Application of chromatographic and spectroscopic techniques in the evaluation of the lipid fraction of animal products

Lipolysis and oxidation of lipids in foods are the major biochemical and chemical processes that cause food quality deterioration, leading to the characteristic, unpalatable odour and flavour called rancidity. In addition to unpalatability, rancidity may give rise to toxic levels of certain compounds like aldehydes, hydroperoxides, epoxides and cholesterol oxidation products. In this PhD study chromatographic and spectroscopic techniques were employed to determine the degree of rancidity in different animal products and its relationship with technological parameters like feeding fat sources, packaging, processing and storage conditions. To achieve this goal capillary gas chromatography (CGC) was employed not only to determine the fatty acids profile but also, after solid phase extraction, the amount of free fatty acids (FFA), diglycerides (DG), sterols (cholesterol and phytosterols) and cholesterol oxidation products (COPs). To determine hydroperoxides, primary products of oxidation and quantify secondary products UV/VIS absorbance spectroscopy was applied. Most of the foods analysed in this study were meat products. In actual fact, lipid oxidation is a major deterioration reaction in meat and meat products and results in adverse changes in the colour, flavour and texture of meat. The development of rancidity has long recognized as a serious problem during meat handling, storage and processing. On a dairy product, a vegetal cream, a study of lipid fraction and development of rancidity during storage was carried out to evaluate its shelf-life and some nutritional features life saturated/unsaturated fatty acids ratio and phytosterols content. Then, according to the interest that has been growing around functional food in the last years, a new electrophoretic method was optimized and compared with HPLC to check the quality of a beehive product like royal jelly. This manuscript reports the main results obtained in the five activities briefly summarized as follows: 1) comparison between HPLC and a new electrophoretic method in the evaluation of authenticity of royal jelly; 2) study of the lipid fraction of a vegetal cream under different storage conditions; 3) study of lipid oxidation in minced beef during storage under a modified atmosphere packaging, before and after cooking; 4) evaluation of the influence of dietary fat and processing on the lipid fraction of chicken patties; 5) study of the lipid fraction of typical Italian and Spanish pork dry sausages and cured hams.

The VIMOS-VLT Deep Survey: the evolution of type-1 AGN

Quasars and AGN play an important role in many aspects of the modern cosmology. Of particular interest is the issue of the interplay between AGN activity and formation and evolution of galaxies and structures. Studies on nearby galaxies revealed that most (and possibly all) galaxy nuclei contain a super-massive black hole (SMBH) and that between a third and half of them are showing some evidence of activity (Kormendy and Richstone, 1995). The discovery of a tight relation between black holes mass and velocity dispersion of their host galaxy suggests that the evolution of the growth of SMBH and their host galaxy are linked together. In this context, studying the evolution of AGN, through the luminosity function (LF), is fundamental to constrain the theories of galaxy and SMBH formation and evolution. Recently, many theories have been developed to describe physical processes possibly responsible of a common formation scenario for galaxies and their central black hole (Volonteri et al., 2003; Springel et al., 2005a; Vittorini et al., 2005; Hopkins et al., 2006a) and an increasing number of observations in different bands are focused on collecting larger and larger quasar samples. Many issues remain however not yet fully understood. In the context of the VVDS (VIMOS-VLT Deep Survey), we collected and studied an unbiased sample of spectroscopically selected faint type-1 AGN with a unique and straightforward selection function. Indeed, the VVDS is a large, purely magnitude limited spectroscopic survey of faint objects, free of any morphological and/or color preselection. We studied the statistical properties of this sample and its evolution up to redshift z 4. Because of the contamination of the AGN light by their host galaxies at the faint magnitudes explored by our sample, we observed that a significant fraction of AGN in our sample would be missed by the UV excess and morphological criteria usually adopted for the pre-selection of optical QSO candidates. If not properly taken into account, this failure in selecting particular sub-classes of AGN could, in principle, affect some of the conclusions drawn from samples of AGN based on these selection criteria. The absence of any pre-selection in the VVDS leads us to have a very complete sample of AGN, including also objects with unusual colors and continuum shape. The VVDS AGN sample shows in fact redder colors than those expected by comparing it, for example, with the color track derived from the SDSS composite spectrum. In particular, the faintest objects have on average redder colors than the brightest ones. This can be attributed to both a large fraction of dust-reddened objects and a significant contamination from the host galaxy. We have tested these possibilities by examining the global spectral energy distribution of each object using, in addition to the U, B, V, R and I-band magnitudes, also the UV-Galex and the IR-Spitzer bands, and fitting it with a combination of AGN and galaxy emission, allowing also for the possibility of extinction of the AGN flux. We found that for 44% of our objects the contamination from the host galaxy is not negligible and this fraction decreases to 21% if we restrict the analysis to a bright subsample (M1450 <-22.15). Our estimated integral surface density at IAB < 24.0 is 500 AGN per square degree, which represents the highest surface density of a spectroscopically confirmed sample of optically selected AGN. We derived the luminosity function in B-band for 1.0 < z < 3.6 using the 1/Vmax estimator. Our data, more than one magnitude fainter than previous optical surveys, allow us to constrain the faint part of the luminosity function up to high redshift. A comparison of our data with the 2dF sample at low redshift (1 < z < 2.1) shows that the VDDS data can not be well fitted with the pure luminosity evolution (PLE) models derived by previous optically selected samples. Qualitatively, this appears to be due to the fact that our data suggest the presence of an excess of faint objects at low redshift (1.0 < z < 1.5) with respect to these models. By combining our faint VVDS sample with the large sample of bright AGN extracted from the SDSS DR3 (Richards et al., 2006b) and testing a number of different evolutionary models, we find that the model which better represents the combined luminosity functions, over a wide range of redshift and luminosity, is a luminosity dependent density evolution (LDDE) model, similar to those derived from the major Xsurveys. Such a parameterization allows the redshift of the AGN density peak to change as a function of luminosity, thus fitting the excess of faint AGN that we find at 1.0 < z < 1.5. On the basis of this model we find, for the first time from the analysis of optically selected samples, that the peak of the AGN space density shifts significantly towards lower redshift going to lower luminosity objects. The position of this peak moves from z 2.0 for MB <-26.0 to z 0.65 for -22< MB <-20. This result, already found in a number of X-ray selected samples of AGN, is consistent with a scenario of “AGN cosmic downsizing”, in which the density of more luminous AGN, possibly associated to more massive black holes, peaks earlier in the history of the Universe (i.e. at higher redshift), than that of low luminosity ones, which reaches its maximum later (i.e. at lower redshift). This behavior has since long been claimed to be present in elliptical galaxies and it is not easy to reproduce it in the hierarchical cosmogonic scenario, where more massive Dark Matter Halos (DMH) form on average later by merging of less massive halos.

