In the area of forensic chemistry, improved or new analysis methods are continually being investigated. One common and powerful technique used in forensic chemistry is wall-coated open-tubular column (WCOT) gas chromatography with electron ionization single quadrupole mass spectrometry (GC-MS). Improvements to and effectiveness of alternatives to this instrumental platform were explored in an array of parallel inquiries. The areas studied included the column for the chromatographic separation, the universal detection method employed, and the fragmentation method used to enhance molecular identification. <br><br>Superfine-micropacked capillary (SFµPC) columns may provide an alternative to commercial packed GC columns and WCOT GC columns that combines the benefits of the larger sample capacity of packed columns and the benefits of the excellent separation capabilities and mass spectrometry (MS) flow rate compatibility of WCOT columns. SFµPC columns suffer from high inlet pressure requirements and prior reported work has required specialized instrumentation for their use. Fabrication of and chromatography with SFµPC GC columns was successfully achieved with typical GC-MS instrumentation and within the flow rate limit of a MS. Additionally, the use of higher viscosity carrier gasses was demonstrated to reduce the required inlet pressure for SFµPC GC columns.<br><br>Recently, a new vacuum ultraviolet spectrometer (VUV) universal detector has been commercialized for GC. The ability of VUV detectors to acquire absorbance spectra from 125 nm to 430 nm poses a potential alternative to MS. As such, GC-VUV provides an exciting potential alternative approach to achieving excellent quantitative and qualitative analysis across a wide range of analytes. The performance of VUV and MS detectors for forensic analysis in terms of quantitative and qualitative analysis was compared. Analysis of alkylbenzenes in ignitable liquids was explored, which can be important evidence from suspected arson fires and are difficult to differentiate with MS. The VUV detector was found to have superior specificity and comparable sensitivity to the MS detector in scan mode.<br><br>Addition of thermolysis (Th) as an orthogonal fragmentation pathway provides the opportunity to increase the differences between MS fragmentation patterns. Fragmentation has been widely established to aid in identification of molecules with MS by providing characteristic fragments at characteristic relative abundances. However, molecules with very similar structures do not result in sizable spectral differences in all cases with typical MS fragmentation techniques. A series of Th units were fabricated and integrated into GC-Th-MS instruments. Th-MS was conducted with the thermally labile nitrate esters across a range of instrumentation and thermal conditions.<br>

<p>The rapid analysis of complex samples using mass spectrometry (MS) provides valuable information in both point-of-care (e.g. drug testing) and laboratory-based applications, including the generation of spectral libraries for classification of biosamples, the identification of biomarkers through large-scale studies, as well as the synthesis and bioactivity assessments of large compound sets necessary for drug discovery. In all these cases, the inherent speed of MS is attractive, but rarely fully utilized due to the widespread use of sample purification techniques prior to analysis. Ambient ionization methodologies can help circumvent this drawback by facilitating high-throughput qualitative and quantitative analysis directly from the complex samples without any need for work-up. For instance, the use of swabs or paper substrates allows for rapid identification, quantification, and confirmation, of drugs of abuse from biofluids or surfaces of forensic interest in a matter of minutes, as described in the first two chapters of this dissertation. Faster analysis can be achieved using an automated desorption electrospray ionization (DESI) platform which allows for the rapid and direct screening of complex-sample microarrays with throughputs better than 1 sample per second, giving access to rich spectral information from tens of thousands of samples per day. The development of the bioanalytical capabilities of this platform, particularly within the context of drug discovery (e.g. bioactivity assays, biosample analysis), is described across most other chapters of this dissertation. The use of DESI, a contactless ambient ionization method developed in our laboratory and whose 20 years of history are overviewed in the introduction of this document, provides an additional advantage as the secondary microdroplets generated through the DESI process act as reaction vessels that can accelerate organic