Síntese e caracterização de nanopartículas vetorizadas para células tumorais com possível aplicação na entrega controlada de fármacos

Dissertação de mestrado em Química Medicinal

Infection by microorganisms on soybean seeds obtained from plants treated with growth regulators

This research, deals with the effects of exogenous growth regulators on infection by microorganisms on soybean (Glycine max cv. Davis) seeds. To study the influence of the chemicals, soybean plants were sprayed with gibberellic acid (GA) 100 ppm, (2-chloroethyl) trimethylammonium chloride (CCC) 2,000 ppm, succinic acid-2,2-dimethy1hydrazide (SADH) 4,000 ppm, indolylacetic acid (IAA) 100 ppm, 2,3,5-triiodobenzoic acid (TIBA) 20 ppm (three applications), and Agrostemin (1g/10 ml/ 3 1). Application of growth regulators did not affect infect ion by microorganisms on soybean seeds. The prominent fungus isolated was Phomopsis sojae. Alternaria and Fusarium spp. were isolated from seeds. The presence of a bacterium on the seeds was observed. The delay in harvest and high humidity increased the number of seeds from which Phomopsis was recovered.

Action of growth regulators on flowering and pod production of soybean cultivar Davis

This research deals with the effects of growth regulators on flowering and pod formation in soybean plant (Glycine max cv. Davis). Under greenhouse conditions, soybean plants were sprayed with 2,3,5-triiodobenzoic acid (TIBA) 20 ppm, Agrostemmin (1g/10 ml/3 l) gibberellic acid (GA) 100 ppm, and (2-chloroethyl) trimethylammonium chloride (CCC) 2,000 ppm. Application of TIBA increased number of flowers. 'Davis' soybean treated with CCC and TIBA presented a tendency to produce a lower number of pods.

Action of growth regulators on production of soybean cultivar Davis

This research deals with the effects of exogenous growth regulators on production of soybean plant (Glycine max cv.. Davis) under greenhouse conditions, At the flower anthesis, 2,3,5-triiodobenzoic acid (TIBA) 20 ppm was applied. Other two applications with TiBA, with intervals of four days, were realized. Before flowering, Agrostemin (1 g/10 ml/3 1), gibberellic acid (GA) 100 ppm, and (2-chloroethyl) trimethylammonium chloride (CCC) 2,000 ppm were applied. It was observed that CCC and TIBA reduced stem dry weight. Soybean plants treated with TIBA reduced weight of pods without seeds , seed number and seed weight.

Action of growth regulators on soybean cultivar Davis

The effects of growth regularots on soybean plant (Glycine max) under greenhouse conditions were studied. Before flower ing, Agrostemmin (1 g/10 ml/3 1), gibberellic acid (GA) 100 ppm, and (2-chloroethyl) trimethylammonium chloride (CCC) 2,000 ppm were applied. At the flower anthesis, 2,3,5 - triio dobenzoic acid (TIBA) 20 ppm was applied. Other two applications with TIBA, with intervals of four days, were realized. Treatment with GA increased plant height while CCC presented a tendency to reduce it. Numbers of leaves, internods, and stems were not affected by the growth regulators.

Efeito da temperatura na germinação de sementes de vigna, feijoeiro e soja

Estudou-se em condições controladas, o efeito das temperaturas de 14, 21 e 28°C na germinação de três cultivares de Vigna unguiculata, Phaseolus vulgaris e Glycine max. As sementes foram acondicionadas em placas de Petri com algodão e papel de filtro , mantidos sob alta umidade. Maior germinação foi observada a 21°C em relação as temperaturas de 14 e 28°C, para os cultivares de vigna, feijoeiro e soja. Melhor germinação foi apresentada pelos cultivares de feijoeiro Carioca e Goiano Precoce, seguidos pelo feijoeiro 'Rosinha' , vigna ' EPACE 1' e soja 'Davis' a 21ºC. Os cultivares de vigna, adaptados a altas temperaturas, apresentaram problemas na germinação a 14ºC. Normalmente a velocidade de emergência do hipocólito revelou-se inferior a velocidade de emergência da radícula.

