Quinones as Electron Acceptors and Redox Mediators for the Anaerobic Biotransformation of Priority Pollutants

Humus is the most abundant organic fraction in the biosphere. It is composed of a complex structure in which recalcitrant polymers prevail with a residence time lasting decades or even centuries. Despite the recalcitrance of humic substances, they have recently been recognized to play an important role on the anaerobic conversion of organic matter by serving as an electron acceptor for microbial respiration. Quinone moieties are the responsible electron-accepting groups accounting for the microbial reduction of humus. Quinones and humus not only serve as terminal electron acceptors for microbial respiration, but they also function as redox mediators during the transfer of electrons in microbial and chemical reactions. In this dissertation the impact of humus and quinone analogues on the anaerobic biotransformation of ecologically important substrates, as well as priority pollutants, was evaluated.Consortia obtained from many different environments including sandy, organic rich, and contaminated sediments, as well as anaerobic and aerobic sludges, showed the capacity for oxidizing a wide variety of ecologically significant substrates, such as lactate and acetate, when the humic model compound, anthraquinone-2,6-disulfonate (AQDS), was provided as a final electron acceptor. AQDS-reducing microorganisms out-competed methanogens for most of the substrates supplied indicating that quinone reduction is a widespread physiological process, which may contribute to important carbon cycling process in many different environments. Quinone and humus reduction was also found in pure cultures of different microorganisms, such as Desulfitobacterium spp. and Methanospirillum hungatei, indicating that the ubiquity of quinone reduction may be due to the wide diversity of microorganisms with the capacity for reducing humic substances. The results also illustrate that phylogenetically distinct microorganisms can channel electrons from anaerobic substrate oxidation via quinone reduction towards the reduction of metal oxides. Quinone respiring microorganisms could also be enriched and immobilized in the microbial community of an anaerobic granular sludge of a upflow anaerobic sludge blanket (UASB) reactor. The feasibility to immobilize quinone-reducing microorganisms can be applied to accelerate the conversion of xenobiotics susceptible to reductive biotransformations such as azo dyes and polychlorinated compounds in continuous bioreactors.The long-term goal of this research was to explore the capacity of humus respiring consortia for oxidizing priority pollutants through the reduction of humic substances. Anaerobic granular sludge originated from different wastewater treatment plants were shown to oxidize phenol and p -cresol coupled to the reduction of AQDS. Both phenolic contaminants were converted to methane in the absence of the humic analogue, but addition of AQDS as an alternative electron acceptor diverted the flow of electrons from methanogenesis towards quinone reduction. Priority pollutants, which were not degraded under methanogenic conditions, could also be mineralized by humus-respiring consortia when humic substances were provided as an electron acceptor. Enriched sediments from different origins readily mineralized uniformly labeled [ <sup>13</SUP>C]toluene to <sup>13</SUP>CO ₂ when humic acids or AQDS were provided as terminal electron acceptors. Negligible recovery of <sup>13</SUP>CO ₂ occurred in the absence of humic substances. Additionally, the electrons in the toluene mineralized were recovered stoichiometrically as reduced humus or AH ₂ QDS (reduced form of AQDS).Humic substances were also shown to accelerate the transfer of reducing equivalents required for the anaerobic conversion of different pollutants containing electron-withdrawing groups. AQDS supplemented at sub-stoichiometric levels in granular sludge incubations enhanced the rate of conversion of carbon tetrachloride (CT) leading to an increased production of inorganic chloride. Negligible dechlorination occurred in sterile controls with autoclaved sludge and considerably less dechlorination was achieved in active controls lacking AQDS. A humus respiring enrichment culture, composed primarily of a Geobacter sp., derived from the same granular sludge was also shown to dechlorinate CT, yielding similar products as the AQDS-supplemented sludge consortium. Addition of catalytic levels of AQDS to a UASB reactor continuously treating the azo dye, acid orange 7 (AO7), also enhanced the biotransformation of this pollutant to the corresponding aromatic amines. High efficiency (&gt;90 %) of decolorization of AO7 occurred even at a hydraulic residence time of 2 hours with a molar ratio of AQDS/AO7 as low as 1/100, whereas 70 % of color removal occurred in the absence of AQDS under the same hydraulic conditions.The evidences provided in this study indicate that humic substances may play an important role on the stabilization of organic matter, as well as on the intrinsic bioremediation of contaminated environments, by serving as a terminal electron acceptor. The application of humic substances for achieving the bioremediation of contaminated aquifers can be considered. Humus and quinones can also be applied in anaerobic reactors to enhance the conversion of priority pollutants containing electron-withdrawing groups