reactions by up to six orders of magnitude, facilitating on-the-fly synthesis of new compounds from arrays of starting materials. Unique implications of this microdroplet chemistry in the prebiotic synthesis of peptides and spontaneous redox chemistry at air-solution interfaces, together with its practical applications to the synthesis of new drug molecules, are also overviewed. The success obtained with the first automated DESI-MS system, developed within the DARPA Make It program, led to increased interest in a new-generation platform which was designed over the past year, as overviewed in the last section of this dissertation, and which is currently being installed for validation prior to the transfer of the technology to NCATS, where we anticipate it will make a significant impact through the consolidation and acceleration of the early drug discovery workflow.</p>

<p>Canines can detect different diseases simply by smelling different biological sample types, including  urine,  breath  and  sweat.  This  has  led  researchers  to  try  and  discovery  unique  volatile  organic compound (VOC) biomarkers. The power of VOC biomarkers lies in the fact that one day they may be able to be utilized for noninvasive, rapid and accurate diagnostics at a point of care using  miniaturized  biosensors.  However,  the  identity  of  the  specific  VOC  biomarkers  must  be  demonstrated before designing and fabricating sensing systems. Through  an  extensive  series  of  experiments,  VOCs  in  urine  are  profiled  by  solid  phase  microextraction (SPME) coupled to gas chromatography-mass spectrometry (GC-MS) to identify biomarkers for breast cancer using murine models. The results from these experiments indicated that  unique  classes  of  urinary  VOCs,  primarily  terpene/terpenoids  and  carbonyls,  are  potential  biomarkers  of  breast  cancer.  Through  implementing  chemometric  approaches,  unique  panels  of  VOCs  were  identified  for  breast  cancer  detection,  identifying  tumor  location,  determining  the  efficacy of dopaminergic antitumor treatments, and tracking cancer progression. Other diseases, including COVID-19 and hypoglycemia (low blood sugar) were also probed to identify volatile biomarkers present in breath samples.  VOC biomarker identification is an important step toward developing portable gas sensors, but  another  hurdle  that  exists  is  that  current  sensors  lack  selectivity  toward  specific  VOCs  of  interest.  Furthermore,  testing  sensors  for  sensitivity  and  selectivity  is  an  extensive  process  as  VOCs  must  be  tested  individually  because  the  sensors  do  not  have  modes  of  chromatographic  separation or compound identification. Another set of experiments is presented to demonstrate that SPME  fibers  can  be  coated  with  materials,  used  to  extract  standard  solutions  of  VOCs,  and  analyzed  by  GC-MS  to  determine  the  performance  of  various  gas  sensing  layers.  In  the  first  of  these  experiments,  polyetherimide  (PEI)  was  coated  onto  a  SPME  fiber  and  compared  to  commercial polyacrylate (PAA) fibers. The second experiment tuned the extraction efficiency of polyvinylidene fluoride (PVDF) - carbon black (CB) composites and showed that they had higher sensitivity  for  urinary  VOC  extraction  relative  to  a  polydimethylsiloxane  (PDMS)  SPME  fiber.  These results demonstrate SPME GC-MS can rapidly characterize and tune the VOC adsorption capabilities of gas sensing layers. </p>

<p dir="ltr">The beautiful complexity of our world is manifested in how macro- and even planetary-scale processes are essentially completely determined and regulated by chemical and physical transformations happening at the micro- and nanoscale. The introduction and subsequent development of optical microscopy methods have provided us with a unique opportunity to visualize, probe, and sometimes even control these processes that are too small to be seen by the human eye by their nature.</p><p dir="ltr">Among the great variety of truly impressive advances in microscopy instrumentation, two techniques stand out in their widespread and usefulness. First of them, fluorescence imaging has completely revolutionized the study of biological specimens and living systems due to its unprecedented single-molecule sensitivity and resolution combined with video-rate imaging capability. On the other hand, chemical imaging in the mid-infrared region provides an unmatched amount of chemical information enabling label-free mapping of the spatial distribution of various classes of biological molecules. However, each of these techniques falls short where the other excels. For example, despite its high resolution and sensitivity, fluorescence imaging does not carry direct chemical information and relies on labeling specificity, while infrared microscopy is diffraction-limited at the resolution of several micrometers and suffers from low penetration depth in aqueous solutions.