Innovative Homogeneous and Heterogeneous Systems for Electrochemically Generated Luminescence Investigations: Towards One-Dimensional ECL

My research PhD work is focused on the Electrochemically Generated Luminescence (ECL) investigation of several different homogeneous and heterogeneous systems. ECL is a redox induced emission, a process whereby species, generated at electrodes, undergo a high-energy electron transfer reaction to form excited states that emit light. Since its first application, the ECL technique has become a very powerful analytical tool and has widely been used in biosensor transduction. ECL presents an intrinsically low noise and high sensitivity; moreover, the electrochemical generation of the excited state prevents scattering of the light source: for all these characteristics, it is an elective technique for ultrasensitive immunoassay detection. The majority of ECL systems involve species in solution where the emission occurs in the diffusion layer near to the electrode surface. However, over the past few years, an intense research has been focused on the ECL generated from species constrained on the electrode surface. The aim of my work is to study the behavior of ECL-generating molecular systems upon the progressive increase of their spatial constraints, that is, passing from isolated species in solution, to fluorophores embedded within a polymeric film and, finally, to patterned surfaces bearing “one-dimensional” emitting spots. In order to describe these trends, I use different “dimensions” to indicate the different classes of compounds. My thesis was mostly developed in the electrochemistry group of Bologna with the supervision of Prof Francesco Paolucci and Dr Massimo Marcaccio. With their help and also thanks to their long experience in the molecular and supramolecular ECL fields and in the surface investigations using scanning probe microscopy techniques, I was able to obtain the results herein described. Moreover, during my research work, I have established a new collaboration with the group of Nanobiotechnology of Prof. Robert Forster (Dublin City University) where I spent a research period. Prof. Forster has a broad experience in the biomedical field, especially he focuses his research on film surfaces biosensor based on the ECL transduction. This thesis can be divided into three sections described as follows: (i) in the fist section, homogeneous molecular and supramolecular ECL-active systems, either organic or inorganic species (i.e., corannulene, dendrimers and iridium metal complex), are described. Driving force for this kind of studies includes the search for new luminophores that display on one hand higher ECL efficiencies and on the other simple mechanisms for modulating intensity and energy of their emission in view of their effective use in bioconjugation applications. (ii) in the second section, the investigation of some heterogeneous ECL systems is reported. Redox polymers comprising inorganic luminophores were described. In such a context, a new conducting platform, based on carbon nanotubes, was developed aimed to accomplish both the binding of a biological molecule and its electronic wiring to the electrode. This is an essential step for the ECL application in the field of biosensors. (iii) in the third section, different patterns were produced on the electrode surface using a Scanning Electrochemical Microscopy. I developed a new methods for locally functionalizing an inert surface and reacting this surface with a luminescent probe. In this way, I successfully obtained a locally ECL active platform for multi-array application.

Continuos Flow Single Cell Separation into Open Microwell Arrays

A novel design based on electric field-free open microwell arrays for the automated continuous-flow sorting of single or small clusters of cells is presented. The main feature of the proposed device is the parallel analysis of cell-cell and cell-particle interactions in each microwell of the array. High throughput sample recovery with a fast and separate transfer from the microsites to standard microtiter plates is also possible thanks to the flexible printed circuit board technology which permits to produce cost effective large area arrays featuring geometries compatible with laboratory equipment. The particle isolation is performed via negative dielectrophoretic forces which convey the particles’ into the microwells. Particles such as cells and beads flow in electrically active microchannels on whose substrate the electrodes are patterned. The introduction of particles within the microwells is automatically performed by generating the required feedback signal by a microscope-based optical counting and detection routine. In order to isolate a controlled number of particles we created two particular configurations of the electric field within the structure. The first one permits their isolation whereas the second one creates a net force which repels the particles from the microwell entrance. To increase the parallelism at which the cell-isolation function is implemented, a new technique based on coplanar electrodes to detect particle presence was implemented. A lock-in amplifying scheme was used to monitor the impedance of the channel perturbed by flowing particles in high-conductivity suspension mediums. The impedance measurement module was also combined with the dielectrophoretic focusing stage situated upstream of the measurement stage, to limit the measured signal amplitude dispersion due to the particles position variation within the microchannel. In conclusion, the designed system complies with the initial specifications making it suitable for cellomics and biotechnology applications.

