CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / Immobilization of enzymes provides advantages such as increased in: stability, level of process control, yield and purity of the final product, in addition to allowing the use of different reactor configurations. In this study, two types of support, an organic polymer (polimetilmetacrilato- PMMA) and other inorganic (silica) were used for the immobilization of lipase Candida antarctica type B (CALB). Different alcohols (ethanol, iso-propanol, n-butanol) were evaluated as agents for optimization of the immobilization process, the concentrations of 5, 10 and 15% (volume of alcohol à 100 / solution volume immobilization). Immobilization of PMMA was conducted using two different protocols here denominated first and second protocol generation. The best results of hydrolytic activity and esterification to the derivative of the first generation was 3,91  0,09 UpNPB/g. of catalyst e 409,5  19,7 Uoleic acid/g. of catalyst, respectively. When using the protocol of the second generation, the best results were obtained when aa immobilization was carried out in the presence of 5% Triton X-100 2 mM, rising rate of the esterification reaction between the derivatives studied was highly significant with a higher ratio of 3,81 times to the derivative prepared with Triton X-100. The CALB was also immobilized for the macroporous silica support (IB S60S - Chiralvision), the activate agent used was Triethoxy(octyl)silane (C8-TEOS). The results observed for immobilization in the presence of 5% n-butanol were the best for both hydrolytic activity, being four times higher than the activity for the immobilized derivative without alcohol, as for the esterification activity was 985,1  5,6 Uoleic acid/g. of catalyst . All observed derivatives supported on silica and PMMA reached a level conversion for equilibrium esterification reaction for a reaction time of 24 h the best results varying between 80 to 90%. / A imobilizaÃÃo das enzimas traz vantagens como: aumento da estabilidade, do nÃvel de controle do processo, do rendimento e da pureza do produto final, alÃm de possibilitar o uso de reatores de diferentes configuraÃÃes. Neste trabalho, dois tipos de suporte, um polÃmero orgÃnico (polimetilmetacrilato- PMMA) e outro inorgÃnico (sÃlica), foram utilizados para a imobilizaÃÃo da lipase do tipo B de Candida antarctica (CALB). Diferentes Ãlcoois (etanol, iso-propanol, n-butanol) foram avaliados como agentes de otimizaÃÃo do processo de imobilizaÃÃo, nas concentraÃÃes de 5, 10 e 15 % (volume de Ãlcool Ã100 / volume da soluÃÃo de imobilizaÃÃo). A imobilizaÃÃo em PMMA foi conduzida atravÃs de dois protocolos diferentes, aqui denominados de protocolo de primeira e segunda geraÃÃo. Os melhores resultados de atividade hidrolÃtica e de esterificaÃÃo para o derivado de primeira geraÃÃo foi de 3,91  0,09 UpNPB/g. de catalisador e 409,5  19,7 UÃc. olÃico/g. de catalisador, respectivamente. Quando se utilizou o protocolo de segunda geraÃÃo, melhores resultados foram obtidos quando a a imobilizaÃÃo foi conduzida na presenÃa de n-butanol 5 % e Triton X-100 2mM; o aumento da velocidade da reaÃÃo de esterificaÃÃo entre os derivados estudados foi bastante significativo, sendo 3,8 vezes maior para o derivado preparado com Triton X-100. A CALB foi imobilizada tambÃm para o suporte sÃlica macroporosa (IB S60S - Chiralvision), o agente ativante utilizado foi o octiltrietoxisilano (C8-TEOS). Os resultados observados para imobilizaÃÃo na presenÃa de n-butanol 5 % foram os melhores tanto para a atividade hidrolÃtica, sendo quatro vezes maior que a atividade para o derivado imobilizado sem Ãlcool, quanto para a atividade de esterificaÃÃo que foi de 985,1  5,6 UÃc. olÃico/g. de catalisador. Todos os derivados observados suportados em PMMA e sÃlica atingiram um patamar de conversÃo do equilÃbrio para reaÃÃo de esterificaÃÃo por um perÃodo de reaÃÃo de 24 h com os melhores resultados variando entre 80 a 90%.