</p><p dir="ltr">This dissertation introduces a novel imaging method designed to combine the advantages of fluorescence imaging and infrared spectroscopy. Fluorescence-detected photothermal mid-IR (F-PTIR) microscopy is presented in <b>chapter 1</b> as a technique enabling sub-diffraction chemically-specific microscopy by detecting local temperature-induced fluctuations in fluorescence intensity to inform on localized mid-infrared absorption. F-PTIR applications in targeted biological microspectroscopy (<b>chapter 1</b>) and pharmaceutical materials (<b>chapters 2 and 3</b>) analysis are demonstrated to highlight the potential of this new method. Furthermore, instrumentation developments relying on modern radiation sources such as dual-comb quantum cascade laser and synchrotron infrared radiation are shown to improve spectral acquisition speed (<b>chapter 4</b>) and spectral coverage (<b>chapter 5</b>), respectively, to extend the application range of F-PTIR.</p>

<p>  </p> <p>Atmospheric aerosols formed through primary emissions, secondary gas-particle formations, and multi-phase chemical processes are composed of solid, semi-solid, or liquid-like particles suspended in the air that have direct implications towards the global radiative balance and human health as air pollutants.  Direct emissions of primary organic aerosols (POA; e.g. soot, BrC) and multi-phase formation of secondary organic aerosols (SOA) from the oxidation of biogenic monoterpene isomers represent two important sources/classes of particulate matter in the atmosphere. Multi-phase chemical processes driving the atmospheric and environmental aging through the photochemistry of iron(III), FeIII in organic aerosol particles and aqueous media drives the multiphase chemistry leading to systematic aging of their chemical composition and modifications to resulting light-absorption properties. The molecular composition, organic structures, physical properties, and sources of emissions are complex requiring development of powerful multi-modal analytical metrology, such as high-resolution mass spectrometry (HRMS) hyphenated with liquid chromatography (LC), photodiode array optical detection, drift tube ion mobility (IM) spectrometry, and desorption and ambient ionization of multi-components mixtures in atmospheric particles using temperature programmed desorption Direct analysis in real time (TPD-DART). Disseminating the molecular-specific composition, chemical and physical properties of complex mixtures in atmospheric organic particles and mixed inorganic/organic systems will help improve our understanding of their formation mechanisms, transformative chemical ageing processes, as well as improved detection of individual components in complex mixtures. </p> <p>     </p> <p>Chapter 1 and 2 of dissertation introduces complexity of atmospheric organic, carbonaceous aerosols, and complex environmental mixtures and discusses analytical metrology, experiments, and data analysis procedures used for detailed molecular-level characterization of mixtures. Chapter 3 the development of a robust analytical method for untargeted screening and determination of the physical and chemical properties (e.g. vapor pressures, enthalpies of sublimation, and saturation mass concentrations) of single components out of complex SOA particles using temperature programmed desorption Direct analysis in real time ionization – high resolution mass spectrometry (TPD-DART-HRMS).  Chapter 4 introduces the use of ion mobility - mass spectrometry (IM-MS) separation and multidimensional characterization of structural isomers in complex SOA mixtures. The chapter discusses the advanced usage of IM-MS to investigate the molecular and structural properties of isomers of alpha-pinene and limonene derived SOA, use of advanced data analysis procedures to resolved complex conformational and structural isomers, and investigate single-molecule structural changes from atmospheric-like ageing in SOA particles using IM-MS.  Chapter 5 discusses the chemical characterization and analysis of individual brown carbon (BrC) chromophores out of mixture of colorless organic carbon constituents and insoluble soot particles generated from controlled flame combustion of ethane fuel, a surrogate system representing gasoline combustion of motor vehicles. The chapter focuses on the quantitative method development and use of state-of-the-art liquid chromatography coupled to photodiode array followed by dopant assisted atmospheric pressure photoionization and HRMS (LC-PDA-HRMS) analysis, followed by conversion to quantitative optical information for comparisons with retrieved literature reports. Chapter 6 examines the complex multiphase photochemical cycling of Fe(III)-citrate, a relevant proxy for [FeIII-carboxylate]2+ complexes in atmospheric water using complementary analytical metrology of optical spectroscopy, LC-PDA-HRMS, oil immersion flow microscopy. Multi-modal datasets from these complementary techniques provide a unique experimental description of various stages of FeIII-citrate photochemistry, elucidate individual components of this reacting system, determine mechanistic insights, and quantify environmental parameters affecting the photochemistry. </p>

