Additively manufactured (AM) iron (Fe)-based scaffolds have been developed as promising biodegradable bone-substituting biomaterials. Multi-material extrusion-based 3D printing has recently yielded Fe-manganese (Mn) alloy-based scaffolds that can resolve ferromagnetism and cytotoxicity associated with Fe-based biomaterials. Herein, we, for the first time, present the findings from in vivo study on extrusion-based AM FeMn-akermanite (Ak) scaffolds for critical-size bone defect repair. The scaffolds comprised Fe, 35 wt% Mn, and 20 or 30 vol% Ak, with microporous struts and 61–63 % porosity. Both scaffolds exhibited mechanical properties within the range of trabecular bone and provided suitable sites for Ca/P deposition during in vitro biodegradation. In vitro cell cultures demonstrated favorable cell responses without negating the osteogenic potential of cells. An in vivo study was conducted in a murine semi-orthotopic subcutaneous model. With this model, 4 bovine bone plugs were implanted subcutaneously with critical-size defects created at their cores. Scaffolds were placed into these critical-size defects to assess biodegradation and bone formation. After 16 weeks, the volume of scaffolds decreased by 6–8 %. The FeMn-20Ak scaffolds retained their yield strength and elastic modulus during the 16 weeks in vivo, whereas the mechanical integrity of FeMn-30Ak scaffolds deteriorated after mechanical push-out tests. Excellent osseointegration of both scaffold groups was apparent. 3D reconstruction of CT images revealed that FeMn-30Ak scaffolds had more newly formed tissue in the macro-pores than FeMn-20Ak. Altogether, our findings demonstrate the potential of AM FeMn-Ak scaffolds as biodegradable bone substitutes, encouraging further in vivo research in a large animal model.