Aim To analyse the antimicrobial and biological properties of a new bioceramic intracanal medicament (Bio‐C Temp), and to compare it with two calcium hydroxide‐based intracanal medicaments (Calen® and UltraCal® XS). Methodology The direct contact and the crystal violet tests were performed to assess the antimicrobial activity of intracanal medicaments against Enterococcus faecalis. The cytocompatibility and the effect of the medication on the biology of the human osteoblast‐like cell line (Saos‐2) were evaluated with methylthiazole tetrazolium (MTT), neutral red, alkaline phosphatase activity and mineralization (alizarin red) assays. The data were analysed using one‐way anova and Tukey’s tests, two‐way anova and Bonferroni’s tests, or Kruskal–Wallis and Dunn’s tests (α = 0.05). Results Bio‐C Temp had significantly less antibacterial activity and biofilm biomass reduction than the other intracanal medicaments (P < 0.05). There was no difference in the viability of Saos‐2 exposed to the various intracanal medicaments, except regarding the 1 : 2 dilution, when the Bio‐C Temp group had significantly lower cell viability than the UltraCal® XS and Calen® groups (P < 0.05). Bio‐C Temp induced significantly greater ALP activity than the other intracanal medicaments (P < 0.05) at day 1. Calen® induced significantly greater deposition of mineralized nodules than the other intracanal medicaments (P < 0.05), and no difference was observed between Bio‐C Temp and UltraCal® XS (P > 0.05). Conclusions Bio‐C Temp had similar cytocompatibility at higher dilutions, and higher or similar induction of ALP activity and deposition of mineralized nodules in comparison with Calen® and UltraCal® XS. However, it had significantly less antibacterial and antibiofilm activity than Calen® and UltraCal® XS.

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