The objective of the present study was to elucidate the mechanism by which glycyrrhizin enhances the antitumor activity of cisplatin in non-small cell lung cancer. Initially, A549 cells were treated with different concentrations of glycyrrhizin (0.25-8 mM) or cisplatin (10-160 µM) for 48 h to investigate the effect of glycyrrhizin combined with cisplatin on A549 cells in vitro. Subsequently, A549 cells were divided into control (untreated), CP (20 µM cisplatin), GL (2 mM glycyrrhizin) and CP + GL (20 µM cisplatin + 2 mM glycyrrhizin) groups to elucidate the underlying mechanism of glycyrrhizin. After 48 h incubation, the viability and colony-forming ability of the cells were assessed using MTT and colony formation assays. Apoptosis levels and cell cycle progression were analyzed using flow cytometry and western blotting was used to evaluate apoptosis- and cell cycle-related proteins. Additionally, comet assays and western blotting were used to evaluate DNA damage and relevant proteins. The results demonstrated both glycyrrhizin and cisplatin individually reduced A549 cell viability in a concentration-dependent manner. Cisplatin demonstrated a lower half-maximal inhibitory concentration (IC50) at higher glycyrrhizin concentrations, with an IC50 value of ~35 µM with 2 mM glycyrrhizin. Furthermore, the combined treatment of glycyrrhizin and cisplatin synergistically reduced cell colony-forming ability, induced apoptosis and arrested the cell cycle at the G2 phase, showing greater efficacy when compared with either treatment individually. In addition, western blotting analysis demonstrated that, in comparison with treatment with cisplatin or glycyrrhizin alone, the combined treatment markedly increased the protein expression levels of B-cell lymphoma 2-associated X protein, cleaved-caspase-3/caspase-3, γH2AX, phosphorylated-checkpoint kinase 1 and phosphorylated-p53/p53, while notably reducing the protein levels of B-cell lymphoma 2, cyclin D1, cyclin-dependent kinase 2 and cyclin-dependent kinase 4. The findings of the present study indicate that glycyrrhizin enhances the antitumor efficacy of cisplatin in non-small cell lung cancer cells by modulating DNA damage and apoptosis.