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White mineral trioxide aggregate (WMTA) is a root canal treatment material, which is known to exhibit a dark brown color when in contact with sodium hypochlorite solution (NaOCl). This study aimed to investigate the effects of NaOCl on the surface properties of WMTA discs and WMTA-induced osteoblastic differentiation of periodontal ligament stem cells (PDLSCs). Mixed WMTA (ProRoot MTA) was filled into the molds to form WMTA discs. These discs were immersed in distilled water (D-WMTA) or 5% NaOCl (Na-WMTA). Their surface structures and Ca 2+ release level was investigated. Moreover, they were cultured with a clonal human PDLSC line (line 1-17 cells). The main crystal structures of Na-WMTA were identical to the structures of D-WMTA. Globular aggregates with polygonal and needle-like crystals were found on D-WMTA and Na-WMTA, which included Ca, Si, Al, C and O. However, many amorphous structures were also identified on Na-WMTA. These structures consisted of Na and Cl, but did not include Ca. NaOCl immersion also reduced Ca 2+ release level from whole WMTA discs. Line 1-17 cells cultured with D-WMTA formed many mineralized nodules and exhibited high expression levels of osteoblast-related genes. However, cells incubated with Na-WMTA generated a small number of nodules and showed low expression levels of osteoblast-related genes. These results indicated that NaOCl reduced Ca 2+ release from WMTA by generating amorphous structures and changing its elemental distribution. NaOCl may also partially abolish the ability of WMTA to stimulate osteoblastic differentiation of PDLSCs., (© 2021. The Author(s).)