Dipartimento di Ingegneria dell'Ambiente - Tesi di Dottorato
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Questa collezione raccoglie le Tesi di Dottorato afferenti al Dipartimento di Ingegneria per l'Ambiente e il Territorio e Ingegneria Chimica dell'Università della Calabria.
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Item <> costruzione di possibili profili di sostenibilità applicati a scala di quartiere.(2013-11-29) Manfredi, Emilia; Pantano, Pietro; Rossi, FrancescoItem Synthesis and characterization of nanomaterials: graphene, silicene and carbon nano- onions(2017-10-20) Osman, Salih Mohamed; Critelli, Salvatore; Caputi, LorenzoThe electronic structure of the graphene/Ni(111) system was investigated by means of electron energy-loss spectroscopy (EELS). A single layer of graphene has been obtained on Ni(111) by dissociation of ethylene. Angle-resolved EEL spectra show a low energy plasmon dispersing up to about 2 eV, resulting from fluctuation of a charge density located around the Fermi energy, due to hybridization between Ni and graphene states. The dispersion is typical of a two-dimensional charge layer, and the calculated Fermi velocity is a factor of ~0.5 lower than in isolated graphene. The interface-π plasmon, related to interband transitions involving hybridized states at the K point of the hexagonal Brillouin zone, has been measured at different scattering geometries. The resulting dispersion curve exhibits a square root behavior, indicating also in this case a two-dimensional character of the interface charge density. As well, it has been shown that it is possible to use EELS in the reflection mode to measure the fine structure of the carbon K-edge in monolayer graphene on Ni(111), thus demonstrating that reflection EELS is a very sensitive tool, particularly useful in cases where the TEM-based ELNES cannot be applied. Clean Ag(111) surface and the two phases of silicene on Ag(111), mixed (4×4, √13×√13R19°, 2√3×2√3R30°) and 2√3×2√3R30, have been studied by XPS, LEED and EELS. EEL spectra of the Ag(111) surface covered by silicene in the (4×4, √13×√13R19°, 2√3×2√3R30°) mixed phase shows a well-defined plasmon peak whose center is located at about 1.75 eV. The 2√3×2√3R30° phase shows EEL spectra that exhibit a peak located at about 0.75 eV loss, which moves clearly towards higher energies with increasing momentum transfer. The typical parabolic dispersion relation obtained from such spectra confirms that the peak is due to a collective excitation which is evidently associated to the silicene layer. These plasmons associated to silicene have never been observed in the past. Our results show that the plasmonic properties of silicene on Ag(111) are strongly dependent on the geometrical arrangement of Si atoms with respect to the substrate. Carbonaceous nanomaterials have been obtained by underwater arc discharge between graphite electrodes. TEM images showed that the resulting particles suspended in water consist of CNOs with other carbonaceous materials such as CNTs and graphene. We observed for the first time the formation of a solid agglomerate on the cathode surface. Raman and TEM studies revealed that the agglomerate is made exclusively of CNOs. The defragmentation of such agglomerate allows to obtain CNOs free of other carbonaceous materials without the complex purification procedures needed for floating nanomaterialsItem Preparation and Characterisation of Photocatalysts for CO2 valorisation in Membrane Reactors(2019-05-10) Pomilla, Francesca Rita; Marcì, Giuseppe; Barbieri, Giuseppe; Molinari, RaffaeleCarbon dioxide is a gas that is constantly exchanged among the atmosphere, oceans, and land surface due to its continuous production and absorption by many microorganisms, plants and animals. These processes tend to balance the CO2 content in the atmosphere; however, since the Industrial Revolution, human activities are perturbing this equilibrium causing global warming and climate change. Due to this problem, an increasing concern has bring the scientific community to devote huge efforts to the CO2 reduction and/or valorisation. The published researches demonstrate that photocatalytic reduction of CO2 in the presence of H2O as reductant is a promising green way to obtain CH4, CO, CH3OH, EtOH, HCHO, acetaldehyde, and other products. Albeit, some aspects should be still improved in the use of the current technology, mainly related to the fact that TiO2, the most known photocatalyst, absorbs light in the ultraviolet region of the electromagnetic spectrum and the use of the visible light is by far desirable. Beside the use of the best light source, also the high recombination hole-electron photogenerated charges (h+ e-) should be reduced or, in the best prospective, suppressed. The aim of the current PhD dissertation is the CO2 reduction by renewable methods, by using sunlight in order to obtain molecules that eventually could be used as fuels. This challenge aims to miming the natural processItem Innovative UV-LED polymerised bicontinuous microemulsion coating for membranes with special emphasis on MBRs(2017-07-11) Schmidt, Slefan-André; Pantano, Pietro; Curcio, Efrem; Gabriele, Bartolo; Figoli, AlbertoThe main objective of this work is the preparation of polymerisable bicontinuous microemulsion (PBM) coatings applied onto commercial membranes for improving the anti-fouling properties and performance, in terms of water flux and foulants rejection. Microstructured and nanostructured materials obtained by PBM have been widely investigated in the course of the last 30 