Abstract: To invest igat e the effect of Hedyotis diffusa extract on apoptosis of human gastric cancer MKN-45 cells. The MKN-45 cells were treated with different concentrations of Hedyotis diffusa extract, and then the quantity and morphology were observed under light microscopy and laser confocal microscopy. The apoptosis rate was detected by flow cytometry. The mRNA and protein expression levels of apoptosis-related genes Bax and Bcl-2 were detected by RT-qPCR and Western Blot. The results showed that after treatment with different concentrations of Hedyotis diffusa extract for 48 hours, the cell volume shrinkage, nucleus cleavage and chromatin morphological changes were observed under the microscope. The apoptosis rate increased with the increase of drug concentration,and the apoptotic rate reached 23.6% after treated with 30 μg/mL of Hedyotis diffusa extract the expression level of Bax gene significantly increased, and Bcl-2 significantly decreased. In summary, Hedyotis diffusa extract can induce apoptosis of human gastric cancer MKN-45 cells, which has potential medical and medicinal value.