LC-MS Based Targeted Isolation of Cyclopeptide Alkaloids and Triterpenoids from Ziziphus jujuba Roots with Development of Dereplication Methods

Abstract

Ziziphus jujuba Mill. is the most economically important species in the family Rhamnaceae. Among hundreds of previously reported constituents, cyclopeptide alkaloids and ceanothane-type triterpenoids have been regarded as chemotaxonomic marker compounds of the Rhamnaceae species, and they have been focused as new drug candidates for their characteristic structures and bioactive potentials. The goal of this study was the development of targeted isolation strategy for bioactive cyclopeptide alkaloids and triterpenoids from Z. jujuba. UHPLC-qTOF-MS based dereplication-guided fractionation was selected as a main platform for this purpose. The analytical methods were optimized for LC-MS profiling of cyclopeptide alkaloids and triterpenic acids in MeOH extracts of Z. jujuba roots, twigs, leaves, and fruits. Based on UV, MS, and MS/MS spectral data, the dereplication methods were developed for cyclopeptide alkaloids and triterpenoids. Cyclopeptide alkaloids were hard to be fully predicted, but many triterpenoids could be tentatively identified with their spectra data. Based on these dereplication results, LC-MS profiles of extracts were compared for their potential as the isolation target. The root extract was selected to be a target for further separation, because of its abundance in both target compound classes. MS-guided fractionation resulted in the isolation of 9 cyclopeptide alkaloids (1-9), 37 triterpenoids (10-46), and 6 phenolic compounds (47-52) from the root extract. Among these 52 compounds, 5 cyclopeptide alkaloids (1-5) and 21 triterpenoids (16-20, 23, 24, 26, 27, 30, 33, and 35-43) were isolated from nature for the first time. In-house LC-MS library was built with the retention time and spectral data of isolated compounds, and it was confirmed that tentatively predicted structures of triterpenoids were correctely matched to their real structures. In addition, it was suggested that dereplication method for cyclopeptide alkaloids could be developed in the further study. Bioactivity of isolated compounds were tested, and as a result, antiviral activity of cyclopeptide alkaloid compounds 2, 3, and 6 against PEDV (EC50 4.49-3.41 μM) and cytotoxicity of triterpenic compounds 10, 22, 28-30, 33, 35, 36, and 41 against HepG2 cell line (IC50 1.93-9.24 μM) were evaluated. In conclusion, it was demonstrated that developed UHPLC-qTOF-MS based derelication method was an efficient strategy for the discovery of bioactive cyclopeptide alkaloids and triterpenoids from Z. jujuba.