Statistical properties of Radio Halos and the re-acceleration model

Galaxy clusters occupy a special position in the cosmic hierarchy as they are the largest bound structures in the Universe. There is now general agreement on a hierarchical picture for the formation of cosmic structures, in which galaxy clusters are supposed to form by accretion of matter and merging between smaller units. During merger events, shocks are driven by the gravity of the dark matter in the diffuse barionic component, which is heated up to the observed temperature. Radio and hard-X ray observations have discovered non-thermal components mixed with the thermal Intra Cluster Medium (ICM) and this is of great importance as it calls for a “revision” of the physics of the ICM. The bulk of present information comes from the radio observations which discovered an increasing number of Mpcsized emissions from the ICM, Radio Halos (at the cluster center) and Radio Relics (at the cluster periphery). These sources are due to synchrotron emission from ultra relativistic electrons diffusing through µG turbulent magnetic fields. Radio Halos are the most spectacular evidence of non-thermal components in the ICM and understanding the origin and evolution of these sources represents one of the most challenging goal of the theory of the ICM. Cluster mergers are the most energetic events in the Universe and a fraction of the energy dissipated during these mergers could be channelled into the amplification of the magnetic fields and into the acceleration of high energy particles via shocks and turbulence driven by these mergers. Present observations of Radio Halos (and possibly of hard X-rays) can be best interpreted in terms of the reacceleration scenario in which MHD turbulence injected during these cluster mergers re-accelerates high energy particles in the ICM. The physics involved in this scenario is very complex and model details are difficult to test, however this model clearly predicts some simple properties of Radio Halos (and resulting IC emission in the hard X-ray band) which are almost independent of the details of the adopted physics. In particular in the re-acceleration scenario MHD turbulence is injected and dissipated during cluster mergers and thus Radio Halos (and also the resulting hard X-ray IC emission) should be transient phenomena (with a typical lifetime <» 1 Gyr) associated with dynamically disturbed clusters. The physics of the re-acceleration scenario should produce an unavoidable cut-off in the spectrum of the re-accelerated electrons, which is due to the balance between turbulent acceleration and radiative losses. The energy at which this cut-off occurs, and thus the maximum frequency at which synchrotron radiation is produced, depends essentially on the efficiency of the acceleration mechanism so that observations at high frequencies are expected to catch only the most efficient phenomena while, in principle, low frequency radio surveys may found these phenomena much common in the Universe. These basic properties should leave an important imprint in the statistical properties of Radio Halos (and of non-thermal phenomena in general) which, however, have not been addressed yet by present modellings. The main focus of this PhD thesis is to calculate, for the first time, the expected statistics of Radio Halos in the context of the re-acceleration scenario. In particular, we shall address the following main questions: • Is it possible to model “self-consistently” the evolution of these sources together with that of the parent clusters? • How the occurrence of Radio Halos is expected to change with cluster mass and to evolve with redshift? How the efficiency to catch Radio Halos in galaxy clusters changes with the observing radio frequency? • How many Radio Halos are expected to form in the Universe? At which redshift is expected the bulk of these sources? • Is it possible to reproduce in the re-acceleration scenario the observed occurrence and number of Radio Halos in the Universe and the observed correlations between thermal and non-thermal properties of galaxy clusters? • Is it possible to constrain the magnetic field intensity and profile in galaxy clusters and the energetic of turbulence in the ICM from the comparison between model expectations and observations? Several astrophysical ingredients are necessary to model the evolution and statistical properties of Radio Halos in the context of re-acceleration model and to address the points given above. For these reason we deserve some space in this PhD thesis to review the important aspects of the physics of the ICM which are of interest to catch our goals. In Chapt. 1 we discuss the physics of galaxy clusters, and in particular, the clusters formation process; in Chapt. 2 we review the main observational properties of non-thermal components in the ICM; and in Chapt. 3 we focus on the physics of magnetic field and of particle acceleration in galaxy clusters. As a relevant application, the theory of Alfv´enic particle acceleration is applied in Chapt. 4 where we report the most important results from calculations we have done in the framework of the re-acceleration scenario. In this Chapter we show that a fraction of the energy of fluid turbulence driven in the ICM by the cluster mergers can be channelled into the injection of Alfv´en waves at small scales and that these waves can efficiently re-accelerate particles and trigger Radio Halos and hard X-ray emission. The main part of this PhD work, the calculation of the statistical properties of Radio Halos and non-thermal phenomena as expected in the context of the re-acceleration model and their comparison with observations, is presented in Chapts.5, 6, 7 and 8. In Chapt.5 we present a first approach to semi-analytical calculations of statistical properties of giant Radio Halos. The main goal of this Chapter is to model cluster formation, the injection of turbulence in the ICM and the resulting particle acceleration process. We adopt the semi–analytic extended Press & Schechter (PS) theory to follow the formation of a large synthetic population of galaxy clusters and assume that during a merger a fraction of the PdV work done by the infalling subclusters in passing through the most massive one is injected in the form of magnetosonic waves. Then the processes of stochastic acceleration of the relativistic electrons by these waves and the properties of the ensuing synchrotron (Radio Halos) and inverse Compton (IC, hard X-ray) emission of merging clusters are computed under the assumption of a constant rms average magnetic field strength in emitting volume. The main finding of these calculations is that giant Radio Halos are naturally expected only in the more massive clusters, and that the expected fraction of clusters with Radio Halos is consistent with the observed one. In Chapt. 6 we extend the previous calculations by including a scaling of the magnetic field strength with cluster mass. The inclusion of this scaling allows us to derive the expected correlations between the synchrotron radio power of Radio Halos and the X-ray properties (T, LX) and mass of the hosting clusters. For the first time, we show that these correlations, calculated in the context of the re-acceleration model, are consistent with the observed ones for typical µG strengths of the average B intensity in massive clusters. The calculations presented in this Chapter allow us to derive the evolution of the probability to form Radio Halos as a function of the cluster mass and redshift. The most relevant finding presented in this Chapter is that the luminosity functions of giant Radio Halos at 1.4 GHz are expected to peak around a radio power » 1024 W/Hz and to flatten (or cut-off) at lower radio powers because of the decrease of the electron re-acceleration efficiency in smaller galaxy clusters. In Chapt. 6 we also derive the expected number counts of Radio Halos and compare them with available observations: we claim that » 100 Radio Halos in the Universe can be observed at 1.4 GHz with deep surveys, while more than 1000 Radio Halos are expected to be discovered in the next future by LOFAR at 150 MHz. This is the first (and so far unique) model expectation for the number counts of Radio Halos at lower frequency and allows to design future radio surveys. Based on the results of Chapt. 6, in Chapt.7 we present a work in progress on a “revision” of the occurrence of Radio Halos. We combine past results from the NVSS radio survey (z » 0.05 − 0.2) with our ongoing GMRT Radio Halos Pointed Observations of 50 X-ray luminous galaxy clusters (at z » 0.2−0.4) and discuss the possibility to test our model expectations with the number counts of Radio Halos at z » 0.05 − 0.4. The most relevant limitation in the calculations presented in Chapt. 5 and 6 is the assumption of an “averaged” size of Radio Halos independently of their radio luminosity and of the mass of the parent clusters. This assumption cannot be released in the context of the PS formalism used to describe the formation process of clusters, while a more detailed analysis of the physics of cluster mergers and of the injection process of turbulence in the ICM would require an approach based on numerical (possible MHD) simulations of a very large volume of the Universe which is however well beyond the aim of this PhD thesis. On the other hand, in Chapt.8 we report our discovery of novel correlations between the size (RH) of Radio Halos and their radio power and between RH and the cluster mass within the Radio Halo region, MH. In particular this last “geometrical” MH − RH correlation allows us to “observationally” overcome the limitation of the “average” size of Radio Halos. Thus in this Chapter, by making use of this “geometrical” correlation and of a simplified form of the re-acceleration model based on the results of Chapt. 5 and 6 we are able to discuss expected correlations between the synchrotron power and the thermal cluster quantities relative to the radio emitting region. This is a new powerful tool of investigation and we show that all the observed correlations (PR − RH, PR − MH, PR − T, PR − LX, . . . ) now become well understood in the context of the re-acceleration model. In addition, we find that observationally the size of Radio Halos scales non-linearly with the virial radius of the parent cluster, and this immediately means that the fraction of the cluster volume which is radio emitting increases with cluster mass and thus that the non-thermal component in clusters is not self-similar.

