Ontogeny and structure of the pericarp and the seed coat of Miconia albicans (Sw.) Triana (Melastomataceae) from " cerrado" , Brazil

Miconia albicans fruit and seed coat ontogeny were described under light microscope. The samples were fixed in formalin-aceto-alcohol (FAA), neutral-buffered formaldehyde solution (NBF) and formalin-ferrous sulphate (FFS) solutions, embedded in plastic resin, sectioned at 10 µm and stained with Toluidine Blue. Specific dyes and/or reagents were used for the microchemical tests. The ovary is semi-inferior and the indehiscent, fleshy globose berries are originated mainly from the development of the inferior portion of the ovary. The immature pericarp is mainly parenchymatous with some sclereids, druse crystal and phenolic-like compounds idioblasts widespread in the mesocarp. In the mature pericarp, the endocarp cells are often collapsed, the mesocarp is thick with cells more or less turgid, and the sclereids, the druses and the phenolic-like compound idioblasts are almost absent. The ovules are anatropous, bitegmic and crassinucellate, and the zig-zag micropyle is formed by both the exostome and the endostome. The mature seed is pyramidal-elongated in shape, exalbuminous and testal. The raphal part occupies about 40% of the seed coat total length and had the mechanical layer derived from its inner layer. The antiraphal side is non-multiplicative and the exotesta, mesotesta and endotesta are differentiated into a sclerotic layer, with the exotesta being the mechanical one. The tegmen is absent.

Polyphenolic cells and their interrelation with cotyledon cells in seven species of Theobroma (Sterculiaceae)

The amount of cotyledon polyphenolic cells varies extensively within the Theobroma species. The polyphenolic compounds of these cells play a protective role and furthermore have an important function in the development of chocolate flavour. The morphology of the polyphenolic cells of the mesophyll is described and the development of these idioblasts in Theobroma cacao L., T. subincanum Mart., T. obovatum Klotzsch ex Bernoulli, T. grandiflorum (Willd. ex Spreng.) K. Schum., T. microcarpum Mart., T. bicolor Bonpl. and T. speciosum Willd. ex Spreng analysed. The total polyphenolic content in the seeds as determined by spectrophotometry showed a variation of about forty times. The alive, transparent polyphenolic cells are scattered throughout the cotiledonary mesophyll. However the polyphenolic cells of T. cacao and T. grandiflorum are also aligned perpendicularly with respect to the mesophyll borders and, in addition, both species display polyphenolic cells with a natural translucent purple colour. All the species analysed contained polyphenolic cells scattered throughout the parenchymal cells and also in a lengthwise association with vascular bundles. In T. bicolor and T. speciosum, the species with the lowest polyphenolic contents, these cells were mostly located around the vascular bundles. Using Scanning Electron Microscopy, the polyphenolic cells demonstrated a complex cytoarchitecture, and after fixing with glutaraldhyde, the polyphenolic secretion was shown to remain as a single unit or was organized into round droplets. Transmission Electron Microscopy displayed immature plastids from young mesophyll cells containing eletron-dense deposits similar to phenolic substances, suggesting that Theobroma plastids are involved in the synthesis of phenolics.

Morphology and histochemistry of glandular trichomes of Cordia verbenacea DC. (Boraginaceae) leaves

The objective of this work was to study, using light and fluorescence microscopy and scanning electron microscopy, the morphology and secretory products of glandular trichomes of Cordia verbenacea DC. (Boraginaceae), known as 'baleeira', a species used in folk medicine as anti-inflammatory, analgesic, anti-ulcerogenic and healing agent. Two classes of glandular trichomes were recognized, globular and reniform. A morphological study of the secretory head and the characterization of the secretory product are also presented. Secretory products of globular trichomes consisted of essential oils, whereas reniform trichomes consisted basically of phenolic compounds such as flavonoids. No pre-established regions for releasing secretory products were found.

