The fragment ions of rhein in total ions mass chromatogram and collision inductive dissociation multi-level mass were studied by electrospray ionization ion trap mass spectrometry(ESI-IT MS).Quantum chemistry calculations were carried out to explore the mass behavior of rhein and the main fragment ions from the molecule.By analysing the characteristic of the fragment ions including geometry parameter,bond dissociation energy,changes of charge,spin density and frontier molecular orbital,the stable structures of the fragment ions with m/z 282.8,256.9,238.9,210.8,192.8,182.8,166.8,and fragmentation pathways could be obtained,the cracking behavior of rhein could be explained systematically by ESI-IT MS.