Parameters and algorithms to evaluate cardiac mechanics by conductance catheter

This work is structured as follows: In Section 1 we discuss the clinical problem of heart failure. In particular, we present the phenomenon known as ventricular mechanical dyssynchrony: its impact on cardiac function, the therapy for its treatment and the methods for its quantification. Specifically, we describe the conductance catheter and its use for the measurement of dyssynchrony. At the end of the Section 1, we propose a new set of indexes to quantify the dyssynchrony that are studied and validated thereafter. In Section 2 we describe the studies carried out in this work: we report the experimental protocols, we present and discuss the results obtained. Finally, we report the overall conclusions drawn from this work and we try to envisage future works and possible clinical applications of our results. Ancillary studies that were carried out during this work mainly to investigate several aspects of cardiac resynchronization therapy (CRT) are mentioned in Appendix. -------- Ventricular mechanical dyssynchrony plays a regulating role already in normal physiology but is especially important in pathological conditions, such as hypertrophy, ischemia, infarction, or heart failure (Chapter 1,2.). Several prospective randomized controlled trials supported the clinical efficacy and safety of cardiac resynchronization therapy (CRT) in patients with moderate or severe heart failure and ventricular dyssynchrony. CRT resynchronizes ventricular contraction by simultaneous pacing of both left and right ventricle (biventricular pacing) (Chapter 1.). Currently, the conductance catheter method has been used extensively to assess global systolic and diastolic ventricular function and, more recently, the ability of this instrument to pick-up multiple segmental volume signals has been used to quantify mechanical ventricular dyssynchrony. Specifically, novel indexes based on volume signals acquired with the conductance catheter were introduced to quantify dyssynchrony (Chapter 3,4.). Present work was aimed to describe the characteristics of the conductancevolume signals, to investigate the performance of the indexes of ventricular dyssynchrony described in literature and to introduce and validate improved dyssynchrony indexes. Morevoer, using the conductance catheter method and the new indexes, the clinical problem of the ventricular pacing site optimization was addressed and the measurement protocol to adopt for hemodynamic tests on cardiac pacing was investigated. In accordance to the aims of the work, in addition to the classical time-domain parameters, a new set of indexes has been extracted, based on coherent averaging procedure and on spectral and cross-spectral analysis (Chapter 4.). Our analyses were carried out on patients with indications for electrophysiologic study or device implantation (Chapter 5.). For the first time, besides patients with heart failure, indexes of mechanical dyssynchrony based on conductance catheter were extracted and studied in a population of patients with preserved ventricular function, providing information on the normal range of such a kind of values. By performing a frequency domain analysis and by applying an optimized coherent averaging procedure (Chapter 6.a.), we were able to describe some characteristics of the conductance-volume signals (Chapter 6.b.). We unmasked the presence of considerable beat-to-beat variations in dyssynchrony that seemed more frequent in patients with ventricular dysfunction and to play a role in discriminating patients. These non-recurrent mechanical ventricular non-uniformities are probably the expression of the substantial beat-to-beat hemodynamic variations, often associated with heart failure and due to cardiopulmonary interaction and conduction disturbances. We investigated how the coherent averaging procedure may affect or refine the conductance based indexes; in addition, we proposed and tested a new set of indexes which quantify the non-periodic components of the volume signals. Using the new set of indexes we studied the acute effects of the CRT and the right ventricular pacing, in patients with heart failure and patients with preserved ventricular function. In the overall population we observed a correlation between the hemodynamic changes induced by the pacing and the indexes of dyssynchrony, and this may have practical implications for hemodynamic-guided device implantation. The optimal ventricular pacing site for patients with conventional indications for pacing remains controversial. The majority of them do not meet current clinical indications for CRT pacing. Thus, we carried out an analysis to compare the impact of several ventricular pacing sites on global and regional ventricular function and dyssynchrony (Chapter 6.c.). We observed that right ventricular pacing worsens cardiac function in patients with and without ventricular dysfunction unless the pacing site is optimized. CRT preserves left ventricular function in patients with normal ejection fraction and improves function in patients with poor ejection fraction despite no clinical indication for CRT. Moreover, the analysis of the results obtained using new indexes of regional dyssynchrony, suggests that pacing site may influence overall global ventricular function depending on its relative effects on regional function and synchrony. Another clinical problem that has been investigated in this work is the optimal right ventricular lead location for CRT (Chapter 6.d.). Similarly to the previous analysis, using novel parameters describing local synchrony and efficiency, we tested the hypothesis and we demonstrated that biventricular pacing with alternative right ventricular pacing sites produces acute improvement of ventricular systolic function and improves mechanical synchrony when compared to standard right ventricular pacing. Although no specific right ventricular location was shown to be superior during CRT, the right ventricular pacing site that produced the optimal acute hemodynamic response varied between patients. Acute hemodynamic effects of cardiac pacing are conventionally evaluated after stabilization episodes. The applied duration of stabilization periods in most cardiac pacing studies varied considerably. With an ad hoc protocol (Chapter 6.e.) and indexes of mechanical dyssynchrony derived by conductance catheter we demonstrated that the usage of stabilization periods during evaluation of cardiac pacing may mask early changes in systolic and diastolic intra-ventricular dyssynchrony. In fact, at the onset of ventricular pacing, the main dyssynchrony and ventricular performance changes occur within a 10s time span, initiated by the changes in ventricular mechanical dyssynchrony induced by aberrant conduction and followed by a partial or even complete recovery. It was already demonstrated in normal animals that ventricular mechanical dyssynchrony may act as a physiologic modulator of cardiac performance together with heart rate, contractile state, preload and afterload. The present observation, which shows the compensatory mechanism of mechanical dyssynchrony, suggests that ventricular dyssynchrony may be regarded as an intrinsic cardiac property, with baseline dyssynchrony at increased level in heart failure patients. To make available an independent system for cardiac output estimation, in order to confirm the results obtained with conductance volume method, we developed and validated a novel technique to apply the Modelflow method (a method that derives an aortic flow waveform from arterial pressure by simulation of a non-linear three-element aortic input impedance model, Wesseling et al. 1993) to the left ventricular pressure signal, instead of the arterial pressure used in the classical approach (Chapter 7.). The results confirmed that in patients without valve abnormalities, undergoing conductance catheter evaluations, the continuous monitoring of cardiac output using the intra-ventricular pressure signal is reliable. Thus, cardiac output can be monitored quantitatively and continuously with a simple and low-cost method. During this work, additional studies were carried out to investigate several areas of uncertainty of CRT. The results of these studies are briefly presented in Appendix: the long-term survival in patients treated with CRT in clinical practice, the effects of CRT in patients with mild symptoms of heart failure and in very old patients, the limited thoracotomy as a second choice alternative to transvenous implant for CRT delivery, the evolution and prognostic significance of diastolic filling pattern in CRT, the selection of candidates to CRT with echocardiographic criteria and the prediction of response to the therapy.