<p> Mass spectrometry imaging (MSI) is an advanced analytical technique that renders spatially defined images of complex label-free samples. Nanospray desorption electrospray ionization (nano-DESI) MSI is an ambient ionization direct liquid extraction technique in which analytes are extracted by means of a continuous liquid flow between two fused-silica capillaries. The droplet generated between the two capillaries is controlled by a delicate balance of solvent flow, solvent aspiration, capillary angles, and distance from the surface. This technique produces reproducible ion images with up to 10 µm resolution and can be used to identify and quantify multiple analytes on a given surface.  This thesis discusses some of the applications of this technique to biological systems, as well as the work done to develop methodology to further improve this technique’s specificity and sensitivity. Herein, applications that push the limits of the current capabilities of nano-DESI are presented, such as the high-resolution imaging of lipid species in skeletal muscle at the single-fiber level, and the quantification of low-abundance drug metabolites.  The second theme of this thesis, developing new capabilities, introduces ion mobility mass spectrometry imaging. This integrated technique increases the selectivity previously possible with MSI. To support these efforts, the work in this thesis has generated data analysis workflows that not only make these experiments possible but also further endeavor to increase sensitivity and combat instrument limitations on mobility resolution. Finally, this thesis present streamlined workflows for tandem MS experiments and modifications to a recently introduced microfluidic variant of the nano-DESI technique. In all, this thesis showcases the current capabilities of the nano-DESI technique and lays the groundwork for future improvements and capabilities.      </p>

<p>  </p> <p>Globally, communities are embracing the cured-in-place-pipe (CIPP) process due to the need to address damaged buried water and sewer pipes. CIPP involves the chemical manufacture of a new plastic pipe inside an existing buried water and sewer pipe, without the need for excavation. The process is popular because it can be 80% less costly than alternative methods and construction workers can be present for hours to not days to weeks. However, as CIPP use has grown, so have the number of hazardous material (HAZMAT) incidents caused by using this practice. Evacuations of daycare centers, schools, homes, healthcare, institutional, and other buildings have been caused. In some cases, chemical exposure victims have required medical assistance and hospital admission. For decades, organizations within the CIPP industry and municipalities have encouraged chemical waste discharge into ambient air, resulting in preventable exposures. Recent work has indicated tons of volatile organic compounds (VOC) may be released during a single CIPP project into the air. Chemicals released include hazardous air pollutants (HAP), carcinogens (CAR), endocrine disrupting chemicals (EDR), and other compounds with little toxicological information. While polymer composites have been manufactured for other applications for more than 50 years, little information exists about what chemicals and materials are used to manufacture CIPPs. As CIPP use has grown along with the number of bystander chemical exposures, concerns about the type, magnitude, and toxicity of chemical emissions from CIPP projects have markedly increased. To reduce the potential for human harm and environmental degradation, a better understanding of CIPP composite chemistry and manufacturing is needed. This dissertation aimed to elucidate the processes that control the composition of waste generated during plastic CIPP manufacture and ascertain how to modify the manufacturing practice to minimize impacts on composite integrity and emission toxicity. </p> <p>Chapter 1 focused on indoor VOC exposure simulation and styrene contamination/ decontamination to evaluate the risk