The challenges and the limitations in Life Cycle Impact Assessment for metal oxide nanoparticles, a case study on nano- TiO2

Life Cycle Assessment (LCA) is a chain-oriented tool to evaluate the environment performance of products focussing on the entire life cycle of these products: from the extraction of resources, via manufacturing and use, to the final processing of the disposed products. Through all these stages consumption of resources and pollutant releases to air, water, soil are identified and quantified in Life Cycle Inventory (LCI) analysis. Subsequently to the LCI phase follows the Life Cycle Impact Assessment (LCIA) phase; that has the purpose to convert resource consumptions and pollutant releases in environmental impacts. The LCIA aims to model and to evaluate environmental issues, called impact categories. Several reports emphasises the importance of LCA in the field of ENMs. The ENMs offer enormous potential for the development of new products and application. There are however unanswered questions about the impacts of ENMs on human health and the environment. In the last decade the increasing production, use and consumption of nanoproducts, with a consequent release into the environment, has accentuated the obligation to ensure that potential risks are adequately understood to protect both human health and environment. Due to its holistic and comprehensive assessment, LCA is an essential tool evaluate, understand and manage the environmental and health effects of nanotechnology. The evaluation of health and environmental impacts of nanotechnologies, throughout the whole of their life-cycle by using LCA methodology. This is due to the lack of knowledge in relation to risk assessment. In fact, to date, the knowledge on human and environmental exposure to nanomaterials, such ENPs is limited. This bottleneck is reflected into LCA where characterisation models and consequently characterisation factors for ENPs are missed. The PhD project aims to assess limitations and challenges of the freshwater aquatic ecotoxicity potential evaluation in LCIA phase for ENPs and in particular nanoparticles as n-TiO2.

Determinazione di radicali OH: sintesi di materiali per la modifica di elettrodi di platino e carbone vetroso

Questa tesi descrive lo sviluppo di un elettrodo modificato con un polimero isolante per la determinazione indiretta del radicale OH. I polimeri testati sono stati polifenolo, polipirrolo e polipirrolo sovraoossidato ed il primo è risultato quello con le migliori prestazioni. Il film di modificante è stato depositato per elettropolimerizzazione del fenolo in ambiente acido, su un elettrodo di carbone vetroso (GC) ed è risultato isolante e perfettamente adeso al GC, impedendo il trasferimento di carica alle più comuni sonde redox. L’attacco dei radicali OH, generati dalla reazione di Fenton o dalla fotolisi di H2O2, rimuove parzialmente il polimero dal GC, ripristinando parzialmente il comportamento conduttore dell’elettrodo. L’entità della degradazione del film polifenolico è stata valutata sfruttando la corrente relativa alla sonda redox Ru(NH3)63+, che rappresenta il segnale analitico per la determinazione del radicale OH. L’elettrodo è stato impiegato per stimare le prestazioni di foto catalizzatori a base di nanoparticelle di TiO2, ottenendo risultati correlati a quelli ricavati da un metodo HPLC. Inoltre esso è stato usato per sviluppare una nuova procedura per la determinazione della capacità di scavenging verso i radicali OH, che è stata applicata all’analisi di composti puri e campioni reali. I risultati erano confrontabili con quelli determinati con metodiche standardizzate, comunemente impiegate per la determinazione della capacità antiossidante. Inoltre è stato condotto uno studio riguardante la modifica di un elettrodo di platino con un idrossido misto a strati a base di cobalto e alluminio (LDH). In particolare si sono valutati gli effetti di diversi pretrattamenti del Pt sulle caratteristiche e prestazioni elettrocatalitiche del film di LDH nei confronti dell’ossidazione di anilina, fenolo e acido salicilico. Questi composti possono essere impiegati come molecole sonda per la determinazione del radicale OH e rivestono interesse da un punto di vista elettroanalitico perché portano facilmente alla passivazione della superficie di Pt.

Hydrodynamic induced by an array of wave energy converters. Experimental and numerical analysis

The thesis analyses the hydrodynamic induced by an array of Wave energy Converters (WECs), under an experimental and numerical point of view. WECs can be considered an innovative solution able to contribute to the green energy supply and –at the same time– to protect the rear coastal area under marine spatial planning considerations. This research activity essentially rises due to this combined concept. The WEC under exam is a floating device belonging to the Wave Activated Bodies (WAB) class. Experimental data were performed at Aalborg University in different scales and layouts, and the performance of the models was analysed under a variety of irregular wave attacks. The numerical simulations performed with the codes MIKE 21 BW and ANSYS-AQWA. Experimental results were also used to calibrate the numerical parameters and/or to directly been compared to numerical results, in order to extend the experimental database. Results of the research activity are summarized in terms of device performance and guidelines for a future wave farm installation. The device length should be “tuned” based on the local climate conditions. The wave transmission behind the devices is pretty high, suggesting that the tested layout should be considered as a module of a wave farm installation. Indications on the minimum inter-distance among the devices are provided. Furthermore, a CALM mooring system leads to lower wave transmission and also larger power production than a spread mooring. The two numerical codes have different potentialities. The hydrodynamics around single and multiple devices is obtained with MIKE 21 BW, while wave loads and motions for a single moored device are derived from ANSYS-AQWA. Combining the experimental and numerical it is suggested –for both coastal protection and energy production– to adopt a staggered layout, which will maximise the devices density and minimize the marine space required for the installation.

