The use of biodiversity as source of new chemical entities against defined molecular targets for treatment of malaria, tuberculosis, and T-cell mediated diseases: a review

The modern approach to the development of new chemical entities against complex diseases, especially the neglected endemic diseases such as tuberculosis and malaria, is based on the use of defined molecular targets. Among the advantages, this approach allows (i) the search and identification of lead compounds with defined molecular mechanisms against a defined target (e.g. enzymes from defined pathways), (ii) the analysis of a great number of compounds with a favorable cost/benefit ratio, (iii) the development even in the initial stages of compounds with selective toxicity (the fundamental principle of chemotherapy), (iv) the evaluation of plant extracts as well as of pure substances. The current use of such technology, unfortunately, is concentrated in developed countries, especially in the big pharma. This fact contributes in a significant way to hamper the development of innovative new compounds to treat neglected diseases. The large biodiversity within the territory of Brazil puts the country in a strategic position to develop the rational and sustained exploration of new metabolites of therapeutic value. The extension of the country covers a wide range of climates, soil types, and altitudes, providing a unique set of selective pressures for the adaptation of plant life in these scenarios. Chemical diversity is also driven by these forces, in an attempt to best fit the plant communities to the particular abiotic stresses, fauna, and microbes that co-exist with them. Certain areas of vegetation (Amazonian Forest, Atlantic Forest, Araucaria Forest, Cerrado-Brazilian Savanna, and Caatinga) are rich in species and types of environments to be used to search for natural compounds active against tuberculosis, malaria, and chronic-degenerative diseases. The present review describes some strategies to search for natural compounds, whose choice can be based on ethnobotanical and chemotaxonomical studies, and screen for their ability to bind to immobilized drug targets and to inhibit their activities. Molecular cloning, gene knockout, protein expression and purification, N-terminal sequencing, and mass spectrometry are the methods of choice to provide homogeneous drug targets for immobilization by optimized chemical reactions. Plant extract preparations, fractionation of promising plant extracts, propagation protocols and definition of in planta studies to maximize product yield of plant species producing active compounds have to be performed to provide a continuing supply of bioactive materials. Chemical characterization of natural compounds, determination of mode of action by kinetics and other spectroscopic methods (MS, X-ray, NMR), as well as in vitro and in vivo biological assays, chemical derivatization, and structure-activity relationships have to be carried out to provide a thorough knowledge on which to base the search for natural compounds or their derivatives with biological activity.

Sucrose hydrolysis by gelatin-immobilized inulinase from Kluyveromyces marxianus var. bulgaricus

The crude cell-free medium from a culture of Kluyveromyces marxianus var. bulgaricus was immobilized in a gelatin-water support, with an immobilization yield of 82.60% for inulinase activity. The optimum pH for both free and immobilized inulinase was the same (3.5) and the optimum temperatures were 55 degrees C for the free and 60 degrees C for the immobilized enzyme. The Arrhenius plots were linear and activation energies were 56.20 (free enzyme) and 20.27 kj/mol K (immobilized enzyme). The kinetic parameters were calculated by Lineweaver-Burk plots and the V-max and K-m were 37.60 IU/mg protein and 61.83 mM for the free inulinase and 31.45 IU/mg protein and 149.28 mM for the immobilized enzyme, respectively. The operational stability of the immobilized inulinase was studied in a continuous fixed-bed column reactor for 33 days, at the end of which the sucrose conversion was 58.12%. (c) 2008 Elsevier Ltd. All rights reserved.

Synergistic effect in the catalytic activity of lipase Rhizomucor miehei immobilized on zeolites for the production of biodiesel

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

Enzymatic toxins from snake venom: structural characterization and mechanism of catalysis

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

Hydrolysis of whey lactose by immobilized β-Galactosidase

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

Amperometric Immunosensor for Chagas' Disease Using Gold CD-R Transducer

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

Quartz Crystal Microbalance monitoring the real-time binding of lectin with carbohydrate with high and low molecular mass

Quartz Crystal Microbalance (QCM) was used to monitor the mass changes on a quartz crystal surface containing immobilized lectins that interacted with carbohydrates. The strategy for lectin immobilization was developed on the basis of a multilayer system composed of Au-cystamine-glutaraldehyde-lectin. Each step of the immobilization procedure was confirmed by FTIR analysis. The system was used to study the interactions of Concanavalin A (ConA) with maltose and Jacalin with Fetuin. The real-time binding of different concentrations of carbohydrate to the immobilized lectin was monitored by means of QCM measurements and the data obtained allowed for the construction of Langmuir isotherm curves. The association constants determined for the specific interactions analyzed here were (6.4 +/- 0.2) X 10(4) M-1 for Jacalin-Fetuin and (4.5 +/- 0.1) x 10(2) M-1 for ConA-maltose. These results indicate that the QCM constitutes a suitable method for the analysis of lectin-carbohydrate interactions, even when assaying low molecular mass ligands such as disaccharides. Published by Elsevier B.V.