Study of the antioxidant fraction in edible olive oils obtained from different technological systems and parameters

Questo lavoro di tesi è stato suddiviso in tre parti. L’argomento principale è stato lo “Studio della componente antiossidante di oli ottenuti da olive mediante l’utilizzo di diversi sistemi e parametri tecnologici”. E’ ben noto come la qualità ossidativa di un olio di oliva dipenda oltre che dalla sua composizione in acidi grassi, dalla presenza di composti caratterizzati da un elevata attività antiossidante, ovvero le sostanze fenoliche. I composti fenolici contribuiscono quindi in maniera preponderante alla shelf life dell’olio extravergine di oliva. Inoltre sono state riscontrate delle forti correlazione tra alcune di queste sostanze e gli attributi sensoriali positivi di amaro e piccante. E’ poi da sottolineare come il potere antiossidante dei composti fenolici degli oli vergini di oliva, sia stato negli ultimi anni oggetto di considerevole interesse, poiché correlato alla protezione da alcune patologie come ad esempio quelle vascolari, degenerative e tumorali. Il contenuto delle sostanze fenoliche negli oli di oliva dipende da diversi fattori: cultivar, metodo di coltivazione, grado di maturazione delle olive e ovviamente dalle operazioni tecnologiche poiché possono variare il quantitativo di questi composti estratto. Alla luce di quanto appena detto abbiamo valutato l’influenza dei fattori agronomici (metodi di agricoltura biologica, integrata e convenzionale) e tecnologici (riduzione della temperatura della materia prima, aggiunta di coadiuvanti in fase di frangitura e di gramolatura, confronto tra tre oli extravergini di oliva ottenuti mediante diversi sistemi tecnologici) sul contenuto in composti fenolici di oli edibili ottenuti da olive (paper 1-3-4). Oltre alle sostanze fenoliche, negli oli di oliva sono presenti altri composti caratterizzati da proprietà chimiche e nutrizionali, tra questi vi sono i fitosteroli, ovvero gli steroli tipici del mondo vegetale, che rappresentano la frazione dell’insaponificabile quantitativamente più importante dopo gli idrocarburi. La composizione quali-quantitativa degli steroli di un olio di oliva è una delle caratteristiche analitiche più importanti nella valutazione della sua genuinità; infatti la frazione sterolica è significativamente diversa in funzione dell’origine botanica e perciò viene utilizzata per distinguere tra di loro gli oli e le loro miscele. Il principale sterolo nell’olio di oliva è il β- sitosterolo, la presenza di questo composto in quantità inferiore al 90% è un indice approssimativo dell’aggiunta di un qualsiasi altro olio. Il β-sitosterolo è una sostanza importante dal punto di vista della salute, poiché si oppone all’assorbimento del colesterolo. Mentre in letteratura si trovano numerosi lavori relativi al potere antiossidante di una serie di composti presenti nell’olio vergine di oliva (i già citati polifenoli, ma anche carotenoidi e tocoferoli) e ricerche che dimostrano invece come altri composti possano promuovere l’ossidazione dei lipidi, per quanto riguarda il potere antiossidante degli steroli e dei 4- metilsteroli, vi sono ancora poche informazioni. Per questo è stata da noi valutata la composizione sterolica in oli extravergini di oliva ottenuti con diverse tecnologie di estrazione e l’influenza di questa sostanza sulla loro stabilità ossidativa (paper 2). E’ stato recentemente riportato in letteratura come lipidi cellulari evidenziati attraverso la spettroscopia di risonanza nucleare magnetica (NMR) rivestano una importanza strategica da un punto di vista funzionale e metabolico. Questi lipidi, da un lato un lato sono stati associati allo sviluppo di cellule neoplastiche maligne e alla morte cellulare, dall’altro sono risultati anche messaggeri di processi benigni quali l’attivazione e la proliferazione di un normale processo di crescita cellulare. Nell’ambito di questa ricerca è nata una collaborazione tra il Dipartimento di Biochimica “G. Moruzzi” ed il Dipartimento di Scienze degli Alimenti dell’Università di Bologna. Infatti, il gruppo di lipochimica del Dipartimento di Scienze degli Alimenti, a cui fa capo il Prof. Giovanni Lercker, da sempre si occupa dello studio delle frazioni lipidiche, mediante le principali tecniche cromatografiche. L’obiettivo di questa collaborazione è stato quello di caratterizzare la componente lipidica totale estratta dai tessuti renali umani sani e neoplastici, mediante l’utilizzo combinato di diverse tecniche analitiche: la risonanza magnetica nucleare (1H e 13C RMN), la cromatografia su strato sottile (TLC), la cromatografia liquida ad alta prestazione (HPLC) e la gas cromatografia (GC) (paper 5-6-7)

Caratterizzazione del legame di molecole di interesse farmaceutico alla sieroalbumina umana mediante biocromatografia e dicroismo circolare