Enzyme-catalyzed biofuel cells would be a great alternative to current battery technology, as they are clean, safe, and capable of using diverse and abundant renewable biomass with high energy densities, at mild reaction conditions. However, currently, three largest technical challenges for emerging enzymatic fuel cell technologies are incomplete oxidation of most fuels, limited power output, and short lifetime of the cell. Synthetic pathway biotransformation is a technology of assembling a number of enzymes coenzymes for producing low-value biocommodities. In this work, it was applied to generate bioelectricity for the first time. Non-natural enzymatic pathways were developed to utilize maltodextrin and glucose in enzymatic fuel cells. Three immobilization approaches were compared for preparing enzyme electrodes. Thermostable enzymes from thermophiles were cloned and expressed for improving the lifetime and stability of the cell. To further increase the power output, non-immobilized enzyme system was demonstrated to have higher power densities compared to those using immobilized enzyme system, due to better mass transfer and retained native enzyme activities. With the progress on pathway development and power density/stability improvement in enzymatic fuel cells, a high energy density sugar-powered enzymatic fuel cell was demonstrated. The enzymatic pathway consisting of 13 thermostable enzymes enabled the complete oxidation of glucose units in maltodextrin to generate 24 electrons, suggesting a high energy density of such enzymatic fuel cell (300 Wh/kg), which was several folds higher than that of a lithium-ion battery. Maximum power density was 0.74 mW/cm2 at 50 deg C and 20 mM fuel concentration, which was sufficient to power a digital clock or a LED light. These results suggest that enzymatic fuel cells via synthetic pathway biotransformation could achieve high energy density, high power density and increased lifetime. Future efforts should be focused on further increasing power density and enzyme stability in order to make enzymatic fuel cells commercially applicable. / Ph. D.

Call number: LD2668 .T4 1984 S87 / Master of Science

The suitability of water hyacinth (Eichornia crassipes) as a viable feedstock for renewable energy production was investigated in this project. Water hyacinth used in this study was harvested from the Vaal River near Parys in the northwest region of the Free State province, South Africa (26°54′S 27°27′E). The wet plants were processed in the laboratory at the North-West University by separating the roots from the leaves and the stems, thus obtaining two separate water hyacinth feedstock. Characterisation of the feedstock showed that the stems and leaves are more suitable for bio-energy production than roots, due to the higher cellulose and hemicellulose content and very low lignin content of the stems and leaves. Water hyacinth was evaluated as feedstock for the production of bio-ethanol gel, bio-ethanol, bio-oil and bio-char. The recovery of water from the wet plants for use in bio-refining or for use as drip-irrigation in agriculture was also investigated. Cellulose was extracted from water hyacinth feedstock to be used as a gelling agent for the production of ethanol-gel fuel. A yield of 200 g cellulose/kg dry feedstock was obtained. The extracted cellulose was used to produce ethanol-gel with varying water content. The gel with properties closest to the SANS 448 standard contained 90 vol% ethanol and 10 vol% water, with 38 wt% cellulose. This gel was found to ignite readily and burn steadily, without flaring, sudden deflagrations, sparking, splitting, popping, dripping or exploding from ignition until it had burned to extinction, as required by SANS 448. The only specifications that could not be met were the viscosity (23,548 cP) and the high waste residue (32 wt%) left after burning. The other major concern is the extremely high costs involved with the manufacturing of ethanol-gel from water hyacinth cellulose. It can be concluded that ethanol-gel cannot be economically produced using water hyacinth as feedstock. Chemical and enzymatic extraction of water from the feedstock, which is stems and leaves or roots, showed that the highest yield of water was obtained using a combination of Celluclast 1.5 L, Pectinex Ultra SP-L and additional de-ionised water. A yield of 0.89 ± 0.01 gwater/gwater in biomass was realised. This is, however, only 0.86 wt% higher than the highest