Produção de anticorpos monoclonais murinos dirigidos contra células-tronco mesenquimais de coelho

Mesenchymal stem cells (MSCs) are a heterogeneous population of cells that proliferate in vitro as plastic-adherent cells, have fibroblast-like morphology and can differentiate into bone, cartilage and fat cells. Therapeutic potential of MSCs have been studied in experimental models, such as rabbit, in Laboratory of Cell Engineering of Botucatu. However, no specific markers have been reported for expanded rabbit MSCs, which hampers the isolation of pure MSC populations by immunophenotypic characterization. Thus, the objective of this study was to produce monoclonal antibodies (mAbs) to rabbit MSCs. MSCs derived from rabbit bone marrow (BM) were isolated, cultured, expanded ex vivo, and immunized into three BALB/c mices, and spleen cells subsequently harvested were used to generate hibridoma cell lines secreting antibodies against MSCs. Hybridoma cells were screened by flow cytometry and antibody-producing cells were subjected to subsequent rounds of retests. MSC1-160 obtained the best positivity for IgG expression and was cloned by limiting dilutions and micromanipulation. Ascitic fluid from ten best clones was purified by affinity chromatography in Protein A-sepharose CL-4B column and purification control was performed by electrophoresis in agarose gels. The purified IgG were tested against rabbit MSCs, obtaining high positivity by flow Cytometry. In conclusion, we developed 10 mAbs, MSC1-160 A20, A30, A41, A47, A55, A60, A63, A69, A81, and A82, that recognize rabbit MSC cell surface antigens showing potential for immunophenotypic characterization of rabbit MSC cell lines

Expressão de SmNPP5a de Schistosoma mansoni em Pichia pastoris

A esquistossomose é causada pelo Schistosoma mansoni e é considerada uma importante doença parasitária que afeta mais de 200 milhões de pessoas em países em desenvolvimento. Estima-se ainda que aproximadamente 600 milhões de pessoas vivam em áreas de risco e que o número de mortes por ano devido a esta parasitose chegue a 200.000. O tratamento existente hoje é eficiente, mas não previne contra re-infecção e contra as formas jovens dos parasitas. O desenvolvimento de uma vacina utilizando antígenos do parasita produzidos de modo recombinantes é o anseio para o controle da esquistossomose. A fosfodiesterase-5 (SmNPP-5a) é uma enzima presente na interface parasita-hospedeiro e acessível ao sistema imune, sendo portanto considerada um potencial candidato vacinal, sua avaliação se faz necessária. A obtenção de uma SmNPP-5a recombinante enzimaticamente ativa é um passo fundamental no processo de avaliação da sua capacidade protetora e determinação da sua função fisiológica para o parasita. Utilizando o sistema de expressão baseados em leveduras metilotróficas, Pichia pastoris, a proteína recombinante pode ser expressa com as modificações pós traducionais necessárias para ter sua estrutura original mantida, característica fundamental para se obter uma proteína enzimáticamente ativa. Para clonagem e expressão da proteína recombinante utilizamos o vetor pPICZαA, pois este possui um sistema de expressão baseado no promotor álcool oxidase (AOX1 e AOX2). Ele possui ainda, a capacidade de adicionar um peptídeo sinal e uma cauda de histidina, com objetivo de secretar a proteína expressa e facilitar sua purificação, respectivamente. A triagem e confirmação dos clones positivos foram feitas por PCR. A análise das amostras de sobrenadantes coletadas com 24, 48 e 72h de estímulo com metanol foi feita por SDS-PAGE e Western blot para verificação da expressão... (Resumo completo, clicar acesso eletrônico abaixo)

Precificação do biogás produzido por autoprodutores, considerando seu transporte por meio de gasodutos existentes

The expansion of the energy matrix is a matter of great importance since the recent problems facing the country in this sector, such as rationing mobilized in 2001. Moreover, there is also concern with global sustainable development. Brazil is producing about 64 million cubic meters of biogas per day, but only a small percentage of this total is harnessed as an energy resource. Compounders are considered potential, for example, landfill energy (LE) and sewage treatment plants (STP), which can be adapted to produce biogas and market them. The work focuses its studies on biogas produced by these small and medium compounders. Are proposed and studied the following alternatives for the utilization of biogas energy: generation and sale of electricity through the installation of a small power station within the landfill, and purification and marketing of gas, whose price will be a consideration on the price of natural gas, taking into account the percentage of methane present in the biogas. Trade ensures that gas plus a destination ecologically appropriate to it, taking advantage of its energy

Uso de sistemas construídos de áreas alagadas (Constructed Wetland System - CWS) tipo ascendente/descendente, para tratamento de efluentes avícolas

