Biomedical subjects

Hermann Stuppner

Publications and source records attributed to Hermann Stuppner.

At least 19 recordsLinked to original sources

PubMed2006

Isogentisin--a novel compound for the prevention of smoking-caused endothelial injury.

The best strategy in the fight against tobacco-induced diseases is prevention. However, more than one billion people around the world are smokers. Most of these people will develop or already suffer from tobacco-induced diseases. In this project, we screened 22 natural alpine plant extracts for their potential to protect human vascular endothelial cells from cigarette smoke-induced cell damage. Extracts from Gentiana lutea (Yellow Gentian) proved to be effective, and were therefore subjected to bio-guided fractionation. Although our analyses suggest that G. lutea contains several active principles, fractions containing isogentisin (1,3-dihydroxy-7-methoxyxanthone), and pure isogentisin, were most effective. In experiments addressing the nature of the mechanism of protection, we were able to show that isogentisin does not directly interfere with cigarette smoke chemicals. Addition of isogentisin to the cells as long as 4.5h after exposure to cigarette smoke chemicals protected endothelial cells from cell death. Finally, detailed analyses of intracellular oxidative stress and protein oxidation suggest that isogentisin promotes cell survival by activating cellular repair functions.

Apoptosis↗

PubMed2006

Venturia inaequalis-inhibiting Diels-Alder adducts from Morus root bark.

In organic apple orcharding there is a continuous need for natural fungicides effective against Venturia inaequalis (Cooke) Winter, the causal agent of apple scab. In this study an in vitro assay is presented for determining the germination inhibitory potential of extracts and pure compounds. From a screening of plant extracts, the methanol extract of Morus root bark revealed distinct V. inaequalis inhibiting qualities, which were subjected to a bioguided fractionation. Among the isolated metabolites [moracins M (1), O/P (2), kuwanon L (3), and sanggenons D (4), B (5), G (6), O (7), E (8), and C (9)] all the Diels-Alder adducts (3-9) showed an antifungal activity with IC50 values between 10 and 123 microM. The in vitro activity of the most active fraction (A5, IC50 39.0 +/- 4.2 microg/mL) was evaluated in vivo, confirming a distinct antifungal activity against V. inaequalis for the tested natural material.

Antifungal Agents↗

PubMed2006

Reaction product analysis by high-performance liquid chromatography-solid-phase extraction-nuclear magnetic resonance Application to the absolute configuration determination of naturally occurring polyyne alcohols.

The absolute configuration of secondary hydroxy functions of seven natural occurring polyyne derivatives has been elucidated by the application of Mosher method of diastereomeric methoxy-2-trifluoromethyl-phenylacetyl (MTPA) ester formation. High-performance liquid chromatography with diode array detection (HPLC-DAD) of the reaction mixture using a water/acetonitrile gradient allowed monitoring the reaction progress. Coupling of high-performance liquid chromatography to solid-phase extraction combined with nuclear magnetic resonance (HPLC-SPE-NMR) was utilized to generate highly reproducible (1)H and (19)F NMR data needed as input for the absolute configuration determination based on the analysis of relative shift differences. Chromatographic peaks of reaction substrates and reaction products bearing less 10mug analyte were trapped on SPE cartridges with the aid of water as makeup solvent. Deuterated chloroform was used to elute and transfer the peak content from the SPE to the 60mul flow cell of a 500MHz NMR spectrometer. For each analyte (1)H NMR spectra were obtained within 15min. Additionally (19)F NMR spectra were recorded for selected analytes in the same timeframe. Based on the obtained NMR data, the absolute configuration of all polyynes under investigation was successfully designated.

Alkaloids↗

PubMed2006

Determination of adenine and pyridine nucleotides in glucose-limited chemostat cultures of Penicillium simplicissimum by one-step ethanol extraction and ion-pairing liquid chromatography.