Negli ultimi anni, un crescente numero di studiosi ha focalizzato la propria attenzione sullo sviluppo di strategie che permettessero di caratterizzare le proprietà ADMET dei farmaci in via di sviluppo, il più rapidamente possibile. Questa tendenza origina dalla consapevolezza che circa la metà dei farmaci in via di sviluppo non viene commercializzato perché ha carenze nelle caratteristiche ADME, e che almeno la metà delle molecole che riescono ad essere commercializzate, hanno comunque qualche problema tossicologico o ADME [1]. Infatti, poco importa quanto una molecola possa essere attiva o specifica: perché possa diventare farmaco è necessario che venga ben assorbita, distribuita nell’organismo, metabolizzata non troppo rapidamente, ne troppo lentamente e completamente eliminata. Inoltre la molecola e i suoi metaboliti non dovrebbero essere tossici per l’organismo. Quindi è chiaro come una rapida determinazione dei parametri ADMET in fasi precoci dello sviluppo del farmaco, consenta di risparmiare tempo e denaro, permettendo di selezionare da subito i composti più promettenti e di lasciar perdere quelli con caratteristiche negative. Questa tesi si colloca in questo contesto, e mostra l’applicazione di una tecnica semplice, la biocromatografia, per caratterizzare rapidamente il legame di librerie di composti alla sieroalbumina umana (HSA). Inoltre mostra l’utilizzo di un’altra tecnica indipendente, il dicroismo circolare, che permette di studiare gli stessi sistemi farmaco-proteina, in soluzione, dando informazioni supplementari riguardo alla stereochimica del processo di legame. La HSA è la proteina più abbondante presente nel sangue. Questa proteina funziona da carrier per un gran numero di molecole, sia endogene, come ad esempio bilirubina, tiroxina, ormoni steroidei, acidi grassi, che xenobiotici. Inoltre aumenta la solubilità di molecole lipofile poco solubili in ambiente acquoso, come ad esempio i tassani. Il legame alla HSA è generalmente stereoselettivo e ad avviene a livello di siti di legame ad alta affinità. Inoltre è ben noto che la competizione tra farmaci o tra un farmaco e metaboliti endogeni, possa variare in maniera significativa la loro frazione libera, modificandone l’attività e la tossicità. Per queste sue proprietà la HSA può influenzare sia le proprietà farmacocinetiche che farmacodinamiche dei farmaci. Non è inusuale che un intero progetto di sviluppo di un farmaco possa venire abbandonato a causa di un’affinità troppo elevata alla HSA, o a un tempo di emivita troppo corto, o a una scarsa distribuzione dovuta ad un debole legame alla HSA. Dal punto di vista farmacocinetico, quindi, la HSA è la proteina di trasporto del plasma più importante. Un gran numero di pubblicazioni dimostra l’affidabilità della tecnica biocromatografica nello studio dei fenomeni di bioriconoscimento tra proteine e piccole molecole [2-6]. Il mio lavoro si è focalizzato principalmente sull’uso della biocromatografia come metodo per valutare le caratteristiche di legame di alcune serie di composti di interesse farmaceutico alla HSA, e sul miglioramento di tale tecnica. Per ottenere una miglior comprensione dei meccanismi di legame delle molecole studiate, gli stessi sistemi farmaco-HSA sono stati studiati anche con il dicroismo circolare (CD). Inizialmente, la HSA è stata immobilizzata su una colonna di silice epossidica impaccata 50 x 4.6 mm di diametro interno, utilizzando una procedura precedentemente riportata in letteratura [7], con alcune piccole modifiche. In breve, l’immobilizzazione è stata effettuata ponendo a ricircolo, attraverso una colonna precedentemente impaccata, una soluzione di HSA in determinate condizioni di pH e forza ionica. La colonna è stata quindi caratterizzata per quanto riguarda la quantità di proteina correttamente immobilizzata, attraverso l’analisi frontale di L-triptofano [8]. Di seguito, sono stati iniettati in colonna alcune soluzioni raceme di molecole note legare la HSA in maniera enantioselettiva, per controllare che la procedura di immobilizzazione non avesse modificato le proprietà di legame della proteina. Dopo essere stata caratterizzata, la colonna è stata utilizzata per determinare la percentuale di legame di una piccola serie di inibitori della proteasi HIV (IPs), e per individuarne il sito(i) di legame. La percentuale di legame è stata calcolata attraverso il fattore di capacità (k) dei campioni. Questo parametro in fase acquosa è stato estrapolato linearmente dal grafico log k contro la percentuale (v/v) di 1-propanolo presente nella fase mobile. Solamente per due dei cinque composti analizzati è stato possibile misurare direttamente il valore di k in assenza di solvente organico. Tutti gli IPs analizzati hanno mostrato un’elevata percentuale di legame alla HSA: in particolare, il valore per ritonavir, lopinavir e saquinavir è risultato maggiore del 95%. Questi risultati sono in accordo con dati presenti in letteratura, ottenuti attraverso il biosensore ottico [9]. Inoltre, questi risultati sono coerenti con la significativa riduzione di attività inibitoria di questi composti osservata in presenza di HSA. Questa riduzione sembra essere maggiore per i composti che legano maggiormente la proteina [10]. Successivamente sono stati eseguiti degli studi di competizione tramite cromatografia zonale. Questo metodo prevede di utilizzare una soluzione a concentrazione nota di un competitore come fase mobile, mentre piccole quantità di analita vengono iniettate nella colonna funzionalizzata con HSA. I competitori sono stati selezionati in base al loro legame selettivo ad uno dei principali siti di legame sulla proteina. In particolare, sono stati utilizzati salicilato di sodio, ibuprofene e valproato di sodio come marker dei siti I, II e sito della bilirubina, rispettivamente. Questi studi hanno mostrato un legame indipendente dei PIs ai siti I e II, mentre è stata osservata una debole anticooperatività per il sito della bilirubina. Lo stesso sistema farmaco-proteina è stato infine investigato in soluzione attraverso l’uso del dicroismo circolare. In particolare, è stato monitorata la variazione del segnale CD indotto di un complesso equimolare [HSA]/[bilirubina], a seguito dell’aggiunta di aliquote di ritonavir, scelto come rappresentante della serie. I risultati confermano la lieve anticooperatività per il sito della bilirubina osservato precedentemente negli studi biocromatografici. Successivamente, lo stesso protocollo descritto precedentemente è stato applicato a una colonna di silice epossidica monolitica 50 x 4.6 mm, per valutare l’affidabilità del supporto monolitico per applicazioni biocromatografiche. Il supporto monolitico monolitico ha mostrato buone caratteristiche cromatografiche in termini di contropressione, efficienza e stabilità, oltre che affidabilità nella determinazione dei parametri di legame alla HSA. Questa colonna è stata utilizzata per la determinazione della percentuale di legame alla HSA di una serie di poliamminochinoni sviluppati nell’ambito di una ricerca sulla malattia di Alzheimer. Tutti i composti hanno mostrato una percentuale di legame superiore al 95%. Inoltre, è stata osservata una correlazione tra percentuale di legame è caratteristiche della catena laterale (lunghezza e numero di gruppi amminici). Successivamente sono stati effettuati studi di competizione dei composti in esame tramite il dicroismo circolare in cui è stato evidenziato un effetto anticooperativo dei poliamminochinoni ai siti I e II, mentre rispetto al sito della bilirubina il legame si è dimostrato indipendente. Le conoscenze acquisite con il supporto monolitico precedentemente descritto, sono state applicate a una colonna di silice epossidica più corta (10 x 4.6 mm). Il metodo di determinazione della percentuale di legame utilizzato negli studi precedenti si basa su dati ottenuti con più esperimenti, quindi è necessario molto tempo prima di ottenere il dato finale. L’uso di una colonna più corta permette di ridurre i tempi di ritenzione degli analiti, per cui la determinazione della percentuale di legame alla HSA diventa molto più rapida. Si passa quindi da una analisi a medio rendimento a una analisi di screening ad alto rendimento (highthroughput- screening, HTS). Inoltre, la riduzione dei tempi di analisi, permette di evitare l’uso di soventi organici nella fase mobile. Dopo aver caratterizzato la colonna da 10 mm con lo stesso metodo precedentemente descritto per le altre colonne, sono stati iniettati una serie di standard variando il flusso della fase mobile, per valutare la possibilità di utilizzare flussi elevati. La colonna è stata quindi impiegata per stimare la percentuale di legame di una serie di molecole con differenti caratteristiche chimiche. Successivamente è stata valutata la possibilità di utilizzare una colonna così corta, anche per studi di competizione, ed è stata indagato il legame di una serie di composti al sito I. Infine è stata effettuata una valutazione della stabilità della colonna in seguito ad un uso estensivo. L’uso di supporti cromatografici funzionalizzati con albumine di diversa origine (ratto, cane, guinea pig, hamster, topo, coniglio), può essere proposto come applicazione futura di queste colonne HTS. Infatti, la possibilità di ottenere informazioni del legame dei farmaci in via di sviluppo alle diverse albumine, permetterebbe un migliore paragone tra i dati ottenuti tramite esperimenti in vitro e i dati ottenuti con esperimenti sull’animale, facilitando la successiva estrapolazione all’uomo, con la velocità di un metodo HTS. Inoltre, verrebbe ridotto anche il numero di animali utilizzati nelle sperimentazioni. Alcuni lavori presenti in letteratura dimostrano l’affidabilita di colonne funzionalizzate con albumine di diversa origine [11-13]: l’utilizzo di colonne più corte potrebbe aumentarne le applicazioni.

