Chemical Characterization and Antimicrobial Activity Evaluation of Natural Oil Nanostructured Emulsions.

The aim of this work was to investigate the antimicrobial activity of nanostructured emulsions based on copaiba (Copaifera langsdorffii) resin-oil, copaiba essential oil, and bullfrog (Rana catesbeiana Shaw) oil against fungi and bacteria related to skin diseases. Firstly, the essential oil was extracted from copaiba resin-oil and these oils, along with bullfrog oil, were characterized by gas chromatography combined with mass spectrometry (GC-MS). Secondly, nanostructured emulsion systems were produced and characterized. The antimicrobial susceptibility assay was performed, followed by the Minimum Inhibitory Concentration (MIC) determination, the bioautography assay, and the antibiofilm determination. Strains of the genera Staphylococcus, Pseudomonas, and Candida were used. The CG-MS analysis was able to identify the components of copaiba resin-oil, copaiba essential oil, and bullfrog oil. The MIC assay in association with the bioautography revealed that some esters of palmitic and oleic acids, a-curcumene, a-himachalene, isothujol, and α-fenchene--probably inhibited some strains. The nanostructured emulsions based on copaiba resin-oil and essential oil improved the antimicrobial activity of the pure oils, especially against Staphylococcus and Candida, resistant to azoles. The bullfrog oil nanostructured emulsion showed a lower antimicrobial effect when compared to the copaiba samples. However, bullfrog oil-based nanostructured emulsion showed a significant antibiofilm activity (p < 0.05). Given the significant antimicrobial and antibiofilm activities of the evaluated oils, it may be concluded that nanostructured emulsions based on copaiba and bullfrog oils are promising candidates for the treatment of infections and also may be used to incorporate other antimicrobial drugs.

Antiproliferative activity of fucan nanogel.

Sulfated fucans comprise families of polydisperse natural polysaccharides based on sulfated L-fucose. Our aim was to investigate whether fucan nanogel induces cell-specific responses. To that end, a non toxic fucan extracted from Spatoglossum schröederi was chemically modified by grafting hexadecylamine to the polymer hydrophilic backbone. The resulting modified material (SNFuc) formed nanosized particles. The degree of substitution with hydrophobic chains was close to 100%, as estimated by elemental analysis. SNFfuc in aqueous media had a mean diameter of 123 nm and zeta potential of -38.3 ± 0.74 mV, as measured by dynamic light scattering. Nanoparticles conserved their size for up to 70 days. SNFuc cytotoxicity was determined using the MTT assay after culturing different cell lines for 24 h. Tumor-cell (HepG2, 786, H-S5) proliferation was inhibited by 2.0%-43.7% at nanogel concentrations of 0.05-0.5 mg/mL and rabbit aorta endothelial cells (RAEC) non-tumor cell line proliferation displayed inhibition of 8.0%-22.0%. On the other hand, nanogel improved Chinese hamster ovary (CHO) and monocyte macrophage cell (RAW) non-tumor cell line proliferation in the same concentration range. The antiproliferative effect against tumor cells was also confirmed using the BrdU test. Flow cytometric analysis revealed that the fucan nanogel inhibited 786 cell proliferation through caspase and caspase-independent mechanisms. In addition, SNFuc blocks 786 cell passages in the S and G2-M phases of the cell cycle.

Freshwater plants synthesize sulfated polysaccharides: heterogalactans from Water Hyacinth (Eicchornia crassipes).

Sulfated polysaccharides (SP) are found mainly in seaweeds and animals. To date, they have only been found in six plants and all inhabit saline environments. Furthermore, there are no reports of SP in freshwater or terrestrial plants. As such, this study investigated the presence of SP in freshwaters Eichhornia crassipes, Egeria densa, Egeria naja, Cabomba caroliniana, Hydrocotyle bonariensis and Nymphaea ampla. Chemical analysis identified sulfate in N. ampla, H. bonariensis and, more specifically, E. crassipes. In addition, chemical analysis, FT-IR spectroscopy, histological analysis, scanning electron microscopy (SEM) and energy-dispersive X-ray analysis (EDXA), as well as agarose gel electrophoresis detected SP in all parts of E. crassipes, primarily in the root (epidermis and vascular bundle). Galactose, glucose and arabinose are the main monosaccharides found in the sulfated polysaccharides from E. crassipes. In activated partial thromboplastin time (APTT) test, to evaluate the intrinsic coagulation pathway, SP from the root and rhizome prolonged the coagulation time to double the baseline value, with 0.1 mg/mL and 0.15 mg/mL, respectively. However, SP from the leaf and petiole showed no anticoagulant activity. Eichornia SP demonstrated promising anticoagulant potential and have been selected for further studies on bioguided fractionation; isolation and characterization of pure polysaccharides from this species. Additionally in vivo experiments are needed and are already underway.

