Applied Science, Faculty of
Engineering, School of (Okanagan)
DSpace
UBCO
Cox, Alison E
Master of Applied Science - MASc
University of British Columbia
Hydrothermal Liquefaction (HTL) is a thermo-chemical method of processing wastewater solids that could be preferential over anaerobic digestion (AD) because it produces no biosolids. Instead, the effluent can be separated into biocrude which can be refined into fuel, hydrochar that can be used for adsorption applications and HTL aqueous, the only waste stream of this process. Unfortunately, HTL aqueous is high in ammonia, phenolics and nitrogen heterocyclic compounds that can inhibit AD if it is used for disposal. Ammonia stripping was tested for pretreatment of HTL aqueous and recovering ammonia. Seven stripping reactor conditions were run, and the best results were seen at 85°C, a pH of 9.3 and 500 mL/min air flow rate. This case achieved ammonia removal of 99.5 ± 0.1% and total phenolic compounds removal of 32 ± 1% in 4 hours. The ammonia stripping was coupled with acid adsorption to produce an ammonium sulphate salt with value as fertilizer. The ideal ammonia stripping conditions produced a salt with a purity of 98.0 ± 0.05% and represented an ammonia recovery of 73.9 ± 0.04%. Semi-continuous flow bench-scale thermophilic anaerobic co-digestion of HTL aqueous with municipal sludge found that 12% of influent chemical oxygen demand (COD) can come from HTL aqueous without inhibition if pretreated with ammonia stripping, while the digester fed 13% of COD from non-pretreated HTL aqueous showed inhibition. Under mesophilic conditions, semi-continuous flow anaerobic co-digestion was successful for digesters fed with either 24% non-pretreated HTL aqueous or 22% pretreated HTL aqueous.
AMMONIA STRIPPING OF HYDROTHERMAL LIQUEFACTION AQUEOUS PHASE FROM MUNICIPAL SLUDGE FOR ANAEROBIC CO-DIGESTION PRETREATMENT AND AMMONIA RECOVERY By Alison E Cox B.Eng., University of Guelph, 2019 A THESIS SUBMITTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF APPLIED SCIENCE in THE COLLEGE OF GRADUATE STUDIES (Civil Engineering) THE UNIVERSITY OF BRITISH COLUMBIA (Okanagan) May 2024 © Alison Cox, 2024 ii The following individuals certify that they have read, and recommend to the College of Graduate Studies for acceptance, a thesis entitled: Ammonia Stripping of Hydrothermal Liquefaction Aqueous Phase from Municipal Sludge for Anaerobic Co-digestion Pretreatment and Ammonia Recovery submitted by Alison Cox in partial fulfillment of the requirements of the degree of Master of Applied Science. Dr. Cigdem Eskicioglu, School of Engineering Supervisor Dr. Nicolas Peleato, School of Engineering Supervisory Committee Member Dr. Alyse Kiesser, School of Engineering Supervisory Committee Member Dr. Sepideh Pakpour, School of Engineering University Examiner Click or tap here to enter text. iii Abstract Hydrothermal Liquefaction (HTL) is a thermo-chemical method of processing wastewater solids that could be preferential over anaerobic digestion (AD) because it produces no biosolids. Instead, the effluent can be separated into biocrude which can be refined into fuel, hydrochar that can be used for adsorption applications and HTL aqueous, the only waste stream of this process. Unfortunately, HTL aqueous is high in ammonia, phenolics and nitrogen heterocyclic compounds that can inhibit AD if it is used for disposal. Ammonia stripping was tested for pretreatment of HTL aqueous and recovering ammonia. Seven stripping reactor conditions were run, and the best results were seen at 85°C, a pH of 9.3 and 500 mL/min air flow rate. This case achieved ammonia removal of 99.5 ± 0.1% and total phenolic compounds removal of 32 ± 1% in 4 hours. The ammonia stripping was coupled with acid adsorption to produce an ammonium sulphate salt with value as fertilizer. The ideal ammonia stripping conditions produced a salt with a purity of 98.0 ± 0.05% and represented an ammonia recovery of 73.9 ± 0.04%. Semi-continuous flow bench-scale thermophilic anaerobic co-digestion of HTL aqueous with municipal sludge found that 12% of influent chemical oxygen demand (COD) can come from HTL aqueous without inhibition if pretreated with ammonia stripping, while the digester fed 13% of COD from non-pretreated HTL aqueous showed inhibition. Under mesophilic conditions, semi-continuous flow anaerobic co-digestion was successful for digesters fed with either 24% non-pretreated HTL aqueous or 22% pretreated HTL aqueous. iv Lay Summary This thesis proposes a disposal method of the aqueous waste product for a new green energy production method. In this method, a liquid petroleum product is produced through heating solids removed from wastewater to 350ºC for 15 minutes in a pressurized container. The research focused on a thermo-chemical pretreatment process to allow disposal of this aqueous waste product in a biological treatment process already found at many wastewater treatment plants. A removal of 79.5 ± 0.1% ammonia, 32 ± 1% phenolic compounds and 38 ± 1% organic acids was achieved during pretreatment. Ammonia removed was captured as a fertilizer salt with a purity of 98%. The pretreated aqueous waste was then tested in a lab-scale version of the biological treatment process proposed for disposal. Results of this testing suggest that the pretreatment was successful in increasing the volume of aqueous waste that can be treated in this treatment process. v Preface The following research was completed by the author at the University of British Columbia School of Engineering, under the supervision of Dr. Cigdem Eskicioglu of the Bioreactor Technology Group (BTG). This includes laboratory experimentation of the ammonia stripping reactors, operation of the anaerobic digesters during the recorded run-time, material characterization, analysis of results and writing of this thesis. Identification of the pretreatment to be researched and preliminary design of the experiment was done by Dr. Parmila Devi (Research Engineer at BTG) and generation of the hydrothermal liquefaction aqueous waste product was assisted by Ph.D. candidates, Alper Basar and Huan Liu, at BTG. The ‘HTL Aqueous Pretreatment’ section of the results chapter has been presented at British Columbia Water Works Association annual conference in May 2023. The presentation was entitled “Hydrothermal Liquefaction Aqueous Detoxification and Resource Recovery through Ammonia Stripping” and was prepared and presented by the author. An abstract has been accepted to the International Water Association World Water Congress and Exhibition in August 2024 in Toronto on the ‘Anaerobic Co-digestion of HTL Aqueous’ section of the results. Preparation is also underway to summarize the research presented in this thesis into a journal article. vi Table of Contents Abstract .................................................................................................................................... iii Lay Summary ........................................................................................................................... iv Preface ....................................................................................................................................... v Table of Contents ..................................................................................................................... vi List of Tables ............................................................................................................................. x List of Figures ......................................................................................................................... xii List of Abbreviations ............................................................................................................. xvi Acknowledgements .............................................................................................................. xvii Dedication ............................................................................................................................ xviii Chapter 1: Introduction .................................................................................................. 1 1.1 Project Objectives ........................................................................................... 1 1.2 Novelty of Research ........................................................................................ 2 1.3 Thesis Sections ............................................................................................... 2 Chapter 2: Literature Review ......................................................................................... 4 2.1 Hydrothermal Liquefaction of Organic Biomass ............................................... 4 2.2 Hydrothermal Liquefaction Aqueous Product Disposal Pathways .................... 5 2.2.1 Recycling in HTL ....................................................................................... 6 2.2.2 Return to Headworks ................................................................................. 6 2.2.3 Sending to Anaerobic Digesters................................................................. 7 2.2.4 Sending to other Anaerobic Reactors ........................................................ 9 2.3 Hydrothermal Liquefaction Aqueous Product Pretreatment Options ................ 9 2.3.1 Carbon Adsorption Pretreatment Methods ................................................. 9 2.3.2 Struvite Precipitation Methods ..................................................................10 2.3.3 Catalytic Hydrothermal Gasification (CHG) Methods ................................10 vii 2.3.4 Biological Pretreatment Methods ..............................................................11 2.3.5 Ammonia Stripping Methods .....................................................................12 2.4 Literature Review Summary ...........................................................................14 Chapter 3: Materials and Methods ...............................................................................15 3.1 Equipment ......................................................................................................15 3.2 HTL Operation to Generate Aqueous Phase ..................................................16 3.3 Ammonia Stripping and Recovery Method .....................................................18 3.4 Anaerobic Co-digestion Method .....................................................................21 3.5 Analytical Characterization Methods ..............................................................26 3.5.1 Total Ammonium Nitrogen (TAN) ..............................................................26 3.5.2 Total Phenolics .........................................................................................28 3.5.3 Volatile Fatty Acids (VFA) .........................................................................29 3.5.4 Total Chemical Oxygen Demand (tCOD) ..................................................29 3.5.5 pH .............................................................................................................30 3.5.6 Total Alkalinity ..........................................................................................30 3.5.7 Total and Volatile Solids (TS and VS) .......................................................30 3.5.8 Total and Volatile Suspended Solids (TSS and VSS) ...............................31 3.5.9 Salt Purity .................................................................................................32 3.5.10 Biogas Volume .......................................................................................33 3.5.11 Biogas composition .................................................................................33 3.5.12 Heavy Metals ..........................................................................................33 3.5.13 Capillary Suction Time ............................................................................34 3.5.14 Fecal Coliforms .......................................................................................35 3.5.15 Nitrogen Heterocyclic Compounds ..........................................................35 3.5.16 Statistical Analysis of Data ......................................................................36 viii Chapter 4: Results and Discussion ..............................................................................37 4.1 HTL Aqueous Pretreatment ............................................................................37 4.1.1 Ammonia Stripping Feed Characterization ................................................37 4.1.2 Ammonia Mass Removal ..........................................................................38 4.1.3 Phenolic Compounds Mass Removal .......................................................41 4.1.4 VFA Mass Removal ..................................................................................42 4.1.5 COD Mass Removal .................................................................................43 4.1.6 Alkalinity Mass Removal ...........................................................................44 4.1.7 Total and Volatile Suspended Solids Mass Removal ................................46 4.1.8 Ammonium Sulphate Salt Analysis ...........................................................46 4.2 Anaerobic Co-digestion of HTL Aqueous .......................................................48 4.2.1 Co-digestion Feed Characterization ..........................................................49 4.2.2 Biogas Production and Composition .........................................................50 4.2.3 Total Ammonium Nitrogen (TAN) ..............................................................56 4.2.4 Volatile Fatty Acids ...................................................................................58 4.2.5 Chemical Oxygen Demand .......................................................................60 4.2.6 Alkalinity and pH .......................................................................................62 4.2.7 Total Phenolic Compounds Concentration ................................................64 4.2.8 Total Solids and Volatile Solids .................................................................66 4.2.9 Heavy Metals ............................................................................................69 4.2.10 Specific Capillary Suction Time ...............................................................72 4.2.11 Fecal Coliform Density ............................................................................73 4.2.12 Nitrogen Heterocyclic Compounds ..........................................................74 Chapter 5: Conclusions ................................................................................................77 5.1 Limitations and Future Research ....................................................................80 ix Bibliography ............................................................................................................................ 81 Appendices ............................................................................................................................. 86 Appendix A Calibration Curves for Colorimetric Tests ..............................................86 Appendix B Scanning Electron Microscope Salt Imaging .........................................88 Appendix C Energy Dispersive Spectroscopy...........................................................92 Appendix D Sample Statistical Analysis Tables ........................................................95 x List of Tables Table 3-1: Equipment inventory for all experiments performed in the BTG laboratory. ..............15 Table 3-2: Ammonia stripping experimental design parameters. ...............................................20 Table 3-3: Ammonia stripping parameters for HTL aqueous pretreatment ahead of anerobic co-digestion. ..................................................................................................................................22 Table 3-4: Bench-scale anaerobic digester conditions ..............................................................23 Table 3-5: Average HTLaq contribution to organic loading rate for each digester condition compared to the flowrate contributions of each co-substance. ..................................................25 Table 3-6: Anaerobic co-digestion experiment sampling schedule. ...........................................26 Table 4-1: Characterization of mixed sludge and HTL aqueous for the ammonia stripping experiment. Parameter values represent the mean concentration of triplicate samples accounting for standard deviation between replicates. ..............................................................37 Table 4-2: ANOVA summary table for analysis of significant mean difference among the ammonia mass removal of ammonia stripping run 1-3 with varying temperatures at 95% confidence (α = 0.05). ...............................................................................................................39 Table 4-3: ANOVA summary table for analysis of significant mean difference among the ammonia mass removal of ammonia stripping runs 3-5 with varying pH setpoints at 95% confidence (α = 0.05). ...............................................................................................................40 Table 4-4: ANOVA summary table for analysis of significant mean difference among the ammonia mass removal of ammonia stripping runs 3,6 and 7 with varying air flow rates at 95% confidence (α = 0.05). ...............................................................................................................40 Table 4-5: Total alkalinity at the start of the ammonia stripping reaction and in the flask effluent along with calculated mass removal during the 3-hour run time. ...............................................45 Table 4-6: Total and volatile suspended solids mass removal from HTLaq during ammonia stripping. ...................................................................................................................................46 xi Table 4-7: CHNS analysis results and purity in the recovered ammonium sulphate salt. ...........47 Table 4-8: Summary of nitrogen recovery from HTLaq during ammonia stripping runs. ............47 Table 4-9: Characterization of co-digestion inoculum. Parameter values represent the mean concentration of triplicate samples accounting for standard deviation between replicates. ........48 Table 4-10: Characterization of mixed sludge, centrate and HTL aqueous for the co-digestion feedstock. Parameter values represent the mean concentration of triplicate samples accounting for standard deviation between replicates. ................................................................................49 Table 4-11: Z-test summary table for analysis of significant mean difference between the specific biogas yield of non-pretreated and pretreated HTLaq fed thermophilic digesters at 95% confidence (α = 0.05). ...............................................................................................................52 Table 4-12: Z-test summary table for analysis of significant mean difference between the specific biogas yield of non-pretreated and pretreated HTLaq fed mesophilic digesters at 95% confidence (α = 0.05). ...............................................................................................................55 Table 4-13: Heavy metal concentrations in anaerobic digester effluents compared to biosolids regulations. ...............................................................................................................................71 Table 4-14: Nitrogen heterocyclic compound concentration in the anaerobic digester effluent. .75 Table 4-15: Nitrogen heterocyclic compound concentrations in non-pretreated and pretreated HTLaq. ......................................................................................................................................76 Table D-1: T-test summary table for analysis of significant mean difference between the COD removal of untreated and pretreated HTLaq fed thermophilic digesters at 95% confidence (α = 0.05). ........................................................................................................................................95 Table D-2: T-test summary table for analysis of significant mean difference between the COD removal of untreated and pretreated HTLaq fed mesophilic digesters at 95% confidence (α = 