Bacteria live in environments featuring various complex chemical concentration gradients. The formation of these gradients is a combined result of mass transfer and bacterial activities, creating a challenge in isolating the effects of specific gradients on bacterial activities. Since the size of a single microbe is 1 to 10 μm, the spatial profiles of the surrounding chemical concentration gradients are at a resolution similar to the size of a single microbe. Moreover, time-dependent bacterial activities could lead to temporal variation in these chemical concentration gradients. Both spatial chemical heterogeneity and temporal chemical variation are important in shaping bacterial activities. The critical roles of gradients on bacterial behavior poses a significant challenge: the coexistence of numerous gradients simultaneously complicates discerning the effects of a specific gradient on a particular bacterial activity. To address this challenge, I established a workflow using electrochemistry to create electrochemical gradients to simulate specific chemical gradients in bacterial environments. Additionally, I leveraged electrode design to control the generated gradients and minimize side effects where electrochemistry could introduce to the bacterial environment. In my first research project (Chapter 2), I fabricated a flow device equipped with a microwire electrode to generate and control oxygen (O2) and hydrogen peroxide (H2O2) gradient to mimic bacterial gradients. The first highlight of this research is that I generated electrochemical H2O2 gradients through an electrocatalyst design to simulate spatial distributions of reactive oxygen species in bacterial environments. By coating the electrodes with Au catalysts to facilitate the two-electron oxygen reduction reaction (2e-ORR), I achieved a spatial profile of H2O2 perpendicular to the electrode surface. The second highlight of this project is that I enabled spatial and temporal control of both electrochemical gradients via electrode morphology design and applying time-dependent external voltages, respectively. This spatiotemporal control strategy achieved spatial control at a precision within 10 μm and temporal control on a second scale, comparable to the spatiotemporal variation of bacterial environments. Moreover, the concentration ranges of O2 (0 to 250 μM) and H2O2 (0 to 40 μM) in the electrochemical gradient, aligned with the conditions of microbiological research. To advance this research, I, together with co-authors Jingyu Wang and Ben Hoar, developed a machine-learning-assisted design strategy for desired O2 and H2O2 gradients, offering a rapid and efficient solution for electrode optimization. I created another novel electrochemical flow device for my second research project (Chapter 3). I introduced two main electrode design strategies to enhance compatibility and convenience in gradient-bacteria research. Firstly, I deposited a layer of porous copolymer on the electrode surface to increase the oxygen reduction reaction (ORR) selectivity and create a “clean” O2 gradient for bacterial research. This innovation was particularly critical in mitigating the interference of external voltages with bacterial redox systems, thus increasing the integrity of experimental results. Based on size exclusion, the coated polymer maintained the ORR current while rejecting the access of larger bacterial redox molecules to the electrode surface. Using a redox molecule produced by Pseudomonas aeruginosa, pyocyanin, as an example, the coated polymer enabled the ORR current to remain around 70% compared to that on the current on a bare electrode while suppressing almost 100% of the redox current of pyocyanin. Secondly, I adopted planar electrode arrays instead of the vertical electrode arrays used in Chapter 2. The planar geometry enabled spatial O2 concentration profile gradient generation both perpendicular and parallel to the electrode surface. This innovation proved instrumental as it facilitated microscopic microbial research while also allowing the replication of gradients within three-dimensional bacterial structures. In summary, this porous polymer coating enabled the creation of a “clean” O2 gradient, allowing investigation of the effect of O2 on microbial behavior. In Chapter 4, I proposed a potential application using electrochemical gradients in microbiological research. I describe utilizing electrochemical O2 gradients to investigate the effect of O2 variation on bacterial adenosine triphosphate concentration ([ATP]) at the single-cell level. I reviewed the published literature on the relationship between O2 concentration and microbial ATP activity. Most previous research is focused on the average effect of O2 on adenosine energy charge (AEC) and [ATP] in a bulk bacteria culture. As such, the effect of O2 on single cells is missing. I proposed an experiment plan to utilize the electrochemical O2 gradient platform to study the effects of O2 on single microbes and explored the feasibility of this plan. I raised several potential concerns or questions that could arise from an electrochemistry-microbiology hybrid system. In summary, my thesis comprehensively explores electrochemical concentration gradients and their implications for microbial behavior. By employing innovative electrode design strategies, I explored the feasibility of using electrochemical concentration gradients for nuanced investigations into the intricate dynamics of bacterial ecosystems. Looking ahead, I believe the fusion of electrochemistry and microbiology holds promise for advancing our understanding of bacterial behavior, especially at the single-cell level and beyond.

