Isoliquiritigenin, a flavonoid extracted from licorice root, has been shown to be active against most cancer cells; however, its antitumor activity is limited by its poor water solubility. The aim of this study was to develop a stable isoliquiritigenin nanosuspension for enhanced solubility and to evaluate its cytostatic activity in A549 cells. The nanosuspension of isoliquiritigenin was prepared through wet media milling with HPC SSL (hydroxypropyl cellulose-SSL) and PVP K30 (polyinylpyrrolidone-K30) as stabilizers, and the samples were then characterized according to particle size, zeta-potential, SEM (scanning electron microscopy), TEM (transmission electron microscopy), DSC (differential scanning calorimetry), XRPD (X-ray powder diffraction), FTIR (Fourier transform infrared spectroscopy), XPS (X-ray photoelectron spectroscopy), and release. The isoliquiritigenin nanosuspension prepared with HPC SSL and PVP K30 had particle sizes of 238.1 ± 4.9 nm and 354.1 ± 9.1 nm, respectively. Both nanosuspensions showed a surface charge of approximately - 20 mV and a lamelliform or ellipse shape. The dissolution of isoliquiritigenin from the 2 nanosuspensions was markedly higher than that of free isoliquiritigenin. studies on A549 cells indicated that the cytotoxicity and cellular uptake significantly improved after treatment with both nanosuspensions in comparison to the isoliquiritigenin solution. Furthermore, cell apoptosis analysis showed a 7.5 - 10-fold increase in the apoptosis rate induced by both nanosuspensions compared with pure drug. However, the cytotoxicity of pure drug and nanosuspension on normal cells (HELF) was lower, which indicated both isoliquiritigenin nanosuspensions have low toxicity to normal cells. Therefore, the isoliquiritigenin nanosuspension prepared with HPC SSL and PVP K30 as stabilizers may be a promising approach to improve the solubility and cytostatic activity of isoliquiritigenin.