Anatomy, ultrastructure and secretion of Hibiscus pernambucensis Arruda (Malvaceae) extrafloral nectary

This paper reports on the extrafloral nectary (EFN) of Hibiscus pernambucensis, a native shrub species occurring in mangrove and restinga along Brazil's coastline. EFNs occur as furrows with a protuberant border on the abaxial surface veins of the leaf blade. Each nectary consists of numerous secretory multicellular trichomes, epidermal cells in palisade-like arrangements and non-vascularized parenchyma tissue. Nectar secretion is prolonged, since secretion starts in very young leaves and remains up to completely expanded leaves. Reduced sugars, lipids, and proteins were histochemically detected in all the nectary cells; phenolic substances were detected in the vacuoles of the epidermal palisade cells and in some secretory trichome cells. The secretory cells that constitute the body of trichomes have large nuclei, dense cytoplasm with numerous mitochondria, dictyosomes, scattered lipid droplets and plastids with different inclusions: protein, lipid droplets or starch grains; vacuoles with different sizes have membranous material, phenolic and lipophilic substances. The palisade cells show thick periclinal walls, reduced cytoplasm with voluminous lipid drops and developed vacuoles. The nectary parenchyma cells contain abundant plasmodesmata and cytoplasm with scattered lipid droplets, mitochondria, plastids with starch grains and endoplasmic reticulum. Mucilage idioblasts are common in the inner nectary parenchyma. Protoderm and ground meristem participate in the formation of EFN. Our data indicate that all nectary regions are involved in nectar production and secretion, constituting a functional unit. Longevity of the extrafloral nectaries is likely associated with the presence of mucilage idioblasts, which increases the capacity of the nectary parenchyma to store water.

Multiple dormancy and maternal effect on Miconia ferruginata (Melastomataceae) seed germination, Serra de Caldas Novas, Goiás, Brazil

Species with mostly asexual reproduction are interesting subjects for germination studies since variation would be more easily linked to environmental factors. Miconia ferruginata DC. is an apomictic treelet in Brazilian cerrado areas on rocky outcrops. Germination of seeds collected from individuals occurring in the Serra de Caldas Novas State Park, Goiás, was studied in three experiments under controlled conditions. Germination characteristics differed among individuals and were correlated with altitude and soil Al content. Seeds from plants growing at lower altitudes, with lower soil aluminium content, presented malformed seeds with absence of embryo which rendered lower, but better synchronized germination. The nested analysis showed that from the total variance, 78.14% for germinability, 54.56% for uncertainty of the germination process, and 68.30% for the quantity of seeds without embryo was attributed to the altitudinal effect. Individuals nested within altitude contributed up to 16.93% for the total variance. It means that there is low variability among individuals of the same altitude and high variability among individuals from different points of the slope, making clear that for the studied population the environmental effect is stronger than the genetic component to determine the seed quality. The testa of the seeds provides a mechanical dormancy which seems to be associated also with phenolic compounds, which help to disperse germination through time. Photoblastism was also registered for seeds of this species.

Leaf and calycine colleters in Odontadenia lutea (Apocynaceae - Apocynoideae - Odontadenieae): their structure and histochemistry

The structure and histochemistry of colleters found on the vegetative and floral apices of Odontadenia lutea are described. Colleters occur on vegetative apices starting at the fourth node, with 68 to 80 colleters being found at each node. Each leaf primordium has only one colleter of axillary origin, 3-5 intra-petiolar, and 12-16 inter-petiolar (intra-stipular). There are four types of colleters: standard, bipartite standard, sessile, and bipartite sessile. Colleters on the reproductive apices alternate with the sepals and are sessile, reduced sessile, tripartite laminar sessile, or asymmetrical. All of the colleters have a central nucleus of parenchymatous cells covered by a palisade uniseriate secretory epidermis and a thin cuticle. Secretory idioblasts were observed in the parenchymatous axis. Vascularization was observed only in standard axillary and laminar colleters. Crystals were observed in the parenchyma of the axillary colleter. Histochemical tests demonstrated that there was no rupturing or distension of the cuticle during the secretion process. Mucilage was identified using the PAS reaction as well as by Mayer's reagent and Ruthenium red staining. The calycine colleters had two distinct secretory phases, the first synthesizing mucilage and the second producing phenolic compounds.

