The crystal structure of the title compound (common name HEPES), C(8)H(18)N(2)O(4)S, has been redetermined at 100 K in order to properly elucidate the protonation state of the HEPES molecule. The piperazine ring has a chair conformation and one of the N atoms in the ring is protonated, which was not previously reported [Gao, Yin, Yang, & Xue (2004). Acta Cryst. E60, o1328-o1329]. The change of protonation state of the nitrogen atom significantly affects the intermolecular interactions in the HEPES crystal. The structure is stabilized by N-H⋯O and O-H⋯O hydrogen bonds and ionic inter-actions, as the title compound in solid state is a zwitterion. HEPES mol-ecules pack in layers that are held together by ionic and weak inter-actions, while a hydrogen-bonded network connects the layers.