0.05). ........................................................................................................................................95 xii List of Figures Figure 3-1: Bench-scale Parr reactor used for HTL treatment of municipal sludge. ...................16 Figure 3-2: Process flow diagram for Annacis Island WWTP solids processing (Coulman, 2022). .................................................................................................................................................17 Figure 3-3: Ammonia stripping and acid adsorption apparatus. .................................................18 Figure 3-4: An image of the ammonium sulphate salt produced during each ammonia stripping run. ...........................................................................................................................................21 Figure 3-5: An image of mesophilic anaerobic digesters setup in 1 L erlenmeyer flasks, tedlar bags for biogas capture, and tubing for feeding and effluent removal. .......................................22 Figure 3-6: Anaerobic co-digestion experimental design. ..........................................................24 Figure 3-7: Image taken during TAN testing, showing the gradient of colour change across the calibration standards prior to being placed in the spectrophotometer. .......................................27 Figure 3-8: Image of microplate of samples being placed into the for total phenolics testing. ....28 Figure 3-9: Ammonium sulphate samples, coated in platinum and palladium, in the SEM/EDS setup. ........................................................................................................................................32 Figure 3-10: Capillary suction timer used for testing dewaterability of digester effluent. ............34 Figure 4-1: Ammonia mass removal from HTLaq during ammonia stripping runs at three different temperatures. ..............................................................................................................38 Figure 4-2: Ammonia mass removal from HTLaq during ammonia stripping runs at three different pH setpoints. ...............................................................................................................39 Figure 4-3: Ammonia mass removal from HTLaq during ammonia stripping runs at three different airflow rates. ................................................................................................................40 Figure 4-4: Total phenolic compounds removal from HTLaq during all ammonia stripping runs. .................................................................................................................................................42 xiii Figure 4-5: Volatile fatty acid removal from HTLaq during all ammonia stripping runs. ..............43 Figure 4-6: Total chemical oxygen demand removal from HTLaq during all ammonia stripping runs. ..........................................................................................................................................44 Figure 4-7: Biogas production of the thermophilic control digester, digester fed non-pretreated HTLaq (HTLaq), and digester fed HTLaq pretreated through ammonia stripping (HTLaqSTRIP). The contribution of non-pretreated and pretreated HTLaq in the feedstock is indicated at the top of the graph. ..............................................................................................................................50 Figure 4-8: Specific biogas yield of the thermophilic control digester, digester fed non-pretreated HTLaq (HTLaq), and digester fed HTLaq pretreated through ammonia stripping (HTLaqSTRIP). The contribution of non-pretreated and pretreated HTLaq in the feedstock is indicated at the top of the graph. ..............................................................................................................................51 Figure 4-9: Biogas production of the mesophilic control digester, digester fed non-pretreated HTLaq (HTLaq), and digester fed HTLaq pretreated through ammonia stripping (HTLaqSTRIP). The contribution of non-pretreated and pretreated HTLaq in the feedstock is indicated at the top of the graph. ..............................................................................................................................53 Figure 4-10: Specific biogas yield of the mesophilic control digester, digester fed non-pretreated HTLaq (HTLaq), and digester fed HTLaq pretreated through ammonia stripping (HTLaqSTRIP). The contribution of non-pretreated and pretreated HTLaq in the feedstock is indicated at the top of the graph. ..............................................................................................................................54 Figure 4-11: Biogas composition in thermophilic anaerobic digesters. ......................................55 Figure 4-12: Biogas composition in mesophilic anaerobic digesters. .........................................56 Figure 4-13: Total ammonium nitrogen (TAN) concentration in the thermophilic anaerobic digester influent and effluent. ....................................................................................................56 Figure 4-14: Total ammonium nitrogen (TAN) concentration in the mesophilic anaerobic digester influent and effluent. ..................................................................................................................57 Figure 4-15: Volatile fatty acid concentration in the thermophilic anaerobic digester influent and effluent. .....................................................................................................................................59 xiv Figure 4-16: Volatile fatty acid concentration in the mesophilic anaerobic digester influent and effluent. .....................................................................................................................................60 Figure 4-17: Total chemical oxygen demand concentration in the thermophilic anaerobic digester influent and effluent. ....................................................................................................61 Figure 4-18: Total chemical oxygen demand concentration in the mesophilic anaerobic digester influent and effluent. ..................................................................................................................61 Figure 4-19: Total alkalinity concentration in the thermophilic anaerobic digester influent and effluent. .....................................................................................................................................63 Figure 4-20: Total alkalinity concentration in the mesophilic anaerobic digester influent and effluent. .....................................................................................................................................63 Figure 4-21: Total phenolics concentration in the thermophilic anaerobic digester influent and effluent. .....................................................................................................................................64 Figure 4-22: Total phenolics concentration in the mesophilic anaerobic digester influent and effluent. .....................................................................................................................................65 Figure 4-23: Total solids concentration in the thermophilic anaerobic digester influent and effluent. .....................................................................................................................................66 Figure 4-24: Total solids concentration in the mesophilic anaerobic digester influent and effluent. .................................................................................................................................................67 Figure 4-25: Volatile solids concentration in the thermophilic anaerobic digester influent and effluent. .....................................................................................................................................67 Figure 4-26: Volatile solids concentration in the mesophilic anaerobic digester influent and effluent. .....................................................................................................................................68 Figure 4-27: Specific capillary suction time of the thermophilic anaerobic digester effluent. ......72 Figure 4-28: Specific capillary suction time of the mesophilic anaerobic digester effluent. ........73 Figure 4-29: Specific fecal coliform concentration in the anaerobic digester effluent. ................74 xv Figure A-1: Total ammonium nitrogen colorimetric test calibration curve. ..................................86 Figure A-2: Total phenolics colorimetric test calibration curve. ..................................................86 Figure A-3: Chemical oxygen demand colorimetric test calibration curve. .................................87 Figure B-1: Run 1 salt at 1,000 times magnification. …………………………………………….... 86 Figure B-2: Run 2 salt at 1,000 times magnification. …………………………………………….... 86 Figure B-3: Run 3 salt at 1,000 times magnification. …………………………………………….... 87 Figure B-4: Run 4 salt at 1,000 times magnification. …………………………………………….... 87 Figure B-5: Run 5 salt at 1,000 times magnification. …………………………………………….... 88 Figure B-6: Run 6 salt at 1,000 times magnification. …………………………………………….... 88 Figure B-7: Run 7 salt at 1,000 times magnification. …………………………………………….... 89 Figure C-1: Run 1 EDS map and element fractions. …………………………………...……….… 90 Figure C-2: Run 2 EDS map and element fractions. …………………………………...……….… 90 Figure C-3: Run 3 EDS map and element fractions. …………………………………...……….… 90 Figure C-4: Run 4 EDS map and element fractions. …………………………………...……….… 91 Figure C-5: Run 5 EDS map and element fractions. …………………………………...……….… 91 Figure C-6: Run 6 EDS map and element fractions. …………………………………...……….… 91 Figure C-7: Run 7 EDS map and element fractions. …………………………………...……….… 92 xvi List of Abbreviations AD Anaerobic digestion ANOVA Analysis of variance BTG Bioreactor technology group CHG Catalytic hydrothermal gasification COD Chemical oxygen demand GAE Gallic acid equivalence GC Gas chromatograph GC/FID Gas chromatograph / Flame Ionization Detector GC/TCD Gas Chromatograph / Thermal Conductivity Detector HTL Hydrothermal liquefaction HTLaq Hydrothermal liquefaction aqueous waste product MPN Most probable number MS Mixed sludge OLR Organic loading rate PS Primary sludge SS Secondary sludge TAN Total ammonium nitrogen tCOD Total chemical oxygen demand TS Total solids TSS Total suspended solids VFA Volatile fatty acid VS Volatile solids VSS Volatile suspended solids WWTP Wastewater treatment plant xvii Acknowledgements I want to thank my Dr. Cigdem Eskicioglu for supervising, supporting, and informing my research and academic growth. I am also so appreciative of my committee members, Dr. Nicolas Peleato, and Dr. Alyse Kiesser. I would also like to thank Negin Azarmina, Alpar Basar and Huan Liu for training me on different BTG laboratory procedures. I am so thankful for BTG staff Jacky Takeuchi and Dr. Parmila Devi for assisting with the setup of my apparatus. And lastly, I would like to give a big thanks to my fellow lab mate Abigail Stokes, for their support in and out of the lab. xviii Dedication To my family who has supported all my academic endeavours, my loving partner who took this adventure across the country with me and all the friends who have made these years ones I will never forget. Chapter 1: Introduction The focus of municipal wastewater treatment plant (WWTP) research is moving towards the recovery of resources to increase the sustainability of communities. The solids removed from the liquid process stream during wastewater treatment are rich in inorganic nutrients which have value as fertilizers and organic compounds with energy generation potential. Energy from municipal WWTP solids can be recovered through the creation of biogas or other forms of biofuel. Currently biogas is the most common energy product produced from WWTP sludge, through anaerobic digestion (AD). AD is a biochemical process where anaerobic microorganisms breakdown organics in sludge, reducing the volume of sludge to be disposed of and releasing methane-rich biogas. There has been great success in producing biogas from AD applications but there is further opportunity to explore biofuel production, which can be achieved through combustion, thermochemical or biochemical methods. An emerging thermochemical method for converting biomass into biofuel is hydrothermal liquefaction (HTL). This could be a more suitable sludge processing method than AD at some WWTPs because the shorter solids retention time could decrease the size of the reactor and the flexibility for high moisture content could reduce dewatering requirements before HTL. 1.1 Project Objectives The goal of the Bioreactor Technology Group (BTG) is to work towards more sustainable WWTPs and energy generation through biochemical and thermochemical treatment. The graduate research project outlined in this thesis works towards this larger goal by: 1. Determining the optimal conditions for pretreatment of HTL aqueous waste product (HTLaq) derived from municipal sludge through ammonia stripping, 2. Identifying the components of HTLaq removed during this pretreatment, 2 3. Investigating nitrogen recovery through acid adsorption and feasibility of generating a valuable product, and 4. Determining the biodegradability of pretreated and untreated HTLaq through anaerobic co-digestion with municipal sludge and dewatering centrate. 1.2 Novelty of Research This work is the first analysis of ammonia stripping for resource recovery and downstream anaerobic digestion enhancement for HTLaq derived from municipal sludge. To the best of the author’s knowledge, this research is the first analysis of any kind of ammonia stripping on the substrate HTLaq. The research area of the effects of HTLaq on biological treatment processes and methods of removing inhibitory compounds is still developing and there are many knowledge gaps to investigate. The research in this thesis provides an extensive characterization of HTLaq before and after ammonia stripping pretreatment, optimizes ammonia stripping conditions of this new emerging substrate and provides long-term anaerobic biodegradability assessment of non-pretreated and pretreated HTLaq. This research also provides new information on the ammonia recovery from HTLaq with purity information on the resulting ammonium sulphate salt. 1.3 Thesis Sections After the introduction section (Chapter 1), this thesis has a review of literature relevant to the development of this project, and for understanding of the experimental results in Chapter 2. Chapter 3 is a description of the equipment and materials used during experimentation, the design of experiment and all analytical characterization methods required to perform the project. Next, the results of the experimental design are presented in Chapter 4 with a discussion on the relevance of these results to the project. The final written section of the thesis (Chapter 5) is a conclusion that summarizes how the results relate to the project objectives, key findings of the experiment and research to be done to further the project objectives. Following the written 3 sections is a bibliography of all sources referenced. This thesis ends with three appendices; example calibration curves for colorimetric tests, ammonia salt images from a scanning electron microscope (SEM) and results of energy dispersive spectroscopy (EDS). 4 Chapter 2: Literature Review The literature review focuses on the emerging technology of hydrothermal liquefaction (HTL) and its potential for use in municipal wastewater treatment plant solids processing applications. The first section of the literature review provides general information of HTL, the second section provides potential disposal options for the aqueous product (HTLaq) created during the process and the third section analyzes potential treatment methods for HTLaq prior to disposal. The final section of Chapter 2 is a review of findings from the literature review. 2.1 Hydrothermal Liquefaction of Organic Biomass HTL involves the pyrolysis of organic feedstock by applying temperatures between 200 and 374ºC and pressures of 15 to 220 bars for a detention time of 1 to 60 minutes (Basar, et al., 2021). The feedstock for HTL can be a variety of types of biomasses, including WWTP sludge, cyanobacteria, algae, and agricultural waste. The extreme conditions in HTL convert the biomass into biocrude, a fuel product that can be used to replace crude oil following pretreatment. Biocrude is rich in hydrocarbons but also can be used as a substitute for gasoline and diesel which are produced from traditional fossil fuels. Biocrude is considered a renewable energy source, and it is predicted that gasoline produced from biocrude will have comparable Energy Return on Investments (EROI) and comparable greenhouse gas (GHG) emissions to the cellulosic ethanol, a current green substitute to gasoline (Liu, et al., 2013). It is also predicted that the diesel substitute produced from biocrude will have a higher EROI and comparable GHG emissions when compared to that of soybean biodiesel. The lifecycle analysis of GHG emissions is lower for biocrude based petroleum products because the biomass feedstock captures carbon. This carbon would be released naturally as the biomass decayed, therefore the burning of biocrude based fuels will have minimal impact on atmospheric GHG levels, in comparison to extracted petroleum products. Extracting and burning petroleum products that 5 would otherwise be trapped within the earth adds carbon into the atmosphere that would not otherwise be there increasing atmospheric GHG levels. The production of biocrude through HTL also produces two waste products: hydrochar and HTLaq. Hydrochar is a solid carbon-rich by-product. This product is created during the hydrothermal carbonization process that also produces biocrude. Having the same source, means optimizing the HTL system for maximum biocrude production would minimize the quantity of hydrochar produced (Basar, et al., 2021). HTLaq is the more toxic by-product with organic and inorganic compounds that can inhibit biological activity. Ketones were present in higher concentrations in HTLaq produced from municipal WWTP sludge than any other feedstocks investigated by Basar et al. (2021). Carboxylic acids were the largest organic constituent in HTLaq from all feedstocks except from cyanobacteria which contains the highest proportion of amides. The study investigated the pathways in the HTL reaction process and found that carboxylic acids are produced from carbohydrates in the feedstock and amides are created from lipids and proteins. This demonstrates the significant effect that feedstock type has on the composition of HTLaq and all other HTL products. The main organic constituents in HTLaq derived from municipal sludge are carboxylic acids, ketones, and amides. The main inorganic contaminants of concern in HTLaq are ammonia and phosphorus. Ammonia concentration is increased during the HTL process because the high temperature converts organic nitrogen in the biomass to ammonia (Zhuang, et al., 2017). The severity of pyrolysis conditions, including high temperature, high pressure and long detention times lead to increased ammonia in the resulting HTLaq. 2.2 Hydrothermal Liquefaction Aqueous Product Disposal Pathways Since the aqueous stream has the highest levels of nutrients and organics of the HTL products, it is the hardest stream to find a safe and sustainable disposal pathway for, which has been the bottleneck for scaling up. To allow for easy disposal, the pretreatment of the HTLaq 6 may be required. One disposal pathway being investigated for HTLaq is to send it to an anaerobic digester for valorisation. Pretreatment of HTLaq may be required to reduce ammonia, phenols and other trace organics inhibitory to AD if the HTLaq is fed in large quantities. For this reason, studies have been performed on co-digesting HTLaq with sludge. Zhou et al. (2015) found that mesophilic anaerobic digestors would fail if the volume percent of HTLaq in the feed exceeded 13.3%. 