A mussel-inspired interlayer of polydopamine (PDA)/ polyethylenimine (PEI) is codeposited on the ultrafiltration substrate to tune the interfacial polymerization of piperazine and trimesoyl chloride for the preparation of thin-film composite (TFC) nanofiltration membranes (NFMs). This hydrophilic interlayer results in an efficient adsorption of piperazine solution in the substrate pores. The solution height increases with the PDA/PEI codeposition time from 45 to 135 min due to the capillary effect of the substrate pores. The prepared TFC NFMs are characterized with thin and smooth polyamide selective layers by ATR/IR, XPS, FESEM, AFM, zeta potential, and water contact angle measurements. Their water permeation flux measured in a cross-flow process increases to two times as compared with those TFC NFMs without the mussel-inspired interlayer. These TFC NFMs also show a high rejection of 97% to Na2SO4 and an salt rejection order of Na2SO4 approximate to MgSO4 > MgCl2 > NaCl.