Purificação, caracterização e análise da atividade bioinseticida, de um inibidor de tripsina em sementes de catanduva (Piptadenia moniliformis)

One Kunitz-type trypsin inhibitors (PmTI) was purified from Piptadenia moniliformis seeds, a tree of the sub-family Mimosoideae, by TCA precipitation, affinity chromatography on immobilized trypsin-Sepharose, DEAE cellulose (ion exchange) and Superose 12 (molecular exclusion) column FPLC/AKTA. The inhibitor has Mr of 25 kDa by SDS-PAGE and chromatography molecular exclusion. The N-terminal sequence of this inhibitor showed high homology with other family Kunitz inhibitors. This also stable variations in temperature and pH and showed a small decrease in its activity when incubated with DDT in the concentration of 100mM for 120 minutes. The inhibition of trypsin by PmTI was competitive, with Ki of 1.57 x10-11 M. The activity of trypsin was effectively inhibited by percentage of inhibition of 100%, among enzymes tested, was not detected inhibition for the bromelain, was weak inhibitor of pancreatic elastase (3.17% of inhibition) and inhibited by 76.42% elastase of neutrophils, and inhibited in a moderate, chymotrypsin and papain with percentage of inhibition of 42.96% and 23.10% respectively. In vitro assays against digestive proteinases from Lepidoptera, Diptera and Coleoptera pests were carried out. Several degrees of inhibition were found. For Anthonomus grandis and Ceratitis capitata the inhibition was 89.93% and 70.52%, respectively, and the enzymes of Zabrotes subfasciatus and Callosobruchus maculatus were inhibited by 5.96% and 9.41%, respectively, and the enzymes of Plodia. interpunctella and Castnia licus were inhibited by 59.94% and 23.67, respectively. In vivo assays, was observed reduction in the development of larvae in 4rd instar of C. capitata, when PmTI was added to the artificial diet, getting WD50 and LD50 of 0.30% and 0.33%, respectively. These results suggest that this inhibitor could be a strong candidate to plant management programs cross transgenic

Purificação, caracterização e avaliação da atividade antiproliferativa de um inibidor de quimotripsina tipo kunitz de semente de Eryhrina velutina

A chymotrypsin inhibitor was purified from Erythrina velutina seeds by ammonium sulphate fractionation, affinities chromatographies on Trypsin-Sepharose, Quimotrypsin-Sepharose and reversed phase C-18 FPLC/AKTA system. The inhibitor, named EvCI, shown molecular mass of 17 kDa, as determined by SDSPAGE. 2D-PAGE showed four isoinhibitors with pI values of 4,42, 4,63, 4,83 and 5,06, with molecular mass of 17 kDa each. The aminoacid sequence of EvCI was determined by MALDI-TOF-MS and showed a high similarity with other Kunitz-type inhibitor of Erythrina variegata. EvCI competitively inhibited chymotrypsin, with Ki of 4 x10-8 M, but did not inhibited trypsin, pancreatic elastase, bromelain and papain. The inhibitory activity of EvCI was stable over wide pH and temperature ranges. In the presence of DTT 100 mM for 120 min, EvCI lost 50 % of activity. Cytotoxicity was studied in HeLa, MDA, HepG2, K562 and PC3 cells after 72-h incubation period. EvCl inhibited HeLa cells growth with an IC50 value of 50 μg/ml. Subsequent studies in HeLa cells analysis of cell death by annexin V/PI double-staining and cell cycle, using flow cytometry. The results provide evidence for a cytostatic activity of EvCl and support further studies on potential application of this inhibitors as an antiproliferative agent in combined therapy against cervical cancer

Obtenção de nanopartículas de hexaferrita de bário pelo método pechini

In this study barium hexaferrite was (general formulae BaFe12O19) was synthesized by the Pechini method under different conditions of heat treatment. Precursors like barium carbonate and iron nitrate were used. These magnetic ceramic, with magnetoplumbite type structure, are widely used as permanent magnet because of its excellent magnetic properties, such as: high Curie temperature, good magnetic anisotropy, high coercivity and corrosion resistance. The samples were characterized by thermal analysis (DTA and TG), X- ray Diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), Scanning Electron Microscopy (SEM) end Vibrating sample Magnetometer (VSM). The results confirm the expected phase, which was reinforced according to our analysis. A single phase powder at relatively high temperatures with particle sizes around 100 nm was obtained. The characteristic magnetic behavior one of the phases has been noted (probably superparamagnetic material), while another phase was identified as a ferrimagnetic material. The ferrimagnetic phase showed vortex configuration with two central and slightly inclined plateaus. In general, increase of heat treatment temperature and time, directly influenced the technological properties of the samples

Proteínas SAG1, SAG2, SAG3 de toxoplasma gondii como perspectivas para o desenvolvimento de protótipo vacinal contra a toxoplasmose

