Size distributions and temporal variations of biological aerosol particles in the Amazon rainforest characterized by microscopy and real-time UV-APS fluorescence techniques during AMAZE-08

As a part of the AMAZE-08 campaign during the wet season in the rainforest of central Amazonia, an ultraviolet aerodynamic particle sizer (UV-APS) was operated for continuous measurements of fluorescent biological aerosol particles (FBAP). In the coarse particle size range (> 1 mu m) the campaign median and quartiles of FBAP number and mass concentration were 7.3x10(4) m(-3) (4.0-13.2x10(4) m(-3)) and 0.72 mu g m(-3) (0.42-1.19 mu g m(-3)), respectively, accounting for 24% (11-41%) of total particle number and 47% (25-65%) of total particle mass. During the five-week campaign in February-March 2008 the concentration of coarse-mode Saharan dust particles was highly variable. In contrast, FBAP concentrations remained fairly constant over the course of weeks and had a consistent daily pattern, peaking several hours before sunrise, suggesting observed FBAP was dominated by nocturnal spore emission. This conclusion was supported by the consistent FBAP number size distribution peaking at 2.3 mu m, also attributed to fungal spores and mixed biological particles by scanning electron microscopy (SEM), light microscopy and biochemical staining. A second primary biological aerosol particle (PBAP) mode between 0.5 and 1.0 mu m was also observed by SEM, but exhibited little fluorescence and no true fungal staining. This mode may have consisted of single bacterial cells, brochosomes, various fragments of biological material, and small Chromalveolata (Chromista) spores. Particles liquid-coated with mixed organic-inorganic material constituted a large fraction of observations, and these coatings contained salts likely from primary biological origin. We provide key support for the suggestion that real-time laser-induce fluorescence (LIF) techniques using 355 nm excitation provide size-resolved concentrations of FBAP as a lower limit for the atmospheric abundance of biological particles in a pristine environment. We also show some limitations of using the instrument for ambient monitoring of weakly fluorescent particles < 2 mu m. Our measurements confirm that primary biological particles, fungal spores in particular, are an important fraction of supermicron aerosol in the Amazon and that may contribute significantly to hydrological cycling, especially when coated by mixed inorganic material.

Synthesis, spectroscopic characterization, photochemical and photophysical properties and biological activities of ruthenium complexes with mono- and bi-dentate histamine ligand

The monodentate cis-[Ru(phen)(2)(hist)(2)](2+) 1R and the bidentate cis-[Ru(phen)(2)(hist)](2+) 2A complexes were prepared and characterized using spectroscopic (H-1, (H-1-H-1) COSY and (H-1-C-13) HSQC NMR, UV-vis, luminescence) techniques. The complexes presented absorption and emission in the visible region, as well as a tri-exponential emission decay. The complexes are soluble in aqueous and non-aqueous solution with solubility in a buffer solution of pH 7.4 of 1.14 x 10(-3) mol L-1 for (1R + 2A) and 6.43 x 10(-4) mol L-1 for 2A and lipophilicity measured in an aqueous-octanol solution of -1.14 and -0.96, respectively. Photolysis in the visible region in CH3CN converted the starting complexes into cis-[Ru(phen)(2)(CH3CN)(2)](2+). Histamine photorelease was also observed in pure water and in the presence of BSA (1.0 x 10(-6) mol L-1). The bidentate coordination of the histamine to the ruthenium center in relation to the monodentate coordination increased the photosubstitution quantum yield by a factor of 3. Pharmacological studies showed that the complexes present a moderate inhibition of AChE with an IC50 of 21 mu mol L-1 (referred to risvagtini, IC50 181 mu mol L-1 and galantamine IC50 0.006 mu mol L-1) with no appreciable cytotoxicity toward to the HeLa cells (50% cell viability at 925 mu mol L-1). Cell uptake of the complexes into HeLa cells was detected by fluorescence confocal microscopy. Overall, the observation of a luminescent complex that penetrates the cell wall and has low cytotoxicity, but is reactive photochemically, releasing histamine when irradiated with visible light, are interesting features for application of these complexes as phototherapeutic agents.

Synthesis, Characterization and Biological Activities of a Bidentate Schiff Base Ligand: N,N '-Bis(1-phenylethylidene)ethane-1,2-diamine and its Transition Metals (II) Complexes

Schiff base ligand: N,N'-bis(1-phenylethylidene)ethane-1,2-diamine (L), was derived from acetophenone and ethylenediamine by condensation and its complexes (1-5) were prepared with Pb2+, Ni2+, Co2+, Cu2+ and Cd2+ metal ions. Their structures were characterized by FAB-MS, IR spectra, elemental analyses and molar conductance. The octahedral geometry of the complexes was proposed by electronic spectra and magnetic moment data. The conductivity data showed that the complexes have non-electrolytic nature. The complexes (1-5) have higher in vitro antimicrobial activity than the Schiff base ligand (L). In the nuclease activity, the complexes cleave DNA as compared to control DNA in the presence of H2O2.