Under specific conditions Penicillium simplicissimum excretes large amounts of organic acids, mainly citrate. As the energetic status of the hyphae might play a role in that respect, we developed a method for the determination of adenine (adenosine triphosphate, adenosine diphosphate, and adenosine monophosphate) and pyridine (nicotinamide adenine dinucleotide and reduced nicotinamide adenine dinucleotide (NADH)) nucleotides in hyphae of P. simplicissimum. An optimum separation of the five compounds in less than 15 min was possible on a C-8 column, utilizing 50 mM aqueous triethylamine-buffer (pH 6.5) and acetonitrile as mobile phase; detection was performed at 254 nm. With the exception of NADH, which could not be determined accurately due to stability problems, the method was sensitive (LOD < or = 0.7 ng on-column), repeatable (sigma(rel) < or = 4.4%), accurate (recovery rates between 97.9 and 104.9%), and precise (intraday variation < or = 9.4%, interday variation < or = 6.2 %). For an optimum extraction of the nucleotides the chemostat samples were directly placed into hot (90 degrees C) 50% ethanol, and shaken for 10 min, followed by evaporation of the solvent and a solid phase extraction cleanup of the redissolved aqueous samples. With this method the nucleotide concentrations in hyphae from a glucose-limited chemostat culture and the respective energy charge were determined. Additionally, the effect of the time lag between sampling and extraction and the effect of a glucose pulse on nucleotide concentrations were determined.

Adenine Nucleotides↗

PubMed2006

Stilbenoids from Tragopogon orientalis.

A phytochemical investigation of Tragopogon orientalis L. (Asteraceae, Cichorieae) yielded the natural products 6''-O-(7,8-dihydrocaffeoyl)-alpha,beta-dihydrorhaponticin, 3'-O-methyl-alpha,beta-dihydrorhaponticin, and (S)-3-(4-beta-glucopyranosyloxybenzyl)-7-hydroxy-5-methoxyphtalide as well as known compounds alpha,beta-dihydrorhaponticin, 3-(4-methoxybenzyl)-5,7-dimethoxyphthalide, p-dihydrocoumaric acid methyl ester, and 1-hydroxypinoresinol-1-O-beta-glucopyranoside. The structures were established by HR mass spectrometry, extensive 1D and 2D NMR spectroscopy, and CD spectroscopy. Moreover, the radical scavenging activities of the major compounds were measured using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay. The chemosystematic impact of the occurrence of stilbene derivatives in T. orientalis is discussed.

Antioxidants↗

PubMed2006

Development and validation of an HPLC/UV/MS method for simultaneous determination of 18 preservatives in grapefruit seed extract.

Grapefruit seed extracts are used in cosmetics, food supplements, and pesticides because of their antimicrobial properties, but suspicions about the true nature of the active compounds arose when synthetic disinfectants such as benzethonium or benzalkonium chloride were found in commercial products. The HPLC method presented herein allows the quality assessment (qualitative and quantitative) of these products for the first time. On the basis of a standard mixture of 18 preservatives most relevant for food and grapefruit products, a method was developed allowing the baseline separation of all compounds within 40 min. Optimum results were obtained with a C-8 stationary phase and a solvent system comprising aqueous trifluoroacetic acid, acetonitrile, and 2-propanol. The assay was fully validated and shown to be sensitive (LOD < or= 12.1 ng on-column), accurate (recovery rates > or = 96.1%), repeatable (sigma(rel) < or = 3.5%), precise (intra-day variation < or = 4.5%, interday variation < or = 4.1%), and rugged. Without any modifications the method could be adopted for LC-MS experiments, where the compounds of interest were directly assignable in positive ESI mode. The quantitative results of several products for ecofarming confirmed previous studies, as seven out of nine specimens were adulterated with preservatives in varying composition. The samples either contained benzethonium chloride (2.5-176.9 mg/mL) or benzalkonium chloride (138.2-236.3 mg/mL), together with smaller amounts of 4-hydroxybenzoic acid esters, benzoic acid, and salicylic acid.

Chromatography, High Pressure Liquid↗

PubMed2006

Apolar chromatography on Sephadex LH-20 combined with high-speed counter-current chromatography. High yield strategy for structurally closely related analytes-Destruxin derivatives from Metarhizium anisopliae as a case study.

A novel high yield isolation procedure for lipophilic cyclic peptide derivatives is presented. Destruxin (dtx) A, B, D, E, and E-diol retrieval from Metarhizium anisopliae culture broth was achieved with a three-step purification protocol. After liquid-liquid extraction column chromatography over Sephadex LH-20 served as enrichment step. High-speed counter-current chromatography (HSCCC) was used for the final purification. Within the first chromatographic step dtx D and dtx E-diol were separated in purities exceeding 90%. The separation of dtx A, B, and E was achieved from an enriched Sephadex LH-20 fraction by a HSCCC protocol using light petroleum-ethyl acetate-methanol-water = 2:5:2:5 (v/v) as eluent system. These derivatives were obtained in purities above 98% and total yields exceeding 40%.