Nuovi catalizzatori per l'idrogenazione, l'idrogenolisi/ring-opening di composti aromatici policiclici

The removal of aromatic hydrocarbons from diesel has received considerable attention after environmental regulations that require petroleum refiners to raise cetane number and to limit aromatics in diesel fuel in order to improve combustion efficiency and reduce particulate and NOx emissions. An alternative is blending with Fischer–Tropsch (FT) gas-to-liquid diesel fuel; however, this option may not be economically viable solution in case of extensive blend. Another alternative is to incorporate in the diesel pool a greater fraction of the so-called light cycle oil (LCO). Due to its high aromatics content and its low cetane number (typically between 20 and 30), the incorporation of LCO may have a negative impact on the quality of diesel. Current technologies for LCO improvement are based on hydrogenation to adjust both sulphur and cetane number but while an important fraction of the aromatics present in LCO can be saturated in a deep hydrogenation process, the cetane number may still be lower than the target values specified in diesel legislations, so further upgrading is needed. An interesting technology for improving the cetane number of diesels and maintaining meanwhile high diesel yields is achieved by combining a complete hydrogenation process with a selective ring opening (SRO) reaction of the naphthenic rings. The SRO can be defined as naphthene ring-opening to form compounds with high cetane number, but without any carbon losses. Controlling the interconversion of six- and five- membered rings via an acid-catalyzed ring-contraction step is also of great importance, since selective conversion of six-membered to five-membered naphthene rings greatly influences ring-opening rates and selectivity. High intrinsic activity may be enhanced by deposition of noble metals on acidic, high surface area supports, because it is possible to arrange close proximity of the metal and acid sites. Moreover, in large-pore supports, the diffusion resistance of liquid reactants into the pores is minimized. In addition to metal centres, the acid sites of support also plays role in aromatics hydrogenation. However, the functions of different kinds of acid sites (Brønsted vs. Lewis acidity), and their optimal concentrations and strengths, remain unclear. In the present study we investigated the upgrading of an aromatic-rich feedstock over different type of metal supported on mesoporous silica-alumina. The selective hydrogenolysis and ring opening of tetrahydronaphthalene (THN or tetralin) was carried out as representative of LCO fractions after deep hydrogenation process. In this regards the aim of this study is to evaluate both the effect of metals and that of the supports characterized by different acid distribution and strength, on conversion and selectivity. For this purpose a series of catalysts were prepared by impregnation. The catalysts were characterized and conversion tests of THN were performed in a lab-scale plant operating in the pressure range from 7.0-5.0 MPa and in the temperature range from 300 to 360°C.

Study of the components of quality in SO2-free wines obtained by innovative vinification protocols. Evaluation of the pre-fermentative addition of lysozyme and oenological tannins.

The research performed during the PhD candidature was intended to evaluate the quality of white wines, as a function of the reduction in SO2 use during the first steps of the winemaking process. In order to investigate the mechanism and intensity of interactions occurring between lysozyme and the principal macro-components of musts and wines, a series of experiments on model wine solutions were undertaken, focusing attention on the polyphenols, SO2, oenological tannins, pectines, ethanol, and sugar components. In the second part of this research program, a series of conventional sulphite added vinifications were compared to vinifications in which sulphur dioxide was replaced by lysozyme and consequently define potential winemaking protocols suitable for the production of SO2-free wines. To reach the final goal, the technological performance of two selected yeast strains with a low aptitude to produce SO2 during fermentation were also evaluated. The data obtained suggested that the addition of lysozyme and oenological tannins during the alcoholic fermentation could represent a promising alternative to the use of sulphur dioxide and a reliable starting point for the production of SO2-free wines. The different vinification protocols studied influenced the composition of the volatile profile in wines at the end of the alcoholic fermentation, especially with regards to alcohols and ethyl esters also a consequence of the yeast’s response to the presence or absence of sulphites during fermentation, contributing in different ways to the sensory profiles of wines. In fact, the aminoacids analysis showed that lysozyme can affect the consumption of nitrogen as a function of the yeast strain used in fermentation. During the bottle storage, the evolution of volatile compounds is affected by the presence of SO2 and oenological tannins, confirming their positive role in scaveging oxygen and maintaining the amounts of esters over certain levels, avoiding a decline in the wine’s quality. Even though a natural decrease was found on phenolic profiles due to oxidation effects caused by the presence of oxygen dissolved in the medium during the storage period, the presence of SO2 together with tannins contrasted the decay of phenolic content at the end of the fermentation. Tannins also showed a central role in preserving the polyphenolic profile of wines during the storage period, confirming their antioxidant property, acting as reductants. Our study focused on the fundamental chemistry relevant to the oxidative phenolic spoilage of white wines has demonstrated the suitability of glutathione to inhibit the production of yellow xanthylium cation pigments generated from flavanols and glyoxylic acid at the concentration that it typically exists in wine. The ability of glutathione to bind glyoxylic acid rather than acetaldehyde may enable glutathione to be used as a ‘switch’ for glyoxylic acid-induced polymerisation mechanisms, as opposed to the equivalent acetaldehyde polymerisation, in processes such as microoxidation. Further research is required to assess the ability of glutathione to prevent xanthylium cation production during the in-situ production of glyoxylic acid and in the presence of sulphur dioxide.

Trasporto di materia in membrane polimeriche e nanocomposite per la separazione di gas

Membrane-based separation processes are acquiring, in the last years, an increasing importance because of their intrinsic energetic and environmental sustainability: some types of polymeric materials, showing adequate perm-selectivity features, appear rather suitable for these applications, because of their relatively low cost and easy processability. In this work have been studied two different types of polymeric membranes, in view of possible applications to the gas separation processes, i.e. Mixed Matrix Membranes (MMMs) and high free volume glassy polymers. Since the early 90’s, it has been understood that the performances of polymeric materials in the field of gas separations show an upper bound in terms of permeability and selectivity: in particular, an increase of permeability is often accompanied by a decrease of selectivity and vice-versa, while several inorganic materials, like zeolites or silica derivates, can overcome this limitation. As a consequence, it has been developed the idea of dispersing inorganic particles in polymeric matrices, in order to obtain membranes with improved perm-selectivity features. In particular, dispersing fumed silica nanoparticles in high free volume glassy polymers improves in all the cases gases and vapours permeability, while the selectivity may either increase or decrease, depending upon material and gas mixture: that effect is due to the capacity of nanoparticles to disrupt the local chain packing, increasing the dimensions of excess free volume elements trapped in the polymer matrix. In this work different kinds of MMMs were fabricated using amorphous Teflon® AF or PTMSP and fumed silica: in all the cases, a considerable increase of solubility, diffusivity and permeability of gases and vapours (n-alkanes, CO2, methanol) was observed, while the selectivity shows a non-monotonous trend with filler fraction. Moreover, the classical models for composites are not able to capture the increase of transport properties due to the silica addition, so it has been necessary to develop and validate an appropriate thermodynamic model that allows to predict correctly the mass transport features of MMMs. In this work, another material, called poly-trimethylsilyl-norbornene (PTMSN) was examined: it is a new generation high free volume glassy polymer that, like PTMSP, shows unusual high permeability and selectivity levels to the more condensable vapours. These two polymer differ each other because PTMSN shows a more pronounced chemical stability, due to its structure double-bond free. For this polymer, a set of Lattice Fluid parameters was estimated, making possible a comparison between experimental and theoretical solubility isotherms for hydrocarbons and alcoholic vapours: the successfully modelling task, based on application of NELF model, offers a reliable alternative to direct sorption measurement, which is extremely time-consuming due to the relevant relaxation phenomena showed by each sorption step. For this material also dilation experiments were performed, in order to quantify its dimensional stability in presence of large size, swelling vapours.

