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BIBLIOTECA DO INSTITUTO DE QUÍMICA
UNICAMP

 
TESE DE DOUTORADO
 
Autor: Soares, Carlos Henrique Lemos
Título: Estudos Mecanísticos da Degradação de Efluentes de Indústrias de Papel e Celulose por Fungos Basidiomicetos Degradadores de Madeira
Ano: 1998
Orientador: Prof. Dr. Nelson Eduardo Durán Caballero
Departamento: Físico-Química
Palavras-chave: Trametes Villosa, Panus crinitus, Lacase
Resumo: O fungo Trametes villosa promoveu a descoloração de 70-80% da intensidade inicial da cor do efluente, no período 35-50 hs de incubação com o mesmo. Concomitantemente ao processo de descoloração, a concentração total de fenóis, medida por dois métodos químicos independentes, método de Folin e método 4-aminoantipirina (4-AAP), decresceu ao longo do período de incubação. Os resultados obtidos pelo método Folin indicaram que o processo de descoloração foi mais rápido do que o consumo de fenóis totais. Tal resultado parece indicar que modificações químicas nas subestruturas dos componentes do efluente, particularmente nas de maior massa molecular, podem contribuir para o processo descoloração, concomitantemente, ou antes mesmo que estas sofram um processo de mineralização total (transformação a CO2). O pH inicial do efluente não foi um fator limitante da ação do fungo, pelo menos para valores menores que 9,5 , para efluentes não tamponados. O efluente tratado com o fungo Panus crinitus apresentou uma taxa de descoloração de 60-70% no período de 45-55 hs. Também neste caso ocorreu a redução acentuada da concentração total de fenóis. A avaliação das atividades enzimáticas extracelulares demonstrou que; o fungo T. villosa produziu quantidades apreciáveis das enzimas lacase, peroxidase totais, lignina peroxidase e de b-glicosidase, superior a todos os demais fungos testados. Já o fungo P. crinitus produziu quantidades ligeiramente inferiores das citadas enzimas, apenas nas primeiras 48 hs de incubação. Para o fungo Phellinus sp não foi detectada nenhuma atividade enzimática relativa a lacase e somente quantidades desprezíveis de atividade para b- glicosidase. Por outro lado, este fungo produziu quantidades moderadas de peroxidases e altas quantidades de Mn-peroxidase, mantidas durante todo período de incubação (20 dias). O fungo Phellinus sp não descoloriu apreciavelmente o efluente. Os resultados obtidos parecem sugerir que a enzima Mn-peroxidase, neste caso, não foi determinante da capacidade de descoloração e biodegradação do efluente. Diferenças significativas puderam ser observadas entre as curvas de distribuição de MM obtidas a 210 e 280 nm. A principal constatação foi a de que a fração de componentes de baixa massa molecular pôde ser melhor caracterizada pela curva obtida a 210 nm, do que pela respectiva curva a 280 nm. As variações observadas nas curvas de distribuição de massas moleculares indicaram que todas as frações; alta. média e baixa-MM, foram degradadas pela ação tanto do fungo Trametes villosa, quanto à do fungo Panus crinitus. Espectros-UV diferenciais de efluentes tipo E1 obtidos durante o período de incubação com o fungo T. villosa revelaram importantes modificações estruturais promovidas por este fungo; no período inicial (0- 48 hs) ocorreu a formação progressiva de um grupamento conjugado com o anel aromático evidenciada pelo aumento acentuado de absorbância a 310 nm. Pelo fato de que tal absorção pode ser suprimida pela redução com borohidreto de sódio, o grupamento intermediário formado deve ser tipo carbonílico. O fungo P. crinitus não apresentou esta característica. A análise dos espectros FT-IR das amostras de efluentes revelou que o processo Kraft de polpação e o processo de branqueamento alteram significativamente a estrutura da lignina residua, contida no efluente, sendo que o conteúdo de estruturas aromáticas, neste caso, é baixo. Também esta técnica, FT-IR, demonstrou a formação de grupo carbornílico com absorção em torno de 1650 cm, decorrente da ação do fungo T. villosa sobre o efluente.
Abstract: The basidiomycete fungi Panus crinitus and Trametes villosa have been studied for their ability to degrade and decolorize Kraft pulp bleaching effluents (E1-stage). The results showed that the white-rot fungi P. crinitus and T. villosa exhibited a decolorization rate about 55-65% and 70-80%, respectively, during 1-2 days. The total phenolic content decreased in agreement with the decolorization rate. The production of lignocellulose-degrading enzymes as lignin-peroxidase (LiP), laccase, manganese peroxidase (MnP), b-glucosidase, glucose oxidase has been evaluated. The ligninolytic enzyme patterns were quite similar, being that laccase and lignin peroxidase were the major extracellular ligninolytic enzymes secreted by both the fungi, T. villosa and P. crinitus. The MnP extracellular activity was not detected under the conditions employed. On the other hand, these fungi exhibited differences in their individual behavior