Extraction and isolation of dictamnine, obacunone and fraxinellone from Dictamnus dasycarpus Turcz. by supercritical fluid extraction and high-speed counter-current chromatography

Supercritical fluid extraction was used to extract active compounds from the Chinese traditional medicinal D. dasycarpus under the pressure of 30 MPa and temperature of 45 ºC. Further separation and purification was established by high-speed counter-current chromatography (HSCCC) with a two-phase solvent system composed of n-hexane-ethyl acetate-methanol-water (1:0.8:1.3:0.9, volume ratio). The separation yielded a total of 47 mg of dictamnine, 24 mg of obacunone and 83 mg of fraxinellone from 1.0 g of the crude extract in one step separation with the purity of 99.2, 98.4 and 99.0%, respectively, as determined by HPLC. The chemical structures of these compounds were identified by ESI-MS, IR, ¹H-NMR and 13C-NMR.

Preparative separation and purification of bufadienolides from ChanSu by high-speed counter-current chromatography combined with preparative HPLC

Eight bufadienolides were successfully isolated and purified from ChanSu by high-speed counter-current chromatography (HSCCC) combined with preparative HPLC (prep-HPLC). First, a stepwise elution mode of HSCCC with the solvent system composed of petroleum ether - ethyl acetate - methanol - water (4:6:4:6, 4:6:5:5, v/v) was employed and four bufadienolides, two partially purified fractions were obtained from 200 mg of crude extract. The partially purified fractions III and VI were then further separated by prep-HPLC, respectively, and another four bufadienolides were recovered. Their structures were confirmed by ESI-MS and ¹H-NMR spectra.

Preparative separation of polyphenols from artichoke by polyamide column chromatography and high-speed counter-current chromatography

An efficient method for the rapid separation and purification of polyphenols from artichoke by polyamide column chromatography in combination with high-speed counter-current chromatography (HSCCC) was successfully built. The crude ethanol extracts from dry artichoke were first pre-separated by polyamide column chromatography and divided in two parts as sample 1 and sample 2. Then, the samples were further separated by HSCCC and yielded 7.8 mg of chlorogenic acid (compound I), 24.5 mg of luteolin-7-O-β-D-rutinoside (compound II), 18.4 mg of luteolin-7-O-β-D-glucoside (compound III), and 33.4 mg of cynarin (compound IV) with purity levels of 92.0%, 98.2%, 98.5%, and 98.0%, respectively, as determined by high-performance liquid chromatography (HPLC) method. The chemical structures of these compounds were identified by electrospray ionization-mass spectrometry (ESI-MS) and nuclear magnetic resonance (NMR).

Preparative separation of C19-diterpenoid alkaloids from Aconitum carmichaelii Debx by pH‑zone-refining counter-current chromatography

The technique of pH-zone-refining counter-current chromatography was successfully applied to preparatively separate three C19-diterpenoid alkaloids from the crude extracts of Aconitum carmichaelii for the first time using a two-phase solvent system of petroleum ether-ethyl acetate-methanol-water (5:5:1:9, v/v/v/v). Mesaconitine (I), hypaconitine (II), and deoxyaconitine (III) were obtained from 2.5 g of the crude alkaloids in a one-step separation; the yields were 4.16%, 16.96%, and 5.05%, respectively. The purities of compounds I, II, and III were 93.0%, 95%, and 96%, respectively, as determined by HPLC. The chemical structures of the three compounds were identified by electrospray ionization mass spectrometry (ESI-MS) and NMR.

Separation and purification of three stilbenes from the radix of Polygonum cillinerve (Nakai) Ohwl by macroporous resin column chromatography combined with high-speed counter-current chromatography

An effective method for the rapid separation and purification of three stilbenes from the radix of Polygonum cillinerve (Nakai) Ohwl by macroporous resin column chromatography combined with high-speed counter-current chromatography (HSCCC) was successfully established. In the present study, a two-phase solvent system composed of chloroform-n-butanol-methanol-water (4:1:4:2, v/v/v/v) was used for HSCCC separation. A one-step separation in 4 h from 150 mg of crude extract produced 26.3 mg of trans-resveratrol-3-O-glucoside, 42.0 mg of pieceid-2"-O-gallate, and 17.9 mg of trans-resveratrol with purities of 99.1%, 97.8%, and 99.4%, respectively, as determined by high-performance liquid chromatography (HPLC). The chemical structures of these compounds were identified by nuclear magnetic resonance (NMR) spectroscopy.

