Analytical techniques to recognize inclusion complexes formation involving monoterpenes and cyclodextrins: A study case with (-) borneol, a food ingredient.

Monoterpenes are non-polar secondary metabolites widely used by industry due to their excellent therapeutic, food-ingredient and cosmetic properties. However, their low solubility in water limits their use. In this sense, cyclodextrins (CDs) have been widely used to solve these technological challenges. Thus, this study aims to use (-)-borneol as a monoterpene model to prepare inclusion complexes between ß-CD and hydroxypropyl-ß-CD (HP-ß-CD) through different ways and characterize them in order to choose the best inclusion method to improve physicochemical properties of monoterpenes. To achieve this goal, the samples were prepared by physical mixture (PM), paste complex (PA) and freeze-drying complex (FD) and then, extensively characterized by thermal analysis, Fourier-transform infrared spectroscopy, scanning electron microscopy, size particle, X-ray diffraction and nuclear magnetic resonance. The physicochemical results showed that freeze-drying was more effective to form inclusion complexes between (-)-borneol with both CDs. This research highlights the importance of recognizing the best method to prepare inclusion complexes, including food additives as (-)-borneol, to achieve better results in food preparations.

Chemical and mechanical changes during leaf expansion of four woody species of dry Restinga woodland.

Young leaves are preferential targets for herbivores, and plants have developed different strategies to protect them. This study aimed to evaluate different leaf attributes of presumed relevance in protection against herbivory in four woody species (Erythroxylum argentinum, Lithrea brasiliensis, Myrciaria cuspidata, and Myrsine umbellata), growing in a dry restinga woodland in southern Brazil. Evaluation of leaf parameters was made through single-point sampling of leaves (leaf mass per area and leaf contents of nitrogen, carbon, and pigments) at three developmental stages and through time-course sampling of expanding leaves (area and strength). Leaves of M. umbellata showed the highest leaf mass per area (LMA), the largest area, and the longest expansion period. On the other extreme, Myrc. cuspidata had the smallest LMA and leaf size, and the shortest expansion period. Similarly to L. brasiliensis, it displayed red young leaves. None of the species showed delayed-greening, which might be related to the high-irradiance growth conditions. Nitrogen contents reduced with leaf maturity and reached the highest values in the young leaves of E. argentinum and Myrc. cuspidata and the lowest in M. umbellata. Each species seems to present a different set of protective attributes during leaf expansion. Myrciaria cuspidata appears to rely mostly on chemical defences to protect its soft leaves, and anthocyanins might play this role at leaf youth, while M. umbellata seems to invest more on mechanical defences, even at early stages of leaf growth, as well as on a low allocation of nitrogen to the leaves. The other species display intermediate characteristics.

Analgesic and anti-inflammatory activity of a crude root extract of Pfaffia glomerata (Spreng) Pedersen.

The anti-inflammatory and antinociceptive effects of the crude hydroalcoholic extract (PE) of Pfaffia glomerata roots was assessed in the carrageenan-induced rat paw edema at the doses of 100, 200 and 300 mg/kg, using different animal models. An anti-inflammatory dose effect response correlation of r=0.997 and Y=11.67x+0.02 was found. At the same doses, the extract-inhibited acetic acid-induced writhing in mice, but no dose response correlation was found. Oral administration of 100 mg/kg of PE and 0.5 mg/kg of dexamethazone inhibited by 29 and 61%, the granulomatous tissue formation (p>0.05), respectively. These results indicate the potential of this plant extract to treat chronic inflammation. At the assayed doses no significant activity was found in the hot plate test, as well as in the cell migration-induced by carrageenan.

Control of resin production in Araucaria angustifolia, an ancient South American conifer.

