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Study of Relevant Factors in the Treatment of Effluents by Fungi for the Degradation of Emerging Contaminants

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Study of Relevant Factors in the Treatment of Effluents by Fungi for the Degradation of Emerging Contaminants Departament d’Enginyeria Química Escola d’Enginyeria Universitat Autònoma de Barcelona Study of relevant factors in the treatment of effluents by fungi for the degradation of emerging contaminants PhD Thesis Marina Badia Fabregat Bellaterra, Setembre 2013 Title: Study of relevant factors in the treatment of effluents by fungi for the degradation of emerging contaminants Carried out by: Marina Badia Fabregat Supervised by: M. Teresa Vicent Huguet and Glòria Caminal Saperas PhD program in Environmental Science and Technology Departament d’Enginyeria Química Escola d’Enginyeria Universitat Autònoma de Barcelona Bellaterra, 2014 This work has been funded by the Spanish Ministries of MICINN (projects CTQ2010-21776- C02, CTM2013-48545-C2 and AIB2010PT-00169) and MMAMRM (project 010/PC08/3-04.1). The author acknowledges the predoctoral fellowship from Universitat Autònoma de Barcelona and the research stay grant from the Generalitat de Catalunya (2012 BE1 00210). Part of this work has been done in collaboration with the Institute of Environmental Assessment and Water Studies (IDAEA) from the Department of Environmental Chemistry, CSIC (Barcelona, Spain), the Catalan Institute for Water Research (ICRA) (Girona, Spain) and the Department of Crystallography, Mineralogy and Mineral Deposits in the University of Barcelona (Barcelona, Spain). Part of the work presented has been performed in research stays at Centre of Biological Engineering (CEB) of University of Minho under the supervision of Dr. Alcina Pereira and Dr. Madalena Alves and at Finnish Forest Research Institute (METLA) under the supervision of Dr. Taina Pennanen and Dr. Hannu Fritze. GLÒRIA CAMINAL SAPERAS, Científica Titula del CSIC a la Unitat de Biocatàlisis Aplicada associada al IIQAB, i M. TERESA VICENT I HUGUET, Professora Titular del Departament d’Enginyeria Química de la Universitat Autònoma de Barcelona, CERTIFIQUEM: Que la llicenciada en Biotecnologia Marina Badia Fabregat ha realitzat sota la nostra direcció, en els laboratoris del Departament d’Enginyeria Química, el treball que amb el títol “Study of relevant factors in the treatment of effluents by fungi for the degradation of emerging contaminants” es presenta en aquesta memòria, la qual constitueix la seva Tesi per a optar al Grau de Doctor per la Universitat Autònoma de Barcelona. I perquè en prengueu coneixement i consti als efectes oportuns, presentem a l’Escola d’Enginyeria de la Universitat Autònoma de Barcelona l’esmentada Tesi, signant el present certificat a: Bellaterra, Setembre 2014 ACKNOWLEDGEMENTS First of all, I would like to thank my thesis supervisors Teresa and Glòria for offering me the opportunity to work in their research group. I sincerely appreciate their advice and encouragement from the beginning until the end. Thank you to all the “Toxics” people, with whom I shared some steps of this thesis: Edu, Marcel, Carles, Carlos, Belén, Rim, Guillem, Andrea and Cesc. Of course, I would also like to express my gratitude to all the “permanent staff” of the group, who in a way or another helped me at different points of my research: Lucia(s), Ernest, Paqui and Montse. I also want to acknowledge the researchers of the chemical analytical groups with whom we have shared projects, without them this work with real effluents would not have been possible. Thus, I would like to specially thank Pablo Gago and Sílvia Díaz from IDAEA-CSIC and Daniel Lucas, Marta Llorca, Meritxell Gros and Sara Rodríguez from ICRA. Thank you also to Dr. Benjamí Piña to provide us with the RYA strains and to his group, in particular to Alba Olivares, for her time, dedication and patience to show me the protocol for RYA assays. Of course, I have a special mention to Ernest and Mònica for making me discover the world of stable isotopes. I would also like to acknowledge the advice from Dr. Petra Bombach about mathematical calculations behind isotope labelling and Dr. Magdalena Grifoll for her kindness on sharing her laboratory for some experimental settings. I would also like to acknowlege the staff of the veterinary hospital of UAB to let us collect the water and thanks also to the maintenance team of the veterinary faculty for their help. Thank you to Jordi Moreno, Michele Stefani, Hèctor Monclús and Joaquim Comas from Lequia (UdG), as well, for providing us with the reverse osmosis concentrate. I would like to aknowledge Dr. Madalena Alves and especially Dr. Alcina Pereira for their dedication at my work during the stay in Braga. Thank you also to all the labmates, especially Marta. Despite it was a short visit, they made me feel like I had been always there. Thanks to Dr. Taina