2.2.1 Recycling in HTL One pathway for HTLaq is to recycle it back into the HTL reactor. Shah et al. (2020) recycled HTLaq through the HTL reactor eight times and found that energy recovery from biocrude increased by 50% but there was also a doubling of the nitrogen concentration in the biocrude. This increased nitrogen would require additional energy and cost in the distillation process to make a usable fuel product. Further investigation is required into whether HTLaq recycling can be achieved without increasing the biochar’s nitrogen concentration before this can be considered as a cost-effective option. There is still a requirement for the disposal of the HTLaq after recirculating eight times. The volume of HTL required for disposal would be reduced by this practice, allowing for greater dilution. The recirculation also had effects on the HTLaq properties. Most inorganics were found to be transferred from the HTLaq to hydrochar through continued recycling, however the concentration of total nitrogen, potassium and sodium in the aqueous phase increased (Shah, et al., 2020). The effect of the changed aqueous phase properties and volume on the potential for disposal to either the WWTP headworks or to anaerobic digesters would need to be further investigated. 2.2.2 Return to Headworks The first option considered for final disposal of HTLaq after the HTL process, is sending it to the headworks of the WWTP. Having the HTLaq sent to the front of the treatment train could 7 have adverse effects on all the downstream treatment processes including biological treatment, filtration, and disinfection. Basar et al. (2021) suggests that given the dilution provided by the plant flowrate, the high carboxylic acid concentration in the HTLaq would not cause an issue in biological treatment. However, Marrone (2016) indicated that the phenolic compounds would be inhibitory to biological treatment without pretreatment of the HTLaq. Another issue in HTLaq being returned to the WWTP headworks is its inorganic constituents, namely nitrogen. If the biological treatment system is designed to decrease nutrients from the level expected in the influent to just below the discharge limits, then adding a high source of nitrogen could lead to regulatory exceedances and nutrients entering the receiving body at an unsustainable rate. The effect of HTLaq on different treatment processes needs additional study. It is important to consider that not only are the levels of inhibitory compounds in the HTLaq dependent on the feedstock, liquification temperature, pressure, and detention time, but also the effect on the WWTP would be dependent on the treatment units utilized, the strength of the existing influent, and the nutrient removal capacity. In most systems, it is likely that pretreatment of the aqueous phase would be required before sending it to the headworks. To send the HTLaq to the headworks of a WWTP with strict nutrient removal requirements it may be advisable to use a microbial fuel cell or struvite precipitation to remove ammonia and phosphorus. These pretreatment methods, provide additional resource recovery by either producing electricity or fertilizer. 2.2.3 Sending to Anaerobic Digesters Another disposal pathway for HTLaq would be to send it to the anaerobic digesters for valorization. This is a practical approach for upcoming pilot projects because many utilities experimenting with new energy recovery methods already have AD capabilities onsite to reduce sludge volume and recover energy through methane. However, requiring valorization in an 8 anerobic digester decreases the feasibility of installing an HTL system at a WWTP that does not already have energy recovery. A benefit of HTL is that it requires a smaller reactor volume than AD and could be an option for WWTPs with a limited footprint, but if a municipality must install a digester for disposal of HTLaq, they may not be able to utilize HTL. There are also benefits of disposal through AD because HTLaq is high in volatile fatty acids (VFAs) which are important for the production of methane in digesters. HTLaq contains compounds that could inhibit AD, including ammonia, phenols, and other trace organics. For this reason, studies have been performed on diluting HTLaq with sludge. Azarmina (2023) found that thermophilic semi-continuous flow digesters could withstand HTLaq contributions to the feedstock up to 4.89% of total chemical oxygen demand (COD). However, at 4.89% COD contribution from HTLaq, the digester experienced 42 days of complete inhibition before beginning to recover. Thermophilic digesters fed 1.21% and 2.74% COD contribution ran without inhibition over the entire 180-day experiment. Zhou et al. (2015) ran batch anaerobic tests to determine the biodegradability of HTLaq derived from swine manure. The study observed inhibition if the volume percent of HTLaq exceeded 13.3%, using HTLaq derived from swine manure. The anaerobic tests diluted HTLaq with supernatant of inoculum sludge which at 13.3% HTLaq by volume would equate to 75.4% of the total COD (tCOD) contribution. The digester was run at 37ºC, falling under the category of mesophilic digestion. These two studies suggest that mesophilic digesters are more tolerant of higher doses of HTLaq, though it is hard to equate anaerobic batch tests to continuous flow operation of digesters. The study by Zhou et al. (2015) also added powdered activated carbon to all tests in attempt to adsorb inhibitory compounds. If VFAs are high in the HTLaq and energy recovery is a priority, then AD could be the ideal volatilization method. 9 2.2.4 Sending to other Anaerobic Reactors Upflow anaerobic sludge blanket reactors are a biological treatment method for soluble substrates, unlike AD which handles a slurry medium. This could allow HTLaq to be processed directly rather than a co-digester with an AD suitable substrate. Upflow anaerobic sludge blanket reactors are high-rate bioreactor systems that utilizes an upward flow pattern to suspend microorganisms that substrate must pass through during the treatment process. A study by Chen, et al. (2020) investigated the effect of HTLaq generated from cornstalk that had a COD of 10 g/L on an upflow anaerobic sludge blanket reactor. This system could handle 61.6% and 45% COD from HTLaq under mesophilic and thermophilic conditions, respectively (Chen, et al., 2020). 2.3 Hydrothermal Liquefaction Aqueous Product Pretreatment Options There are various studies on methods for pretreatment and disposing of HTLaq to allow for larger scale testing of energy recovery through the HTL process. However, HTLaq is a complex matrix of organic and inorganic compounds that differs depending on the feedstock and process conditions. This makes it difficult to generalize which disposal pathway will work best for every WWTP and which pretreatment methods will be required. 2.3.1 Carbon Adsorption Pretreatment Methods Adsorption can be used as a method for removing metals and organic compounds from liquids. Adsorption with a carbon-based media is when toxins come out of solution and attach themselves to the surface of the solid media the fluid is being passed through. The large surface area can also act as a site for reactions that further remove toxins. Two common adsorbents are granular activated carbon and powdered activated carbon, which were both found to spontaneously adsorb copper and phenolic compounds from HTLaq (Adedeji & Jahan, 2023). The same study found that boron and COD could also be adsorbed but external energy would have to added to the system. 10 Metals and phenolic compounds can be removed from HTLaq through adsorption by hydrochar, the solid waste product of HTLaq (Marx, et al., 2021). Marx et al. (2021) investigated using hydrochar to improve the quality of HTLaq produced from synthesized organic waste created based on the South Africa State of Waste report (Department of Environmental Affairs, 2018). The study focused on removing alkali and alkaline earth metals and phenolic compounds, as they are challenging to safely dispose. Removal of 91-94% of measured alkali and alkaline earth metals, 100% of the quaiacol, phenol and resorcinol compounds and 61% removal of vanillyl alcohol was achieved. Phenolic compounds form stronger hydrogen bonds with the hydrochar and were preferentially adsorbed, requiring a higher quantity of hydrochar for the removal of alkali and alkaline earth metals (Marx, et al., 2021). Some benefits of carbon adsorption are that it removes a variety of contaminants, and it may utilize the waste product hydrochar as an adsorbent. 2.3.2 Struvite Precipitation Methods Inorganic nutrients such as ammonia and phosphorus can be chemically precipitated out of solution. Struvite precipitation is a method used for nutrient recovery from the liquid treatment train of WWTPs. Ovsyannikova et al. (2020) experimented with the effectiveness of struvite precipitation at recovery of nutrients from HTLaq produced from municipal sludge and microalgae (Spirulina). The recovery rates for HTLaq derived from municipal sludge were 99% and 79.4% for phosphorus and ammonia, respectively. The method was less successful for Spirulina derived HTLaq, with 66.5% phosphorus and 19.4% ammonia recovered. Ovsyannikova et al. (2020) indicated that simpler methods for isolating the phosphorous would not create a usable fertilizer due to the concentration of heavy metals in the HTLaq. 2.3.3 Catalytic Hydrothermal Gasification (CHG) Methods HTLaq can also be further processed using catalytic hydrothermal gasification (CHG) to decrease inhibitory compounds. CHG is another thermochemical process that produces biogas 11 (containing methane) rather than the biocrude that HTL creates. When the two processes are used in series, they were shown to reduce COD in all HTLaq feedstocks to CHG (HTLaq produced from primary, secondary and post digested sludge) by greater than 99.9% and decreases the total solids (TS) by between 94 and 99% (Marrone, 2016). Prior to the HTLaq entering the CHG process, it was pretreated using an ion exchange column which reduced sulphates. Sulphates are a poison for the ruthenium-based catalyst used in CHG and decreasing the level of sulphates in the CHG feed reduces the frequency of catalyst replacement. Benefits to this CHG treatment method is that further energy is recovered, and solids removed. MicroBio Engineering (2019) indicated that sending the HTLaq produced in their study back to the WWTP’s headworks would introduce phenolic and aromatic compounds which the plant was not designed to handle and implied that the CHG process would remove these compounds as its waste product was safe to discharge to headworks. The levels of phenolic compounds were not indicated in the report for HTLaq or the CHG process effluent. The ammonia concentration in the HTLaq increased after the CHG process which could also pose a problem for disposal to headworks, but an opportunity to recover it as a fertilizer. CHG could be a valuable HTL process addition as it provides additional resource recovery and solids removal. 2.3.4 Biological Pretreatment Methods One method for pretreatment of HTLaq is through the use of microbial fuel cells. In these systems electricity is generated by electrons being transferred from one tank (cathode), where compounds are being reduced by microorganisms, to another tank (anode) of oxidized compounds (Shanmugam, et al., 2017). If the group of microorganisms chosen is fueled by the inhibitory organics or nutrients present in HTLaq then it can improve the quality of the aqueous through the oxidation reactions. 12 Kumar (2021) studied nutrient removal from HTLaq using four different stains of oleaginous microalgae and co-cultivation with yeast. Chlorella minutissima with co-cultivation with yeast was the microalgae strain that had the highest removal of nitrogen and phosphorus at 91% and 85% respectively (Kumar, et al., 2021). The HTL experiments in this study were run under isothermal conditions with the catalyst TiO2. Biocrude was extracted using organic solvent-assisted method and Soxhlet method. The Soxhlet extraction method produced the highest yield of 20.7% on a mass basis. Nitrogen was the limiting nutrient in the microalgae growth so Chlorella minutissima may be effective in treating HTLaq even at high levels of ammonia. Though nutrients were adsorbed by the microalgae, the biomass growth did not exceed that produced in the control medium (Bold’s Basal Medium). Another successful microorganism was Tetraselmis sp. however it had to be diluted to a level that 50% of the nutrients were coming from HTLaq (Das, et al., 2020). Another study utilizing Desmodesmus sp. found that the biomass growth was inhibited by HTLaq (Garcia Alba, et al., 2013). Lui et. al. (2024) treated HTLaq of sewage sludge using three different bacterial strains and four fungal strains with up to 44% increase in removal efficiency of COD during biological treatment, when the HTLaq was pretreated using ammonia stripping. The ammonia stripping also increased removal efficiencies of VFAs, phenolics and N-heterocyclic compounds in some biological treatment setups. All ammonia stripping was run overnight at 50ºC and a pH of 12 with the only sample characterization data collected for HTLaq before and after stripping being COD, total nitrogen, and total ammonium nitrogen (TAN). Literature focusing on ammonia stripping as treatment for HTLaq are investigated in the next subsection (2.3.5). 2.3.5 Ammonia Stripping Methods Ammonia gas is a weak base meaning that it will accept hydrogen ions if present, for example in acidic environments. With an extra hydrogen ion, this compound becomes ammonium, that dissolves more readily into water. Increasing the pH or temperature of the 13 solution shifts the equilibrium and increases the tendency of ammonium (ionized) to convert to ammonia (gaseous) and leave the liquid phase. Ammonia stripping is a treatment process that volatizes ammonium to remove it from a liquid stream. Ammonia stripping can be done through chemical stripping by adjusting the pH, thermal stripping by increasing the temperature or a combination by increasing both parameters. Thermal stripping can be achieved by directly heating the liquid being treated or having it come in contact with hot air (Tao & Ukwuani, 2015). A study by Adghim et al. (2022) found that 78% ammonia removal of hydrolysed poultry manure could be achieved over 6 hours of stripping at a pH of 10, temperature of 55⁰C and air flow rate of 300 L air/L feedstock/hour. This study used a water bath to heat the samples and Ca(OH)2 for increasing pH. Adghim et al. (2022) also conducted biochemical methane potential tests which showed that ammonia stripping relieved inhibition to anaerobic biological treatments and increased methane potential by 200% compared to nontreated post-hydrolysis poultry manure. Ammonia gas generated during stripping should be captured as it can be harmful to the atmosphere by contributing to smog. Its release can also negatively impact surrounding ecosystems because if ammonia gas encounters acidic pollutants, such as nitric or sulphuric acids, it will form ammonia salts which can sublimate and fall to the ground or water body below (Krupa, 2003). Instead, ammonia can be captured by running the off gas through an adsorption column with an acid solution that will produce a neutral ammonium salt. A study by Sigurnjak et al. (2019) found that ammonium nitrate (NH4NO3) and ammonium sulphate ((NH4)2SO4) salts, produced from ammonia stripping of animal manure, produced similar crop yields and no additional risk of nitrate leaching to currently used synthetic nitrogen fertilizers. 14 2.4 Literature Review Summary HTL is a budding area of research with lots of interest in its effectiveness for converting different high-moisture organic wastes into fuel and how to safely dispose of all products generated in the process. Many studies have been generated on the impact of HTLaq on different disposal pathways and methods to pretreat HTLaq to lessen that impact. AD was chosen as the process unit to dispose of HTLaq created from municipal WWTP sludge for this work due to the potential to recover more energy from the remaining organics and thermophilic digesters being available on the study site of the collaborating municipality. However, due to existing literature showing that thermophilic digesters can only withstand 4.89% COD from HTLaq, analysis of a HTLaq pretreatment method would be required (Azarmina, 2023). Ammonia stripping was chosen as the pretreatment method as it had not yet been tested on HTLaq, but is well cited for removing ammonia, which can be inhibitory to AD, from other substrates. 15 Chapter 3: Materials and Methods The methods used to conduct the research of HTL, the pretreatment of its aqueous phase, and downstream anaerobic co-digestion with municipal sludge, and the equipment and materials required to perform them are outlined in this chapter. 3.1 Equipment The research was carried out in the University of British Columbia Okanagan Campus’s BTG laboratory. The equipment, tabulated in Table 3-1, was required for the completion of the methods detailed in the remaining sections of the material and methods chapter. Table 3-1: Equipment inventory for all experiments performed in the BTG laboratory. Click or tap here to enter text. Equipment Type Manufacturer Analytical balance XA105DU Mettler Toledo Centrifuge (with large rotor) Sorvall Legend XT Thermo Fisher Scientific Centrifuge (with small rotor) Legend micro-21 Thermo Scientific Parr reactor 4570 HT/HP Parr Instrument Company Gravity oven Isotemp 625G Fisher Scientific Muffle furnace W-13 Paragon Industries Hot plate CIMAREC Thermo Scientific Flow monitor FM-112-08-SA-VN Omega Dual channel pH/ion meter XL25 Thermo Fisher Scientific Spectrophotometer GENESYSTM 10S UV-VIS Thermo Fisher Corporation Microplate reader SynergyTM HT Multi-Detection Bio Tek Instruments Gas chromatograph (GC) 7890A GC with Flame Ionization Detector (GC/FID) Agilent Technologies Benchtop homogenizer mixer PT 10-35 GT KinematicaTM PolytronTM Shaker incubator Innova ® 44 Rotary Shaking Incubator New Brunswick Scientific Gas flowmeter M-0.5SCCM Alicat Gas chromatograph 7820A GC with Thermal Conductivity Detector (GC/TCD) Agilent Technologies Gas chromatograph 8860 GC 5977B-Mass Spectrometer Agilent Technologies Microwave Digestion System Ethos EZ Milestone Capillary suction timer MODEL 440 Fann 16 3.2 HTL Operation to Generate Aqueous Phase Before the pretreatment of HTLaq experiment, the HTL reactor had to be run to generate sufficient volume of HTL aqueous to be used as ammonia stripping feedstock. The Parr HTL reactor (Figure 3-1) was run three times using a 50% thickened screened primary sludge (PS) and 50% thickened waste secondary sludge (SS) by weight, sampled from Annacis Island WWTP on August 4th, 2022. The two streams were collected separately because the plant mixed sludge (MS) stream varies in PS and SS contribution and HTLaq characterization is affected by these fluctuations. The ratio of PS and SS was chosen based on the average annual production volume from each stream in 2019 to be representative of average conditions. For HTLaq used for semi-continuous flow co-digestion, MS was collected from the plant to account for variation in characteristics over the run time. Figure 3-1: Bench-scale Parr reactor used for HTL treatment of municipal sludge. 17 Annacis Island WWTP run by Metro Vancouver uses a grit chamber and primary sedimentation in primary treatment and a trickling filter, solids contact tank and secondary sedimentation in secondary treatment. The plant also performs tertiary treatment in the form of chlorine disinfection. The location of PS, SS and MS collection in the treatment process can be identified in the process flow diagram below (Figure 3-2). Figure 3-2: Process flow diagram for Annacis Island WWTP solids processing (Coulman, 2022). At the BTG lab, once the sludge streams were mixed, the resulting 8.0 L was centrifuged first at 3,600 xg for 10 minutes using 750 mL bottles, then the solids portion was again centrifuged at 20,000 ×g for 10 minutes in 50 mL tubes. The centrate was saved for characterization. The dewatered cake with a TS content of 17.2 ± 0.2% by weight was separated into three 500 g batches for the HTL reactor and run at 350ºC for 15 minutes each. The reactor temperature of 350ºC was chosen based on continuous flow HTL reactor work done by Pacific Northwest National Laboratory and The U.S. Department of Energy (Snowden-Swan, et al., 2021). The ideal 15-minute run time was selected based on previous work completed by the BTG, using Annacis Island WWTP sludge. This chosen time accounts for the warmup time of the Parr batch reactor (Liu, et al., 2022). After the HTL reactor is cooled down, the reactor 18 was opened, and effluent was then filtered using a 0.45 µm filter to separate the HTLaq from the solid products (hydrochar and biocrude). All HTLaq collected from the three runs was combined to make one uniform feedstock for all the ammonia stripping runs. Hydrochar and biocrude were outside the scope of this thesis. 