Development and characterization of microsatellite markers for turmeric (Curcuma longa)

P>Curcuma longa L. is a sterile, triploid, vegetatively-propagated crop cultivated mainly in Southeast Asia. When dried rhizomes are ground, the resulting yellow powder is used by the food industry as a natural food dye. Moreover, many pharmacological compounds have broadened the commercial application of the crop. However, conventional breeding is difficult and hence, improvement has been limited to germplasm selection. To better utilize the germplasm collections and facilitate genotype selection, a total of 17 polymorphic microsatellite loci were developed using a CT/GT/CTT enriched genomic library. All microsatellites resulted in amplified PCR products, showing a banding pattern of 2-11 polymorphic bands per locus, enabling genotype discrimination. These results can be used in further studies aimed at characterizing C. longa genetic resource collections and also to improve breeding strategies.

Development and characterization of microsatellite markers from Guarea guidonia (Meliaceae), a tree species from different habitats within the Brazilian Atlantic forest

The tree species Guarea guidonea (Meliaceae) belongs to a predominantly tropical family, being largely found in natural or anthropic forest fragments within the Brazilian Atlantic Forest. Aiming to develop future studies on the genetic structure of plant species from forests fragments, eleven microsatellite markers were developed for Guarea guidonia, based on the analysis of 45 individuals from natural populations of three different fragments within the forest-anthropic edge, interior fragment and natural edge. Only eight loci showed to be polymorphic and the number of alleles ranged from two to four (mean of 2.50). All populations showed almost the same level of genetic diversity (mean H(e) = 0.3775). These loci will be useful for population genetics studies on Guarea guidonea, providing information for the conservation and management of this species.

Development and characterization of microsatellite markers for castor (Ricinus communis L.), an important oleaginous species for biodiesel production

Castor (Ricinus communis L.) is an important oleaginous plant from both economic and social points of view. The seeds contain an oil with excellent properties for industrial uses. This paper presents the main results of a study aiming to develop microsatellite markers for castor. Twelve new polymorphic microsatellite markers were isolated and characterized in 38 genotypes accessions from the castor germplasm of the Brazilian Agricultural Research Company (EMBRAPA). Knowledge on the genetic diversity of castor can be used to gain a better understanding on genetic diversity conservation, and germplasm management, guiding breeding programs and conservation strategies.

Development and characterization of microsatellite markers for Psychotria tenuinervis (Rubiaceae), a shrub species from the Atlantic forest, and primers transferability from Coffea

The shrub species Psychotria tenuinervis (Rubiaceae) is native to the Brazilian Atlantic forest and is largely found within natural and disturbed forest fragments. Aiming to develop studies on population genetic structure of forest fragment species, eigth microsatellite markers were developed for P. tenuinervis. Also, 15 loci already developed for Coffea (Rubiaceae) were tested for transferability to this species. We utilized 45 individuals from natural populations of three different fragments-anthropic edge, interior fragment and natural edge, within the Brazilian Atlantic forest. The average number of alleles per locus was 2.5 (two-four alleles/locus). These loci will be useful for future population genetic studies aiming to the conservation and management of this species.

Attainment and Characterization of Ternary Complexes of Simvastatin-Cyclodextrin-Hydrossoluble Polymers

The purpose of this study was to attain and characterize ternary complexes of simvastatin, beta-cyclodextrin (beta CD) and different polymers, and then select those that lead to a greater increase in drug solubility. The complexes were prepared with the co-evaporation method and the polymers used were polyethylene glycol 1500, polyethylene glycol 4000, povidone, copovidone, crospovidone, maltodextrin and hydroxypropyl methyl cellulose. The characterization of complexes was carried out through aqueous solubility, DSC and TG. There was an increase in solubility for all the complexes prepared with beta CD and the different polymers, but only when crospovidone and maltodextrin were used was there a significant difference observed between the solubility of the physical mixture and that of the complex. The DSC curves indicate that the non-complexed drug is even in the sample of the complex with higher solubility, thus none of the polymers was able to achieve a total complexation of the drug.

Development and Characterization of L-Alanyl-L-Glutamine Containing Pellets employing Extrusion-Spheronization Method and Drying Process in Fluidized Bad Equipment.