Noctuidae-induced plant volatiles: current situation and prospects

Noctuids are phytophagous lepidopterans with some species causing significant damage to agriculture. The host plants, in turn, have developed defense mechanisms to cope with them, for instance chemical defenses. In this study we review the literature on plant volatiles induced by noctuids, and discuss the methodologies used to induce the production of volatiles that are usually employed in plant defense mechanisms. Future prospects involving this line of research in pest control are also discussed.

Genetic linkage map of Phaseolus vulgaris and identification of QTLs responsible for resistance to Xanthomonas axonopodis pv. phaseoli

Common bean (Phaseolus vulgaris) cultivars with a high degree of resistance to Xanthomonas axonopodis pv. phaseoli (Xap) are not available in Brazil. Despite many studies, a low degree of resistance to Xap continues to exist due to its complex genetic inheritance, which is not well known. The objectives of this research were to complement a common bean genetic map based on the cross between a susceptible genotype 'HAB-52' and a resistant genotype 'BAC-6', and to map and analyze genomic regions (quantitative trait loci – QTLs) related to Xap resistance. Eleven linkage groups were determined using 143 RAPD markers, covering 1,234.5 cM of the genome. This map was used to detect QTLs associated with Xap resistance on leaves and pods. The averages of disease severity on leaves (represented by the transformed disease index – TDI) and pods (represented by the diameter of lesion on pods – DLP) were added to the data of the linkage map. Five TDI QTLs and only one LDP QTL were detected. The TDI QTLs were placed in the A, B, G and J linkage groups, with phenotypic variations ranging from 12.7 to 71.6%. The DLP QTL explained 12.9% of the phenotypic variation and was mapped in a distinct linkage group. These results indicate that there are different genes involved in the control of resistance on leaves and pods.

Dielectric spectroscopy and thermally stimulated discharge current in PEEK film

The complex permittivity of films of polyether ether ketone (PEEK) has been investigated over a wide range of frequency. There is no relaxation peak in the range of 1Hz to 10(5) Hz but in the low-frequency side (10-4 Hz) there is an evidence of a peak that also can be observed by thermally stimulated discharge current measurements. That peak is related with the glass transition temperature (Tg) of the polymer. The activation energy of the relaxation was found to be 0.44 eV, similar to that of several synthetic polymers. Space charges are important in the conduction mechanism as shown by discharging transient.

Etiology and epidemiology of Pythium root rot in hydroponic crops: current knowledge and perspectives

The etiology and epidemiology of Pythium root rot in hydroponically-grown crops are reviewed with emphasis on knowledge and concepts considered important for managing the disease in commercial greenhouses. Pythium root rot continually threatens the productivity of numerous kinds of crops in hydroponic systems around the world including cucumber, tomato, sweet pepper, spinach, lettuce, nasturtium, arugula, rose, and chrysanthemum. Principal causal agents include Pythium aphanidermatum, Pythium dissotocum, members of Pythium group F, and Pythium ultimum var. ultimum. Perspectives are given of sources of initial inoculum of Pythium spp. in hydroponic systems, of infection and colonization of roots by the pathogens, symptom development and inoculum production in host roots, and inoculum dispersal in nutrient solutions. Recent findings that a specific elicitor produced by P. aphanidermatum may trigger necrosis (browning) of the roots and the transition from biotrophic to necrotrophic infection are considered. Effects on root rot epidemics of host factors (disease susceptibility, phenological growth stage, root exudates and phenolic substances), the root environment (rooting media, concentrations of dissolved oxygen and phenolic substances in the nutrient solution, microbial communities and temperature) and human interferences (cropping practices and control measures) are reviewed. Recent findings on predisposition of roots to Pythium attack by environmental stress factors are highlighted. The commonly minor impact on epidemics of measures to disinfest nutrient solution as it recirculates outside the crop is contrasted with the impact of treatments that suppress Pythium in the roots and root zone of the crop. New discoveries that infection of roots by P. aphanidermatum markedly slows the increase in leaf area and whole-plant carbon gain without significant effect on the efficiency of photosynthesis per unit area of leaf are noted. The platform of knowledge and understanding of the etiology and epidemiology of root rot, and its effects on the physiology of the whole plant, are discussed in relation to new research directions and development of better practices to manage the disease in hydroponic crops. Focus is on methods and technologies for tracking Pythium and root rot, and on developing, integrating, and optimizing treatments to suppress the pathogen in the root zone and progress of root rot.

