The clerodane diterpene casearin J induces apoptosis of T-ALL cells through SERCA inhibition, oxidative stress, and interference with Notch1 signaling.

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematologic malignancy that preferentially affects children and adolescents. Over 50% of human T-ALLs possess activating mutations of Notch1. The clerodane diterpene casearin J (CJ) is a natural product that inhibits the sarcoendoplasmatic reticulum calcium ATPase (SERCA) pump and induces cell death in leukemia cells, but the molecular mechanism of cytotoxicity remains poorly understood. Here we show that owing to SERCA pump inhibition, CJ induces depletion of the endoplasmic reticulum calcium pools, oxidative stress, and apoptosis via the intrinsic signaling pathway. Moreover, Notch1 signaling is reduced in T-ALL cells with auto-activating mutations in the HD-domain of Notch1, but not in cells that do not depend on Notch1 signaling. CJ also provoked a slight activation of NF-κB, and consistent with this notion a combined treatment of CJ and the NF-κB inhibitor parthenolide (Pt) led to a remarkable synergistic cell death in T-ALL cells. Altogether, our data support the concept that inhibition of the SERCA pump may be a novel strategy for the treatment of T-ALL with HD-domain-mutant Notch1 receptors and that additional treatment with the NF-κB inhibitor parthenolide may have further therapeutic benefits.

IκBα glutathionylation and reduced histone H3 phosphorylation inhibit eotaxin and RANTES.

Airway smooth muscle cells (ASMCs) secrete eotaxin and RANTES (regulated on activation, normal T-cell expressed and secreted) in response to tumour necrosis factor (TNF)-α, which is inhibited by the nuclear factor (NF)-κB inhibitor dimethylfumarate (DMF). NF-κB/IκB (inhibitor of NF-κB) glutathionylation and changes in chromatin remodelling can inhibit NF-κB activity. In this study, we determined whether NF-κB/IκB glutathionylation and reduced histone H3 phosphorylation might underlie the inhibitory effect of DMF on NF-κB activity, and eotaxin and RANTES secretion. Primary human ASMCs were treated with DMF, diamide and/or glutathione (GSH) ethylester (OEt) prior to TNF-α stimulation and were subsequently analysed by ELISA, electrophoretic mobility shift assay, immunofluorescence, co-immunoprecipitation or immunoblotting. DMF reduced intracellular GSH and induced IκBα glutathionylation (IκBα-SSG), which inhibited IκBα degradation, NF-κB p65 nuclear entry and NF-κB/DNA binding. In addition, DMF inhibited the phosphorylation of histone H3, which was possibly mediated by the inhibitory effect of DMF on mitogen- and stress-activated protein kinase (MSK)-1. However, p38 mitogen-activated protein kinase (MAPK), extracellular signal-regulated kinase MAPK and MAPK phosphatase-1, upstream of MSK-1, were not inhibited by DMF. Importantly, DMF-mediated effects on NF-κB, histone H3, eotaxin and RANTES were reversed by addition of GSH-OEt. Our data suggest that DMF inhibits NF-κB-dependent eotaxin and RANTES secretion by reduction of GSH with subsequent induction of IκBα-SSG and inhibition of histone H3 phosphorylation. Our findings offer new potential drug targets to reduce airway inflammation in asthma.

In vitro cytotoxic activity of abietane diterpenes from Peltodon longipes as well as Salvia miltiorrhiza and Salvia sahendica.

Phytochemical investigations of the n-hexane extract from the roots of Peltodon longipes (Lamiaceae) resulted in the isolation of 12 known abietane diterpenes (1-12). Structures were established on the basis of one and two dimensional nuclear magnetic resonance spectroscopic data ((1)H and (13)C, COSY, HSQC and HMBC), electron ionization mass spectrometric analysis (EIMS) as well as comparison with data from literature. These compounds, as well as eight known diterpenes (13-19) from Salvia miltiorrhiza, and two from Salvia sahendica (20 and 21) were evaluated for their cytotoxic effects in human pancreatic (MIAPaCa-2) and melanoma (MV-3) tumor cell lines using the MTT assay. Tanshinone IIa (13), 7α-acetoxyroyleanone (1), 1,2-dihydrotanshinone (16) and cryptotanshinone (14) had the highest cytotoxic effects in MIAPaCa-2, displaying IC(50) of 1.9, 4.7, 5.6, and 5.8 µM, respectively. Structure-activity relationships of abietane diterpenoid quinones are discussed.