The shale gas revolution has boosted the United States economy ever since the early 2000s. Its surge in shale gas production has led to an abundance of the light alkanes including methane (CH4), ethane (C2H6), and propane (C3H8), presenting both opportunities and challenges in terms of their conversion into valuable products.CH4 is the primary component of shale gas. Its emission is indeed driving global climate change. Developing an efficient chemical transformation pathway for CH4 provides an applicable solution to use CH4 in shale gas and meanwhile reduce its emission into atmosphere, yet it is still one of the remaining ‘grand challenges’ in chemistry, especially for the one that occurs at ambient conditions. The key resides on the discovery of new catalytic systems to break the very strong C-H bond within a CH4 molecule. The major part of this dissertation focuses on the development of metal complexes as the homogeneous electrocatalysts by which the injected CH4 can be directly converted into functionalized product in liquid phase. At a relatively “wild” guess from the classic hard-soft acid-base (HSAB) theory, we speculated that a high-valent transition metal, divalent silver (AgII) radical, as a soft class (b) Lewis acid based on Pearson’s classification, is a suitable candidate for CH4 activation due to methyl (CH3) moiety’s relatively low value of chemical hardness. Inspired by such notion, we reported that electrochemically generated AgII readily functionalizes CH4 into methyl bisulfate (CH3OSO3H) as a precursor of methanol (CH3OH) at ambient conditions in concentrated sulfuric acid (98% H2SO4). The second work further provides chemical insight that metal-bound bisulfate ligand (–OSO3H) introduced by 98% H2SO4 could be redox-active toward CH4 with our reported molecular catalyst vanadium (V)-oxo dimer (V2V,V). While CH4 is often used for energy production, the extraction and utilization of other non-negligible components including C2H6 has diversified the applications and markets for shale gas products. To expand our knowledge base in the emerging field of electrocatalytic alkane functionalization, we further discovered and developed the direct electrochemical coupling of C2H6 at ambient conditions, where all of the gas products can be easily separated from the cheap and green aqueous electrolytes and transported to the downstream chemical production. This work also highlights electrolyte engineering to avoid the common use of 98% H2SO4 electrolyte, which may inspire environmentally benign electrocatalysis for alkane activation. Altogether, these three independent works show the full potential of electrocatalytic ambient light alkane functionalization for shale gas utilization on the chemical industry in the near future. Throughout my time as a graduate student, with the understanding of scientific knowledge, technological applications, and practical tasks, I have demonstrated how the combination of electrochemistry, radical chemistry, organometallics, and homogeneous catalysis with instrumentation skills can provide fresh perspectives on fundamental chemistry and chemical engineering. The use of electrogenerated radicals to C-H activation is a distinctive phenomenon, offering a direct way to overcome kinetic inertness and facilitate innovative reactivity pathways of small molecules.

Multi-step processes, such as catalytic cycles, proceed through one or more intermediates that may participate in unwanted side reactions, leading to inefficiency and waste. Further, these intermediates may have a short shelf life and/or pose a safety concern. Thus, the development of methods to carry out multi-step processes to generate and utilize intermediates in one pass is of great desire. Biology manages its complex reaction network of multi-step, multi-enzyme processes by numerous means, namely spatial control via compartmentalization. By controlling where certain processes occur and the diffusion of key intermediates between reaction sites, biology efficiently carries out multiple concurrent reaction sequences efficiently with minimal competing pathways. For example, carboxysomes enhance the rate of CO2 fixation by co-encapsulating carbonic anhydrase and ribose 1,5-bisphosphate carboxylase-oxygenase while excluding deactivating oxygen (O2). Inspired by spatial control in biology, my research seeks to adapt such methods of spatial control to construct efficient multi-step, multi-catalyst organometallic and electrochemical processes. In this manner, commodity chemicals can be produced from abundant feedstocks while obviating intermediate isolation and work up. Electrochemistry has emerged within the last few decades at the forefront of small molecule activation, particularly of environmental pollutants such as CO2, and organometallic chemistry is apt to further utilize products of electrochemical small molecule activation owing to decades of rich literature in homogeneous catalyst development. However, it is highly likely that catalyst - catalyst, catalyst - substrate, or substrate - substrate interference may impede the integration of multiple processes. Potential undesired interference may be circumvented by spatially separating while co-localizing the electro- and organometallic (or any type) catalysts in one reactor system allowing the transport of intermediates between them. The projects outlined below demonstrate the critical role spatial control and mass transport have in constructing efficient multi-step, multi-catalyst electro- and/or organometallic processes.