years. The interest in microemulsion lies mainly in the possibility of dissolving larger amounts of oil and water by using polymerisable and non-polymerisable surfactants. By polymerising the bicontinuous microemulsion it is possible to produce transparent porous polymeric solids [Gan et al. (1995), Gan and Chew (1997)]. This thesis represents the follow-up of the work done by Galiano et al. (2015) and Deowan et al. (2016). Galiano et al. (2015) developed the PBM composition that based on a non-polymerisable surfactant (DTAB) and another polymerisable surfactant (AUTEAB). In their work the PBM was polymerised by redox initiators leading to a process that is very difficult to up-scale for a commercial application. Critical issues were, the polymerisation time (at least 20 minutes), and the reproducibility of the coating. Therefore, it is the aim of this work to develop another polymerisation technique that increases the polymerisation speed and allows the easy reproduction of membranes with defined properties. The polymerisation by photoinitiators excited by UV-light represents a promising possibility for this requirement as it has the potential of decreasing the polymerisation time down to a few seconds. Several photoinitiators were selected for their compatibility with the PBM, and studied for their conversion rate efficiency (section 5.1.5). As there is a wide range of potential UV-light sources available, several technologies are studied for their coating performance (section 5.1.3). Subsequent to that, experiments were done in order to define the ideal photoinitiator type and concentration while polymerising onto glass plates. The coating onto commercial membranes is studied deeply for e.g. different casting knife thickness or ambient temperature (section 5.2.2 As the polymerisation under inert conditions is expected to increase the polymerisation speed, experiments are done, both under inert and non-inert conditions. The final membrane, coated under the optimum conditions, is further characterised for their permeability under different conditions like transmembrane pressure (TMP), model foulant experiments and a fixed volume flow (section 5.2.3). Further characterisation is done by contact angle, SEM, AFM (section 5.2.5 to 5.2.7). The prepared PBM membranes are foreseen to be finally applied for model textile dye wastewater treatment by Membrane BioReactor (MBR) technology. According to the previous results of Deowan et al. (2016) higher permeate quality through increased COD, TOC, dyestuffs removal efficiency and stronger anti-fouling properties are expected. Consequently, lower operation/maintenance costs due to reduced necessary aeration for scouring purposes and reduced membrane cleaning cycles as well as less membrane replacement are of special interest for commercial applications. In the previous work of Deowan et al. (2016) a lab scaled MBR with a single membrane housing was used. As of the biocenosis of the bacteria inside the reactor tank, a comparison of the membrane performance of the PBM and commercial membrane is difficult to achieve. Therefore an existing MBR system was redesigned to allow the simultaneous run of a commercial and a PBM coated membrane (section 4.1). As the revamp requires also additional sensors, the data acquisition needs to be adapted as well. To assure the proper function of the MBR the system was running for long term with two commercial membranes using a model textile wastewater (see 5.3.2). Finally the PBM coated membrane was compared with a commercial one for their performance in the MBR. Initial experiments for the water permeability are done as preparation for future work (see 5.3.3).Item Dynamic operation and control of cell culture environments in bioreactors for bioartificial liver application(2017-07-11) Naghib, Seyed Danial; Pantano, Pietro; Di Renzo, Alberto; Curcio, Efrem; Di Maio, Francesco Paolo; De Bartolo, LoredanaOn the global scale, liver diseases are severe public health problems, with the incidences of end-stage liver disease (ESLD) rising annually. Isolated hepatocytes represent a good model of liver metabolism because they are able to perform the full range of functions. In recent years, biochemical and biotechnological engineering have been applied to the culture of human and animal hepatocyte cells, which requires the design, operation, and control of complex appropriate bioreactors. In this work, the predictable, stable and durable operation of two types of bioartificial reactors for cell cultures is investigated. The thesis is divided into the following two parts. Part I: Fluidized bed bioreactor Fluidized-bed-based biomedical devices acting as bioartificial liver, in which cells are trapped and encapsulated into appropriated fluidized beads, have proved effective solutions to many respects. However, the bioreactor performance is significantly affected by the hydrodynamics and mass transfer, not well characterized yet for most aspects. In the present work, the intrinsic and fluidization properties of alginate beads as encapsulation medium for hepatic cells are carefully analyzed experimentally using two rigs at different scales. Appropriate alginate beads were prepared and characterized in terms of size distribution and density. Expansion properties were evaluated for free alginate beads (i.e. without hepatic cells) using saline (Ringer) solutions as fluidization medium. Bed expansion tests over a wide range of voidage values have been conducted in a 1-cm diameter column, used for perfusion during in vitro experiments, as well as in