Effective water removal from bio-oil is very important for upgrading bio-oil quality via esterification because water strongly inhibits the conversion and simultaneously lowers the heating value. In the present study, ZSM-5 zeolite membranes are applied to make bio-oil pervaporation dehydration more efficient. An investigation of the permeation performance demonstrated that water mainly permeated through ZSM-5 membranes via intercrystalline pores, whereas intracrystalline pores in the membrane were almost completely blocked because of intensive membrane fouling. The above result showed that tuning water transport paths was significantly important when designing high-performance ZSM-5 membranes for bio-oil pervaporation. A decrease in membrane fabrication time successfully created loose membrane structures with abundant intercrystalline pores and resulted in highly selective and stable ZSM-5 membranes with significantly improved permeation flux in bio-oil dehydration. These robust ZSM-5 membranes show great potential for bio-oil refining in industrial applications.