Comprehensive 1H and 13C NMR assignment of 13 protobassic acid saponins

This study aimed to carry out complete ¹H and ¹³C NMR assignment of 13 protobassic acid saponins, including arganins A–C ( 1 – 3 ) and F ( 4 ), butyrosides B–D ( 5 – 7 ), tieghemelin ( 8 ), 3′-O-glucosyl-arganin C ( 9 ), Mi-saponins A–C ( 10 – 12 ), and mimusopsin ( 13 ), recorded in methanol-d₄. This was accomplished by the analysis of high-resolution one-dimensional (1D) NMR (¹H and ¹³C), two-dimensional (2D) NMR (¹H–¹H COSY, HSQC, and HMBC), and selectively excited 1D TOCSY spectra. Before this study, ¹H and ¹³C NMR data of arganins A–C ( 1 – 3 ) and F ( 4 ) were partially assigned. Our effort leads to their complete assignment, especially the glycon residue, and revises some reported data. Some revisions of the ¹H and ¹³C NMR data in the glycon part of butyroside C ( 6 ), tieghemelin ( 8 ), Mi-saponin A ( 10 ), and mimusopsin ( 13 ) were made. Those data of butyrosides B and D ( 5 & 7 ) and Mi-saponin B ( 11 ), which had not been recorded in methanol-d₄, are provided. In addition, the ¹H and ¹³C NMR data of Mi-saponin C ( 12 ) are reported for the first time. These data, being recorded in methanol-d₄, should be more friendly for use as a reference for identifying the related triterpenoid saponins.     

New Triterpenoid Glycosides from the Roots of Camellia oleifera Abel

Five new triterpenoid saponins, oleiferosides I–M ( 1 – 5 , resp.) were isolated from the roots of Camellia oleifera Abel . Their structures were elucidated by a combination of 1D‐ and 2D‐NMR spectroscopy, mass spectrometry, and chemical methods. All the compounds were identified as oleanane‐type saponins with sugar moieties linked to C(3) of the aglycone. In addition, cytotoxic activities of these saponins were evaluated against four human tumor cell lines (A549, B16, BEL‐7402, and MCF‐7) by using the 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide (MTT) in vitro assay. All of the compounds showed significant cytotoxic activities against the tested cell lines.     

Neuroprotective effect of Brahmi,an ayurvedic drug against oxidative stress induced by methyl mercury toxicity in rat brain mitochondrial-enriched fractions

The present study evaluates the neuroprotective effect of Brahmi against methyl mercury (MeHg) toxicity. The results demonstrated that MeHg-decreased mitochondrial viability in MTT assay and IC₅₀ value was found to be 2.5 μg/mL. However, Brahmi at 250 μg/mL concentration effectively prevented mitochondrial damage caused by MeHg in MTT assay. Our results also demonstrated MeHg significantly inhibited catalase enzyme activity, glutathione content and increased the level of thiobarbituric acid reactive substances in mitochondrial-enriched fractions of rat brain. These alterations were prevented by preincubation with Brahmi. In addition, Brahmi reverted glutathione level to normal that was depleted by MeHg, confirming its chelating effect, one of the molecular mechanisms that underlie protection against oxidative damage. Our study also focused on total phenolic and flavonoid contents of Brahmi, and it was found to contain significant amount of phenols and flavonoids. The presence of saponins was detected by HPLC which might be responsible for neuroprotection against MeHg.     

Two New Cytotoxic Spirostane‐Steroidal Saponins from the Roots of Bletilla striata

Two new spirostane‐steroidal saponins, bletilnoside A ( 1 ) and bletilnoside B ( 2 ), together with five known compounds, 3 – 7 , were isolated from the roots of Bletilla striata (Thunb .) Reichb . F. The structures of the new compounds were determined based on their 1D‐ and 2D‐NMR spectral data. The isolated compounds 1 – 7 were tested for cytotoxicity against four human tumor cells (A549, SK‐OV‐3, SK‐MEL‐2, and HCT15) in vitro using a sulforhodamin B bioassay, and compounds 1, 2 , and 5 showed significant cytotoxicities against all tested tumor cell lines with IC₅₀ values ranging from 3.98±0.16 to 12.10±0.40 μM .     