a 10-cm diameter column close to human size bioreactor, in the latter case at two temperatures: ambient (20°C) and human body (37°C) conditions. Full fluid-dynamic characterization of the alginate beads is conducted, including expansion data, terminal velocity measurements, and velocity-voidage plots and their elaboration in terms of Richardson-Zaki parameters. Part II: Hollow fiber membrane bioreactor Due to their structure affine to the physiological environment in vivo, hollow fibre membrane bioreactors in crossed configuration can provide favourable conditions for the cell behaviour and metabolism. Specific devices have been proposed in recent years with very promising potential for applications. To be able to develop bioartificial systems that operate effectively and for the long term, in addition to handling the biological complexities, fluid dynamics and transport phenomena require an advance model, careful control, and appropriate automation strategies. Tight control of the culturing environment and strategies for dealing with some inherently unsteady changes of conditions in a membrane bioreactor is investigated by developing and implementing a new hydrodynamic dual control system for an existing bioreactor prototype. The experimental implementation of the sensors-controllers-actuators system is complemented by the development of a transient mathematical model of the instrumented bioreactor, in which the membrane unit is treated as a three-compartment model. A four-input/seven-state transient model of the bioreactor is obtained, able to describe the time evolution of the flowrates, the extra-capillary space liquid level and the oxygen concentration across the system. The selection of appropriate sensors and the manipulated control variables is discussed. Bioreactor dynamic simulation and control is carried out within the Matlab/Simulink environment and Matlab is also used as a platform for the experimental data digital acquisition and control logic implementation (e.g. controller tuning), allowing both for flexibility with testing of different control schemes and for direct comparison of simulated and experimental values. Different experiments with selected input changes were carried out under idealized conditions and using water as perfusing medium. The applied stimuli served to mimick causes of previously observed bioreactor malfunctions (e.g. high sensitivity to liquid level variations during prolonged cell culturing experiments) and check the control system efficacy and efficiency. Finally, the developed control system is utilized during a prolonged experiment of multi-cell culture within the membrane bioreactor, demonstrating the reliable, continuous and successful cultivation for nearly one month time. The set of results collected during the present work allows to achieve new insight into the operation and reliability of bioreactors for application as bioartificial devices, by improving the capacity to predict their behaviour and better design their structure as well as by enhancing the control over the cell culture environment conditionsItem Development of Tailored Hydrogel Composite Membranes for Application in Membrane Contactors(2017-07-11) Majidi Salehi, Shabnam; Pantano, Pietro; Curcio, Efrem; Di Profio, Gianluca; Fontananova, EnricaThis work was performed during the period from November 2013 to May 2015 in the Institute on Membrane Technology (ITM-CNR) at the University of Calabria (UNICAL), under supervision of Prof. Efrem Curcio, Dr. Gianluca Di Profio and Dr. Enrica Fontananova, from May 2015 to December 2015 at Universidade Nova de Lisboa (UNL), under supervision of Prof. Joao Crespo and from March 2016 to September 2016 at the University of Chemistry and Technology (ICT) Prague, under supervision of Dr. Eng. Vlastmil Fila. The main objective of this study was to design and develop tailored hydrogel composite membranes for application in membrane contactors, in particular, membrane distillation and membrane crystallization. Among various methods for membrane surface functionalization, surface photo-initiated graft polymerization technique (at UNICAL) and surface coating by incorporating nanoparticles (at UNL) were investigated to fabricate tailored hydrogel composite membranes In the first year at the University of Calabria, various hydrogel composite membranes were prepared by using photo-initiated polymerization method. The possibility of fine tuning the porosity and the chemical nature of hydrogels, were implemented with the preparation of composites containing diverse hydrogel components (monomer and cross-linker) and ratio among them. The selection of hydrogel components was based on the possibility to obtain homogeneous and stable composites by using specific polymeric porous membranes as supports. The resulting composite membranes were characterized by electron scanning microscopy, surface chemistry analysis, swelling degree, ion exchange capacity and water contact angle measurements Furthermore, virgin and hydrogel composite membranes were used in membrane distillation and crystallization experiments and the performance improvement was evaluated. As a result, higher water-transfer flux and enhanced ion rejection than traditional MD membranes was observed in MD treatment of saline solutions. When such HCMs used in membrane assisted crystallization of carbonate calcium (biomineralization), a wide range of crystal morphologies, most of them displaying a polycrystalline or mesocrystalline structure, was obtained in a great variety of experimental conditions. We