In this study, melatonin/2-hydroxypropyl-beta-cyclodextrin (HP beta CD) inclusion complex was prepared by using microwave irradiation in a very short time. Phase-solubility studies indicated a linear increase in the solubility of melatonin with HP beta CD demonstrating Higuchi's AL-type phase solubility profiles and the formation of 1:1 stoichiometric inclusion complexes. Also, the ability of inclusion complex formation was promoted at microwave conditions compared to room conditions. The structure of inclusion complex was determined by FTIR, DSC, XRD, H-1 NMR and C-13 NMR studies. Both of pure melatonin and melatonin/HP beta CD inclusion complex were loaded into the chitosan scaffolds that were prepared by freeze-drying method and the effect of released melatonin on human osteosarcoma cells (MG-63) was investigated in vitro. Approximately 9 mM melatonin concentration caused time dependent cell death by reducing the proportion of the cells in the G2/M phase rather than S phase. In conclusion, melatonin/HP beta CD inclusion complex loaded chitosan scaffolds could be considered as an alternative system for the human osteosarcoma therapy. (C) 2015 Elsevier B.V. All rights reserved.