3.3 Ammonia Stripping and Recovery Method Chemical stripping was run for 4 hours to remove ammonia from a 100 mL HTLaq sample whose derivation is outlined in Section 3.2. The four-hour run time was selected after completing nine preliminary tests when the ammonia stripping apparatus was run between two and seven hours. In these initial tests run time, pH levels and apparatus design were adjusted to create the experimental design. Chemical stripping was performed using a round bottom flask submerged in a water bath placed on a hot plate with magnetic mixing (Figure 3-3). Figure 3-3: Ammonia stripping and acid adsorption apparatus. 19 In one neck of the flask, a plug holds a hollow metal rod and a temperature probe connected to the hot plate to monitor the temperature of the sample. The hollow metal rod end was a corkscrew shape with holes to allow for air to bubble into the aqueous sample. The air then flowed into the condenser attached to the neck at the top of the flask. The condenser reduced the moisture from the system air prior to entering the acid column. In another neck of the flask, a 13 mL pipette tip was placed into the HTLaq so that sampling could be done with a syringe during stripping without air leaving the system. The remaining round bottom flask neck was sealed with a plug to ensure all air leaving the system flows through the condenser and into the acid scrubber. The temperatures, pH levels, and air flow rates were tested to determine the ideal ammonia stripping conditions and are displayed in Table 3-2. The pH setpoint was achieved by adding 10 N NaOH to the HTLaq in the round bottom flask during sample addition. The lowest pH setpoint was chosen as 9.3 as this is pKa of ammonium. The pKa represents the pH at which 50% of ammonium would dissociate into H+ and free ammonia nitrogen (the conjugate base and the form able to be stripped from the sample) if the acid-base relationship was the only factor in ammonium dissociation. The amount of free ammonia nitrogen however, is also affected by temperature so the equation in 3.5.1 Total Ammonium Nitrogen is needed to determine the maximum possible percentage of ammonia that can be removed from the system for every pH and temperature combination for the reactor. The air flow rates to be tested were chosen based on the range available on the flow meter and the setpoint on the flowmeter was monitored throughout the four-hour run time. The temperature of the HTLaq sample was increased and maintained using a hot plate with a temperature probe attachment. The temperature range of 65 – 85ºC was chosen because the HTLaq would come out of a full-scale continuous flow HTL reactor at 70ºC (Marrone, 2016). Using this temperature range provides 20 information concerning if the HTLaq should be given time to cool, prior to entering a full-scale ammonia stripping facility or if it may need to be further heated. Table 3-2: Ammonia stripping experimental design parameters. *Optimum temperature and pH were determine based on the highest ammonia removal of earlier runs. Samples of 3 mL were taken hourly from the HTLaq in the round bottom flask during the four-hour stripping run so that TAN, phenols, VFAs, tCOD and pH could be measured throughout the run time. These parameters, as well as total alkalinity, were also measured in the feedstock, reactor effluent and at time zero. Time zero was when the sample had reached the setpoint temperature. At this time, 10 mL was removed from the apparatus to allow for initial alkalinity measurement as well as additional replicates for VFA analysis. Immediately after the sample was removed for characterization, the air was switched on and the rate was set based on the run setpoint. Total suspended solids (TSS) and volatile suspended solids (VSS) were only measured in the HTLaq prior to and at the end of the stripping run due to the large volume required to measure these parameters. For ammonia recovery, an acid scrubber was prepared using a 500 mL glass graduated cylinder and adding 310 mL of 0.1 N sulfuric acid (H2SO4). This was used to precipitate the ammonia stripped from the HTLaq. An air stone bubble diffuser was used to increase surface area for adsorption of ammonia from the air stream into the sulfuric acid. After the run time was Run no. Temperature (oC) pH Air flow (L/min) 1 65 10.3 0.5 2 75 10.3 0.5 3 85 10.3 0.5 4 Optimum T* 9.8 0.5 5 Optimum T 9.3 0.5 6 Optimum T Optimum pH* 0.3 7 Optimum T Optimum pH 0.1 21 complete, the contents of the acid scrubber were transferred to a shallow dish and left to dry at room temperature (22ºC), as shown in Figure 3-4. The salt generated from each run was weighed and tested for purity. Figure 3-4: An image of the ammonium sulphate salt produced during each ammonia stripping run. 3.4 Anaerobic Co-digestion Method The six anaerobic digesters used in the co-digestion experiment were fed with different mixtures of mixed sludge (MS), centrate and HTLaq (Figure 3-5). As mentioned in Section 3.2, the MS was obtained from the Annacis Island WWTP and collected after the primary and secondary sludge streams meet. The volume of PS and SS in this stream can vary for each batch (delivered monthly) but onsite digesters would need to be able to handle this fluctuation. As explained earlier in Section 3.2, MS was used in the HTL runs needed to produce the HTLaq for the digester feedstock and the centrate collected during dewatering for HTL feed was saved for digester feedstock. 22 Figure 3-5: An image of mesophilic anaerobic digesters setup in 1 L erlenmeyer flasks, tedlar bags for biogas capture, and tubing for feeding and effluent removal. Non-pretreated and pretreated HTLaq were both used in different feedstocks to compare biodegradability. In the non-pretreated HTLaq no additional measures were taken after filtration at 0.45 µm to separate the products of HTL. The pretreated HTLaq had an additional step of ammonia stripping (described in Section 3.3) performed prior to co-digestion. For co-digestion, the run conditions for the HTLaq pretreatment are listed in Table 3-3 and were chosen in partnership with Metro Vancouver based on the results of the ammonia stripping experiment (Runs 1-7, Table 3-2) with consideration of the operational cost and complexity of chemical additions compared to heating at a facility that produces biogas. Table 3-3: Ammonia stripping parameters for HTL aqueous pretreatment ahead of anerobic co-digestion. Run 4 Operational Parameters Sample Volume 110 mL Temperature 85°C pH 9.8 Time 3 hours Air Flow 0.5 L/min Stir Speed 500 rpm 23 The anaerobic co-digestion experiment was performed to compare the biodegradability of pretreated and non-pretreated HTLaq under thermophilic and mesophilic conditions. Laboratory scale digesters were operated in 1 L Erlenmeyer flasks, with a port in the neck (Figure 3-5). A rubber stopper with two glass tubes inserted through it was placed on the mouth of each flask and sealed with caulking. One of the glass tubes was long enough to extend nearly to the bottom of the flask and the other was cut short to remain in the air space. Tubing was fastened to both of the glass tubes and the flask port to provide access to digester for biogas sampling, effluent removal and feeding while maintaining anaerobic conditions. More information on digester operation is provided in table Table 3-4. Table 3-4: Bench-scale anaerobic digester conditions Total Volume 1 L Effective Volume 0.5 L Flow Rate 25 mL/day Solids Retention Time 20 days Mesophilic Temperature 35ºC Thermophilic Temperature 55ºC Six anaerobic co-digestion digesters were run to assess the biodegradability of HTLaq pretreated through ammonia stripping for a year. Both digester temperatures had a control digester only fed MS, a digester fed with non-pretreated HTLaq, centrate and MS, and a digester fed with pretreated HTLaq, centrate and MS. The digesters being fed HTLaq were started at a level known to be non-inhibitory and then were transitioned to a higher level of organic loading coming from HTLaq. Under thermophilic conditions, the contribution of HTLaq was chosen from experimentation with similar co-substrates by Azarmina, (2023) who found that digesters with 5% COD from HTLaq were able to stabilize after a period of inhibition while digesters with 13% COD from HTLaq did not recover. Mesophilic digesters were found to be 24 more resistant to HTLaq co-digestion, so this experiment started at 13% COD from HTLaq and increased to 22% COD from HTLaq. Figure 3-6 illustrates the conditions tested for all six digesters with the labels indicated the percentage of organic loading rate (OLR) coming from HTLaq in term of mass of COD. Figure 3-6: Anaerobic co-digestion experimental design. The testing setpoints of HTLaq contribution were originally designated based on percentage of the daily solids stream being diverted away from the anaerobic digesters and into a HTL reactor, as indicated in Table 3-5. For example, in the potential case where the Annacis Island WWTP installs a HTL reactor that can take 25% of their daily MS production, for every 25 mL of influent fed to the anaerobic digesters, 18.9 mL would be the non-diverted MS, 1 mL would be centrate from dewatering the diverted MS, and 5.1 mL would be HTLaq. 25 Table 3-5: Average HTLaq contribution to organic loading rate for each digester condition compared to the flowrate contributions of each co-substance. Digester Tag Distribution of daily MS between HTL and AD (%) Contribution of co-substrates to total flowrate to AD (25 mL/d) OLR (kg COD/m3d) HTLaq contribution to digester OLR HTL AD MS (mL/d) HTLaq (mL/d) Centrate (mL/d) 0%HTLaq Control 0 100 25.0 0.0 0.0 2.5 ± 0.1 0% 5%HTLaq 25 75 18.9 1.0 5.1 2.3 ± 0.1 5% 13%HTLaq 50 50 12.7 2.0 10.3 1.9 ± 0.1 13% 24%HTLaq 75 25 6.4 3.0 15.6 1.4 ± 0.1 24% 5%HTLaq Stripped 25 75 19.0 0.9 5.1 2.4 ± 0.1 5% 13%HTLaq Stripped 50 50 12.8 1.9 10.4 1.8 ± 0.1 13% 22%HTLaq Stripped 75 25 6.5 2.8 15.7 1.4 ± 0.1 22% Since each shipment of MS and aqueous waste from each batch of HTL have a different COD, the HTLaq contribution to OLR could not be calculated straight from the volumes of each co-substance and instead the digester tags designated in Table 3-5 are averages for the run time of that condition. While running the digesters, the flowrates of co-substrates designated in this table dictated digester feeding. Effluent was removed from the digesters daily using a syringe to maintain the anerobic condition and this effluent was used to carry out the sampling outlined in Table 3-6. Digesters were also fed daily based on their co-digestion conditions at which time biogas production was measured from the tedlar bag attached to the digester (Section 3.5.10). When this daily feeding/removal was complete, the digesters were stored in incubator shakers set to 95 rpm. 26 Table 3-6: Anaerobic co-digestion experiment sampling schedule. Parameter Frequency Sample Location Biogas production volume Daily Digester headspace Biogas composition (CH4, CO2, O2, and N2) Weekly Digester headspace Total ammonium nitrogen (TAN) Weekly Digester influent/effluent Total phenolic compounds Weekly Digester influent/effluent Volatile fatty acids (VFAs) Weekly Digester influent/effluent Total chemical oxygen demand (tCOD) Weekly Digester influent/effluent pH Weekly Digester influent/effluent Total alkalinity Weekly Digester influent/effluent Total/volatile solids Weekly Digester influent/effluent Capillary suction time Twice Digester effluent Gas chromatography – mass spectroscopy Once Digester effluent and HTLaq Fecal coliforms Once Digester effluent Heavy metals Twice Digester effluent HTLaq contributions were increased in two steps to not shock the system and digesters were considered to be at steady-state after three solids retention times (60 days). All weekly sampled parameters had 10 data points collected after steady-state was reached to calculate the average values. 3.5 Analytical Characterization Methods Many characterization methods were required to determine the level of treatment provided by ammonia stripping and its effect on the biodegradability of HTLaq. 3.5.1 Total Ammonium Nitrogen (TAN) A colorimetric method used for determining the TAN concentration of different samples (American Public Health Association, 2005). Preparation of the sample differed based on the type of sample. Samples high in solids, such as sludge and digester effluent, were first centrifuged at 12,000 rpm for 15 minutes. Samples with low solids, such as HTLaq, were not required to be centrifuged. Samples were then diluted using type I water in volumetric flasks so 27 the TAN concentration would be within the range of the calibration standards. The calibration standards used were between 0.1 and 7.5 mg/L TAN (Figure 3-7). Figure 3-7: Image taken during TAN testing, showing the gradient of colour change across the calibration standards prior to being placed in the spectrophotometer. In a 10 mL test tube, 0.61 mL of diluted sample or calibration standard was mixed with 2.0 mL of complexing solution, 0.86 mL of alkaline phenol, 0.01 mL of 5% (wt/wt) sodium hypochlorite and 0.36 mL of 500 mg/L nitroferricyanide. The test tubes were then placed in a dark place for one hour so that the blue-coloured compound indophenol has time to be produced depending on the amount of ammonia present in the sample. After an hour, the absorbance in each test tube was measured using the GENESYSTM 10S UV-VIS spectrophotometer set to 640 nm to detect the blue of indophenol. The absorbance of the samples was compared to that of the calibration standard solutions using a calibration curve (an example is shown in Appendix A, Figure A-1). TAN is comprised of free ammonia nitrogen (NH3) and ammonium nitrogen (NH4) with free ammonia nitrogen being the more inhibitory to AD due to its reactivity with methanogenic bacteria (Melbinger, 1971). Free ammonia nitrogen (mg NH3-N/L) can be calculated from TAN (mg/L) using Equation 3-1 if the temperature (K) and pH of the sample are known, with higher temperatures and pHs leading to an increase in ammonium converting to ammonia. 𝐹𝑟𝑒𝑒 𝐴𝑚𝑚𝑜𝑛𝑖𝑎 Nitrogen (𝑚𝑔 𝑁𝐻ଷ − 𝑁/𝐿) = TAN ∗ ቆ1 + ଵ଴ష೛ಹଵ଴షቀబ.బవబభఴ శ మళమవ.వమ೅ ቁቇିଵ [3-1] 28 3.5.2 Total Phenolics Total phenolics concentration was measured using a colorimetric method by the SynergyTM HT microplate multi-detection reader (Singleton, 1965). Samples were centrifuged (12,000 xg, 15 minutes) if necessary and then all samples were filtered using a 0.2 µm nylon filter. HTLaq samples were diluted ten times with type I water to remain in the range of the calibration standard solutions which ranged from 0-500 mg/L gallic acid equivalence (GAE). MS and digester effluent samples were found to be in a suitable range without dilution. In small tubes suitable for mixing, 0.1 mL of sample or standard were added, 0.5 mL of ten times diluted Folin-Ciocalteu reagent and 0.4 mL of 75 g/L sodium carbonate (Na2CO3). All work with Folin reagent was done under a fume hood. The solution was mixed and then transferred to a microplate with 0.2 mL was added to each well in the plate. The microplate was then inserted into the microplate reader with instructions to heat for five minutes at 40ºC and then shake for five seconds prior to reading the absorbance at 735 nm (Figure 3-8). Figure 3-8: Image of microplate of samples being placed into the for total phenolics testing. The Folin-Ciocalteu reagent contains phosphotungstic and phosphomolybdic acid which, when reduced in the presence of phenolic compounds, produced tungsten and molybdenum oxides respectively. These oxides have a dark blue colour that has a peak absorbance between 725 and 750 nm depending on which compound is reduced more. The absorbance of the samples was then compared to the absorbance of the standards using a calibration curve (an example of the calibration curve can be found in Appendix A, Figure A-2) to determine the concentration of total phenolic compounds (mg GAE/L). 29 3.5.3 Volatile Fatty Acids (VFA) VFA concentrations were tested using the Agilent 7890A GC/Flame Ionizer Detector and an internal standard of iso-butyric acid (Ackman, 1972). Iso-butyric acid is included in every sample in a known amount as an internal standard. The instrument is calibrated using the internal standard technique, and calibration standards are prepared using the same concentration of internal standard as the samples. Every batch of samples, one calibration check standard is also run that has 2,000 mg/L of acetic, propionic and butyric acids. First, samples were centrifuged (12,000 xg, 15 minutes) as necessary and then filtered with a 0.45 µm nylon filter. HTLaq samples were diluted ten times with type I water as they are higher in VFAs than other samples. Next, 0.5 mL of the prepared sample or calibration check standard and 0.5 mL of 20 g/L iso-butyric acid was added to a glass autosampler vial and then the vials were capped. The vials were transferred to the GC/FID autosampler and analysed. The Chemstation software calculated the concentration of each VFA using a calibration table with internal standard quantitation. 3.5.4 Total Chemical Oxygen Demand (tCOD) A colorimetric method was used to calculate tCOD with the GENESYSTM 10S UV-VIS spectrophotometer (American Public Health Association, 2005). The samples were first diluted with reverse osmosis (RO) treated water to be within the range of the calibration standard solutions. The calibration standards ranged from 0-700 mg/L COD. Then 2.5 mL of sample or standard, 3.5 mL of digestion solution and 1.5 mL of COD catalyst solution were added to a test tube with secure caps and mixed. The digestion solution contains mercury sulphate (HgSO4) and potassium chromate (K2Cr2O7) and the the COD catalyst solution is silver sulphate (Ag2SO4). Both solutions contain a high concentration of sulfuric acid which needs to be handled in a fume hood with appropriate personal protective equipment until the caps are closed. The test tubes were then placed in a heating block at 30 150ºC for three hours. The outside of the test tubes were then wiped with a Kimwipe and absorbance at 600 nm was tested in the UV spectrophotometer. The absorbances of the samples were compared to that of the calibration standards using a calibration curve to determine the sample concentrations (an example COD calibration curve can be found in Appendix A, Figure A-3). 3.5.5 pH The dual-channel AccumetTM pH meter XL25 was used to measure pH. A pH probe attachment is submerged in the sample. 3.5.6 Total Alkalinity A chemical titration of sample with 0.1 N sulfuric acid was used to determine total alkalinity (American Public Health Association, 2005). Using a 25 mL burette, 0.1 N sulfuric acid was slowly added into a beaker containing a recorded volume of the sample. A pH probe was placed into the beaker, which was gently swirled, so that acid could be added until a pH of 4.6 was reached. The volume of acid used was recorded using the starting and ending volume in the burette. The total alkalinity was calculated using the Equation 3-2 below, where V is volume of the acid (Va) or sample (Vs) in mL, and Na is the normality of the acid used (0.1 N). 𝑇𝑜𝑡𝑎𝑙 𝑎𝑙𝑘𝑎𝑙𝑖𝑛𝑖𝑡𝑦 (௠௚ େୟେ୓య௅) = ହ଴,଴଴଴ ௏ೌ ேೌ௏ೞ [3-2] 3.5.7 Total and Volatile Solids (TS and VS) The weight percentage of solids can be found by comparing the weight of dried solids to the total weight of the sample (American Public Health Association, 2005). The percentage of those solids that are volatile and non-volatile can then be determined by measuring the weight of solids remaining after ashing the sample. First, the mass of crucibles was recorded using an XA105DU analytical balance and then sample was added to each crucible. The weight of the crucible and sample together was 31 recorded before the crucible was placed in an oven at 105ºC for at least twelve hours to evaporate any moisture. The mass was then recorded again and labelled as ‘dry weight plus crucible’ before ashing any of the solids that are volatile in a muffle furnace at 550ºC for 30 minutes. After the crucible was cooled it was weighed a final time. TS and Volatile solids (VS) by percentage (weight/weight) can be calculated using Equation 3-3 and Equation 3-4, respectively. 