yield obtained (0.87 ± 0.01 gwater/gwater in biomass) using only Pectinex Ultra SP-L and de-ionised water. It is recommended to use only Pectinex Ultra SP-L and de-ionised water at a pH of 3.5 and a temperature of 40°C. Using one enzyme instead of two reduces operating costs and simplifies the chemical extraction process. The extracted water, both filtered and unfiltered, was not found to be suitable for domestic use without further purification to reduce the total dissolved solids (TDS), potassium and manganese levels. Both the unfiltered and filtered water were, however, found to be suitable for industrial and agricultural purposes, except for the high TDS levels. If the TDS and suspended particle level can be reduced, the extracted water would be suitable for domestic, industrial and agricultural use. The potential fermentation of the sugars derived from the water hyacinth, using ultrasonic pretreatment, was investigated. Indirect ultrasonic treatment (ultrasonic bath) proved to be a better pretreatment method than direct sonication (ultrasonic probe). The optimum sugar yield for the ultrasonic bath pretreatment with 5% NaOH was found to be 0.15 g sugar/g biomass (0.47 g sugar/g available sugar) using an indirect sonication energy input of 27 kJ/g biomass. The optimum sugar yield is lower than those reported in other studies using different pretreatment methods. Theoretically a maximum of 0.24 g ethanol can be obtained per g available sugar. This relates to an ethanol yield of 0.08 g ethanol/kg wet biomass. The low yield implies that ethanol production from water hyacinth is not economically feasible. The production of bio-oil and bio-char from water hyacinth through thermochemical liquefaction of wet hyacinth feedstock was investigated. An optimum bio-char yield of 0.55 g bio-char/g biomass was achieved using an inert atmosphere (nitrogen) at 260°C and the stems and leaves as feedstock. With the roots as feedstock a slightly lower optimum yield of 0.45 g bio-char/g biomass was found using a non-reducing atmosphere (carbon monoxide) at 280°C. The bio-oil yield was too low to accurately quantify. As water is required during thermochemical liquefaction, it was found unnecessary to dry the biomass to the same extent as was the case with the pretreatment and fermentation of the water hyacinth, making this a more feasible route for biofuel production. Bio-char produced through liquefaction of roots as the feedstock and leaves and stems as the other feedstock had a higher heating value (HHV) of 10.89 ± 0.45 MJ/kg and 23.31 ± 0.45 MJ/kg respectively. Liquefaction of water hyacinth biomass increased the HHV of the feedstock to a value comparable to that of low grade coal. This implies a possible use of water hyacinth for co-gasification. The most effective route for bio-energy production in the case of water hyacinth was found to be thermochemical liquefaction (12.8 MJ/kg wet biomass). Due to the high production costs involved, it is recommended to only use water hyacinth as a feedstock for biofuel production if no alternative feedstock are available. / MIng (Chemical Engineering), North-West University, Potchefstroom Campus, 2014

The suitability of water hyacinth (Eichornia crassipes) as a viable feedstock for renewable energy production was investigated in this project. Water hyacinth used in this study was harvested from the Vaal River near Parys in the northwest region of the Free State province, South Africa (26°54′S 27°27′E). The wet plants were processed in the laboratory at the North-West University by separating the roots from the leaves and the stems, thus obtaining two separate water hyacinth feedstock. Characterisation of the feedstock showed that the stems and leaves are more suitable for bio-energy production than roots, due to the higher cellulose and hemicellulose content and very low lignin content of the stems and leaves. Water hyacinth was evaluated as feedstock for the production of bio-ethanol gel, bio-ethanol, bio-oil and bio-char. The recovery of water from the wet plants for use in bio-refining or for use as drip-irrigation in agriculture was also investigated. Cellulose was extracted from water hyacinth feedstock to be used as a gelling agent for the production of ethanol-gel fuel. A yield of 200 g cellulose/kg dry feedstock was obtained. The extracted cellulose was used to produce ethanol-gel with varying water content. The gel with properties closest to the SANS 448 standard contained 