Stensel, H. DLust, Mariko2014-04-302015-12-142014-04-302014Lust_washington_0250E_12874.pdfhttp://hdl.handle.net/1773/25440Thesis (Ph.D.)--University of Washington, 2014The discharge of the synthetic estrogen, 17α-ethinylestradiol (EE2), in wastewater treatment plant (WWTP) effluents is an environmental concern as this compound can alter the reproductive system of aquatic wildlife at low ng/L concentrations. The impact of EE2 at such low concentrations indicates the need to identify activated sludge (AS) process designs that minimize WWTP effluent EE2 concentrations. An EE2 fate and transformation model was developed based on the following mechanisms: (1) EE2 production from deconjugation of EE2-3-sulfate (EE2-3S), a conjugated form excreted from humans, (2) EE2 removal from biodegradation by heterotrophic biomass growing on other substrates, and (3) EE2 removal from sorption to activated sludge. These mechanisms were incorporated into the International Water Association (IWA) Activated Sludge Model No. 2d (ASM2d) to model the fate of EE2 across aerobic and biological nutrient removal (BNR) AS systems. The model was successfully calibrated and evaluated using lab-scale aerobic and BNR AS sequencing batch reactors (SBRs) fed primary effluent. A sensitivity analysis predicted effluent EE2 concentrations were most sensitive to the biodegradation rate coefficient, the influent biodegradable chemical oxygen demand (bCOD) to EE2 ratio, and the aerobic SRT and were less sensitive to the deconjugation rate coefficient and the solid-liquid partitioning coefficient. EE2 biodegradation kinetics were further investigated using lab-scale SBRs at 20°C fed synthetic wastewater. Three sets of reactor experiments were conducted using different municipal AS plant seed sources and with solids retention times (SRTs) ranging from 8 to 13 days. Significant EE2 biodegradation occurred only under aerobic conditions. Pseudo first-order biodegradation rate coefficients (k_b) normalized to the reactor volatile suspended solids (VSS) concentration ranged from 4 to 22 L/g VSS-d, 4 to 19 L/g VSS-d, and 3 to 20 L/g VSS-d for aerobic, anaerobic/aerobic, and anoxic/aerobic AS processes, respectively. Enriched denitrifying communities selected by anoxic-only operation did not degrade EE2 under anoxic or aerobic conditions. The variation in EE2 k_b values suggests there is a high degree of uncertainty in this value when predicting process performance. Experiments were conducted with an EE2-degrading isolate, Rhodococcus equi, to examine its biodegradation kinetics, EE2 degradation inhibition, and application of a transposon mutagenesis technique to identify genes involved in EE2 degradation. R. equi degraded EE2 during exponential growth on glucose with a relatively low k_b of 3.4 L/g VSS-d at 27°C, and did not degrade EE2 during the stationary growth phase. Clotrimazole inhibited EE2 degradation, suggesting a cytochrome p450 mono-oxygenase may be involved in EE2 degradation. Transposome mutagenesis coupled with a yeast estrogen screen (YES) assay to screen mutants was successfully applied to isolate an R. equi mutant with lost EE2-degrading ability. This technique and/or mutant may be useful for future research aimed at elucidating the EE2 degradation pathway(s).application/pdfen-USCopyright is held by the individual authors.Environmental engineeringcivil engineeringThesis