Steroids from Helleborus caucasicus reduce cancer cell viability inducing apoptosis and GRP78 down-regulation.

Helleborus caucasicus (Ranunculaceae) is an endemic plant of the Caucasian flora, widely distributed in West Georgia. Biological activities for the extracts of some Helleborus species including H. caucasicus have been reported. In this work we found that butanolic extract of the underground parts of H. caucasicus and isolated compounds decreased cell viability in vitro on cancer cell line of lung origin (Calu-1) in a concentration-dependent manner, compared to the normal cell line. In particular, we identified that furostanol derivative (25S)-22α,25-epoxyfurost-5-ene-3ß,11ß,26-triol 26-O-ß-d-glucopyranoside (5), 20-hydroxyecdysone (6), and 3ß,5ß,14ß-trihydroxy-19-oxo-bufa-20,22-dienolide 3-O-α-l-rhamnopyranoside, known as deglucohellebrin (7) exerted a strong cytotoxic effect on the same cells and on other cancer cell lines (HepG2 and Caco-2) reducing the S-phase entry (compound 6) and inducing cell apoptosis associated with activation of caspase-3 (compound 7). Moreover we demonstrated that 6 and 7 significantly decreased protein expression of GRP78, a general ER-stress marker, suggesting pro-apoptotic functions. These findings indicated that selected compounds from H. caucasicus are potential interesting agents in anti-cancer therapy.

Steroidal glycosides from the leaves of Ruscus colchicus: isolation and structural elucidation based on a preliminary liquid chromatography-electrospray ionization tandem mass spectrometry profiling.

An HPLC-ESIMS(n) method, based on high-performance liquid chromatography coupled to electrospray positive ionisation multistage ion trap mass spectrometry, has been used as an effective tool to rapidly identify and guide the isolation of target saponins from the ethanol extract of the leaves of Ruscus colchicus Y. Yeo. Twenty-two steroidal glycosides, including seventeen furostanol, four spirostanol and one cholestane glycosides, were online identified. Subsequently, compounds were isolated and their structures were established by the extensive use of 1D- and 2D-NMR experiments. The structures identified by MS were fully consistent with those elucidated by NMR data. Sixteen steroidal glycosides, including thirteen furostanol, two spirostanol and one cholestane glycosides, were identified along with four known furostanol and two spirostanol glycosides. The saponin profile shows that the furostanol glycosides are the main constituents of R. colchicus extract, unlike the other Ruscus species, for which the spirostanol derivatives generally are reported as the major compounds. Moreover, for the first time a cholestane glycoside has been isolated from R. colchicus.

Steroidal glycosides from the underground parts of Helleborus caucasicus.

Four polyhydroxylated and polyunsaturated furostanol glycosides (1-4), named caucasicosides A (1), B (2), C (3) and D (4), were isolated from the MeOH extract of the underground parts of Helleborus caucasicus, along with four spirostanol derivatives, a furostanol glycoside, a furospirostanol glycoside, 20-hydroxyecdysone and the bufadienolides hellebrigenin and deglucohellebrin. The structures of 1-4 were elucidated as furosta-5,20(22),25(27)-triene-1beta,3beta,11alpha,26-tetrol 26-O-beta-D-glucopyranoside (1), 26-O-beta-D-glucopyranosylfurosta-5,20(22),25(27)-triene-1beta,3beta,11alpha,26-tetrol 3-O-alpha-L-rhamnopyranosyl-(1-->2)-beta-D-glucopyranoside (2), 26-O-beta-d-glucopyranosyl-22alpha-methoxyfurosta-5,25(27)-diene-1beta,3beta,11alpha,26-tetrol 3-O-alpha-L-rhamnopyranosyl-(1-->2)-beta-D-glucopyranoside (3), 26-O-beta-D-glucopyranosylfurosta-5,20(22),25(27)-triene-1beta,3beta,26-triol 3-O-beta-D-xylopyranosyl-(1-->3)-alpha-L-rhamnopyranosyl-(1-->2)-4-O-sulfo-alpha-L-arabinopyranoside (4). Structure elucidation was accomplished through the extensive use of 1D- and 2D NMR experiments including 1H-1H (COSY, 1D-TOCSY) and 1H-13C (HSQC, HMBC) spectroscopy along with ESI-MS and HR-ESI-MS. The aglycones of 1-4 have never been reported before.