纤细薯蓣甾体皂甙的分离鉴定

从江西庐山采集的纤细薯蓣根茎中分到两个三糖皂甙(A,B),通过乙酰化、酸水解、红外、质谱和碳谱等鉴定,皂甙 A 是薯蓣皂甙(dioscin),皂甙 B 是纤细皂甙(gracillin),二者得率之比约为3:1。     

Identification and Isolation of Steroidal Saponins of Dioscorea zingiberensis Wright

Two diosgenin trisaccharides (A,B) and two furostanolic tetrasacharides of diosgenin (C, D) had been isolated from Dioscorea zingiberensis wright. By means of acetylation, acid hydrolysis, enzymolysis, IR, MS, 1H-NMR and 13C-NMR, saponin A was proved to be a new compound of structure: diosgenin-3-O- [ β-D-glucopyranosyl (1 → 2) ]-O- [ α-L-rhamnopyra- nosyl (1→3)]-O-β-D-glucopyranoside was provisionally named zingiberenin A; saponin B may be suggested as the steric isomer of gracillin; saponin C was identified as 26-O-β-D- glucopyranoside of A and saponin D as 26-O-β-D-glucopyranoside of B. The last two components named proto-zingiberenin A and proto-zingiberenin B respectively.     

Study on the Structure of a New Triterpene Saponin B from Polygala japoniea Houtt

A new triterpene saponin B has been isolated from the earial parts of Polygala japonica Houtt in folk-lore medicine. Its molecular: C48H78O20, m.p. 199–202℃, [α]D23+30.0 (C, 0.5, CH3OH). Acidic hydrolysis of this saponin gave a sapogenin (2α, 3α, 24-tri-hydroxyolean-12-ene-28-oic acid) and D-glucose. The structure of saponin B was elucidated as 28-O- [β-D-glucopyranosyl (1→2) -β-D-glucopyranosyl (1→2) -β-D-glucopy- ranosyl] 2α, 3α, 24-trihydroxyolean-12-ene-28-oic acid mainly by 13C-NMR, MS and some chemical transfomations.     

Two new triterpene saponins from the anti-inflammatory saponin fraction of Ilex pubescens root

The saponin fraction from the ethanolic extracts of the root of Ilex pubescens Hook. et Arn. (Ilexaceae) was found to exhibit potent anti-inflammatory effects on carrageenan-induced paw edema in rats. Two novel triterpene saponins, pubescenosides C and D (1 and 2, resp.), together with five known saponins were isolated from this saponin fraction. The structures of 1 and 2 were elucidated as (20beta)-3-O-[beta-D-glucopyranosyl-(1-->2)-beta-D-xylopyranosyl]ursa-12,18-dien-28-oic acid 28-O-beta-D-glucopyranosyl ester, and (20beta)-3-O-[alpha-L-rhamnopyranosyl-(1-->2)-beta-D-glucopyranosyl-(1-->2)-beta-D-xylopyranosyl]ursa- 12,18-dien-28-oic acid 28-O-beta-D-glucopyranosyl ester, respectively, on the basis of chemical and spectroscopic data. Five known saponins isolated from the saponin fraction were identified as ilexsaponin B(1), B(2), B(3), A(1), and chikusetsusaponin IV(a).     

Comparative studies of acidic and enzymatic hydrolysis for production of soyasapogenols from soybean saponin

Abstract

Soyasapogenols, aglycones of soyasaponins, can be produced from crude soybean saponin extract by acid or enzymatic hydrolysis. Soyasapogenol B is known to have hepatoprotective, antimutagenic, antivirus, and anti-inflammatory activities. Hydrolysis of soyabean saponin extract for 72 h with 2 M HCl in methanol gave three soyasapogenols, namely: soyasapogenol D, soyasapogenol B₁ and soyasapogenol A. However, the microbial hydrolysis of soybean saponin extract by Aspergillus terreus led to isolation of soyasapogenol B as a major product. A systematic evaluation of the effect of key operational parameters on the microbial transformation was performed. Maximum production of soyasapogenol B (about 152.3 mg/50ml) was observed using 1.5% (w/v) soybean saponin and 1.5% (w/v) glucose, 32°C after 72 h at pH 7 using phosphate buffer. Under these optimal conditions, the cells’ bioconversion efficiency increased from 20.5 to 85.3%. The isolation of soyasapogenols was performed using chromatographic methods and their structures identified on the basis of spectroscopic tools.     