𝑇𝑜𝑡𝑎𝑙 𝑠𝑜𝑙𝑖𝑑𝑠 ቀ% ௚௚ቁ = ୢ୰୷ ୵ୣ୧୥୦୲ ୮୪୳ୱ ୡ୰୳ୡ୧ୠ୪ୣ (୥) ି ୡ୰୳ୡ୧ୠ୪ୣ ୵ୣ୧୥୦୲ (୥)ୱୟ୫୮୪ୣ ୵ୣ୧୥୦୲ ୮୪୳ୱ ୡ୰୳ୡ୧ୠ୪ୣ (୥) ି ୡ୰୳ୡ୧ୠ୪ୣ ୵ୣ୧୥୦୲ (୥) ∗ 100 [3-3] 𝑉𝑜𝑙𝑎𝑡𝑖𝑙𝑒 𝑠𝑜𝑙𝑖𝑑𝑠 ቀ% ௚௚ቁ = ୢ୰୷ ୵ୣ୧୥୦୲ ୮୪୳ୱ ୡ୰୳ୡ୧ୠ୪ୣ (୥) ି ୠ୳୰୬୲ ୵ୣ୧୥୦୲ ୮୪୳ୱ ୡ୰୳ୡ୧ୠ୪ୣ (୥)ୱୟ୫୮୪ୣ ୵ୣ୧୥୦୲ ୮୪୳ୱ ୡ୰୳ୡ୧ୠ୪ୣ (୥) ି ୡ୰୳ୡ୧ୠ୪ୣ ୵ୣ୧୥୦୲ (୥) ∗ 100 [3-4] 3.5.8 Total and Volatile Suspended Solids (TSS and VSS) The percentage of total and volatile suspended solids can be found by drying and burning a filter that catches suspended solids from the sample. Glass fiber filters of 0.45 µm pore size were prepared by running RO water through them and then drying them in a muffle furnace at 550ºC for 30 minutes. These filters were then used to filter a measured volume of sample. The filters were then place in labelled pre-weighed aluminum dishes. To determine the TSS, the filters with the aluminum dishes were placed in an oven set at 110ºC for 12 hours. These were allowed to cool and the weight of the dish, with dried solid on the filter was measured and recorded. After this, the dish and filter were put into the muffle furnace at 550ºC for 30 minutes to ash the solid material. After allowing the dish and filter to cool, the weight of the dish and ashed filter was recorded. Equation 3-5 and Equation 3-6 below were used to determine the TSS and VSS respectively, by percentage (weight/volume). 𝑇𝑜𝑡𝑎𝑙 𝑠𝑢𝑠𝑝𝑒𝑛𝑑𝑒𝑑 𝑠𝑜𝑙𝑖𝑑𝑠 ቀ% ௚௠௅ቁ = (ୢ୰୷ ୵ୣ୧୥୦୲ ୭୤ ୤୧୪୲ୣ୰ ା ୢ୧ୱ୦) ି (ୢ୧ୱ୦ ା ୤୧୪୲ୣ୰ ୵ୣ୧୥୦୲)ୱୟ୫୮୪ୣ ୴୭୪୳୫ୣ∗ 100 [3-5] 𝑉𝑜𝑙𝑎𝑡𝑖𝑙𝑒 𝑠𝑢𝑠𝑝𝑒𝑛𝑑𝑒𝑑 𝑠𝑜𝑙𝑖𝑑𝑠 ቀ% ௚௠௅ቁ = (ୢ୰୷ ୵ୣ୧୥୦୲ ୭୤ ୤୧୪୲ୣ୰ ାୢ୧ୱ୦)ି (ୠ୳୰୬୲ ୵ୣ୧୥୦୲ ୭୤ ୤୧୪୲ୣ୰ ା ୢ୧ୱ୦)ୱୟ୫୮୪ୣ ୴୭୪୳୫ୣ∗ 100 [3-6] 32 3.5.9 Salt Purity To determine the ammonium sulphate salt purity, carbon hydrogen nitrogen sulfur (CHNS) elemental analysis was undertaken on the salt sample. The CHNS analysis was completed by a third-party laboratory. The purity of the salt (%) was then calculated using Equation 3-7 and the ammonia recovered (%) from ammonia stripping run using Equation 3-8. . 𝑃𝑢𝑟𝑖𝑡𝑦 (%) = 1 − ଶ଼ଵଷଶ+ ௡௜௧௥௢௚௘௡ ௪௘௜௚௛ %ଵ଴଴ ∗ 100 [3-7] 𝐴𝑚𝑚𝑜𝑛𝑖𝑎 𝑅𝑒𝑐𝑜𝑣𝑒𝑟𝑦 ቀ% ௚௚ቁ = ௉௨௥௜௧௬ (%) ∗ ௌ௔௟௧ ெ௔௦௦ (௚) ∗ యలభయమ஺௠௠௢௡௜௔ ெ௔௦௦ ௜௡ ோ௘௔௖௧௢௥ (௚) ∗ 100 [3-8] Scanning electron microscope (SEM) imaging and energy dispersive spectroscopy (EDS) at FILTER facility were also used to view the structure of the salt (Appendix B) and determine the elements present (Appendix C), respectively. The EDS did not provide accurate information on salt purity as the size of each site tested was small and it did not give the percent of the compound in the overall mass but rather related the frequency of each compound the test could detect which did not include hydrogen. EDS however did provide useful insights into what elements are present in salt impurities. For these tests, the salt was placed on carbon tape and coated in 80% platinum and 20% palladium before being placed in the microscope (Figure 3-9). Figure 3-9: Ammonium sulphate samples, coated in platinum and palladium, in the SEM/EDS setup. 33 3.5.10 Biogas Volume Biogas volume was measured using an Alicat M-0.5SCCM flowmeter that was connected to the anaerobic digester’s tedlar bag. The bag, which collects biogas produced by the digester was removed using clamps on either side of the tubing connector piece. When the external biogas measurement pump is turned on in the forward direction the tedlar bag is emptied and the volume of the air flowing out of it is measured. That volume was then normalized to standard temperature of 0ºC and pressure of 1 atm. The volume was also adjusted based on the time between the last volume measurement to ensure every reading give the biogas that would be produced in 24 hours. 3.5.11 Biogas composition Gas chromatography with a thermal conductivity detector (GC/TCD) was used to determine the composition of gas produced by the digesters (Huyssteen, 1967). A gas tight syringe with a needle tip was used to puncture the tubing that connects the digester headspace to the expandable tedlar bag, 0.5 mL of gas sample was collected using the syringe. The syringe tip was then changed to an injection tip and inserted into the rubber septa on the injection port of the Agilent 7820A GC/TCD. The plunger on the syringe was pushed down as the ‘start’ button was hit on the GC/TCD and the syringe tip was quickly removed from the injection port. The GC detector was set to a temperature of 70ºC, and the helium was used as the carrier gas. The carrier gas was pushed through the packed column at 25 mL/min. 3.5.12 Heavy Metals Heavy metal concentration in the digester effluent was tested at a third-party laboratory using an Agilent Tech 8900 ICP-MS (inductively coupled plasma mass spectrometry). The samples were prepared using microwave digestion at 180ºC for 20 minutes at BTG lab. The microwave tubes were filled with 1 mL of sample or standard, 8 mL of 69% nitric acid and 2 mL of 38% hydrochloric acid. The tubes were then tightly secured closed and fixed to the apparatus 34 that rotates to ensure equal digestion. After microwaving the acidic solution plus sample and allowing the solution to cool, the sample was serially diluted to a final dilution of 750 times and sent to the external laboratory in a 15 mL plastic centrifuge tube. 3.5.13 Capillary Suction Time The dewaterability of the digester effluent was tested using the capillary suction time method (American Public Health Association, 2005). The base of the capillary suction time apparatus is an acrylic pad that has metal pins to secure a piece of filter paper in place. A second acrylic pad sits atop of the pins and has a circular hole in the center for a metal cylinder to fit through and sit on the filter paper below. The top pad is connected to the fann440 capillary suction timer, displayed in Figure 3-10, with two probes at 15.9 mm and one probe at 22.2 mm from the center of the two pads. Figure 3-10: Capillary suction timer used for testing dewaterability of digester effluent. The samples were given time to warm to room temperature (22ºC). In this method, 6.4 mL of digester effluent sample was syringed delicately into the metal cylinder and the time between 35 the water in the sample spreading to a radius of 15.9 mm and 22.2 mm was detected based on when there is conduction between the probes. The capillary suction time can be used to compare how readily water can be separated from an effluent and therefore how easily it can be dewatered. When samples with different TS concentrations were analyzed, capillary suction time was normalized, and specific values were reported as seconds/%TS of sample. 3.5.14 Fecal Coliforms Digested samples were sent out to a third-party laboratory for fecal coliform testing. It is important for coliform testing that the digester effluent is tested within 24 hours of sampling. Results are provided in most probable number (MPN) of coliform colonies per 100 mL sample. 3.5.15 Nitrogen Heterocyclic Compounds The analysis for nitrogen heterocyclic compounds was conducted using a GC mass spectrometer. Digester effluent samples were first prepared by centrifuging for 10 minutes at 20,000 xg. The resulting centrate (around 50 mL) was transferred to a 250 mL glass bottle and 10 mL of dichloromethane was measured into the bottle. The bottle was then capped and placed on a platform shaker, set to 250 rpm, for 30 minutes. The sealed glass bottle was then left overnight so that they separated into water and dichloromethane layers. A long glass pipet was then used to transfer the bottom layer into a glass test tube. From there, 1 mL of the extracted sample was mixed with 40 µL of 50 µg/mL internal standard in each of the triplicate autosampler vials. Another triplicate set was made at a 10 times dilution with dichloromethane. Samples of HTLaq (non-pretreated and pretreated) were filtered at 0.45 µm pore size with a nylon filter to remove any formed solids. The extraction was performed in a similar manner but at a smaller scale, with 1 mL of aqueous sample mixed in a glass test tube with 2 mL of dichloromethane. Dilutions were made in autosampler vials at ten and fifty times with dichloromethane. The samples were then analyzed using an Agilent 8860 GC/ 5977B-Mass Spectrometer and an Agilent 30 m HP5MS column. The GC mass spectrometer was set to an 36 injector temperature of 250ºC, an oven program was used, and the MSD full scan mode was set to 40 to 400 amu. The instrument was calibrated using stock standards and the internal standard technique. Sample results were determined using the calibrations and multiple dilutions of the sample extracts. 3.5.16 Statistical Analysis of Data Statistical analysis was used when it was necessary to determine if there was a significant difference between the means of different data sets. All tests were run using Microsoft Excel with α = 0.05 which determines significant difference with 95% confidence. The type of analysis was chosen based on the number of treatments being tested and the size of the data set. When comparing three different data sets, for example determining if the ammonia removal was significantly different for three ammonia stripping runs all with different run temperatures, analysis of variance (ANOVA) was used. When analyzing data sets with only two treatments a t-test or z-test was used depending on the size of the data set. For data sets with less than 30 results a t-test was used and for over 30 results a z-test was used. 37 Chapter 4: Results and Discussion The following chapter presents and discusses the results of laboratory experiments. This chapter is divided into two sections; Section 4.1 HTL Aqueous Pretreatment and Section 4.2 Anaerobic Co-digestion of HTL Aqueous. 4.1 HTL Aqueous Pretreatment This section focuses on finding the ideal conditions for ammonia stripping with a goal of removing the most ammonia by mass. The success of different run conditions was determined by comparing the characterization of the influent and effluent. The second section in this chapter, Section 4.2, focuses on whether ammonia stripping is a suitable pretreatment method based on its biodegradability compared to non-pretreated HTLaq. The run condition used in the second section is based on the results from the optimization in this section. 4.1.1 Ammonia Stripping Feed Characterization The makeup of HTLaq varies with feed parameters so the characterization of the mixed sludge of 50% PS and 50% SS by mass that was used to generate the HTLaq is given in Table 4-1 below. Table 4-1: Characterization of mixed sludge and HTL aqueous for the ammonia stripping experiment. Parameter values represent the mean concentration of triplicate samples accounting for standard deviation between replicates. Parameter Mixed Sludge HTL Aqueous TAN (mg/L) 511 ± 24 4,500 ± 140 Total phenolics (mg GAE/L) 69.9 ± 0.8 1,640 ± 35 VFA (mg/L) 2,550 ± 340 4,620 ± 210 tCOD (mg/L) 10,100 ± 1,650 67,200 ± 2,400 pH 5.54 ± 0.3 7.7 ± 0.1 Total alkalinity (mg CaCO3/L) 1020 ± 73 13,900 ± 140 TS (%) (wt/wt) 3.99 ± 0.09 VS (%) (wt/wt) 3.47 ± 0.03 TSS (%) (wt/vl) 0.199 ± 0.0004 VSS (%) (wt/vl) 0.117 ± 0.0004 38 HTLaq is much higher in TAN and total phenolics than MS which means that HTLaq can be inhibitory to biological treatment processes. HTLaq is also higher in tCOD and VFAs which feed biological activity and can help increase methane generation. As HTLaq is the waste product separated from biocrude and hydrochar after filtering using 0.45 µm filter, there should not be any remaining TSS in this stream. However, it has been observed that HTLaq forms precipitates over time and even grows noticeably darker in colour. Mass removal (%) is used for analysis of much of the results moving forward using the concentrations in the feed HTLaq given in Table 4-1 and by using the known 110 mL sample volume. 4.1.2 Ammonia Mass Removal Ammonia removal from HTLaq was the primary objective of ammonia stripping and used to determine the optimum temperature, pH, and air flow rate. In the first three runs (Run 1, Run 2, Run 3), pH and air flow rate were held constant so that the effect of temperature on ammonia removal could be observed (Figure 4-1). Figure 4-1: Ammonia mass removal from HTLaq during ammonia stripping runs at three different temperatures. The ammonia removal shown in Figure 4-1 indicates that increasing the temperature of the sample (HTLaq) increases the amount of ammonia removed during the stripping period. The ideal temperature was determined to be 85ºC and was therefore used for the remainder of the 60657075808590951000 1 2 3 4Mass Removal (%)Run Time (hours)Run 1: 65°C, 10.3, 0.5 L/minRun 2: 75°C, 10.3, 0.5 L/minRun 3: 85°C, 10.3, 0.5 L/min39 ammonia stripping runs. An ANOVA test was conducted to validate that the mean ammonia removal was significantly affected by change in temperature. It was found that at a 95% confidence interval, there is a significant difference between the ammonia removal among runs with an F-value magnitude (103) far above the F-critical (5.14). A summary of the ANOVA parameters can be found in Table 4-2. Table 4-2: ANOVA summary table for analysis of significant mean difference among the ammonia mass removal of ammonia stripping run 1-3 with varying temperatures at 95% confidence (α = 0.05). Source of Variation Sum of Squares Degrees of Freedom Mean of Squares F-value P-value F-critical Between Groups 0.00261 2 0.00131 103 2.26E-05 5.14 Within Groups 7.61E-05 6 1.27E-05 Total 0.00269 8 In Run 4 and Run 5, temperature and airflow were held constant so the effect of pH change on ammonia removal could be observed. These two runs are compared to Run 3 and are shown in Figure 4-2 below. The figure shows that ammonia removal during ammonia stripping increased with pH. The highest removal was observed in Run 3 with a pH of 10.3, which was used as the ideal pH for the remainder of the runs. Figure 4-2: Ammonia mass removal from HTLaq during ammonia stripping runs at three different pH setpoints. An ANOVA analysis showed that there was a significant difference among the mean ammonia mass removal in runs where pH was varied. The F-value and F-critical value can be 20304050607080901000 1 2 3 4Mass Removal (%)Run Time (hours)Run 3: 85°C, 10.3, 0.5 L/minRun 4: 85°C, 9.8, 0.5 L/minRun 5: 85°C, 9.3, 0.5 L/min40 found in Table 4-3. Since the F-value is larger than F-critical, the null hypothesis, that the ammonia mass removal between runs was not significantly different, was rejected. Table 4-3: ANOVA summary table for analysis of significant mean difference among the ammonia mass removal of ammonia stripping runs 3-5 with varying pH setpoints at 95% confidence (α = 0.05). Source of Variation Sum of Squares Degrees of Freedom Mean of Squares F-value P-value F-critical Between Groups 0.0727 2 0.0363 50 2.06E-10 5.14 Within Groups 4.3E-05 6 7.16E-06 Total 0.0727 8 Run 6 and Run 7 had constant temperature and pH. To be able to assess the effect of air flow rate on ammonia removal, these runs were also compared to Run 3 in Figure 4-3. Figure 4-3: Ammonia mass removal from HTLaq during ammonia stripping runs at three different airflow rates. The figure shows that ammonia removal increased with faster air flow with 0.5 L/min yielding the most removal. An ANOVA concluded that there was a significant difference in mean ammonia mass removal among ammonia stripping runs with different air flow rates (Table 4-4). Table 4-4: ANOVA summary table for analysis of significant mean difference among the ammonia mass removal of ammonia stripping runs 3,6 and 7 with varying air flow rates at 95% confidence (α = 0.05). Source of Variation Sum of Squares Degrees of Freedom Mean of Squares F-value P-value F-critical Between Groups 0.0187 2 0.00936 1350 1.10E-08 5.14 Within Groups 4.18E-05 6 6.96E-06 Total 0.0188 8 505560657075808590951000 1 2 3 4Mass Removal (%)Run Time (hours)Run 3: 85°C, 10.3, 0.5 L/minRun 6: 85°C, 10.3, 0.3 L/minRun 7: 85°C, 10.3, 0.1 L/min41 All three parameters analyzed (temperature, pH and airflow rate) showed significant ammonia removal when increased. This is to be expected as, increasing the pH promotes the shift of ammonium to ammonia gas, which increases the ammonia available for mass transfer and increasing the temperature or airflow rate increases the mass transfer coefficient of ammonia (Liu, et al., 2015). The best run was Run 3 (85°C, 10.3, 0.5 L/min) with 99.5 ± 0.09% ammonia removal. The lowest ammonia removal was experienced when the pH was at its lowest setpoint which suggests that pH has the highest impact on ammonia removal. Evaluating the p-values of the three parameters also identifies that pH has the largest variation between treatments, and therefore the greatest impact on ammonia removal within the ranges tested. Adghim et. al. (2022) also found pH to have a greater impact than temperature on ammonia removal in post-hydrolysis sludge, but interestingly found that increased temperature had more improvement on the methane potential indicating higher biodegradability. This is why ammonia removal should not be used alone to evaluate the best ammonia stripping conditions. From subsections 4.1.2 to 4.1.8, the effect of ammonia stripping on other HTLaq characteristics is discussed to determine the ideal conditions for HTLaq pretreatment and resource recovery. 4.1.3 Phenolic Compounds Mass Removal Phenolic compound removal for all runs is presented in Figure 4-4. In some cases, there was an increase in phenolic compounds during stripping which is denoted as a negative removal. Ammonia stripping literature reviewed did not analyze phenolic compounds to confirm the trend of them being created at lower operational temperatures. However, in a study of HTL pathways it was found that the organics in MS can lead to the creation of phenols through hydrolysis of lignin/humic substances or oxidation of carbohydrate derivatives (Liu, et al., 2022). More research is needed on the pathways of HTLaq components to phenols through temperature and pH adjustment to conclude how phenolic compounds are being created during ammonia stripping. 42 Figure 4-4: Total phenolic compounds removal from HTLaq during all ammonia stripping runs. Total phenolic compounds were removed up to 32 ± 1 % at the conditions of Run 5 (85°C, pH of 9.3, 0.5 L/min). This indicates that more phenolics are removed when the pH of the solution is lower. 4.1.4 VFA Mass Removal The mass removal of VFAs throughout ammonia stripping for all runs is graphed in Figure 4-5. Results show that to reduce the amount of VFAs removed from the HTLaq sample, the pH and temperature should be low. The air flow rate had minimal effect on VFA removal, with the three runs with the highest VFA removal being the ones with the highest pH and temperature setpoints and only the air flow rate varying. Run 6 and Run 7 had the highest removal of VFAs with 91 ± 1% and 90 ± 0.4% removed, respectively. -60-40-2002040600 1 2 3 4Mass Removal (%)Run Time (hours)Run 1: 65°C, 10.3, 0.5 L/minRun 2: 75°C, 10.3, 0.5 L/minRun 3: 85°C, 10.3, 0.5 L/minRun 4: 85°C, 9.8, 0.5 L/minRun 5: 85°C, 9.3, 0.5 L/minRun 6: 85°C, 10.3, 0.3 L/minRun 7: 85°C, 10.3, 0.1 L/min43 Figure 4-5: Volatile fatty acid removal from HTLaq during all ammonia stripping runs. 4.1.5 COD Mass Removal For the COD removal, HTLaq was sampled every hour during ammonia stripping and the results are displayed in Figure 4-6 below. To reduce the COD loss during ammonia stripping the best conditions were to keep the temperature, pH and air flow rate low. Within the pH range of 9.3-10.3, pH changes seem to have the smallest effect on COD removal with the conditions of the two top removal runs being consistent other than their pH setpoints. The runs with the highest COD mass removal were Run 4 and Run 3 with 30 ± 1% and 29 ± 2% removals, respectively after a runtime of 4 hours. For all runs at all times measured, COD decreased in the system during stripping. This differs from the results in sections 4.1.3 and 4.1.4, which showed that some operational conditions resulted in an increase in phenolic compounds or an initial increase in VFAs, respectively. Since phenolic compounds and VFAs are organics, they should impact the tCOD of the HTLaq. One explanation for the discrepancy could be that organics are changing form during stripping, not being added to the system, so they do not greatly impact the overall COD. -60-40-200204060801000 1 2 3 4Mass Removal (%)Run Time (hours)Run 1: 65°C, 10.3, 0.5 L/minRun 2: 75°C, 10.3, 0.5 L/minRun 3: 85°C, 10.3, 0.5 L/minRun 4: 85°C, 9.8, 0.5 L/minRun 5: 85°C, 9.3, 0.5 L/minRun 6: 85°C, 10.3, 0.3 L/minRun 7: 85°C, 10.3, 0.1 L/min44 Figure 4-6: Total chemical oxygen demand removal from HTLaq during all ammonia stripping runs. Another possibility for the discrepancy between COD, VFA and phenolic values, is that the relative COD provided by these organics are insignificant compared to the tCOD of HTLaq. The COD equivalence of acetic acid, propionic acid and butyric acid measured to represent VFAs is known to be 1.067, 1.514 and 1.818 g COD/g acid respectively (Rossle & Pretorius, 2001). Using these values, the VFA contribution to the feed HTLaq tCOD was found to be 5.96 ± 0.3 g/L out of the 67.2 ± 2.4 gg/L (around 9%). The two stripping runs where VFAs were initially increased (Run 1 and Run 5) are the runs with the lowest COD removal at time zero. This shows that though the trends between VFA and COD do not fully correlate between operational conditions the VFA creation did have an effect on the COD levels. 