Efeitos do alumínio sobre o desenvolvimento e a composição mineral de quatro leguminosas forrageiras tropicais

Quatro leguminosas forrageiras tropicais (Galactia striata (Jacq) Urb. , Glycine wightii Verde. cv. Tinaroo, Macvoptilium atropurpureus cv. Siratro e Stylosanthes guianensis (Avbl) Swartz cv. IRI 1022) foram cultivadas em solução nutritiva recebendo doses de alumínio de 0,5 , 10 e 20 ppm, com o objetivo de se verificarem os efeitos das doses crescentes do elemento sobre as díferentes espécies. Submetendo-se os dados de produção de matéria seca e de concentrações de macro e micronutrientes à análise estatística, observou-se o seguinte: a) O estilosantes e a galactia mostraram-se mais tolerantes. A galactia apresentou a máxima produção em presença de 5 ppm de Alumínio, não sendo afetada pelas concentrações mais elevadas. O estilosantes apresentou a máxima produção em presença de 20 ppm de Alumínio. b) A soja e o siratro mostram-se sensíveis, notando-se uma queda na produção de matéria seca em presença de 20 ppm de Alumínio na solução. A soja mostrou sintomas mais graves de toxidez, apresentando seu sistema radicular totalmente danificado na concentração mais alta de Alumínio na solução. c) A adição de Alumínio provocou uma diminuição nas concentrações de fósforo, cálcio e magnésio nas raízes ou partes aéreas das plantas e de manganês nas raízes e uma elevação na concentração de Alumínio nas raízes.

Efeitos do manganês sobre o desenvolvimento e a composição mineral de quatro leguminosas forrageiras tropicais

Quatro leguminosas forrageiras tropicais (Galactia striata (Jacq) Urb; Glycine wightii cv. Tinaroo, Macroptilium atropurpureum cv. Siratro e Stylosanthes guianensis cv. IRI 1022) foram cultivadas em solução nutritiva recebendo doses de manganês de 0, 5, 10 e 20 ppm, com o objetivo de se verificarem os efeitos das doses crescentes do elemento sobre as diferentes espécies. Submetendo-se os dados de produção de matéria seca e de concentrações de macro e micronutrientes à análise estatística, observou-se o seguinte: a) O estilosantes foi o que se mostrou mais tolerante, não apresentando variação significativa na produção de matéria seca com o aumento dos teores de manganês. Não se observaram sintomas severos nas folhas ou nas raízes. A galactia também mostrou-se tolerante, apresentando queda significativa de produção em presença de 100 ppm de manganês e sentomas mais severos de toxidez que os apresentados pelo estilosantes. b) O Siratro e a soja mostraram-se sensíveis ao excesso de manganês. A soja mostrou a mínima produção em presença de 25 ppm do elemento não havendo modificação significativa nas concentrações mais elevadas. O siratro apresentou a mínima produção de matéria seca total em presença de 50 ppm de manganês. Os sintomas de toxidez apresentados pela soja foram mais severos que os apresentados pelas plantas de siratro. c) A adição de manganês provocou uma diminuição nas concentrações de cálcio, potássio e magnésio nas partes aéreas ou raízes das plantas estudadas; elevação dos teores de zinco e cobre nas aéreas ou raízes e de manganês na planta toda.

The use of triphenyltetrazolium chloride in the study of dehydrogenase activity of Brucellae

Experiments for the investigation of dehydrogenase activity of washed cells of a strains of Br. abortus and another of Br. suis in presence of different single added substrates are reported. The activity was measured as the amount of formazan produced by the reduction of 2, 3, 5-triphenyltetrazolum chloride acting as a hydrogen ions acceptor, at pH 7.0. In a general manner the dehydrogenase activity of Br. suis was much more intense than that of Br. abortus (fig. 5). In the conditions of the experiments Br. abortus oxidized L-arabinose, D-galactose, D-glucose, glycerol, D-xylose, DL-alanine, D-fructose, and D-sorbitol. Brucella suis oxidized D-xylose, L-arabinose, D-glucose, D-galactose, DL-alanine, sodium acetate, maltose, glycine, D-fructose, and D-sorbitol. Glycerol was oxidized by Br. abortus but its oxidation by Br. suir was very slight. Sodium acetate and maltose were intensely oxidized by Br. suir but not by Br. abortus. The sites of more intense enzymatic acitivity were seen as small red colored round granules located in one pole of the cells.

