High-performance liquid chromatographic analysis of saponin compounds in Bupleurum falcatum.
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A mixture of saponin compounds (saikosaponin c, a, and d) in the 70% ethanol extract of a powdered sample of Bupleuri radix are analyzed by an Inertsil ODS-3 C(18) column at a flow rate of 1.0 mL/min and detection wavelength of 203 nm. Well resolved chromatograms of saikosaponin c, a, and d are obtained with a gradient elution of acetonitrile-water from 40:60 (v/v) to 50:50 (v/v). The total time required for a single analysis is approximately 20 min. Calibration curves for saikosaponin c, a, and d are linear up to 2.5 mg/mL. The coefficient of variability values for saikosaponins in the extract are below 4%, and the recoveries for saikosaponin c, a, and d are 95.2 +/- 1.1, 96.5 +/- 0.9, and 96.2 +/- 1.0%, respectively. The changes in saikosaponin contents for a two-year growth of Bupleurum falcatum are measured by the established high-performance liquid chromatography method. (+info)
Effect of saikosaponin, a triterpene saponin, on apoptosis in lymphocytes: association with c-myc, p53, and bcl-2 mRNA.
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1. The mechanisms involved in the apoptotic effect of saikosaponin-d, a triterpene saponin from Bupleurum falcatum L., were studied in human CEM lymphocytes and compared with those of dexamethasone (3 x 10(-7) M). 2. Saikosaponin-d (10(-8) to 10(-5) M) inhibited the serum-stimulated [(3)H]-thymidine incorporation in a concentration-dependent manner. Dexamethasone also inhibited serum-stimulated [(3)H]-thymidine incorporation. 3. Cell viability was unaffected by saikosaponin-d until 10(-5) - 10(-4) M. Dexamethasone significantly reduced the number of viable cells. 4. Following saikosaponin-d (10(-5) - 10(-4) M) treatment, flow cytometry analysis of propidium iodide-stained cells showed a significant increase in the percentage of cells in the apoptotic region. Dexamethasone also significantly increased the percentage of apoptotic cells. The supravital exposure to propidium iodide and annexin V labelling demonstrated that saikosaponin-d (10(-5) - 10(-4) M) induced apoptosis as well as necrosis. 5. The apoptotic effect of saikosaponin-d (3 x 10(-6) - 10(-4) M) was also demonstrated by TUNEL analysis and DNA laddering. The percentage of apoptotic cells induced by saikosaponin-d (3 x 10(-6) - 10(-5) M) was unaffected by the presence of Z-VAD-FMK, indicating that saikosaponin-d-induced apoptosis may not be mediated by caspase activity. However, the percentage of apoptotic cells induced by dexamethasone was significantly reduced by the presence of Z-VAD-FMK. 6. Levels of c-myc, p53, and bcl-2 mRNA were analysed by the reverse transcription-polymerase chain reaction. Levels of c-myc and p53 mRNA were significantly increased, while the level of bcl-2 mRNA was decreased, by saikosaponin-d (10(-5) M) treatment. Dexamethasone did not significantly change the expression of these genes. 7. It is suggested that the apoptotic effect of saikosaponin-d may be partly mediated by increases in c-myc and p53 mRNA levels accompanied by a decrease in bcl-2 mRNA level. (+info)
How to distinguish garlic from the other Allium vegetables.
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The establishment of international monographs for herbs is in progress. Here, we propose both a marker compound and a method for its analysis for the identification of garlic bulbs and their products. The constituents in 26 kinds of fresh edible parts of Allium vegetables and three types of garlic preparations were analyzed. Sulfur compounds are the most characteristic constituents in garlic, but manufacturing processes of garlic products dramatically affect these constituents. Thus, no sulfur compound could be specified as a universal marker of identification applicable for any type of garlic. On the other hand, garlic contains other characteristic compounds, namely, saponins. After analyzing Allium vegetables and garlic preparations, we concluded that sapogenins, especially beta-chlorogenin, may be a viable candidate for identifying and distinguishing garlic from other Allium vegetables. (+info)
Purification of saponin compounds in Bupleurum falcatum by solvent partitioning and preparative LC.
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Saponin compounds (saikosaponin c, a, and d) in Bupleurum falcatum were partially purified by solvent partitioning of the herbal extract using diethyl ether, distilled water, n-butanol, and acetone. After separation of the saponins by preparative LC, the purity of each saikosaponin was more than 94%. The identities of purified individual saikosaponins were confirmed by TLC, analytical LC, and fast-atom bombardment mass spectrometry. (+info)
Triterpenoid sapogenin from Anthyllis vulneraria L.
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Triterpene pentacyclic sapogenin, 3beta,22beta,24-trihydroxy-olean-12-ene, soyasapogenol B, was isolated for the first time from the Anthyllis vulneraria L.. Identification and structure assignment was performed by 1H and 13C NMR spectroscopy. (+info)
Chemistry and cancer preventing activities of ginseng saponins and some related triterpenoid compounds.
