Background & Objective: Spray drying techniques are used to produce inhaled powders of enzymes and proteins without denaturation. The surface free energy data of these powders is used to reduce the time of formulation development. During storage, the surface free energy of the inhaled protein powders could change and so their performance will be affected. Many factors affect the limit of this change. Storage temperature is the critical factor. Material& methods: A spray drying method was optimized to prepare inhaled spray dried lysozyme powders. Inverse gas chromatography was utilized to measure the surface free energy of the prepared powders pre and post-storage to detect the effect of storage temperature on the surface free energy. Differential Scanning Calorimetry and biological activity tests were used to evaluate the integrity of lysozyme conformation after storage at different storage temperature. Results: Increasing the storage temperature decreased both dispersive and specific surface components of the spray-dried lysozyme powders significantly (ANOVA: P < 0.001) up to 12% and up to 31 % after storage for one year at 40 °C, respectively. The overall decrease in the polar components resulted in a reduction of the surface basicity. These decreases in the surface energetics would be due to the aggregation of denatured lysozyme molecules at the powder surface via hydrophobic regions rich with electrons. The same results were found after storage for one year at room temperature, but the extents of changes were smaller. Conclusion: It is advisable to store protein powders at 5°C to inhibit the changes in their surface free energy which is the crucial factor affecting their performance.