Araucaria angustifolia is an ancient slow-growing conifer that characterises parts of the Southern Atlantic Forest biome, currently listed as a critically endangered species. The species also produces bark resin, although the factors controlling its resinosis are largely unknown. To better understand this defence-related process, we examined the resin exudation response of A. angustifolia upon treatment with well-known chemical stimulators used in fast-growing conifers producing both bark and wood resin, such as Pinus elliottii. The initial hypothesis was that A. angustifolia would display significant differences in the regulation of resinosis. The effect of Ethrel(®) (ET - ethylene precursor), salicylic acid (SA), jasmonic acid (JA), sulphuric acid (SuA) and sodium nitroprusside (SNP - nitric oxide donor) on resin yield and composition in young plants of A. angustifolia was examined. In at least one of the concentrations tested, and frequently in more than one, an aqueous glycerol solution applied on fresh wound sites of the stem with one or more of the adjuvants examined promoted an increase in resin yield, as well as monoterpene concentration (α-pinene, ß-pinene, camphene and limonene). Higher yields and longer exudation periods were observed with JA and ET, another feature shared with Pinus resinosis. The results suggest that resinosis control is similar in Araucaria and Pinus. In addition, A. angustifolia resin may be a relevant source of valuable terpene chemicals, whose production may be increased by using stimulating pastes containing the identified adjuvants.

Distinct effects of auxin and light on adventitious root development in Eucalyptus saligna and Eucalyptus globulus.

Adventitious rooting is essential for vegetative propagation of woody species. We studied the effects of auxin and light on the development of adventitious roots in cuttings obtained from seedlings of Eucalyptus saligna Smith and E. globulus Labill in an attempt to characterize the adventitious rooting process and identify factors controlling rhizogenesis. Root development was scored as rooting percentage, root density (roots per rooted cutting), mean rooting time and root length. In both species, rooting time was reduced in the presence of auxin. Cuttings from 2-month-old E. saligna seedlings were responsive to lower auxin concentrations than comparable cuttings from E. globulus seedlings. Cuttings from 3-month-old E. saligna seedlings rooted promptly and rooting was not significantly affected by light conditions. In contrast, rooting of cuttings from 3-month-old E. globulus seedlings exhibited recalcitrant behavior and no roots were formed if illuminated during the root formation phase. Effective root regeneration of E. globulus cuttings was obtained by a 4-day exposure to 10 mg l(-1) IBA and culture in darkness during the root formation step. Loss of rooting capacity with seedling age was more pronounced in E. globulus than in E. saligna. The possibility of switching adventitious rooting off and on by manipulating light regime and exogenous auxin supply in E. globulus, and the constitutive nature of rooting in E. saligna may provide useful models for examining the rooting process at the biochemical and molecular levels in Eucalyptus.

Cell culture of Taxus as a source of the antineoplastic drug taxol and related taxanes.

Callus cultures of Taxus cuspidata and Taxus canadensis were induced using different tissue explants including green and red arils, seed contents, young stems and needles. Callus derived from stem segments displayed the best growth in defined media. The culture medium was supplemented with reducing agents and phenolic-binding compounds to inhibit callus darkening and subsequent growth reduction. T. cuspidata explant growth was affected by different concentrations and ratios of 2,4-D and kinetin. Callus tissues of T. cuspidata were extracted for taxol after 2 months in culture and analysed by HPLC. The presence of taxol (0.020 +/- 0.005% of the extracted dry weight) was indicated based on retention time, U.V. spectra, peak purity as assessed by photo-diode array spectroscopy and compared with an authentic taxol standard, as well as by 1H-NMR analysis. Suspension cultures of T. cuspidata were established from the callus cultures, subsequently immobilized onto glass fiber mats, and maintained as immobilized cultures for 6 months. The immobilized cell cultures also produced taxol at levels up to 0.012 +/- 0.007% of the extracted dry weight.

Improved growth and taxol yield in developing calli of Taxus cuspidata by medium composition modification.

Cell culture of Taxus spp. represents a potential alternative source of taxol and related taxanes used in cancer chemotherapy. We have analyzed the effect of different culture media components on growth and production of taxol in developing callus cultures of T. cuspidata. Several sequential modifications were made to the basal B5 medium, which included addition and/or variation in the concentration of sucrose, B5 organic supplements, gibberellic acid, 36 combinations of 2,4-D/kinetin ratios, media salts and organic supplements, phenylalanine, casein hydrolysate and medium pH. The experiments were conducted during a 55 day-growth period followed by taxane extraction and analysis. Significant increases in taxol yield and growth over basal medium grown calli were observed with some of the modified media.

Distinct modes of adventitious rooting in Arabidopsis thaliana.

