Climate change, water stress, and rapid population growth have increased the need to manage water resources through innovative sustainable technologies. Decentralized systems such as membrane treatment trains have become increasingly important to provide high volumes of potable water to millions of people. Pressure-driven membrane systems have dominated separation processes due to their low cost, small footprint, ease of operation, and high permeate quality. Conventionally, pressure-driven membranes are classified into microfiltration (MF), ultrafiltration (UF), nanofiltration (NF), and reverse osmosis (RO). MF and UF membranes operate under low pressure (< 7 bar, <~100 psi). They can separate a variety of large particles such as bacteria, natural organic matter, suspended solids, and colloids. In contrast, NF and RO membranes are more energy-intense due to operating at high pressures (7 – 80 bar, ~100 – 1200 psi) and can remove small molecules such as ions, pharmaceuticals, and heavy metals. Fouling is a primary challenge with membranes that compromises the membrane performance, increases energy consumption, and reduces the membrane lifetime. Many strategies are used to address fouling, such as pre-treatment (pH adjustment, screening, coagulation), membrane modification (chemical and morphological properties), and membrane cleaning (physical, chemical). However, such strategies increase operational expenditures, produce waste products that can impact the environment, and negatively impact membrane lifetimes. Recently, electrically conductive membranes (ECMs) have been introduced to address the challenges with traditional membranes. They contain conductive surfaces that offer self-cleaning and antifouling properties across the surface in response to electrical potential externally applied to them. ECMs are advantageous as compared to traditional membranes because (a) they are more effective in treating foulants as they target foulants at the membrane/solvent interface, (b) they are more economical and environmentally friendly as they reduce the need for chemical consumption, and (c) they can be responsive to fouling conditions as their antifouling mechanisms can be easily manipulated by changing the applied current type (positive, negative, direct current, alternating current) to match the foulant. ECMs have been formed from all categories of membranes (MF, UF, NF, MD, FO, and RO) with a range of applications. Despite the remarkable progress in demonstrating their excellent antifouling performance, there are many hurdles to overcome before they can be commercialized. Two of these are (a) a fundamental understanding of their underlying mechanisms, (b) surface materials that can withstand extreme chemical and electrical conditions. In this work, we have comprehensively discussed antifouling mechanisms with respect to surface polarization and elaborated on the impact of electrically-induced mechanisms on four major fouling categories. i.e., biofouling, organic fouling, mineral scaling, and oil wetting. In addition, we characterized surface properties of a common electrically conductive composite membrane composed of carbon nanotubes (CNTs) and polyvinyl alcohol (PVA). We then investigated the impact of cross-linkers in CNT/PVA network on transmembrane flux, electrical conductivity, hydrophilicity, and surface roughness. In addition, we proposed standard, practical, and straightforward methodologies to detect and quantify the electrochemical, physical, and mechanical stability of ECMs, using chronoamperometry and cyclic voltammetry, an evaluation of polymer leaching from membranes, and micro mechanical scratch testing, respectively. Our methods can readily be extended to all membrane-based separation processes and different membrane materials (carbonaceous materials, ceramics, metal-based, and polymers). To demonstrate the antifouling properties of ECMs, we challenged ECMs with mixed-bacterial cultures in a flow-through system. Although ECMs showed high rejection, comparable flux, and excellent self-cleaning performance