Liposomes were used to encapsulate rifampicin (RIF) as an alternative formulation for delivery to the respiratory tract. Factors affecting the stability of liposomes containing RIF were determined. Four liposome suspensions were prepared, containing different millimole ratios of cholesterol (CH) and soybean L-infinity-phosphatidylcholine (SPC) by the chloroform film method, followed by freeze-drying. Cryo-transmission electron microscopy, photon correlation spectroscopy, (2)H and (31)P solid-state nuclear magnetic resonance were used to characterize the liposome suspensions. Differential scanning calorimetry and X-ray diffraction were used to examine the properties of the powder formulations. The powder was dispersed through an Andersen cascade impactor to evaluate the performance of the aerosolized powder. The liposomes were a mixture of 200-300 nm unilamellar and multilamellar vesicles. Higher CH content in the liposome formulation resulted in a smaller change in size distribution with time, and higher CH content was associated with an increase in the (2)H NMR splitting, indicative of an increase in order of the lipid acyl chains. Furthermore, the SS-NMR results indicated that RIF was located between the acyl chains of the phospholipid bilayer and associated with CH molecules. Fifty percent encapsulation of RIF was obtained when the lipid content was high (SPC 10 mM: CH 10 mM). Mannitol was found to be a suitable cryoprotectant, which is attributed to its crystallinity, and use of mannitol gave particles with a mass median aerodynamic diameter of less than 5 microm. In terms of chemical stability, RIF in dry powder formulations was considerably more stable when compared to RIF aqueous solutions and RIF liposomal suspensions.