Development and Characterization of L-Alanyl-L-Glutamine Containing Pellets employing Extrusion-Spheronization Method and Drying Process in Fluidized Bad Equipment"". In this work, five formulations of L-alanyl-L-glutamine (glutamine dipeptide) containing pellets with different drug concentration were developed and evaluated: F1 (9.07%); F2 (17.70%); F3 (27.98%); F4 (37.74%) e F5 (47.53%). Pellets were prepared by extrusion-spheronization method and, further, dried in fluidized bad equipment. The following assays were carried out with the batches obtained: granulometry, friability, true density and morphologic analysis. Between the five formulations evaluated, pellets obtained from F3 present best yield (75.80%), most uniform particle size distribution (89.67% of pellets with size in the range of 0.80 to 1.18), most high true density (2.1634 g/ml) and best aspect (1.0795 +/- 0.0410). Due to these features, pellets obtained from F3 were considered adequate to further polymeric coating process in order to produce a multiparticulate system to prolong L-alanyl-L-glutamine release.

Emulsified systems based on glyceryl monostearate and potassium cetyl phosphate: Scale-up and characterization of physical properties

introducing a pharmaceutical product on the market involves several stages of research. The scale-up stage comprises the integration of previous phases of development and their integration. This phase is extremely important since many process limitations which do not appear on the small scale become significant on the transposition to a large one. Since scientific literature presents only a few reports about the characterization of emulsified systems involving their scaling-up, this research work aimed at evaluating physical properties of non-ionic and anionic emulsions during their manufacturing phases: laboratory stage and scale-up. Prototype non-ionic (glyceryl monostearate) and anionic (potassium cetyl phosphate) emulsified systems had the physical properties by the determination of the droplet size (D[4,3 1, mu m) and rheology profile. Transposition occurred from a batch of 500-50,000 g. Semi-industrial manufacturing involved distinct conditions: intensity of agitation and homogenization. Comparing the non-ionic and anionic systems, it was observed that anionic emulsifiers generated systems with smaller droplet size and higher viscosity in laboratory scale. Besides that, for the concentrations tested, augmentation of the glyceryl monostearate emulsifier content provided formulations with better physical characteristics. For systems with potassium cetyl phosphate, droplet size increased with the elevation of the emulsifier concentration, suggesting inadequate stability. The scale-up provoked more significant alterations on the rheological profile and droplet size on the anionic systems than the non-ionic. (C) 2008 Elsevier B.V. All rights reserved.

Preparation and characterization of ethanol-treated silk fibroin dense membranes for biomaterials application using waste silk fibers as raw material

The possibility of producing valued devices from low cost natural resources is a subject of broad interest. The present study explores the preparation and characterization of silk fibroin dense membranes using waste silk fibers from textile processing. Morphology, crystallinity, thermal resistance and cytotoxicity of membranes as well as the changes on the secondary structure of silk fibroin were analyzed after undergoing treatment with ethanol. Membranes presented amorphous patterns as determined via X-ray diffraction. The secondary structure of silk fibroin on dense membranes was either random coil (silk I) or p-sheet (silk II), before and after ethanol treatment, respectively. The sterilized membranes presented no cytotoxicity to endothelial cells during in vitro assays. This fact stresses the material potential to be used in the fabrication of biomaterials, as coatings of cardiovascular devices and as membranes for wound dressing or drug delivery systems. (C) 2010 Elsevier Ltd. All rights reserved.

Batch purification of high-purity lysozyme from egg white and characterization of the enzyme modified by PEGylation

PEGylation is one of the most promising and extensively studied strategies for improving the pharmacological properties of proteins as well as their physical and thermal stability. Purified lysozyme obtained from hen egg white by batch mode was modified by PEGylation with methoxypolyethyleneglycol succinimidyl succinato (mPEG-SS, MW 5000). The conjugates produced retained full enzyme activity with the substrate glycol chitosan, independent of degree of enzyme modification, although lysozyme activity with the substrate Micrococcus lysodeikticus was altered according to the degree of modification. The conjugate with a low degree of modification by mPEG-SS retained 67% of its enzyme activity with the M. lysodeikticus substrate. The mPEG-SS was also shown to be a highly reactive polymer. The effects of pH and temperature on PEGylated lysozymes indicated that the conjugate was active over a wide pH range and was stable up to 50 degrees C. This conjugate also showed resistance to proteolytic degradation, remained stable in human serum, and displayed greater antimicrobial activity than native lysozyme against Gram-negative bacteria.