Laparoscopy in uterine cervical cancer. Current state and literature review

Cervical cancer remains the most frequent gynecological tumor in Brazil and other developing countries. Minimally invasive techniques, especially laparoscopy, have been increasingly employed in such tumors. This article aims to describe the main applications of laparoscopy in the treatment and staging of cervical cancer. In the early stages, it is possible to provide a fertility-preserving surgery in the form of radical trachelectomy and, in a study protocol, the function-preserving surgery, avoiding parametrectomy and the associated morbidity. A fully laparoscopic radical hysterectomy is fairly standard in the literature and has the tendency to become the standard of care in early cases, for patients who want to bear no more children. In advanced stages, minimally invasive surgery can offer ovarian transposition, with intent to prevent actinic castration, without upsetting the time for the start of radiotherapy and chemotherapy. Staging laparoscopic surgery, including pelvic and para-aortic lymphadenectomy, has been the subject of studies, since it has the potential to modify the extension of radiotherapy depending on the extent of lymph node spread.

Welding current effect on diffusible hydrogen content in flux cored arc weld metal

The application of flux cored arc welding (FCAW) has increased in manufacturing and fabrication. Even though FCAW is well known for its good capability in producing quality welds, few reports have been published on the cause of the relatively high diffusible hydrogen content in the weld metal and its relation with the ingredients used in the wire production and with the welding parameters (mainly welding current). This paper describes experiments where data obtained from weld metal diffusible hydrogen analysis, metal droplet collection, and high-speed recording of metal droplet transfer were used to evaluate the effect of welding current on diffusible hydrogen content in the weld metal. The results from gas chromatography analysis showed that weld metal hydrogen content indeed increased with welding current. A polynomial regressional analysis concluded that hydrogen increase with current was better described by a linear function with proportional constant of approximately 0.7 or 70%. Different from the GMA welding transfer behavior, statistical analysis showed only a small increase in metal droplet size with increasing current. The metal transfer mode remained in the globular range for currents between 100 and 150 A. The most surprising findings were with the high-speed cinematography recording. Observing the high speed movies, it was possible to see that at low current, "unmelted" flux sporadically touched the weld pool but at higher current, the flux remained touching the weld pool during the whole time of droplet formation and transfer. It is believed that since the flux has ingredients that contain hydrogen, hydrogen passes through the arc undisturbed, going to the weld bead intact and increasing the hydrogen content in the weld metal. Another important observation is regarding to droplet size. Droplet size increased with increasing current because forces from decomposed gases from the flux could sustain the droplets, retarding their transfer and allowing them to grow.

Comparative analysis of leaf cell-wall polysaccharides of Dialypetalanthus fuscescens and Bathysa meridionalis: evidence of biochemical similarities between Dialypetalanthaceae and Rubiaceae-Cinchonoideae

Dialypetalanthus fuscescens is an Amazonian endemic species with problematic taxonomic position. This neotropical rainforest tree belongs to the monospecific Dialypetalanthaceae. In the present work, we analysed the leaf cell-wall polysaccharide composition of Dialypetalanthus fuscescens and compared it to that of Bathysa meridionalis (Rubiaceae-Cinchonoideae). Glycosyl composition and glycosyl-linkage analysis indicated that both species have similar cell wall composition. Arabinogalactans were the major component of the pectic polysaccharides and xylans, although being reported in minor amounts in dicots, were found to be the predominant hemicellulosic polysaccharide in cell walls of both species. These findings are in agreement with previous data on cell wall composition reported for Rubiaceae and corroborate the current suggestion of the possible link between this family and Dialypetalanthaceae.