The literature describes different rooting protocols for Arabidopsis thaliana as models to study adventitious rooting, and results are generally perceived as comparable. However, there is a lack of investigations focusing on the distinct features, advantages and limitations of each method in the study of adventitious rooting with both wild-type (WT) ecotypes and their respective mutants. This investigation was undertaken to evaluate the adventitious rooting process in three different experimental systems, all using A. thaliana, analysing the same rooting parameters after transient exposure to auxin (indole-3-acetic acid) and control conditions: excised leaves, de-rooted plants and etiolated seedlings. The founding tissues and sites of origin of roots differed depending on the system used, whereas all rooting patterns were of the direct type (i.e., without callus formation). None of the systems had an absolute requirement for exogenous auxin, although rooting was enhanced by this phytohormone, with the exception of de-rooted plants, which had adventitious rooting strongly inhibited by exogenous auxin. Root elongation was much favoured in isolated leaves. Auxin-overproducing mutants could not be used in the detached leaf system due to precocious senescence; in the de-rooted plant system, these mutants had a WT-like rooting response, whereas the expression of the 'rooty' phenotype was only evident in the etiolated seedling system. Adventitious rooting of etiolated WT seedlings in the presence of exogenous auxin was inhibited by exogenous flavonoids, which act as auxin transport inhibitors; surprisingly, the flavonoid-deficient mutant chs had a lower rooting response compared to WT. Although Arabidopsis is an excellent model system to study adventitious rooting, physiological and developmental responses differed significantly, underlining the importance of avoiding data generalisation on rooting responses derived from different experimental systems with this species.

Immunoadjuvant and anti-inflammatory plant saponins: characteristics and biotechnological approaches towards sustainable production.

Saponins can be classified as triterpenoid (C30) or steroidal (C27), based on their carbon nucleus (aglycone). Sugar residues are linked to the aglycone, conferring an amphiphilic nature on these molecules, which is relevant for their biological activities. Saponins include a large variety of molecules that find several applications in pharmacology. Saponins have been shown to display immunoadjuvant, anti-inflammatory, antiplatelet, hypocholesterolemic, antitumoral, anti-HIV, antibacterial, insecticide, fungicide and anti-leishmanial activities. Anti-inflammatory medicines are increasingly demanded to treat various forms of arthritis in aging and obese populations and to help reduce the doses and duration of conventional corticotherapy with less side effects and without immunosuppression. The vaccine market for both human and veterinary uses is close to US$ 15 billion, progressively inflated by the recurrent threat of global pandemics.This paper provides an overview of recent advances (main focus on the last five years) on plant saponins that show anti-inflammatory and/or immunoadjuvant activities: source plants, isolation procedures, mechanism of action and biotechnological approaches towards sustainable production of bioactive saponins. Special attention is given to ginseng and Quillaja saponins. Strategies based on plant cultivation, cell and tissue culture, elicitation, and metabolic engineering for improved production of saponins are described. Future directions for research in the field and strategies to overcome bottlenecks are also discussed.

Psychollatine, a glucosidic monoterpene indole alkaloid from Psychotria umbellata.

A monoterpene indole alkaloid, psychollatine ( 1), was isolated from Psychotria umbellata leaves. Its structure was characterized by interpretation of spectroscopic data and by comparison of its NMR data with those of croceaine A ( 2) from Palicourea crocea. The configuration of psychollatine ( 1) was established by NOE difference and circular dichroism (CD) techniques, while its conformation was evaluated through molecular modeling studies and NMR coupling constants.

Distribution and amounts of taxol in different shoot parts of Taxus cuspidata.

Different fresh shoot parts of male and female plants of Taxus cuspidata were extracted and analysed for taxol concentration by high performance liquid chromatography (HPLC). Extracted parts included: young needles (first 10 top needle pairs of 30 cm long branches), old needles (last 10 needle pairs of 30 cm long branches), green bark, dark bark (with intense secondary growth), young wood (originally surrounded by green bark), wood (originally surrounded by dark bark), young stems (surrounded by green bark and devoid of needles), and mature male cones. Dichloromethane extracts were analysed by HPLC and diode array spectroscopy. Taxol identification was done by retention time, U.V. spectra, and spiking with an authentic taxol standard; 1H-NMR analysis was done for needle extracts. All parts except male cones had measurable amounts of taxol; no effect of plant sex on taxol levels of the plant parts analysed was observed. Results indicated that the bark accounted for almost all the taxol present in stems devoid of needles. Needles showed the highest levels of taxol (overall average of 0.035 +/- 0.006% of the extracted dry weight), significantly higher than those displayed by dark bark samples (0.012 +/- 0.001% of the extracted dry weight). Different needle post-harvesting procedures were evaluated in relation to taxol yields, 96 h dark incubation at -12 degrees C and 96 h dark incubation at 25 degrees C under vacuum gave taxol yields equivalent to those of freshly extracted samples. However, results obtained for 96 h dark incubation at 60 degrees C indicated some extent of taxol degradation.

