Glioblastoma multiforme (GBM) is the most common and lethal primary brain cancer with an average survival time of 15 months. GBM is considered incurable with even the most aggressive multimodal therapies and is characterized by near universal recurrence. Irreversible electroporation (IRE) is a cellular ablation method currently being investigated as a therapy for a variety of cancers. Application of IRE involves insertion of electrodes into tissue to deliver pulsed electric fields (PEFs), which destabilize the cell membrane past the point of recovery, thereby inducing cell death. While this treatment modality has numerous advantages, the lack of selectivity for malignant cells limits its application in the brain where damage to healthy tissue is especially deleterious. In this dissertation we hypothesize that a form of IRE therapy, high-frequency IRE (H-FIRE), may be able to act as a selective targeted therapy for GBM due to its ability to create an electric field inside a cell to interact with altered inner organelles. Through a comprehensive investigation involving experimental testing combined with numerical modeling, we have attained results in strong support of this hypothesis. Using tissue engineered hydrogels as our platform for therapy testing, we demonstrate selective ablation of GBM cells. We develop mathematical models that predict the majority of the electric field produced by H-FIRE pulses reach the inside of the cell. We demonstrate that the increased nuclear to cytoplasm ratio (NCR) of malignant GBM cells compared to healthy brain—evidenced in vivo and in in vitro tissue mimics—is correlated with greater ablation volumes and thus lower electric field thresholds for cell death when treated with H-FIRE. We enhance the selectivity achieved with H-FIRE using a molecularly targeted drug that induces an increase in NCR. We tune the treatment pulse parameters to increase selective malignant cell killing. Finally, we demonstrate the ability of H-FIRE to ablate therapy-resistant GBM cells which are a focus of many next-generation GBM therapies. We believe the evidence presented in this dissertation represents the beginning stages in the development of H-FIRE as a selective therapy to be used for treatment of human brain cancer. / Ph. D.

Glioblastoma Multiforme (GBM) is a highly aggressive and prevalent brain tumor with an average 5-year survival rate of approximately 6.9%. Its complex pathophysiology, characterized by the capacity to invade surrounding tissues beyond the visible tumor margin, intratumor heterogeneity, hypoxic core, and the presence of the blood-brain barrier (BBB) that restricts the penetration of large therapeutic agents, all pose formidable challenges for effective therapeutic intervention. The standard of care for GBM has thus far exhibited limited success, and patients often face a poor prognosis. Electroporation-based therapies, such as irreversible electroporation (IRE), have emerged as promising alternatives to conventional treatments. By utilizing high amplitude pulsed electric fields, IRE is able to permeabilize cells, disrupt the BBB, and induce non thermal ablation of soft tissues. However, IRE is oftentimes accompanied by undesirable secondary effects such as muscle contractions, complex anesthetic protocols, and susceptibility to electrical heterogeneities, which have impeded its clinical translation. To address these limitations, high-frequency IRE (H-FIRE) was developed. H-FIRE employs short bursts of bipolar pulses, similar in duration to the cell charging time constant, enabling the desired tissue ablation while minimizing nerve excitation and muscle contractions. Additionally, H-FIRE reduces susceptibility to electrical heterogeneities, allowing for more predictable treatment volumes, thus enhancing the feasibility of clinical translation. This dissertation investigates H-FIRE for targeting malignant gliomas while looking into improved efficacy when administering the therapy in conjunction with other treatment forms and technologies. Specifically, the presented work focuses on several key areas: (1) determining the effect of pulsing protocol and geometric configuration selection on the biological outcomes from electroporation; (2) using a tumor bearing rodent glioma model to evaluate the effects of H-FIRE as a standalone therapy and as a combinatorial therapy with liposomal doxorubicin; (3) investigating the effects of waveform shape on biological outcomes; (4) utilizing real-time Fourier Analysis SpecTroscopy (FAST) to accurately model rises in temperature during treatment; and (5) modifying real-time FAST methods to determine treatment endpoints for safe and effective ablation volumes. / Doctor of Philosophy / Glioblastoma Multiforme (GBM) is one of the deadliest tumors, with an overall five-year survival rate of approximately 6.9%. Unfortunately, it also holds the position of being the most prevalent malignant brain tumor, constituting nearly 50.1% of all primary malignant brain tumor diagnoses. Despite its widespread occurrence, there has been limited success in improving survival rates. The tumor's infiltrative nature and its location behind the blood-brain barrier (BBB), which often screens out large drug molecules like chemotherapeutics, contribute significantly to these unfavorable treatment outcomes. This dissertation explores the potential of high-frequency irreversible electroporation (H-FIRE) as a solution to these challenges. H-FIRE employs bursts of pulsed electric fields to induce nanoscale defects in the cell membrane. The response to these defects may involve temporary pores that facilitate the uptake of therapeutic molecules into the cell, or larger and longer lasting pores that disrupt cell homeostasis, ultimately leading to cell death. Furthermore, this pulsed field therapy has shown success in enabling molecules to bypass the BBB. Thus, this dissertation aims to elucidate the various biophysical phenomena associated with H-FIRE, shedding light on how to manipulate treatment protocols to maximize BBB disruption and enhance therapy when used in conjunction with combinatorial agents. Additionally, this work aims to further develop technologies to provide real-time feedback, ensuring the safe and effective delivery of the treatment. Through these efforts, this dissertation aspires to offer valuable insights into optimizing H-FIRE for the treatment of malignant gliomas and advancing the understanding of combinatorial therapies in this specific context.

This work investigates the use of irreversible electroporation (IRE) for tissue engineering applications and as a cancer ablation therapy. IRE uses short, high-intensity electric pulses to create pores in a cell's membrane and disrupt its stability. At a certain energy level, damage to the cell becomes too great and it leads to cell death. The particular mechanisms that drive this response are still not completely understood. Thus, further characterization of this behavior for cell death induced by pulsed electric fields (PEFs) will advance the understanding of these types of therapies and encourage their use to treat unresectable tumors that can benefit from the non-thermal mechanism of action which spares critical blood vessels and nerves in the surrounding area. We evaluate the response to PEFs by different cell types through experimental testing combined with computer simulations of these treatments. We show that IRE can be used to kill a specific type of bacteria that produce cellulose which can be used as an implantable material to repair damaged tissues. By killing these bacteria at particular times and locations during their cellulose production, we can create conduits in the overall structure of this material for the transport of oxygen and nutrients to the cells within the area after implantation. The use of tissue models also plays a key role in the investigation of various cancer treatments by providing a controlled environment which can mimic the state of cells within a tumor. We use tumor models comprised of a mix of collagen and cancer cells to evaluate their response to IRE based on the parameters that induce cell death and the time it takes for this process to occur. The treatment of prostate and pancreatic cancer cells with standard monopolar (only positive polarity) IRE pulses resulted in different time points for a full lesion (area of cell death) to develop for each cell type. These results indicate the presence of secondary processes within a cell that induce further cell death in the border of the lesion and cause the lesion to increase in size several hours after treatment. The use of high-frequency irreversible electroporation (H-FIRE)--comprised of short bursts of high-intensity, bipolar (both positive and negative polarity) pulses--can selectively treat cancer cells while keeping healthy cells in the neighboring areas alive. We show that H-FIRE pulses can target tumor-initiating cells (TICs) and late-stage, malignant cancer cells over non-malignant cells using a mouse ovarian cancer model representative of different stages of disease progression. To further explore the mechanisms that drive this difference in response to IRE and H-FIRE, we used more complex tumor models. Spheroids are a type of 3D cell culture model characterized by the aggregation of one or more types of cells within a single compact structure; when embedded in collagen gels, these provide cell-to-cell contact and cell-to-matrix adhesion by interactions of cells with the collagen fibers (closely mimicking the tumor microenvironment). The parameters for successful ablation with IRE and H-FIRE can be further optimized with the use of these models and the underlying mechanisms driving the response to PEFs at the cellular level can be revealed. / Ph. D. / This work investigates the use of irreversible electroporation (IRE) for tissue engineering applications and as a cancer ablation therapy. IRE uses short, high-intensity electric pulses to create pores in a cell’s membrane and disrupt its stability. At a certain energy level, damage to the cell becomes too great and it leads to cell death. The particular mechanisms that drive this response are still not completely understood. Thus, further characterization of this behavior for cell death induced by pulsed electric fields (PEFs) will advance the understanding of these types of therapies and encourage their use to treat unresectable tumors that can benefit from the non-thermal mechanism of action which spares critical blood vessels and nerves in the surrounding area. We evaluate the response to PEFs by different cell types through experimental testing combined with computer simulations of these treatments. We show that IRE can be used to kill a specific type of bacteria that produce cellulose which can be used as