Morinda citrifolia is a well-known herbal medicine that might play an important role in the promotion of bone healing. Objective: To investigate the effect of Morinda citrifolia leaf on enhancing bone healing by examining alkaline phosphatase (ALP) activity and mineralized nodule formation of human periodontal ligament (hPDL) cells in vitro. Method: The hPDL cells from healthy donors were cultured in DMEM complete medium. Cells were treated with 2.5% (w/v) Morinda citrifolia leaf extract as experimental group, with 2 mM β-glycerophosphate and 50µg/ml ascorbic acid as positive control, and with complete medium as negative control. After 6 weeks, cells were assayed for ALP activity and the formation of mineralized nodules was tested with Alizarin red S and X-ray diffraction analysis. Result: Morinda citrifolia leaf extract significantly enhanced the ALP activity and induced matrix mineralization that was positively stained with Alizarin red S. X-ray diffraction analysis revealed that the composition of these mineralized nodules was rich in calcium and phosphorus. Conclusion: Our results suggest that Morinda citrifolia leaf extract is the suitable natural product for inducing bone regeneration in periodontal disease.