Production and characterization of sponge-dough bread using scalded rye

In this work, flatbed scanning, instrumental texture analysis, spectrophotometric color determination (L*, a*, b*), moisture and specific volume measurements were used to evaluate the effects of the addition of rye flour or rye flakes, yeast and boiling water in different amounts in sponge-dough rye bread production. The treatments changed significantly (P < 0.05) the crumb cell area (mm(2)), cell diameter (mm), cell perimeter (mm), texture parameters and light reflectance (L*, a*, b*). Scalding process could be used to produce new textures and color of baked products.

Purification and characterization of Ts15, the first member of a new alpha-KTX subfamily from the venom of the Brazilian scorpion Tityus serrulatus

Voltage-gated potassium channel toxins (KTxs) are basic short chain peptides comprising 23-43 amino acid residues that can be cross-linked by 3 or 4 disulfide bridges. KTxs are classified into four large families: alpha-, beta-, gamma- and kappa-KTx. These peptides display varying selectivity and affinity for K(v) channel subtypes. In this work, a novel toxin from the Tityus serrulatus venom was isolated, characterized and submitted to a wide electrophysiological screening on 5 different subtypes of Nay channels (Na(V)1.4; Na(V)1.5; Na(V)1.6; Na(V)1.8 and DmNa(V)1) and 12 different subtypes of Kv channels (K(V)1.1 - K(V)1.6; K(V)2.1; K(V)3.1; K(V)4.2; K(V)4.3; Shaker IR and ERG). This novel peptide, named Ts15, has 36 amino acids, is crosslinked by 3 disulfide bridges, has a molecular mass of 3956 Da and pI around 9. Electrophysiological experiments using patch clamp and the two-electrode voltage clamp techniques show that Ts15 preferentially blocks K(V)1.2 and K(V)1.3 channels with an IC(50) value of 196 +/- 25 and 508 +/- 67 nM, respectively. No effect on Na(V) channels was observed, at all tested concentrations. Since Ts15 shows low amino acid identity with other known KTxs, it was considered a bona fide novel type of scorpion toxin. Ts15 is the unique member of the new alpha-Ktx21 subfamily and therefore was classified as alpha-Ktx21.1. (C) 2011 Elsevier Ltd. All rights reserved.

Preparation and characterization of D, L-PLA loaded 17-beta-Estradiol valerate by emulsion/evaporation methods

PLA microparticles containing 17-beta-estradiol valerate were prepared by an emulsion/evaporation method in order to sustain drug release. This system was characterized concerning particle size, particle morphology and the influence of formulation and processing parameters on drug encapsulation and in vitro drug release. The biodegradation of the microparticles was observed by tissue histological analysis. Scanning electron microscopy and particle size analysis showed that the microparticles were spherical, presenting non-aggregated homogeneous surface and had diameters in the range of 718-880 nm (inert microparticles) and 3-4 mu m (drug loaded microparticles). The encapsulation efficiency was similar to 80%. Hormone released from microparticles was sustained. An in vivo degradation experiment confirmed that microparticles are biodegradable. The preparation method was shown to be suitable, since the morphological characteristics and efficiency yield were satisfactory. Thus, the method of developed microparticles seems to be a promising system for sustained release of 17-beta-estradiol.