an implantable material to repair damaged tissues. By killing these bacteria at particular times and locations during their cellulose production, we can create conduits in the overall structure of this material for the transport of oxygen and nutrients to the cells within the area after implantation. The use of tissue models also plays a key role in the investigation of various cancer treatments by providing a controlled environment which can mimic the state of cells within a tumor. We use tumor models comprised of a mix of collagen and cancer cells to evaluate their response to IRE based on the parameters that induce cell death and the time it takes for this process to occur. The treatment of prostate and pancreatic cancer cells with standard monopolar (only positive polarity) IRE pulses resulted in different time points for a full lesion (area of cell death) to develop for each cell type. These results indicate the presence of secondary processes within a cell that induce further cell death in the border of the lesion and cause the lesion to increase in size several hours after treatment. The use of high-frequency irreversible electroporation (H-FIRE)—comprised of short bursts of high-intensity, bipolar (both positive and negative polarity) pulses—can selectively treat cancer cells while keeping healthy cells in the neighboring areas alive. We show that H-FIRE pulses can target tumor-initiating cells (TICs) and late-stage, malignant cancer cells over non-malignant cells using a mouse ovarian cancer model representative of different stages of disease progression. To further explore the mechanisms that drive this difference in response to IRE and H-FIRE, we used more complex tumor models. Spheroids are a type of 3D cell culture model characterized by the aggregation of one or more types of cells within a single compact structure; when embedded in collagen gels, these provide cell-to-cell contact and cell-to-matrix adhesion by interactions of cells with the collagen fibers (closely mimicking the tumor microenvironment). The parameters for successful ablation with IRE and H-FIRE can be further optimized with the use of these models and the underlying mechanisms driving the response to PEFs at the cellular level can be revealed.

Les champs électriques pulsés (CEP) offrent un réel intérêt dans le cadre de la traite des microalgues. En effet, ils permettent l'extraction sélective des composés hydrosolubles ou l'utilisation de solvants biocompatibles pour récolter les molécules hydrophobes. La viabilité des microalgues peut ainsi être conservée. L'objectif de ce travail de thèse est de définir des conditions de traitement permettant à la fois l'extraction des composés d'intérêt et le maintien de la viabilité des microalgues. Le modèle d'étude est la microalgue verte Haematococcus pluvialis. Au stade végétatif, celle-ci contient près d'un tiers de son poids de matière sèche en protéines et en conditions stressantes, elle accumule de l'astaxanthine, un caroténoïde à haute valeur ajoutée.L'application de CEP de 1 kV.cm-1 permet de collecter 50 % des protéines extractibles par broyage avec des microbilles. La mesure de cinq paramètres biologiques suite à ce traitement a mis en évidence que les cellules retrouvaient un état physiologique comparable à celui de microalgues non traitées au bout de 72 h. Cette condition de traitement constitue donc un bon compromis entre l'extraction des protéines et la survie des microalgues, ce qui renforce la faisabilité d'une traite de microalgues par CEP.Dans le cadre de l'extraction de l'astaxanthine, la paroi très résistante des kystes constitue le principal verrou à lever. Une optimisation des conditions de traitement (en particulier de la force des impulsions) et du stade cellulaire traité représentent les deux principales perspectives à étudier pour que l'utilisation des CEP dans le cadre de l'extraction de l'astaxanthine soit pertinente. / Pulsed electric fields (PEF) offer a real interest for microalgae milking. Indeed, they allow the selective extraction of water-soluble compounds or the use of biocompatible solvents to harvest the hydrophobic molecules. The viability of microalgae can thus be maintained. The aim of this PhD thesis work is to define the treatment conditions allowing both the extraction of compounds of interest and the maintenance of the microalgae viability. The biological model is the green microalga Haematococcus pluvialis. At the vegetative stage, it contains nearly one third of its dry matter weight in proteins and under stressful conditions, it accumulates astaxanthin, a high added value carotenoid.The application of PEF of 1 kV.cm-1 allows to collect 50% of the proteins extractable by bead milling. The measurement of five biological parameters highlights that treated cells recover a physiological state comparable to that of untreated microalgae after 72h. This treatment condition constitutes therefore a good compromise between the protein extraction and the survival of the microalgae, which reinforces the feasibility of microalgae milking by PEF.Within the context of astaxanthin extraction, the high resistance of the cell wall of the cysts constitutes the main limitation. Optimization of the treatment conditions (particularly pulse strength) and the cellular stage to treat represent the two main perspectives to study for the use of PEF for astaxanthin extraction to be relevant.