demonstrated that this composite provides the opportunity to fine control the delivery of additives to the gel network through the porous structure of both support membrane and hydrogel layer, thus affecting crystallization kinetics, and crystal morphologies In the second year of the study at Universidade Nova de Lisboa, hydrogel composite membranes with tailored surface roughness and patterning were designed to examine the influence of the topography of such composite membranes on the growth of protein crystals. Iron oxide nanoparticles (NPs) were used as topographical designers providing a good control of membrane surface roughness and patterning. Surface morphology and topography of the prepared membranes were characterized using electron scanning microscopy, profilometry analysis and contact angle measurements. Finally, their performance was evaluated in the crystallization of Lysozyme used as a model protein and the effect of surface chemistry and topography on the heterogeneous nucleation of lysozyme crystals was investigated. We demonstrated that roughness influences crystallization, but we also show that excessive roughness may be deleterious, since it increases the number of crystals formed at the expenses of crystal size. Therefore, there is an optimum value of roughness for the formation of a low number of well-faced crystals with a larger size In the third year at the University of Chemistry and Technology Prague, the modeling of membrane crystallization was studied. The main goal of this work was to develop general model of membrane crystallization process. The development of this model involved the fundamental theories and models in membrane process and crystallization engineering, especially the models described the mass and heat transfers in membrane module and the crystal size distribution (CSD) determined by both nucleation and crystal growth processes based on the concept of the population balance equation. The experimental results of this study, allows to achieve new insight to fabricate and develop the novel hydrogel composite membranes with proper properties and novel functionality for application in membrane distillation and membrane crystallization processesItem Decoration and Characterization of Carbon-based nanomaterial for third generation photovoltaic devices(2018-03-16) Imbrogno, Alessandra; Critelli, Salvatore; Bonanno, Assunta; Macario, Anastasia; El Khakani, AlìThe PhD project is oriented on the synthesis and characterization of carbonbased nanomaterial and their eventual decoration with pulsed laser deposition technique for the developing of advanced nanomaterial suitable for photovoltaic application, in particular in DSSC devices. The dye sensitized solar cells belong to the third generation of photovoltaic devices, and are mainly composed of two electrodes deposited on FTO conductive glasses: the photoanode is also called \working electrode" (WE) and it is made of a thin lm of TiO2 deposited on a conductive FTO glass and sensitized by an organic dye, while the cathode, called \counter electrode" (CE), is made of a thin lm of platinum sputtered on a conductive FTO glass. The space between these two electrodes is lled with an electrolyte solution composed of a redox couple. The great advantage of these solar devices respect to traditional silicon-based solar devices is the relatively easy fabrication processes and the use of materials that are abundant on Earth. However, their conversion e ciency is still unsatisfactory, with conversion e ciency that barely reach the 18% for the solid-type DSSC and the 10% for the liquid-type of DSSC. The main issues that a ect the photovoltaic e ciency in DSSC are the dye deterioration, the high e-/h+ recombination in TiO2-dye substrate, the contact resistance between CE and electrolyte, and the degradation of the platinum counter electrode due to the electrolyte solution. During the last two decades many e orts have been made to resolve these issues, and some advances have been made by modify both the working and the counter electrodes. This Ph.D. project is focused on improving the materials used in both electrodes in liquid-type DSSC by using carbon nanomaterials. In particular, for what concern the counter electrode, the expensive platinum was substituted with multi walled carbon-nanotubes (MWCNT) decorated with metal nanoparticles that ensured both a good resistance to the corrosive action of the electrolyte solution and a highly rough surface that improved the catalysis of the redox reaction, resulting in a improvement of the photovoltaic performance of the DSSC device. For what concern the working electrode, instead, this Ph.D project was focused on the insertion of di erent carbon-based nanomaterials as multiwalled carbon nanotubes and graphene inside the TiO2 thin lm to reduce the loss of electron due to the e-/h+ recombination. Even in this case, the results showed interesting improvements of the photovoltaic e ciency of the DSSC device. All the experiments were conducted in both University of Calabria (Italy) and Institut National de la Recherche Scienti que (Canada).Item Mass and momentum transfer in membrane-based bioartificial liver systems(2017-07-11) Khakpour, Shervin; Pantano, Pietro; De Bartolo, Loredana; Curcio, EfremLiver failure, caused by acute or chronic end-stage liver disease (ESLD) imposes a significant disease burden worldwide. Chronic liver disease and cirrhosis is ranked as 12th cause of death in the United States and 4th in middle-aged adults. Researchers in Mayo Clinic report liver-related mortality as 8th