Neocandenatone, an isoflavan-cinnamylphenol quinone methide pigment from Dalbergia congestiflora

A purple pigment neocandenatone (vestitol[6-->9";7O-->7"]obtusaquinone) was isolated from the heartwood of campincerán (Dalbergia congestiflora), an endemic Mexican tree. The isoflavan-cinnamyl phenol quinone methide structure of this compound was elucidated by HRMS, IR, and 1H and 13C NMR spectroscopic analysis, including 2D experiments (COSY, NOESY, HMQC and HSQC).     

Dalnigrin, a neoflavonoid marker for the identification of Brazilian rosewood (Dalbergia nigra) in CITES enforcement

International trade in Brazilian rosewood, Dalbergia nigra (Vell.) Allemão ex Benth., is regulated by the Convention on International Trade in Endangered Species of Wild Fauna and Flora (CITES). One problem in enforcing these regulations is the difficulty in distinguishing the wood of D. nigra from that of a closely-related but unregulated species, Dalbergia spruceana Benth. Using LC-MS to analyse methanol extracts of xylaria specimens, we identified a chemical marker for D. nigra heartwood, and determined its structure as the neoflavonoid 6-hydroxy-7-methoxy-4-(4-methoxyphenyl)-2H-1-benzopyran-2-one (4′-O-methylmelanettin; dalnigrin), using spectroscopic techniques. Dalnigrin was present in all nine available heartwood specimens of D. nigra, but it was not detected in extracts of 59 other heartwood samples representing 15 species of Dalbergia, including D. spruceana. Five other phenolic compounds were also isolated from D. nigra heartwood and similarly identified as the neoflavonoids 3′-hydroxymelanettin, melanettin, melannein and dalbergin, and the isoflavone caviunin. In extracts of D. spruceana heartwood, pseudobaptigenin was identified by LC-MS to be a major phenolic component that was not detected in wood extracts of D. nigra. We conclude that chemical analysis, in combination with anatomical investigation, can provide persuasive evidence to support the positive identification of untreated heartwood of D. nigra.     

Isoflavanoids of dalbergia Oliveri

The heartwood of Dalbergia oliveri has yielded 11 natural products of which two are new 3-phenylcoumarins. The structures of the extractives have been examined by physical methods and in addition the assigned structures have been confirmed by synthesis. The oxygenation pattern relating the structures of chalcone, isoflavone, pterocarpans, 3-arylcoumarins, coumestones and the isoflavan and isoflavanone suggests a common biosynthetic origin.     

Estrogenic constituents of the heartwood of Dalbergia parviflora

From the heartwood of Dalbergia parviflora, five compounds, dalparvin A (1), B (2), C (3), dalparvinol C (4), and neokhriol A (5), along with 11 known compounds, kenusanone G (6), cajanin (7), sophorol (8), alpinetin (9), hesperetin (10), 3'-O-methylorobol, odoratin, (2R)(3R)-2,3-trans 7-hydroxy-5-methoxydihydroflavonol, (6aR, 11aR)-3,8-dihydroxy-9-methoxypterocarpan, (6aR, 11aR)- vesticarpan, and methyl-3,4-dihydroxy-2-methoxybenzoate were isolated and characterized. Isolates were evaluated for their cell proliferation stimulatory activity against MCF-7, T-47D, and BT20 human breast cancer cell lines. Along with 7-10, two compounds 2 and 3 stimulated not only MCF-7, but also T-47D human breast cancer cell proliferation. Compound 6 had activity only against MCF-7 cells, and the activity of 7 was more than equivalent to that of daidzein. On the other hand, none of the isolates had any significant effects on BT20 cell proliferation, and these results indicated that the stimulative activity of these compounds was not general to any cell proliferations. Furthermore, these compounds were tested in the estrogen-responsive transient luciferase reporter assay.     

Arylbenzofurans from Dalbergia parviflora

The isolation of two polysubstituted arylbenzofurans from the heartwood of Dalbergia parviflora is described. Their structures were elucidated mainly by spectroscopic techniques (UV, IR, ¹H and ¹³C NMR). They were named parvifuran (5-hydroxy-6-methoxy-3-methyl-2-phenylbenzofuran) and isoparvifuran (5-hydroxy-6-methoxy-2-methyl-3-phenylbenzofuran) and are the first compounds of this class isolated from a Dalbergia species.     

Induction of Cr(VI) reduction activity in an Anoxybacillus strain under heat stress: a biochemical and proteomic study

A bacterial strain, designated as TSB-6, was isolated from the sediments of a Tantloi (India) hot spring at 65 °C. The strain showed 98% 16S rRNA gene sequence similarity with Anoxybacillus kualawohkensis strain KW12 and was found to grow optimally at 37 °C. However, growing cells, cell suspensions, and cell-free extracts from 65 °C cultures showed higher Cr(VI) reduction activities when assayed at either 37 or 65 °C than those obtained from 37 °C cultures. On fractionation of extracts from cells grown at 65 °C, the chromate reductase activity assayed at 65 °C was found mostly in the soluble fraction. When log-phase cells growing at 37 °C were shifted to 65 °C, the stressed cells produced larger quantities of reactive oxygen species. Consequently, growth of the cells was retarded, but specific Cr(VI) reduction activity increased. 2D gel electrophoresis followed by MALDI-TOF MS/MS identified the proteins whose expression level changed as a result of heat stress. The upregulated set included proteins involved in cellular metabolism of sugar, nucleotide, amino acids, lipids and vitamins, oxidoreductase activity, and protein folding. The downregulated proteins are also involved in cellular metabolism, DNA binding, and environmental signal processing.     

