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A rapid ultra-performance liquid chromatography-electrospray ionization tandem mass spectrometric (UPLC-ESI-MS/MS) method is developed for the qualitative identification of constituents in the flower buds of seven Lonicera species. The optimal condition of separation and detection were achieved on an AcQuity UPLC BEH C18 column with a gradient elution with acetonitrile and 0.1% acetic acid within 17 min. Among the 33 constituents detected, 6 caffeoylquinic acids (including caffeic acid), 8 flavonoids and 8 iridoid glycosides were characterized based on their fragmentation patterns in collision-induced dissociation (CID) experiments and/or by comparison with standard compounds. In addition, to statistically establish the correlation and discrimination of the Lonicera species, principle component analysis (PCA) was applied in this study. Lonicera samples were divided into well-defined groups directly related to their species based on PCA in terms of the log transformed relative contents of the major caffeoylquinic acids (including caffeic acid) as the variables. All of results indicated that the method presented here is able to classify the sample species and to reveal characteristic details of the chemical constituents of different Lonicera species.

The aim of the present study was to investigate the mechanism of hyperoside protecting ECV-304 cells against tertbutyl hydroperoxide (TBHP)-induced injury. ECV-304 cell viability was measured by MTT assay. Cellular morphologic changes were observed using phase contrast microscopy. The genotoxic effects of TBHP and the protective ability of hyperoside were assessed by the Comet test. Lipid peroxidation was measured by HPLC method. The cellular redox status was determined from GSH/GSSG ratios. Apoptosis was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assay. Western blot analysis was used to evaluate the levels of cytochrome c, p53, SIRT1, Bax and Bcl-2 expression. The results showed that 128 µmol/l hyperoside could effectively protect TBHP-treated ECV-304 cells from death, increase superoxide dismutase activity and significantly decrease malondialdehyde production. Hyperoside was effective in protecting against the induction of oxidized DNA bases and redox state alterations induced by TBHP. Furthermore, the release of proapoptotic cytochrome c from mitochondria was reduced by hyperoside, which increased the expression of antiapoptotic SIRT1 and inhibited the translocation of Bax from cytoplasm to mitochondria. Taken together, these results indicate that hyperoside is effective in protecting against the oxidative damage induced by TBHP. The mechanism of hyperoside protecting against ECV-304 cell apoptosis by TBHP is related with resuming mitochondrial function and regulating the expression of SIRT1 and Bcl-2 family members.