Isolation and Identification of Steroidal Saponins from Dioscorea panthaica Prain et Burkill

Two steroidal saponins have been isolated from the rhizomes of Dioscorea panthaica Prain et Burkill. collected from Sichuan province. They were identified as 3-0-{α-L- rhamnopyranosyl (1→4)- [α-L-rhamnopyranosyl (1→2) ]-β-D-glucopyranosyl } -diosgenin (dioscin) and 3-O-{β-D-glucopyranosyl (1→3)-[α-L-rhamnopyranosyl (1→2)]-β-D-glucopyranosyl}-diosgenin (gracillin) on the basis of mp., m. mp., TLC. acetylation, acid hydrolysis, IR, MS, and 13C NMR.     

Synthesis of monomethylated dioscin derivatives and their antitumor activities

All possible eight monomethylated dioscin derivatives (1-8) were synthesized. Their inhibitory activities against P388 and A-549 cells were determined, and the results indicate that six of the eight hydroxyls of dioscin are the 'key polar groupings' for tumor inhibitory activities.     

Liquid chromatography tandem mass spectrometry in study of the pharmacokinetics of six steroidal saponins in rats

A rapid, simple and sensitive high-performance liquid chromatography tandem mass spectrometry method was developed and validated for simultaneous determination of six main steroidal saponins in Paris polyphylla in rat plasma. Ginsenoside Rg3 was selected as the internal standard (IS). Plasma samples were pretreated with protein precipitation, and the separation was achieved on a reverse phase Agilent poroshell120 EC-C18 column using a gradient mobile phase system of acetonitrile–water containing 0.1% formic acid. The triple quadruple mass spectrometer was set in negative electrospray ionization mode and multiple reaction monitoring (MRM) was used for six steroidal saponins quantification. The precursors to produce ion transitions monitored for polyphyllin I, polyphyllin II, polyphyllin VI, polyphyllin VII, dioscin, gracillin and IS were m/z 899.5 > 853.4, 1059.5 > 1013.5, 783.4 > 737.4, 1075.5 > 1029.5, 913.5 > 867.4, 929.5 > 883.4 and 819.5 > 783.4, respectively. The intra- and inter-day precisions (RSD%) were less than 13% and the average extraction recoveries ranged from 85% to 97.0% for each analyte. Six steroidal saponins were proved to be stable during sample storage, preparation and analytical procedures. The established method was employed for simultaneous quantification and successfully used for the first time for the pharmacokinetics evaluation of the six main compounds after intragastric administration of P. polyphylla extract in Sprague–Dawley rats.     

Proteomic approach to study the cytotoxicity of dioscin (saponin)

Dioscin, extracted from the root of Polygonatum zanlanscianense pamp, exhibits cytotoxicity towards human myeloblast leukemia HL-60 cells. Proteomic analysis revealed that the expression of mitochondrial associated proteins was substantially altered in HL-60 cells corresponding to the dioscin treatment, suggesting that mitochondria are the major cellular target of dioscin. Mitochondrial functional studies validated that mitochondrial apoptotic pathway was initiated by dioscin treatment. Changes in proteome other than mitochondrial related proteins implicate that other mechanisms were also involved in dioscin-induced apoptosis in HL-60 cells, including the activity impairment in protein synthesis, alterations of phosphatases in cell signaling, and deregulation of oxidative stress and cell proliferation. Current study of protein alterations in dioscin-treated HL-60 cells suggested that dioscin exerts cytotoxicity through multiple apoptosis-inducing pathways.     

Synthesis, cytotoxicity, and hemolytic activity of 6'-O-substituted dioscin derivatives

Dioscin derivatives (1-12) with a variety of substitutions at the 6'-OH of the chacotriosyl residue and the 3',6'-anhydrosaponin derivatives (26, 30, and 32) were synthesized. All these derivatives showed much lower cytotoxicity than that of the parent dioscin, while their hemolytic activities were partially retained depending on the various 6'-O-substitutions.     

