A novel chiral acetylenic monomer derived from cholic acid was synthesized and structurally characterized. The monomer underwent precipitation polymerization in tetrahydrofuran/n-heptane mixed solvent with [Rh(nbd)Cl]2 as catalyst. Without adding porogen, porous microspheres were successfully prepared in a high yield (>80 wt %). The formation mechanism of the porous structure was proposed. Circular dichroism and UV–vis absorption spectra demonstrated that the porous microspheres possessed optical activity. The optical activity was originated in the chiral helical conformations of substituted polyacetylene forming the microspheres. The porous microspheres were further used as specific chiral additive to induce enantioselective crystallization of racemic BOC-alanine, in which BOC-l-alanine was preferentially induced forming rod-like crystals with e.e. of 69%. This strongly indicates the significant potential applications of the porous microspheres in chiral technologies. The present study also provides a new approach to prepare chiral porous polymer microspheres.