by using a more comprehensive definition accounting for other aspects of liver disease as well. Currently, liver transplantation remains the conventional treatment for ESLD as the only medically proven method to promote patient’s health. To avoid the problem of inadequate donor organs and yet provide a comprehensive range of liver functions, cell-based therapies have been actively under investigation to potentially provide a substitute for transplantation, or a temporary support for liver failure patients. Studies on the latter aim has led to development of extracorporeal bioartificial liver (BAL) devices. Hepatic cell cultures are exploited for different applications in liver disease studies, drug toxicity testing, and bioartificial liver (BAL) devices. However, development of such systems is often hindered by the peculiar characteristics and intricate requirements of primary hepatocytes, challenging their prolonged functionality and viability in vitro. Despite the development of various 3D cell culture systems using perfused bioreactors, mass transfer properties still remain a critical and controversial topic, especially oxygen supply as the limiting and modulating factor The aim of this work is to enhance and optimize a prototype hollow fiber membrane bioreactor (HFMBR) providing efficient mass transfer for nutrient provision and catabolite removal, promoting prolonged viability and functionality of hepatocytes. In this bioreactor, two bundles of hollow fibers are employed in a crossed configuration: one bundle for supplying the oxygenated medium, and the other for removing the medium from the extra-capillary space. Optimization of the operational culture conditions to enforce an in vivo-like microenvironment is an intrinsic part of the process that requires a clear understanding of the in vitro cellular microenvironment. Oxygen transport in a convection-enhanced, crossed-configuration HFMBR hosting hepatocyte spheroids was investigated through mass transfer modelling using COMSOL Multiphysics®, a specialized, commercial finite-element software. The permeability of hollow fibers (hydraulic, albumin solution) was evaluated experimentally, showing significant, irreversible decrease in the permeance of the membranes due to protein absorption during culture period. Bioreactor’s hydrodynamics was investigated through residence time distribution analysis, by which a portion of the bioreactor was diagnosed as stagnant region. Finally, oxygen diffusion through the medium and the effect of different conditionings on the oxygen sensor’s readings in multi-well plates were studied. Mass transfer in static culture systems – both as a monolayer and as spheroids – was evaluated using a diffusion-reaction model numerically solved for oxygen (steady-state study) and urea (time-dependent study). In addition to the size and number of spheroids, sufficiency of oxygen supply to cells also depended on their distribution (the distance between them) and the amount of culture medium in each well. A convection-diffusion-reaction time distribution analysis, by which a portion of the bioreactor was diagnosed as stagnant region. Finally, oxygen diffusion through the medium and the effect of different conditionings on the oxygen sensor’s readings in multi-well plates were studied. Mass transfer in static culture systems – both as a monolayer and as spheroids – was evaluated using a diffusion-reaction model numerically solved for oxygen (steady-state study) and urea (time-dependent study). In addition to the size and number of spheroids, sufficiency of oxygen supply to cells also depended on their distribution (the distance between them) and the amount of culture medium in each well. A convection-diffusion-reaction time distribution analysis, by which a portion of the bioreactor was diagnosed as stagnant region. Finally, oxygen diffusion through the medium and the effect of different conditionings on the oxygen sensor’s readings in multi-well plates were studied. Mass transfer in static culture systems – both as a monolayer and as spheroids – was evaluated using a diffusion-reaction model numerically solved for oxygen (steady-state study) and urea (time-dependent study). In addition to the size and number of spheroids, sufficiency of oxygen supply to cells also depended on their distribution (the distance between them) and the amount of culture medium in each well. A convection-diffusion-reaction time distribution analysis, by which a portion of the bioreactor was diagnosed as stagnant region. Finally, oxygen diffusion through the medium and the effect of different conditionings on the oxygen sensor’s readings in multi-well plates were studied. Mass transfer in static culture systems – both as a monolayer and as spheroids – was evaluated using a diffusion-reaction model numerically solved for oxygen (steady-state study) and urea (time-dependent study). In addition to the size and number of spheroids, sufficiency of oxygen supply to cells also depended on their distribution (the distance between them) and the amount of culture medium in each well. A convection-diffusion-reaction model was developed to describe momentum and mass transfer in the bioreactor, in which the influential parameters were parametrized through implementation of applicable correlations. The model was numerically solved for two different types of geometries: (i) single-spheroid model using a periodic/symmetric unit cell within the bioreactor to locally represent the system decreasing the computational complexity of the model, (ii) miniaturized bioreactor