Sinonasal tract seromucous adenocarcinomas: a report of 12 cases.

Sinonasal seromucous adenocarcinomas may originate from the surface epithelium or from the submucosal glands. We reviewed the clinicopathologic material from 12 patients with sinonasal tract seromucous adenocarcinomas at the University of Texas M. D. Anderson Cancer Center (Houston, TX). There were nine men and three women age 30 to 87 years (mean age, 56.3 years). The clinical presentation included nasal obstruction, nasal mass, and epistaxis. Eight tumors were located in the nasal cavity, three in the ethmoidal sinuses, and one involved the nasal cavity and ethmoid. Histologically, in nine cases the neoplastic glands were lined by a single cell type, arranged back to back without intervening stroma and often inducing desmoplastic reaction. The remaining three tumors also had a cribriform and papillary pattern. All patients were treated by surgical resection. Three patients had recurrences, which occurred at 36, 36, and 48 months after initial therapy. Their treatment involved surgery and irradiation. Eleven patients are alive and free of disease at 36 to 108 months after diagnosis. One patient died 48 months after diagnosis of another cause. Sinonasal tract seromucous adenocarcinomas arise purely from submucosal seromucous glands. The diagnosis is facilitated by their anatomic location, the absence of tumor within the mucosal surface epithelium, and the striking similarity to terminal tubules of the seromucous glands.

Kinetics of taxol production, growth, and nutrient uptake in cell suspensions of Taxus cuspidata.

Cell culture of Taxus cuspidata may represent an alternative to extraction of bark as a source of taxol and related taxanes. Cell suspensions of a cell line of T. cuspidata were grown for 44 days in shake flasks containing B5C2 medium. Throughout the growth cycle, fresh and dry weight accumulation, taxol yield on a dry weight basis, taxol accumulation in the medium, pH and pigmentation variation in the medium, as well as the uptake of sucrose, glucose, fructose, nitrate, and inorganic phosphate from the culture medium were examined. The results showed that the growth was relatively slow (doubling times of 17 and 20 days for fresh and dry weight, respectively), and taxol accumulation in the cells was non-growth related (higher in the stationary phase) and at relatively low levels (up to 4 mug/g of the extracted dry weight). Taxol concentration in the medium had two peaks: one during the early (0.4mug/mL) and another during the late (0.1-mug/mL) parts of the growth cycle. On a volumetric basis, the average total amount of taxol produced during the stationary phase (day 38) was 0.15 mug/mL, of which approximately 66% was in the medium and 34% was in the cells. Total carbohydrate uptake was closely associated with the increase in dry biomass. Sucrose was apparently extracellularly hydrolyzed after the first 6 days of culture; glucose was used before fructose. Nitrate was assimilated throughout the growth cycle, but phosphate was absorbed within the first week of culture. The pH variation showed an initial drop followed by a trend toward alkalinization for most of the growth period. Dark pigmentation in the medium increased progressively, particularly during the stationary phase. (c) 1994 John Wiley & Sons, Inc.

Evaluation of the trypanocidal and leishmanicidal in vitro activity of the crude hydroalcoholic extract of Pfaffia glomerata (Amarathanceae) roots.

Three different concentrations (1, 10 and 50 microg/ml) of lyophilized hydroalcoholic crude extract of Pfaffia glomerata roots were assayed in vitro against strains of Trypanosoma cruzi (Y) and Leishmania braziliensis. It was observed that P. glomerata hydroalcoholic extract was relatively active within the tested concentrations for L. (V) braziliensis, but inactive against T. cruzi. Despite the fact that both protozoans belong to the Trypanosomatidae family, we suggest that the difference observed for activity should be related to the biological differences between the two parasite species.