Molluscicidal saponins from Gundelia tournefortii

From the roots of Gundelia tournefortii seven saponins have been isolated mainly by DCCC. The main saponins (A and B) were characterized, mainly by ¹³C and ¹H NMR spectroscopy, as oleanolic acid 3-O-(2-[α-l-arabinopyranosyl(1 → 3) -β-d-gentiotriosyl(1 → 6) -β-d-glucopyranosyl]gb-d-xylopyranoside) (saponin A) and oleanolic acid 3-O-(2-[α-l-arabinopyranosyl] (1 → 3)-β-d-gentiobiosyl (1 → 6)-β-d-glucopyranosyl β-d-xylopyranoside) (saponin B). The other saponins are also derived from oleanolic acid and contain more sugar units. The saponin mixture and the saponins A and B possess strong molluscicidal activity against the schistosomiasis transmitting snail Biomphalaria glabrata.     

Optimization of cell permeabilization for multiparametric flow cytometric analysis with lectin staining

BACKGROUND: This study was undertaken in mice to develop a reproducible procedure of cell permeabilization, allowing intracellular protein staining by immunofluorescence (i.e., Bcl-2) without losing surface labeling especially for lectins (i.e., B220 and peanut agglutinin [PNA]). This article reports results obtained with different permeabilization protocols. METHODS: Lymphoid cells were extracted and prepared from Peyer's patches and spleen. After surface labeling using anti-B220-Cy-chrome and PNA-biotin/streptavidin-phycoerythrin, we comparatively tested three permeabilization protocols: saponin 0.3%, methanol 70%, and the commercial kit Dako Intrastain. Final Bcl-2 staining was performed and cells were analyzed by flow cytometry. RESULTS: With 0.3% saponin as the permeabilization reagent, a significant loss of lectin labeling was observed when comparing mono PNA and triple (i.e. , B220-PNA-Bcl-2) staining (74.8% and 22.5% positive cells, respectively). Quality of PNA staining was conserved with Intrastain when comparing multiparametric versus monoparametric stainings (82. 4% of positive cells versus 78.3%, respectively). Intrastain preserved scatter characteristics (69.9% of total cells in the lymphocyte gate with Intrastain versus 13.7% with saponin 0.3% and 20.9% with methanol 70%). This protocol has been used for a preliminary multiparametric analysis in order to quantify Bcl-2 expression in PNA/B220-positive cells. CONCLUSION: This protocol may be useful to assess simultaneously lectin cell surface labeling and intracellular target staining.     

Fungicidal properties of two saponins from Capsicum frutescens and the relationship of structure and fungicidal activity

Two steroidal saponins have been purified from cayenne pepper (Capsicum frutescens). Both have the same steroidal moiety but differ in the number of glucose moieties: the first saponin has four glucose moieties (molecular mass 1081 Da) and the second contains three glucose moieties (molecular mass 919 Da). Solubility in aqueous solution is less for the saponin containing three glucose moieties than for the one containing four glucose moieties. The larger saponin was slightly fungicidal against the nongerminated and germinating conidia of Aspergillus flavus, A. niger, A. parasiticus, A. fumigatus, Fusarium oxysporum, F. moniliforme, and F. graminearum, whereas, the second saponin (molecular mass 919 Da) was inactive against these fungi. Results indicate that the absence of one glucose molecule affects the fungicidal and aqueous solubility properties of these similar molecules.     