Identification of rosmarinic acid as the major active constituent in Cordia americana.

ETHNOPHARMACOLOGICAL RELEVANCE: Preparation from leaves of Cordia americana have been widely used in traditional medicine in South Brazil to treat wounds and various inflammations. AIM OF THE STUDY: The objective of this work was to identify the effective compounds in the ethanolic extract prepared from the leaves of Cordia americana, which is used in traditional South Brazilian medicine as anti-inflammatory and wound healing remedy. MATERIALS AND METHODS: Isolation and structure elucidation techniques were performed in order to identify the compounds of Cordia americana and HPLC analysis was used for the quantification. The major constituent and the ethanolic extract were investigated for inhibition of 5-lipoxygenase, p38alpha MAPK, TNFalpha release and NF-kappaB as well as in the fibroblast scratch assay. RESULTS: Rosmarinic acid (1) was identified as the major compound with an amount of 8.44% in the ethanolic extract of the leaves of Cordia americana. The ethanolic extract as well as (1) exhibited the highest inhibitory effects on 5-lipoxygenase (IC(50)=0.69 and 0.97microg/mL, resp., IC50 of BWA4C as reference: 0.3microM) and p38alpha (IC50=3.25 and 1.16microg/mL, resp., IC50 of SB203580 as reference: 0.046microM) and moderate inhibitory effects on TNFalpha release. Slight effects were observed in the fibroblast scratch assay. CONCLUSIONS: This study increases our knowledge on the effective compound in Cordia americana and supports its use in traditional medicine. We demonstrated for the first time pharmacological effects of Cordia americana and we provide evidences for a crucial role of rosmarinic acid as the major key player.

Reseda luteola L. extract displays antiproliferative and pro-apoptotic activities that are related to its major flavonoids.

Reseda luteola L. has been used as a dye due to its high luteolin content since ancient times. However, no pharmacological studies have been performed with Reseda extracts so far. Here, we have assessed antiproliferative and apoptosis-inducing effects of the Reseda extract RF-40. It contains 40% flavonoids, primarily luteolin, but also luteolin-7-O-glucoside and apigenin. RF-40 and the isolated flavonoids dose-dependently inhibited cell proliferation and induced apoptotic oligonucleosomes in PHA-stimulated peripheral blood mononuclar cells. These effects were not due to cytotoxicity as shown with a luminometric ATP assay. Dose-response curves of RF-40 and the isolated flavonoids were similar, with luteolin being the most effective isolated flavonoid. Comparison of RF-40 to its major flavonoids revealed that the pharmacological effects of the extract can mostly be attributed to luteolin. We conclude that Reseda extract is an interesting raw material not only for dyeing purposes but also for further pharmacological investigation.

Determination of the wound healing effect of Calendula extracts using the scratch assay with 3T3 fibroblasts.

UNLABELLED: PHARMACOLOGICAL RELEVANCE: Presentation of the scratch assay as a convenient and inexpensive in vitro tool to gain first insights in the wound healing potential of plant extracts and natural compounds. AIM OF THE STUDY: The present study deals with the optimization of the scratch assay which can be used as an in vitro model for quantification of fibroblast migration to and proliferation into the wounded area. It is suitable for the first evaluation of the wound re-epithelialization potential of crude herbal extracts, isolated compounds and pharmaceutical preparations. As a proof of concept three preparations from traditional medicinal plants were investigated. MATERIALS AND METHODS: Swiss 3T3 albino mouse fibroblasts were used in monolayers and platelet derived growth factor as positive control. Hexane and ethanolic extracts from Calendula officinalis and Matricaria recutita, Hypericum oil as well as the triterpenoids faradiol myristate and palmitate were studied. To differentiate between proliferation and migration antimitotic mitomycin C was added. RESULTS: Both extracts of Calendula officinalis stimulated proliferation and migration of fibroblasts at low concentrations, e.g. 10 microg/ml enhanced cell numbers by 64.35% and 70.53%, respectively. Inhibition of proliferation showed that this effect is mainly due to stimulation of migration. Faradiol myristate and palmitate gave comparable stimulation rates at an almost 50 microg/ml concentration, indicating that they contribute partially, but not most significantly to the wound healing effects of Calendula preparations. Extracts from Matricaria recutita were only moderately active. Hypericum oil was cytotoxic at concentrations higher than 0.5 microg/ml. CONCLUSIONS: The scratch assay in the present form can be used as a promising scientific approach and platform to differentiate between plant extracts known for their wound healing and their anti-inflammatory properties.