of occupant exposure during CIPP installation. Styrene is a common monomer used in many CIPP resins and can be discharged into the air at CIPP worksites. A review of prior incidents revealed that CIPP waste (liquid, organic chemicals, etc.) could enter nearby buildings through multiple routes including windows, doors, or heating, ventilation, and air conditioning outdoor air intakes. When CIPP is manufactured inside a sanitary sewer pipe, waste can enter buildings through sewer laterals of nearby buildings and through foundation cracks. Study results showed that plumbing seal backflows in bathrooms caused by sewer repair work are hydraulically possible: the minimum pressure required to displace water in the plumbing trap was estimated to be 0.995 kPa and 8.85 kPa for a sink and toilet, separately. These pressures are much lower than those applied by the contractor during the sewer lining (up to 193.05 kPa). Based on the indoor exposure events, the dissipation potential of vapors, as well as the hydraulic calculations, indoor air chemical contamination and decontamination profiles were also examined. A mass balance model of chemical vapor dispersion was developed. Modeling results revealed that bathroom exhaust fan operation during a CIPP project can increase the indoor styrene concentration by enhancing the inflow of styrene-containing air from the sink and toilet. However, the styrene concentration decreased as air leaked across the bathroom door due to reduced suction in the plumbing. Based on incident reviews, chemical magnitudes, and modeling results it was concluded that CIPP waste discharge should be treated as hazardous material discharge, because of its threat to human health. Actions are needed to reduce waste generation and contain the waste, so it does not leave the worksite. Chapter 2 aimed to determine the manufacturing conditions that most influence chemical residual left in the thermally manufactured CIPP. Bench-scale testing of multiple styrene- and non-styrene composites revealed the manufacturing conditions (curing time, temperature, initiator loading) necessary to produce a high integrity composite while minimizing chemical residual and air emissions. Even though the VOC loading of the non-styrene resin (4 wt.%) was much less than that of styrene resin (39 wt.%), the non-styrene resin did contain HAP, EDR, CAR compounds including ethylbenzene, 2-ethylhexanoic acid, methacrylic acid, styrene, toluene, and <em>m</em>-xylene. Study results also revealed that by changing initiator loading a drastic reduction in the amount of styrene (-42 wt.%) and styrene oxide (-33 wt.%) residual left in the newly manufactured composite was achieved. Discoveries prompted a new hypothesis that this decreased residual also prompted a decreased amount of VOCs emitted into the air. The explanation is that this occurs because that a greater amount of the monomer styrene was incorporated into the resin during polymerization and not permitted to enter the air. Despite decades of polymer composite use, this study provides a new fundamental understanding of composite chemicals and techniques for reducing air pollutant emissions during plastic composite manufacture. In Chapter 3, the complexity of organic vapor chemicals found in the air during thermal heating of CIPP composites was explored and quantified. The emission rate of a popular monomer, styrene, was quantified from the materials before, during, and after composite manufacture. Scaling up bench-scale results, 1.9 to 14 US tons and 0.18 to 1.35 US tons of VOCs (0.05 to 0.36 US tons and 0.001 to 0.007 US tons of styrene) were estimated to be emitted during curing of styrene- and non-styrene CIPPs (i.e., typically 1-3 m of diameter pipes). By modifying standard air sampling methods, previously undetectable chemicals associated with CIPP manufacture were found in the styrene-laden air. These include acetophenone, benzaldehyde, phenol, and 1,3,5-trimethylbenzene. Results have immediate relevance to improved air monitoring for public and worker safety. Further, results can be used to examine the cumulative health and environmental risks of the CIPP pollutant mixtures. Chapter 4 focused on identifying CIPP technology/knowledge gaps and feedback from health officials from multiple state and federal agencies. Through this study, a public health workgroup was assembled to include disciplinary experts and 13 federal, state, and city health agencies and public health associations. Building on dialogue with U.S. health officials, the state of knowledge pertaining to CIPP chemical exposures, mitigation, and response actions was reviewed. Topics included 1) CIPP manufacturing process and waste; 2) sewers and buildings; 3) chemical exposure and health; 