that EAELD could significantly inhibit the proliferation of NCin a concentration gradient dependent manner. The low and high concentration groups were 20 μg/mL（IC₃₀）and 40 μg/mL（IC₅₀），respectively. The RT-qPCR results showed that the EAELD high concentration group significantly promoted the gene expression of miR-1（P<0.05）and HIF-1αshowed a downward trend，without statistically significant difference. Dual-luciferase gene reporter assay showed that the fluorescence intensity of wild type（WT）＋mimics group was significantly lower than that of WT＋negative control（mimics NC）group（P<0.05），and there was no significant difference in the fluorescence intensity of mutant（MUT）＋mimics group compared with that of MUT＋mimics NCgroup. When high and low concentrations of EAELD were applied to IN，the MTT results showed that compared with that of NCgroup，the inhibitory effect of EAELD on IN group was significantly reduced（P<0.05）. Seahorse energy metabolism results showed that compared with those of NC group，the ratio of mitochondrial oxygen consumption to glycolysis（P<0.01，P<0.05），glycolytic potential（P<0.01），basal respiration value，proton leakage oxygen consumption（P<0.01，P<0.05），proton leakage oxygen consumption（P<0.01，P<0.05），mitochondrial maximum oxygen consumption capacity and spare respiratory capacity（P<0.01）were inhibited in the high and low concentration groups of Liujunzi Decoction. The high concentration group could inhibit basal glycolysis，compensate glycolysis，non-mitochondrial oxygen consumption and ATPsynthesis oxygen consumption（P<0.01）. Compared with those of NC＋Liujunzi Decoction low concentration group，basal glycolysis，mitochondrial to basal glycolysis ratio，glycolytic potential（P<0.05）and oxygen consumption for ATPsynthesis（P<0.01）were significantly increased in IN＋Liujunzi Decoction low concentration group. Compared with those of NC＋Liujunzi Decoction high concentration group，basal glycolysis，compensatory glycolysis（P<0.05），basal respiratory capacity，proton leakage oxygen consumption，maximum mitochondrial oxygen consumption and non-mitochondrial oxygen consumption（P<0.01）were significantly increased in IN＋Liujunzi Decoction high concentration group. WB results showed that compared with NCgroup，EAELD high and low concentration groups significantly decreased the protein relative expression levels of HK2（P<0.01），GLUT1（P<0.05，P<0.01）and MCT1（P<0.01，P<0.05）. The relative protein expressions of PKM2（P<0.05），MCT2 and MCT4（P<0.01）in the high concentration of EAELD group were significantly decreased. Compared with those of NC＋Liujunzi Decoction low concentration group，the expressions of HK2，GLUT1，PKM2（P<0.01），p-PKM2，MCT1 and MCT2（P<0.05）in IN＋Liujunzi Decoction low concentration group were significantly increased. Compared with those of NC＋Liujunzi Decoction high concentration group，the expressions of HK2，GLUT1，PKM2（P<0.01），p-PKM2，MCT1 and MCT2（P<0.05）in IN＋Liujunzi Decoction low concentration group were significantly increased. IN＋Liujunzi Decoction high concentration group could significantly promote the expressions of HK2，GLUT1，PKM2 and MCT4（P<0.05）.

EAELD can inhibit the energy metabolism of esophageal cancer cell line TE-1 by down-regulating the expressions of GLUT1，PKM2，p-PKM2，HK2，MCT1，MCT2 and MCT4，and the mechanism is partly related to miR-1/HIF-1αaxis.