The separation of metallic ions in particular heavy metallic ions in aqueous solution is a great challenge in membrane science. Herein, a simple and facile strategy -Ca2+-incorporated layer-by-layer (LbL) assembly and CO2-induced biomineralization was developed and applied to the preparation of nacrelikestructured multilayered PEI/PSS-CaCO3 nanocomposite membrane on PAN substrate. These nanocomposite membranes showed higher rejection (>= 99.3%) and permeable flux ( > 49.1 L m-2 MPa-1 h(-1)) for divalent metallic ions such as Mg2+, Cu2+,Ca2+, Ni2+, Zn2+, and Cd2+ in aqueous solution compared to the multilayer PEI/PSS or PEI/PSS-CaCO3 membrane prepared by alternate soaking process. High rejection and flux can be attributed to the following three aspects: (i) The incorporation of Ca2+ into PEI and PSS greatly reduces the formation of crosslinked structure of PEI and PSS and probably induces the ordered arrays of amine and sulfonate groups from PEI and PSS on membrane; (ii) Due to limited Ca2+ ions source and spatial limitation of crosslinking of PEI/PSS phase the nanostructured CaCO3 was in-situ formed after CO2-induced biomineralization and created some additional short paths (the bulgy section of PEI/PSS layer) that are beneficial to the permeability of membrane; (iii) Due to the calcification of incorporated Ca2+ ions the formed H ions makes amine and sulfonate groups further protonation that improves surface hydrophilicity of membrane. On the basis of a long-term performance stability, high rejection and flux, the nacrelike-structured multilayered PEI/PSSCaCO3 nanocomposite membrane can be used as a promising membrane in industry for the separation of metallic ions in aqueous solution. (C) 2015 Elsevier B.V. All rights reserved.