4) chemical risk assessment; 5) risk communication. This study helped establish relationships among federal, state, and city officials to improve public health response. Additionally, a primer for CIPP chemical fate and transport, as well as assisting in identifying and prioritizing public health information needs was developed. Identification and prioritization of current public health knowledge gaps and proposed practices for reducing exposures to the public and workers were reported. CIPP-related bench and research results throughout the dissertation can serve as an important basis for environmental policy and public health guidelines on the prevention and mitigation aspects of environmental and human health impacts resulting from CIPP manufacturing practices.</p>

<p dir="ltr">Atmospheric organic aerosols (OA) have diverse compositions and undergo complex reactions and transformations within the atmosphere, leading to profound impacts on air quality, climate, and atmospheric chemistry. In particular, these aerosols play an important role in Earth's effective radiative forcing (ERF) through interactions with solar radiation, absorbing and scattering sunlight and terrestrial radiation. These interactions result in a warming and cooling effect on the climate, respectively. This dissertation seeks to unravel the intricate molecular characteristics of atmospheric OA, focusing specifically on its light-absorbing components, known as ‘Brown Carbon’ (BrC), and aims to comprehend its dynamic interplay within the atmosphere. The research employs state-of-the-art multi-modal mass spectrometry techniques to investigate atmospheric OA derived from the combustion of fossil fuels and biomass burning. Through a combination of controlled laboratory experiments and real-world sample analyses, these works provide molecular-level insights crucial for source apportionment and predictive modeling of OA fate. Chapter 2 details the instrumentation and data analysis methods, laying a robust foundation for subsequent chapters.</p><p dir="ltr">Chapter 3 delves into the investigation of smoldering-phase biomass burning organic aerosols (BBOA) and introduces an innovative fractionation method for high-level molecular characterization, targeted to streamline source apportionment of BBOA. This chapter also presents an extensive assessment of particle-to-gas partitioning of BBOA, providing valuable information for modeling atmospheric lifetimes and fate. In Chapter 4, a comparative analysis of BBOA from wild and agricultural fires is conducted, employing advanced molecular characterization techniques. Chapter 5 showcases the synergistic use of multi-modal mass spectrometry techniques to probe the chemical evolution of individual BBOA components. Finally, Chapter 6 examines the molecular analysis of secondary OA (SOA) generated from the photooxidation of a fossil-fuel proxy.</p><p dir="ltr">The comprehensive molecular-level studies presented contribute essential insights for climate modeling, aiding in resolving uncertainties associated with OA's impact on global ERF. The research not only challenges existing analytical methods but also introduces novel approaches for obtaining relevant information about atmospheric OA components. Overall, this work advances our understanding of the intricate dynamics of atmospheric aerosols, facilitating more accurate climate predictions and addressing uncertainties surrounding their net radiative impact.</p>

<p dir="ltr">Ion mobility spectrometry (IMS), with its unparalleled ability to separate and filter ions based on their overall size before channeling them into a Mass Spectrometer, has placed itself as a cornerstone of the modern Analytical Chemistry field. IMS provides an orthogonal separation, aiding in the identification and analysis processes of various compounds. While there have been many inventions for ion mobility (IM) devices with exponential growth in the separation capability in the past few years, there is very little emphasis on the theoretical explanation. For example, most modern IMS devices often use a high ratio of electric field to gas concentration (E/n) as it provides better separation capabilities. However, the interaction between ion and gas at such E/n cannot be explained by current IM theories as they ignore several critical factors such as the increase in ion’s energy due to energetic collisions, the energy loss/transferred in the internal degree of freedoms, and change in the ion’s structure, requiring empirical data to identify ions after separation. The thesis presented here contributes towards bridging this gap by elucidating the complex interplay of forces and interactions that govern the ion separation process, thereby explaining on how these mechanisms can be further exploited for refined separation and advancing the computational approach to identify the separated ion.