under application of electrical potential, understanding the relationship between applied electrical currents and antifouling mechanisms demands a well-controlled investigation. To this end, we quantified the impact of electrochemically-induced acidic conditions, alkaline conditions, and H2O2 concentration on model bacteria, Escherichia Coli. We first quantified the electrochemical potential of CNT-based ECMs in generating stressors such as protons, hydroxyl ions, and H2O2, under a range of applied electrical currents (± 0-150 mA). Next, these individual stressors with identical magnitude were imposed on E. Coli cells and biofilms in batch and flow-through systems, respectively. This thesis guides researchers to understand the underlying antifouling mechanisms associated with ECMs, how to match the mechanisms to the application of ECMs, and offers benchmarks for making practical ECMs. / Thesis / Doctor of Philosophy (PhD)

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Organelles are crucial compartments in living cells for carrying out biological events, and cells normally employ compartmentalization to spatially manage their cellular material transport, signaling, and metabolic processes. Engineering biomimetic nanoreactors to replicate biological processes has attracted a lot of interest in recent years. pH-responsive and photo-crosslinked polymersomes, for example, as synthetic vesicles, have tuable membrane permeability and mechanical stability, and may be utilized to build artificial organelles by encapsulating bioactive molecules in their cavity. Most existing reports of stimuli-responsive polymersomes for enzymatic cascade reactions are based on a simple mix of two types of polymersomes loaded with different enzymes, whereas cells process multi-enzyme catalytic systems in which intracellular biological reactions are carried out by combining two or more enzymes in the same organelle. In fact, the most of sophisticated biological functions and features of cells are based on self-organization, the coordination and connection between their cell organelles determines their key functions. Therefore, spatially ordered and controllable self-assembly of polymersomes to construct clusters to simulate complex intracellular biological functions has attracted widespread attention. Here, a simple one-step copper-free click strategy is present to crosslink nanoscale pH-responsive and photo-crosslinked polymersomes (less than 100 nm) to micron-level clusters (more than 90% in 0.5-2 µm range). Various influencing factors in the clustering process and subsequent purification methods were studied to obtain optimal clustered polymeric vesicles. Even if co-clustering the separately loaded polymeric vesicles with different enzymes (glucose oxidase and myoglobin), the overall permeability of the clusters can still be regulated through tuning the pH values on demand. Compared with the conventional enzyme cascade reaction through simple blending polymersomes, the rate of enzymatic cascade reaction increased significantly due to the interconnected complex microstructure established. The connection of catalytic nano-compartments into clusters confining different enzymes of a cascade reaction provide an excellent platform for the development of artificial systems mimicking natural organelles or cells. Although pH-responsive polymersomes present a good membrane permeability in response to alternate pH values and good stability in swelling/shrinking behavior owing to the photo-crosslinked membrane, they are still insufficient to simulate more complex biological activity. The intrinsic pH values for molecules transport are always acidic, whereas the majority of cellular action occurs at physiological pH levels. Due to the closed membrane, the enzyme reaction cannot be carried out efficiently under simulated physiological conditions (pH 7.4). To generate a permeable membrane at a physiological pH value, a new stimulus element must be introduced into existing polymersomes. To self-assemble pH- and light-responsive as well as