Evaluation of a Continuous Quantification Method of Apoptosis and Necrosis in Tissue Cultures

In tissue-engineering and other life sciences, there is a growing need for real-time, non-destructive information on apoptosis and necrosis in 2D and 3D tissue cultures. Previously, propidium iodide was applied as a fluorescent marker for monitoring necrosis. In the current study this technique was extended with a fluorescent apoptosis marker, YO-PRO-1, to discriminate between both stages of cell death. The main goal was to evaluate the performance of YO-PRO-1 and propidium iodide during monitoring periods of up to 3 days. Apoptosis was induced in C2C12 cultures and the numbers of YP-positive and PI-positive nuclei were counted in time. The performance of the dual staining was evaluated with a metabolic measure and a probe intensity study. Cell metabolism was unaffected during the first 24 h of testing. In conclusion, the YP/PI dual staining method was found to be a powerful tool in obtaining real-time spatial information on viability in cell and tissue culture without culture disruption.     

Two ellagitannins from Punica granatum heartwood

In the course of studies on polyphenol metabolism in Punica granatum heartwood two new ellagitannins, diellagic acid rhamnosyl (1-->4) glucopyranoside and 5-O-galloylpunicacortein D were isolated and characterized together with four known tannin metabolites, punicacortein D, punicalin, punicalagin and 2-O-galloylpunicalin. Structures of the isolated compounds were established by chromatography, chemical degradation, UV and 1D/ 2D 1H/13C NMR spectroscopy.     

Phenolic compounds from Dalbergia odorifera

Six new phenolic compounds (1–6), along with five known phenolics (7–11) were isolated from the heartwood of Dalbergia odorifera T. Chen. (Leguminosae). Their structures were determined by spectroscopic techniques including MS, UV, IR, 1D and 2D NMR. Bioassay results showed that compound 1 exhibited cytotoxicity against SGC-7901 and BEL-7402 tumor cell lines, while compounds 3, 5 and 10 showed antibacterial activities against Ralstonia solanacearum.     

Neoflavanoids of Dalbergia cultrata

In addition to the known neoflavanoids (S)-4-methoxydalbergione, dalbergin, and stevenin, the heartwood of Dalbergia cultrata Grah. contains a new 3,3-diphenylprop-l-ene (Ia). The assigned structure is based on data acquired mainly by physical methods. The softwood of D. ferruginea Roxb. ¹ yielded sitosterol and friedelin.     

Feedback regulation of beta-thujaplicin production and formation of its methyl ether in a suspension culture of Cupressus lusitanica

Suspension cell cultures of Cupressus lusitanica produce beta-thujaplicin, a tropolone found mostly in Cupressaceae heartwood. The factors controlling beta-thujaplicin accumulation in this cell culture system were investigated. Initial cell density of the cultures did not affect beta-thujaplicin levels, though initial addition of beta-thujaplicin suppressed its de novo production. When beta-thujaplicin accumulation reached a certain level (ca. 40 mg/l) in the medium, the cultures seemed to cease beta-thujaplicin production. However, beta-thujaplicin productivity was restored when the beta-thujaplicin-containing medium was exchanged for fresh medium; the formation of 2-methoxy-6-(methylethyl)cyclohepta-2,4,6-trien-1-one, an isomer of methylated beta-thujaplicin, in medium was also observed. These results suggest that beta-thujaplicin synthesis was regulated by product feedback mechanism in this cell line, and that excess accumulation of beta-thujaplicin is relieved by conversion of beta-thujaplicin to its methyl ether.     

3D spheroid cultures improve the metabolic gene expression profiles of HepaRG cells

3D (three-dimensional) cultures are considered to be an effective method for toxicological studies; however, little evidence has been reported whether 3D cultures have an impact on hepatocellular physiology regarding lipid or glucose metabolism. In the present study, we conducted physiological characterization of hepatoma cell lines HepG2 and HepaRG cells cultured in 3D conditions using a hanging drop method to verify the effect of culture environment on cellular responses. Apo (Apolipoprotein)B as well as albumin secretion was augmented by 3D cultures. Expression of genes related to not only drug, but also glucose and lipid metabolism were significantly enhanced in 3D cultured HepaRG spheroids. Furthermore, mRNA levels of CYP (cytochrome P450) enzymes following exposure to corresponding inducers increased under the 3D condition. These data suggest that this simple 3D culture system without any special biomaterials can improve liver-specific characteristics including lipid metabolism. Considering that the system enables high-throughput assay, it may become a powerful tool for compound screening concerning hepatocellular responses in order to identify potential drugs.     

