Objective: Today, esophageal cancer (EC) has become one of the most common cancer types in China. Therefore, new drug and therapeutic strategies are urgently needed to improve postoperative survival rate of patients with EC. As a food additive, several lines of evidence have shown that citric acid can be served as glycolysis suppressor to inhibit growth of some tumor cells. However, little is known about the effect of this organic acid on the growth of human esophageal carcinoma cell line, EC109. Materials and Methods: In this experimental study, cell proliferation rate was determined using MTT assay. Apoptotic morphological changes were evaluated by fluorescent microscopy using Hoechst 33258 staining. Cell apoptosis rate and mitochondrial membrane potential (MMP) were detected using flow-cytometry. Effect of citric acid on cellular membrane permeability was assessed by measuring lactate dehydrogenase (LDH) activity, using LDH assay kit. Results: Compared to the control group, there was a marked decrease in cells proliferation when the cells were treated with higher citric acid concentrations (800, 1600 mu g/ml). Typical apoptotic morphology of EC109 cells was observed upon treatment with citric acid, such as chromatin condensation and appearance of apoptotic body. Cell apoptotic indexes were significantly increased (P<0.01) after treatment with citric acid at the concentration of 400-1600 mu g/ml. Extracellular LDH activity and loss of MMP in all of the treated groups were significantly higher than control (P<0.05), in a dose-dependent manner. Conclusion: These results suggest that citric acid prevents EC109 cell growth by inhibiting cell proliferation and inducing apoptosis, which perhaps offers some theoretical guidance for its application in EC treatment.