Potent apoptotic effects of saponins from Liliaceae plants in L1210 cells.

We isolated eight saponins, a hexacyclic lanosterol tetraglycoside (1), a 27-norlanosterol tetraglycoside (2) and six spirostanol oligoglycosides (3-8), from the plants of the family Liliaceae. In murine leukaemic L1210 cells, saponins 5 and 7 at a concentration of 1 microM showed potent cytotoxic activity and the activities were in the following decreasing order: 5, 7, 1, 3, 2, 8, 4, 6. At a concentration of 10 microM, not only 5 and 7 but also 3 and 8 markedly caused cell death. The flow cytometric analysis indicated that 7 and 8 caused a concentration- and time-dependent apoptosis of L1210 cells (EC50 value = approximately 5 microM). The morphological observation using a light microscope revealed that both 7 and 8 induced shrinkage in cell soma and chromatin condensation, suggesting apoptotic cell death. Moreover, in agarose gel electrophoretic analysis, a typical apoptotic DNA ladder pattern was observed after treatment with both 7 and 8. These results suggest that 7 and 8 caused the death of L1210 cells through the apoptotic process. These compounds may become powerful pharmacological tools for studying the molecular mechanism of apoptosis.

Cytotoxic activities and structure-cytotoxic relationships of steroidal saponins.

We have systematically examined the cytotoxic activities of the steroidal saponins mainly isolated from the Liliaceae plants against HL-60 human promyelocytic leukemia cells and found several structure-activity relationships. Some steroidal saponins evaluated in the assay system showed considerable cytotoxic activities, which were almost as potent as that of etoposide used as a positive control. The activities were found to be sensitive to the monosaccharides constituting the sugar moieties and their sequences, as well as to the structures of the aglycons.

Macrophage-oriented cytotoxic activity of novel triterpene saponins extracted from roots of Securidaca inappendiculata.

It is recognized that macrophages in peripheral tissues often proliferate under pathological conditions such as tumors, inflammation and atherosclerosis. Because the growth state of macrophages is believed to be a factor regulating the pathological process of the diseases, substances that regulate macrophage growth or survival may be useful for disease control. In this paper, we identified the activity inhibiting macrophage growth in a hot water extract of roots of Securidaca inappendiculata. The extract markedly inhibited macrophage colony-stimulating factor (M-CSF/CSF-1)-induced growth of macrophages, whereas it exerted a less potent effect on growth of Concanavalin A (Con A)-stimulated thymocytes or M-CSF-stimulated bone marrow cells. The inhibition of macrophage growth was caused by a cytotoxic effect rather than a cytostatic effect. Cell death was due to the induction of apoptosis, as judged by staining with terminal deoxynucleotidyl transferase-mediated d-UTP nick end labelling (TUNEL). The cytotoxic activity seemed to be specific to peripheral macrophages; it showed a weak effect on the growth and survival of tumor cell lines including a macrophage-like cell line, J-774.1. Moreover, the saponin fraction induced apoptotic cell death of macrophages only when they were stimulated by M-CSF; it did not affect the viability of macrophages cultured without M-CSF or with granulocyte/macrophage-CSF. We determined the structures of the two active triterpene saponin compounds in the fraction, named securioside A and securioside B having a 3,4-dimethoxycinnamic group which is essential for the cell death-inducing activity. They are believed to be the primary compounds of new drugs for the treatment of pathological states in which macrophage proliferation occurs.

New bisdesmosidic triterpene saponins from the roots of Pulsatilla chinensis.

Further phytochemical analysis aimed at the triterpene saponin constituents of the roots of Pulsatilla chinensis has resulted in the isolation of four new bisdesmosidic triterpene saponins whose aglycons are based on the lupane skeleton (1-4), together with three known saponins (5-7). The structures of the new compounds were determined by spectroscopic analysis and acid-catalyzed hydrolysis.

Five new polyoxygenated cholestane bisdesmosides from the bulbs of Galtonia candicans.

Two new cholestane bisdesmosides (1, 2) based upon (22S)-cholest-5-ene-3 beta,16 beta,22-triol with an acetyl group at the sugar moiety and three new ones (3-5) based upon (22S)-cholest-5-ene-1 beta,3 beta,16 beta,22-tetrol, along with a known cholestane glycoside, were isolated from the bulbs of Galtonia candicans. The structures of the new compounds were determined by spectroscopic analysis and chemical transformations.