With the surge of intermittent, renewable electricity, the storage of excessive electricity and reduction of CO2 or N2 into value-added chemicals is of great significance for a sustainable society. One viable route that fulfills such a target is to construct a hybrid inorganic-biological system that converts electricity into chemical energy and reduces CO2/N2 into commodity chemicals. In this general approach, water is split into H2 and O2 by renewable electricity and the yielded H2 is consumed by microbes as a reducing equivalent for CO2/N2 reduction. The throughput of this system is limited by the poor gas solubility in the aqueous environment. The control and design of the gas environment in these hybrid systems is crucial to achieving high throughput and sustainable reactions. To this end, my thesis has focused on combining perfluorocarbon (PFC) nanoemulsions with the hybrid system to enhance H2 or O2 gas solubility and optimize reaction performance. My first research project focused on combining PFC nanoemulsions with a hybrid biological inorganic CO2 fixation system to enhance the faradaic efficiency of acetic acid production (Chapter 2). PFC nanoemulsions are biologically inert and have reported high H2 gas solubilities. We determined that the nanoemulsions are compatible with the acetogenic microbe strain Sporomusa ovata and the electrochemical water-splitting catalysts used in the hybrid CO2 fixation system. When 2.5 % (v/v) PFC nanoemulsion was combined with the hybrid system we observed approximately 90 % or greater faradaic efficiency at all tested current densities. After 4 days, we were able to achieve an average acetic acid titer of 6.4 � 1.1 g�L−1 (107 mM), which equates to one of the highest reported productivities of 1.1 mM�h−1. With this observed enhancement, we decided to explore the change in the gas environment made by the addition of PFC nanoemulsions. We performed experiments to understand the mechanism of the observed enhancement when PFC nanoemulsions were added to the hybrid CO2 fixation system (Chapter 3). Experiments of flow cytometry were performed using fluorescently tagged PFC nanoemulsions to probe the association of the S. ovata with the nanoemulsions. We found a non-specific binding interaction between the nanoemulsion and the bacteria that could result in an increase in the local H2 concentration or an increase in the rate of H2 transfer to the microbes. Rotating disk electrode (RDE) experiments were performed as an electrochemical surrogate to understand the H2 transfer kinetics. Our experiments revealed that there was 3.5 times increase in the kinetic current density with the addition of the nanoemulsion. Additionally, we found that the local H2 concentration was 1.2 times the calculated bulk H2 concentration. This suggests that the addition of the PFC nanoemulsions resulted in a local increase in H2 concentration and an increase in the rate of transfer of the reducing equivalent to the microbe. We theorized that the addition of PFC nanoemulsions to other hybrid systems could result in similar enhancement and an ability to control the gas environment. With the success of the PFC nanoemulsion in the hybrid CO2 fixation system, my research shifted to focus on using the nanoemulsions with a hybrid N2 fixation system to enhance ammonia production and self-sufficiency (Chapter 4). The hybrid N2 fixation reaction utilizes Xanthobacter autotrophicus, a microaerobic bacterial strain that reduces N2 to NH3 through its nitrogenase enzyme. This has previously been accomplished through a precise supplied gas mixture, since the electrochemically generated O2 is poorly soluble in the aqueous solution. We added the PFC nanoemulsions to this system to utilize the electrochemically generated O2 and enhance the throughput of the N2 fixation system. Our experiments demonstrated that the use of PFC nanoemulsions drastically reduced the loss of cell viability in a supplied anaerobic atmosphere and led to a 3-fold increase in Faradaic efficiency of the system during 5-days of running. We performed mechanistic studies to understand the O2 gas-PFC nanoemulsion environment within the hybrid system (Chapter 5). Confocal microscopy images of the PFC nanoemulsions interacting with the microbes revealed binding and complete coverage of the microbe surface by the PFC nanoemulsions. Experiments of RDE were performed to ascertain the O2 transfer kinetics between the PFC nanoemulsions and the microbes. We found that there was a 20-fold increase in the kinetic O2 reduction rate when the PFC nanoemulsions were added. Assuming a Langmuir adsorption model of the PFC nanoemulsion binding to the microbe, we were able to use flow cytometry to determine the maximum number of nanoemulsions that could adsorb to the microbe surface and the equilibrium constant for the adsorption. Our results showed that the number of nanoemulsions that could bind to a given microbe or particle is dependent on the size of the particle, but the equilibrium constant is more influenced by the surface charge, with the -COOH functionalized microspheres having the largest binding affinity. In conclusion, my thesis research was able to integrate PFC nanoemulsions with hybrid biological inorganic systems to enhance throughput and alter the gas environment in the systems. The ability to understand and alter the gas environment using these PFC nanoemulsions can allow for other aqueous systems with poor gas transfer rates or gas restrictions to function with enhanced benefits. This dissertation also serves as groundwork for continued research in customizing the PFC nanoemulsion to achieve the optimal binding and gas environment.