An overview of lignin metabolism and its effect on biomass recalcitrance

Lignin, after cellulose, is the second most abundant biopolymer on Earth, accounting for 30% of the organic carbon in the biosphere. It is considered an important evolutionary adaptation of plants during their transition from the aquatic environment to land, since it bestowed the early tracheophytes with physical support to stand upright and enabled long-distance transport of water and solutes by waterproofing the vascular tissue. Although essential for plant growth and development, lignin is the major plant cell wall component responsible for biomass recalcitrance to industrial processing. The fact that lignin is a non-linear aromatic polymer built with chemically diverse and poorly reactive linkages and a variety of monomer units precludes the ability of any single enzyme to properly recognize and degrade it. Consequently, the use of lignocellulosic feedstock as a renewable and sustainable resource for the production of biofuels and bio-based materials will depend on the identification and characterization of the factors that determine plant biomass recalcitrance, especially the highly complex phenolic polymer lignin. Here, we summarize the current knowledge regarding lignin metabolism in plants, its effect on biomass recalcitrance and the emergent strategies to modify biomass recalcitrance through metabolic engineering of the lignin pathway. In addition, the potential use of sugarcane as a second-generation biofuel crop and the advances in lignin-related studies in sugarcane are discussed.

Antioxidant activity of the microalga Spirulina maxima

Spirulina maxima, which is used as a food additive, is a microalga rich in protein and other essential nutrients. Spirulina contains phenolic acids, tocopherols and ß-carotene which are known to exhibit antioxidant properties. The aim of the present study was to evaluate the antioxidant capacity of a Spirulina extract. The antioxidant activity of a methanolic extract of Spirulina was determined in vitro and in vivo. The in vitro antioxidant capacity was tested on a brain homogenate incubated with and without the extract at 37oC. The IC50 (concentration which causes a 50% reduction of oxidation) of the extract in this system was 0.18 mg/ml. The in vivo antioxidant capacity was evaluated in plasma and liver of animals receiving a daily dose of 5 mg for 2 and 7 weeks. Plasma antioxidant capacity was measured in brain homogenate incubated for 1 h at 37oC. The production of oxidized compounds in liver after 2 h of incubation at 37oC was measured in terms of thiobarbituric acid reactant substances (TBARS) in control and experimental groups. Upon treatment, the antioxidant capacity of plasma was 71% for the experimental group and 54% for the control group. Data from liver spontaneous peroxidation studies were not significantly different between groups. The amounts of phenolic acids, a-tocopherol and ß-carotene were determined in Spirulina extracts. The results obtained indicate that Spirulina provides some antioxidant protection for both in vitro and in vivo systems.

Antioxidant and free radical scavenging effects in extracts of the medicinal herb Achyrocline satureioides (Lam.) DC. ("marcela")

Achyrocline satureioides (Lam.) DC. (Compositae) is a medicinal herb used in Argentina, Uruguay, Brazil and Paraguay for its choleretic, antispasmodic and hepatoprotective properties. The presence of the flavonoid quercetin and its derivatives, and of different phenolic acids such as caffeic, chlorogenic and isochlorogenic acids in the aerial parts of this plant has led us to study the antioxidant activity of its extracts using different bioassays. The inhibition of luminol-enhanced chemiluminescence by the aqueous and methanolic extracts was used to show that their total reactive antioxidant potential index (TRAP; in µM Trolox equivalents) was 91.0 ± 15.4 and 128.1 ± 20.1 µM, respectively, while the total antioxidant reactivity index (TAR) was calculated to be 1537 ± 148 and 1910 ± 171 µM. Only the methanolic extract was capable of reducing iron (II)-dependent DNA damage. Lipid peroxidation was assessed by two different methods. The aqueous extract reduced hydroperoxide-initiated chemiluminescence in rat liver homogenates at all concentrations in a dose-dependent manner, with a calculated IC50 = 225 µg/ml, while the methanolic extract was only effective at higher concentrations (100 and 1000 µg/ml). Both aqueous and methanolic extracts were capable of reducing the production of thiobarbituric acid reactive substances (TBARS) in rat liver homogenates, with an IC50 >1000 µg/ml. The results obtained suggest that the extracts of A. satureioides possess significant free radical scavenging and antioxidant activity in vitro, a fact that should encourage future in vivo studies.

Novel donors of nitric oxide derived of S-nitrosocysteine possessing antioxidant activities

Novel S-nitrosothiols possessing a phenolic function were investigated as nitric oxide (NO) donors. A study of NO release from these derivatives was carried out by electron spin resonance (ESR). All compounds gave rise to a characteristic three-line ESR signal in the presence of the complex [Fe(II)(MGD)2], revealing the formation of the complex [Fe(II)(MGD)2(NO)]. Furthermore, tests based on cytochrome c reduction were performed in order to study the ability of each phenolic disulfide, the final organic decomposition product of S-nitrosothiols, to trap superoxide radical anion (O2-). This study revealed a high reactivity of 1b and 3b towards O2-. For these two compounds, the respective inhibitory concentration (IC) 50 values were 92 µM and 43 µM.