model. The single-spheroid model was used to carry out a systematic parametric study to evaluate the effect of different parameters – oxygen tension (Co,sat), perfusion rate (QBR), hollow fiber spacing (δHF), spheroid diameter (Dsph), Michaelis-Menten kinetics for oxygen uptake (Vmax, Km) and porosities of the spheroid (εcc) and the membrane (εm) – on dissolved oxygen concentration (DOC) profile. Dimensionless numbers were defined for in-depth analysis of oxygen transfer and how each parameter can affect that. Among the operational conditions, Co,sat was found much more influential than QBR. Due to the mild advection added, the extra-spheroid resistances to diffusive mass transfer, i.e. the membrane (governed by εm) remains an important factor. However, εcc was found as a key intrinsic property strongly affecting intra-spheroid DOC profile. Maintaining physiological DOC range in large spheroids (Dsph=400μm) with different porosities was investigated in the single-spheroid model. Regulation of DOC profile was more manageable in spheroids with higher εcc, which lead to feasibility of achieving physiological oxygen concentrations. Low-porosity spheroids demonstrated a sharper concentration gradient, challenging sufficient oxygen supply to cells. Temporal shrinkage of spheroids due to rearrangement of cells changes the microstructure of the spheroid, the effect of which was numerically studied and proved to adversely affect the transport properties and consequently the DOC profile inside the spheroid. In the end, values from an experimental study were incorporated into the model to analyze the cellular microenvironment during the experiment, and the predictive capacity of the model regarding the outcome. Miniaturized bioreactor model was developed to reduce the computational cost while providing a more realistic model for the bioreactor. Another major advantage of this approach is capacitating investigation of the fluid dynamics inside the bioreactor. Notable DOC drop along the lumina of the supplying bundle was observed, consistent with the DOC gradient in the extra-capillary space along the supplying bundle. Having retentate flow in the hollow fibers significantly reduced these gradients and improved oxygen supply to the cells. Oxygen transfer was not noticeably affected by different flow patterns realized through using both bundles supplying or both removing the medium. However, minimization of the stagnant region had in fact a negative influence on oxygen supply. The miniaturized bioreactor model was also modified based on the experimental results for comparison with the single-spheroid model and the actual bioreactor, showing better compatibility with the real case.Item Performance of hollow fiber membrane bioreactor as a bioartificial liver(2017-07-11) Magdy Ahmed, Haysam Mohamed; Pantano, Pietro; De Bartolo, Loredana; Curcio, EfremC'è una crescente necessità di sviluppare un dispositivo bioartificiale di tipo epatico da utilizzare sia in applicazioni in vitro, per la sperimentazione della tossicità di molecole da parte delle aziende farmaceutiche, e sia in applicazioni cliniche per supportare pazienti con insufficienza epatica in attesa di trapianto di organo. A tale scopo è stato realizzato un bioreattore a membrana a fibre cave incrociate adoperante cellule epatiche umane in grado di favorire il mantenimento a lungo termine di epatociti. Il bioreattore è costituito da due fasci di membrane a fibre cave (HFM), uno deputato all’alimentazione e l’altro alla rimozione di cataboliti e prodotti specifici cellulari. I due fasci di fibre sono assemblati in una configurazione incrociata ed alternata in modo da stabilire una distanza l’una dall’altra di 250 μm. Questa configurazione del bioreattore delinea tre compartimenti separati: due compartimenti all’interno del lumen delle fibre cave dove il mezzo di coltura fluisce e un compartimento extraluminale dove le cellule sono coltivate. I compartimenti intraluminali ed extraluminale comunicano tra di loro attraverso i pori della parete di membrana. Il mezzo che fluisce nel lumen delle fibre di alimentazione permea nel compartimento cellulare, dove i cataboliti ed i metaboliti prodotti dalle cellule vengono rimossi dalle fibre cave deputate all’allontanamento dei molecole di sintesi e di scarto cellulari. In questo dispositivo le membrane a fibre cave consentono la compartimentalizzazione delle cellule in un microambiente controllato a livello molecolare ed il trasporto selettivo di molecole verso e dal compartimento cellulare proteggendo le cellule da eventuali sforzi di taglio. Inoltre, le membrane, grazie alla loro geometria intrinseca, offrono un'ampia superficie per l'adesione e la crescita delle cellule in un volume ridotto. Epatociti umani rappresentano una fonte cellulare ottimale da utilizzare nelle terapie che sono basate sull’uso di cellule, in quanto riflettono più da vicino le condizioni in vivo. In vivo gli epatociti sono altamente proliferativi all'interno del loro microambiente. Tuttavia, quando sono isolati dal loro microambiente e coltivati in vitro, perdono rapidamente le loro funzioni specifiche. Pertanto, è di importanza fondamentale la realizzazione di modelli in vitro in grado di mantenere gli epatociti vitali e funzionali per lungo tempo. Un aspetto critico è la la scarsa disponibilità di epatociti umani per cui occorre prendere in considerazione fonti cellulari alternative. Gli studi effettuati in questi ultimi anni indicano come una delle migliori fonti cellulari alternativa agli epatociti le cellule staminali, poiché queste