Synthesis and pharmacokinetics of strontium fructose 1,6-diphosphate (Sr-FDP) as a potential anti-osteoporosis agent in intact and ovariectomized rats

A novel strontium compound has been synthesized by the reaction of fructose-1,6-diphosphate with strontium (Sr-FDP). The compound was characterized and confirmed with elemental analyses and spectroscopic (IR, NMR) methods. The pharmacokinetic profiles of Sr-FDP were investigated in Sprague-Dawley rats following oral administration at a dose of 110, 220, and 440 mg/kg respectively. Pharmacokinetic differences were also compared in intact rats and ovariectomized rats with and without estrogen supplement. Strontium concentrations in plasma, urine, tissue and feces were determined by graphite furnace atomic absorption spectroscopy (GFAAS). The results showed that Sr-FDP was absorbed rapidly with T_max < 1 h in all the groups with AUC_0-∞ proportional to the oral dose. The pharmacokinetic profiles were characterized by long half-life, a large apparent volume of distribution. The highest Sr concentration was observed in the bone at 6 h, and the level of Sr decreased close to the baseline in heart, liver, spleen, lung, intestine, brain and kidney after 12 h. The cumulative amounts of Sr over 96 h were found to be ~ 3% in urine, but ~ 70% in feces suggesting that the parent drug was mainly excreted from the intestine. The C_max and AUC_0-∞ of Sr-FDP in ovariectomized rats were significantly decreased compared to those in intact rats, and this trend was ameliorated by using 17-beta-estradiol (E₂) treatment in the ovariectomized rats.     

The synthesis of gracillin and dioscin: two typical representatives of spirostanol glycosides

Two representative spirostanol saponins that have the typical structure for the sugar moiety, diosgenyl alpha-L-rhamnopyranosyl-(1-->2)-[beta-D-glucopyranosyl-(1-->3)]-beta-D-glucopyranoside (gracillin) and diosgenyl alpha-L-rhamnopyranosyl-(1-->2)-[alpha-L-rhamnopyranosyl-(1-->4)]-beta-D-glucopyranoside (dioscin), were easily synthesized by a general approach. A procedure using guanidine for the selective deblocking of acetyl while retaining benzoyl protecting groups is described.     

iTRAQ‐based proteomic analysis of dioscin on human HCT‐116 colon cancer cells

Dioscin shows various pharmacological effects. However, its activity on colorectal cancer is still unknown. The present work showed that dioscin significantly inhibited cell proliferation on human HCT‐116 colon cancer cells, and affected Ca²⁺ release and ROS generation. The content of nitric oxide (NO) and its producer inducible NO synthase (iNOS) associated with DNA damage and aberrant cell signaling were assayed using the kits. DNA damage and cell apoptosis caused by dioscin were also analyzed through single‐cell gel electrophoresis and in situ terminal deoxynucleotidyl transferase dUTP nick‐end labeling assays. The results showed that dioscin increased the levels of NO and inducible NO synthase. The comet length in dioscin‐treated groups was much longer than that of the control group, and the number of terminal deoxynucleotidyl transferase dUTP nick‐end labeling positive cells (apoptotic cells) was significantly increased by the compound (p < 0.01). Furthermore, dioscin caused mitochondrial damage and G2/M cell cycle arrest through transmission electron microscopy and flow cytometry analysis, respectively. To study the cytotoxic mechanism of dioscin, an iTRAQ‐based proteomics approach was used. There were 288 significantly different proteins expressed in response to dioscin, which were connected with each other and were involved in different Kyoto Encyclopedia of Genes and Genomes pathways. Then, some differentially expressed proteins involved in oxidative phosphorylation, Wnt, p53, and calcium signaling pathways were validated by Western blotting and quantitative real‐time PCR assays. Our work elucidates the molecular mechanism of dioscin‐induced cytotoxicity in colon cancer cells, and the identified targets may be useful for treatment of colorectal cancer in future.     

中药薯蓣皂苷对NLRP3炎性体的作用

目的 检测中药薯蓣皂苷是否可以抑制粪肠球菌脂磷壁酸（LTA）所诱导的NLRP3炎性体的活化。方法 选用中药薯蓣皂苷作为实验药物,作用于小鼠巨噬细胞RAW264.7,NLRP3炎性体相关因子mRNA的表达用Real-time qPCR方法检测。通过免疫荧光染色检测薯蓣皂苷对NF-κB的表达情况,流式细胞仪检测其对ROS的表达情况。结果 Real-time qPCR试验显示,中药薯蓣皂苷在LTA存在下,可以明显降低NLRP3、Caspase-1及IL-1β的mRNA表达。免疫荧光染色及流式细胞仪的检测证实,其对NLRP3的抑制主要是通过抑制NF-κB的活化及ROS的释放而实现。结论 薯蓣皂苷可有效抑制NLRP3炎性体的表达,其机制是通过抑制NF-κB信号通路及ROS的释放而实现。薯蓣皂苷可以作为临床难治性牙髓根尖周病治疗的候选药物。     