Topical application of solubilized Reseda luteola extract reduces ultraviolet B-induced inflammation in vivo.

We investigated the skin tolerance and anti-inflammatory potential of a nanoparticular solubilisate of a luteolin-rich Reseda extract (s-RE) in two independent studies in vivo. Reseda luteola extract containing 40% flavonoids was solubilized with polysorbate, resulting in product micelles with a diameter of 10 (+/-1.5)nm. Standardized inflammation was induced by irradiating test areas on the back of healthy volunteers with defined doses of ultraviolet B (UVB). In the first study different concentrations of s-RE were tested in 10 volunteers to evaluate dose-dependency of anti-inflammatory effects of s-RE. In the second randomized, double-blind, placebo-controlled study a defined concentration of s-RE (2.5%w/w) was tested in 40 volunteers in comparison to the vehicle (glycerol) and hydrocortisone (1%w/w). s-RE dose-dependently reduced UVB-induced erythema when applied 30 min before irradiation. To a lesser extent, topical application of s-RE after irradiation also reduced UVB-induced erythema. s-RE was as effective as hydrocortisone, whereas the vehicle had no effect. Occlusive application of s-RE on non-irradiated test sites did not cause any skin irritation. Due to excellent skin tolerance combined with potent anti-inflammatory properties s-RE bears potential especially for the prevention but also for the treatment of inflammatory skin conditions such as UV-induced erythema.

Dimethylfumarate inhibits NF-{kappa}B function at multiple levels to limit airway smooth muscle cell cytokine secretion.

The antipsoriatic dimethylfumarate (DMF) has been anecdotically reported to reduce asthma symptoms and to improve quality of life of asthma patients. DMF decreases the expression of proinflammatory mediators by inhibiting the transcription factor NF-kappaB and might therefore be of interest for the therapy of inflammatory lung diseases. In this study, we determined the effect of DMF on platelet-derived growth factor (PDGF)-BB- and TNFalpha-induced asthma-relevant cytokines and NF-kappaB activation by primary human asthmatic and nonasthmatic airway smooth muscle cells (ASMC). Confluent nonasthmatic and asthmatic ASMC were incubated with DMF (0.1-100 microM) and/or dexamethasone (0.0001-0.1 microM), NF-kappaB p65 siRNA (100 nM), the NF-kappaB inhibitor helenalin (1 microM) before stimulation with PDGF-BB or TNFalpha (10 ng/ml). Cytokine release was measured by ELISA. NF-kappaB, mitogen and stress-activated kinase (MSK-1), and CREB activation was determined by immunoblotting and EMSA. TNFalpha-induced eotaxin, RANTES, and IL-6 as well as PDGF-BB-induced IL-6 expression was inhibited by DMF and by dexamethasone from asthmatic and nonasthmatic ASMC, but the combination of both drugs showed no glucocorticoid sparing effect in either of the two groups. NF-kappaB p65 siRNA and/or the NF-kappaB inhibitor helenalin reduced PDGF-BB- and TNFalpha-induced cytokine expression, suggesting the involvement of NF-kappaB signaling. DMF inhibited TNFalpha-induced NF-kappaB p65 phosphorylation, NF-kappaB nuclear entry, and NF-kappaB-DNA complex formation, whereas PDGF-BB appeared not to activate NF-kappaB within 60 min. Both stimuli induced the phosphorylation of MSK-1, NF-kappaB p65 at Ser276, and CREB, and all were inhibited by DMF. These data suggest that DMF downregulates cytokine secretion not only by inhibiting NF-kappaB but a wider range of NF-kappaB-linked signaling proteins, which may explain its potential beneficial effect in asthma.

Biological studies on Brazilian plants used in wound healing.

AIM OF THE STUDY: n-Hexanic and ethanolic extracts from twelve plants (Brugmansia suaveolens Brecht. et Presl., Eupatorium laevigatum Lam., Galinsoga parviflora Cav., Iresine herbstii Hook., Kalanchöe tubiflora Hamet-Ahti, Petiveria alliacea L., Pluchea sagittalis (Lam.) Cabrera, Piper regnellii DC., Schinus molle L., Sedum dendroideum Moç et Sessé ex DC., Waltheria douradinha St. Hill., Xanthium cavanillesii Schouw.) used in traditional South Brazilian medicine as wound healing agents were investigated in various biological assays, targeting different aspects in this complex process. MATERIALS AND METHODS: The extracts were investigated on NF-kappaB DNA binding, p38alpha MAPK, TNF-alpha release, direct elastase inhibition and its release as well as on caspase-3. Fibroblasts migration to and proliferation into the wounded monolayers were evaluated in the scratch assay, the agar diffusion test for antibacterial and the MTT assay for cytotoxic effects. RESULTS: The hydrophilic extracts from Galinsoga parviflora, Petiveria alliacea, Schinus molle, Waltheria douradinha and Xanthium cavanillesii as well as the lipophilic extract of Waltheria douradinha turned out to be the most active ones. CONCLUSIONS: These results increase our knowledge on the wound healing effects of the investigated medicinal plants. Further studies are necessary to find out the effective secondary metabolites responsible for the observed effects.