90 vol% ethanol and 10 vol% water, with 38 wt% cellulose. This gel was found to ignite readily and burn steadily, without flaring, sudden deflagrations, sparking, splitting, popping, dripping or exploding from ignition until it had burned to extinction, as required by SANS 448. The only specifications that could not be met were the viscosity (23,548 cP) and the high waste residue (32 wt%) left after burning. The other major concern is the extremely high costs involved with the manufacturing of ethanol-gel from water hyacinth cellulose. It can be concluded that ethanol-gel cannot be economically produced using water hyacinth as feedstock. Chemical and enzymatic extraction of water from the feedstock, which is stems and leaves or roots, showed that the highest yield of water was obtained using a combination of Celluclast 1.5 L, Pectinex Ultra SP-L and additional de-ionised water. A yield of 0.89 ± 0.01 gwater/gwater in biomass was realised. This is, however, only 0.86 wt% higher than the highest yield obtained (0.87 ± 0.01 gwater/gwater in biomass) using only Pectinex Ultra SP-L and de-ionised water. It is recommended to use only Pectinex Ultra SP-L and de-ionised water at a pH of 3.5 and a temperature of 40°C. Using one enzyme instead of two reduces operating costs and simplifies the chemical extraction process. The extracted water, both filtered and unfiltered, was not found to be suitable for domestic use without further purification to reduce the total dissolved solids (TDS), potassium and manganese levels. Both the unfiltered and filtered water were, however, found to be suitable for industrial and agricultural purposes, except for the high TDS levels. If the TDS and suspended particle level can be reduced, the extracted water would be suitable for domestic, industrial and agricultural use. The potential fermentation of the sugars derived from the water hyacinth, using ultrasonic pretreatment, was investigated. Indirect ultrasonic treatment (ultrasonic bath) proved to be a better pretreatment method than direct sonication (ultrasonic probe). The optimum sugar yield for the ultrasonic bath pretreatment with 5% NaOH was found to be 0.15 g sugar/g biomass (0.47 g sugar/g available sugar) using an indirect sonication energy input of 27 kJ/g biomass. The optimum sugar yield is lower than those reported in other studies using different pretreatment methods. Theoretically a maximum of 0.24 g ethanol can be obtained per g available sugar. This relates to an ethanol yield of 0.08 g ethanol/kg wet biomass. The low yield implies that ethanol production from water hyacinth is not economically feasible. The production of bio-oil and bio-char from water hyacinth through thermochemical liquefaction of wet hyacinth feedstock was investigated. An optimum bio-char yield of 0.55 g bio-char/g biomass was achieved using an inert atmosphere (nitrogen) at 260°C and the stems and leaves as feedstock. With the roots as feedstock a slightly lower optimum yield of 0.45 g bio-char/g biomass was found using a non-reducing atmosphere (carbon monoxide) at 280°C. The bio-oil yield was too low to accurately quantify. As water is required during thermochemical liquefaction, it was found unnecessary to dry the biomass to the same extent as was the case with the pretreatment and fermentation of the water hyacinth, making this a more feasible route for biofuel production. Bio-char produced through liquefaction of roots as the feedstock and leaves and stems as the other feedstock had a higher heating value (HHV) of 10.89 ± 0.45 MJ/kg and 23.31 ± 0.45 MJ/kg respectively. Liquefaction of water hyacinth biomass increased the HHV of the feedstock to a value comparable to that of low grade coal. This implies a possible use of water hyacinth for co-gasification. The most effective route for bio-energy production in the case of water hyacinth was found to be thermochemical liquefaction (12.8 MJ/kg wet biomass). Due to the high production costs involved, it is recommended to only use water hyacinth as a feedstock for biofuel production if no alternative feedstock are available. / MIng (Chemical Engineering), North-West University, Potchefstroom Campus, 2014

The study herein described involved the detection of pyridine N-oxide reductase activity in cell-free extracts of E. coli 9723, the determination of co-factors necessary for the enzymatic process, a study of the optimum conditions for enzyme catalysis, and a general characterization of the enzyme.

