This study describes a method for separation and purification of polyphenolics of ethyl acetate extract (EAE) of Trapa bispinosa Roxb. pericarps (TBP) by macroporous resins. The abilities of static absorption and desorption of seven kinds of macroporous resins were studied. The best macroporous resin D101 was selected. The dynamic absorption and desorption conditions were optimized as follows: 5 mg/mL of the sample concentration, 1 mL/min of the flow rate of sample loading, 70% of ethanol as the eluent, 4 mL/min of the elution flow rate and 5 BV of the elution volume. After enrichment by D101 macroporous resin, the content of total polyphenols increased from 478 mg GAE/g in freeze dried extracts to 825 mg GAE/g in purified samples by 1.73 times, and the recovery was 59.43%. In the scale-up test, the recovery and purification rate did not change with the increase of macroporous resin, indicating that the optimized conditions were suitable for large-scale production.