Les microalgues présentent un vrai potentiel d’innovation dans les principaux secteurs industriels tel que l’énergie, l’agroalimentaire, la cosmétique et la santé. Elles sont considérées comme étant la solution privilégiée pour répondre aux besoins énergétiques futurs et ainsi permettre une transition des énergies fossiles vers les énergies renouvelables. Néanmoins, les systèmes de production à grande échelle à partir de microalgues nécessitent encore des améliorations afin de les rendre économiquement compétitifs et durables tout en préservant l’environnement.Ainsi, l’objectif de cette thèse consiste à évaluer une nouvelle voie pour l’extraction de composés d’intérêt à partir de microalgues et de caractériser leur performance en termes d’efficacité d’extraction. L’utilisation combinée de champs électriques pulsés, et de compressions mécaniques (à travers un système microfluidique dédié) en tant que prétraitements à l’extraction de composés lipidiques, riches en énergie, produits par la microalgue Chlamydomonas reinhardtii, a donc été étudiée. Les mécanismes mis en jeu, à l’échelle de la cellule, ont été mis en évidence.Ce projet de thèse s’est déroulé dans le contexte d’une collaboration entre les laboratoires SATIE de l’ENS Paris-Saclay et LGPM de CentraleSupélec Paris-Saclay.Les résultats obtenus ont permis de confirmer le potentiel des technologies utilisées dans l’amélioration du rendement d’extraction de l’huile algale. Cette étude démontre notamment le rôle important de la paroi cellulaire de l’algue en tant qu’obstacle à une extraction optimale. Une étude approfondie de sa réponse physiologique aux prétraitements et aux conditions de stress est proposée. / Microalgae have a real potential in the innovation of the main industrial sectors such as energy, food, cosmetics and health. They are considered as a promising solution to meet future energy needs and thus enable a transition from fossil to renewable energies. Nevertheless, large scale production systems using microalgae still need improvements to become economically competitive and sustainable while preserving the environment.Thus, the aim of this thesis is to evaluate an innovative approach for the extraction of compounds of interest from microalgae and characterize their performance in terms of extraction efficiency. The effect of combining pulsed electric fields and mechanical compressions (through a dedicated microfluidic system) as pretreatments for the extraction of lipids, energy-rich compounds produced by the microalga Chlamydomonas reinhardtii, was therefore studied. The mechanisms involved, at the cellular scale, were highlighted.This project took place in the context of a collaboration between the laboratories SATIE of ENS Paris-Saclay and LGPM of CentraleSupélec Paris-Saclay.The obtained results have confirmed the potential of the technologies to improve the algal oil extraction. Furthermore, this study demonstrates the important role of the algae’s cell wall as an obstacle to an optimal extraction. A comprehensive study of the microalgae’s physiological response to pretreatments and stress conditions is proposed.