The first project focused on the preparation, characterization, and application of dual emission quantum dot encoded mesoporous silica microparticles. The quantum dots were added in precisely controlled ratios and were stably encapsulated within the pores of the silica. Several experiments were performed to test the superior stability of the quantum dot-silica composites over dye-loaded silica particles. The composite particles exhibited very high fluorescence, were functionalized with antibodies, and were used as signal transducers for the detection of a protein expressed by breast cancer cells. The second project focused in more detail on the detection capabilities of the quantum dot-silica composites. Three different types of quantum dot-silica composites were prepared. Each type was loaded with a separate type of quantum dot with distinct emission wavelengths and was functionalized with separate antibodies for detection of three different breast cancer biomarkers. These three composite sensors were used together for the simultaneous detection of each of the breast cancer markers. The initial strategy utilized the direct detection method in which the antigen is nonspecifically adsorbed to a glass plate. An improved second strategy was more sensitive and used a capture antibody which was covalently bound to a glass plate to immobilize the antigen. The third project focused on the preparation and application of magnetic, fluorescent human serum albumin nanoparticle composites. A fluorescent drug analogue and iron oxide nanoparticles were encapsulated into 100 nm human serum albumin nanoparticles. The advantage of these composite particles is that they could be used as a theranostic tool which could target, detect, and treat diseased tissue in a single application. Release of the drug analogue from the nanocomposites was achieved by addition of proteolytic enzymes that are expressed or overexpressed in cancer cells. The temporal release of the fluorescent drug analogue was measured as a function of enzyme concentration. The amount of drug released was directly proportional to enzyme concentration.

Orientador: Mario de Oliveira Neto / Resumo: A busca por fontes de energias renováveis vêm ganhando cada vez mais importância, pois, com a eminente escassez dos combustíveis fósseis, que são as principais fontes energéticas utilizadas mundialmente e ao mesmo tempo com os problemas ambientais causados por esses combustíveis, o etanol de segunda geração (2G) surge como uma alternativa de energia renovável já que é obtido a partir de biomassa lignocelulósica. Entretanto, as tecnologias para a conversão desta biomassa em açúcares fermentáveis ainda possuem fatores limitantes. Um dos grandes problemas do processo de obtenção do etanol 2G está relacionado ao custo das enzimas celulolíticas. Sendo assim, desenvolver estratégias para a recuperação dessas enzimas é necessário para assegurar a viabilidade econômica do processo de conversão de biomassa. Diversas técnicas estão sendo estudadas e colocadas em prática para promover a recuperação, reciclagem e imobilização enzimática. Entretanto, nenhuma das metodologias utilizadas demonstram, de maneira geral, alta eficiência, custo baixo, rapidez e manutenção da atividade enzimática. A partir da lignina, subproduto da indústria de papel e celulose, pode-se sintetizar o lignocresol. Este, apresenta grandes expectativas de interação enzimática uma vez que apresenta características físico-químicas favoráveis à adsorção. Frente a isto, propusemos a produção deste composto para promover a interação com enzimas para promover recuperação enzimática. A síntese de lignocresol, oriundo da lig... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The search for renewable energy sources is gaining increasing importance because, with the imminent shortage of fossil fuels, which are the main energy sources used worldwide and at the same time with the environmental problems caused by these fuels, second-generation ethanol (2G) arises as a renewable energy alternative since it is obtained from lignocellulosic biomass. However, the technologies for the conversion of this biomass to fermentable sugars still have limiting factors. One of the major problems in the process of obtaining 2G ethanol is related to the cost of cellulolytic enzymes. Thus, developing strategies for the recovery of these enzymes is necessary to ensure the economic viability of the biomass conversion process. Several techniques are being studied and put into practice to promote recovery, recycling, and enzymatic immobilization. However, none of the methodologies used shows, in general, high efficiency, low cost, fastness and maintenance of the enzymatic activity. From lignin, a byproduct of the pulp and paper industry, lignocresol can be synthesized. This one presents high expectations of enzymatic interaction since it presents physicochemical characteristics favorable to the adsorption. In view of this, we proposed the production of this compound to promote interaction with enzymes to promote enzymatic recovery. The lignocresol synthesis from Pinus spp. Kraft lignin was made by adapting to the phase separation system methodology. In this process, more ... (Complete abstract click electronic access below) / Doutor

