To investigate the effect of ginkgolide B on the proliferation and apoptosis of human lung adenocarcinoma (A549) cells and the regulatory effect of IκB kinase/nuclear factor κB (IKK/NF-κB) signaling pathway. A549 cells were cultured in vitro and divided into control group (same volume DMSO solvent) and different concentrations (4, 8, 16, 32 μmol/L) of ginkgolide B group. After 24 hours of intervention, cell counting kit-8 (CCK-8) was used to detect cell viability to screen the optimal concentration of ginkgolide B for subsequent experiments. Then A549 cells were divided into control group, ginkgolide B group (16 μmol/L ginkgolide B), positive drug group (0.4 mg/L adriamycin), inhibitor group (16 μmol/L ginkgolide B+10 μmol/L IKK/NF-κB pathway inhibitor BAY11-7082) and activator group (16 μmol/L ginkgolide B+1 μmol/L IKK/NF-κB pathway activator Prostratin), intervention for 24 hours. 5-ethynyl-2′-deoxyuridine (EdU) was used to measure cell proliferation. Hoechst 33258 staining kit was used to detect cell apoptosis. Western blot (WB) was used to detect the expression levels of CyclinD1, Caspase-3 and IKK/NF-κB pathway-related proteins. According to the results of CCK-8 experiment, 16 μmol/L ginkgolide B was selected for subsequent experiments. Compared with the control group, the proliferation rate of A549 cells and the protein expression levels of CyclinD1, p-NF-κB p65/NF-κB p65 and p-IκBα/IκBα in the ginkgolide B group and the positive drug group significantly decreased (P<0.05), and the apoptosis rate and Caspase-3 protein expression level significantly increased (P<0.05). Compared with ginkgolide B group, cell proliferation rate, CyclinD1, p-NF-κB p65/NF-κB p65 and p-IκBα /IκBα ratio significantly decreased in inhibitor group (P<0.05), while apoptosis rate and Caspase-3 protein expression level significantly increased (P<0.05). In the activator group, cell proliferation rate, CyclinD1, p-NF-κB p65/NF-κB p65 and p-IκBα/IκBα ratios significantly increased (P<0.05), while apoptosis rate and Caspase-3 protein expression levels significantly decreased (P<0.05). Ginkgolide B can inhibit the proliferation and induce apoptosis of A549 cells by regulating IKK/NF-κB pathway signaling.