Microarray analysis reveals influence of the sesquiterpene lactone parthenolide on gene transcription profiles in human epithelial cells.

Sesquiterpene lactones are known for their anti-inflammatory activity which has been proven in various assays on DNA, mRNA and protein level. Here we report on the change in the gene expression profile in TNF-alpha stimulated human 293 cells after treatment with parthenolide using a cDNA microarray analysis. Twenty-one of 7028 genes were found to be up- and 18 down-regulated. They encode for chemoattractants, immune system proteins, glycoproteins, metabolism, serine proteinases, and transcription factors. Confirmatory analyses were carried out using quantitative real-time RT-PCR (TaqMan). Additional studies with selected genes revealed the concentration-dependent influence of parthenolide on the expression of these genes.

Primary mouse hepatocytes for systems biology approaches: a standardized in vitro system for modelling of signal transduction pathways.

Complex cellular networks regulate regeneration, detoxification and differentiation of hepatocytes. By combining experimental data with mathematical modelling, systems biology holds great promises to elucidate the key regulatory mechanisms involved and predict targets for efficient intervention. For the generation of high-quality quantitative data suitable for mathematical modelling a standardised in vitro system is essential. Therefore the authors developed standard operating procedures for the preparation and cultivation of primary mouse hepatocytes. To reliably monitor the dynamic induction of signalling pathways, the authors established starvation conditions and evaluated the extent of starvation-associated stress by quantifying several metabolic functions of cultured primary hepatocytes, namely activities of glutathione-S-transferase, glutamine synthetase, CYP3A as well as secretion of lactate and urea into the culture medium. Establishment of constant metabolic activities after an initial decrease compared with freshly isolated hepatocytes showed that the cultured hepatocytes achieve a new equilibrium state that was not affected by our starving conditions. To verify the highly reproducible dynamic activation of signalling pathways in the in vitro system, the authors examined the JAK-STAT, SMAD, PI3 kinase, MAP kinase, NF-kappaB and Wnt/beta-catenin signalling pathways. For the induction of gp130, JAK1 and STAT3 phosphorylation IL6 was used, whereas TGFbeta was applied to activate the phosphorylation of SMAD1, SMAD2 and SMAD3. Both Akt/PKB and ERK1/2 phosphorylation were stimulated by the addition of hepatocyte growth factor. The time-dependent induction of a pool of signalling competent beta-catenin was monitored in response to the inhibition of GSK3beta. To analyse whether phosphorylation is actually leading to transcriptional responses, luciferase reporter gene constructs driven by multiple copies of TGFbeta-responsive motives were applied, demonstrating a dose-dependent increase in luciferase activity. Moreover, the induction of apoptosis by the TNF-like cytokine Fas ligand was studied in the in vitro system. Thus, the mouse hepatocyte in vitro system provides an important basis for the generation of high-quality quantitative data under standardised cell culture conditions that is essential to elucidate critical hepatocellular functions by the systems biology approach.

Topic utilization of sesquiterpene lactone from Milleria quinqueflora on treatment of bothropic envenomation in rabbits

Estudou-se o efeito terapêutico da lactona sesquiterpênica (SL), 4,15-Epoxy-miller-9-Z-enolide, na lesão local do envenenamento botrópico experimental. Utilizaram-se três grupos de coelhos inoculados com 1.0æg de veneno de Bothrops alternatus e tratados com solução NaCl (0,85 por cento) (grupo I), SL diluída em glicerol (0,5 por cento) (grupo II) e SL diluída em vaselina (0,5 por cento) (grupo III). Todos os animais foram avaliados nos tempos 30min e 1, 2, 24, 30, 48, 54, 72, 96, 120 e 148h quanto ao grau de edema, diâmetro do halo hemorrágico e presença de necrose local. Os animais do grupo II apresentaram os menores valores de grau de edema e halo hemorrágico com desaparecimento em 54h. Apesar de a necrose ter ocorrido em todos os animais, o diâmetro também foi menor no grupo II, quando comparado com os outros grupos. A SL, extraída da Milleria quinqueflora, possui efeito antiinflamatório, que é importante no tratamento local do envenenamento botrópico.

