Separation and partitioning of Green Fluorescent Protein from Escherichia coli homogenate in poly(ethylene glycol)/sodium-poly(acrylate) aqueous two-phase systems

The partitioning of Green Fluorescent Protein (GFP) in poly(ethylene glycol)/Na-poly(acrylate) aqueous two-phase systems (PEG/NaPA-ATPS) has been investigated. The aqueous two-phase systems are formed by mixing the polymers with a salt and a protein solution. The protein partitioning in the two-phase system was investigated at 25 degrees C. The concentration of the GFP was measured by fluorimetry. It was found that the partitioning of GFP depends on the salt type, pH and concentration of PEG. The data indicates that GFP partitions more strongly to the PEG phase in presence of Na2SO4 relative to NaCl. Furthermore, the GFP partitions more to the PEG phase at higher pH. The partition to the PEG phase is strongly favoured in systems with larger tie-line lengths (i.e. systems with higher polymer concentrations). The molecular weight of PEG is important since the partition coefficient (K) of GFP gradually decreases with increasing PEG size, from K ca. 300-400 for PEG 400 to K equal to 1.19 for PEG 8000. A separation process was developed where GFP was separated from a homogenate in two extraction steps: the GFP is first partitioned to the PEG phase in a PEG 3000/NaPA 8000 system containing 3 wt% Na2SO4, where the K value of GFP was 8. The GFP is then re-extracted to a salt phase formed by mixing the previous top-phase with a Na2SO4 solution. The K-value of GFP in this back-extraction was 0.22. The total recovery based on the start material was 74%. (c) 2008 Elsevier B.V. All rights reserved.

Plasmid DNA partitioning and separation using poly(ethylene glycol)/poly(acrylate)/salt aqueous two-phase systems

Phase diagrams of poly(ethylene glycol)/polyacrylate/Na2SO4 systems have been investigated with respect to polymer size and pH. Plasmid DNA from Escherichia coil can depending on pH and polymer molecular weight be directed to a poly(ethylene glycol) or to a polyacrylate-rich phase in an aqueous two-phase system formed by these polymers. Bovine serum albumin (BSA) and E. coil homogenate proteins can be directed opposite to the plasmid partitioning in these systems. Two bioseparation processes have been developed where in the final step the pDNA is partitioned to a salt-rich phase giving a total process yield of 60-70%. In one of them the pDNA is partitioned between the polyacrylate and PEG-phases in order to remove proteins. In a more simplified process the plasmid is partitioned to a PEG-phase and back-extracted into a Na2SO4-rich phase. The novel polyacrylate/PEG system allows a strong change of the partitioning between the phases with relatively small changes in composition or pH. (C) 2012 Elsevier B.V. All rights reserved.

Protein partitioning in poly(ethylene glycol)/sodium polyacrylate aqueous two-phase systems

The partition of hemoglobin, lysozyme and glucose-6-phospate dehydrogenase (G6PDH) in a novel inexpensive aqueous two-phase system (ATPS) composed by poly(ethylene glycol) (PEG) and sodium polyacrylate (NaPA) has been studied. The effect of NaCl and Na2SO4, pH and PEG molecular size on the partitioning has been studied. At high pH (above 9), hemoglobin partitions strongly to the PEG-phase. Although some precipitation of hemoglobin occurs, high recovery values are obtained particularly for lysozyme and G6PDH. The partitioning forces are dominated by the hydrophobic and electrochemical (salt) effects, since the positively charged lysozyme and negatively charged G6PDH partitions to the non-charged PEG and the strongly negatively charged polyacrylate enriched phase, respectively. (c) 2007 Elsevier B.V. All rights reserved.

Phase Diagrams of the Aqueous Two-Phase Systems of Poly(ethylene glycol)/Sodium Polyacrylate/Salts

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Synthesis, characterization and solution properties of amphiphilic N-isopropylacrylamide-poly(ethylene glycol)-dodecyl methacrylate thermosensitive polymers

Terpolymers of N-isopropylacrylamide, dodecyl methacrylate (DOMA) and poly(ethylene glycol) (PEG) methacrylate, were synthesized by random copolymerization, and the composition was controlled to achieve systems having different thermosensitivities. H-1 NMR spectra and gel permeation chromatography (GPC) were employed to characterize the different samples obtained. The solution properties were studied by employing spectrophotometry, fluorescence, and dynamic light scattering techniques. The chemical compositions in the final terpolymers are close to those in the feed. The polymers exhibited cloud point temperatures (T-es) varying from 17 to 52 degrees C. Micropolarity studies using I-1/I-3 ratio of the vibronic bands of pyrene show the formation of amphiphilic aggregates capable of incorporating hydrophobic drugs as the polymer concentration is increased. The critical aggregation concentration (CAC) increases from 3.6 x 10(-3) to 1 x 10(-2) g/l with the PEG content varying from 5 to 35 mol%. Anisotropy measurements confirm the results obtained by pyrene fluorescence and show that the aggregates resulting from intermolecular interactions present different organizations. The hydrodynamic diameters (Dh) of the aggregates determined by dynamic light scattering (DLS) vary from 40 to 150 nm depending on the terpolymer composition. The T-cs and Dh values decreased with the ionic strength, and this behavior was attributed to the dehydration of the polymeric micelles. The capacity of solubilization of the aggregates was evaluated by employing pyrene, and the obtained results confirm the ability to incorporate hydrophobic molecules. (c) 2005 Elsevier B.V All rights reserved.

