The separation of acetylene (C 2 H 2 ) from carbon dioxide (CO 2 ) and methane (CH 4 ) is of great significance, but remains challenging, because of their similar physicochemical properties, and it currently has received substantial research interest by using adsorptive separation based on metal−organic frameworks (MOFs). Herein, a new microporous Cu-MOF, [Cu(5-OH-IPA 2− )-(DPYA)(H 2 O)] (BUT-318, where 5-OH-IPA 2− = 5-hydroxyisophthalate and DPYA = 4,4′-dipyridylamine) has been synthesized successfully under solvothermal conditions, which exhibits excellent separation performance for C 2 H 2 /CO 2 and C 2 H 2 /CH 4 gas mixtures. The presence of Cu(II) open metal sites and different Lewis base sites (−OH and −NH) make the activated BUT-318 efficiently bind C 2 H 2 and exhibit high adsorption capacity under low pressure (24.52 and 9.1 cm 3 g −1 under 0.01 bar, at 273 and 298 K, respectively). The corresponding IAST (ideal adsorbed solution theory) selectivity was 9.8 and 244.3 at 273 K and 1 bar for an equimolar C 2 H 2 /CO 2 and C 2 H 2 /CH 4 mixture. The separation performance and reusability under dynamic conditions were also confirmed by column breakthrough experiments, making BUT-318a a promising candidate for the practical C 2 H 2 separation.