regarding to their LiP/laccase activity ratio, b-glucosidase activity levels, overall peroxidase activity levels and in the temporal profile of enzyme liberation into the extracellular media. Both fungi secreted substantial quantities of extracellular laccase (120-418 U 1) and b-glucosidase (30-145 U 1), a moderate rate a generic peroxidases, lignin peroxidase, and traces of manganese peroxidase. Laccase presented a maximum activity at 36 hs, for T. villosa, and at 50 hs for P. crinitus, after incubation, and it decreased slowly in the subsequent period. Peroxidases e b-glucosidases activities decreased during the degradation period.Glucose oxidase and oxalic acid oxidase activities have also been evaluated but the results were not conclusive. While some authors related the importance of manganese peroxidase activity on decolorization of the effluent by other fungi, we could not observe this effect in this case. It does not seem to be responsible for the decolorization or degradation of effluent by T. villosa and P. crinitus. Studies of changes of molecular-size distribution during the effluent incubation period with the fungi was carried out by size exclusion HPLC and the elution was concomitantly monitored at 210 nm and 280 nm. Significative differences could be verified in this case. We could observe that not only the two elution profiles, at 210 and 280 nm, were partially different, but, curiously, it was also evident that the formation of lower-MW intermediate compounds, during the biodegradation process, could be better characterized at 210 nm than at 280 nm. The fungi could efficiently degrade all the fractions of different molecular-mass, however, the individual degradation rate for each fraction was different. The present results showed that the depolymerizing pathway was preferential in the T. villosa and P. crinitus case. As laccase has been refereed as an enzyme which the polymerizing ability is predominant in vitro, a presence of a natural mediator for laccase depolimerization capability could be pointed out, on speculative-terms. The higher-molecular-weight fractions were chemically modified prior to their depolymerization and the decolorization process seems to be more correlated to this modification. Laccase may play an importante role in this case, however, synergistic effects from other enzymes cannot be ruled out. Minor polymerization reactions could be also found for both fungi. Bleached Kraft pulp effluent displays no typical differential UV spectrograms as for lignin and lignin related compounds. It showed maximum peak at about 220, 276, 310 nm and above 350 nm. Kinetic studies of the decolorization by reducing agents as sodium borohydride enhanced the presence of different carbonyl groups conjugated with -OH phenolic groups. The modifications observed during the biodegradation of the effluent indicated the formation of intermediary carbonyl groups, including the quinoid type, which were consumed in the final phase of the degradation. The differential spectrograms in the final phase were similar to those of the phenolic acids as observed for model compounds. Another important observation comes from the analysis of extracts obtained from the solvent extraction. The results suggested that the spectrogram observed for the effluent was a composition of absorbancies of the different molecular mass fractions contained in it. The structural changes during the biodegradation of E1-Kraft pulp bleaching effluent by basidiomycete fungi, Trametes villosa and Panus crinitus, were also characterized by using the FT-IR method. The major chemical modifications of E1- effluent observed during this biodegradation process can be summarized as follows; i) In despite of the fact that the lignin-derivated compounds contained in the untreated effluent showed a lower aromatic nuclei content than normal lignin (1500 cm region), and phenolic hydroxyl group content, both substructures were intensively degraded by fungal treatment. ii) The formation of a new type of intermediary conjugated carbonyl, which was removed upon reduction with sodium borohydride, and carboxyl groups were found (1630-1660 cm region) only in the T. villosa case, and was absent in the P. crinitus. Fungal attack changes the absorption maximum of original carbonyl absorption band in the untreated effluent spectra to higher wavenumbers. iii) The aliphatic moieties in the organic material dissolved in the effluent seem to be degraded faster than the aromatic ones. iv) The relative content of syringyl units or guaiacyl ring condensed structures into the treated effluent sample apparently increased (increased intensity at 1326 cm) during the incubation period with the T. villosa fungus, which may indicate a relative selectivity of its ligninolitic system. The same effect was not observed in the P. crinitus case.
Arquivo (Texto Completo): vtls000189128.pdf ( tamanho: 3,66MB )

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