〔Abstract〕 Objective To explore the effect of C2-ceramide, one of the sphingomyelin substances, on apoptosis of non-small cell lung cancer cells(A549 and PC9). Methods Non-small cell lung cancer cell lines(A549 and PC9) were cultured, total proteins were extracted and Western blot was performed to detect the expression of apoptotic protein Caspase-3 and cleaved Caspase-3 in the two cells. CCK-8 colorimetric method was used to screen drug concentration. Hoechst 33258 apoptosis staining was used to detect apoptosis. The apoptosis rate was detected by flow cytometry, and the expression of apoptotic protein Caspase-3 was detected by RT-PCR. Results The cell viability was about 70% when ceramide was treated at 50 μmol/L. Compared with the control group, the expression of apoptotic proteins increased in the ceramide group(P<0.05).Flow cytometry and apoptosis staining showed that the rate of apoptosis increased in the ceramide treatment group compared with the control group(P<0.05). mRNA detection at gene level showed increased the expression of apoptotic protein Caspase-3(P<0.05). Conclusion C2-ceramide can promote the apoptosis of non-small cell lung cancer cells, thus providing a new therapeutic target for clinical lung cancer chemotherapy.