New Steroidal Saponin of Stem and Leaf of Fritillaria ussuriensis Maxim

A new steroidal saponin, pingpeisaponin, was isolated from stem and leaf of Fritillaria ussuriensis Maxim by column chromatographic technique. On the basis of the IR, MS, 1H NMR and 13C NMR spectra of pingpei saponin, the structure has been established as 24α-hydroxyl diosgenin-3-O-α-L-rhomno-pyranosyl-(l-2)-β-D-glucopyranoside.     

Histamine pharmacokinetics in tumor and host tissues after bolus-dose administration in the rat

In this study, we estimated interstitial histamine concentrations in normal and malignant tissues after a single intravenous (i.v.) injection of 0.5 mg/kg histamine dihydrochloride in the rat. The microdialysis technique was used to collect interstitial fluid from subcutis, liver and a NGW adenocarcinoma. Histamine was absorbed with equal efficiency to all tissues (t 1/2 AB 3.9-7.7 minutes) but maximum concentration (Cmax; nmol/l) of histamine was higher in liver (2,388 +/- 357) than in subcutis (951 +/- 125) (p < 0.01) and subcutaneous tumor (523 +/- 140) (p = 0.01) and, moreover, Cmax in liver tumor (1,752 +/- 326) was higher than in subcutaneous tumor (p = 0.01). The tl/2 elimination was significantly longer in subcutis and subcutaneous tumor than in liver and liver tumor. Area under the curve (AUC; mmol-min/l) for histamine was significantly lower in subcutaneous tumor (9.8 +/- 2.3) than in liver (17.6 +/- 1.9) (p = 0.03) and liver tumor (15.8 +/- 1.8) (p = 0.03). Local tissue blood flow as assessed by the 14C-ethanol method was not significantly altered by the histamine administration. In conclusion, after an i.v. injection of histamine dihydrochloride a higher maximum concentration and AUC of histamine was reached in liver and liver tumor than in subcutaneous tissues.     

New cytotoxic dihydrochalcone and steroidal saponins from the aerial parts of Sansevieria cylindrica Bojer ex Hook

A new dihydrochalcone, 2‘,4‘-dihydroxy-3‘-methoxy-3,4-methylenedioxy-8-hydroxymethylene dihydrochalcone 1 and two new steroidal saponins, (25S)-ruscogenin-1-O-α-l-rhamnopyranosyl-(1 → 2)-β-d-glucopyranoside 2, (25S)-ruscogenin-3-O-α-l-rhamnopyranosyl-(1 → 4)-β-d-glucopyranoside 3, together with three known steroidal saponins (25S)-ruscogenin-3-O-β-d-glucopyranoside 4, (25S)-ruscogenin-1-O-α-l-rhamnopyranosyl-(1 → 2)-[β-d-xylopyranosyl-(1 → 3)]-α-l-arabinopyranoside 5 and (25R)-26-O-β-d-glucopyranosyl-furost-5-ene-1β,3β,22α,26-tetrol-1-O-α-L-rhamnopyranosyl-(1 → 2)-[β-d-xylopyranosyl-(1 → 3)]-α-l-arabinopyranoside 6 were isolated from the aerial parts of Sansevieria cylindrica. The structures of the new compounds were established by UV, IR, EI-MS, HR-ESI–MS as well as 1D (¹H,¹³C and DEPT-135) and 2D (HSQC, HMBC and TOCSY) NMR spectral analysis. The isolated compounds 1-6 were assayed for in vitro cytotoxicities against the three human tumor cell lines HT116, MCF7 and HepG2. Compound 1 showed a moderate cytotoxicity against MCF7. Compounds 2, 3 and 6 exhibited moderate cytotoxicities against the three used cell lines and compound 5 showed marked cytotoxicities against all used cell lines.     