In vitro antioxidant, anticoagulant and antimicrobial activity and in inhibition of cancer cell proliferation by xylan extracted from corn cobs.

Xylan is one of most abundant polymer after cellulose. However, its potential has yet to be completely recognized. Corn cobs contain a considerable reservoir of xylan. The aim of this work was to study some of the biological activities of xylan obtained from corn cobs after alkaline extraction enhanced by ultrasonication. Physical chemistry and infrared analyses showed 130 kDa heteroxylan containing mainly xylose:arabinose: galactose:glucose (5.0:1.5:2.0:1.2). Xylan obtained exhibited total antioxidant activity corresponding to 48.5 mg of ascorbic acid equivalent/g of xylan. Furthermore, xylan displayed high ferric chelating activity (70%) at 2 mg/mL. Xylan also showed anticoagulant activity in aPTT test. In antimicrobial assay, the polysaccharide significantly inhibited bacterial growth of Klebsiella pneumoniae. In a test with normal and tumor human cells, after 72 h, only HeLa tumor cell proliferation was inhibited (p < 0.05) in a dose-dependent manner by xylan, reaching saturation at around 2 mg/mL, whereas 3T3 normal cell proliferation was not affected. The results suggest that it has potential clinical applications as antioxidant, anticoagulant, antimicrobial and antiproliferative compounds.

Heterofucans from the brown seaweed Canistrocarpus cervicornis with anticoagulant and antioxidant activities.

Fucan is a term used to denominate a family of sulfated polysaccharides rich in sulfated l-fucose. We extracted six fucans from Canistrocarpus cervicornis by proteolytic digestion followed by sequential acetone precipitation. These heterofucans are composed mainly of fucose, glucuronic acid, galactose and sulfate. No polysaccharide was capable of prolonging prothrombin time (PT) at the concentration assayed. However, all polysaccharides prolonged activated partial thromboplastin time (aPTT). Four sulfated polysaccharides (CC-0.3/CC-0.5/CC-0.7/CC-1.0) doubled aPTT with only 0.1 mg/mL of plasma, only 1.25-fold less than Clexane, a commercial low molecular weight heparin. Heterofucans exhibited total antioxidant capacity, low hydroxyl radical scavenging activity, good superoxide radical scavenging efficiency (except CC-1.0), and excellent ferrous chelating ability (except CC-0.3). These results clearly indicate the beneficial effect of C. cervicornis polysaccharides as anticoagulants and antioxidants. Further purification steps and additional studies on structural features as well as in vivo experiments are needed to test the viability of their use as therapeutic agents.

Heterofucan from Sargassum filipendula induces apoptosis in HeLa cells.

Fucan is a term used to denominate a family of sulfated polysaccharides rich in sulfated l-fucose. Heterofucan SF-1.5v was extracted from the brown seaweed Sargassum filipendula by proteolytic digestion followed by sequential acetone precipitation. This fucan showed antiproliferative activity on Hela cells and induced apoptosis. However, SF-1.5v was not able to activate caspases. Moreover, SF-1.5v induced glycogen synthase kinase (GSK) activation, but this protein is not involved in the heterofucan SF-1.5v induced apoptosis mechanism. In addition, ERK, p38, p53, pAKT and NFκB were not affected by the presence of SF-1.5v. We determined that SF-1.5v induces apoptosis in HeLa mainly by mitochondrial release of apoptosis-inducing factor (AIF) into cytosol. In addition, SF-1.5v decreases the expression of anti-apoptotic protein Bcl-2 and increased expression of apoptogenic protein Bax. These results are significant in that they provide a mechanistic framework for further exploring the use of SF-1.5v as a novel chemotherapeutics against human cervical cancer.

Antioxidant and antiproliferative activities of heterofucans from the seaweed Sargassum filipendula.

Fucan is a term used to denominate a type of polysaccharide which contains substantial percentages of l-fucose and sulfate ester groups. We obtained five heterofucans from Sargassum filipendula by proteolytic digestion followed by sequential acetone precipitation. These heterofucans are composed mainly of fucose, glucose, glucuronic acid, galactose and sulfate. These fucans did not show anticoagulant activity in PT and aPTT tests. Their antioxidant activity was evaluated using the follow tests; total antioxidant capacity, scavenging hydroxyl and superoxide radicals, reducing power and ferrous ion [Fe(II)] chelating. All heterofucans displayed considerable activity, especially SF-1.0v which showed the most significant antioxidant potential with 90.7 ascorbic acid equivalents in a total antioxidant capacity test and similar activity when compared with vitamin C in a reducing power assay. The fucan antiproliferative activity was performed with HeLa, PC3 and HepG2 cells using MTT test. In all tested conditions the heterofucans exhibited a dose-dependent effect. The strongest inhibition was observed in HeLa cells, where SF-1.0 and SF-1.5 exhibited considerable activity with an IC50 value of 15.69 and 13.83 µM, respectively. These results clearly indicate the beneficial effect of S. filipendula polysaccharides as antiproliferative and antioxidant. Further purification steps and additional studies on structural features as well as in vivo experiments are needed to test the viability of their use as therapeutic agents.

