The aim of this study was to prepare two types of emulsions, bufadienolides-loaded nano-emulsion (BU-NE) and submicro-emulsion (BU-SE) which were separately prepared by ultrasonic-high-pressure homogenization (UHPH) and high-pressure homogenization (HPH) methods, and to try to stabilize the colloid systems by lyophilization. The lyoprotective effects of cryoprotectant carbohydrates during the freeze-thawing and freeze-drying cycles on the emulsions were investigated in detail. The lyophilized products were characterized with regard to their appearance and particle size by transmission electron microscopy (TEM), scanning electron microscopy (SEM), photon correlation spectroscopy (PCS) and zeta potential. The median diameter, polydispersity index (PI) and zeta potential of BU-NE and BU-SE were 43.5 nm versus 161.4 nm, 0.100 versus 0.143 and -19.7 to -26.2 mV versus -29.4 to -35.3 mV, respectively. With the same drug content (0.28 mg mL(-1)) BU-SE exhibited a higher entrapment efficiency than BU-NE. The optimum cryoprotectant for BU-NE and BU-SE was maltose (20%) and trehalose (20%), respectively. The median diameters (95.7 and 168.5 nm) of the rehydrated BU-NE and BU-SE were slightly increased. For both of them, the bufadienolides entrapment efficiency was reduced whereas the drug content was not. The lyophilized BU-NE and BU-SE powders were stable over a period up to 3 months with no change in visual appearance, reconstitution ability, particle size distribution and drug concentration. This shows that freeze-drying could be a promising method to stabilize the emulsions.