Irreversible electroporation (IRE) is a non-thermal tissue ablation modality in which electrical pulses are used to generate targeted disruption of cellular membranes. Clinically, IRE is administered by inserting one or more needles within or around a region of interest, then applying a series of short, high amplitude pulsed electric fields (PEFs). The treatment effect is dictated by the local field magnitude, which is quite high near the electrodes but dissipates exponentially. When cells are exposed to fields of sufficient strength, nanoscale "pores" form in the membrane, allowing ions and macromolecules to rapidly travel into and out of the cell. If enough pores are generated for a substantial amount of time, cell homeostasis is disrupted beyond recovery and cells eventually die. Due to this unique non-thermal mechanism, IRE generates targeted cell death without injury to extracellular proteins, preserving tissue integrity. Thus, IRE can be used to treat tumors precariously positioned near major vessels, ducts, and nerves. Since its introduction in the late 2000s, IRE has been used successfully to treat thousands of patients with focal, unresectable malignancies of the pancreas, prostate, liver, and kidney. It has also been used to decellularize tissue and is gaining attention as a cardiac ablation technique. Though IRE opened the door to treating previously inoperable tumors, it is not without limitation. One drawback of IRE is that pulse delivery results in intense muscle contractions, which can be painful for patients and causes electrodes to move during treatment. To prevent contractions in the clinic, patients must undergo general anesthesia and temporary pharmacological paralysis. To alleviate these concerns, high-frequency irreversible electroporation (H-FIRE) was introduced. H-FIRE improves upon IRE by substituting the long (~100 µs) monopolar pulses with bursts of short (~1 µs) bipolar pulses. These pulse waveforms substantially reduce the extent of muscle excitation and electrochemical effects. Within a burst, each pulse is separated from its neighboring pulses by a short delay, generally between 1 and 5 µs. Since its introduction, H-FIRE burst waveforms have generally been constructed simply by choosing the duration of constitutive pulses within the burst, with little attention given to this delay. This is quite reasonable, as it has been well documented that pulse duration plays a critical role in determining ablation size. In this dissertation, we explore the role of these latent periods within burst waveforms as well as their interaction with other pulse parameters. Our central hypothesis is that tuning the latent periods will allow for improved ablation size with reduced muscle contractions over traditional waveforms. After gaining a simple understanding of how pulse width and delay interact in vitro, we demonstrate theoretically that careful tuning of the delay within (interphase) and between (interpulse) bipolar pulses in a burst can substantially reduce nerve excitation. We then analyze how pulse duration, polarity, and delays affect the lethality of burst waveforms toward determining the most optimal parameters from a clinical perspective. Knowing that even the most ideal waveform will require slightly increased voltages over what is currently used clinically, we compare the clinical efficacy of two engineered thermal mitigation strategies to determine what probe design modifications will be needed to successfully translate H-FIRE to the clinic while maintaining large, non-thermal ablation volumes. Finally, we translate these findings in two studies. First, we demonstrate that burst waveforms with an improved delay structure allow for enhanced safety and larger ablation volumes in vivo. And finally, we examine the efficacy of H-FIRE in spontaneous canine liver tumors while also comparing the ablative effect of H-FIRE in tumor and non-neoplastic tissue in a veterinary clinical setting. / Doctor of Philosophy / Cancer is soon to become the most common cause of death in the United States. In 2023, approximately 2 million new cases of cancer will be diagnosed, leading to roughly 650 thousand lost lives. Interestingly, about half of newly diagnosed cancers are caught in the early stages before the disease has spread throughout the body. With effective local intervention, these patients could potentially be cured of their malignancy. Surgical removal of the tumor is the gold standard, but it is often not possible due to tumor location, patient comorbidities, or organ health status. In some instances, focal thermal ablation with radiofrequency or microwave energy can be performed when resection is not possible. These treatments entail the delivery of thermal energy through a needle electrode, which causes local tissue damage through coagulation (cooking) of the tissue. However, thermal ablation destroys tissue indiscriminately, meaning that any nearby blood vessels or neural components will also be damaged, which precludes thousands of patients from treatment each year. Irreversible electroporation (IRE) was introduced to overcome these challenges and provide a treatment option for patients diagnosed with otherwise untreatable tumors. IRE uses pulsed electric fields to generate nanoscale pores in cell membranes, which lead to a homeostatic imbalance and cell death. Because IRE is a membrane-based effect, it does not rely on thermal effects to generate cellular injury, which allows it to be administered to tumors that are adjacent to critical tissue structures such as major nerves and vasculature. Though IRE opened the door to treating otherwise inoperable tumors, procedures are technically challenging and require specialized anesthesia protocols. High-frequency irreversible electroporation (H-FIRE) was introduced by our group roughly a decade ago to simplify the procedure through the use of an alternate pulsing strategy. These higher frequency pulses offer several advantages such as limiting muscle contractions and reducing the risk of cardiac interference, both of which were concerns with IRE. However, H-FIRE ablations have been limited in size, and there is limited knowledge regarding the optimal pulsing strategy needed in order to maximize the ratio of therapeutic benefits to undesirable side effects like muscle stimulation and Joule heating. In this dissertation, we sought to understand how different pulse parameters affect these outcomes. Using a combination of computational, benchtop, and in vivo experiments, we comprehensively characterized the behavior of user-tunable pulse parameters and identified optimal methods for constructing H-FIRE protocols. We then translated our findings in a proof-of-principle study to demonstrate the ability of newly introduced waveform designs to increase ablation size with H-FIRE. Overall, this dissertation improves our understanding of how H-FIRE waveform selection affects clinical outcomes, introduces a new strategy for maximizing therapeutic outcomes with minimal side effects, and provides a framework for selecting parameters for specific applications.