Quantitative method of viral pollution determination for large volume of water using ferric hydroxide gel impregnated on the surface of glassfibre cartridge

Quantitative method of viral pollution determination for large volume of water using ferric hydroxide gel impregnated on the surface of glassfibre cartridge filter. The use of ferric hydroxide gel, impregnated on the surface of glassfibre cartridge filter enable us to recover 62.5% of virus (Poliomylitis type I, Lsc strain) exsogeneously added to 400 liters of tap-water. The virus concentrator system consists of four cartridge filters, in which the three first one are clarifiers, where the contaminants are removed physically, without significant virus loss at this stage. The last cartridge filter is impregnated with ferric hydroxide gel, where the virus is adsorbed. After the required volume of water has been processed, the last filter is removed from the system and the viruses are recovered from the gel, using 1 liter of glycine/NaOH buffer, at pH 11. Immediately the eluate is clarified through series of cellulose acetate membranes mounted in a 142mm Millipore filter. For the second step of virus concentration, HC1 1N is added slowly to the eluate to achieve pH 3.5-4. MgC1, is added to give a final concentration of 0.05M and the viruses are readsorbed on a 0.45 , porosity (HA) cellulose acetate membrane, mounted in a 90 mm Millipore filter. The viruses are recovered using the same eluent plus 10% of fetal calf serum, to a final volume of 3 ml. In this way, it was possible to concentrate virus from 400 liters of tap-water, into 1 liter in the first stage of virus concentration and just to 3 ml of final volume in a second step. The efficiency, simplicity and low operational cost, provded by the method, make it feasible to study viral pollution of recreational and tap-water sources.

Resting metabolic rate and protein turnover in apparently healthy elderly Gambian men.

Body composition, resting energy expenditure (REE), and whole body protein metabolism were studied in 26 young and 28 elderly Gambian men matched for body mass index during the dry season in a rural village in The Gambia. REE was measured by indirect calorimetry (hood system) in the fasting state and after five successive meals. Rates of whole body nitrogen flux, protein synthesis, and protein breakdown were determined in the fed state from the level of isotopic enrichment of urinary ammonia over a period of 12 h after a single oral dose of [15N]glycine. Expressed in absolute value, REE was significantly lower in the elderly compared with the young group (3.21 +/- 0.07 vs. 4.04 +/- 0.07 kJ/min, P < 0.001) and when adjusted to body weight (3.29 +/- 0.05 vs. 3.96 +/- 0.05 kJ/min, P < 0.0001) and fat-free mass (FFM; 3.38 +/- 0.01 vs. 3.87 +/- 0.01 kJ/min, P < 0.0001). The rate of protein synthesis averaged 207 +/- 13 g protein/day in the elderly and 230 +/- 13 g protein/day in the young group, whereas protein breakdown averaged 184 +/- 13 g protein/day in the elderly and 203 +/- 13 g protein/day in the young group (nonsignificant). When values were adjusted for body weight or FFM, they did not reveal any difference between the two groups. It is concluded that the reduced REE adjusted for body composition observed in elderly Gambian men is not explained by a decrease in protein turnover.

Immuno-reactive proteins from Mycobacterium immunogenum useful for serodiagnosis of metalworking fluid hypersensitivity pneumonitis.

Metalworking fluid-associated hypersensitivity pneumonitis (MWF-HP) is a pulmonary disease caused by inhaling microorganisms present in the metalworking fluids used in the industrial sector. Mycobacterium immunogenum is the main etiological agent. Among the clinical, radiological and biological tools used for diagnosis, serological tests are important. The aim of this study was to identify immunogenic proteins in M. immunogenum and to use recombinant antigens for serological diagnosis of MWF-HP. Immunogenic proteins were detected by two-dimensional Western blot and candidate proteins were identified by mass spectrometry. Recombinant antigens were expressed in Escherichia coli and tested by enzyme-linked immunosorbent assay (ELISA) with the sera of 14 subjects with MWF-HP and 12 asymptomatic controls exposed to M. immunogenum. From the 350 spots visualized by two-dimensional gel electrophoresis with M. immunogenum extract, 6 immunogenic proteins were selected to be expressed as recombinant antigens. Acyl-CoA dehydrogenase antigen allowed for the best discrimination of MWF-HP cases against controls with an area under the receiver operating characteristics (ROC) curve of 0.930 (95% CI=0.820-1), a sensitivity of 100% and a specificity of 83% for the optimum threshold. Other recombinant antigens correspond to acyl-CoA dehydrogenase FadE, cytosol aminopeptidase, dihydrolipoyl dehydrogenase, serine hydroxymethyltransferase and superoxide dismutase. This is the first time that recombinant antigens have been used for the serodiagnosis of hypersensitivity pneumonitis. The availability of recombinant antigens makes it possible to develop standardized serological tests which in turn could simplify diagnosis, thus making it less invasive.

A New Large-DNA-Fragment Delivery System Based on Integrase Activity from an Integrative and Conjugative Element.