Sulfate-Dependent Interspecies H(2) Transfer between Methanosarcina barkeri and Desulfovibrio vulgaris during Coculture Metabolism of Acetate or Methanol

We compared the metabolism of methanol and acetate when Methanosarcina barkeri was grown in the presence and absence of Desulfovibrio vulgaris. The sulfate reducer was not able to utilize methanol or acetate as the electron donor for energy metabolism in pure culture, but was able to grow in coculture. Pure cultures of M. barkeri produced up to 10 mumol of H(2) per liter in the culture headspace during growth on acetate or methanol. In coculture with D. vulgaris, the gaseous H(2) concentration was 相似文献   

Effect of glycine on the cell yield and growth rate of Escherichia coli: evidence for cell-density-dependent glycine degradation as determined by (13)C NMR spectroscopy.

Addition of selected amino acids could be a means to improve production of recombinant proteins in industrial processes. We found that glycine increased the maximum specific growth rate of Escherichia coli from 0.67 to 0.78 h(-1), and the cell yield from 0.57 to 0.98 g dry weight per g substrate, when supplemented to batch cultures in a glucose-mineral medium. Maximum effect occurred at pH 6.8, at a glycine concentration of 6-12 mmol l(-1), and at cell densities below 1.15 g dry weight l(-1) (0D(610).3). When glycine was added to a culture at a cell density of 1.15 g l(-1) or above, no growth promoting effect of glycine was seen. The 'glycine effect' was not due to CO(2) produced by the glycine cleavage system (GCV), and the lack of effect at higher cell densities was not masked by acetate accumulation, but coincided with increased acetate production. The metabolism of glycine was further investigated in cultures supplied with [2-(13)C] labelled glycine, and the redistribution of label in the [1-(13)C], [2-(13)C], and [1,2-(13)C] isotopomeres of excreted acetate was analysed by 13C NMR. The NMR data revealed that very little degradation of glycine occurred at cell densities below 1.15 g l(-1). Simultaneously the biosynthesis of serine and glycine was repressed as judged by the absence of [2-(13)C] acetate, implying that added glycine was used as a source of glycine, serine, one-carbon units, and threonine. At cell densities above 1.15 g l(-1), 53% of the consumed glycine carbon was excreted as acetate. Degradation of glycine was associated with an increased uptake rate, cleavage by GCV, and degradation of both glycine-derived serine, and glucose-derived serine to pyruvate. This switch in metabolism appears to be regulated by quorum sensing.     

Aspergillus candidus is a newly recognized source of sphaeropsidin A: Isolation,semi-synthetic derivatization and anticancer evaluation

This report details a search for alternative strains that produce the diterpenoid sphaeropsidin A (SphA) among A. candidus strains from the USDA Northern Regional Research Laboratories Culture Collection. We identified two strains that produced SphA using a limited set of test media. An initial scaled-up fermentation of NRRL 313 and isolation effort led to the procurement of sufficient quantities of SphA to prepare five semi-synthetic analogues (1–5) and evaluate their anticancer effects against glioblastoma cells D423 and Gli56 grown in 2D and 3D cultures. Although, the effectiveness of the synthetic analogues varied depending on the cell line and the type of cell culture, compound 5, bearing an aromatic ring at C16, displayed a stronger toxicity towards both D423 and Gli56 cell lines in 2D cultures and D423 spheroids in 3D culture than either SphA or compounds 1–4.     

Simultaneous quantitative measurement of luciferase reporter activity and cell number in two- and three-dimensional cultures of hepatitis C virus replicons

Hepatitis C virus (HCV) is a global health problem and an important human pathogen. The development of cell culture models for HCV infection has been difficult to accomplish, primarily because HCV is very sensitive to the host cell state. Future models will require the use of three-dimensional (3D) cultures that model the host cell state and environment more accurately. Higher information content screens for anti-HCV therapeutics will also involve 3D cell cultures. Here we report a method for screening cell models for HCV replication that involves normalizing luciferase reporter activity based on cell number in two-dimensional (2D) and 3D HCV replicon cultures. Human hepatoma cells stably replicating luciferase-containing HCV replicons were cultured in 2D monolayer culture and 3D spheroid culture. Optimization of cell lysis was performed so that cell lysates could be used to quantify both luciferase activity and cellular DNA content. Cellular DNA content was quantified using Hoechst 33258 dye and was converted to cell number. The method is straightforward, reproducible, and sensitive down to 5000 cells. This method enables low-throughput but high-information content screening of HCV replicons, with the potential for high-throughput screening in a variety of 3D cultures and cocultures.     

Coelobillardin,an iridoid glucoside dimer from Coelospermum billardieri

An isoflavanoid (6-->2) neoflavonoid dimer and a 4',5',7-trihydroxy-2'-methoxyisoflavone, both as the acetate derivatives were isolated from the heartwood of Dalbergia nitidula. Their structures were established by spectroscopic methods.