4.1.6 Alkalinity Mass Removal Total alkalinity at ammonia stripping reaction time zero (0-hour) can differ from the HTLaq feedstock alkalinity (13.9 g CaCO3/L, in Section 4.1.1) due to the effect of NaOH addition to reach the pH setpoint, which is added to the sample prior to heating. More NaOH addition is 051015202530350 1 2 3 4Mass Removal (%)Run Time (hours)Run 1: 65°C, 10.3, 0.5 L/minRun 2: 75°C, 10.3, 0.5 L/minRun 3: 85°C, 10.3, 0.5 L/minRun 4: 85°C, 9.8, 0.5 L/minRun 5: 85°C, 9.3, 0.5 L/minRun 6: 85°C, 10.3, 0.3 L/minRun 7: 85°C, 10.3, 0.1 L/min45 required for runs with higher pH setpoint and runs with lower temperatures to achieve the required pH. The removals in all previous subsections of 4.1 HTL Aqueous Pretreatment are based on the difference in mass of each parameter in the initial sample versus the mass remaining in the flask at the sampling time or experiment conclusion (accounting for sample removed for time-series testing). The removal for alkalinity, displayed in Table 4-5, was only calculated over the 3-hour runtime using the difference between the 0-hour and effluent sample due to large impact of NaOH addition. This better displays the effect of the ammonia stripping itself but will not account for reactions occurring while the sample is being heated to the temperature setpoint, though there is no airflow provided prior to the 0-hour sample being taken to assist the removal of volatilized chemicals out of the reactor. Table 4-5: Total alkalinity at the start of the ammonia stripping reaction and in the flask effluent along with calculated mass removal during the 3-hour run time. Run No. NaOH addition (mL) Alkalinity at 0 hr (g CaCO3/L) Alkalinity in Effluent* (g CaCO3/L) Alkalinity Mass Removal* (%) 1: 65°C, 10.3, 0.5 L/min 4.0 28.9 17.3 ± 0.07 45.6 ± 0.2 2: 75°C, 10.3, 0.5 L/min 3.5 26.8 15.1 ± 0.24 50.3 ± 0.8 3: 85°C, 10.3, 0.5 L/min 3.4 25.3 14.7 ± 0.84 48.4 ± 0.7 4: 85°C, 9.8, 0.5 L/min 2.4 22.7 8.89 ± 0.09 65.9 ± 0.3 5: 85°C, 9.3, 0.5 L/min 1.4 19.1 7.07 ± 0.15 67.0 ± 0.7 6: 85°C, 10.3, 0.3 L/min 3.4 27.1 13.0 ± 0.17 55.5 ± 0.6 7: 85°C, 10.3, 0.1 L/min 3.4 26.4 15.2 ± 0.03 46.6 ± 0.1 *Mean values are provided for duplicate samples with a range provided using the standard error. Alkalinity in the 0-hour sample was higher than the initial HTLaq (13.9 g CaCO3/L) in all cases indicating that NaOH addition increases alkalinity. Comparing the 0 hour to effluent alkalinity concentration, it is clear that alkalinity decreases during the stripping run. For most runs, the effluent alkalinity was still higher than in the non-pretreated HTLaq but for runs 4 and 5 with less NaOH addition, the alkalinity removed during stripping was enough to have an overall lower alkalinity in the pretreated HTLaq. 46 4.1.7 Total and Volatile Suspended Solids Mass Removal The removal or production of TSS and VSS during ammonia stripping runs is given in Table 4-6. The TSS and VSS in the ammonia stripping feedstock was very low, 0.199 ± 0.0004% and 0.117 ± 0.0004% respectively, so any reduction or addition of solids in the reactor had large impacts on removal. Table 4-6: Total and volatile suspended solids mass removal from HTLaq during ammonia stripping. Run No. TSS Removal (%) VSS Removal (%) 1: 65°C, 10.3, 0.5 L/min - 0.1 ± 0.4 6.4 ± 3.0 2: 75°C, 10.3, 0.5 L/min 55 ± 4.2 51 ± 3.2 3: 85°C, 10.3, 0.5 L/min 7.6 ± 5.1 - 2.9 ± 7.1 4: 85°C, 9.8, 0.5 L/min 29 ± 0.3 15 ± 1.7 5: 85°C, 9.3, 0.5 L/min 19 ± 2.8 2.5 ± 5.2 6: 85°C, 10.3, 0.3 L/min 5.5 ± 0.4 - 9.0 ± 0.8 7: 85°C, 10.3, 0.1 L/min 25 ± 1.1 5.3 ± 0.2 The run with the highest TSS and VSS removal was Run 2, which has over 50% reduction of both parameters. Run 1 was the only increase in TSS and two runs, Run 3 and Run 6, had an increase in VSS during the stripping run. No clear trend emerges between suspended solids results and ammonia stripping setpoints. It is possible that the large variation in removals in due to experimental error given that the highest mass calculated for solids remaining after drying and ashing was 0.010 g and 0.004 g, respectively. This is relatively small compared to the mass of dish and filter which was 1.38 g on average. 4.1.8 Ammonium Sulphate Salt Analysis The percentage of carbon, hydrogen, nitrogen, and sulfur found in salts recovered from HTLaq during each ammonia stripping run can be found in Table 4-7 below. The highest salt purity was achieved in Run 6 with 98.1 ± 0.01% purity. Run 1 had the lowest purity with only 85.3 ± 0.12% of the salt mass being ammonium sulphate. 47 Table 4-7: CHNS analysis results and purity in the recovered ammonium sulphate salt. Sample Carbon (%) Hydrogen (%) Nitrogen (%) Sulfur (%) Salt Purity* (%) Run 1 Salt 0 3.45 6.51 23.2 85.3 ± 0.12 Run 2 Salt 1.77 5.71 17.7 22.7 96.5 ± 0.47 Run 3 Salt 1.55 4.99 14.2 24.0 93.0 ± 0.30 Run 4 Salt 1.30 5.96 18.2 23.5 97.0 ± 1.5 Run 5 Salt 0.00 5.95 19.3 23.1 98.0 ± 0.05 Run 6 Salt 1.52 6.03 19.3 23.7 98.1 ± 0.01 Run 7 Salt 1.00 5.96 19.1 23.5 97.9 ± 0.03 *CHNS analysis was done in duplicate and the difference between the mean and each data point was given as the error. Table 4-8 shows that Run 1 also had the lowest ammonia recovery at only 34.0 ± 0.05% which may explain why this salt took longer to dry compared to the rest of the runs, since the absorption column solution would still be acidic there was less ammonia neutralizing the sulfuric acid to create ammonium sulphate. Table 4-8: Summary of nitrogen recovery from HTLaq during ammonia stripping runs. Run No. Setpoints Ammonia Mass Removal (%) Salt Purity (%) Ammonia Recovery (%) Temp (°C) pH Air Flow (L/min) 1 65 10.3 0.5 95.4 ± 0.6 85.3 ± 0.1 34.0 ± 0.05 2 75 10.3 0.5 97.1 ± 0.2 96.5 ± 0.5 92.7 ± 0.5 3 85 10.3 0.5 99.5 ± 0.1 93.0 ± 0.3 78.2 ± 0.3 4 85 9.8 0.5 97.4 ± 0.4 97.0 ± 1.5 94.6 ± 1.5 5 85 9.3 0.5 79.5 ± 0.1 98.0 ± 0.05 73.9 ± 0.04 6 85 10.3 0.3 98.5 ± 0.2 98.1 ± 0.01 94.1 ± 0.01 7 85 10.3 0.1 89.4 ± 0.4 97.9 ± 0.03 84.3 ± 0.02 Another salt analysis performed was by EDS which identified that some of the impurities in the ammonium sulphate salt were calcium, sodium, zirconium, potassium, carbon, and magnesium. More information on the elements found on tested sites of the salt can be found in Appendix C. During EDS analysis, SEM images were also taken of the ammonium salts from each run which can be found in Appendix B. 48 4.2 Anaerobic Co-digestion of HTL Aqueous The anaerobic digesters for the co-digestion experiment were operated prior to the recording of experimental data shown in this section. Characterization of the inoculum initially used for co-digesters in this research is presented in Table 4-9. This gave the digesters, being fed with HTLaq, time to acclimatize to this feed. The control and HTLaq digesters under thermophilic conditions were run for over 400 days for a previous study by another BTG member prior to the start of my experimentation (Azarmina, 2023). The third digester used in thermophilic testing was built up from the effluent of the original HTLaq digester. Once this new digester was stable on day 138 of experimentation, the two identically operated HTLaq digesters were mixed together and equally distributed between two digester flasks. On this day one of the digesters was switched to pretreated HTLaq. Table 4-9: Characterization of co-digestion inoculum. Parameter values represent the mean concentration of triplicate samples accounting for standard deviation between replicates. Response Parameter Thermophilic Control Thermophilic 5%HTL Mesophilic Control Mesophilic 13%HTL TAN (mg/L) 1,760 ± 46 1,580 ± 6 1,560 ± 23 1,390 ± 42 Total phenolics (mg GAE/L) 158 ± 0.3 190 ± 3 53.3 ± 0.6 173 ± 2 VFA (g/L) 3.37 ± 0.19 2.73 ± 0.06 3.34 ± 0.16 2.61 ± 0.21 tCOD (g/L) 22.8 ± 1.6 19.1 ± 0.1 22.6 ± 0.3 15.5 ± 0.4 pH 7.73 ± 0.21 7.86 ± 0.16 6.99 ± 0.10 7.19 ± 0.06 Total alkalinity (g CaCO3/L) 6.30 ± 0.29 6.07 ± 0.25 5.36 ± 0.03 6.18 ± 0.42 TS (%) (wt/wt) 1.77 ± 0.03 1.49 ± 0.07 1.67 ± 0.03 1.24 ± 0.09 VS (%) (wt/wt) 1.29 ± 0.03 1.15 ± 0.03 1.28 ± 0.02 0.956 ± 0.041 A similar approach was taken for the mesophilic digesters, which were from another study by BTG members (Basar, et al., 2024). The mesophilic digesters were run for over 180 days prior to the beginning of this project. A full characterization was done for the digesters at the first 49 non-pretreated/pretreated HTLaq feeding condition before increasing the contribution of non-pretreated/pretreated HTLaq in the feed to the final testing condition. 4.2.1 Co-digestion Feed Characterization The feedstock makeup for the co-digestion experiment was dependant on the contribution of MS and the three other co-substrates. The mixing ratios of the co-substrates and resulting average COD contribution for each digester condition was previously presented in Table 3-5. The feedstock is also dependent on changes to MS based on operational changes and WWTP influent variations. Over a year of experimentation, the MS and resulting centrate and HTLaq characterization fluctuated, as shown in the standard deviations in Table 4-10, which simulated the feed variations full-scale digesters would experience at a WWTP. Table 4-10: Characterization of mixed sludge, centrate and HTL aqueous for the co-digestion feedstock. Parameter values represent the mean concentration of triplicate samples accounting for standard deviation between replicates. Parameter Mixed Sludge Centrate HTL Aqueous HTL Aqueous Stripped TAN (mg/L) 689 ± 213 914 ± 164 2,520 ± 247 74.3 ± 18 Total phenolics (mg GAE/L) 103 ± 12 104 ± 13 1,580 ± 111 461 ± 118 VFA (g/L) 2.34 ± 0.81 3.28 ± 0.69 7.95 ± 0.23 1.90 ± 0.37 tCOD (g/L) 51.2 ± 3.7 15.9 ± 4.4 61.6 ± 4.2 52.8 ± 4.0 pH 5.52 ± 0.16 5.39 ± 0.17 5.87 ± 0.31 8.86 ± 0.72 Total alkalinity (g CaCO3/L) 1.42 ± 0.28 1.15 ± 0.17 4.14 ± 0.72 7.47 ± 0.48 TS (%) (wt/wt) 3.42 ± 0.39 0.754 ± 0.26 0.794 ± 0.26 0.414 ± 0.10 VS (%) (wt/wt) 3.01 ± 0.29 0.587 ± 0.19 0.745 ± 0.27 0.288 ± 0.12 HTLaq is more concentrated than MS in organics especially phenolic compounds as well as having higher TAN and alkalinity. HTLaq, as well as centrate added to co-digestion reactors, is much lower in solids and therefore result in a much thinner feedstock. 50 4.2.2 Biogas Production and Composition The thermophilic anerobic digesters were operated for 322 days and the daily biogas production is displayed in Figure 4-7. Although all 3 digesters had the same SRT of 20 days throughout the operation, OLRs ranged from 1.8-2.5 kg COD/m3d corresponding to the HTL contribution from 0-13%. Figure 4-7: Biogas production of the thermophilic control digester, digester fed non-pretreated HTLaq (HTLaq), and digester fed HTLaq pretreated through ammonia stripping (HTLaqSTRIP). The contribution of non-pretreated and pretreated HTLaq in the feedstock is indicated at the top of the graph. In this graph, the control digester has a higher average biogas production throughout the experiment which can be explained by the higher OLR of 2.5 kg COD/m3d from MS compared to the HTLaq digester with an OLR of 1.9-2.3 kg COD/m3d from HTLaq and centrate with less tCOD than MS. Therefore, the HTLaq digester has less potential to generate methane and the biogas production will be lower even when not inhibited. This also explains the decrease in biogas production after the HTLaq contributions to the feed was increased on day 167 and 172 of the experiment. However, since the HTLaq and HTLaq stripped digesters have similar 0501001502002503003504004505000 20 40 60 80 100 120 140 160 180 200 220 240 260 280 300 320 340Biogas Production (mL/d)Run Time (d)ControlHTLaqHTLaq Stripped8%HTLaq 13%HTLaq8%HTLaqSTRIP 12%HTLaqSTRIP5%HTLaqSTRIP5%HTLaq5%HTLaq51 influent CODs, the theoretical methane yields do not explain the difference between these two digesters. This difference that arises after the condition change is likely due to inhibition in the non-pretreated HTLaq digester which suggests that the pretreatment of HTLaq improves biodegradability in anaerobic conditions. To focus on efficiency of biogas generation per organics fed rather than total biogas production Figure 4-8 displays specific biogas yield. Figure 4-8: Specific biogas yield of the thermophilic control digester, digester fed non-pretreated HTLaq (HTLaq), and digester fed HTLaq pretreated through ammonia stripping (HTLaqSTRIP). The contribution of non-pretreated and pretreated HTLaq in the feedstock is indicated at the top of the graph. When reviewing daily biogas production normalized for influent COD it can be observed that at the initial condition all digesters have a similar efficiency of converting organics to biogas with an average daily production of approximately 0.24 mL/g COD fed/day. This changes after around day 210 of testing when both HTLaq digesters have a drop in specific production, likely a delayed response to the condition change on day 172. Though HTLaq stripped digester experiences a period of reduced specific production, it recovers to the original production 0.00.10.20.30.40.50 20 40 60 80 100 120 140 160 180 200 220 240 260 280 300 320 340Specific Biogas Yield (mL/g COD fed/d)Run Time (d)ControlHTLaqHTLaq Stripped8%HTLaq 13%HTLaq8%HTLaqSTRIP 12%HTLaqSTRIP5%HTLaq5%HTLaq5%HTLaqSTRIP52 efficiency. The non-pretreated HTLaq digester however is not able to recover from its drop in production after the fed is changed to 13% of COD contribution to the feed coming from HTLaq. A Z-test was chosen to determine if the specific biogas yield was significantly different between the digesters fed non-pretreated and pretreated HTLaq due to the large sample size (92 daily biogas measurements after steady-state was reached on day 232 of testing). A summary of the results can be found in Table 4-11. Table 4-11: Z-test summary table for analysis of significant mean difference between the specific biogas yield of non-pretreated and pretreated HTLaq fed thermophilic digesters at 95% confidence (α = 0.05). Thermophilic 13%HTLaq Thermophilic 13%HTLaqStrip Mean 0.0955 0.2043 Known Variance 0.00065 0.0015 Observations 92 92 Hypothesized Mean Difference 0 z-value -22.5 z-critical (Two-Tail) 1.64 To reject the null hypothesis that there was no significant difference between the mean specific biogas yield of the non-pretreated and pretreated HTLaq digesters, the z-value would have an absolute value greater than 1.64 (z-critical). The z-value for the digesters was -22.5 therefore it can be concluded that there is a significant difference between the specific biogas yield means. The digester fed pretreated HTLaq had a higher mean specific biogas yield, meaning that pretreatment of HTLaq improved biogas yield. The daily biogas production was also tested at mesophilic conditions and is displayed in Figure 4-9 where the pretreated and non-pretreated HTLaq digesters start at 13% COD fed coming from HTLaq (as explained in Figure 3-6). The non-pretreated HTLaq digester increases to 24% COD contribution as indicated on the graph but the HTLaq stripped digester only reaches 22% COD contribution from treated HTLaq due to the reduction of COD during ammonia stripping. For mesophilic digesters, while control had an OLR of 2.5 kg COD/m3d from MS (same as its thermophilic counterpart), HTLaq and HTLaqStripped digesters had OLR ranges of 1.4-1.9 kg COD/m3d at the HTLaq contribution range of 13-24%. 53 The mesophilic HTLaq digester does not appear to have the same inhibition problem as the thermophilic HTLaq digester with the pretreated and non-pretreated HTLaq digesters performing similarly throughout the experiment. The specific biogas production of these mesophilic digesters with respect to COD is given in Figure 4-10. The specific biogas production of the mesophilic digesters when stable was similar to the thermophilic digesters, around 0.25 mL/g COD fed/day. The digester fed stripped HTLaq had no biogas yield values for the period between day 254 and day 282 due to a tedlar bag leak. Once the bag was replaced biogas production was shown to similar to digester fed HTLaq. A bag leak also occurred in the control digester from day 78 to 126. These values were removed from the graphs and any statistical analysis as they do not reflect the condition of the digesters. 0501001502002503003504004505000 20 40 60 80 100 120 140 160 180 200 220 240 260 280 300 320 340Biogas Production (mL/d)Run Time (d) ControlHTLaqHTLaq Strippedbag leak18%HTLaq 24%HTLaq18%HTLaqSTRIP 22%HTLaqSTRIP13%HTLaqSTRIP13%HTLaq13%HTLaqbag leakFigure 4-9: Biogas production of the mesophilic control digester, digester fed non-pretreated HTLaq (HTLaq), and digester fed HTLaq pretreated through ammonia stripping (HTLaqSTRIP). The contribution of non-pretreated and pretreated HTLaq in the feedstock is indicated at the top of the graph. 54 Figure 4-10: Specific biogas yield of the mesophilic control digester, digester fed non-pretreated HTLaq (HTLaq), and digester fed HTLaq pretreated through ammonia stripping (HTLaqSTRIP). The contribution of non-pretreated and pretreated HTLaq in the feedstock is indicated at the top of the graph. A Z-test was completed to determine if the specific biogas yield was significantly different between the mesophilic digesters fed non-pretreated and pretreated HTLaq and a summary of the results can be found in Table 4-12. The null hypothesis, that there was no significant difference between the mean specific biogas yield of the mesophilic digesters fed 24% pretreated and 22% non-pretreated HTLaq after steady-state was reached (day 232), was rejected as the z-value for the digesters of 5.90 has an absolute value greater than the z-critical (1.64) for a 95% confidence interval. This means there is a significant difference between the specific biogas yields, and it can be concluded that pretreatment of HTLaq reduces biogas production in the mesophilic digesters though neither reactor showed signs of inhibition. 0.00.10.20.30.40 20 40 60 80 100 120 140 160 180 200 220 240 260 280 300 320 340Specific Biogas Yield (mL/g COD fed/d)Run Time (d) ControlHTLaqHTLaq Strippedbag leak18%HTLaq 24%HTLaq18%HTLaqSTRIP 22%HTLaqSTRIP13%HTLaqSTRIP13%HTLaq13%HTLaqbag leak55 Table 4-12: Z-test summary table for analysis of significant mean difference between the specific biogas yield of non-pretreated and pretreated HTLaq fed mesophilic digesters at 95% confidence (α = 0.05). Mesophilic 24%HTLaq Mesophilic 22%HTLaqStrip Mean 0.226 0.199 Known Variance 0.00083 0.00049 Observations 85 45 Hypothesized Mean Difference 0 z-value 5.90 z-critical (Two-Tail) 1.64 To understand the energy generation potential of the digesters, the percentage of methane in the biogas as well as the amount of biogas being produced needs to be analyzed. The distribution of methane (CH4), carbon dioxide (CO2), oxygen (O2) and nitrogen (N2) for thermophilic digesters is displayed in Figure 4-11. After initial results were gathered the HTLaq contributions were increased, and ten biogas composition tests were performed at steady-state. Figure 4-11: Biogas composition in thermophilic anaerobic digesters. The methane percentage in biogas was quite variable between weeks with standard deviations between 3% and 7% for all digesters. Biogas composition for mesophilic digesters is displayed in Figure 4-12. The methane percentage in biogas produced by each digester were similar to each other. For this reason, it cannot be concluded that there is a difference in 01020304050607080901001100%HTLaq 5%HTLaq 5%HTLaqStrip 13%HTLaq 12%HTLaqStripFraction of Biogas (%)CH4 CO2 O2 N256 methane percentage in any digester. The mesophilic 24%HTLaq and 22%HTLaqStrip digester which had average methane percentage of 66 ± 4% and 55 ± 5% methane, respectively. Figure 4-12: Biogas composition in mesophilic anaerobic digesters. 