cellule sono ampiamente disponibili possiedono in vitro un’elevata capacità proliferativa e possono essere differenziate in epatociti. A differenza delle cellule provenienti da animali e delle linee cellulari, le cellule staminali non costituiscono un rischio di trasmissione virale zoonotica o tumorigenicità. In questo lavoro, il bioreattore a membrana è stato ottimizzato al fine di creare condizioni di coltura per aggregati cellulari come sferoidi e per sistemi organotipici tridimensionali (co-coltura di epatociti e cellule non parenchimali) che garantiscano il mantenimento a lungo termine della funzionalità dei costrutti epatici umani. A tal proposito, le funzioni specifiche epatiche come l'urea, la sintesi dell'albumina e la biotrasformazione di farmaci sono state valutate nel bioreattore. I cambiamenti morfologici cellulari sono stati analizzati utilizzando il microscopio elettronico a scansione ed il microscopio confocale a scansione laser. Inoltre, il consumo di ossigeno delle cellule poste in coltura nel bioreattore è stato continuamente monitorato nel tempo al fine di assicurare un adeguato approvvigionamento di ossigeno. Gli sferoidi epatici umani, posti in coltura nello spazio extracapillare del bioreattore sono andati incontro ad un processo di fusione che ha portato alla formazione di strutture di maggiore dimensione simili a microtessuti. La fusione degli sferoidi è stata osservata sia tra le fibre che intorno alle fibre simulando il processo che avviene in vivo. Questo modello di coltura, grazie alle sue caratteristiche tridimensionali e all'aumentata interazione cellulare, così come avviene in vivo, ha favorito il mantenimento a lungo termine della vitalità e delle diverse funzioni specifiche epatiche come la sintesi di albumina ed urea ed il metabolismo xenobiotico. Allo stesso modo, nel sistema organotipico, le cellule si riorganizzano formando strutture tissutali simili a quelle del tessuto epatico in vivo. Questo è stato reso possibile grazie al piastramento sequenziale sulle membrane di cellule non parenchimali e parenchimali che hanno formato strutture stratificate tridimensionali simili a quelli in vivo. Il bioreattore che è stato ottimizzato in questo lavoro di tesi fornisce un microambiente di coltura ben controllato da un punto di vista molecolare attraverso l'alimentazione continua di sostanze nutritive, di cui una delle più importanti è l'ossigeno, e la rimozione di cataboliti. Ciò è stato confermato dai risultati relativi alla misura della concentrazione di ossigeno nel mezzo di coltura sia nella corrente in ingresso che in uscita dal bioreattore. In entrambi i modelli di coltura, l'approvvigionamento di ossigeno nel bioreattore è risultato essere sufficiente e significativamente maggiore a quello osservato in condizioni di coltura statica. Inoltre, una nuova fonte di cellule staminali, ovvero le cellule staminali mesenchimali derivate dal fegato, è stata utilizzata: le cellule sono state differenziate con successo in epatociti dopo 24 giorni di coltura, sia nel sistema statico che nel bioreattore. Tuttavia, il bioreattore ha mostrato una migliore capacità di mantenere la vitalità delle cellule e di differenziare le cellule staminali mesenchimalinel fenotipo epatico, come dimostrato dall'aumento dell'espressione genica di marcatori epatici specifici (ad es. albumina ed il fattore nucleare epatico alfa-4) e dalle velocità di sintesi di urea e albumina. Il prototipo di bioreattore realizzato su scala di laboratorio ha mantenuto con successo e funzionalmente attivi gli epatociti umani coltivati come sferoidi e in co-coltura con cellule non parenchimali per quasi un mese. Un aspetto importante è stato il differenziamento epatico delle cellule staminali mesenchimali, che rappresentano una potenziale fonte di cellule alternativa agli epatociti umani primari. Tutti questi risultati sono stati ottenuti utilizzando solo cellule umane, che convalidano le prestazioni del dispositivo che è stato sviluppato come sistema epatico bioartificiale da utilizzare in vitro. Questo bioreattore su scala di laboratorio ha un elevato potenziale applicativo cha va dagli studi in vitro delle malattie epatiche agli studi di tossicità a lungo termine. Inoltre, può essere realizzato su scala clinica ed applicato come fegato biartificiale per sostituire le funzioni epatiche di pazienti affetti da insufficienza epatica in attesa di trapianto.Item Preparation of mixed matrix membranes for water treatment(2017-07-11) Grosso, Valentina; Panano, Pietro; Drioli, Enrico; Fontananova, Enrica; Di Profio, Gianluca; Curcio, Efrem; Gabriele, BartoloThe treatment of wastewater and its reuse is a very important topic in industrial processes. This because not only avoids drawing on natural resources, but also enables a significant reduction in the amount of wastewater discharged into the natural environment. Wastewater can also be used for various purposes where drinking water quality is not mandatory, including agricultural irrigation, the cleaning of industrial equipment, the watering of green spaces, and street maintenance, etc. In fact, the water reuse has become essential in all areas in the world that suffer from water shortages [1]. Different methods are used for wastewater treatment. These processes can be to divide in: primary, secondary and tertiary treatment. Primary treatment (screening, filtration, centrifugation, sedimentation, coagulation, gravity and flotation method) includes preliminary purification processes of a physical and chemical nature while secondary treatment deals with the biological