Antioxidant and antimicrobial activities of Bauhinia racemosa L. stem bark

The present study was carried out to evaluate the antioxidant and antimicrobial activities of a methanol extract of Bauhinia racemosa (MEBR) (Caesalpiniaceae) stem bark in various systems. 1,1-Diphenyl-2-picryl-hydrazyl (DPPH) radical, superoxide anion radical, nitric oxide radical, and hydroxyl radical scavenging assays were carried out to evaluate the antioxidant potential of the extract. The antioxidant activity of the methanol extract increased in a concentration-dependent manner. About 50, 100, 250, and 500 µg MEBR inhibited the peroxidation of a linoleic acid emulsion by 62.43, 67.21, 71.04, and 76.83%, respectively. Similarly, the effect of MEBR on reducing power increased in a concentration-dependent manner. In DPPH radical scavenging assays the IC50 value of the extract was 152.29 µg/ml. MEBR inhibited the nitric oxide radicals generated from sodium nitroprusside with an IC50 of 78.34 µg/ml, as opposed to 20.4 µg/ml for curcumin. Moreover, MEBR scavenged the superoxide generated by the PMS/NADH-NBT system. MEBR also inhibited the hydroxyl radical generated by Fenton's reaction, with an IC50 value of more than 1000 µg/ml, as compared to 5 µg/ml for catechin. The amounts of total phenolic compounds were also determined and 64.7 µg pyrocatechol phenol equivalents were detected in MEBR (1 mg). The antimicrobial activities of MEBR were determined by disc diffusion with five Gram-positive, four Gram-negative and four fungal species. MEBR showed broad-spectrum antimicrobial activity against all tested microorganisms. The results obtained in the present study indicate that MEBR can be a potential source of natural antioxidant and antimicrobial agents.

Plant-Herbivore Interaction in a Fragmented Landscape: Local Adaptation and Inbreeding

Reciprocal selection between interacting species is a major driver of biodiversity at both the genetic and the species level. This reciprocal selection, or coevolution, has led to the diversification of two highly diverse and abundant groups of organisms, flowering plants and their insect herbivores. In heterogeneous environments, the outcome of coevolved species interactions is influenced by the surrounding community and/or the abiotic environment. The process of adaptation allows species to adapt to their local conditions and to local populations of interacting species. However, adaptation can be disrupted or slowed down by an absence of genetic variation or by increased inbreeding, together with the following inbreeding depression, both of which are common in small and isolated populations that occur in fragmented environments. I studied the interaction between a long-lived plant Vincetoxicum hirundinaria and its specialist herbivore Abrostola asclepiadis in the southwestern archipelago of Finland. I focused on mutual local adaptation of plants and herbivores, which is a demonstration of reciprocal selection between species, a prerequisite for coevolution. I then proceeded to investigate the processes that could potentially hamper local adaptation, or species interaction in general, when the population size is small. I did this by examining how inbreeding of both plants and herbivores affects traits that are important for interaction, as well as among-population variation in the effects of inbreeding. In addition to bi-parental inbreeding, in plants inbreeding can arise from self-fertilization which has important implications for mating system evolution. I found that local adaptation of the plant to its herbivores varied among populations. Local adaptation of the herbivore varied among populations and years, being weaker in populations that were most connected. Inbreeding caused inbreeding depression in both plants and herbivores. In some populations inbreeding depression in herbivore biomass was stronger in herbivores feeding on inbred plants than in those feeding on outbred ones. For plants it was the other way around: inbreeding depression in anti-herbivore resistance decreased when the herbivores were inbred. Underlying some of the among-population variation in the effects of inbreeding is variation in plant phenolic compounds. However, variation in the modification of phenolic compounds in the digestive tract of the herbivore did not explain the inbreeding depression in herbivore biomass. Finally, adult herbivores had a preference for outbred host plants for egg deposition, and herbivore inbreeding had a positive effect on egg survival when the eggs were exposed to predators and parasitoids. These results suggest that plants and herbivores indeed exert reciprocal selection, as demonstrated by the significant local adaptation of V. hirundinaria and A. asclepiadis to one another. The most significant cause of disruption of the local adaptation of herbivore populations was population connectivity, and thus probably gene flow. In plants local adaptation tended to increase with increasing genetic variation. Whether or not inbreeding depression occurred varied according to the life-history stage of the herbivore and/or the plant trait in question. In addition, the effects of inbreeding strongly depended on the population. Taken together, inbreeding modified plant-herbivore interactions at several different levels, and can thus affect the strength of reciprocal selection between species. Thus inbreeding has the potential to affect the outcome of coevolution.