Pennanen and Dr. Hannu Fritze not only for their supervision in the lab but also for taking care of me in Finland. I would also like to thank Dr. Tero Tuomivirta. Of course, thank you to all Puimala people, who made those days a great experience. Finally, thanks to Lara to propose me to go to her former lab at METLA for my research stay, it was a good piece of advice. I would also like to acknowledge the rest of the members of the Chemical Department of UAB: professors, technicians, secretaries and colleagues, for making these years more pleasant. Special thanks to everybody from “Dimecres de Pastís” for all the good moments inside and outside the Department. Thank you too to my friends and Marcel’s family, with many of whom we have shared the same troubles and joys that theses entail. And, last but not least, thank you to the three most important persons in my life and who gave me more support during all this time and that, inevitably, have also suffered the consequences. To my parents and Marcel, thank you for everything! LIST OF CONTENTS ABSTRACT ........................................................................................................ 7 RESUM ............................................................................................................. 9 LIST OF ABBREVIATIONS ............................................................................. 11 Chapter 1. General introduction ................................................ 15 1.1. Emerging contaminants ......................................................................... 15 1.1.1. From priority pollutants to emerging contaminants ............................... 15 1.1.2. Sources of emerging contaminants ...................................................... 18 1.1.3. Occurrence and fate in the environment ............................................... 22 1.1.4. Environmental toxic effects and risk assessment ................................. 24 1.1.5. Treatments to degrade emerging contaminants ................................... 27 1.2. Biotechnological applications of white-rot fungi, and Trametes versicolor in particular .................................................................................. 32 1.2.1. Why white-rot fungi? ............................................................................. 32 1.2.2. Use of WRF for bioremediation ............................................................ 36 1.2.3. Fungal biodegradation of individual emerging contaminants ................ 37 1.2.4. Scale-up to bioreactors and treatment of real wastewater .................... 40 1.3. Use of molecular tools for treatment assessment ........................... 42 1.3.1. Use of stable isotopes to assess fate of pollutants ............................... 43 1.3.2. Use of molecular techniques for fungal treatment assessment ............ 44 Chapter 2. Objectives ................................................................. 49 Chapter 3. List of articles and author’s contribution ............... 53 Chapter 4. Materials and methods ............................................ 59 4.1. Microorganisms ...................................................................................... 60 4.1.1. Fungus .................................................................................................. 60 1 LIST OF CONTENTS 4.1.2. Recombinant yeasts ............................................................................. 60 4.2. Chemical compounds and reagents ..................................................... 61 4.2.1. Selected contaminants ......................................................................... 61 4.2.2. Chemical products and reagents .......................................................... 61 4.3. Wastewater effluents .............................................................................. 63 4.3.1. Reverse osmosis concentrate .............................................................. 63 4.3.2. Veterinary hospital wastewater effluent ................................................ 64 4.4. Media and culture conditions ................................................................ 64 4.4.1. Media for Trametes versicolor, culture conditions and pellets production ............................................................................................. 64 4.4.2. Media for RYA assays .......................................................................... 66 4.4.3. Growth and maintainance of Saccharomyces cerevisiae strains .......... 67 4.5. Degradation experiments of selected contaminants ........................... 68 4.5.1. Degradation of selected contaminants in serum bottles ....................... 68 4.5.2. Degradation of selected contaminants in bioreactors ........................... 69 4.5.3. Experiments with purified laccase ........................................................ 70 4.5.4. Experiments with cytochrome
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