Food lipids: their effects on quality and oxidative stability of animal tissues and emulsions

Lipolysis and oxidation of lipids in foods are the major biochemical and chemical processes that cause food quality deterioration, leading to the characteristic, unpalatable odour and flavour called rancidity. In addition to unpalatability, rancidity may give rise to toxic levels of certain compounds like aldehydes, hydroperoxides, epoxides and cholesterol oxidation products. In this PhD study chromatographic and spectroscopic techniques were employed to determine the degree of lipid oxidation in different animal products and its relationship with technological parameters like feeding fat sources, packaging, processing and storage conditions. To achieve this goal capillary gas chromatography (CGC) was employed not only to determine the fatty acids profile but also, after solid phase extraction, the amount of sterols (cholesterol and phytosterols) and cholesterol oxidation products (COPs). To determine hydroperoxides, primary products of oxidation and quantify secondary products UV/VIS absorbance spectroscopy was applied. Beef and pork meat in this study were analysed. In actual fact, lipid oxidation is a major deterioration reaction in meat, meat products and results in adverse changes in the colour, flavour, texture of meat and develops different compounds which should be a risk to human health as oxysterols. On beef and pork meat, a study of lipid fraction during storage was carried out to evaluate its shelf-life and some nutritional features life saturated/unsaturated fatty acids ratio and sterols content, in according to the interest that has been growing around functional food in the last years. The last part of this research was focused on the study of lipid oxidation in emulsions. In oil-in-water emulsions antioxidant activity of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) was evaluated. The rates of lipid oxidation of 1.0% stripped soybean oil-in-water emulsions with DOPC were followed by monitoring lipid hydroperoxide and hexanal as indicators of primary and secondary oxidation products and the droplet surface charge or zeta potential (ζ) of the emulsions with varying concentrations of DOPC were tested. This manuscript reports the main results obtained in the three activities briefly summarized as follows: 1. study on effects of feeding composition on the photoxidative stability of lipids from beef meat, evaluated during storage under commercial retail conditions; 2. evaluation of effects of diets and storage conditions on the oxidative stability of pork meat lipids; 3. study on oxidative behavior of DOPC in stripped soybean oil-in-water emulsions stabilized by nonionic surfactant.

Emerging contaminants in agricultural ecosystems: impact of selected pharmaceutical on water and soil ecology and pratical implications

Pharmaceuticals are useful tools to prevent and treat human and animal diseases. Following administration, a significant fraction of pharmaceuticals is excreted unaltered into faeces and urine and may enter the aquatic ecosystem and agricultural soil through irrigation with recycled water, constituting a significant source of emerging contaminants into the environment. Understanding major factors influencing their environmental fate is consequently needed to value the risk, reduce contamination, and set up bioremediation technologies. The antiviral drug Tamiflu (oseltamivir carboxylate, OC) has received recent attention due to the potential use as a first line defence against H5N1 and H1N1 influenza viruses. Research has shown that OC is not removed during conventional wastewater treatments, thus having the potential to enter surface water bodies. A series of laboratory experiments investigated the fate and the removal of OC in surface water systems in Italy and Japan and in a municipal wastewater treatment plant. A preliminary laboratory study investigated the persistence of the active antiviral drug in water samples from an irrigation canal in northern Italy (Canale Emiliano Romagnolo). After an initial rapid decrease, OC concentration slowly decreased during the remaining incubation period. Approximately 65% of the initial OC amount remained in water at the end of the 36-day incubation period. A negligible amount of OC was lost both from sterilized water and from sterilized water/sediment samples, suggesting a significant role of microbial degradation. Stimulating microbial processes by the addition of sediments resulted in reduced OC persistence. Presence of OC (1.5 μg mL-1) did not significantly affect the metabolic potential of the water microbial population, that was estimated by glyphosate and metolachlor mineralization. In contrast, OC caused an initial transient decrease in the size of the indigenous microbial population of water samples. A second laboratory study focused on basic processes governing the environmental fate of OC in surface water from two contrasting aquatic ecosystems of northern Italy, the River Po and the Venice Lagoon. Results of this study confirmed the potential of OC to persist in surface water. However, the addition of 5% of sediments resulted in rapid OC degradation. The estimated half-life of OC in water/sediment of the River Po was 15 days. After three weeks of incubation at 20 °C, more than 8% of 14C-OC evolved as 14CO2 from water/sediment samples of the River Po and Venice Lagoon. OC was moderately retained onto coarse sediments from the two sites. In water/sediment samples of the River Po and Venice Lagoon treated with 14C-OC, more than 30% of the 14C-residues remained water-extractable after three weeks of incubation. The low affinity of OC to sediments suggests that the presence of sediments would not reduce its bioavailability to microbial degradation. Another series of laboratory experiments investigated the fate and the removal of OC in two surface water ecosystems of Japan and in the municipal wastewater treatment plant of the city of Bologna, in Northern Italy. The persistence of OC in surface water ranged from non-detectable degradation to a half-life of 53 days. After 40 days, less than 3% of radiolabeled OC evolved as 14CO2. The presence of sediments (5%) led to a significant increase of OC degradation and of mineralization rates. A more intense mineralization was observed in samples of the wastewater treatment plant when applying a long incubation period (40 days). More precisely, 76% and 37% of the initial radioactivity applied as 14C-OC was recovered as 14CO2 from samples of the biological tank and effluent water, respectively. Two bacterial strains growing on OC as sole carbon source were isolated and used for its removal from synthetic medium and environmental samples, including surface water and wastewater. Inoculation of water and wastewater samples with the two OC-degrading strains showed that mineralization of OC was significantly higher in both inoculated water and wastewater, than in uninoculated controls. Denaturing gradient gel electrophoresis and quantitative PCR analysis showed that OC would not affect the microbial population of surface water and wastewater. The capacity of the ligninolytic fungus Phanerochaete chrysosporium to degrade a wide variety of environmentally persistent xenobiotics has been largely reported in literature. In a series of laboratory experiments, the efficiency of a formulation using P. chrysosporium was evaluated for the removal of selected pharmaceuticals from wastewater samples. Addition of the fungus to samples of the wastewater treatment plant of Bologna significantly increased (P < 0.05) the removal of OC and three antibiotics, erythromycin, sulfamethoxazole, and ciprofloxacin. Similar effects were also observed in effluent water. OC was the most persistent of the four pharmaceuticals. After 30 days of incubation, approximately two times more OC was removed in bioremediated samples than in controls. The highest removal efficiency of the formulation was observed with the antibiotic ciprofloxacin. The studies included environmental aspects of soil contamination with two emerging veterinary contaminants, such as doramectin and oxibendazole, wich are common parasitic treatments in cattle farms.