Ce travail de thèse porte sur l’étude de l’effet du traitement par champs électriques pulsés (CEP) sur différents procédés de séchage et de friture à partir de produits végétaux (pommes de terre et carottes). Les interactions entre différents modes de séchage et de prétraitement ont été étudiées. L’impact du traitement par CEP et du pré-séchage par l’air chaud ou pré-séchage par le vide sur la cinétique de friture et sur la qualité des produits frits ont été analysés. Le prétraitement par CEP entraîne une électro-perméabilisation des membranes cellulaires, ce qui favorise l’accélération des cinétiques de perte en masse (humidité). Les résultats montrent que le temps de séchage a été réduit significativement dans tous les procédés étudiés (séchage par l’air chaud, séchage par microonde, séchage sous vide). L’avantage du traitement par CEP se manifeste également par une diminution au niveau de la température interne du produit séché. Cette température basse présente un avantage notable dans la préservation des composés sensibles à la chaleur (caroténoïdes…). La couleur des échantillons prétraités par CEP puis séchés, gardent mieux la coloration initiale et présente une déviation réduite en termes de couleur après réhydratation. En ce qui concerne le procédé de friture, l’application du traitement par CEP montrent un avantage en termes de temps de friture mais également en termes de la teneur en huile absorbée. En effet, cette teneur en huile est moins élevée pour le cas des échantillons traités électriquement comparés aux échantillons non traités. La combinaison du CEP et du pré-séchage à l’air chaud (ou du pré-séchage sous vide) montre une réduction importante du temps de friture et également en terme de teneur en huile absorbée. / This research project focuses on the effect of pulsed electric field (PEF) treatment on various drying and frying processes from plant products (potatoes and carrots). Interactions between different drying modes and pretreatment have been studied. The impact of PEF treatment and pre-drying by hot air or pre-drying by vacuum drying on frying kinetics and the quality of fried products were analyzed. PEF pretreatment results in electro-permeabilization of the cell membranes, which favors the acceleration of mass transfer processes. The results showed that the drying time was significantly reduced in all processes (hot air drying, microwave drying, vacuum drying). The advantage of the PEF treatment was also manifested by a decrease of the internal temperature of the product during drying. This lower temperature has a significant advantage in the preservation of heat-sensitive compounds (carotene, etc.). The dried sample pretreated by PEF could better retain the initial product color and had a reduced color deviation after rehydration. In regards to the frying process, the application of the PEF treatment showed not only an advantage in terms of the frying time but also in terms of oil content absorbed. The oil content of PEF treated sample was lower compared to untreated ones. Moreover, the combination of the PEF pretreatment and hot air pre-drying (or vacuum pre-drying) showed a synergistic efficiency on frying time and also in terms of oil content absorbed.