Outward potassium current oscillations in macrophage polykaryons: extracellular calcium entry and calcium-induced calcium release

Outward current oscillations associated with transient membrane hyperpolarizations were induced in murine macrophage polykaryons by membrane depolarization in the absence of external Na+. Oscillations corresponded to a cyclic activation of Ca2+-dependent K+ currents (IKCa) probably correlated with variations in intracellular Ca2+ concentration. Addition of external Na+ (8 mM) immediately abolished the outward current oscillations, suggesting that the absence of the cation is necessary not only for their induction but also for their maintenance. Oscillations were completely blocked by nisoldipine. Ruthenium red and ryanodine reduced the number of outward current cycles in each episode, whereas quercetin prolonged the hyperpolarization 2- to 15-fold. Neither low molecular weight heparin nor the absence of a Na+ gradient across the membrane had any influence on oscillations. The evidence suggests that Ca2+ entry through a pathway sensitive to Ca2+ channel blockers is elicited by membrane depolarization in Na+-free medium and is essential to initiate oscillations, which are also dependent on the cyclic release of Ca2+ from intracellular Ca2+-sensitive stores; Ca2+ ATPase acts by reducing intracellular Ca2+, thus allowing slow deactivation of IKCa. Evidence is presented that neither a Na+/Ca2+ antiporter nor Ca2+ release from IP3-sensitive Ca2+ stores participate directly in the mechanism of oscillation

Absence of linkage between MHC and a gene involved in susceptibility to human schistosomiasis

Six hundred million people are at risk of infection by Schistosoma mansoni. MHC haplotypes have been reported to segregate with susceptibility to schistosomiasis in murine models. In humans, a major gene related to susceptibility/resistance to infection by S. mansoni (SM1) and displaying the mean fecal egg count as phenotype was detected by segregation analysis. This gene displayed a codominant mode of inheritance with an estimated frequency of 0.20-0.25 for the deleterious allele and accounted for more than 50% of the variance of infection levels. To determine if the SM1 gene segregates with the human MHC chromosomal region, we performed a linkage study by the lod score method. We typed for HLA-A, B, C, DR and DQ antigens in 11 informative families from an endemic area for schistosomiasis in Bahia, Brazil, by the microlymphocytotoxicity technique. HLA-DR typing by the polymerase chain reaction with sequence-specific primers (PCR-SSP) and HLA-DQ were confirmed by PCR-sequence-specific oligonucleotide probes (PCR-SSOP). The lod scores for the different q values obtained clearly indicate that there is no physical linkage between HLA and SM1 genes. Thus, susceptibility or resistance to schistosomiasis, as defined by mean fecal egg count, is not primarily dependent on the host's HLA profile. However, if the HLA molecule plays an important role in specific immune responses to S. mansoni, this may involve the development of the different clinical aspects of the disease such as granuloma formation and development of hepatosplenomegaly.

Impaired beta-adrenergic response and decreased L-type calcium current of hypertrophied left ventricular myocytes in postinfarction heart failure

Infarct-induced heart failure is usually associated with cardiac hypertrophy and decreased ß-adrenergic responsiveness. However, conflicting results have been reported concerning the density of L-type calcium current (I Ca(L)), and the mechanisms underlying the decreased ß-adrenergic inotropic response. We determined I Ca(L) density, cytoplasmic calcium ([Ca2+]i) transients, and the effects of ß-adrenergic stimulation (isoproterenol) in a model of postinfarction heart failure in rats. Left ventricular myocytes were obtained by enzymatic digestion 8-10 weeks after infarction. Electrophysiological recordings were obtained using the patch-clamp technique. [Ca2+]i transients were investigated via fura-2 fluorescence. ß-Adrenergic receptor density was determined by [³H]-dihydroalprenolol binding to left ventricle homogenates. Postinfarction myocytes showed a significant 25% reduction in mean I Ca(L) density (5.7 ± 0.28 vs 7.6 ± 0.32 pA/pF) and a 19% reduction in mean peak [Ca2+]i transients (0.13 ± 0.007 vs 0.16 ± 0.009) compared to sham myocytes. The isoproterenol-stimulated increase in I Ca(L) was significantly smaller in postinfarction myocytes (Emax: 63.6 ± 4.3 vs 123.3 ± 0.9% in sham myocytes), but EC50 was not altered. The isoproterenol-stimulated peak amplitude of [Ca2+]i transients was also blunted in postinfarction myocytes. Adenylate cyclase activation through forskolin produced similar I Ca(L) increases in both groups. ß-Adrenergic receptor density was significantly reduced in homogenates from infarcted hearts (Bmax: 93.89 ± 20.22 vs 271.5 ± 31.43 fmol/mg protein in sham myocytes), while Kd values were similar. We conclude that postinfarction myocytes from large infarcts display reduced I Ca(L) density and peak [Ca2+]i transients. The response to ß-adrenergic stimulation was also reduced and was probably related to ß-adrenergic receptor down-regulation and not to changes in adenylate cyclase activity.