4.2.3 Total Ammonium Nitrogen (TAN) TAN was measured in the AD feedstock and effluent for each thermophilic digester (Figure 4-13). Though, HTLaq is high in TAN (2,520 ± 21 mg/L) compared to MS it does not greatly impact TAN in the 5%HTLaq digester feedstock due to the small volume incorporated. Figure 4-13: Total ammonium nitrogen (TAN) concentration in the thermophilic anaerobic digester influent and effluent. 01020304050607080901001100%HTLaq 13%HTLaq 13%HTLaqStrip 24%HTLaq 22%HTLaqStripFraction of Biogas (%)CH4 CO2 O2 N2 - 200 400 600 800 1,000 1,200 1,400 1,600 1,8000%HTLaq 5%HTLaq 5%HTLaqStrip 13%HTLaq 12%HTLaqStripTAN (mg/L)Feed Effluent57 With a feedstock of 13% HTLaq by COD, the influent TAN is increased but there is still little effect on the effluent TAN concentration (which also describes the concentrations in the digesters because of continuous stirred tank reactor nature of the reactor). The TAN concentration in the digesters is generally higher than that of the influent because ammonia is produced during the breakdown of proteins during AD. Figure 4-14 displays the effect of different HTLaq feedstocks on mesophilic digesters. In the final condition, with higher HTLaq contributions, there is an increase in TAN concentrations in the feedstocks which have HTLaq while the HTLaqStrip feedstocks remain at similar levels to the control MS feed. The digester TAN concentrations do not vary significantly between conditions or feeds which suggests that none of the digesters are experiencing inhibition due to high TAN levels. Figure 4-14: Total ammonium nitrogen (TAN) concentration in the mesophilic anaerobic digester influent and effluent. Free ammonia nitrogen has been shown to be inhibitory to mesophilic AD at levels of 150 mg NH3-N/L (McCarty & McKinney, 1961). Typically, thermophilic cultures are thought to be more sensitive due to their decreased microbial diversity but Gallert and Winter (1997) found that thermophilic digesters could handle twice the free ammonia nitrogen as their mesophilic - 200 400 600 800 1,000 1,200 1,400 1,600 1,800 2,0000%HTLaq 13%HTLaq 13%HTLaqStrip 24%HTLaq 22%HTLaqStripTAN (mg/L)Feed Effluent58 counterparts while still digesting proteins. This study indicated that thermophilic digesters would begin experiencing inhibition at levels of 250 mg NH3-N/L with aceticlastic methanogens experiencing the most inhibition. Given the higher temperature of thermophilic digesters, a lower TAN concentration is required to reach the same level of free ammonia nitrogen concentration as mesophilic digesters. The mesophilic digester with the highest TAN concentration was the 13%HTLaq digester at 1,570 ± 164 mg/L which at a pH of 7.3 gives a free ammonia nitrogen concentration of 33.9 ± 3.6 mg NH3-N/L. All mesophilic digesters were below the inhibitory level of 150 mg NH3-N/L. The thermophilic digester with the highest TAN concentration was the control digester at 1,620 ± 92 mg/L which also had the highest pH at 7.8 giving a free ammonia nitrogen 317 ± 18 mg NH3-N/L. This is above the inhibition level seen in literature but the control digester ran smoothly throughout the year-long experiment. The thermophilic 13%HTLaq digester condition however, had a free ammonia nitrogen below the inhibition level (155 ± 19 mg NH3-N/L) but showed other signs of inhibition. These results suggest that neither the TAN nor the more toxic form of free ammonia nitrogen are the cause of inhibition in the thermophilic 13%HTLaq digester. 4.2.4 Volatile Fatty Acids Total VFAs were measured in the digesters in terms of acetic, propionic, and butyric acid, and the results for the thermophilic digesters can be found in Figure 4-15 and mesophilic digesters in Figure 4-16. The average VFA concentrations found in co-substrates can be found in 4.2.1 Co-digestion feedstock characterization. These average values would suggest that increasing the contribution of centrate and HTLaq would increase the VFA concentration in feed, with only HTLaqStrip (1.90 ± 0.37 g/L) having a lower VFA concentration than MS (2.34 ± 0.81 g/L). It is also important to note that VFA concentration in WWTP MS is variable between shipments and can vary with seasonal or operational changes. 59 Figure 4-15: Volatile fatty acid concentration in the thermophilic anaerobic digester influent and effluent. For this experiment, VFA concentration in MS and the resulting centrate and HTLaq decreased throughout the experimental run. Since, only three digesters were operated at each temperature condition (a control, HTLaq and HTLaqStrip) the results for different percentage of COD contribution from HTLaq or HTLaqStrip are results from the effluent of same digesters at different times during the year of experimentation. The seasonal change in VFA concentrations explains why digesters with higher contributions from co-substrates with higher average VFA concentrations had lower influent VFA concentrations than those with lower contributions from these co-substrates. The lack of VFA reduction in the 13%HTLaq digester suggests AD inhibition was experienced when the HTLaq contribution was increased from 5% COD coming from HTLaq to 13% under thermophilic conditions (Figure 4-15). The inhibition is likely being experienced during acetogenesis, the phase of AD where propionic and butyric acid are converted to acidic acid, because of the minimal butyric acid reduction and the increase in propionic acid levels (Metcalf and Eddy, Inc., 2014). Acidic acid or the conjugate base of acetate is an intermediate product in the creation of methane so VFAs not being reduced in 13%HTLaq digester explains 050010001500200025003000350040004500InfluentEffluent InfluentEffluent InfluentEffluent InfluentEffluent InfluentEffluentVFA (mg/L)Acetic Acid Propionic Acid Butyric Acid0%HTLaq 5%HTLaq 5%HTLaqStrip 13%HTLaq 12%HTLaqStrip60 the decrease in biogas production discussed in section 4.2.2. The minimal reduction in acetic acid suggests that archaea involved in the methanogenesis phase were also inhibited. Figure 4-16: Volatile fatty acid concentration in the mesophilic anaerobic digester influent and effluent. All mesophilic digesters had large reductions in VFAs suggesting no digesters were inhibited. No conclusions can be drawn on the effect of HTLaq contribution or pretreatment. 4.2.5 Chemical Oxygen Demand The tCOD of the thermophilic digester influent and effluent is graphed in Figure 4-17. When HTLaq contribution to the thermophilic feedstock increased from 5% COD to 13% COD from HTLaq the COD reduction decreased from 52 ± 6% to 22 ± 9%. The digesters receiving stripped HTLaq did not see the same impact with the COD reduction of 46 ± 7% at 5%HTLaqStrip and 55 ± 9% at 12%HTLaqStrip. A t-test comparing 13%HTLaq and 12%HTLaqStrip digesters was done to determine if stripping HTLaq prior to co-digestion could improve the COD reduction at the increased COD contribution. The analysis revealed that the t-critical to show significant difference in the mean COD removal was 2.11 and the t-value between the digesters was -7.38. Since the absolute value of the t-value is greater than the t-critical it can be concluded that pretreatment of HTLaq significantly reduced the COD removal, again suggesting inhibition in this thermophilic digester. 0100020003000400050006000InfluentEffluent InfluentEffluent InfluentEffluent InfluentEffluent InfluentEffluentVFA (mg/L)Acetic Acid Propionic Acid Butyric Acid0%HTLaq 13%HTLaq 13%HTLaqStrip 24%HTLaq 22%HTLaqStrip61 Summary tables for the t-tests, calculated for thermophilic and mesophilic digesters, can be found in Appendix D Sample Statistical Analysis Tables. The impact of increased HTLaq on COD reduction for mesophilic digesters is shown in Figure 4-18. The lowest average COD removals for the mesophilic digesters was seen in the digesters being fed HTLaq with 55 ± 6% removal at 13%HTLaq and 52 ± 4% at 24%HTLaq compared to the control digester of 60 ± 5%. Figure 4-18: Total chemical oxygen demand concentration in the mesophilic anaerobic digester influent and effluent. - 10,000 20,000 30,000 40,000 50,000 60,0000%HTLaq 5%HTLaq 5%HTLaqStrip 13%HTLaq 12%HTLaqStriptCOD (mg/L)FeedEffluent - 10,000 20,000 30,000 40,000 50,000 60,0000%HTLaq 13%HTLaq 13%HTLaqStrip 24%HTLaq 22%HTLaqStriptCOD (mg/L)FeedEffluentFigure 4-17: Total chemical oxygen demand concentration in the thermophilic anaerobic digester influent and effluent. 62 The digesters being fed stripped HTLaq had an increase in COD removal (from 66 ± 3% to 74 ± 7%) when HTLaq contribution to feedstock COD was increased from 13% to 22%. Though influent tCOD decreased in the 24%HTLaqStrip and 22%HTLaqStrip mesophilic digesters, when HTLaq contribution was increased, the COD reduction during digestion remained similar to the previous feed conditions. A t-test performed between 24%HTLaqStrip and 22%HTLaqStrip found no significant difference between the mean COD removal with a t-value, 0.185, within the t-critical range of -2.26 to 2.26 for the test. This leads to the null hypothesis, that the mean COD removal is insignificantly different between the mesophilic digesters being fed non-pretreated and pretreated HTLaq, being validated. 4.2.6 Alkalinity and pH All the mesophilic digesters had an average pH of 7.3 ± 0.02 with the control digester showing the most variation (between 6.9 and 8.2 pH throughout the year of operation). The average pH of the thermophilic digesters ranged from 7.5 (13%HTLaq) and 7.8 (0%HTLaq). The control digester was also the most variable for the thermophilic digesters with a pH range of 7.1 to 8.1. Given that alkalinity is the capacity of a substance to resist decreases in pH, a lower pH will likely result in a lower alkalinity. This is because the more acidic a substance is, the less its able to neutralize further acid addition without dropping the pH further. The total alkalinity is presented in Figure 4-19 for the thermophilic digester feed and effluent. The alkalinity in the digesters is much higher than in the influent because alkalinity is generated by AD and all the feedstock co-substrates are more acidic than the digester effluents with the exception of stripped HTLaq which is added in small volumes (co-substrate pHs are given in Table 4-10). It is understandable that pH increases during AD as VFAs are broken down. 63 Figure 4-19: Total alkalinity concentration in the thermophilic anaerobic digester influent and effluent. The alkalinity to and from most of the digesters remained relatively consistent after the increase in HTLaq contribution. There was a decrease in effluent alkalinity in the thermophilic HTLaq digester after HTLaq contribution to OLR was changed from 5% to 13% of COD. This drop in digester alkalinity is likely due to the VFAs not being reduced, leaving the digester more acidic. The results for mesophilic digesters are displayed in Figure 4-20. Figure 4-20: Total alkalinity concentration in the mesophilic anaerobic digester influent and effluent. Under mesophilic conditions, all digesters had similar influent and effluent alkalinity. Influent alkalinity is slightly higher in digesters fed pretreated HTLaq compared to non-010002000300040005000600070000%HTLaq 5%HTLaq 5%HTLaqStrip 13%HTLaq 12%HTLaqStripAlkalinity (mg CaCO3/L)Feed Effluent010002000300040005000600070000%HTLaq 13%HTLaq 13%HTLaqStrip 24%HTLaq 22%HTLaqStripAlkalinity (mg CaCO3/L)Feed Effluent64 pretreated. This result is linked to stripped HTLaq having the highest alkalinity of the co-substrates. 4.2.7 Total Phenolic Compounds Concentration The concentration of phenolic compounds found in the thermophilic digester influent and effluent is displayed in Figure 4-21. Phenolic compounds have long been known to be inhibitory to AD with inhibition seen when phenolics reach 1 g/L in the digester (He, et al., 2022). Another study by Madigou et. al. (2016), found that the inhibition threshold for phenolic compounds could be increased to 1.9 g/L if digester microbiota were given time to acclimatize to the increase level of phenolics. Phenolics are a grouping of compounds, and the quantity of individual compounds can have an effect on when inhibition occurs. Fedorak and Hrudey (1984) found that 2,5-, 3,4- and 3,5-dimethylphenols were inhibitory at 500 mg/L but cresols were inhibitory at 1,000 mg/L. Figure 4-21: Total phenolics concentration in the thermophilic anaerobic digester influent and effluent. At higher percentages of HTLaq in the feedstock, there is an increase in phenolic compounds in the feed and effluent of digesters fed with HTLaq compared to control. In the digesters fed with pretreated HTLaq, the feed has less phenolic compounds than non-pretreated 0501001502002500%HTLaq 5%HTLaq 5%HTLaqStrip 13%HTLaq 12%HTLaqStripTotal Phenolics (mg GAE/L)FeedEffluent65 HTLaq but the effluent concentration of phenolics is not much different than non-pretreated HTLaq digesters. The concentration of phenolic compounds found in the mesophilic digester influent and effluent is displayed in Figure 4-22. Figure 4-22: Total phenolics concentration in the mesophilic anaerobic digester influent and effluent. The mesophilic control (0%HTLaq) and 24%HTLaq digesters seemed to provide a reduction of phenolic compounds. The control digester had a lower effluent phenolics concentration than the other digesters, while the 24%HTLaq digester had increased phenolic compounds in the influent but an effluent concentration of phenolic compounds similar to the other mesophilic digesters. None of the digesters at either temperature had total phenolics concentrations in their influent or effluent above any of the inhibitory levels found in the review of literature. Given the long acclimatization period of the digesters being fed HTLaq and the relatively low phenolics concentration compared to inhibitory levels, it is unlikely that the inhibition in the thermophilic HTLaq digesters was a result of inhibition from phenolic compounds. 0501001502002503000%HTLaq 13%HTLaq 13%HTLaqStrip 24%HTLaq 22%HTLaqStripTotal Phenolics (mg GAE/L) FeedEffluent66 4.2.8 Total Solids and Volatile Solids The percentage of total solids in the thermophilic anaerobic digester influent and effluent is displayed in Figure 4-23 below. Under initial conditions the TS removal of the three digesters was between 44 and 51%, with the control digester having the highest average removal and the digester fed stripped HTLaq having the lowest. As the volume of MS decreased, centrate increased and HTLaq increased in the feedstocks, the influent solids were decreased (as expected based on Table 4-10). Though the influent TS in the thermophilic HTLaq digester decreased as the HTLaq contribution increased, the effluent TS is consistent between HTLaq contributions leaving the digester effluent with around 66% of the initial solids (only an average of 34% TS removal). The thermophilic digester being fed stripped HTLaq however, had a slight increase in TS removal with 53% solids removed at an HTLaqStrip contribution of 12%. The percentage of total solids in the mesophilic digesters is presented in Figure 4-24. 012340%HTLaq 5%HTLaq 5%HTLaqStrip 13%HTLaq 12%HTLaqStripTS (% wt/wt)FeedEffluentFigure 4-23: Total solids concentration in the thermophilic anaerobic digester influent and effluent. 67 Figure 4-24: Total solids concentration in the mesophilic anaerobic digester influent and effluent. The mesophilic digesters had average TS removals between 38 and 49% with the highest removal achieved by the control digesters and the lowest removal seen in the digester fed 22%HTLaqStrip. The solids present in thermophilic AD feedstock and effluent are mostly volatile organics so the graph of VS (Figure 4-25) of the digesters is similar to the TS graph presented earlier in this section. Figure 4-25: Volatile solids concentration in the thermophilic anaerobic digester influent and effluent. Most of the digesters maintained stable reduction of VS during the switch to the final feeding condition, except for the thermophilic HTLaq digester. The VS in the feedstock to this 012340%HTLaq 13%HTLaq 13%HTLaqStrip 24%HTLaq 22%HTLaqStripTS (% wt/wt)FeedEffluent012340%HTLaq 5%HTLaq 5%HTLaqStrip 13%HTLaq 12%HTLaqStripVS (% wt/wt)FeedEffluent68 digester decreased from 2.5 ± 0.2 to 1.8 ± 0.1% VS (wt/wt) when HTLaq increased from 5 to 13%, yet the effluents were very similar at 1.1 ± 0.2% VS (wt/wt). This confirms the inhibition of cultures beneficial for organic solids destruction in this thermophilic digester. T-tests were performed to determine if there was a statistically significant difference between the mean VS removal of digester fed non-pretreated and pretreated HTLaq. For the thermophilic digesters, the t-critical value for a 95% confidence interval was 1.78 and the t-value was -8.8 for the VS removal values of 13%HTLaq and 12%HTLaqStrip digesters at steady-state. Since the absolute value of the t-value was greater than that of t-critical, it can be concluded that the means are significantly different and pretreatment of HTLaq increased VS removal in the thermophilic AD test. The percentage of volatile solids in the mesophilic digesters is presented in Figure 4-26. Figure 4-26: Volatile solids concentration in the mesophilic anaerobic digester influent and effluent. Under mesophilic conditions, the mean VS removal of digesters was between 48 and 60%, with the control having the highest removal and the digesters fed non-pretreated HTLaq having the lowest. T-tests were also performed on the final conditions of mesophilic digesters fed non-pretreated and pretreated HTLaq. It was determined that there was no statistically significant difference between the mean VS removal of 24%HTLaq and 22%HTLaqStrip 012340%HTLaq 13%HTLaq 13%HTLaqStrip 24%HTLaq 22%HTLaqStripVS (% wt/wt)FeedEffluent69 digesters, so no conclusions can be made on the effect of HTLaq pretreatment of VS removal during mesophilic co-digestion. 4.2.9 Heavy Metals A third-party laboratory completed inductively coupled plasma mass spectrometry analysis of 64 different metals. The results provided either gave the concentration of that metal for each of the duplicate samples or indicated that the concentration of the metal was below the provided detection limit. The metals that are relevant to land application regulations for the Metro Vancouver area are presented in Table 4-13. In the case that the metal was not present in detectable concentrations, the detection limit of the analysis for that metal was given as a ceiling value under concentration. The maximum allowable metals concentration for Class B biosolids is indicated by British Columbia’s Organic Matter Recycling Regulation (British Columbia, 2018). Class A biosolids level is dictated by the Government of Canada’s Safety standards for fertilizer and supplements for a land application rate of 4400 kg/ha per year (Canadian Food Inspection Agency, 2023). In the initial round of metals testing (March 12, 2023 effluent), the Thermo0%HTLaq, Thermo5%HTLaqStrip, Meso13%HTLaq and Meso13%HTLaqStrip digesters would only be eligible for Class B biosolids because of their specific copper concentrations. However, the thermophilic 0%HTLaq digester would not be eligible for Class B biosolids status due to the specific zinc concentration exceeding the threshold by 40 ug/g dry weight. During the second round of testing (June 8, 2023 effluent) all digesters met Class A biosolids requirements. This shows the variation in biosolids quality depending on the feedstock characterization. Both sets of samples provide no indication that the HTLaq co-digestion would increase the metals concentration in the biosolids. This is consistent with literature on HTL because most of the metals (>69%) in the HTL feed are captured in hydrochar (Liu, et al., 2023). The study found that HTLaq had a lower metals concentration than the MS it was derived from for all metals 70 relevant to biosolids regulations for British Columbia. Furthermore, a HTLaq co-digestion study found the trend that increasing the HTLaq contribution to the digester feed resulted decreased metals concentration in the digester effluent (Azarmina, 2023). It can therefore be concluded that co-digestion with HTLaq will decrease metals concentration in the resulting biosolids, by diluting the metals concentration in MS. 71 Table 4-13: Heavy metal concentrations in anaerobic digester effluents compared to biosolids regulations. Label TS (dry g/kg) Chromium (ug/g dry) Cobalt (ug/g dry) Nickel (ug/g dry) Copper (ug/g dry) Zinc (ug/g dry) Arsenic (ug/g dry) Selenium (ug/g dry) Molybdenum (ug/g dry) Cadmium (ug/g dry) Mercury (ug/g dry) Lead (ug/g dry) 2023-03-12 Effluent Thermo 0%HTLaq 14.7 97 ± 5 < 7 * 36 ± 3 822 ± 16 1,890 ± 22 < 15 < 19 18 ± 1 3 ± 0.1 < 17 44 ± 1 Thermo 5%HTLaq 16.6 69 ± 0.2 < 6 28 ± 1 622 ± 24 1,310 ± 19 < 13 < 18 < 16 2 ± 0.3 < 16 41 ± 12 Thermo 5%HTLaq Stripped 13.9 99 ± 7 5 ± 1 34 ± 1 837 ± 31 1,850 ± 99 < 16 < 21 < 18 4 ± 0.3 < 18 45 ± 2 Meso 0%HTLaq 15.7 90 ± 4 < 7 33 ± 1 839 ± 11 1,790 ± 18 < 14 < 18 < 16 3 ± 0.1 < 16 42 ± 2 Meso 13%HTLaq 11.5 75 ± 1 < 9 25 ± 0.3 825 ± 8 1,350 ± 1 < 19 < 25 < 22 2 ± 0.2 < 22 29 ± 1 Meso 13%HTLaq Stripped 11.9 70 ± 1 < 9 22 ± 1 821 ± 14 1,280 ± 10 < 18 < 24 < 21 2 ± 0.2 < 21 29 ± 0.3 2023-06-08 Effluent Thermo 0%HTLaq 17.5 280 ± 217 < 6 134 ± 112 646 ± 1 1,110 ± 53 < 12 < 16 < 15 2 ± 0.3 < 15 25 ± 1 Thermo 13%HTLaq 18.9 46 ± 2 < 6 14 ± 1 507 ± 58 733 ± 79 < 11 < 15 < 13 1 ± 0.2 < 13 16 ± 2 Thermo 12%HTLaq Stripped 11.6 63 ± 4 < 9 22 ± 4 647 ± 0.3 1,050 ± 81 < 19 < 24 < 22 2 ± 0.2 < 22 21 ± 3 Meso 0%HTLaq 19.3 67 ± 2 < 5 23 ± 1 616 ± 2 1,240 ± 14 < 11 < 15 < 13 2 ± 0.04 < 13 27 ± 1 Meso 24%HTLaq 9.19 71 ± 1 < 11 22 ± 1 746 ± 40 977 ± 37 < 24 < 31 < 28 2 ± 0.2 < 28 25 ± 5 Meso 22%HTLaq Stripped 10.6 59 ± 4 < 10 15 ± 0.4 646 ± 19 741 ± 20 < 21 < 27 < 24 < 1 < 24 16 ± 3 Class A biosolids 1060 151 181 757 1,868 75 14 20 20 5 505 Class B biosolids 1060 150 180 2200 1,850 75 14 20 20 15 500 * “<” indicates that the sample was below the detection limit for that metal on both replicates. 