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More than 25 dammarane-type tetracyclic triterpenoid saponins have been isolated from ginseng, the root and rhizome of Panax ginseng C.A. Meyer (Araliaceae). The genuine sapogenins of those saponins, 20(S)-protopanaxa-diol and -triol, were identified as 20(S) 12beta-hydroxy-and 20(S) 6alpha,12beta-dihydroxy-dammarenediol-II, respectively. There are two types of preparations from ginseng: white ginseng prepared by drying after peelling off and red ginseng prepared by steaming and drying. Some partly deglycosylated saponins such as ginsenoside Rh-1, Rh-2, and Rg-3 are obtained from red ginseng as artifacts produced during steaming. Several workers studied the metabolic transformation by human intestinal bacteria after oral administration of ginsenoside Rb-1 and Rb-2 and found that the stepwise deglyco-sylation yielded compound K and finally 20(S)-protopanaxadiol. Ginsenoside Rg-1 was converted into 20(S)-protopanaxatriol via ginsenoside Rh-1. Yun et al. in Korea conducted the epidemiological case-control studies of ginseng and suggested its cancer preventing activities. Kitagawa et al. demonstrated in vitro that ginsenosides, especially 20(R)-ginsenoside Rg-3, specifically inhibited cancer cell invasion and metastasis. Azuma et al. found that ginsenoside Rb-2 inhibited tumor angiogenesis, and Kikuchi et al. reported that ginsenoside Rh-2 inhibited the human ovarian cancer growth in nude mice. Recently, ginsenoside Rg-3 was produced as an anti-angiogenic anti-cancer drug in China. The aforementioned reports suggest that less glycosylated protopanaxadiol derivatives are effective in cancer prevention. Apart from Ginseng tetracyclic triterpenoid saponins, some oleanane-type pentacyclic triterpenoid compounds showed the anti-carcinogenic activity in the two-stage anti-cancer-promotion experiments in vitro and in vivo. (+info)
Characteristics and mechanism of enzyme secretion and increase in [Ca2+]i in Saikosaponin(I) stimulated rat pancreatic acinar cells.
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AIM: This investigation was to reveal the characteristics and mechanism of enzyme secretion and increase in [Ca2+]i stimulated by saikosaponin(I) (SA(I)) in rat pancreatic acini. METHODS: Pancreatic acini were prepared from male Wistar rats. Isolated acinar cells were suspended in Eagle's MEM solution. After adding drugs, the incubation was performed at 37 degrees for a set period of time. Amylase of supernatant was assayed using starch-iodide reaction. Isolated acinar single cell was incubated with Fura-2/AM at 37 degrees, then cells were washed and resuspended in fresh solution and attached to the chamber. Cytoplasm [Ca2+]i of a single cell was expressed by fluorescence ratio F340/F380 recorded in a Nikon PI Ca2+ measurement system. RESULTS: Rate course of amylase secretion stimulated by SA(I) in rat pancreatic acini appeared in bell-like shape. The peak amplitude increased depended on SA(I) concentration. The maximum rate responded to 1 x 10(-5)mol/L SA(I) was 13.1-fold of basal and the rate decreased to basal level at 30 min. CCK-8 receptor antagonist Bt(2)-cGMP markedly inhibited amylase secretion stimulated by SA(I) and the dose-effect relationship was similar to that by CCK-8. [Ca2+]i in a single acinar cell rose to the peak at 5 min after adding 5 x 10(-6)mol/L SA(I) and was 5.1-fold of basal level. In addition, there was a secondary increase after the initial peak. GDP could inhibit both the rate of amylase secretion and rising of [Ca2+]i stimulated by SA(I) in a single pancreatic acinar cell. CONCLUSION: SA(I) is highly efficient in promoting the secretion of enzymes synthesized in rat pancreatic acini and raising intracellular [Ca2+]i. Signaling transduction pathway of SA(I) involves activating special membrane receptor and increase in cytoplasm [Ca2+]i sequentially. (+info)
Prevention of growth and metastasis of murine melanoma through enhanced natural-killer cytotoxicity by fatty acid-conjugate of protopanaxatriol.
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Ginsenosides, the glycosides of Panax ginseng, are metabolized (deglycosylated) by intestinal bacteria after oral administration. 20(S)-Protopanaxatriol (M4) is the main bacterial metabolite of protopanaxatriol-type ginsenosides and mediates their antitumor effects. To clarify the mechanism of the M4-mediated antitumor effect, the antitumor activity and metabolism of M4 was examined, using the C57BL/6 mice implanted with B16-BL6 melanoma. The chronic oral administration of M4 inhibited the growth of B16-BL6 melanoma at the implanted site. Analyses using TLC, HPLC, MS and NMR suggest that orally administered M4 was absorbed from the small intestine into the mesenteric lymphatics followed by the rapid esterification of M4 with fatty acids and its accumulation in the tissues including the liver and lung. The administration of M4 prior to the intravenous injection of B16-BL6 cells abrogated the enhanced lung metastasis in the mice pretreated with 2-chloroadenosine more effectively than in those pretreated with anti-asialo GM1. The esterified M4 (EM4) did not directly affect tumor growth in vitro, whereas it stimulated splenic NK cells to become cytotoxic to tumor cells. These results indicate that the antitumor activity of M4 is based on the NK cell-mediated tumor lysis enhanced by EM4. (+info)