</p><p dir="ltr">To explain the ion-gas interaction under high E/n, this research extends the Two-Temperature Theory (2TT) up to the fourth order approximation. The central idea of the 2TT is to solve moments of the Boltzmann equation for the ion’s velocity distribution involving ion-gas collisions. The research shows a decreasing error between each subsequent approximations, indicating convergence. This advancement is demonstrated through the development and application of our in-house program, IMoS, and validated against experimental data for small ions in monoatomic gases. This research also justifies the mechanisms of increasing and decreasing mobility as the electric field is increased by explaining the interplay between the interaction potential and the collision energy.</p><p dir="ltr">Subsequent chapters investigate the impact of internal degrees of freedom (rotational and vibrational) on ion mobility. This includes pioneering work with the Structures for Lossless Ion Manipulations (SLIM) device to separate isotopomers, alongside computational advancements in simulating these effects, leading to the development of IMoS 2.0. In IMoS 2.0 software an ion is placed in a virtual drift tube with electric field, where it is free to rotate and translate upon collision. The research notably uncovers the role of rotational degrees of freedom in isotopomer separation, a previously underexplored area.</p><p dir="ltr">To ascertain the effect of the vibrational DoF and differentiate from the ion’s structural expansion and heating resulting from energetic collisions, a combined simulation of ion mobility and molecular dynamics (IM-MD) was performed. This analysis revealed that structural expansion plays a dominant role for the cause of deviation at high E/n, to such an extent that the vibrational DoF (or inelastic collisions) can normally be disregarded. Moreover, the research also indicates that using a combination of IM-MD simulation, one can identify accurate gas-phase structure of the ion at any temperature from a pool of probable structures.</p><p dir="ltr">Guided by these conclusions, the research now takes a significant step forward by aiming to accurately characterize protein structures in the gas phase using IM-MD simulation. Traditional MD simulations provide larger structures since the force field is not optimized for the gas-phase simulation. To address this, a biasing force towards the center of the protein is applied, compressing it. This method efficiently explores multiple feasible configurations, including those obscured by energy barriers. This strategy generated structures that closely align with the experimental evidence.</p>

<p dir="ltr">Mass spectrometry imaging (MSI) is a powerful technique for visualizing the distribution of molecules within biological samples. Advancements in MSI instrumentation and computational tools have enabled the impactful applications of this technique across various fields including clinical research, drug discovery, forensics, microbiology, and natural products. Nanospray desorption electrospray ionization (nano-DESI), an ambient localized liquid extraction ionization technique, has proven valuable to the MSI community. Nano-DESI has been used for imaging of various molecules in biological samples including drugs, metabolites, lipids, N-linked glycans, and proteins.</p><p dir="ltr">My research has been focused on expanding the sensitivity and specificity of nano-DESI for biomolecular imaging. One of the newly developed methods employs ammonium fluoride NH₄F as a solvent additive to enhance the sensitivity of nano-DESI for the analysis of lipids in negative ionization mode. Secondly, methods were developed for the spatial mapping of isobaric and isomeric species in biological tissues by implementing nano-DESI MSI on a triple quadrupole (QqQ) mass spectrometer. This work used multiple reaction monitoring (MRM) mode of a QqQ with unit mass resolution to separate isobaric lipid species that require high mass resolving power and imaging of isomeric low-abundance species in tissue sections. Next, I demonstrate nano-DESI as a liquid extraction technique for imaging of N-linked glycans within biological tissue sections. Lastly, the spatial distribution of eicosanoids and specialized pro-resolving mediators (SPMs) in a mouse model for acetaminophen-induced liver injury (AILI) provides insights into the inflammation and resolution phases of AILI. Collectively, these developments have advanced the sensitivity, chemical specificity, and molecular coverage of nano-DESI for imaging of different classes of molecules in biological tissues.</p>