Electrochemical surface modification of Single walled carbon nanotubes and graphene-based electrodes for (bio)sensing applications

Sensors are devices that have shown widespread use, from the detection of gas molecules to the tracking of chemical signals in biological cells. Single walled carbon nanotube (SWCNT) and graphene based electrodes have demonstrated to be an excellent material for the development of electrochemical biosensors as they display remarkable electronic properties and the ability to act as individual nanoelectrodes, display an excellent low-dimensional charge carrier transport, and promote surface electrocatalysis. The present work aims at the preparation and investigation of electrochemically modified SWCNT and graphene-based electrodes for applications in the field of biosensors. We initially studied SWCNT films and focused on their topography and surface composition, electrical and optical properties. Parallel to SWCNTs, graphene films were investigated. Higher resistance values were obtained in comparison with nanotubes films. The electrochemical surface modification of both electrodes was investigated following two routes (i) the electrografting of aryl diazonium salts, and (ii) the electrophylic addition of 1, 3-benzodithiolylium tetrafluoroborate (BDYT). Both the qualitative and quantitative characteristics of the modified electrode surfaces were studied such as the degree of functionalization and their surface composition. The combination of Raman, X-ray photoelectron spectroscopy, atomic force microscopy, electrochemistry and other techniques, has demonstrated that selected precursors could be covalently anchored to the nanotubes and graphene-based electrode surfaces through novel carbon-carbon formation.

Tailoring properties and functionalities of TiO2 and Ag nanoparticles involved in surfaces engineering processes

The functionalization of substrates through the application of nanostructured coatings allows to create new materials, with enhanced properties. In this work, the development of self-cleaning and antibacterial textiles, through the application of TiO2 and Ag based nanostructured coatings was carried out. The production of TiO2 and Ag functionalized materials was achieved both by the classical dip-padding-curing method and by the innovative electrospinning process to obtain nanofibers doped with nano-TiO2 and nano-Ag. In order to optimize the production of functionalized textiles, the study focused on the comprehension of mechanisms involved in the photocatalytic and antibacterial processes and on the real applicability of the products. In particular, a deep investigation on the relationship between nanosol physicochemical characteristics, nanocoating properties and their performances was accomplished. Self-cleaning textiles with optimized properties were obtained by properly purifying and applying commercial TiO2 nanosol while the studies on the photocatalytic mechanism operating in self-cleaning application demonstrated the strong influence of hydrophilic properties and of interaction surface/radicals on final performance. Moreover, a study about the safety in handling of nano-TiO2 was carried out and risk remediation strategies, based on “safety by design” approach, were developed. In particular, the coating of TiO2 nanoparticles by a SiO2 shell was demonstrated to be the best risk remediation strategy in term of biological response and preserving of photoreactivity. The obtained results were confirmed determining the reactive oxygen species production by a multiple approach. Antibacterial textiles for biotechnological applications were also studied and Ag-coated cotton materials, with significant anti-bacterial properties, were produced. Finally, composite nanofibers were obtained merging biopolymer processing and sol-gel techniques. Indeed, electrospun nanofibers embedded with TiO2 and Ag NPs, starting from aqueous keratin based formulation were produced and the photocatalytic and antibacterial properties were assessed. The results confirmed the capability of electrospun keratin nanofibers matrix to preserve nanoparticle properties.

Non-thermal emission in High Frequency Peaked blazars towards the Square Kilometer Array era

In this thesis work we will explore and discuss the properties of the gamma-ray sources included in the first Fermi-LAT catalog of sources above 10 GeV (1FHL), by considering both blazars and the non negligible fraction of still unassociated gamma-ray sources (UGS, 13%). We perform a statistical analysis of a complete sample of hard gamma-ray sources, included in the 1FHL catalog, mostly composed of HSP blazars, and we present new VLBI observations of the faintest members of the sample. The new VLBI data, complemented by an extensive search of the archives for brighter sources, are essential to gather a sample as large as possible for the assessment of the significance of the correlation between radio and very high energy (E>100 GeV) emission bands. After the characterization of the statistical properties of HSP blazars and UGS, we use a complementary approach, by focusing on an intensive multi-frequency observing VLBI and gamma-ray campaign carried out for one of the most remarkable and closest HSP blazar Markarian 421.