The purpose of this study was to investigate the effect of four sterilization methods (Steam autoclave, Hydrogen peroxide plasma, Ethylene oxide, and Gamma sterilization) on the surface chemistry and in vitro bioactivity of polycrystalline pseudowollastonite (psW). psW samples obtained by solid-state reaction sintering were sterilized and soaked in Kokubo et al.'s proposed simulated body fluid (SBF) up to 30 days. The sterilization procedure was found to result in no significant chemical changes in the surface of the samples. On the other hand, a Ca/P layer, of different thickness, identified as hydroxyapatite (HA) like, was developed on all the samples after soaking, although the Ethylene oxide-sterilized samples present a non-homogeneous and similar to 68% thinner HA layer. The psW samples before soaking were analyzed by X-ray diffraction, raman spectroscopy, and scanning electron microscopy (SEM). The interfacial reaction product was examined by SEM fitted with an energy-dispersive X-ray analyzer. Additionally, changes in ionic concentrations at the psW/SBF interface were measured.