Discovery and Structure Activity Relationship of Small Molecule Inhibitors of Toxic β-Amyloid-42 Fibril Formation

Increasing evidence implicates Aβ peptides self-assembly and fibril formation as crucial events in the pathogenesis of Alzheimer disease. Thus, inhibiting Aβ aggregation, among others, has emerged as a potential therapeutic intervention for this disorder. Herein, we employed 3-aminopyrazole as a key fragment in our design of non-dye compounds capable of interacting with Aβ42 via a donor-acceptor-donor hydrogen bond pattern complementary to that of the β-sheet conformation of Aβ42. The initial design of the compounds was based on connecting two 3-aminopyrazole moieties via a linker to identify suitable scaffold molecules. Additional aryl substitutions on the two 3-aminopyrazole moieties were also explored to enhance π-π stacking/hydrophobic interactions with amino acids of Aβ42. The efficacy of these compounds on inhibiting Aβ fibril formation and toxicity in vitro was assessed using a combination of biophysical techniques and viability assays. Using structure activity relationship data from the in vitro assays, we identified compounds capable of preventing pathological self-assembly of Aβ42 leading to decreased cell toxicity.     

Neuroprotective and anticonvulsant effects of organic and conventional purple grape juices on seizures in Wistar rats induced by pentylenetetrazole

Epilepsy is the most common neurological disorder worldwide. Studies have shown that recurrent seizures may increase the concentration of reactive oxygen species, which can lead to oxidative stress and neuronal damage. These seizures result in substantial deleterious effects on an individual's health. Organic and conventional grape juices are rich in polyphenols, compounds with important antioxidant activity. However, these juices could have differences in their polyphenol content. The aim of this study was to investigate the neuroprotective and anticonvulsant effects of organic and conventional grape juice treatments in Wistar rats against pentylenetetrazole (a convulsant drug)-induced damage. In addition, we evaluated potential behavioral changes in rats treated with the juices and the polyphenolic profile of those samples. Animals (n=16 in each group) received treatment with saline, organic or conventional grape juice for 17 days. On the eighteenth day, behavioral changes were evaluated by an open field test. Afterwards, half of the rats from each group received pentylenetetrazole and were observed for 30 min to evaluate possible seizure characteristics. The animals were subsequently killed by decapitation and their hippocampus, cerebellum and cerebral cortex tissues were isolated. The results of this study showed that neither organic nor conventional grape juice altered the behavior parameters, and no statistical differences were observed in the seizure characteristics of the groups. Nevertheless, both juice types were able to protect from lipid and protein oxidative damage, decrease nitric oxide content and increase enzymatic (superoxide dismutase and catalase) and non-enzymatic (sulfhydryl protein) antioxidant defenses in brain tissues following pentylenetetrazole-induced seizures. In general, organic juice showed superior results in each test, probably due to its higher polyphenol content relative to conventional juice. These results indicate that grape juices can provide further insight into natural neuroprotective compounds and may lead to the development of new therapeutic strategies for epileptic patients.     

Orgenic plants: Gene‐manipulated plants compatible with organic farming

Based on recent advances in plant gene technology, I propose to develop a new category of GM plants, orgenic plants, that are compatible with organic farming. These orgenic plants do not contain herbicide resistance genes to avoid herbicide application in agriculture. Furthermore, they either contain genes that are naturally exchanged between species, or are sterile to avoid outcrossing if they received a transgene from a different species. These GM plants are likely to be acceptable to most sceptics of GM plants and facilitate the use of innovative new crops.     

Physiological responses to salt stress in young umbu plants

Soil salinity affects plant growth and development due to harmful ion effects and water stress caused by reduced osmotic potential in the soil solution. In order to evaluate the effects of salt stress in young umbu plants, research was performed in green house conditions at the Laboratory of Plant Physiology at Federal Rural University of Pernambuco, Brazil. Growth, stomatal behaviour, water relations, and both inorganic and organic solutes were studied aiming for a better understanding of the responses of umbu plants to increasing salinity. Plants were grown in washed sand with Hoagland and Arnon nutrient solution with 0, 25, 50, 75, and 100 mM NaCl. Growth, leaf water potential, transpiration, and diffusive resistance were evaluated. Na⁺, K⁺, Cl⁻, soluble carbohydrates, and free amino acid contents were measured in several plant organs. Most variables were affected with salinity above 50 mM NaCl showing decreases in: number of leaves, plant height, stems diameter, and dry masses, and increases in root-to-shoot ratio. Reductions in ψ_pd were observed in plants grown under 75 and 100 mM NaCl. All salt levels above zero increased Na⁺ and Cl⁻ contents in leaves. However, K⁺ content was not affected. Na⁺ and Cl⁻ in stems and roots reached saturation in treatments above 50 mM NaCl. Organic solute accumulation in response to salt stress was not observed in umbu plants. These results suggest that umbu plants tolerate salt levels up to 50 mM NaCl without showing significant physio-morphological alterations.     

Overexpression of prefoldin from the hyperthermophilic archaeum Pyrococcus horikoshii OT3 endowed Escherichia coli with organic solvent tolerance

Prefoldin is a jellyfish-shaped hexameric chaperone that captures a protein-folding intermediate and transfers it to the group II chaperonin for correct folding. In this work, we characterized the organic solvent tolerance of Escherichia coli cells that overexpress prefoldin and group II chaperonin from a hyperthermophilic archeaum, Pyrococcus horikoshii OT3. The colony-forming efficiency of E. coli cells overexpressing prefoldin increased by 1,000-fold and decreased the accumulation of intracellular organic solvent. The effect was impaired by deletions of the region responsible for the chaperone function of prefoldin. Therefore, we concluded that prefoldin endows E. coli cells by preventing accumulation of intracellular organic solvent through its molecular chaperone activity.     