Antioxidant and antimicrobial activities of bitter and sweet apricot (Prunus armeniaca L.) kernels

Abstract The present study describes the in vitro antimicrobial and antioxidant activity of methanol and water extracts of sweet and bitter apricot (Prunus armeniaca L.) kernels. The antioxidant properties of apricot kernels were evaluated by determining radical scavenging power, lipid peroxidation inhibition activity and total phenol content measured with a DPPH test, the thiocyanate method and the Folin method, respectively. In contrast to extracts of the bitter kernels, both the water and methanol extracts of sweet kernels have antioxidant potential. The highest percent inhibition of lipid peroxidation (69%) and total phenolic content (7.9 ± 0.2 µg/mL) were detected in the methanol extract of sweet kernels (Hasanbey) and in the water extract of the same cultivar, respectively. The antimicrobial activities of the above extracts were also tested against human pathogenic microorganisms using a disc-diffusion method, and the minimal inhibitory concentration (MIC) values of each active extract were determined. The most effective antibacterial activity was observed in the methanol and water extracts of bitter kernels and in the methanol extract of sweet kernels against the Gram-positive bacteria Staphylococcus aureus. Additionally, the methanol extracts of the bitter kernels were very potent against the Gram-negative bacteria Escherichia coli (0.312 mg/mL MIC value). Significant anti-candida activity was also observed with the methanol extract of bitter apricot kernels against Candida albicans, consisting of a 14 mm in diameter of inhibition zone and a 0.625 mg/mL MIC value.

Inhibition of cyclooxygenase activity by standardized hydroalcoholic extracts of four Asteraceae species from the Argentine Puna

We determined the anti-inflammatory activity of standardized extracts of four medicinal plant species (Baccharis incarum, B. boliviensis, Chuquiraga atacamensis, Parastrephia lucida) that grow in the Argentine Puna (3800 m above sea level) and that are used to reduce oxidative stress and alleviate gout and arthritic pain. The extracts of plant aerial parts were standardized in terms of total phenolic compounds and flavone/flavanone content and free radical scavenging activity. All extracts showed high phenolic compound concentration (0.5-1.6 mg/mL), mainly flavones and flavonols (0.1-0.8 mg/mL). The extracts showed hydrogen donating ability (DPPH and ABTS) and reactive oxygen species scavenging activity (O2●-, OH-, H2O2). The ability of the extracts to inhibit cyclooxygenase enzymes (COX-1 and COX-2) was determined by calculating percent inhibition of PGE2 production measured by enzyme immunoassay. All extracts inhibited both enzymes with IC50 values of 2.0 to 16.7 µg/mL. The anti-inflammatory activity of B. incarum and C. atacamensis extracts was higher than that of B. boliviensis and P. lucida. The IC50 values obtained for indomethacin were 0.11 and 0.78 µM for COX-1 and COX-2, respectively. The present results are consistent with the anecdotal use of these species in phytotherapic preparations.

Rosemary (Rosmarinus officinalis): a study of the composition, antioxidant and antimicrobial activities of extracts obtained with supercritical carbon dioxide

Rosemary leaf extracts were obtained by supercritical fluid extraction (SFE) and Soxhlet extraction. Their chemical compositions were evaluated by GC-MS. The extracts were analyzed for compounds reported in the literature as showing antimicrobial and antioxidant activities. The rosemary extracts were tested with regard to antioxidant (DPPH radical scavenging and total phenolic content - Folin-Denis reagent), antibacterial (Gram-positive bacteria - Staphylococcus aureus ATCC 25923 and Bacillus cereus ATCC 11778 - and Gram-negative bacteria - Escherichia coli ATCC 25922 and Pseudomonas aeruginosa ATCC 27853) and antifungal (Candida albicans) activities. Antioxidant, antibacterial and antifungal activities of the SFE extracts were confirmed.