Cytotoxicological aspects of organic arsenic compounds contained in marine products using the mammalian cell culture technique

Arsenobetaine, arsenocholine, trimethylarsine oxide and tetramethylarsonium iodide, which are contained in marine fishery products, were examined for their potencies on cell growth inhibition, chromosomal aberration and sister chromatid exchange (SCE). Arseno- betaine, the major water-soluble organic arsenic compound in marine animals, exhibited very low cytotoxicity towards mammalian cells. This compound showed no cell growth inhibition at a concentration of 10 mg cm⁻³ and the cytotoxicity was lower than 1/14 000th of that of sodium arsenite and 1/1600th of that of sodium arsenate towards BALB/c 3T3 cells. The chromosomal aberrations caused by arsenobetaine at a concentration of 10 mg cm⁻³ consisted mainly of chromatid gaps and chromatid breaks, but in this concentration chromosomal breakage owing to its osmotic pressure is likely to be considerable. No SCE was observed at a concentration of 1 mg cm⁻³. Arsenocholine and trimethylarsine oxide also showed no cell growth inhibited at a concentration of 10 mg cm⁻³. However, tetramethylarsonium iodide inhibition the growth of BALB/c 3T3 at a concentration of 8 mg cm⁻³. These compounds exhibited a low ability to induce chromosomal aberrations at a concentration range of 2–10 mg cm⁻³ and no SCE was observed at a concentration of 1.0 mg cm⁻³. These results suggested that the major and minor organic arsenic compounds contained in marine fishery products are much less cytotoxic inorganic arsenic, methylarsonic acid and dimethylarsinic acid. © 1998 John Wiley & Sons, Ltd.     

Enhanced bacoside production in shoot cultures of Bacopa monnieri under the influence of abiotic elicitors

The effect of different abiotic elicitors [jasmonic acid, copper sulphate (CuSO₄) and salicyclic acid] at varying concentrations on the stimulation of biomass and bacoside production in in vitro Bacopa monnieri shoot culture was studied. A systematic study conducted over a period of 35 days indicated that the maximum bacoside production (6.74 mg g^? 1 dry weight (DW)) was obtained after a lag of 7 days and thereafter, the content decreased gradually to again increase at 28 days (5.91 mg g^? 1 DW). Therefore, elicitation experiments were carried out over a period of 3, 6 and 9 days. The shoot cultures treated with 45 mg L^? 1 of CuSO₄ exhibited the highest bacoside content of 8.73 mg g^? 1 DW (～1.42-fold higher) than in control cultures (6.14 mg g^? 1 DW). This study indicates the effectiveness of abiotic elicitation on bacoside production in in vitro shoot cultures of this medicinally important herb known for its memory-enhancing properties.     

New Triterpenoid Saponins from the Roots of Gypsophila paniculata L.

The seven new triterpenoid saponins 1 – 7 were isolated from the roots of Gypsophila paniculata L. Their structures were established by 1D ‐ and 2D‐NMR techniques, HR‐MS, and acid hydrolysis. The isolated compounds include 3,28‐O‐bidesmosides with or without a 4‐methoxycinnamoyl group (see 1 vs. 2 and 3 ), and 3‐O‐monoglucosides 4 – 7 . All isolated saponins 1 – 7 and their aglycones were evaluated for their α‐glucosidase inhibition activity. Compound 1 showed inhibitory activity against yeast α‐glucosidase with an IC₅₀ value of 100.9±3.3 μM , whereas compounds 2 – 7 were inactive.     

Four New 13,28‐Epoxyoleanane Saponins from Lysimachia lobelioides

Four new oleanane saponins, lobelioidosides A–D ( 1 – 4 , resp.), all endowed with 16‐oxo and a 23‐OH group, as well as with a 13,28‐epoxy bridge as a common moiety, have been isolated from the 75% EtOH extract of the whole plant of Lysimachia lobelioides. Their structures were elucidated on the bases of MS, ¹H‐ and ¹³C‐NMR, HMQC, HMBC, and ¹H,¹H‐COSY data analysis.     

Triterpene Saponins from Entada phaseoloides

Four oleanane‐type triterpene saponins, phaseoloideside A–D ( 1 – 4 ), were isolated from the seed kernels of Entada phaseoloides, along with rheediinoside B. The structures of the four new compounds were established by 2D‐NMR spectroscopic methods, HR‐ESI‐MS analysis, and chemical degradation. Phaseoloideside D ( 4 ) showed cytotoxic activity against the Eca‐109 cell line with an IC₅₀ value of 28.0 μM .     