Artificial photosynthesis, the biomimetic approach to converting sunlight's energy directly into chemical fuels, offers an attractive way to address the need for a clean, renewable source of energy. In plants, chloroplasts store the sun's energy using a system of integrated photosynthetic nanostructures including light-absorbing pigments, electron-transport chains, and chemical catalysts. However, neither such integration of nanostructures nor energy conversion efficiency suitable for practical applications has been achieved in artificial photosynthesis. In this context, the subject of my graduate research is to develop an integrated system using nanowire-based nanostructures to imitate natural photosynthesis. This centers on two themes: (1) constructing novel integrated nanostructures for solar-to-fuel conversion, and (2) developing next-generation materials and catalysts for improved photoelectrochemical (PEC) performance.

Although natural photosynthesis organizes its active components at the nanometer scale to better control the process of energy conversion, this level of integration had not been realized until recently for artificial photosynthesis. Here the construction of a nanowire-based integrated system to realize such a nanoscopic control is demonstrated.

Since all of the processes in PEC relate to the interfaces among semiconductor light-absorbers, electrocatalysts, and the electrolyte, the first to realize an integrated nanosystem was to under how photo-excited carriers would transfer within these interfaces. By using kelvin probe force microscopy (KPFM), the local electrostatic potential of an asymmetric nanowire composed of Si and a TiO2 shell, which was covered in a layer of water. Different local potentials were observed in dark and under illumination, which provides the knowledge that the heterojunction of Si and TiO2 could function as a Z-scheme system for solar water splitting.

After obtaining this piece of information, we moved forward to develop an integrated nanosystem for artificial photosynthesis. Taking the concept of Z-scheme, we used Si and TiO2 nanowires as building blocks to construct a tree-shaped heterostructure. In this structure, the positions of the reduction and oxidation components were pre-defined to mimic the spatial control found in chloroplasts. The integrated standalone device splits H2O into H2 and O2 under simulated sunlight, with an efficiency of solar-to-fuel conversion comparable to that of natural photosynthesis. This first demonstration paves the way for using nano-sized building blocks to achieve efficient solar-to-fuel conversion.

The nanowire-based integration allows individual building blocks to be replaced with newly developed ones. In this dissertation advanced building blocks for artificial photosynthesis is also demonstrated.

We have explored new materials and methods to improve the energy conversion efficiency of semiconductor light-absorbers. Solution-phase synthesis of III-V semiconductor nanowires was successfully demonstrated for photocatalytic reactions, and the nanowires' electronic properties could be fine-tuned to fit the needs of device integration. Also we demonstrated enhancement of the photoanodic activity of hematite (Fe2O3) using the surface plasmon resonance of exquisitely controlled Au nanostructures.

Additionally, new electrocatalysts suitable for practical applications are developed. We first looked into the lower limit of platinum (Pt) loading as a catalyst for the H2 evolution reaction (HER). Using the atomic layer deposition (ALD) technique, it is possible to quantitatively controlled the Pt loading down to about 0.2% of a monolayer (~10 ng/cm2), which is sufficient for some PEC applications. Cobalt sulfide, an earth-abundant catalyst, was also synthesized by electrodeposition. It acted as a HER catalyst in water at neutral pH and could be coupled with a Si photocathode for solar H2 production. Moreover we are developing an effective CO2 reduction catalyst of near unity selectivity for acetate production, which could be added into the integrated nanostructure.¬¬¬

In conclusion, my graduate research focuses on the integration of nanowire-based structures to achieve more efficient artificial photosynthesis. This is demonstrated in this dissertation not only at system level for an intergrated nanostructure, but also at component level for advanced building blocks. This research can serve as a foundation for the efforts of other researches in the field of artificial photosynthesis.