Ascomycota↗

PubMed2006

In vivo efficacy of different extracts of Edelweiss (Leontopodium alpinum Cass.) in animal models.

Dichloromethane, methanolic and CO(2) extracts of the aerial parts and roots of Edelweiss (Leontopodium alpinum Cass.) were investigated for their anti-inflammatory and analgesic effects after oral administration. The highest activity in rat's paw edema assay was found for the lipophilic extracts of the aerial plant parts (dose 200 mg/kg), exhibiting a swelling reduction of 72% (CO(2)-extract) and 80% (DCM-extract), respectively. Histological evaluation of the treated paws showed a significant reduction of the inflammatory response in the pre-treated specimens. On the contrary in the acetic acid-induced writhing test the dichloromethane extract of the root extract exhibited more pronounced analgesic effects than the extracts of the aerial parts, suggesting a different pattern of active compounds. As far as gastrointestinal effects are concerned, oral administration of aerial parts (hDCM 200 mg/kg) to mice induces a highly significant inhibition in gastrointestinal propulsion probably related to the presence of so far unknown compounds. Moreover, the antioxidant capacity of some extracts was studied in order to establish a possible correlation with anti-inflammatory properties.

Analgesics↗

PubMed2006

Altitudinal variation of secondary metabolite profiles in flowering heads of Arnica montana cv. ARBO.

The altitudinal variation on the contents of secondary metabolites in flowering heads of Arnica montana was assessed. Plants of A. montana cultivar ARBO were grown in nine experimental plots at altitudes between 590 and 2230m at Mount Patscherkofel near Innsbruck/Austria. The total contents of sesquiterpene lactones and flavonoids were not positively correlated with the altitude of the growing site. However, the proportion of flavonoids with vicinal free hydroxy groups in ring B to flavonoids lacking this feature significantly increased with elevation. Additionally, the level of caffeic acid derivatives also positively correlated with the altitude of the growing site. In particular amounts of 1-methoxyoxaloyl-3,5-dicaffeoylquinic acid significantly increased in higher sites and samples from the summit region contained 85% more of this compound than samples from valley sites. These results are discussed with regards to chemosystematic studies comparing samples collected in different altitudes as well as in the light of a UV-B protective and radical scavenging function of phenolics and their significance for plant life in environments with elevated UV-B radiation.

Altitude↗

PubMed2006

Taspine: bioactivity-guided isolation and molecular ligand-target insight of a potent acetylcholinesterase inhibitor from Magnolia x soulangiana.

A bioactivity-guided approach was taken to identify the acetylcholinesterase (AChE, EC 3.1.1.7) inhibitory agent in a Magnolia x soulangiana extract using a microplate enzyme assay with Ellman's reagent. This permitted the isolation of the alkaloids taspine (1) and (-)-asimilobine (2), which were detected for the first time in this species. Compound 1 showed a significantly higher effect on AChE than the positive control galanthamine and selectively inhibited the enzyme in a long-lasting and concentration-dependent fashion with an IC(50) value of 0.33 +/- 0.07 muM. Extensive molecular docking studies were performed with human and Torpedo californica-AChE employing Gold software to rationalize the binding interaction. The results suggested ligand 1 to bind in an alternative binding orientation when compared to galanthamine. While this is located in close vicinity to the catalytic amino acid triad, the 1-AChE complex was found to be stabilized by (i) sandwich-like pi-stacking interactions between the planar aromatic ligand (1) and the Trp84 and Phe330 of the enzyme, (ii) an esteratic site anchoring with the amino side chain, and (iii) a hydrogen-bonding network.

Alkaloids↗

PubMed2006

Oxindole alkaloids from Uncaria tomentosa induce apoptosis in proliferating, G0/G1-arrested and bcl-2-expressing acute lymphoblastic leukaemia cells.