photo-crosslinked polymersomes, a single azobenzene unit is used as a junction molecule between the hydrophilic and hydrophobic segments of block copolymer. To compare light utilization, block copolymers based on donor-acceptor-substituted azobenzene junction and ether-substituted azobenzene junction were prepared. Besides, the photo-isomerization of novel macroinitiators, block copolymers and polymersomes was also studied to get responsive wavelength ranges of light. The dye release experiments proven the hydrophobic dye on the membrane of polymersomes can release from the membrane under light irradiation. Despite the fact that blue light (400-500 nm) has a higher release efficiency than UV light (365 nm) and ether-substituted azobenzene polymersomes have a slightly higher release efficiency than donor-acceptor-substituted azobenzene polymersomes, the mechanism is still unknown due to the different power of light sources. Furthermore, based on the results of light-driven enzyme reaction, more experiments are required to confirm the light-induced membrane permeability, such as photo-oxidation of substrates and photo-induced deactivation of enzyme. But in general, photo-induced membrane disorder does squeeze the tiny cargo out of the membrane. The single azobenzene unit as the linkage between hydrophilic and hydrophobic block induced membrane pertubation proposes a novel concept in which a trace of azobenzene unit can affect cargo mobility on the membrane of polymersomes and even propagate the fluidity of water molecules to the entire membrane, thereby resulting in membrane permeability. This approach offers a unique framework for the development of biomimetic behaviors under physiological simulated conditions.:Part I Fundamentals 1 Theoretical Background 1.1 Polymersomes 1.1.1 Polymersomes Formation 1.1.2 Self-Assembly Principles of Amphiphilic Block Copolymers (BCPs) 1.1.3 Preparation Methods of Polymersomes 1.1.4 Cargo Loading in Polymersomes 1.2 Clustering Methods of Synthetic Vesicle 1.2.1 Host-Guest Interaction 1.2.2 DNA Hybridization 1.2.3 Copper-Free Click Chemistry 1.3 Stimuli-Responsive Polymersomes with Controllable Membrane Permeability 1.3.1 pH-Responsive Polymersomes 1.3.2 Light-Responsive Polymersomes 2 Motivation and Aim Part II Experiments 3 Materials and Methods 3.1 Materials 3.2 Analytical Methods 4 Clustered pH-Responsive Polymersomes for Enzymatic Cascade Reaction 4.1 Synthetic Methods and Characterization of Block Copolymer (BCP) for Self- Assembly of Polymersomes 4.1.1 Synthesis of Poly(Ethylene Glycol) (PEG) Macroinitiator 4.1.2 Synthesis of Photo-Crosslinker 4.1.3 Synthesis of BCP with Different Terminal Groups 4.1.4 Synthesis of Bis-BCN Poly(ethylene glycol) Crosslinker (BisBCN-PEG) 4.2 Formation of Empty and Loaded Psomes-N3 4.2.1 Formation and Photo-Crosslinking of Empty-Psomes-N3 4.2.2 Preparation of Cy5 Labeled BSA (BSA-Cy5) 4.2.3 Preparation of RhB Labeled Myo (Myo-RhB) 4.2.4 Preparation of Cy5 Labeled GOx (GOx-Cy5) 4.2.5 Formation and Photo-Crosslinking of Loaded Psomes-N3 4.3 Preparation and Purification of Clustered Empty-Psomes-N3 II 4.3.1 Preparation of Clustered Empty-Psomes-N3 at Different Conditions 4.3.2 Optimized Preparation of Clustered Empty-Psomes-N3 4.3.3 Purification Method of Clustered Empty-Psomes-N3 4.3.4 DLS Measurement of the Empty-Psomes-N3 in the Supernatant 4.3.5 Quantification of Removed Psomes-N3 after Centrifugal Purification 4.4 Preparation and Purification of Clustered Enzyme-Psomes-N3: Enzymatic Cascade Reaction 4.4.1 Preparation of Clustered GOx or Myo Loaded Psomes-N3 (GOx-Psomes-N3 or Myo-Psomes-N3) 4.4.2 Enzyme Activity of Myo Samples 4.4.3 Enzyme Activity of GOx Samples 4.5 Preparation and Purification of Co-Clustered Enzyme-Psomes-N3: Enzymatic Cascade Reaction 4.5.1 Preparation of Co-Clustered Myo/GOx-Psomes-N3 4.5.2 Enzyme Activity of Co-Clustered Myo/GOx-Psomes-N3 Samples 5 Light-Driven Enzyme Reaction Based on pH-Responsive Polymersomes 5.1 Synthetic Methods and Characterization of Block Copolymers with Single Azobenzene Unit 5.1.1 Synthesis