New Dammarane‐Type Saponins from the Rhizomes of Panax japonicus

Phytochemical investigation of the rhizomes of Panax japonicus C. A. Meyer (Araliaceae) resulted in the isolation of two new dammarane‐type triterpenoid saponins, yesanchinoside R₁ ( 1 ) and yesanchinoside R₂ ( 2 ), together with one new natural product, 6′′′‐O‐acetylginsenoside Re ( 3 ). In addition, 25 known compounds, including 23 triterpenoid saponins, 4 – 26 , β‐sitosterol 3‐O‐β‐D ‐glucopyranoside ( 27 ), and ecdysterone ( 28 ), were also identified. The known saponins 12, 15 , and 18 – 22 were reported for the first time from the title plant. Their structures were elucidated on the basis of detailed spectroscopic analyses, including 1D‐ and 2D‐NMR techniques, as well as acidic hydrolysis.     

Genotoxic effects of cadmium chloride on human amniotic fluid cells cultured in vitro

In this study we report the results of cytogenetic tests, namely a search for chromosome aberrations (CA) and sister chromatid exchanges (SCEs), performed on human amniotic fluid cells cultured and treated with Cadmium chloride. The cells from primary cultures were exposed to CdCl2 at 1 microM and 10 microM for 24 h. At the higher dose, no metaphases were scored and at the lower dose (1 microM) no effects were evident on cell proliferation, and no chromosome aberrations were found. In the subsequent experiments we used cells from subcultures exposed to 1 microM and 5 microM CdCl2. At the 5 microM dose was evident the induction of chromatid breaks, while the frequency of sister chromatid exchanges shows a small increase, not statistically significant at the dose of 1 microM. In this study we positively demonstrated that amniotic fluid cells grown in vitro are reliable for testing various mutagenic or teratogenic substances. With regard to cadmium treatment results, it is evident a clastogenic effect of cadmium chloride but not a significant induction of SCEs.     

Three Novel Triterpenoids from the Aerial Part of Helwingia chinensis

Three novel triterpenoids named chinenols A – C ( 1 – 3 ), together with two known compounds, glutin‐5‐en‐3β‐ol ( 4 ) and daucosterol ( 5 ), were isolated from the aerial part of Helwingia chinensis Batal . The structures of 1 – 3 were determined on the basis of their HR‐EI‐MS, IR, ¹H‐ and ¹³C‐NMR (DEPT), and 2D (HMQC, HMBC, NOESY) data. Compounds 1 – 3 and 5 showed inhibition activity in an antibacterial assay.     

A sensitive HPLC–MS/MS method for the simultaneous determination of anemoside B4, anemoside A3 and 23‐hydroxybetulinic acid: Application to the pharmacokinetics and liver distribution of Pulsatilla chinensis saponins

Pulsatilla chinensis saponins, the major active components in the herb, have drawn great attention as potential hepatitis B virus infection and hepatoma treatments. Here, a sensitive and accurate HPLC–MS/MS method was established for simultaneous determination of three saponins – anemoside B4, anemoside A3 and 23‐hydroxybetulinic acid – in rat plasma and liver, and fully validated. The method was successfully applied to a pharmacokinetics and liver distribution study of P. chinensis saponins. Consequently, 23‐hydroxybetulinic acid, with an extremely low content in the P. chinensis saponins, exhibited the highest exposure in the liver and in sites before and after hepatic disposition, namely, in the portal vein plasma and systemic plasma, followed by anemoside B4, which showed the highest content in the herb, whereas anemoside A3 displayed quite limited exposure. The hepatic first‐pass effects were 71% for 23‐hydroxybetulinic acid, 27% for anemoside B4 and 37% for anemoside A3, corresponding to their different extents of liver distribution. To our knowledge, this is the first investigation on the liver first‐pass effect and distribution of P. chinensis saponins to date. These results also provide valuable information for the understanding of the pharmacological effect of P. chinensis saponins on liver diseases.     

Six New Steroidal Saponins from Helleborus thibetanus

Six steroidal saponins, including five spirostanol glycosides, 1 – 5 , and one furostanol glycoside 1‐sulfonate, 6 , previously unknown in nature, together with three known compounds, 7 – 9 , were isolated from dried roots and rhizomes of Helleborus thibetanus. Their structures were elucidated by extensive 1D‐ and 2D‐NMR experiments, along with IR and HR‐ESI‐MS data, as well as the results of acid hydrolysis. Compounds 1 – 5 possessed a C(25)?C(27) bond and were glycosylated at HO? C(1), which was unusual in steroidal saponins.     