treatment of wastewater. In tertiary treatment process wastewater is converted into good quality water that can be used for different types of purpose, i.e. drinking, industrial, medicinal etc. supplies [2]. The complexity of industrial processes, the variety of pollutants and the limitation of a single operation, has led to the need for more complex processes and especially to a combination of processes. Membranes technologies falls on the tertiary water treatment technologies and are actually the most effective separation processes and they are still in rapid development creating new prospects of their applications in clean technologies [3]. The utilization of membrane operations as hybrid systems, i.e. in combination with other conventional techniques or integrated with different membrane operations is considered the way forward for more rationale applications [4]. The possibilities of redesigning various industrial cycles by combining various membrane operations have been studies and in some case realized with a low environmental impact and a low energy consumption [5]. Different processes can be used in various steps of a hybrid system, depending from the size of the pollutants to be removed. Microfiltration (MF) and ultrafiltration (UF) membrane processes, can be used as pre-treatment, while nanofiltration (NF) and reverse osmosis (RO) can used in the final step of the integrated system to remove particles with smaller dimensions (Chapter 1) The membranes have different morphological characteristics that affect their performance. The study of all the conditions which modulate these characteristics is a crucial point in the choice of membranes to be used in the various separation processes. Therefore, it is important to investigate about new materials and new types of membranes, like as mixed matrix membrane (MMM). MMM is a heterogeneous membrane consisting of inorganic fillers embedded in a polymeric matrix and can be made into flat sheets and hollow-fiber. Nevertheless, the selection of membrane configuration is greatly dependent on the application and therefore the production of MMMs in useful configuration is undoubtedly a crucial point in membrane development [6]. Also, the selection of inorganic fillers depends of desired membrane performance and their use. More attention was focus on the interesting characteristic of carbonanotubes (CNTs) (chapter 2). CNTs themselves have remarkable electrical, thermal, and mechanical properties. These nanotubes have the structure of a rolled-up graphene sheet with smaller diameter. Multiwalled carbonanotube (MWCNTs) were used to prepare MMMs for wastewater treatment. Different compounds, as additives in the polymeric membranes were used in high percentage; in the case of MWCNTs was observed as a low amount can change the membrane morphologies, mechanical and transport properties. A crucial point was the choice of membrane materials. Two type, hydrophilic poly(imide) (PI) and hydrophobic poly(vinylidenfluoride) (PVDF) were choose for membrane materials to produce MMMs. Another important point in this study was the use of functionalized MWCNTs that provide a good dispersion in the casting solution first, and in the polymeric matrix after phase separation. The main limitation in the use of CNTs is their poor dispersion in the main solvents used for the preparation of membranes. The functionalization has been proven an efficient method to overcome this limitation improving the compatibility with the polymer matrix. The presence of polar groups on the carbon nanotubes can reduces their tendency to aggregate by van der Waals interactions, while forming hydrogen bonds and electron donor/acceptor interactions with the polymer. Low percentages of CNTs were used for the preparation of membranes. These percentages were sufficient to improve better performance to modified membranes. PI was select as polymeric materials because combine easy processability in the form of membranes, with a high chemical and thermal stability, over a wide range of operative conditions. Three different PI polymers were used to prepared porous asymmetric membrane by non-solvent induced phase separation (NIPS): a homopolymer (Matrimid) and two co-polymers (Lenzing P84 and Torlon). The effect of membrane preparation conditions on the membrane morphology and transport properties, were investigate. Moreover, mixed matrix based on co-polyimide P84 and functionalized multiwalled carbon nanotubes (oxidized and aminated MWCNTs) were prepared. The different polymeric membranes were compared in the rejection of organic dyes, as model of organic pollutant present in wastewater (chapter 3). To investigate about the influence of functional groups on the MWCNTs for their interaction with polymeric matrix, three different type of functionalized MWCNTs (oxidized, amined and aminated) were dispersed also in polymeric hydrophobic PVDF membranes. PVDF was selected as polymeric materials of its outstanding properties: excellent chemical resistance and hydrolytic stability; high mechanical strength and stability over a broad pH range; polymorphism (main crystalline phases are: α, β, γ, δ and ε) [7]. The aim was to tailor the interactions with the polymeric matrix in order to realize high performing composite film with improved performance. Bovine serum albumin (BSA) protein was select as compound to evaluate the membrane performance. In particular, the antifouling properties and the permeation flux of mixed matrix membranes, were evaluate as well as thermal and structural and mechanical properties (chapter 4).