Moments of physiological transit time distributions and the time course of drug disposition in the body

Characterizing the tissue distribution kinetics of drugs by physiological and physico-chemical parameters and using a circulatory model the time course of blood concentration after intravenous injection is predicted for linear pharmacokinetic systems. The interrelationships between the first three (zero to second) moments of the distribution functions of organ transfer times, circulation times and residence times of drug molecules in the body are described. Utilizing literature data the model is applied to the analysis of lidocain kinetics in humans.     

Steroidal saponins and homoisoflavanone from the aerial parts of Sansevieria cylindrica Bojer ex Hook.

Phytochemical study on the methanolic extract of Sansevieria cylindrica aerial parts lead to the isolation, characterization and structure elucidation of a new steroidal saponin, 1β-hydroxy-kryptogenin-1-O-α-l-rhamnopyranosyl-(1 → 2)-α-l-arabinopyranoside (1), a new homoisoflavanone, (3S)-3,7-dihydroxy-8-methoxy-3-(3′,4′-methylenedioxybenzyl) chroman-4-one (2) and the known saponin alliospiroside A (3). To the best of our knowledge, the genin 1β-hydroxy-kryptogenin is reported here for the first time. The structures of the new compounds were determined by UV, IR, EIMS, HRESIMS together with 1D (¹H and ¹³C) and 2D (HSQC and HMBC) NMR spectral analysis. The isolated compounds 1–3 were tested for their radical scavenging activity (DPPH). Compound 2 exhibited activity compared to that of ascorbic acid as a standard. The cytotoxicity of the isolated compounds and the standard doxorubicin was tested against the three human tumor cell lines HT116, MCF-7 and PC-3. The results showed that the isolated compounds were inactive.     

Structural characterization of a new steroidal saponin from Agave angustifolia var. Marginata and a preliminary investigation of its in vivo antiulcerogenic activity and in vitro membrane permeability property

A new furostane steroidal saponin was isolated from the leaves of Agave angustifolia var. marginata. On the basis of chemical conversions and spectroscopic analyses, its structure was established as 3-[O-β-d-glucopyranosyl-(1→3)-O-β-d-glucopyranosyl-(1→3)-O]-[O-6-deoxy-α-l-mannopyranosyl-(1→4)-β-d-xylopyranosyl-(1→2)-O-β-d-glucopyranosyl-(1→4)-β-d-galactopyranosyl)oxy]-(3β,5α,22α,25R)-26-(β-d-glucopyranosyloxy)-22-methoxy-furostane (1). Results of preliminary biological investigations indicated that compound 1 showed significant protective effects against induced gastric ulcers using in vivo experimental models and demonstrated negligible toxicity on membrane integrity in the in vitro assays.     

Ultrastructural changes and diosgenin content in cell suspensions of <Emphasis Type="Italic">Trigonella foenum-graecum</Emphasis> L. by ethylene treatment

This paper studies the effect of different concentrations of ethephon (an ethylene releasing compound) on the level of the steroidal sapogenin diosgenin in cell suspensions of Trigonella foenum-graecum L. The results reveal that diosgenin synthesis and/or accumulation was stimulated by 5 ppm ethephon treatment, an increase of 126% being observed, while concentrations of 25 ppm and 50 ppm reduced the levels of this secondary compound. Changes in the isopentenyl diphosphate isomerase activity, increased cell diameter, decreased cell packing (with all the ethephon concentrations assayed), increased cytoplasmic density (5 ppm ethephon treatment) and alterations in the membrane structures (25 and 50 ppm ethephon treatments) were also observed.     

Fetal and maternal tissue distribution of the new fluoroquinolone DW-116 in pregnant rats