Stabilization of water-in-oil emulsion by <Emphasis Type="Italic">Rhodococcus opacus</Emphasis> B-4 and its application to biotransformation

Rhodococcus opacus B-4, which has recently been isolated as an organic solvent-tolerant bacterium, stabilized water-in-oil (w/o) emulsions by inhibition of droplet coalescence when the cells were dispersed in 90% (v/v) organic solvents. Confocal microscopy revealed that many bacterial cells assembled at the interface between oil and water droplets, though free cells were also detectable at the inside of water droplets. Bacterial cells in the w/o emulsion were capable of utilizing both a water-soluble (glucose) and an oil-soluble substrate (oleic acid) as an energy source. Availability of the w/o emulsion as an immobilized cell system in organic solvents was demonstrated using production of indigo from indole and production of o-cresol from toluene as model conversions. When glucose and oleic acid were simultaneously supplied as energy sources, the w/o emulsion culture of R. opacus B-4 produced indigo and o-cresol at levels of 0.217 and 2.12 mg ml⁻¹, respectively, by 12 h.     

Composting of poultry manure and wheat straw in a closed reactor: optimum mixture ratio and evolution of parameters

The main objectives of this work were to investigate the evolution of some principal physico-chemical properties (temperature, carbon dioxide, oxygen, ammonia, pH, electrical conductivity, organic matter) and microbial population (mesophilic and thermophilic bacteria and fungi) during composting poultry manure with wheat straw in a reactor system, and to evaluate the optimum mixture ratio for organic substrate production. The experiments were carried out in four small laboratory reactors (1 l) and one large reactor (32 l) under adiabatic conditions over 14 days. During the process the highest temperature was 64.6°C, pH varied between 7.40 and 8.85, electrical conductivity varied between 3.50 and 4.31 dS m⁻¹ and the highest value of organic matter (dry weight) degradation was 47.6%. Mesophilic bacteria and fungi predominated in the beginning, and started the degradation with generation of metabolic heat. By increasing the temperature in reactors, the number of thermophilic microorganisms also increased, which resulted in faster degradation of substrate. The application of a closed reactor showed a rapid degradation of manure/straw mixture as well as a good control of the emissions of air polluting gases into atmosphere. The results showed that the ratio of manure to straw 5.25:1 (dry weight) was better for composting process than the other mixture ratios.     

Biocatalytic properties of native and immobilized subtilisin 72 in aqueous-organic and low water media

Subtilisin 72 was immobilized on cryogel of poly(vinyl alcohol), the macroporous carrier prepared by the freeze-thaw-treatment of concentrated aqueous solution of the polymer. The obtained biocatalyst was active and stable in aqueous, aqueous-organic, as well as in low water media. The stability of immobilized biocatalyst was substantially higher than that of native enzyme in all mixtures especially in aqueous buffer containing 5–8 M Urea and in acetonitrile/60–90%DMF mixtures. The ability of native and immobilized subtilisin to catalyze peptide bond formation between Z-Ala-Ala-Leu-OMe and Phe-pNA was studied in non-aqueous media. Considerable enzyme stabilization in acetonitrile/90%DMF mixture, induced by the immobilization, resulted in higher product yield (57%) than in case of native subtilisin suspension (32%). Detailed study of synthesis reaction revealed that notable increase in product yield could be reached using increase in both substrate concentrations up to 200 mM.     

Interaction of nitrogen source and light intensity on the growth and photosynthesis of the brown tide alga Aureococcus anophagefferens

High ratios of dissolved organic nitrogen (DON) to dissolved inorganic nitrogen (DIN) have been suggested to favor the growth of the brown tide alga Aureococcus anophagefferens. DON could provide a particular advantage in low light levels, as occur when blooms induce self-shading. We examined the effects of varying DON:DIN ratios on the photosynthetic abilities of cultured Aureococcus at two light intensities, 93 and 17 μmol photons m⁻² s⁻¹. Glutamic acid and urea were used as DON sources, and the remainder of the nitrogen was added as nitrate.In experiments examining Aureococcus growth with varying ratios of DON_Glu:DIN_Nitrate at two light intensities in batch culture, higher growth rates and biomass were observed in treatments containing DIN than in those with DON only, which contrasts with the results of previous studies. In semi-continuous growth experiments with varying DON_Urea:DIN_Nitrate ratios, low light cultures with urea had higher growth rates than those without urea. Also, the effective target area for light absorption per cell and photosystem II efficiency were greater for the low light cultures of each nutrient treatment, particularly when DON:DIN mixtures (33 and 67% N_Urea) were used. The same pattern was seen in the maximum photosynthetic rates per cell in the light-saturated (P_m^cell) and in the initial slope (α^cell) of the PE (photosynthesis versus irradiance) curve, and in PON, POC and chlorophyll a cell⁻¹. This indicates that the ability of Aureococcus to acclimate to low light conditions may be enhanced by the presence of both organic and inorganic nitrogen sources. These results suggest that Aureococcus physiology and photosynthesis are different during growth on a mixture of urea-N and nitrate than when either nitrogen source is present alone. Results of this study suggest that Aureococcus may not respond to all DON substrates in the same way, and that mixtures of DON and DIN may provide for higher photosynthetic rates, especially at low light. Our results did not, however, support earlier suggestions that growth on DON alone provides the brown tide alga with a large advantage at low light levels.