During the past few decades, numerous plasmid vectors have been developed for cloning, gene expression analysis, and genetic engineering. Cloning procedures typically rely on PCR amplification, DNA fragment restriction digestion, recovery, and ligation, but increasingly, procedures are being developed to assemble large synthetic DNAs. In this study, we developed a new gene delivery system using the integrase activity of an integrative and conjugative element (ICE). The advantage of the integrase-based delivery is that it can stably introduce a large DNA fragment (at least 75 kb) into one or more specific sites (the gene for glycine-accepting tRNA) on a target chromosome. Integrase recombination activity in Escherichia coli is kept low by using a synthetic hybrid promoter, which, however, is unleashed in the final target host, forcing the integration of the construct. Upon integration, the system is again silenced. Two variants with different genetic features were produced, one in the form of a cloning vector in E. coli and the other as a mini-transposable element by which large DNA constructs assembled in E. coli can be tagged with the integrase gene. We confirmed that the system could successfully introduce cosmid and bacterial artificial chromosome (BAC) DNAs from E. coli into the chromosome of Pseudomonas putida in a site-specific manner. The integrase delivery system works in concert with existing vector systems and could thus be a powerful tool for synthetic constructions of new metabolic pathways in a variety of host bacteria.

A corneal dystrophy associated with transforming growth factor beta-induced Gly623Asp mutation an amyloidogenic phenotype.

PURPOSE: To present the light and electron microscopic findings of a unique corneal dystrophy never before described in a German family carrying the Gly623Asp Mutation of the TGFBI gene with late clinical onset. DESIGN: Experimental study. PARTICIPANTS: Four affected and 6 nonaffected family members. METHODS: Slit-lamp examination, photographic documentation, and isolation of genomic DNA from peripheral blood leucocytes obtained from each family member examined. Exons 3, 4, 5, and 11 to 14 of the TGFBI gene were amplified and sequenced in these family members. Five corneal buttons of 3 affected siblings were excised at the time of penetrating keratoplasty. Light and electron microscopic examination were performed including immunohistochemistry with antibodies against keratoepithelin (KE) 2 and 15. MAIN OUTCOME MEASURES: Clinical and histologic characteristics of corneal opacification in affected patients and presence of coding region changes in the TGFBI gene. RESULTS: The specimens showed destructive changes in Bowman's layer and the adjacent stroma. Patchy Congo red-positive amyloid deposits were found within the epithelium in 1 cornea, in Bowman's layer and in the anterior stroma of all specimens also showing KE2, but not KE15, immunostaining. Electron microscopy revealed deposits mainly located in the anterior stroma and Bowman's layer and in small amounts in the basal area of some epithelial cells. The destroyed areas were strongly Alcian blue-positive, the Masson Trichrome stain proved mainly negative for the deposits. All affected but none of the unaffected family members had a heterozygous missense mutation in exon 14 of the TGFBI gene (G-->A transition at nucleotide 1915) replacing glycin by aspartic acid amino acid (Gly623Asp) at position 623 of the KE protein. CONCLUSIONS: In contrast with the patient carrying the Gly623Asp mutation of the TGFBI gene described by Afshari et al, our cases presented with Salzmann's nodular degeneration-like clinical features and their specimens contained KE2-positive amyloid. The reason for this now "meeting the expectation histologic phenotype" is unclear. The histologic findings emphasize that this is a unique corneal dystrophy, which shares no clinical characteristics with Reis-Bücklers' dystrophy and should be treated as a distinct entity. FINANCIAL DISCLOSURE(S): The authors have no proprietary or commercial interest in any materials discussed in this article.

Whole body protein metabolism and resting energy expenditure in pregnant Gambian women.

Whole body protein metabolism and resting energy expenditure (REE) were measured at 11, 23, and 33 wk of pregnancy in nine pregnant (not malnourished) Gambian women and in eight matched nonpregnant nonlactating (NPNL) matched controls. Rates of whole body nitrogen flux, protein synthesis, and protein breakdown were determined in the fed state from the level of isotope enrichment of urinary urea and ammonia during a period of 9 h after a single oral dose of [15N]glycine. At regular intervals, REE was measured by indirect calorimetry (hood system). Based on the arithmetic end-product average of values obtained with urea and ammonia, a significant increase in whole body protein synthesis was observed during the second trimester (5.8 +/- 0.4 g.kg-1.day-1) relative to values obtained both for the NPNL controls (4.5 +/- 0.3 g.kg-1.day-1) and those during the first trimester (4.7 +/- 0.3 g.kg-1.day-1). There was a significant rise in REE during the third trimester both in the preprandial and postprandial states. No correlation was found between REE after meal ingestion and the rate of whole body protein synthesis.