Human development and population growth during the twentieth century increased the water demand, tripling its consumption between 1950 and 1990. As the water streams were polluted; and as water is the source of minerals and also regulates vital functions, it becomes the vehicle of transmission and consequently spreads many diseases. Probably, the industries are the major responsible for this pollution when they dump untreated effluents to water streams, saturating the already insufficient net of sanitation facilities polluting water and soil. An effective treatment has been established with low cost in Europe and the United States, through constructed systems on wetlands Constructed Wetland Systems - CWSs, gradually used in other countries in the last three decades. Lately, we observe a continuous growth in Brazilian poultry business, and poultry industry showed greatest dynamism in the country, following the global market. Pondering this information and the efficiency of such treatment, this work aimed to study prototypes, in a laboratory scale, simulating ascending and descending types of CWSs, vegetated with aquatic macrophytes Eichhornia crassipes and the uses of aggregates and soil, to treat industrial wastewater from slaughterhouses and aviary. We conducted the initial characterization of the effluent to have an idea of its constituents and to scale the system and the continuous flow. Furthermore, we characterized the soil to be used in this system. The collects are periodically made in the refrigeration industry FRICOCK FRIGORIFICAÇÃO AVICULTURA INDÚSTRIA E COMÉRCIO LTDA. for local treatment simulation. The effluent that was treated with 12 prototypes of CWSs are analyzed with some frequency. The results of these reviews were compared to the effluent coming from the industry... (Complete abstract click electronic access below)

Utilização de biossurfactante e surfactante químico para dessorção de óleo presente em vermiculita hidrofobizada: estudo comparativo

A number of methods and products have been developed in order to eliminate or reduce the negative effects that hydrocarbons cause to the environment, including hydrophobic expanded vermiculite, used in oil residue filtering systems at gas stations. However, upon adsorbing organic compounds, the vermiculite is no longer used and is sent to landfills. The aim of the present study was to wash granular and powdered vermiculite containing oil lubricant in its pores with distilled water and solutions of 0.1% SDS surfactant and rhamnolipids, with the aim of removing the lubricant and the possibility of reusing the mineral. The greatest amount of lubricant removal was obtained through washing with 0.1% SDS and both granulometric forms. This may be associated to the industrial purification received by the surfactant. However, the biosurfactant is ecologically more viable due to its low toxicity and ease of degradability. In the readsorption tests, greatest adsorption was obtained with the granular vermiculite washed in SDS solution. In order to enable the reuse of the mineral, further tests are needed to enhance desorption/adsorption efficiency.

Biorremoção do corante acid blue 29, usando serragem (Pinus sp.) como biossorvente

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Análise eletroforética de proteínas recombinantes obtidas de Pichia pastoris

A glicerol quinase é uma proteína tetramérica que cataliza a fosforilação do glicerol a glicerol-3-fosfato, composta por quatro subunidades - sua análise eletroforética fornecerá apenas uma banda se for constatada a sua pureza num gel não desnaturante - e sua reação com o glicerol é dependente de magnésio e de ATP. A eletroforese de proteínas utiliza como suporte um gel de poliacrilamida, que é formado pela polimerização de monômeros de acrilamida ao longo da sua cadeia e ligações cruzadas de cadeias pela inclusão de um co-monômero bifuncional apropriado, usualmente a N,N’ – metileno-bis-acrilamida ou apenas Bis . A eletroforese de proteínas mais comum é a que utiliza SDS para um gel de poliacrilamida desnaturante. O SDS é um sal chamado Dodecil Sulfato de Sódio que tem por característica se ligar a cadeia proteína nos resíduos de aminoácidos apolares, deixando com a carga negativa, sendo assim, toda proteína fica com carga total negativa, migrando para o anodo na eletroforese. As técnicas de purificação utilizadas foram ultrafiltração e precipitação com ácido tricloro acético, etanol gelado e sulfato de amônio. A dupla precipitação com etanol resultou na recuperação de maior quantidade de proteínas. A coloração do gel com prata foi mais sensível do que Comassie blue. O gel de eletroforese mostrou quatro bandas, correspondentes às quatro subunidades da glicerol quinase quando revelados com prata em gel de SDS-PAGE.