Role of cysteine residues of p65/NF-kappaB on the inhibition by the sesquiterpene lactone parthenolide and N-ethyl maleimide, and on its transactivating potential.

Sesquiterpene lactones (SLs) are potent anti-inflammatory substances. It was previously shown that the anti-inflammatory effect could be partly explained by the inhibition of the transcription factor NF-kappaB. Whether they inhibit the DNA binding of NF-kappaB, the activation of the IkappaB-kinase, or both is still a matter of debate. The data supporting these hypotheses were obtained using different cell systems. In this contribution we analyzed the mechanism of the sesquiterpene lactone-mediated inhibition using different cell systems, and showed that in all the cell lines analyzed, SLs inhibited both NF-kappaB binding and the IkappaB-kinase, but that the former played a more preponderant role in the inhibition. These results again confirm the importance of cysteine 38 in the inhibition and regulation of NF-kappaB's function. Moreover, we compared the selectivity of the SL parthenolide with that of N-ethyl maleimide (NEM). We showed that NEM directly alkylated p65 as well as p50 of NF-kappaB, whereas SLs possess a selectivity towards p65. Finally, we studied the transactivating properties of various p65 mutants, to analyze the effect of exchanged cysteine residues in the DNA binding domain of NF-kappaB/p65 on its function and demonstrated that the transactivating potential of the mutants did not correlate with their DNA binding strenght.

Structure activity relationship studies of cinnamic acid derivatives as inhibitors of human neutrophil elastase revealed by ligand docking calculations.

Structure-activity relationship of cinnamic acid derivatives as inhibitors of the human neutrophil elastase is reported. Comparison of the inhibitory concentrations (IC50 values) with the results of the ligand docking calculations revealed that the structure element of the aromatic ortho-dihydroxy groups combined with a lipophilic residue seems to be a prerequisite for an optimal binding within the active site.

[Arnica: new insights on the molecular mode of action of a traditional medicinal plant]. / Arnika: Neue Erkenntnisse zum Wirkungsmechanismus einer traditionellen Heilpflanze.

Preparations from Arnica flowers have been used in traditional medicine since a long time for the treatment of inflammatory diseases. Sesquiterpene lactones are considered as their main active compounds. Previously, it was shown that these natural products attack inflammatory processes at a very central point by inhibiting the transcription factors NF-kappa B and NF-AT at micromolar concentrations. Both transcription factors regulate the transcription of genes encoding for many inflammatory mediators. Thus, these new insights on their molecular mode of action are an important contribution for a better understanding of the antiinflammatory activity of preparations from Arnica. First clinical studies show that they can support the treatment of rheumatic diseases. The agreed use is important to avoid undesirable side effects.

Sesquiterpene lactones as inhibitors of human neutrophil elastase.

Human neutrophil elastase (HNE) is a serine protease that has been implicated in the abnormal turnover of connective tissue proteins and has been described as an important pathogenic factor in several inflammatory diseases such as rheumatoid arthritis or cystic fibrosis. Here we investigated 17 sesquiterpene lactones (SLs) for their ability to inhibit human neutrophil elastase in an in vitro assay. Podachaenin was the most active compound with an IC(50) value of 7 microM. SLs do not covalently bind to the amino acids of the catalytic triad, thus differing from other elastase inhibitors with a lactone moiety. In contrast to most other biological activities of SLs HNE inhibition is not mediated by alpha,beta-unsaturated carbonyl functions. Ligand binding calculations using the X-ray structure of HNE and the program FlexX revealed structural elements which are a prerequisite for their inhibitory activity.

Inhibition of inflammatory cytokine production and lymphocyte proliferation by structurally different sesquiterpene lactones correlates with their effect on activation of NF-kappaB.