We investigated maternal and fetal tissue distribution of DW-116, a newly developed fluoroquinolone with a broad antibacterial spectrum against both G(+) and G(-) bacteria, in pregnant rats. After oral administration of [14C]-DW-116 (labeled 1 mg and unlabeled 500 mg/kg) to female rats on the 18th day of gestational, groups of three rats were killed at various time points up to 24 h, and plasma and tissues were collected, processed and analyzed. [14C]-DW-116 was rapidly absorbed, and distributed into the maternal and fetal tissues, and it declined in a biphasic manner with elimination half-lives (t(1/2)) of 10-15 h and mean residence times (MRT(0-24 h)) of 4-9 h. The radioactivity in most tissues of both dams and fetus reached its peak within 1 h and radioactivity levels of up to 10-25% of the peak level were maintained until 24 h after dosing. Among various tissues, the radioactivity in the maternal lungs was the highest (27 times that of plasma) at the C(max). Radioactivity in other tissues including liver, kidney, heart, lung, brain, spleen, mammary gland, placenta, ovary and uterus was higher than that in the maternal plasma (one- to three-fold). The tissue-to-plasma partition coefficient (K(p), AUC(0-24 h,tissue)/AUC(0-24 h,plasma)) of [14C]-DW-116 in maternal tissues was highest in the lung (K(p)=3.7), followed by the spleen (2.2), kidney (2.0), liver (1.8), heart (1.5), placenta (1.3), brain (1.3), ovary (1.1), uterus (1.1), and mammary gland (1.0). The tissue-to-plasma partition coefficient values in fetal tissues were heart (K(p)=2.2), kidney (2.1), liver (1.9), lung (1.6) and brain (1.4). When lactating rats were given a single oral dose of [14C]-DW-116, the radioactivity was rapidly secreted into the milk with K(p) of 1.7 at T(max) (0.5 h). These results indicate that DW-116 or its related metabolite(s) rapidly cross the blood-placenta and blood-milk barrier, extensively distribute into the fetal tissues, and are eliminated from the body in a prolonged manner. This study sheds insights into the maternal and fetal tissue distribution of DW-116 and will be useful for assessing both therapeutic and toxicological relevance of DW-116 in pregnant subjects.     

Direct Biotransformation of Dioscin into Diosgenin in Rhizome of Dioscorea zingiberensis by Penicillium dioscin

Diosgenin is an important precursor for synthesis of more than 200 steroidal hormone medicines. Rhizome of Dioscorea zingiberensis C. H. Wright (RDZ) contained the highest content of diosgenin in Dioscorea plant species. Diosgenin is traditionally extracted by acid hydrolysis from RDZ. However, the acid hydrolysis process produces massive wastewater which caused serious environment pollution. In this study, diosgenin extraction by direct biotransformation with Penicillium dioscin was investigated. The spawn cultivation conditions were optimized as: Czapeks liquid culture medium without sugar and agar (1,000 ml) + 6.0 g dioscin/6.0 g DL, 30 °C, 36 h; solid fermentation of RDZ: mycelia/RDZ of 0.05 g/kg, 30 °C, 50 h; the yield of diosgenin was over 90 %. Spawn cultivation was crucial for the direct biotransformation. In the spawn cultivation, amount and ratio of dioscin/DL were the key factors to promote biotransformation activity of P. dioscin. This biotransformation method was environment-friendly, simple and energy saving, and might be a potential substitute for acid hydrolysis in diosgenin extraction industry.     

云南永善产蜘蛛抱蛋的甾体成分

从云南省永善县产的百合科植物蜘蛛抱蛋（Ａｓｐｉｄｉｓｔｒａ ｅｌａｔｉｏｒ）根状茎中分离得到８个甾体化合物。其中６个为已知的甾体化合物,分别鉴定为（２５Ｓ）－３β－ｈｙｄｒｏｘｙ－ｓｐｉｒｏｓｔ－５－ｅｎｅ ３－Ｏ－β－Ｄ－ｇｌｕｃｏｐｙｒａｎｏｓｙｌ（１→）「β－Ｄ－ｘｙｌｏｐｙｒａｎｏｓｙｌ（１→３）」－β－Ｄ－ｇｌｕｃｏｐｙｒａｎｏｓｙｌ（１→４）－β－Ｄ－ｇａｌａｃｔｏｐｙｒａｎｏｓｉｄｅ     

Concise synthesis and antitumor activities of trisaccharide steroidal saponins

The naturally derived trisaccharide steroidal saponin 1 and structurally modified derivatives 2 and 3 bearing the same sarsasapogenin aglycon were synthesized concisely via a direct transglycosylation strategy. The antitumor activities of the synthetic trisaccharide saponins 1–3 and their corresponding α-isomers 1a–3a were preliminarily evaluated against human gastric adenocarcinoma cell (MKN-45) and human epithelial cervical cancer cell (HeLa) by CCK-8 assay.