Abstract: AIM To investigate the relative bioavailability of pulmonary-delivered insulin lipid suspension (INS-LIP-SP) in normal Wistar rats. METHODS INS-LIP-SP were prepared by two different methods and then delivered to the rat lung using an intratracheal instillation method. Blood glucose levels and INS concentrations in serum were determined by glucose oxidase method and radioimmunoassay method, respectively. The relative pharmacological bioavailability (F%) and relative bioavailability (F%) of INS-LIP-SP were calculated from the area above the curve (AAC) and the area under the curve (AUC) compared with subcutaneous injection of INS solution. RESULTS The mean particle diameter, span of dispersity and entrapment efficiency of INS-LIP-SP prepared by a membrane-formed with sonication method and a reversed phase evaporation method were 1.91 μm, 0.94 and 16.45% and 2.08 μm, 1.28 and 39.51%, respectively. The values of F% and F% of both INS-LIP-SP were up to 37% and 32%, separately, and there was a statistically significant difference between INS-LIP-SP and INS solution. However, there was no significant difference between the two INS-LIP-SP and the physical mixture of INS solution and blank liposomes. CONCLUSION The results showed that INS-LIP-SP could achieve higher bioavailability following pulmonary delivery to rats.