Anticoagulant, antioxidant and antitumor activities of heterofucans from the seaweed Dictyopteris delicatula.

In the present study, six families of sulfated polysaccharides were obtained from seaweed Dictyopteris delicatula by proteolytic digestion, followed by acetone fractionation and molecular sieving on Sephadex G-100. Chemical analyses demonstrated that all polysaccharides contain heterofucans composed mainly of fucose, xylose, glucose, galactose, uronic acid, and sulfate. The fucans F0.5v and F0.7v at 1.0 mg/mL showed high ferric chelating activity (∼45%), whereas fucans F1.3v (0.5 mg/mL) showed considerable reducing power, about 53.2% of the activity of vitamin C. The fucan F1.5v presented the most prominent anticoagulant activity. The best antiproliferative activity was found with fucans F1.3v and F0.7v. However, F1.3v activity was much higher than F0.7v inhibiting almost 100% of HeLa cell proliferation. These fucans have been selected for further studies on structural characterization as well as in vivo experiments, which are already in progress.

Antioxidant and Antiproliferative Activities of Leaf Extracts from Plukenetia volubilis Linneo (Euphorbiaceae).

Plukenetia volubilis Linneo, or Sacha inca, is an oleaginous plant from the Euphorbiaceae family. The aim of this work was to perform a chemical and biological analysis of different leaf extracts from P. volubilis such as aqueous extract (AEL), methanol (MEL), ethanol (EEL), chloroform (CEL), and hexane (HEL). Thin layer chromatography analysis revealed the presence of phenolic compounds, steroids, and/or terpenoídes. Furthermore, the antioxidant activities were analyzed by in vitro assays and their effects on cell lineages by in vivo assays. The Total Antioxidant Capacity (TCA) was expressed as equivalent ascorbic acid (EEA/g) and it was observed that the extracts showed values ranging from 59.31 to 97.76 EAA/g. Furthermore, the DPPH assay values ranged from 62.8% to 88.3%. The cell viability assay showed that the extracts were able to reduce viability from cancer cells such as HeLa and A549 cells. The extracts MEL and HEL (250 µg/mL) were able to reduce the proliferation of HeLa cells up to 54.3% and 48.5%, respectively. The flow cytometer results showed that these extracts induce cell death via the apoptosis pathway. On the other hand, the extracts HEL and AEL were able to induce cell proliferation of normal fibroblast 3T3 cells.

Biological activities of the sulfated polysaccharide from the vascular plant Halodule wrightii

A sulfated polysaccharide (SPSG) was successfully isolated from seagrass Halodule wrightii Asch., Cymodoceaceae, and its antioxidant and anticoagulant activities were investigated. The data presented here showed that the SPSG is a 11 kDa sulfated heterogalactan with a sulfatation degree of 20.63 percent and it also contains glucose and xylose. SPSG antioxidant activities were evaluated using several in vitro assays and the anticoagulant activity was evaluated by aPTT and PT tests. These assays suggested that the SPSG possessed remarkable antioxidant properties in different in vitro assays and an outstanding anticoagulant activity 2.5-fold higher than that of heparin Clexane® in the aPTT test. This data represents the first reported on the sulfated polysaccharide biological activities from seagrass. These results indicate that SPSG can be considered in the future as a drug utilized in treating diseases from these systems.

Evaluation of acute and subchronic toxicity of a non-anticoagulant, but antithrombotic algal heterofucan from the Spatoglossum schröederi in Wistar rats

Fucan is a term used to denominate a family of sulfated polysaccharides rich in L-fucose. The brown alga Spatoglossum schröederi, Dictyotaceae, synthesizes three heterofucans named A, B, and C. Fucan A is a non-anticoagulant heterofucan which possesses potent antithrombotic (in vivo) and antiproliferative (in vitro) activities. However, its toxicity in vivo has not been determined. The present study examined the acute and subchronic toxicity of the fucan A in Wistar rats after subcutaneous administration. After that, the animals were killed and examined. The results showed in the acute study that fucan A did not cause general adverse effects and mortality in the concentrations 0, 20, 100, 1000, and 2000 µg/g body weight per rat for seven days. Regarding the subchronic study, the data showed that the fucan A did not cause any change in hematological and biochemistry parameters, as well as in the morphology, and in the size of the rat's organs analyzed at a concentration of 20 µg/g body weight per rat during a 62-day period. In conclusion, this study indicates this heterofucan is a compound with potential pharmacological value that has no toxicity in vivo.