Polymethylmethacrylate-based bone cements are widely used for fixation of joint replacements. To improve the long-term outcome, bioactive bone cements are aspired to advance the bone-cement interface. This study evaluated the in vivo properties of a new polymethylmethacrylate-based bioactive bone cement with addition of amphiphilic phosphorylated 2-hydroxyethylmethacrylate. Previous in vitro studies confirmed bioactive properties in cell culture, as well as unchanged mechanical properties are tests according to ISO 5833:2002.Three different variations of the cement (polymethylmethacrylate + phosphorylated 2-hydroxyethylmethacrylate, polymethylmethacrylate + phosphorylated 2-hydroxyethylmethacrylate + CaCl2 and polymethylmethacrylate + phosphorylated 2-hydroxyethylmethacrylate + CaCl2 + Na2CO3) were compared to conventional polymethylmethacrylate cement. To evaluate the properties under load-bearing conditions, a spacer prosthesis was implanted into the femoral diaphysis of 24 rabbits. Additionally, a cement plug was installed into the proximal tibia. After three months, polished sections with Giemsa surface staining were prepared. The bioactivity was determined using the bone affinity index.The sections showed a good osseointegration of the bioactive bone cement without cement cracks under load-bearing conditions. Regarding the bone affinity index, the bioactive bone cement revealed a significantly higher value in the proximal tibia (25.9-37.7%) and around the spacer prosthesis (36.8-58.9%) compared to the conventional polymethylmethacrylate cement (12.8-17.0%).The results confirm the in vivo bioactivity of this bone cement. The absence of cement cracks indicates a sufficient mechanical stability to fix prostheses with this bioactive cement, but for a final assessment long-term tests are necessary.