The increasingly stringent environmental regulations on hazardous wastes has encouraged the search for innovative solutions for the remediation of contaminated wastewaters. In this field, bioremediation is seen as an attractive solution due to its reputation as a low cost, environmentally friendly and publicly acceptable treatment technology. The present research program explores new strategies for the bio-treatment of dyecontaminated wastewaters produced by textile industries. Two kind of processes have been specifically analysed: biodegradation and biosorption based processes. Their effectiveness was tested on three textile wastewater models (Acid, Direct, Reactive), by assessing and comparing their performances in terms of decolourisation ability, COD and toxicity reduction of the treated effluent. The inactivated biomasses of three Mucorales fungi have been tested as potential candidates for dye biosorption. They displayed good sorption capabilities towards all the tested synthetic effluents, giving rise to decolourisation percentages up to 94% and decrease in COD up to 58%, coupled to a significant toxicity reduction. Biodegradation based processes, have been explored from a multiplicity of perspectives, by testing the degrading performances of whole-cell as well as enzyme-based biosystems, in both free and immobilized forms. The white-rot fungi Pleurotus ostreatus and Phanerochaete chrysosporium and their oxidative enzymes were the object of this investigation. Both fungi showed good decolourisation and detoxification capabilities towards the model wastewaters, exhibiting different specificities. As far enzymatic biosystems, the study has been specifically focused on laccases from P. ostreatus. Optimal conditions for laccase production by P. ostreatus have been defined and an extra-cellular enzyme mixtures, with known laccase isoenzyme composition, was produced and tested for its decolourisation potential, showing the best performances toward Acid wastewater model. The same laccase preparation was successfully immobilized on perlite, a siliceous inert carrier, and preliminary tested for the conversion of the reactive dye Remazol Brilliant Blue R. The immobilization process was optimized with reference to the immobilization yield and to the dye adsorption capacity of the solid biocatalyst. Stability and catalytic parameters of immobilized laccases, in comparison with those observed for free enzyme, were also assessed. A parallel session of the project was aimed at identifying and at characterizing new members of P. ostreatus laccase gene family. This research pathway was ultimately targeted at enriching the knowledge of this complex enzymatic family as well as at developing new bio-catalysts for wastewater treatment. The existence of a laccase gene clustering was demonstrated in P. ostreatus and three new laccase genes were cloned, thus enlarging the panel of these biocatalysts up to seven members. A “laccase subfamily” consisting of three laccase genes was identified. cDNAs coding for two of the new laccases were isolated and expressed in yeasts in order to characterize the recombinant proteins. A peculiar intron-exon structure was revealed for one of the new laccases, along with a high instability of the recombinant enzyme due to lability of its copper ligand.
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