Comparison of saponin composition and content in wild soybean (Glycine soja Sieb. and Zucc.) before and after germination

Eight wild soybean accessions with different saponin phenotypes were used to examine saponin composition and relative saponin quantity in various tissues of mature seeds and two-week-old seedlings by LC–PDA/MS/MS. Saponin composition and content were varied according to tissues and accessions. The average total saponin concentration in 1?g mature dry seeds of wild soybean was 16.08?±?3.13?μmol. In two-week-old seedlings, produced from 1?g mature seeds, it was 27.94?±?6.52?μmol. Group A saponins were highly concentrated in seed hypocotyl (4.04?±?0.71?μmol). High concentration of DDMP saponins (7.37?±?5.22?μmol) and Sg-6 saponins (2.19?±?0.59?μmol) was found in cotyledonary leaf. In seedlings, the amounts of group A and Sg-6 saponins reduced 2.3- and 1.3-folds, respectively, while DDMP?+?B?+?E saponins increased 2.5-fold than those of mature seeds. Our findings show that the group A and Sg-6 saponins in mature seeds were degraded and/or translocated by germination whereas DDMP saponins were newly synthesized.     

Determination of saponins in the kernel cake of Balanites aegyptiaca by HPLC-ESI/MS

The kernel cake produced from Balanites aegyptiaca fruit of Israeli origin was analysed for its saponin constituents using high-performance liquid chromatography-mass spectrometry (HPLC-MS). The HPLC was equipped with a reversed-phase C18 column and a refractive index detector (RID), and elution was isocratic with methanol and water (70:30). The MS system was equipped with electrospray ionisation (ESI). Nine compounds were chromatographically separated, their masses were determined in the negative ion mode and subsequent fragmentation of each component was carried out. From the nine components, six saponins with molecular masses of 1196, 1064, 1210, 1224, 1078 and 1046 Da were identified, with the compound of mass 1210 Da being the main saponin (ca. 36%). Saponins with masses of 1224 and 1046 Da have not been previously reported in B. aegyptiaca. In all saponins, diosgenin was found to be the sole aglycone. This study shows that HPLC-ESI/MS is a quick and reliable technique for characterizing the saponins from kernel cake of B. aegyptiaca.     

异柱五加叶中齐墩果烷型皂苷的研究(英文)

利用柱色谱从异柱五加 (Acanthopanax sieboldianus forma albeofolium Yook) 叶的甲醇提取液中分离出四个齐墩果烷型皂苷类化合物.通过波谱方法(1H NMR、13C NMR、2D NMR和FAB-MS)鉴定它们分别为kalopanax-saponin B (1)、acanthopanax saponin CP3(2)、kalopanax-saponin A (3) 和sieboldianoside A (4).     

Preparative separation of the saponin lancemaside a from Codonopsis lanceolata by centrifugal partition chromatography

Introduction. Lancemaside A is a saponin that inhibits decreases in blood testosterone level and thus prevents or ameliorates symptoms associated with male climacteric disorder. Our initial attempt to preparative isolation of lancemaside A from the saponin fraction of Codonopsis lanceolata roots by a preparative HPLC did not give a clear result. Objective. To develop a simple and efficient method for the preparative isolation of lancemaside A from the hot water extract of C. lanceolata roots using centrifugal partition chromatography (CPC). Methodology. The saponin fraction obtained from the hot water extract of C. lanceolata roots was used as the sample for preparative‐scale separation of lancemasides by CPC using n‐hexane:n‐butanol:methanol:0.1% aqueous formic acid (3:4:1:6, v/v) as the two‐phase solvent system. The upper phase (organic phase) of the two‐phase solvent system was used as the mobile phase, and 0.5 g of saponin fraction was applied for separation by CPC. Each fraction that was separated by CPC was analysed by HPLC, and the fractions containing each of the separated compounds were pooled together, and then were purified by simple preparative HPLC. Results. The demonstrated separation sequence, hot water extraction, DIAION HP‐20 column chromatography, CPC and preparative HPLC, yielded lancemaside A, foetidissimoside A and astersaponin Hb in their pure forms. Conclusion. The simple and efficient method for the preparative isolation of lancemaside A along with two other saponins, foetidissimoside A and astersaponin Hb, from the saponin fraction of C. lanceolata was established using CPC.     