72 4.2.10 Specific Capillary Suction Time The specific capillary suction time is charted in Figure 4-27 to compare dewaterability of thermophilic digestate samples. For thermophilic digesters, HTLaq addition to feedstock does not significantly impact CST and therefore dewaterability because the water separates from solids in the same time frame. The CST was also measured after HTLaq contribution was increased and steady-state was reached at the final condition. Due to the variability in this test, not many conclusions could be drawn about dewaterability under co-digestion conditions. The specific capillary suction time is charted in to compare dewaterability in mesophilic digesters in Figure 4-28. In the control digesters, mesophilic had a longer CST meaning it would be more difficult to dewater than thermophilic. This trend was also seen in the mesophilic HTLaq digester compared to the thermophilic HTLaq digester. This result is unexpected because in wastewater treatment plant operation it is commonly seen that mesophilic digesters dewater more readily than thermophilic digesters. The CST analysis was completed on other thermophilic and mesophilic digesters in the laboratory and those digesters followed the trend of water leaving the mesophilic digesters 01002003004005006007008000%HTLaq 5%HTLaq 5%HTLaqStrip 13%HTLaq 12%HTLaqStripSpecific CST (s/%TS)Figure 4-27: Specific capillary suction time of the thermophilic anaerobic digester effluent. 73 quicker (lower CST). The CST analysis was replicated on multiple days and with different instruments, and it seems that the digesters used in this research differ from typical dewaterability trends but the cause for this is unknown. Figure 4-28: Specific capillary suction time of the mesophilic anaerobic digester effluent. 4.2.11 Fecal Coliform Density Fecal coliforms are important for determining the biosolids class for land application of the digester solids. Digestate fecal coliform densities are compared to regulatory levels in Figure 4-29. Specific fecal coliform levels are required to be below 2,000,000 MPN/g TS to classify for Class B biosolids and below 1,000 MPN/g TS for Class A biosolids in British Columbia (Goverment of British Columbia, 2022). All thermophilic digesters have specific fecal coliforms in the Class A biosolids range, with the addition of HTLaq as a co-substrate not significantly impacting the number of colonies in the digestate sample. In the mesophilic condition, the control digester had higher fecal coliform levels than the test was able to detect for all three replicates even with a 10 times dilution applied. The test was only able to indicate that the fecal coliforms were greater than 242,000 per 100 mL sample. With the digester TS given in Section 4.2.7 and a dilution of 10 (required to get triplicate 100 mL samples from the small digesters) the specific fecal coliforms are > 1,450 MPN/g TS. This 01002003004005006007008009000%HTLaq 13%HTLaq 13%HTLaqStrip 24%HTLaq 22%HTLaqStripSpecific CST (s/%TS)74 disqualifies the mesophilic control digester from being a Class A biosolids but is inconclusive of if it would be categorized as Class B or not. In the mesophilic digesters, the effluent coliforms decreased through the addition of HTLaq in the feedstock and further reduced by pretreatment of the HTLaq. Figure 4-29: Specific fecal coliform concentration in the anaerobic digester effluent. 4.2.12 Nitrogen Heterocyclic Compounds The variety of nitrogen heterocyclic compounds that could be detected by the GC mass spectrometer in the effluent digester samples are displayed in Table 4-14. Very little nitrogen heterocyclic compounds were detected in the control digesters, with only less than 0.71 ± 0.01 mg/L of indole presenting in the scan. Digesters being fed with HTLaq as one of the co-substrates had more detectible compounds including pyrazines and pyridines. In thermophilic digesters, the highest detected concentration was methyl-pyrazine at 32.5 ± 0.05 and 26.1 ± 0.16 mg/L in non-pretreated and pretreated HTLaq fed digesters, respectively. In the mesophilic digesters, 3-methyl-cyclopentanone had the highest concentration (18.9 ± 0.02 mg/L) in 24%HTLaq digester and 2,3-dimethyl-pyrazine had the 1101001,00010,000100,0001,000,00010,000,0000%HTLaq 5%HTLaq 5%HTLaq Stripped 0%HTLaq 13%HTLaq 13%HTLaq StrippedSpecific Fecal Coliforms (MPN/gTS)Thermophilic Digesters Mesophilic Digesters Class B Biosolids Class A BiosolidsAD00 was over 1,450 MPN/gTSfor all three replicates75 highest concentration (4.49 ± 0.22 mg/L) in 22%HTLaqStrip digester. For all compounds the effluent concentration was lower in the digesters where HTLaq was pretreated before feeding. Table 4-14: Nitrogen heterocyclic compound concentration in the anaerobic digester effluent. Compound Name Concentration in Thermophilic Digester Effluent (mg/L) Concentration in Mesophilic Digester Effluent (mg/L) 0%HTLaq 13%HTLaq 12%HTLaqStrip 0%HTLaq 24%HTLaq 22%HTLaqStrip Pyrazines methyl-pyrazine ND* 32.5 ± 0.05 26.1 ± 0.16 ND 0.35 ± 0.06 0.14 ± 0.14 ethyl-pyrazine ND 5.13 ± 0.15 2.95 ± 0.22 ND 0.76 ± 0.45 0.19 ± 0.03 2,3-dimethyl-pyrazine ND 4.16 ± 0.06 3.57 ± 0.26 ND 6.46 ± 0.09 4.49 ± 0.22 trimethyl-pyrazine ND 2.17 ± 0.16 1.68 ± 0.24 ND 3.71 ± 0.21 2.56 ± 0.40 2,3-diethyl-5-methyl-pyrazine ND ND ND ND 0.07 ± 0.06 0.048 ± 0.001 Pyridines 2-methyl-pyridine ND 1.02 ± 0.10 0.25 ± 0.04 ND 5.20 ± 1.21 0.21 ± 0.07 2-ethyl-pyridine ND 0.12 ± 0.01 0.01 ± 0.01 ND 0.69 ± 0.05 0.005 ± 0.009 5-ethyl-2-methyl-pyridine ND 0.133 ± 0.003 0.001 ± 0.001 ND 0.58 ± 0.07 0.032 ± 0.0007 Other cyclohexanone ND 0.13 ± 0.02 0.048 ± 0.004 ND 0.55 ± 0.28 0.07 ± 0.03 3-methyl-cyclopentanone ND 4.11 ± 0.10 1.34 ± 0.09 ND 18.9 ± 0.02 2.81 ± 0.20 2,6-lutidine ND 0.27 ± 0.01 0.062 ± 0.002 ND 1.47 ± 0.08 0.03 ± 0.02 4-phenyl-2-butanone ND 0.03 ± 0.008 0.019 ± 0.004 ND ND ND indole 0.71 ± 0.01 1.93 ± 0.08 1.00 ± 0.14 0.78 ± 0.01 0.40 ± 0.06 0.003 ± 0.001 *ND = not detected (below the instruments detection limit) The non-pretreated and pretreated HTLaq of the final HTL run for semi-continuous flow digester feeding was tested for nitrogen heterocyclic compounds and the results are displayed on Table 4-15. The stripped HTLaq results are based on the Run 4 operational parameters (85°C, 9.8, 0.5 L/min). For every compound, the pretreated HTLaq sample had a lower concentration than the non-pretreated sample. Whenever possible, when at least two replicates from both HTLaq and stripped HTLaq samples had detectable concentrations, a t-test was performed to determine if the mean concentration of the specific compound before and after 76 ammonia stripping was statistically different (at 95% confidence). In all cases, the t-tests revealed that there was a significant removal of the compound during ammonia stripping. The compound with the highest concentration in both HTLaq samples was methyl-pyrazine. Table 4-15: Nitrogen heterocyclic compound concentrations in non-pretreated and pretreated HTLaq. Compound Name Concentration in HTLaq (mg/L) Concentration in Stripped HTLaq (mg/L) Significant Removal Pyrazines methyl-pyrazine 903 ± 0.5 641 ± 2 Yes (p* = 0.03) ethyl-pyrazine 105 ± 0.6 44.4 ± 0.01 Yes (p = 0.0001) 2,3-dimethyl-pyrazine 61.9 ± 3.4 59.1 ± 0.8 Yes (p = 0.0008) trimethyl-pyrazine 31.1 ± 0.2 23.9 ± 1.7 Yes (p = 0.009) 2,3-diethyl-5-methyl-pyrazine ND ND Pyridines 2-methyl-pyridine 34.6 ± 0.4 1.34 ± 0.04 Yes (p = 0.004) 2-ethyl-pyridine 3.21 ± 0.01 ND 5-ethyl-2-methyl-pyridine 9.87 ± 0.2 ND Other cyclohexanone 13.2 ± 0.1 0.602 3-methyl-cyclopentanone 33.2 ± 1.0 ND 2,6-lutidine 15.7 ± 0.1 ND 4-phenyl-2-butanone ND ND Indole ND ND *p in a t-test indicates probability that the difference between the means shown in two columns is by chance and there is no significant difference between the populations. If the p-value is less than 0.05, then there is 95% confidence that the mean values in columns are different. The clear reduction in nitrogen heterocyclic compounds during ammonia stripping and reduced presence of these compounds in digesters fed pretreated versus non-pretreated HTLaq make a strong case that these compounds were the cause of inhibition in the thermophilic 13%HTLaq digester while the 12%HTLaqStrip digester was not inhibited. This could indicate that ammonia stripping pretreatment could enhance thermophilic AD in substrates high in nitrogen heterocyclic compounds. 77 Chapter 5: Conclusions The research in this thesis was working towards the goal of designing more sustainable WWTPs by making HTL a feasible new solids processing and energy recovery solution. The research focused on the treatment of HTLaq, as the treatment/disposal of this byproduct is an obstacle to bringing this technology full-scale. The disposal pathway investigated was co-digesting HTLaq with mixed sludge after a pretreatment of ammonia stripping and acid adsorption, for fertilizer generation. In the first part of the study focusing on HTLaq pretreatment optimization, seven runs of ammonia stripping were completed, and characterization was completed to identifying the components of HTLaq removed focusing on TAN. The initial TAN concentration of the HTLaq was 4,580 ± 15 mg/L and was reduced to 32 ± 6 mg/L in the most successful run (85°C, pH of 10.3, 0.5 L/min). The seven runs suggested that pH has the largest impact on ammonia removal and temperature the smallest (Section 4.1.2). In most runs, phenolic compounds were also removed, with Run 5 (85°C, pH of 9.3, 0.5 L/min) removing the most phenolics at 32 ± 1%. Though removal of inhibitory compounds is the goal of ammonia stripping, given the chosen disposal pathway of AD, it is ideal to have as many VFAs remain in the stripped effluent to be converted into methane and not starve the microorganisms. Run 5 had the least VFA removal with 38 ± 1% of VFAs leaving the aqueous. Since ammonia removal was the main objective during the experimental design, 85°C, a pH of 10.3, and 0.5 L/min flowrate were chosen as the ideal conditions. Once all characterisation was done and accounting for the cost and operation complexity of large amounts of chemical addition Run 4 (85°C, pH of 9.8, 0.5 L/min) was chosen to test biodegradability through anaerobic co-digestion. Ammonia stripping was chosen for pretreatment in part because of the potential for ammonia recovery through acid adsorption and the potential of reducing use of synthetic 78 fertilizers. The purity of ammonium sulphate salts produced in all runs was between 85 and 98% with Run 4 achieving the most recovery at 95 ± 1% of digester ammonia transferred to salt. In the second part of the study, biodegradability of HTLaq was assessed. Finding a suitable treatment stream for the HTLaq is barrier in bringing HTL to full scale at WWTPs and any energy or nutrient recovery benefits will not be realized if HTLaq can not be pretreated to a quality not inhibitory to biological treatment units already onsite. The biodegradability of pretreated HTLaq was compared to non-pretreated HTLaq through bench-scale anaerobic co-digestion with mixed sludge and dewatering centrate. This was to determine if pretreatment could increase the contribution of HTLaq to OLR that an anaerobic digester could operate at. Under thermophilic conditions, the digester fed pretreated HTLaq showed less inhibition as the percentage of HTLaq fed increased (from 5% to 12% of total COD fed). Under mesophilic conditions, neither the non-pretreated nor pretreated HTLaq digesters showed inhibition even at 24% and 22% COD from HTLaq, indicating higher tolerance than their thermophilic counterparts. If both digesters are uninhibited, then the pretreatment of HTLaq slightly decreases the amount of biogas produced as ammonia stripping reduces the COD. Increasing the contribution of HTLaq that can be co-digested increases the feasibility of implementing this system at WWTPs for pilot- or full-scale operation. Successful implementing of HTL for municipal WWTPs could lead to a future with more green energy and less municipal biosolids sent to the environment. Beyond energy recovery, WWTP operators are very concerned with process changes that impact the dewaterability of biosolids or that could change the biosolids land application classification. For biosolids classification concerns, heavy metals and fecal coliform density were tested on the effluent of all digesters in this study. HTLaq is lower in heavy metals than MS so, heavy metal concentrations were lower in digesters fed HTLaq compared to control digesters. The extreme conditions in HTL result in HTLaq not being biologically active and 79 resulted in the mesophilic digesters being fed HTLaq having lower fecal coliform densities than the control digesters. All thermophilic digesters had very low fecal coliform densities, which is expected at the higher temperature. Pretreatment of HTLaq did not impact the results for either of these biosolids parameters. The capillary suction time tests did not suggest that co-digestion with non-pretreated or pretreated HTLaq would reduce the dewaterability of digester effluent. Overall, the results of these biosolids tests did not indicate that pretreatment with ammonia stripping would improve the biosolids production or classification but also did not identify any issues with co-digestion of non-pretreated HTLaq. After observing that the ammonia levels in the inhibited thermophilic HTLaq digester were consistent to the uninhibited digesters, it could be concluded that ammonia levels were not the limiting factor in increasing HTLaq contribution to co-digestion. The thermophilic digester fed pretreated HTLaq however did have better performance at 13% COD supplied from HTLaq. Another compound, potentially a phenolic or nitrogen heterocyclic compound, being removed during the ammonia stripping process could be inhibitory to AD. The concentration of all nitrogen heterocyclic compounds detected in HTLaq were decreased during stripping and the concentration for almost all tested compounds was lower in the effluent of digesters that were fed the pretreated HTLaq compared to the non-pretreated. For future operation, the optimal conditions for ammonia stripping of HTLaq should be switched to Run 5 conditions (85°C, pH of 9.3, 0.5 L/min) for having the highest removal of phenolics, lowest removal of VFAs and lowest demand for chemicals. The downside of this condition would be the reduction in nutrient recovery and production of value-added product. The ammonium sulphate salt produced Run 5 was the second most pure at 98.0 ± 0.05% but the recovery was still low at 73.9 ± 0.04% due to the Run 5 having the lowest ammonia removal of any run at 79.5 ± 0.1%. 80 5.1 Limitations and Future Research A limitation to this work on HTLaq pretreatment is that the composition of HTLaq is variable dependent on the feedstock characteristics. Though trends can be identified for the effect of pH, temperature, and airflow on the efficiency of ammonia stripping, the removal efficiency or effluent concentration cannot be guaranteed in this complex aqueous product. With the current level of HTL literature available, there would need to be system-specific assessments performed prior to constructing even an HTL pilot system at any WWTP. More research is required into the contents of HTLaq from different feedstocks and varying HTL conditions to determine which components of HTLaq are inhibitory to biological processes. If the components of HTLaq which are most inhibitory to WWTP processes can be identified, then HTL conditions could be chosen that produce HTLaq that can be safely disposed of or at least identify which pretreatment methods are required depending on the chosen disposal pathway. Nitrogen heterocyclic compounds were found to be reduced during ammonia stripping, but further work could be done on the optimum stripping conditions to remove the most inhibitory compounds. Future work is also required on the effects of HTLaq on the liquid train of WWTPs to understand if disposal to the headworks is suitable in WWTPs without AD. 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Figure A-2: Total phenolics colorimetric test calibration curve. y = 92.219x + 0.3896R² = 0.996801002003004005006007008000.00 2.00 4.00 6.00 8.00Absorbance Concentration (mg/L)y = 5.2564x + 115.23R² = 0.99420500100015002000250030000 100 200 300 400 500 600Absorbance Concentration (mg/L)87 Figure A-3: Chemical oxygen demand colorimetric test calibration curve. y = 0.4308x + 1.3333R² = 0.99980501001502002503003500 200 400 600 800Absorbance Concentration (mg/L)88 Appendix B Scanning Electron Microscope Salt Imaging Figure B-1: Run 1 salt at 1,000 times magnification. Figure B-2: Run 2 salt at 1,000 times magnification. 89 Figure B-4: Run 4 salt at 1,000 times magnification. Figure B-3: Run 3 salt at 1,000 times magnification. 90 Figure B-5: Run 5 salt at 1,000 times magnification. Figure B-6: Run 6 salt at 1,000 times magnification. 91 Figure B-7: Run 7 salt at 1,000 times magnification. 92 Appendix C Energy Dispersive Spectroscopy Figure C-3: Run 3 EDS map and element fractions. Figure C-2: Run 2 EDS map and element fractions. Figure C-1: Run 1 EDS map and element fractions. 93 Figure C-5: Run 5 EDS map and element fractions. Figure C-4: Run 4 EDS map and element fractions. Figure C-6: Run 6 EDS map and element fractions. 94 Figure C-7: Run 7 EDS map and element fractions. 95 Appendix D Sample Statistical Analysis Tables T-tests are statistical analysis tool suitable for comparing two data sets, and determining if the means are significantly different. In this research t-tests were used to determine if anaerobic digesters fed with untreated or pretreated HTLaq had significantly different COD and VS removal. T-tests were also used to determine if ammonia stripping pretreatment of HTLaq significantly reduced the concentration of a variety of nitrogen heterocyclic compounds. Summary tables of the parameters required to run a t-test and the resulting values are displayed in Table D-1 and Table D-2 for COD removal from thermophilic and mesophilic respectively. Table D-1: T-test summary table for analysis of significant mean difference between the COD removal of untreated and pretreated HTLaq fed thermophilic digesters at 95% confidence (α = 0.05). Thermophilic 13%HTLaq Thermophilic 12%HTLaqStrip Mean 0.216105 0.542729 Variance 0.007399 0.011357 Observations 9 10 Hypothesized Mean Difference 0 Degrees of Freedom 17 t-value -7.38169 t-critical (Two-Tail) 2.109816 Table D-2: T-test summary table for analysis of significant mean difference between the COD removal of untreated and pretreated HTLaq fed mesophilic digesters at 95% confidence (α = 0.05). Mesophilic 24%HTLaq Mesophilic 22%HTLaqStrip Mean 0.521863 0.51398 Variance 0.001928 0.012538 Observations 8 8 Hypothesized Mean Difference 0 Degrees of Freedom 9 t-value 0.185385 t-critical (Two-Tail) 2.262157
Thesis/Dissertation
eng
Civil Engineering
Vancouver : University of British Columbia Library
Attribution-NonCommercial-NoDerivatives 4.0 International
Graduate
Eskicioglu, Cigdem
Ammonia stripping of hydrothermal liquefaction aqueous phase from municipal sludge for anaerobic co-digestion pretreatment and ammonia recovery
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