Abstract: The structures of phenylethanoid glycosides in plant extract of Cistanches deserticola were successfully analyzed and identified by electrospray ionization multi-stage mass spectrometry (ESI-MSⁿ) in negative mode, some interesting neutral losses were found, which helped us to confirm the structures of compounds without tedious separation and purification procedures, then the fragmentation mechanisms were explored. The identification of the phenylethanoid glycosides in plant extract of Cistanche deserticola was based on the comparison of molecular ions, and the fragment ions obtained by MSⁿ experiments with those of the authentic standards and the data was reported in the literatures.