Natural products are still an untapped source of promising lead compounds for the generation of antineoplastic drugs. Here, we investigated for the first time the antiproliferative and apoptotic effects of highly purified oxindole alkaloids, namely isopteropodine (A1), pteropodine (A2), isomitraphylline (A3), uncarine F (A4) and mitraphylline (A5) obtained from Uncaria tomentosa, a South American Rubiaceae, on human lymphoblastic leukaemia T cells (CCRF-CEM-C7H2). Four of the five tested alkaloids inhibited proliferation of acute lymphoblastic leukaemia cells. Furthermore, the antiproliferative effect of the most potent alkaloids pteropodine (A2) and uncarine F (A4) correlated with induction of apoptosis. After 48 h, 100 micromol/l A2 or A4 increased apoptotic cells by 57%. CEM-C7H2 sublines with tetracycline-regulated expression of bcl-2, p16ink4A or constitutively expressing the cowpox virus protein crm-A were used for further studies of the apoptosis-inducing properties of these alkaloids. Neither overexpression of bcl-2 or crm-A nor cell-cycle arrest in G0/G1 phase by tetracycline-regulated expression of p16INK4A could prevent alkaloid-induced apoptosis. Our results show the strong apoptotic effects of pteropodine and uncarine F on acute leukaemic lymphoblasts and recommend the alkaloids for further studies in xenograft models.

Alkaloids↗

PubMed2005

Quantification of Fumaria officinalis isoquinoline alkaloids by nonaqueous capillary electrophoresis-electrospray ion trap mass spectrometry.

A capillary electrophoresis (CE) method using non-aqueous (NA) separation solutions combined with an ion trap mass spectrometer (MS and MS/MS) as detection device is presented for the separation, identification and quantification of isoquinoline alkaloids from Fumaria officinalis. The best results were obtained with a mixture of acetonitrile-methanol (9:1, v/v) containing 60mM ammonium acetate and 2.2M acetic acid as running electrolyte and an applied voltage of 30 kV. Electrospray MS measurements were performed in the positive ionization mode with isopropanol-water (1:1, v/v) as sheath liquid at a flow rate of 3 microl/min. Alkaloids were detected as [M+H](+)-ions and showed typical fragmentation patterns in MS/MS experiments. The developed assay was used for the quantification of seven isoquinoline alkaloids representing different structural subtypes in Fumariae herba extracts and F. herba containing phytopharmaceuticals.

Alkaloids↗

PubMed2005

Comparative molecular and phytochemical investigation of Leontodon autumnalis (Asteraceae, Lactuceae) populations from Central Europe.

Previous analyses of Leontodon autumnalis L. revealed the existence of two chemotypes. In the current study molecular and phytochemical methods were combined to investigate 24 Central European populations of L. autumnalis. The focus of this study was the correlation of molecular and phytochemical characters at the intraspecific level. DNA fingerprint profiles of 183 individuals were obtained by random amplified polymorphic DNA (RAPD) providing 77 molecular markers. Contents of phenolics and sesquiterpenoids of flowering heads and sub-aerial parts were quantified by HPLC-DAD analyses. HPLC results were evaluated by principal component analysis. Geographic distribution of the two detected chemotypes partially overlapped. Phylogenetic groupings displayed in an unrooted neighbor-joining tree calculated from the RAPD data matrix were correlated with the geographical origin of the plant material. However, genetic profiles neither correlated with the two chemotypes nor with the morphologically based subspecies of L. autumnalis recognized by some authors. The presented data imply that the morphotypes are of multiple origins or due to different ecological growing conditions rather than genetically determined and that phytochemical races are induced by a limited number of genetical differences, which might have occurred independently in different lineages of the L. autumnalis group.

Asteraceae↗

PubMed2005

Analysis of phenolic glycosides and saponins in Primula elatior and Primula veris (primula root) by liquid chromatography, evaporative light scattering detection and mass spectrometry.

This paper describes the first liquid chromatographic method suitable for the simultaneous determination of bioactive compounds, saponins and phenolic glycosides, present in Primula elatior and Primula veris, including the NMR data of primulaverin and primeverin. Optimum separations were obtained with a Synergi 4mum Fusion RP 80A column, using 0.025% TFA in water and 5% acetonitrile in methanol as mobile phase. Saponins were detected by evaporative light scattering detection (ELSD), whereas the phenolic glycosides were monitored by UV at 210 nm. The method was validated for repeatability (sigma(rel) or=97.1%) and sensitivity (LOD <or=22 ng (UV) and <or=38 ng (ELSD) on-column, respectively). LC-MS experiments in negative APCI mode allowed a final peak assignment. Both Primula species could easily be differentiated by their saponin pattern. The total saponin content was highest in P. veris roots (max. 14.9%), the aerial parts or P. elatior contained significantly less amounts; primeverin (0.64-1.42%) showed to be the most dominant phenolic glycoside.

Chromatography, Liquid↗

PubMed2005

High-performance liquid chromatography-diode array detection assay for the detection and quantification of the Beauveria metabolite oosporein from potato tubers.