The application of non-thermal processes pulsed electric fields (PEF) and cross-flow micro-filtration (CFMF) continuous to be studied with the purpose of controlling microorganisms in milk. Trends suggesting increased adoption include the study of Food Safety Objectives as a safety criterion, the promotion of sustainable processing, and the implementation of hurdle strategies. While the advance of gentle processing is counteracted by the risk of enhanced resistance due to microbial stress response, several techniques can be applied to quantitatively assess its impact. The objective of this project was to evaluate the effectiveness of microbial inactivation by PEF and CFMF at various steps of milk processing including shelf-life, its comparison with high temperature short time (HTST) pasteurization, and the quantitative assessment of the cross protection resistance to PEF of Escherichia coli O157:H7. Some differences in mesophilics inactivation were observed in milks (fat contents between 1.1% and 3.1%). Increasing the PEF inlet temperature decreased the treatment time by three or two-fold. The combination of CFMF/PEF yielded similar microbial reductions as CFMF/HTST. Higher inactivation of the coliforms was achieved in homogenized cream (12% fat) compared to non-homogenized. The linear relation between electrical conductivity and nutrient content (fat and solids content) was established. In a parallel study the PEF/CFMF sequence resulted in higher inactivation of mesophilics compared to CFMF/PEF and HTST. The shelf life was acceptable for CFMF/PEF and HTST after 7 days, while enterics and psychrotrophs grew more after PEF/CFMF, thermodurics did after HTST. The growth and stress of Escherichia coli O157:H7 in lactose containing broths was monitored by absorbance and fluorescence expression of stress reporters. Growth was explained using a secondary model, and stress response using mechanistic and probabilistic models. PEF inactivation was evaluated following the Weibull distribution after the cells reached stationary phase or maximum fluorescence expression. Similar resistances were observed within the cells grown in lactose broth at 10, 25 or 40°C, as within stressed cells (starved or cold shocked). Cells grown at 45 °C were more resistant compared to the cells grown in acid, high salt concentration while the ones grown at cold temperatures were the weakest. / Dairy Farmers of Ontario, Natural Sciences and Engineering Research Council.

Electrons are weakly attracted to liquid helium due to the small but finite polarizability of helium atoms. However, they cannot enter the liquid unless their energy is more than 1 eV, due to the Pauli exclusion principle. As a result, electrons are bound perpendicular to the surface but are free to move parallel to the surface i.e., they form a two-dimensional electron system (2DES). If the electron density of the 2DES is increased above a critical value ( 1013 per m2) the surface becomes un-stable leading to the formation of charged bubbles known as multielectron bubbles (MEBs). In MEBs the electrons are confined to the inner bubble surface and hence we have a 2DES on a curved surface. The critical density of electrons on the bulk surface is too low to study the quantum dominated phases of the 2DES. In contrast, due to the enormous surface defects and impurities, the electronic density of 2DES in semiconductors cannot be lowered below 1015 per m2, which is high enough such that the 2DES is always in a quantum liquid phase. Alternatively, the possibility of varying the electron density over a wide range and the effects of curvature implies that MEBs can be used to probe new phases of 2DES like Wigner crystallization with strong electron-ripplon coupling, quantum melting, superconductivity etc. In this thesis the experiments done on MEBs in liquid helium are described. In the initial experiments we generated MEBs which were observed to shrink in size. We saw a difference in their collapse behaviour: MEBs in super fluid helium though initially bigger in size collapse much faster than MEBs generated in normal fluid. The vapour present in the MEBs cannot condense fast in normal fluid due to the lower thermal conductivity. In subsequent experiments, we could trap these MEBs, generated in normal fluid and stabilised by their vapour content, in a linear Paul trap. We measured the charge and radius of these trapped MEBs by analysing their dynamics. Interestingly, the stably trapped MEBs were found not to lose charge as they shrink and disappear in hundreds of milliseconds, implying that the charge density inside increases at least two orders of magnitude from the initial value. MEBs so trapped can be used to study the properties of 2DES in the high electron density limit where the quantum confinement energy dominates. Later, we measured the charge of the MEB with respect to time when it was held on a solid substrate. We propose a charge loss mechanism as the tunneling of electrons across a thin lm of helium formed between the MEB and the substrate. We estimated the density of electrons on this thin lm by using a numerical model. We found that the maximum electron density (about a few 1015 per m2) which could be held on a thin lm is limited by tunneling. Moreover, the substrate surface roughness did not affect the charge loss due to the microscopic contact of MEBs with the substrate, resolving the complications in charge loss observed in previous experiments on macroscopic thin films on metallic substrates. Finally, we describe the experiments and the results on the stability of MEBs generated in super fluid helium. Highly charged MEBs (with more than 104 electrons which have an equilibrium radius that is easily visible) are found to be unstable against fission into smaller bubble showing a type of electro-hydrodynamic instability. However, the stability of bubbles with radius less than our detection limit ( 1 m) is still an open question.