Preparation and characterization of chitosan-treated alginate microparticles incorporating all-trans retinoic acid

Chitosan treated alginate microparticles were prepared with the purpose of incorporating all-trans retinoic acid (ATRA) using an inexpensive, simple and fast method, enhancing dermal localization and sustaining the release of ATRA into the skin. Microparticles characterization, drug-polymer interaction, release profile and in vitro skin retention were investigated. Microparticles presented spherical shape and drug loading capacity of 47%. The drug content of these microparticles was affected by ATRA concentration and by the solvent used and it was more weakly affected by chitosan concentration. The release of ATRA was also affected by chitosan concentration. Microparticles prepared with 0.4% chitosan (w/w) resulted in drug release with a more sustained profile. The results of in vitro retention studies showed that chitosan treated alginate microparticles decreased the drug retention in the stratum corneum (SC), where occur the skin irritation, but maintained the ATRA concentration in the deeper skin layers, where occur the pathologies treated with ATRA. Then, the microparticles developed in this work can be a good candidate to improve the topical therapy with retinoid.

The isolation and characterization of virulence factors of Cryptococcus spp. from saprophytic sources in the city of Ribeirao Preto, Sao Paulo, Brazil

Yeasts of the Cryptococcus genus are distributed in nature associated to animal and vegetal organic residues. Occasionally, species other than C. neoformans may be responsible for infectious diseases in human and animals. This study aims to determine the occurrence of Cryptococcus species in the atmosphere and bird droppings in the city of Ribeirao Preto, Sao Paulo, Brazil, and to evaluate three virulence factors: capsule formation, growth at 37 degrees C and melanin production. We analyzed 86 environmental samples (54 droppings and 32 air). Of the 41 strains isolated, 15 were C. neoformans var. neoformans (12 droppings and 3 air), 15 C. albidus (12 droppings and 3 air), 9 C. laurentii (7 droppings and 2 air) and 2 C. uniguttulatus (from droppings). Capsules were produced by 93.3% of C. neoformans var. neoformans, 66.7% of C. albidus, 88.9% of C. laurentii and 50% (1/2) of C. uniguttulatus. All, strains of C. neoformans, 20% of C. albidus and 44.4% of C. laurentii were able to grow at 37 degrees C. The melanin production on DOPA agar was verified in C. neoformans (93.3%), C. albidus (26.7%) and C. laurentii (66.7%). We concluded that different Cryptococcus species coexist in the same ecological niche and they are able to produce virulence factors. (C) 2007 Elsevier GmbH. All rights reserved.

Prevalence and characterization of Enterococcus spp. isolated from Brazilian foods

Enterococci can be used in the food industry as starter or probiotic cultures. However, enterococci are also implicated in severe multi-resistant nosocomial infections. In this study, the prevalence of enterococci in selected Brazilian foodstuffs (raw and pasteurized milk, meat products, cheeses and vegetables) was evaluated. Phenotypic and PCR protocols were used for species identification. Tests for production of gelatinase, haemolysin, bacteriocin and bile salt hydrolysis were done with all enterococci isolates, whereas molecular determination of virulence markers (genes esp, gel, ace, as, efaA, hyl and cylA) and antibiotic resistance was checked only for Enterococcus faecium and Enterococcus faecalis isolates. The antibiotic-resistant isolates were assayed for biofilm formation and adhesion to mammalian cells. From the 120 food samples analyzed, 52.5% were positive for enterococci, meat and cheese being the most contaminated. E. faecium was the predominant species, followed by E. faecalis, E. casseliflavus and Enterococcus gallinarum. Phenotypic tests indicated that 67.7% of isolates hydrolyzed bile salts, 15.2% produced bacteriocin, 12.0% were beta-hemolytic and 18.2% produced gelatinase. Antibiotic resistance (gentamicin, tetracycline and erythromycin) and genes encoding for virulence traits were more frequent in E. faecalis than in E. faecium. Three E. faecium isolates were resistant to vancomycin. Among antibiotic-resistant isolates, 72.4% of E. faecalis were able to form biofilm and 13.8% to adhere to Caco-2 cells. Antibiotic-resistant E. faecalis and E. faecium isolates were grouped by RAPD-PCR and a scattered distribution was noted, indicating that resistance was not related to a particular clone. The spread of virulence/resistance traits in isolates of the two species and different RAPD-types suggest the pathogenic potential of both species. By contrast, the recovery of bacteriocinogenic E. faecium isolates with no virulence traits suggests their potential for biotechnological applications. In conclusion, our results showed that enterococci from Brazilian foods present important dualist aspects for food safety. (C) 2008 Elsevier Ltd. All rights reserved.