A high-performance liquid chromatography-diode array detection (HPLC-DAD) assay is described for the detection and quantification of the secreted Beauveria brongniartii metabolite oosporein from potato tubers. Analyte recovery was achieved with a Britton-Robinson buffer system at pH 5.5 diluted with methanol 3:7 (v/v) (BR5.5-MeOH). An internal standard protocol using 2-iodobenzoic acid was established to minimize analytical error. The resulting assay, using a binary solvent gradient with acidic modifiers and detecting the metabolite at 287 nm, showed a limit of detection (LOD) of 2.4 mg oosporein/kg potato tubers. The oosporein content of potato tuber samples obtained from a field trial using the biological pest control B. brongniartii formulation Melocont-Pilzgerste in up to five-fold higher doses (250 kg Melocont-Pilzgerste/ha) as recommended per year was found to be below the established LOD.

Ascomycota↗

PubMed2005

Distribution of a new rosmarinic acid derivative in Eryngium alpinum L. and other Apiaceae.

The roots of Eryngium alpinum L. (Apiaceae) demonstrated radical scavenging properties toward the 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical in a TLC autographic assay. Isolation of the bioactive compounds allowed the identification of R-(+)-3'-O-beta-D-glucopyranosyl rosmarinic acid, a new rosmarinic acid derivative. The quantitative determination of the antioxidant capacity of the compound by chemoluminescence demonstrated half the activity of R-(+)-rosmarinic acid. To find another source richer in this compound, a chemotaxonomic study was conducted. The higher content was found in the aerial parts of Sanicula europeae L., also belonging to the Saniculoideae subfamily. Although present in most of the Eryngium species, this compound was not detected in Imperatoria ostruthium L., Pimpinella peregrina L., and Levisticum officinalis L. species from the Apioideae subfamily and Hydrocotyle asiatica L. from the Hydrocotyloideae subfamily. The results indicate that the new derivative R-(+)-3'-O-beta-D-glucopyranosyl rosmarinic acid is a potential chemotaxonomic marker of the Saniculoideae subfamily.

Antioxidants↗

PubMed2005

Polyacetylenes from the Apiaceae vegetables carrot, celery, fennel, parsley, and parsnip and their cytotoxic activities.

A dichloromethane extract of root celery yielded falcarinol, falcarindiol, panaxydiol, and the new polyacetylene 8-O-methylfalcarindiol. The structure of the new compound was established by one- and two-dimensional (1D and 2D) NMR, mass spectrometry, and optical rotation data. Nonpolar extracts of roots and bulbs of carrots, celery, fennel, parsley, and parsnip were investigated for their content of polyacetylenes by high-performance liquid chromatography with diode array detection (HPLC-DAD). All five species contained polyacetylenes, although carrots and fennel only in minor amounts. Additionally, the cytotoxicity of the four polyacetylenes against five different cell lines was evaluated by the annexin V-PI assay. Falcarinol proved to be the most active compound with a pronounced toxicity against acute lymphoblastic leukemia cell line CEM-C7H2, with an IC(50) of 3.5 micromol/L. The possible chemopreventive impact of the presented findings is discussed briefly.

Acetylene↗

PubMed2005

Development of a sensitive high-performance liquid chromatography-diode array detection assay for the detection and quantification of the beauveria metabolite oosporein from submerged culture broth and bio-control formulations.

A sensitive high-performance liquid chromatography-diode array detection (HPLC-DAD) assay is described for the detection and quantification of the Beauveria metabolite oosporein from fungal culture broth and two biocontrol agent formulations. In all cases, analyte recovery was achieved with a Britton-Robinson buffer system at pH 5.5 diluted with methanol 3:7 (v/v) (BR5.5-MeOH). The HPLC-DAD assay, using a binary solvent gradient with acidic modifiers and detecting the metabolite at 287 nm, showed linearity over 3 orders of magnitude and a limit of detection (LOD) of 6.0 +/- 2.3 microg of oosporein/L of BR5.5-MeOH. The oosporein content of the representative fungal culture broth samples and two Beauveria formulations (Melocont-Pilzgerste and Melocont-WP) was found to be 504.7 +/-13.6 mg of oosporein/L of culture filtrate, 7.4 +/- 0.6 mg of oosporein/kg of Melocont-Pilzgerste, and 38.2 +/- 1.3 mg of oosporein/kg of Melocont-WP with recovery rates of 93 +/- 2, 99 +/- 8, and 92 +/- 3%, respectively.

Benzoquinones↗