Cholestane glycosides from the bulbs of Galtonia candicans and their cytotoxicity.

Further search for cytotoxic compounds contained in the bulbs of Galtonia candicans (Liliaceae) led to the isolation of four potent cytotoxic cholestane glycosides (1-4) based upon 3beta,16beta,17alpha-trihydroxycholest-5-en-22-one, three of which (2-4) have not been reported previously. A new cholestane bisdesmoside (5) and a new rearranged cholestane glycoside (6) were also isolated. The structural assignment of the new constituents was carried out by spectroscopic analysis and a few chemical transformations.

Pharmacological studies of geissoschizine methyl ether, isolated from Uncaria sinensis Oliv., in the central nervous system.

The pharmacological properties of geissoschizine methyl ether, isolated from Uncaria sinensis Oliv., were analyzed in vitro and in vivo using mice central serotonin neurons. In the in vitro experiment, geissoschizine methyl ether inhibited [3H]8-hydroxy-2-(di-n-propylamino)tetralin) ([3H]8-OH-DPAT) (K(i)=0.8 microM), [3H]mesulergine (K(i)=0.9 microM) and [3H]ketanserin (K(i)=1.4 microM), but had less affinity toward [3H]prazosin (K(i) > 10 microM) and [3H]spiperone (K(i) >15 microM) binding to mouse brain membranes. The in vivo studies showed that geissoschizine methyl ether dose-dependently reduced 5-hydroxy-L-tryptophan (I-5-HTP) plus clorgyline-induced head twitch response without inhibiting the I-5-HTP plus clorgyline and 8-OH-DPAT-induced head weaving. On the other hand, geissoschizine methyl ether also decreased the rectal temperature of mice (hypothermic response) in a dose-dependent manner. These results suggest that geissoschizine methyl ether possesses mixed 5-HT(1A) receptor agonist/5-HT(2A/2C) receptor antagonist activities and inhibits the head twitch response by blocking the 5-HT(2A) receptors, and possibly, at least in part, by stimulating the 5-HT(1A) receptors in the central nervous system.

Cytotoxic activity of saponins from Camassia leichtlinii against human oral tumor cell lines.

Five steroidal saponins from Camassia leichtlinii showed higher cytotoxicity against human oral squamous cell carcinoma cells HSC-2, as compared to normal human gingival fibroblasts HGF. The tumor specificity of saponins varied considerably from sample to sample, but was generally higher than that of tannins, flavonoids and prenylated compounds such as geranylgeraniol and vitamin K2 (MK-2). Agarose gel electrophoresis showed that the saponins failed to induce internucleosomal DNA fragmentation, but produced large DNA fragments in HSC-2 cells, whereas two saponin samples (compounds 1 and 5) induced internucleosomal DNA fragmentation in human promyelocytic leukemic HL-60 cells. In contrast to epigallocatechin gallate or gallic acid, the cytotoxic activity of saponins was not significantly affected by metals (Co2+, Cu2+, Fe3+) or by antioxidants (sodium ascorbate, N-acetyl-L-cysteine, catalase). Furthermore, the saponins did not produce radicals (detected by ESR spectroscopy) nor oxidation potential (measured by NO monitor). These data suggest that an oxidation-mediated mechanism is not involved in the cytotoxicity induced by the steroidal saponins.

Triterpene glycosides from the roots of Sanguisorba officinalis.

Five triterpene glycosides were isolated from the MeOH extract of Sanguisorba officinalis (Rosaceae) roots, as confirmed by detailed analysis of their 1H, 13C, and two-dimensional NMR data, and by the results of hydrolytic cleavage. Three known triterpenes and six known triterpene glycosides were also isolated and identified. The isolated compounds were evaluated for their cytotoxic activities against HSC-2 cells and HGF.

Steroidal glycosides from the bulbs of Camassia leichtlinii and their cytotoxic activities.

Phytochemical analysis of the bulbs of Camassia leichtlinii (Liliaceae) resulted in the isolation of six new spirostanol saponins, a new furostanol saponin, a cholestane glucoside, and four known steroidal saponins. The structures of the new saponins were determined by detailed analysis of their spectral data, including two-dimensional NMR spectroscopy, and by the results of hydrolytic cleavage. Cytotoxic activities of the isolated compounds against human oral squamous cell carcinoma (HSC-2) cells and normal human gingival fibroblasts (HGF) are also reported.