Abstract A successful post‐surgical implant is associated with accelerated recovery periods, involving the efficient regeneration of lost or non‐viable tissue and a reduction in microbial growth. Alternatively, the long‐term success of an implant is guided by the selection of an engineered biomimetic material that is biocompatible, non‐biodegradable, and stable at the site of implantation, without invoking any non‐essential or undesirable biological responses. The potential for developing an injectable bone substitute (IBS) was investigated here. In particular, carrageenan (CG) and nano‐hydroxyapatite (nHA) injectable composites were fabricated by chemical cross‐linking, and the in vitro behavior of mammalian cells and bacteria on the IBS surface structures were evaluated. Formulations consisting of 1%, 1.5%, and 2.5% CG and 60% nHA by weight were then evaluated for their interactions with human osteoblasts (or bone forming cells). MTS viability testing indicated that osteoblast adhesion and viability on the IBS were excellent and uniform among various formulation types. Bacteria assays were also performed to assess antimicrobial functions on the CG/nHA composite against both Gram‐negative and Gram‐positive strains. A higher CG content, as found in some samples, correlated with improved Pseudomonas aeruginosa growth inhibition, although other bacteria strains appeared unaffected by the IBS. In summary, this study highlights CG/nHA composites as innovative biomaterials that should be further studied for reduced bacteria activity and promoted osteoblast responses which was achieved without using pharmaceutical drugs. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 2984–2993, 2018.