Characterization of quillaja bark extracts and evaluation of their purity using liquid chromatography–high resolution mass spectrometry

In the course of development of semi-preparative liquid chromatographic methods for the isolation of individual quillaja saponins from Quillaja saponaria (L.), some commercially available quillaja bark extracts revealed a distinctive and characteristic pattern of additional peaks in the chromatogram that could not be attributed to saponins commonly present in quillaja. To identify these peaks, analytical procedures based on HPLC coupled with high resolution MS detection were optimized which allowed the identification of the additional saponins Mi saponin A, Mi saponin B, Mi saponin C, madhucoside A and madhucoside B. These compounds are known to be the main saponins of the Indian plant Madhuca longifolia (L.). Tandem MS experiments were performed for the unambiguous assignment of the sapogenin. Madhuca saponins yielded a characteristic fragment of protobassic acid, whereas quillaja saponins showed a fragment of quillaic acid as expected. In addition, samples from madhuca seed kernels were analysed to verify the origin of the characteristic chromatographic peak pattern observed frequently in commercially available quillaja bark extracts.     

Probing cytoskeleton modulation by optical biosensors

This paper reported the use of resonant waveguide grating biosensors for studying the cytoskeleton structure in cells. This was achieved by measuring the changes in mass within the bottom portion of cells upon exposure to saponin in the absence and presence of cytoskeleton modulators. Treatment of Chinese hamster ovary cells with saponin led to a dose-dependent and dynamic mass changes. When a higher concentration of saponin (> 60 microg/ml) was used, a net loss in mass was observed. This is probably resulted from the diffusion of soluble intracellular materials away from the bottom portion of cells after pore formation in the cell plasma membranes by saponin. The pretreatment of cells with actin disruption agents, cytochalasin B and latrunculin A, led to significantly increased loss in cell mass induced by either 75 or 125 microg/ml saponin. These results suggested that optical biosensors provide an attractive means to study the cytoskeleton structure and screen modulators that affect the cytoskeleton structure.     

Studies on the Steroidal Saponins of Dioscorea tokoro Makino Collected from Zhejiang

Six steroidal saponins were isolated from rhizome of Dioscorea tokoro Makino (collected from Zhejiang). By means of acetylation, acid hydrolysis, IR, MS, and 13C-NMR etc., five of them were identical with yononin (A) (yield 0.0069%), dioscin (C) (0.04%), gracillin (D) (0.03%), protodioscin (E) (0.5%), protogracillin (F) (0.2%), respectively. Because of low yield, saponin B was not identified.     

浙江山萆薢甾体皂甙的研究

从山萆薢根茎分到6个甾体皂甙,通过乙酰化、酸水解,红外、质谱和碳谱等鉴定,分别为约诺皂甙(A),薯蓣皂甙(C),纤细皂甙(D),原薯蓣皂甙(E),原纤细皂甙(F);皂甙 B 因得率极低,未能完成鉴定。     

Simultaneous estimation of saponin contents from soybean seed using Fourier transform infrared spectroscopy and high-performance liquid chromatography analysis

The aim of this study is to determine the feasibility of Fourier transform infrared (FT-IR) spectroscopy for simultaneous determination of saponin contents in different soybean cultivars. In cross validation between predicted content of saponin by PLS regression modeling from FT-IR spectra and measured content by HPLC, total saponin contents were predicted with good accuracy (R ² ≥ 0.71). In external validation, saponin group Ab (R ² = 0.88), saponin DDMP-group βg (R ² = 0.85), total saponin group B (R ² = 0.88), and total saponin content (R ² = 0.87) were predicted with good accuracy, while prediction for saponin group Aa (R ² = 0.58), saponin group Bb′ (R ² = 0.58), and total saponin group A (R ² = 0.25) had relatively lower accuracy. Considering these results, we suggest that the PLS prediction system for saponin contents using FT-IR analysis could be applied as a novel screening tool for high yielding lines in soybean breeding.     