Sulfanyl Radical Addition to Alkynes: Revisiting an Old Reaction to Enter the Novel Realms of Green Chemistry, Bioconjugation, and Material Chemistry

In the last decade considerable attention has been devoted to the rewarding use of Green Chemistry in various synthetic processes and applications. Green Chemistry is of special interest in the synthesis of expensive pharmaceutical products, where suitable adoption of “green” reagents and conditions is highly desirable. Our project especially focused in a search for new green radical processes which might also find useful applications in the industry. In particular, we have explored the possible adoption of green solvents in radical Thiol-Ene and Thiol-Yne coupling reactions, which to date have been normally performed in “ordinary” organic solvents such as benzene and toluene, with the primary aim of applying those coupling reactions to the construction of biological substrates. We have additionally tuned adequate reaction conditions which might enable achievement of highly functionalised materials and/or complex bioconjugation via homo/heterosequence. Furthermore, we have performed suitable theoretical studies to gain useful chemical information concerning mechanistic implications of the use of green solvents in the radical Thiol-Yne coupling reactions.

Effetti idrodinamici prodotti dal costume tecnico sul nuotatore

L’introduzione dei costumi tecnici nel nuoto ha portato miglioramenti senza precedenti sulla prestazione. I miglioramenti nella velocità di nuoto sono stati attribuiti dalla letteratura a riduzioni nelle resistenze idrodinamiche sul nuotatore. Tuttavia, gli effetti specifici dovuti all’utilizzo di questo tipo di costume non sono ancora completamente chiariti. Questa tesi aveva l’obiettivo di indagare gli effetti del costume tecnico sul galleggiamento statico, sulla posizione del corpo e sulla resistenza idrodinamica in avanzamento passivo. Nello studio preliminare sono stati misurati la spinta idrostatica, i volumi polmonari dinamici e la circonferenza toracica di 9 nuotatori che indossavano un costume tradizionale o un costume tecnico in gomma sintetica. Indossare il costume tecnico ha determinato una riduzione significativa del galleggiamento statico, e la compressione toracica causata da questo tipo di costume potrebbe avere una relazione con la significativa riduzione dei volumi polmonari misurati quando il nuotatore indossa questo tipo di costume. Un successiva analisi prevedeva il traino passivo di 14 nuotatori che mantenevano la miglior posizione idrodinamica di scivolamento indossando un costume tradizionale, tecnico in tessuto e tecnico in gomma. La posizione del corpo in avanzamento è stata misurata con un’analisi cinematica. La resistenza passiva indossando i costumi tecnici è risultata significativamente minore per entrambi i costumi tecnici rispetto alla prova con costume tradizionale. L’analisi condotta attraverso modelli di regressione lineari ha mostrato che una parte della riduzione della resistenza passiva era legata a proprietà intrinseche dei costumi tecnici. Tuttavia, anche l’area di impatto frontale determinata dall’inclinazione del tronco del soggetto in scivolamento e l’inclinazione degli arti inferiori hanno mostrato una marcata influenza sulla resistenza idrodinamica passiva. Pertanto, la riduzione di resistenza idrodinamica durante lo scivolamento passivo effettuato con costume tecnico da nuoto è attribuibile, oltre all’effetto del materiale di composizione del costume, ad una variazione della posizione del corpo del nuotatore.

Constraints on neutrino mass fraction using Redshift Space Distortions

Redshift Space Distortions (RSD) are an apparent anisotropy in the distribution of galaxies due to their peculiar motion. These features are imprinted in the correlation function of galaxies, which describes how these structures distribute around each other. RSD can be represented by a distortions parameter $\beta$, which is strictly related to the growth of cosmic structures. For this reason, measurements of RSD can be exploited to give constraints on the cosmological parameters, such us for example the neutrino mass. Neutrinos are neutral subatomic particles that come with three flavours, the electron, the muon and the tau neutrino. Their mass differences can be measured in the oscillation experiments. Information on the absolute scale of neutrino mass can come from cosmology, since neutrinos leave a characteristic imprint on the large scale structure of the universe. The aim of this thesis is to provide constraints on the accuracy with which neutrino mass can be estimated when expoiting measurements of RSD. In particular we want to describe how the error on the neutrino mass estimate depends on three fundamental parameters of a galaxy redshift survey: the density of the catalogue, the bias of the sample considered and the volume observed. In doing this we make use of the BASICC Simulation from which we extract a series of dark matter halo catalogues, characterized by different value of bias, density and volume. This mock data are analysed via a Markov Chain Monte Carlo procedure, in order to estimate the neutrino mass fraction, using the software package CosmoMC, which has been conveniently modified. In this way we are able to extract a fitting formula describing our measurements, which can be used to forecast the precision reachable in future surveys like Euclid, using this kind of observations.

Biochar characterization for its environmental and agricultural utilization. Occurrence, distribution and fate of labile organic carbon and polycyclic aromatic hydrocarbons

In this thesis the potential risks associated to the application of biochar in soil as well the stability of biochar were investigated. The study was focused on the potential risks arising from the occurrence of polycyclic aromatic hydrocarbons (PAHs) in biochar. An analytical method was developed for the determination of the 16 USEPA-PAHs in the original biochar and soil containing biochar. The method was successfully validated with a certified reference material for the soil matrix and compared with methods in use in other laboratories during a laboratory exercise within the EU-COST TD1107. The concentration of 16 USEPA-PAHs along with the 15 EU-PAHs, priority hazardous substances in food, was determined in a suite of currently available biochars for agricultural field applications derived from a variety of parent materials and pyrolysis conditions. Biochars analyzed contained the USEPA and some of the EU-PAHs at detectable levels ranging from 1.2 to 19 µg g-1. This method allowed investigating changes in PAH content and distribution in a four years study following biochar addition in soils in a vineyard (CNR-IBIMET). The results showed that biochar addition determined an increase of the amount of PAHs. However, the levels of PAHs in the soil remained within the maximum acceptable concentration for European countries. The vineyard soil performed by CNR-IBIMET was exploited to study the environmental stability of biochar and its impact on soil organic carbon. The stability of biochar was investigated by analytical pyrolysis (Py-GC-MS) and pyrolysis in the presence of hydrogen (HyPy). The findings showed that biochar amendment significantly influence soil stable carbon fraction concentration during the incubation period. Moreover, HyPy and Py-GC-MS were applied to biochars deriving from three different feedstock at two different pyrolysis temperatures. The results evidenced the influence of feedstock type and pyrolysis conditions on the degree of carbonisation.