Remoção de matéria orgânica e nitrogênio de efluentes provenientes de abatedouros de bovinos por meio de reator aeróbio operando em bateladas sequenciais com adição de coagulante químico

The volume of liquid effluent generated in cattle slaughterhouses is quite high and cannot be released untreated in water bodies due to its high pollution load of predominantly organic origin. To minimize the environmental impacts of its industrial wastewater and meet the local environmental legislation, abattoirs shall make the treatment of these effluents. The present work aims to develop the study of a reactor by sequential batch pilot scale, in order to optimize their performance in treating wastewater from a cattle slaughterhouse. The treatment system used was developed and installed in the Laboratory of Wastewater Treatment, in Faculty of Science and Technology UNESP, Presidente Prudente campus. The procedure used followed the operation of sequential batch reactors, in which all processes and treatment operations occurring sequentially in a single unit, by establishing specific operating cycles, which comprise the following separated phases: aerobic reaction, anoxic reaction, sedimentation and emptying. Aiming to improve the quality of treatment was planned the addition of coagulant Poly Aluminum Chloride (PAC) in the reactor, by determining their optimal dosage by Jar-test trials. Were prepared four steps with specific operating cycles: step one or acclimatization (10 hour of aeration, one hour and 30 minutes of sedimentation and 30 minutes for exchanging the effluent); step 2 (6 hours of aeration or aerobic phase, 4 hours and 45 minutes of stirring or anoxic phase and 1 hour and 15 minutes for sedimentation and exchange effluent); step 3 (2 hours and 30 minutes of aeration, 8 hours and 15 minutes of stirring and 1 hour and 15 minutes for sedimentation and exchange) and step 4 (2 hours of aeration, 8 hours and 45 minutes of stirring and 1 hour and 15 minutes for sedimentation and exchange)... (Complete abstract click electronic access below)

Desenvolvimento de kit para quantificação enzimática de etanol

A dosagem do etanol durante a produção de medicamentos, bebidas e combustíveis tem se mostrado bastante importante para o aumento de rendimento dos processos e qualidade dos produtos. Embora existam vários métodos para determinar a concentração do etanol, os enzimáticos são os mais interessantes por serem rápidos, precisos, de fácil manuseio e baixo custo. O objetivo geral desse trabalho foi a elaboração de um Kit de quantificação enzimática de etanol, utilizando a enzima álcool desidrogenase I recombinante de S. cerevisiae (Adh1). Para isso, o gene codificador da enzima álcool desidrogenase I (ADH1) de levedura foi clonado no vetor pET28a para expressão em E. coli (BL21). Em seguida, a produção e solubilidade da enzima álcool desidrogenase I foi determinada, sendo definida uma condição ideal de produção e purificação da proteína recombinante por cromatografia de afinidade. A atividade enzimática da álcool desidrogenase I purificada foi determinada através de ensaios já padronizados na literatura (UV), bem como sua estabilidade em diversas condições de tempo e temperatura. Os resultados obtidos aqui demonstraram que a enzima obtida apresentou boa atividade biológica in vitro, com sensibilidade na detecção de no mínimo 2,3 x 10–4 g/L de etanol em 2 minutos. Além disso, a proteína purificada teve sua atividade preservada quando estocada a - 20°C durante 120 dias, características que possibilitarão que a enzima álcool desidrogenase I recombinante, purificada pela metodologia apresentada nesse trabalho, seja utilizada em um kit de quantificação enzimática de etanol rápido, preciso e de baixo custo

Estudo da tratabilidade de água pluvial utilizando coagulante natural a base de tanino visando fins não potáveis

This paper aimed to study the treatability of rainwater collected in Rio Claro, SP, using natural coagulant tannin based. The tannin was compared with other too primary coagulants, namely ferric chloride and corn starch. Assays were performed in laboratory scale using equipment known as Jarteste and aimed to study the conditions most suitable dosage and coagulation pH. First tests were conducted to compare the coagulating and determination of the conditions to be used in the following tests, taking as a criterion the greatest efficiency for water treatment. Also performed were performed tests to construct the tannin coagulation diagram, using the optimal concentration found in the previous phase, by varying the dosage of coagulant and the pH of clotting in order to determine the isoefficiency regions for apparent color and turbidity. Finally, tests were performed cyclically filtration with filter paper. Preliminary tests indicated that the vegetable tannin concentration 3% (m/m) from the commercial solution was presented the best set of results (84.3% removal of apparent color and 82.51% removal of turbidity) compared to ferric chloride (66.25% removal efficiency and apparent color of turbidity 67.82%) and starch (73.68% removal efficiency apparent color and turbidity to 67.19%). The coagulation diagrams indicate that the best region to work with vegetable tannin 3% is where the coagulant dosage ranges from 15 mg/L to about 37.5 mg/L and coagulation pH ranging from about 6.5 to 7.5. The cyclic filtration showed to be not efficient for the best dosage of coagulant, due to the process of clogging of the filter paper, but proved to be efficient for other dosages. Thus, for the waters studied, the results indicate that the natural coagulant based tannin is an alternative to conventional coagulants, possessing benefits of technical and environmental

Resolução de problema de valor no contorno associado ao projeto de reatores de fluxo disperso empregados no tratamento de efluentes