Toxoplasmosis is a zoonosis of worldwide distribution caused by the protozoan Toxoplasma gondii, triggering dangerous complications in immunocompromised patients and pregnant women, as well as having great economic impact for the livestock. So far the control of toxoplasmosis is made primarily by chemotherapy. However, most drugs used routinely have some limitations. In order to control this disease, several research groups, including ours, has been working to develop a medical-veterinary vaccine based on parasite antigens, vectors and protocols of immunization. In this study were implemented and standardized methodologies for amplification and cloning of recombinant immunogens in the system for the development of a prototype vaccine, based on the surface antigens of T. gondii and recombinant adenovirus encoding these antigens. Genes encoding BAG1, GRA2 and SAG1 proteins were amplified. We established a strategy for cloning SAG1, SAG2, SAG3 and TgAMA1- genes in recombinant system. The genes encoding SAG1 and SAG2 were cloned and their sequences showed high similarity with sequences from GenBank. The virtual translation of these proteins showed polymorphisms in the amino acid sequence, which can be correlated with levels of antigenicity. Simultaneously, the adenovirus encoding the SAGs (HAdSAGs) were expanded, purificated and characterizated. Immunization of C57bl/6 mice, using viral supernatant was not enought to elicit immune responses at high levels, being required HAdSAGs titration for future immunizations. Therefore, this study allowed the cloning of the two genes important for the development of a prototype vaccine. Besides, implementations methodologies that permit advancements in the development of a vaccine against toxoplasmosis using adenovirus to express proteins of the parasite

Obtenção de ferrita de cobalto através de dois métodos de síntese: método de complexação combinando EDTA/Citrato e método hidrotérmico

In this work it was synthesized and characterized the cobalt ferrite (CoFe2O4) by two methods: complexation combining EDTA/Citrate and hydrothermal investigating the influence of the synthesis conditions on phase formation and on the crystallite size. The powders were mainly characterized by x-ray diffraction. In specific cases, it was also used scanning electron microscopy (SEM), energy dispersive spectroscopy (EDS), x-ray fluorescence (XRF) and isotherms of adsorption and desorption of nitrogen (BET method). The study of the crystallite size was based on the interpretation of x-ray diffractograms obtained and estimated by the method of Halder-Wagner-Scherrer and Langford. An experimental design was made in order to assist in quantifying the influence of synthesis conditions on the response variables. The synthesis parameters evaluated in this study were: pH of the reaction medium (8, 9 and 10), the calcination temperature (combined complexation method EDTA/Citrate 600°C, 800°C and 1000°C), synthesis temperature (hydrothermal method 120°C, 140°C and 160°C), calcination time (combined complexation method EDTA/Citrate - 2, 4 and 6 hours) and time of synthesis (hydrothermal method 6, 15 and 24 hours). By the hydrothermal method was possible to produce mesoporous powders with high purity, with an average crystallite size up to 7 nm, with a surface area of 113.44 m²/g in the form of pellets with irregular morphology. By using the method of combined complexation EDTA/Citrate, mesoporous powders were produced with greater purity, crystallite size up to 22nm and 27.95 m²/g of surface area in the form of pellets with a regular morphology of plaques. In the experimental design was found that the hydrothermal method to all the studied parameters (pH, temperature and time) have significant effect on the crystallite size, while to the combined complexation method EDTA/Citrate, only temperature and time were significant

Hidrólise e caracterização de poliacrilamida hidrofobicamente modificada: avaliação da aplicabilidade na recuperação de petróleo

The hydrolysis reaction in alkaline conditions of the commercial polymer poly(acrylamide-co-metacrylate of 3,5,5-trimethyl-hexane) called HAPAM, containing 0.75 % of hydrophobic groups, was carried out in 0.1 M NaCl and 0.25M NaOH solutions, varying the temperature and reaction time. The polymers were characterized by 1H and 13C Nuclear Magnetic Resonance (NMR), Elemental Analysis and Size Exclusion Chromatography (SEC). The values of the hydrolysis degree were obtained by 13C NMR. The viscosity of HAPAM and HAPAM-10N-R solutions was evaluated as a function of shear rate, ionic strength and temperature. At high polymer concentration (Cp), the viscosity of HAPAM solutions increased with the ionic strength and decreased with the temperature. The viscosity of HAPAM-10N-R solutions increased significantly in distilled water, due to repulsions between the carboxylate groups. At high Cp, with the increase of ionic strength and temperature, occurred a decrease of viscosity, due to mainly the high hydrolysis degree and the low amount of hydrophobic groups. These results indicated that the studied polymers have properties more suitable for the application in Enhanced Oil Recovery (EOR) in low salinity and moderate temperature reservoirs