Contortisiliosides A–G: Isolation of Seven New Triterpene Bisdesmosides from Enterolobium contortisiliquum and Their Cytotoxic Activity

Seven new bisdesmosidic triterpene saponins, with up to eight monosaccharides, which were given the trivial names contortisiliosides A–G ( 1 – 7 ), were isolated from Enterolobium contortisiliquum. The structures of the new saponins were determined on the basis of extensive spectroscopic and chromatographic analyses of both intact and acid‐hydrolyzed compounds. The isolated saponins were evaluated for their cytotoxic activities against BAC1.2F5 mouse macrophages, EL‐4 mouse lymphoma cells, and L‐929 mouse fibroblasts. Whereas contortisiliosides A ( 1 ) and C ( 3 ) were moderately cytotoxic to both BAC1.2F5 macrophages and EL‐4 cells, and contortisiliosides D–G ( 4 – 7 ) did not show any apparent cytotoxic activities against the three cell lines, contortisilioside B ( 2 ) exhibited selective cytotoxic activity against BAC1.2F5 mouse macrophages, with an IC₅₀ value of 3.4 μM . The macrophage death caused by 2 was shown to be neither necrotic nor apoptosis‐inducing based on the unique morphological change of the killed cells, whose cytosols were transformed into large vacuoles, and according to the TUNEL assay.     

Six New Triterpene Saponins with a 21,23‐Lactone Skeleton from Gynostemma pentaphyllum

The six new triterpene saponins 1 – 6 with a 21,23‐lactone skeleton were isolated from the MeOH extract of the aerial parts of Gynostemma pentaphyllum. Their structures were elucidated by 1D‐ and 2D‐NMR‐spectra interpretation as well as by chemical degradation.     

Characterization of three saponins from a fraction using 1D DOSY as a solvent signal suppression tool. Agabrittonosides E–F. Furostane Saponins from Agave brittoniana Trel. spp. Brachypus

A careful NMR analysis, especially by 1D TOCSY and 1D ROESY, of a refined saponin fraction allowed us to determine the structure of three saponins from a polar extract of Agave brittoniana Trel. spp. Brachypus leaves. The use of 1D DOSY for the suppression of the solvent signal was useful to obtain the chemical shifts of anomeric signals. A full assignment of the ¹H and ¹³C spectral data for the new saponins, agabrittonosides E–F (1–2) and the well‐known Karatavioside C (3) and their methoxyl derivatives, is reported. The structures were established using a combination of 1D and 2D (¹H, ¹H‐COSY, TOCSY, ROESY, g‐HSQC, g‐HMBC and g‐HSQC‐TOCSY) NMR techniques and ESI–MS. In addition, the methoxylation of these furostane saponins in the presence of MeOH was studied. Copyright © 2010 John Wiley & Sons, Ltd.     

Steroidal Saponins from the Rhizomes of Smilacina henryi

Five steroidal saponins, namely henryiosides A–E ( 1 – 5 ), were isolated from the EtOH extract of the rhizomes of Smilacina henryi. Their structures were elucidated by the extensive use of 1D‐ and 2D‐NMR experiments, along with HR‐MALDI‐MS analysis and the results of acid hydrolysis. The aglycones of henryiosides A–E possess a C(7)?C(8) or C(9)?C(11) bond and were not previously found in saponins.     

A Pair of New Nortriterpene Saponin Epimers from the Roots of Gypsophila oldhamiana

A pair of new oleanane‐type nortriterpene saponin epimers, neogypsoside A ( 1 ) and B ( 2) (Fig. 1) with neogypsogenin A ( 3 ) and neogypsogenin B ( 4 ) as the two new aglycons, as well as the two known triterpene saponins 5 and 6 (Fig. 1), were isolated from the roots of Gypsophila oldhamiana. Their structures were determined by analysis of their NMR data. A possible biogenetic pathway to the nortriterpene saponins 1 and 2 is proposed (Scheme 2).     

Total assignment of 1H and 13C NMR spectra of three triterpene saponins from roots of Silene vulgaris (Moench) Garcke

The assignments of ¹H and ¹³C NMR spectra for three new triterpene saponins from Silene vulgaris (gypsogenin 3‐O‐glucuronide, quillaic acid 3‐O‐glucuronide, and gypsogenin 3‐O‐glycoside) are reported. In addition to 1D NMR methods, 2D NMR techniques (COSY, HSQC, HMBC, and HSQC‐TOCSY) were used for the assignments. Copyright © 2002 John Wiley & Sons, Ltd.