Subcutaneous tissue reaction to castor oil bean and calcium hydroxide in rats

Castor oil bean cement (COB) is a new material that has been used as an endodontic sealer, and is a candidate material for direct pulp capping. Objective: The purpose of this study was to evaluate the biocompatibility of a new formulation of COB compared to calcium hydroxide cement (CH) and a control group without any material, in the subcutaneous tissue of rats. Material and Methods: The materials were prepared, packed into polyethylene tubes, and implanted in the rat dorsal subcutaneous tissue. Animals were sacrificed at the 7th and 50th days after implantation. A quantitative analysis of inflammatory cells was performed and data were subjected to ANOVA and Tukey's tests at 5% significance level. Results: Comparing the mean number of inflammatory cells between the two experimental groups (COB and CH) and the control group, statistically significant difference (p=0.0001) was observed at 7 and 50 days. There were no significant differences (p=0.111) between tissue reaction to CH (382 inflammatory cells) and COB (330 inflammatory cells) after 7 days. After 50 days, significantly more inflammatory cells (p=0.02) were observed in the CH group (404 inflammatory cells) than in the COB group (177 inflammatory cells). Conclusions: These results demonstrate that the COB cement induces less inflammatory response within long periods.

Light emitting diodes containing Langmuir-Blodgett films of copolymer of a poly(p-phenylene-vinylene) derivative and poly(octaneoxide)

The properties of Langmuir and Langmuir-Blodgett (LB) films from a block copolymer with polyethylene oxide and phenylene-vinylene moieties are reported. The LB films were successfully transferred onto several types of substrates, with sufficient quality to allow for evaporation of a metallic electrode on top of the LB films to produce polymer light emitting diodes (PLEDs). The photoluminescence and electroluminescence spectra of the LB film and device were similar, featuring an emission at ca. 475 nm, from which we could infer that the emission mechanisms are essentially the same as in poly(p-phenylene) derivatives. Analogously to other PLEDs the current versus voltage characteristics of the LB-based device could be explained with the Arkhipov model according to which charge transport occurs among localized sites. The implications for nanotechnology of the level of control that may be achieved with LB devices will also be discussed.

Nanopore size growth and ultrafiltration performance of SnO2 ceramic membranes prepared by sol-gel route

Supported ceramic membranes have been produced by the sol-casting procedure from aqueous colloidal suspensions prepared by the sol-gel route. Coatings on a tubular alumina support have been successfully performed leading to crack free layers. Samples have been sintered at 400, 500 and 600 degreesC, and the effect of heating treatment on the nanostructure and on the ultrafiltration properties are analyzed. The characterization has been done by high resolution scanning electron microscopy, nitrogen adsorption-desorption isotherms, water permeation and cut-off determination using polyethylene glycol standard solutions. The micrographs have revealed that grains and pore size increase with the temperature, whereas their shape remains invariant. This results is in agreements with N-2 adsorption-desorption analyses, which have revealed that the mean pore size diameter increases from 4 to 10 nm as the sintering temperature increases from 400 to 600 degreesC, while the total porosity remains constant. Furthermore, the tortuosity, calculated from water permeability, is essentially invariant with the sintering temperatures. The membranes cut-off, determined with a retention rate equal to 95%, are 3500, 6500 and 9000 g . mol(-1) for 400, 500 and 600 degreesC, respectively, showing that the permeation properties of SnO2 ultrafiltration membranes can easily be controlled by sintering condition.

Desenvolvimento e caracterização de suportes porosos de polietileno de ultra alto peso molecular (PEUAPM) para utilização como biomaterial para reposição e regeneração óssea

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

An efficient microwave-assisted hydrothermal synthesis of BaZrO3 microcrystals: growth mechanism and photoluminescence emissions

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

CuO urchin-nanostructures synthesized from a domestic hydrothermal microwave method

This letter reports the synthesis of CuO urchin-nanostructures by a simple and novel hydrothermal microwave method. The formation and growth of urchin-nanostructures is mainly affected by the addition of polyethylene glycol (PEG). The hierarchical malachite particles are uniform spheres with a diameter of 0.7-1.9 mu m. CuO urchin-nano structures were characterized by X-ray diffraction (XRD), field-emission scanning electron microscopy (FEG-SEM) and nitrogen adsorption (BET). The specific surface area of the CuO nanostructured microspheres was about 170.5 m(2)/g. A possible mechanism for the formation of such CuO urchin-nanostructures is proposed. (c) 2007 Elsevier Ltd. All rights reserved.

