Herein, two highly stable β-CD and α-Alkenyl-β-CD (α-Alk-β-CD) monolithic columns were fabricated by copolymerizing β-CD and α-Alk-β-CD with GMA onto the pretreated capillary by a simple one-pot strategy. 1H NMR, SEM, FT-IR, TGA and BET surface area analyses were applied to characterize the prepared monolithic columns. The monoliths were evaluated on the enantioseparations of antibiotic drugs, antihistamine drugs, anticholinergic agents, antifungal drugs and pesticides. In contrast to the native β-CD monolith, the derived α-Alk-β-CD reveals significantly improved enantiodiscrimination performance for the analytes especially for pesticides. Through run-to-run (n = 6), day-to-day (n = 9), and column-to-column (n = 9) investigations, the RSDs values were all lower than 3.72 %. Molecular simulation reveled that the chiral recognition between α-Alk-β-CD and chiral drugs was an exothermic process mainly governed by interaction hydrogen bindings, resulting in different binding strength between a pair of isomers compounds and α-Alkenyl-β-CD, and then leading to different migration and finally achieving effective chiral separation.