O microrreator faz parte de conjunto de dispositivos de uma nova e promissora tecnologia, que podem ser chamados de micro fabricados, atuante em campos como a da química, biológica, farmacêutica, engenharia química e biotecnologia. Trata-se de um dispositivo que possibilita reação química, tais como os reatores convencionais, mas com dimensões menores, com canais na escala micrométrica. A tecnologia de miniaturização de dispositivos para reações químicas vem se expandindo promovendo uma importante evolução, com microssistemas que abrange dispositivos mais eficazes, com configuração e geometrias específicas e menor consumo de energia, onde reações com elevadas taxas de transporte podem ser usadas para muitas finalidades diferentes, tais como, reações rápidas, mistura, reações sensíveis à temperatura, temperatura de homogeneização, ou até mesmo precipitação de nano partículas. Devido sua escala ser extremamente reduzida em relação à escala macro, oferecem um sistema que permite uma investigação do processo em um curto espaço de tempo, sendo muito útil para o rastreio de substratos, enzimas, condições de reação, bem como a determinação de parâmetros cinéticos. O presente trabalho teve por objetivo estudar a biodegradação enzimática de 2,4,6-Triclorofenol, com a utilização das enzimas Lacase e Soybean Peroxidase em microrreator da Syrris com volume de 250 ?l, que permite o estudo de cinéticas muito rápidas. Para as análises de degradação utilizou-se duas enzimas, a Lacase em concentrações de 0,05; 0,1 e 0,2 mg/ml; e a Soybean Peroxidase em concentrações de 0,0005; 0,001 e 0,002 mg/ml com a adição de Peróxido de Hidrogênio. Através dos ensaios realizados obteve-se dados experimentais da reação enzimática, possibilitando a verificação da taxa inicial de reação e sua cinética. Posteriormente, realizou-se as análises em simulação utilizando os dados experimentais, que através de um sistema de EDOs estimando inicialmente as constantes cinéticas k1, k2 e k3 usando a ferramenta ESTIMA, onde apresentaram duas respostas, uma resposta típica de mínimos quadrados, e a outra resposta que a velocidade inicial, que foi melhor representada pelos parâmetros obtidos. O método empregado na degradação do substrato, o microrreator mostrou-se eficiente, permitindo a detecção de baixo consumo de substrato para a determinação da taxa inicial, em curto tempo de residência. Perante os ensaios realizados com Lacase e Soybean Peroxidase, o microrreator é também um equipamento eficaz na repetitividade e na reprodutibilidade dos dados obtidos em diferentes concentrações. / The microreactor is part of a set of devices in a new and promising technology, which can be called micro manufactured, active in fields such as chemical, biological, pharmaceutical, chemical engineering and biotechnology. It is a device that enables chemical reactions, such as conventional reactors, but with smaller dimensions, in the micrometer scale channels. Miniaturization technology devices for chemical reactions is expanding promoting an important development, with microsystems covering most effective devices, configuration and specific geometries and lower power consumption, where reactions with high transportation fees can be used for many different purposes such as fast reactions, mixing, temperature sensitive reactions, homogenization temperature or even precipitation of nanoparticles. Because of its scale is greatly reduced compared to the macro scale, provide a system which allows an investigation of the process in a short time, being very useful for screening for substrates, enzymes, reaction conditions, and the determination of kinetic parameters. One of the advantages of using microreactors is that this equipment requires small amounts of reagents for performing a catalytic reaction of action, and is very important when dealing with enzyme as a catalyst. This study aimed to study the enzymatic biodegradation of 2,4,6-Trichlorophenol with the use of laccase and Soybean Peroxidase enzymes in microreactor Syrris with volume of 250 ?l, which allows the study of very fast kinetics. For degradation analyzes were used two enzymes, laccase concentrations of 0.05; 0.1 and 0.2 mg / ml; and Soybean peroxidase at concentrations of 0.0005; 0.001 and 0.002 mg / ml with the addition of Hydrogen Peroxide. Through trials was obtained experimental data from enzyme reaction, allowing the verification of the initial reaction rate and its kinetics. Later, there was the analysis simulation using the experimental data, which through a system of ODEs initially estimating the rate constants k1, k2 and k3 using the ESTIMA tool, which had two answers, a typical response of least squares, and another answer to the initial rate, which was best represented by the parameters obtained. The method used in substrate degradation, the microreactor was efficient, allowing low substrate consumption detection for determining the initial rate in the short residence time. Before the tests with Laccase and Soybean Peroxidase, the microreactor is also an effective equipment in the repeatability and reproducibility of the data obtained at different concentrations.

