The enantionteric separation of a series of 13 new chiral polycycles has been examined on both cyclodextrin-based and macrocyclic glycopeptide chiral stationary phases (CSPs) using HPLC in the normal phase, reversed phase, and polar organic modes. The most effective chiral selectors for the enantiorneric separation of these analytes are the 2,3-dimethyl-beta-cyclodextrin (Cyclobond I-2000 DM) and hydroxypropyl-,beta-cyclodextrin (Cyclobond I-2000 RSP). The other Cyclobond-type and Chirobiotic (macrocyclic glycopeptide) CSPs only show enantioselectivity for a few of the racemic polycycles. The effects of mobile phase composition and analyte structure on chiral recognition and separation are considered.