Dans cette thèse, nous proposons et étudions des systèmes d’exposition en vue d’explorer les effets biologiques sanitaire et thérapeutique des ondes électromagnétiques sur le vivant. Nous proposons une antenne micro-onde pour l’ablation thermique des tumeurs cancéreuses du foie à 2.45 GHz. Son originalité réside en ses dimensions miniatures et la possibilité de l’insérer dans le foie par voie endoscopique. Pour cette antenne, un débit d’absorption spécifique (DAS) supérieur à 50 W/kg/W inc a montré une zone exposée de 1-cm de diamètre. Nous proposons ensuite une cellule transverse électromagnétique (TEM) avec une ouverture fermée par un matériau transparent conducteur l’Indium tin oxyde (ITO). Cette cellule TEM peut être utilisée pour évaluer les effets sanitaires potentiels des signaux de télécommunications sans fils. Ce système permet l’observation microscopique en temps réel du milieu biologique exposé, à travers son ouverture fermée par l’ITO. L’influence de la présence de l’ouverture et de la couche d’ITO sur le DAS dans le milieu exposé a été évaluée. Les valeurs du DAS obtenues à 1.8 GHz dans le milieu exposé dans la cellule TEM avec l'ouverture fermée ou non par l’ITO étaient de 1.1 W/kg/W inc et 23.6 W/kg/W inc, respectivement. Une excellence homogénéité du DAS a été obtenue dans le milieu en présence de l’ITO. Enfin, nous proposons plusieurs dispositifs spécifiques pour l’exposition des cellules biologiques aux champs électriques pulsés nanosecondes de haute intensité (nsPEFs). Les effets biologiques des nsPEFs sont utilisés pour des applications dans le domaine médical et en biotechnologie. Nous proposons deux dispositifs à électrodes en contact direct avec le milieu biologique et trois dispositifs à électrodes isolées. Nous démontrons l’adaptation de ces dispositifs aux impulsions courtes de durée 3-ns et la capacité de ceux à électrodes en contact à fournir des champs intenses de l’ordre de quelques MV/m. Nous illustrons aussi l’importance des dispositifs isolés pour délivrer des impulsions ultracourtes. / In this thesis, we propose and study exposure systems to explore healthy and therapeutic biological effects of EM signals. We propose a microwave antenna for thermal ablation of liver tumors at 2.45 GHz. Its original feature consists in its reduced dimensions that permits the endoscopic insertion in the zone to be treated. For this antenna, a specific absorption rate (SAR) greater than 50 W/kg/W inc showed an exposed zone of 1-cm diameter. We propose a transverse electromagnetic cell (TEM) with an aperture sealed with a transparent conducting material Indium tin oxide (ITO).This TEM cell can be used to study the potential effects of wireless communication systems on biological cells. This delivery device allows real-time observation of biological cells during exposure across the aperture sealed with ITO. The effect of the aperture and the ITO layer presence on the SAR in the exposed sample was evaluated. The SAR values obtained at 1.8 GHz in the sample exposed in the TEM cell with the sealed or non-sealed aperture of 20-mm diameter were 1.1 W/kg/W inc and 23.6 W/kg/W inc, respectively. An excellent homogeneity of SAR was achieved in the medium in the presenceof ITO. Finally, we propose several devices for the exposure of biological medium to nanosecond pulsed electric field with high intensity (nsPEFs). The biological effect of nsPEFs are used in biotechnology and medicine. We propose two devices with electrodes in direct contact with the biological medium and three devices with isolated electrodes. We demonstrate their adaptation for 3-ns duration pulses and the suitability of those with electrodes in contactwith the biological medium to provide high intensities fields in the order of several MV/m. We demonstrate the importance of the isolated devices for delivering ultrashort pulses.