of Block Copolymer with Donor-Acceptor-Substituted Azobenzene Linkage between Hydrophilic and Hydrophobic Segments (BCP-DA-Azo) 5.1.2 Synthesis of Block Copolymer with Ether Substituted Azobenzene Linkage between Hydrophilic and Hydrophobic Segments (BCP-Azo) 5.2 Photo-Isomerization of Macroinitiator and Block Copolymer with Azobenzene Linkage 5.2.1 Photo-Isomerization of PEG-DA-Azo Macroinitiator Based on Blue Light Irradiation or UV Irradiation 5.2.2 Photo-Isomerization of PEG-Azo Macroinitiator Based on Blue Light Irradiation or UV Irradiation 5.2.3 Photo-Isomerization of BCP-DA-Azo (-) Based on Blue Light Irradiation or UV Irradiation 5.2.4 Photo-Isomerization of BCP-Azo (-) Based on Blue Light Irradiation or UV Irradiation 5.3 Formation and Characterization of Polymersomes with Azobenzene 5.3.1 Self-Assembly of Polymersomes with Azobenzene III 5.3.2 Photo-Isomerization of Psomes-DA-Azo (-) Based on Blue Light Irradiation or UV Irradiation 5.3.3 Photo-Isomerization of Psomes-Azo (-) Based on Blue Light Irradiation or UV Irradiation 5.3.4 Photo-Crosslinking of Polymersomes with Azobenzene 5.3.5 DLS Measurement of Photo-Crosslinked Polymersomes with Azobenzene through pH Titration 5.3.6 Photo-Stability of Polymersomes with Azobenzene 5.3.7 In-Situ Loaded Nile Red in Non-Photo-Crosslinked Polymersomes with Azobenzene (NR-Psomes-DA-Azo (+) or NR-Psomes-Azo (+)) 5.3.8 In-Situ Loaded Myo in Photo-Crosslinked Polymersomes with Azobenzene (Myo-Psomes-DA-Azo (+) or Myo-Psomes-Azo (+)) 5.4 Light-Induced Dye Release from Polymersomes with Azobenzene 5.4.1 Fluorescence Photobleaching of Nile Red under Blue Light or UV Irradiation 5.4.2 Nile Red Release under Blue Light or UV Irradiation 5.5 Light-Driven Enzyme Reaction Based on Polymersomes with Azobenzene Part III Results and Discussions 6 Clustered pH-Responsive Polymersomes for Enzymatic Cascade Reaction 6.1 Aim and Strategy 6.2 Photo-Crosslinked and pH-Responsive Polymersomes 6.2.1 Synthesis and Characterization of Block Copolymers (BCPs) 6.2.2 Formation and Characterization of Polymersomes 6.3 Preparation and Purification of Clustered Empty-Psomes-N3 6.3.1 Key Parameters of Clustering Process 6.3.2 Purification Methods of Clustered Empty-Psomes-N3 6.4 Preparation and Purification of Clustered Empty-Psomes-N3 and Enzyme-Psomes- N3 90 6.4.1 Formation and Characterization of Enzyme in-Situ Loaded Psomes-N3 (Enzyme- Psomes-N3) 6.4.2 Enzyme Location in Polymersomes 6.4.3 Deeper Characterization of Clustered Empty-Psomes-N3 and Clustered Enzyme- Psomes-N3 6.5 Clustered Enzyme-Psomes-N3 for Enzymatic Cascade Reaction 6.5.1 Influence of Enzyme Activity on Clustering Condition IV 6.5.2 Mixed Enzyme-Psomes-N3 for Enzymatic Cascade Reaction 6.5.3 Co-Clustered Enzyme-Psomes-N3 for Enzymatic Cascade Reaction 6.6 Summary 7 Light-Driven Enzyme Reaction Based on pH-Responsive Polymersomes 7.1 Aim and Strategy 7.2 Preparation and Characterization of Light-Responsive Polymersomes 7.2.1 Synthesis and Characterization of BCP with Different Types of Azobenzene Unit 7.2.2 Self-Assembly and Photo-Crosslinking of Light-Responsive Polymersomes 7.2.3 Characterization of Photo-Crosslinked Light-Responsive Polymersomes 7.3 Photo-Isomerization of Azobenzene Containing Polymeric Macromolecules and Vesicles 7.3.1 Photo-Isomerization of Azobenzene Containing PEG Macroinitiators 7.3.2 Photo-Isomerization of Azobenzene Containing BCPs and Polymersomes 7.4 Light-Driven Dye Release from Polymersomes with Azobenzene at Simulated Physiological Conditions 7.4.1 Characterization of In-Situ Nile Red Loaded Polymersomes 7.4.2 Light-Driven Dye Release from Polymersomes at Simulated Physiological Conditions 7.5 Light-Induced Enzyme Reaction in Polymersomes with Azobenzene at Simulated Physiological Conditions 7.5.1 Characterization of Polymersomes in-Situ Loaded Myoglobin 7.5.2 Light-Induced Enzyme Reaction in Polymersomes at Simulated Physiological Conditions 7.6 Summary 8 Conclusion and Outlook Reference List of Figures List of Tables List of Abbreviations and Symbols Appendix Acknowledgements Versicherung