Direct Determination of Copper,Lead and Cadmium in Tea Infusions by Flameless Atomic Absorption Spectrometry

Two kinds of “group A saponin,” Aa and Ab, are present as the main constituent in soybean seed. The saponin composition and content in F₁ and F₂ seeds derived from the cross parents of Aa and Ab types were analyzed. The “group A saponin” was of Aa–Ab type in all the F₁ seeds, and the ratio of Aa type : Aa–Ab type : Ab type was 1:2:1 in the F₂ seeds. From these results, it appears that Aa and Ab were controlled by codominant allelic alternatives at a single locus. An investigation of the saponin composition of the seed hypocotyls of 18 wild lines revealed some lines in which “group A saponin” was absent.     

Saponin 1 Induces Apoptosis and Suppresses NF-κB-Mediated Survival Signaling in Glioblastoma Multiforme (GBM)

Saponin 1 is a triterpeniod saponin extracted from Anemone taipaiensis, a traditional Chinese medicine against rheumatism and phlebitis. It has also been shown to exhibit significant anti-tumor activity against human leukemia (HL-60 cells) and human hepatocellular carcinoma (Hep-G2 cells). Herein we investigated the effect of saponin 1 in human glioblastoma multiforme (GBM) U251MG and U87MG cells. Saponin 1 induced significant growth inhibition in both glioblastoma cell lines, with a 50% inhibitory concentration at 24 h of 7.4 µg/ml in U251MG cells and 8.6 µg/ml in U87MG cells, respectively. Nuclear fluorescent staining and electron microscopy showed that saponin 1 caused characteristic apoptotic morphological changes in the GBM cell lines. Saponin 1-induced apoptosis was also verified by DNA ladder electrophoresis and flow cytometry. Additionally, immunocytochemistry and western blotting analyses revealed a time-dependent decrease in the expression and nuclear location of NF-κB following saponin 1 treatment. Western blotting data indicated a significant decreased expression of inhibitors of apoptosis (IAP) family members,(e.g., survivin and XIAP) by saponin 1. Moreover, saponin 1 caused a decrease in the Bcl-2/Bax ratio and initiated apoptosis by activating caspase-9 and caspase-3 in the GBM cell lines. These findings indicate that saponin 1 inhibits cell growth of GBM cells at least partially by inducing apoptosis and inhibiting survival signaling mediated by NF-κB. In addition, in vivo study also demonstrated an obvious inhibition of saponin 1 treatment on the tumor growth of U251MG and U87MG cells-produced xenograft tumors in nude mice. Given the minimal toxicities of saponin 1 in non-neoplastic astrocytes, our results suggest that saponin 1 exhibits significant in vitro and in vivo anti-tumor efficacy and merits further investigation as a potential therapeutic agent for GBM.     

Cloning and characterization of saponin hydrolases from Aspergillus oryzae and Eupenicillium brefeldianum

We purified saponin hydrolases from Aspergillus oryzae PF1224 and Eupenicillium brefeldianum PF1226. It was confirmed that the enzymes from A. oryzae PF1224 (Sda1) and E. brefeldianum PF1226 (Sde1) are glycoproteins with molecular masses of 82 and 90 kDa respectively. The deduced amino acid sequences of each enzyme from the cloned genes (sda1 or sde1) showed approximately 50% homology with that of the saponin hydrolase Sdn1 from Neocosmospora vasinfecta var. vasinfecta PF1225 (DDBJ accession no. AB110615). When sda1 and sde1 were expressed in the host Trichoderma viride under the control of the cellobiohydrolase I gene promoter, recombinant proteins were secreted with molecular masses of 77 and 67 kDa respectively. These recombinant enzymes hydrolyzed soyasaponin I to soyasapogenol B and triose, and its substrate specificities for glycosides were similar to that of Sdn1, but the specific activities of these enzymes were lower than that of Sdn1.