Many sesquiterpene lactones (Sls) are known to possess anti-inflammatory activities. To gain further insight into their structure-activity relationships and the molecular mechanism of action, four germacranolide sesquiterpene lactones which differ in the skeleton and the number of reactive centers (4beta,15-epoxy-miller-9E-enolide (1), 15-acetoxy-eremantholide B (2), a mixture of 15-(isovaleroyl)/15-(2-methyl-butyryl)-2alpha-acetoxy-miguanin (3), and 15-(2-hydroxy)-isobutyryloxy-micrantholide (4)) were investigated for their effect on production of proinflammatory cytokines (interleukin-1beta [IL-1beta], IL-6, and tumor necrosis factor-alpha [TNF-alpha]) as well as proliferation of concanavalin A (Con A) and lipopolysaccharide (LPS)-stimulated mouse lymphocytes. Compounds 1 and 3 which possess an alpha-methylene-gamma-lactone function and a conjugated carbonyl group induced a half-maximal inhibition of cytokine synthesis in adherent mouse peritoneal exudate cells at micromolar concentrations (IC(50) 0.69-1.70 microM), while compound 4 which contains only an alpha-methylene-gamma-lactone residue was less active (IC(50) > or 38 microM). Interestingly, compound 2, which carries only a conjugated keto group, displayed a potency similar to those of the bifunctional compounds 1 and 3. All four Sls suppressed proliferation of murine lymphocyte at IC(50) concentrations between 0.22 and 5.03 microM. The rank order of potency was 1 = 2 > 3 > 4. Generally, the growth of LPS-stimulated cells was more strongly influenced than those of Con A-activated lymphocytes. This effect was particularly pronounced with 4. Inhibitory concentrations correlated well with those necessary for inhibition of the transcription factor nuclear factor kappaB (NF-kappaB) observed in a previous investigation. Therefore, it can be assumed that NF-kappaB may be involved in the suppressive effect of Sls on cytokine production and lymphocyte proliferation.

Cysteine 38 in p65/NF-kappaB plays a crucial role in DNA binding inhibition by sesquiterpene lactones.

Sesquiterpene lactones (SLs) have potent anti-inflammatory properties. We have shown previously that they exert this effect in part by inhibiting activation of the transcription factor NF-kappaB, a central regulator of the immune response. We have proposed a molecular mechanism for this inhibition based on computer molecular modeling data. In this model, SLs directly alkylate the p65 subunit of NF-kappaB, thereby inhibiting DNA binding. Nevertheless, an experimental evidence for the proposed mechanism was lacking. Moreover, based on experiments using the SL parthenolide, an alternative mode of action has been proposed by other authors in which SLs inhibit IkappaB-alpha degradation. Here we report the construction of p65/NF-kappaB point mutants that lack the cysteine residues alkylated by SLs in our model. In contrast to wild type p65, DNA-binding of the Cys(38) --> Ser and Cys(38,120) --> Ser mutants is no longer inhibited by SLs. In addition, we provide evidence that parthenolide uses a similar mechanism to other SLs in inhibiting NF-kappaB. Contrary to previous reports, we show that parthenolide, like other SLs, inhibits NF-kappaB most probably by alkylating p65 at Cys(38). Although a slight inhibition of IkappaB degradation was detected for all SLs, the amount of remaining IkappaB was too low to explain the observed NF-kappaB inhibition.

3-Desoxyanthocyanidins from Arrabidaea chica.

The new 3-desoxyanthocyanidins 6,7,3'-trihydroxy-5,4'-dimethoxy-flavylium and 6,7,3',4'-tetrahydroxy-5-methoxy-flavylium and the known 6,7-dihydroxy-5,4'-dimethoxy-flavylium (Carajurin) were isolated by bioguided fractionation from the leaves of Arrabidaea chica, with transcription factor NF-kappaB as target. The structure of Carajurone was revised to be 6,7,4'-trihydroxy-5-methoxy-flavylium. Additionally, the flavone acacetin was found. All structures were mainly established on the basis of MS- and NMR data (1H, 1H-1H COSY and partly 13C, GHMQCR and GHSQCR). Carajurin, which failed to give a positive result in the DPPH TLC assay completely inhibited NF-kappaB, but not NF-AT at a 500 microM concentration.

Germacranolides from Mikania guaco.

Fourteen novel sesquiterpene lactones of the germacranolide type have been isolated from the aerial parts of Mikania guaco: six costunolide, two melampolide and six germacra-4-trans,10(14),11(13)-trien-12.6alpha-olide derivatives. Except for one compound all the others possess a carbonyl function at C-9. Eight were obtained in the form of four isomer pairs which were difficult to separate. Structure elucidation was based on mass and ID and 2D NMR measurements. Low energy conformations were obtained by quantum mechanical calculations. Pyrrolizidine alkaloids could not be detected.