In biology, chemical species (such as oxygen (O2)) and their local concentrations are highly regulated. One example is the reduction of dinitrogen (N2) to ammonia by aerobic bacteria. In this scenario, the O2 sensitive nitrogenase enzyme provides electrons for N2 reduction through aerobic respiration. The tandem reactions of N2 fixation and aerobic respiration are only possible due to the buildup of a chemical O2 concentration gradient where the O2 sensitive nitrogenase is situated in an O2-free region and aerobic respiration in an O2-rich region. Inspired by biological microscopic chemical gradients, my research aims to use electricity and nanomaterials to mimic these natural phenomena and apply them to synthetic organometallic chemistry to offer new pathways in carbon maintenance. Homogeneous organometallic complexes are often utilized for small molecule activation because they offer a delicate synthetic control and can be characterized extensively, but can suffer from air or moisture sensitivity. Therefore, the marriage of traditional organometallic chemistry with electricity and nanoscience can help reconcile this incompatibility by offering new reactivity or unforeseen reaction routes. The projects described below illustrate the spatiotemporal control we can achieve over electroactive chemical species to induce novel catalytic routes and difficult-to-achieve transformations.In my first research project (Chapter 2), I developed an electricity-powered catalytic system mediated by a RhII metalloradical complex for the conversion of methane (CH4) to methanol (CH3OH) with air as the terminal oxidant. The challenge here is that the RhII complex kinetically reacts with O2 faster than with CH4. I reconciled this incompatibility by introducing a nanowire electrode, which, when combined with a reducing potential, created a steep O2 gradient within the wire array while also regenerating the RhII complex. Thus, C‒H activation occurred anaerobically while CH3OH synthesis proceeded aerobically together in one catalytic cycle. Moreover, the reaction rate of CH4 activation increased 220,000-fold within the wire array, achieving a turnover number of 52,000 in 24 hours. Additionally, other light alkanes (such as ethane, propane, and toluene) were used as substrates and oxidation to primary alcohols was acheived in all cases. Building off the work presented in Chapter 2, the work described in Chapter 3 uncovers the identity and the role of the terminal oxidant responsible for generating CH3OH from the methylated Rh complex. While O2 reduction at the silicon nanowire array working electrode creates an anaerobic region within the wire array in air, it simultaneously generates the reactive oxygen species necessary for alcohol formation. The utilization of electron paramagnetic resonance spectroscopy identified the electrogenerated oxidant as superoxide and reactions with a selective chromophore quantified the rate of superoxide generation. While superoxide is the immediately generated species at the electrode, it is presumed that chemical and/or further electrode reactions convert superoxide into a hydroperoxyl or hydroperoxide species, which is ultimately responsible for the oxidation reaction. Chapter 4 investigates the role of the nanowire array’s efficacy in creating a microscopic compartment for the RhII metalloradical-assisted CH4 activation. While quantitative analysis of microscopic compartmentalization is typically applied to biochemical reactions, in this work we translated this quantitative understanding to organometallic cascade reactions in terms of reaction efficiency. We analyzed the reaction efficiency of our nanowire array compartment as the ratio of product molecules leaving the nanowire array to substrate molecules that enter the wire array compartment. In order to properly quantify this reaction metric, we utilized the experimental data obtained in Chapter 3. In conclusion, we found good agreement between our experimental results and the results from our semi-quantitative kinetic model. The final chapter of my dissertation (Chapter 5) aims to translate the O2 gradient observed previously to an electrochemical carbon monoxide (CO) gradient, from carbon dioxide (CO2) reduction, to control the microstructure of synthetic polyketones. The ultimate goal of this project is to integrate, in one pot, the electroreduction of CO2 to CO and the palladium (Pd) assisted copolymerization of CO and ethylene (C2H4). While polyketone synthesis typically follows a perfectly alternating pattern (1:1 stoichiometry between CO and C2H4), recent work has focused on utilizing Pd catalysts with asymmetric ligand frameworks to induce a non-alternating structure. The goal of this effort is to reduce the CO content (through extra C2H4 insertions) in order to improve the degradability of the polymer while maintaining its durability. However, non-alternating copolymerization still requires modulation of both the monomer feed ratio (C2H4 relative to CO) and reaction temperature to control the extent of CO insertion. Based on the previously observed chemical O2 gradient, our hypothesis here is that electricity and a nanowire array will allow us to spatiotemporally control the local concentration of CO and ultimately its incorporation into the polymer. During my graduate career, I have shown that interfacing electrochemistry and nanomaterials with homogeneous catalysis can offer new insight into fundamental chemistry and push the boundaries of what was thought was possible with traditional organometallic chemistry. The utilization of electrochemically generated concentration gradients is a unique phenomenon that can provide an avenue to bypass inherent incompatibilities and enable novel reactivities.