Os compostos fenólicos, quando em altas concentrações na água, são poluentes de alta periculosidade, difíceis de serem eliminados até mesmo por métodos convencionais, como por exemplo, a biorremediação microbiológica dos lodos ativados e os métodos físico-químicos. Sabendo da presença dos fenóis em muitos efluentes industriais se faz necessário o estudo de novos processos para o tratamento desses efluentes que possuam aplicações eficazes e, sobretudo, ecologicamente corretas. A presente pesquisa visou a degradação de poluentes fenólicos em soluções aquosas através da oxidação enzimática com tirosinase imobilizada em quitosana. A contribuição tecnológica relaciona-se com a possibilidade de aplicação do estudo em escala industrial em processos de tratamento de efluentes industriais que contenham poluentes fenólicos. Fenóis como o 4-clorofenol, 4-cresol, 4-nitrofenol e o próprio fenol, foram utilizados em ensaios de oxidação enzimática em vaso agitado, em regime dinâmico, em processo de batelada, temperatura de 45°C e três faixas de concentração enzimática (40, 60 e 80 U/ml). Além disso, foram testadas três formas de quitosana utilizadas na imobilização enzimática e aplicadas nos processos de oxidação de fenol: esferas, flocos e micro partículas de quitosana. A enzima tirosinase foi eficiente na degradação de fenol, 4-cresol e 4-clorofenol, diminuindo consideravelmente a concentração destes poluentes nas soluções aquosas. Porém, a tirosinase não oxidou significativamente o 4-nitrofenol. Verificou-se que o efeito de alguns substituintes do anel aromático do fenol e a posição desse substituinte no anel aromático, exerce influência direta na atividade da enzima durante as reações oxidativas envolvendo os compostos fenólicos. Embora o presente estudo tenha apresentado bons resultados na remoção de alguns compostos fenólicos em soluções aquosas por meio da oxidação de tais poluentes pela enzima tirosinase imobilizada em quitosana, o complexo enzima-suporte não apresentou a mesma eficiência nos ensaios subseqüentes, nos quais foi estudada a reutilização da enzima imobilizada. / The phenolic compounds when in high concentrations in water are extremely dangerous pollutants and difficult to be eliminated even by conventional methods such as the microbiological bioremediation with activated sludge and physico-chemical methods. Phenols are present in many industrial effluents and is necessary to develop new and effective processes for the treatment of that effluents and, if possible, environmentally friendly. This research aimed the degradation of phenolic pollutants in aqueous solutions by enzymatic oxidation with tyrosinase immobilized in chitosan. Phenols such as 4-chlorophenol, 4-cresol, 4-nitrophenol and phenol, had been used in assays of enzymatic oxidation in an agitated reactor, in dynamic regime, in a batch process, temperature of 45°C and three values of enzymatic concentration ( 40, 60 and 80 U/ml). In addition, three forms of chitosan had been used in enzymatic immobilization and applied in the processes of oxidation of phenol: pellets, flakes and small particles of chitosan. Tyrosinase was efficient in the degradation of phenol, 4-cresol and 4-chlorophenol, reducing significantly the concentration of these pollutants in aqueous solutions. However, tyrosinase did not oxidized 4-nitrophenol. It was verified that the effect of some substitutes and their position in the aromatic ring has a direct influence on the activity of the enzyme during the oxidative reactions involving phenolic compounds. Although this study has shown good results in the removal of some phenolic compounds in aqueous solutions through the oxidation of such pollutants by the tyrosinase immobilized on chitosan, the enzyme-support did not present the same efficiency in subsequent assays, in which we studied the reuse of the immobilized enzyme.