Survival of Ucides cordatus (Decapoda: Ocypodidae) megalopae during transport under different conditions of density and duration

Target areas for Ucides cordatus (Linnaeus, 1763) restocking programs are often located far from the laboratory where larval rearing is developed. During translocation, the larvae are submitted to highly stressful conditions due to handling, packing, and transport activities. The aim of the present study was to assess the mortality rates of U. cordatus megalopae caused by different transportation procedures. Megalopae at loading densities of 50, 150, and 300 ind.L-1 were packed in double polyethylene 12 x 25 cm plastic bags with 200 ml of marine water at salinity 30. The bags were filled with oxygen at a proportion of 1:2 parts of water and sealed tightly. The trepidations during transport were simulated by the use of a shaker device (800 vibrations/minute) over periods of three and six hours inside a dark container. The survivorship rates of larvae after simulation were compared to those obtained in control groups, which consisted of plastic vials with megalopae at a loading density of 50 ind.L-1 maintained at rest. Immediately after the two transport simulations, there was no significant difference in survivorship between the treatments and the control. However, 24 hours after simulation some of the tested densities resulted in significantly lower survivorships. The results demonstrated that U. cordatus megalopae can tolerate six hours of shaking during transportation, at high densities with minimal mortality.

Avaliação das alterações em polpa de manga 'Tommy-Atkins' congeladas

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Avaliação da qualidade de polpa de goiaba 'Paluma' armazenada a -20ºC

Neste estudo, goiabas da cultivar 'Paluma' foram cortadas em duas partes e submetidas a diferentes processos de conservação antes do armazenamento a -20ºC. No primeiro processo a polpa dura foi triturada, acondicionada em sacos de polietileno, com espessura de 40 µm, congelada e armazenada a -20°C°, e no segundo processo a polpa dura foi cortada ao meio, branqueada em água quente a 98ºC por quatro minutos, seca e acondicionada em sacos de polietileno, com espessura de 40 µm, congelada e armazenada a -20ºC. A qualidade da polpa foi avaliada através do pH, teor de ácido ascórbico, sólidos solúveis totais, acidez total titulável, aparência, textura e coloração. As duas formas de conservação mostraram-se adequadas para preservação da polpa com boa aparência até 18 semanas. Houve redução no conteúdo de ácido ascórbico, principalmente na polpa triturada. A textura e o sabor da polpa foram afetados pelo tempo de armazenamento. Os dois procedimentos, como técnicas de preservação de goiabas pelas indústrias de alimentos, podem ser usados durante o período de safra, sendo uma boa alternativa para evitar perdas pós-colheita.

Armazenagem em atmosfera modificada passiva de carambola azeda (Averrhoa carambola L.) cv. 'Golden Star'

Este trabalho avaliou o efeito das embalagens de polietileno de baixa densidade (PEBD), em diferentes espessuras, no prolongamento da vida útil pós-colheita de carambolas cv. 'Golden Star'. Os frutos foram colhidos fisiologicamente maturos, sendo selecionados pela ausência de defeitos e acondicionados nas embalagens de PEBD, constituindo os seguintes tratamentos: T1 - controle (sem embalagem); T2 - PEBD de 6 mim; T3 - PEBD de 10 mim; T4 - PEBD de 15 mim. Os frutos foram armazenados a 12 +/- 0,5 ºC e 95 +/- 5% de UR, por 45 dias, e mais 5 dias a 22 +/- 3ºC e 72 +/- 5% de UR. Vinte e quatro horas após a colheita e a cada nove dias, amostras eram retiradas do armazenamento refrigerado (AR), mantidas por 12 horas em condições ambiente (22 +/- 3ºC e 72 +/- 5% UR) e analisadas quanto à firmeza de polpa (FP), à perda de massa fresca, à coloração da epiderme, aos sólidos solúveis totais (SST), à acidez total titulável (ATT) e à ocorrência de distúrbios fisiológicos. Realizou-se também uma análise sensorial ao final do experimento. A maior firmeza de polpa e acidez total titulável, o melhor padrão de coloração, o menor conteúdo de sólidos solúveis totais, a ausência de manchas e podridões, e a melhor aceitabilidade pelos julgadores foram obtidos com os frutos acondicionados em embalagens de 10 mim.