The objective of this work was to develop a numerical method to solve boundary value problems concerning to the use of dispersion model for describing the hydraulic behavior of chemical or biological reactors employed in the wastewater treatment. The numerical method was implemented in FORTRAN language generating a computational program which was applied to solve cases involving reaction kinetics of both integer and fractional orders. The developed method was able to solve the proposed problems evidencing to be a useful tool that provides more accurate design of wastewater treatment reactors

Estudo sobre a condição de coleta e tratamento de esgoto sanitário do município de Ubatuba-SP

The actual work broaches a study about the collection and treatment of sanitary sewage from the municipality of Ubatuba in its operational aspects and the tariff policy from the service. Themes related to the subject like the history of collection and treatment of sewage, its development in Brazil and its importance for public health and environmental preservation, as well as historical and geographical aspects of Ubatuba/SP, are present in the literature review. The methodology consists in guided visit in the colletion and treatment of sewage facilities and interview with technical managers. With the technical data obtained, it is made a discussion of the results and possible recommendations

Otimização de um método cromatográfico de análise de pesticidas organoclorados em amostras de peixes

Considering that persistent organic pollutants (POPs) including synthetic substances belonging to differents organic functions, including organochlorine pesticides (OCPs), which are the most persistent and bioaccumulative, with high toxicity to humans and animals. Accordingly environmental and biological monitoring is necessary, in order to have greater control regarding the irresponsible use of these products. Though there are several analytical methodology reported in the literature to make determinations of this pesticide, they present some difficulties, requiring several steps to make the clean up of the sample. The proposed project aims to optimize a new analytical method that allows to perform the extraction of organochlorine pesticides in fish tissues, employing acetone as solvent assisted by ultrasound bath, making the method more quickly and not requiringfurther steps to purify the sample. Were analyzed the recoveries of pesticides in study in samples of tilapia average values: Heptachlor=84,7±9,8%; Aldrin=87,3±3,5%; Endosulfan=101,4±18,6%; DDE= 90,5± 3,3%, Endrin=102,2±13,8%., DDD=92,1±19,9%; DDT=90,8±6,8%. The real samples showed values granted for our legislation and low values of error

Desidratação de lodo proveniente de ETA através de tubos geotexteis

Currently exists a growing concern for the preservation of the environment Around the world, the environmental awareness in Brazil has strengthened during the past two decades. This concern in Brazil arises from the creation of mechanisms of supervision and punishment on the part of the environmental agencies. In order to meet the pertinent legislation many public and private companies have performed dehydration of waste generated in the process of treatment of industrial effluents, waste water and water treatment in order to reduce transport and disposal costs. The use of geotextile tubes has proven technically and economically feasible to be applied in various situations from water treatment to mining tailings. This work presents the solution adopted for an environmental liability in water treatment plant through the use of geotextile tubes to reduce the water content of the disposal. We evaluated the size distribution curves, Atterberg limits, and chemical composition of the residue. Found high concentrations of aluminum in the waste which would characterize environmental pollution if disposed off directly in nature with no treatment

Produção de polpa solúvel a partir de Eucalyptus urograndis

The wood of the gender Eucalyptus occupies a prominence place in relation to other cellulose sources, due to its chemical composition, its low cost, abundance and availability. The dissolving pulp obtaining occurs basically starting from three stages: prehydrolysis of the wood following by cooking and finally by the bleaching process. In the dissolving pulp production is necessary a pretreatment stage for the hemicelluloses removal. Among the chemical cooking processes, the Kraft process is the most used for cellulosic pulps production starting from eucalyptus, in the additive presence or not. The anthraquinone uses in the pulping process has as main purposes increasing the delignification rate as well as protecting the pulp regarding the degradation of the polysaccharide chains. The ECF bleaching process consists of the purification of the pulp through bleaching agents, being excepted the elementary chlorine. The dissolving pulp is a pulp of high a-cellulose content and purity destined to the cellulose derivatives production as carboxymethyl cellulose, nitrocellulose and microcrystalline cellulose. The production of a certain product, as well as its final application, is determined by the chemical properties of the pulp. The present work had as purpose producing cellulosic pulp with kappa number around 9 destined to the production of dissolving pulp in way to evaluate the use of the anthraquinone (AQ) in the Kraft process according to the chemical properties of the obtained pulps. It has been still intended developing an effective technology of bleaching for the high purity and quality dissolving pulp production. The prehydrolysis was accomplished in laboratory rotational autoclave for 30 minutes at 170ºC in the water:wood rate 3,5:1 (L:kg). The Kraft/AQ pulping were accomplished in laboratory rotational autoclave being initially modified the active alkali load (10, 13, 16 and 19%) at sulfidity of 25% ...