Electrochemical and spectroscopic characterization of screen-printed gold-based electrodes modified with self-assembled monolayers and Tc85 protein

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

Photodegradation of sulfamethoxazole in various aqueous media: Persistence, toxicity and photoproducts assessment

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Double-Tagging Polymerase Chain Reaction with a Thiolated Primer and Electrochemical Genosensing based on Gold Nanocomposite Sensor for Food Safety

A novel material for electrochemical biosensing based on rigid conducting gold nanocomposite (nano-AuGEC) is presented. Islands of chemisorbing material (gold nanoparticles) surrounded by nonreactive, rigid, and conducting graphite epoxy composite are thus achieved to avoid the stringent control of surface coverage parameters required during immobilization of thiolated oligos in continuous gold surfaces. The spatial resolution of the immobilized thiolated DNA was easily controlled by merely varying the percentage of gold nanoparticles in the composition of the composite. As low as 9 fmol (60 pM) of synthetic DNA were detected in hybridization experiments when using a thiolated probe. Moreover, for the first time a double tagging PCR strategy was performed with a thiolated primer for the detection of Salmonella sp., one of the most important foodborne pathogens affecting food safety. Ibis assay was performed by double-labeling the amplicon during the PCR with a -DIG and -SH set of labeled primers. The thiolated end allows the immobilization of the amplicon on the nano-AuGEC electrode, while digoxigenin allows the electrochemical detection with the antiDIG-HRP reporter in the femtomole range. Rigid conducting gold nanocomposite represents a good material for the improved and oriented immobilization of biomolecules with excellent transducing properties for the construction of a wide range of electrochemical biosensors such as immunosensors, genosensors, and enzymosensors.

A label-free immunosensor based on recordable compact disk chip for early diagnostic of the dengue virus infection

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Glucose metabolism in rats submitted to skeletal muscle denervation

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

Metabolismo glicídico em ratos submetidos a desnervação do músculo esquelético e ao exercício de natação

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

Utilização do nitrogênio da palha de milho e de adubos verdes pela cultura do milho

A qualidade dos resíduos vegetais de culturas comerciais e adubos verdes influencia a taxa de mineralização/imobilização de N e o respectivo aproveitamento desse nutriente pelas subseqüentes culturas. Com o objetivo de avaliar a utilização do N mineralizado da parte aérea e do sistema radicular da crotalária (Crotalaria juncea) e milheto (Pennisetum americanum) e da palha de milho, marcados com 15N, realizou-se um estudo, em casa de vegetação, no Centro de Energia Nuclear na Agricultura - CENA/USP, Piracicaba (SP), em vasos com 5 kg de solo de um Latossolo Vermelho distroférrico. Foram utilizados seis tratamentos e quatro repetições, dispostos num delineamento inteiramente casualizado, compreendendo: T1 = palha de milho-15N (parte aérea, exceto os grãos) (80 mg kg-1 de N no solo); T2 = raiz de milheto-15N (30 mg kg-1 de N no solo); T3 = parte aérea de milheto-15N (80 mg kg-1 de N no solo); T4 = raiz de crotalária-15N (30 mg kg-1 de N no solo); T5 = parte aérea de crotalária-15N (80 mg kg-1 de N no solo) e T6 = tratamento sem adição de fonte orgânica de N. Para os tratamentos que receberam raiz marcada com 15N, adicionou-se parte aérea sem marcação isotópica na mesma quantidade que naqueles que receberam parte aérea marcada, e vice-versa. As raízes foram incorporadas ao solo e a parte aérea adicionada sobre a superfície. Para avaliar absorção de N da palha de milho-15N (7,35 Mg ha-1, equivalente a 56 kg ha-1 de N) pela cultura do milho, realizou-se também um experimento de campo, na mesma área em que foi coletado solo para o experimento de casa de vegetação. A quantidade de N no milho proveniente da crotalária (111,80 mg vaso-1 N ) foi superior à do milheto (30,98 mg vaso-1 N ), que foi superior à da palha de milho (11,80 mg vaso-1N ). A crotalária proporcionou maior absorção de N e produtividade de matéria seca ao milho. O aproveitamento pelo milho do N da parte aérea da crotalária foi superior ao do N do sistema radicular, mas não houve diferença para o N do milheto. A absorção do N dos restos culturais de milho pela cultura do milho, no campo, foi de 4,1 % da quantidade inicial.

Dinâmica do nitrogênio no solo em razão da calagem e adubação nitrogenada, com palha na superfície

Em sistemas com semeadura direta, a calagem tem sido aplicada superficialmente, causando excesso de calcário nos primeiros centímetros do perfil do solo, onde também é aplicado o N em cobertura e a densidade de comprimento radicular é alta. O objetivo deste trabalho foi estudar a dinâmica do N e do pH do solo em razão da calagem em superfície e aplicação de N em cobertura, na presença de palha. Plantas de algodão (Gossypium hirsutum) foram cultivadas, por 60 dias, em vasos que receberam calagem superficial ou incorporada e aplicação de doses equivalentes a 0, 50, 100 e 150 kg ha-1 de N em cobertura, na forma de sulfato de amônio; os vasos apresentavam, na superfície, o equivalente a 4 t ha-1 de palha de milheto. A calagem aumentou a mineralização e a nitrificação do N no solo, independentemente do modo de aplicação de calcário, aumentando a sua disponibilidade à planta e também a possibilidade de lixiviação. A absorção de N pela planta, além de sua imobilização pela massa microbiana do solo, neutralizou o efeito do adubo nitrogenado amoniacal no pH do solo. O algodoeiro respondeu melhor à calagem incorporada do que à superficial, mas esta resposta não foi causada por diferenças na disponibilidade de nitrogênio.