Synthesis, Biological Evaluation, And Molecular Modeling of Chalcone Derivatives As Potent Inhibitors of Mycobacterium tuberculosis Protein Tyrosine Phosphatases (PtpA and PtpB)

Tuberculosis (TB) is a major infectious disease caused by Mycobacterium tuberculosis (Mtb). According to the World Health Organization (WHO), about 1.8 million people die from TB and 10 million new cases are recorded each year. Recently, a new series of naphthylchalcones has been identified as inhibitors of Mtb protein tyrosine phosphatases (PTPs). In this work, 100 chalcones were designed, synthesized, and investigated for their inhibitory properties against MtbPtps. Structure-activity relationships (SAR) were developed, leading to the discovery of new potent inhibitors with IC50 values in the low-micromolar range. Kinetic studies revealed competitive inhibition and high selectivity toward the Mtb enzymes. Molecular modeling investigations were carried out with the aim of revealing the most relevant structural requirements underlying the binding affinity and selectivity of this series of inhibitors as potential anti-TB drugs.

Synthesis, biological evaluation and molecular docking studies of 3-(triazolyl)coumarin derivatives: Effect on inducible nitric oxide synthase

A series of 3-(triazolyl)-coumarins were synthesized and tested as anti-inflammatory agents. It was possible to infer that these compounds do not alter the interaction of LPS with TLR-4 or TLR-2, as the intracellular pathways involved in the TNF-alpha secretion and COX-2 activity were not affected. Nevertheless, the compounds inhibited iNOS-derived NO production, without affecting the eNOS activity. The outcome of the docking studies showed that it pi center dot center dot center dot pi interactions with the heme group are important for the iNOS inhibition, thus making compound 3c a promising lead. Moreover, the efficacy of this compound was visualized by the reduced number of neutrophils in the LPS-inflamed subcutaneous tissue. Together, biological and docking data show that triazolyl-substituted coumarins, that can act on iNOS, are a good scaffold to be explored. (C) 2012 Elsevier Masson SAS. All rights reserved.

A tellurium-based cathepsin B inhibitor: Molecular structure, modelling, molecular docking and biological evaluation

The crystallographically determined structure of biologically active 4,4-dichloro-1,3-diphenyl-4-telluraoct-2-en-1-one, 3, shows the coordination geometry for Te to be distorted psi-pentagonal bipyramidal based on a C2OCl3(lone pair) donor set. Notable is the presence of an intramolecular axial Te center dot center dot center dot O (carbonyl) interaction, a design element included to reduce hydrolysis. Raman and molecular modelling studies indicate the persistence of the Te center dot center dot center dot O(carbonyl) interaction in the solution (CHCl3) and gasphases, respectively. Docking studies of 3' (i.e. original 3 less one chloride) with Cathepsin B reveals a change in the configuration about the vinyl C = C bond. i.e. to E from Z (crystal structure). This isomerism allows the optimisation of interactions in the complex which features a covalent Te-SGCys29 bond. Crucially, the E configuration observed for 3' allows for the formation of a hypervalent Te center dot center dot center dot O interaction as well as an O center dot center dot center dot H-O hydrogen bond with the Gly27 and Glu122 residues, respectively. Additional stabilisation is afforded by a combination of interactions spanning the S1, S2, S1' and S2' sub-sites of Cathepsin B. The greater experimental inhibitory activity of 3 compared with analogues is rationalised by the additional interactions formed between 3' and the His110 and His111 residues in the occluding loop, which serve to hinder the entrance to the active site. (C) 2012 Elsevier B.V. All rights reserved.

Use of solvents and environmental friendly materials for applications in Green Chemistry

The present Thesis studies three alternative solvent groups as sustainable replacement of traditional organic solvents. Some aspects of fluorinated solvents, supercritical fluids and ionic liquids, have been analysed with a critical approach and their effective “greenness” has been evaluated from the points of view of the synthesis, the properties and the applications. In particular, the attention has been put on the environmental and human health issues, evaluating the eco-toxicity, the toxicity and the persistence, to underline that applicability and sustainability are subjects with equal importance. The “green” features of fluorous solvents and supercritical fluids are almost well-established; in particular supercritical carbon dioxide (scCO2) is probably the “greenest” solvent among the alternative solvent systems developed in the last years, enabling to combine numerous advantages both from the point of view of industrial/technological applications and eco-compatibility. In the Thesis the analysis of these two classes of alternative solvents has been mainly focused on their applicability, rather than the evaluation of their environmental impact. Specifically they have been evaluated as alternative media for non-aqueous biocatalysis. For this purpose, the hydrophobic ion pairing (HIP), which allows solubilising enzymes in apolar solvents by an ion pairing between the protein and a surfactant, has been investigated as effective enzymatic derivatisation technique to improve the catalytic activity under homogeneous conditions in non conventional media. The results showed that the complex enzyme-surfactant was much more active both in fluorous solvents and in supercritical carbon dioxide than the native form of the enzyme. Ionic liquids, especially imidazolium salts, have been proposed some years ago as “fully green” alternative solvents; however this epithet does not take into account several “brown” aspects such as their synthesis from petro-chemical starting materials, their considerable eco-toxicity, toxicity and resistance to biodegradation, and the difficulty of clearly outline applications in which ionic liquids are really more advantageous than traditional solvents. For all of these reasons in this Thesis a critical analysis of ionic liquids has been focused on three main topics: i) alternative synthesis by introducing structural moieties which could reduce the toxicity of the most known liquid salts, and by using starting materials from renewable resources; ii) on the evaluation of their environmental impact through eco-toxicological tests (Daphnia magna and Vibrio fischeri acute toxicity tests, and algal growth inhibition), toxicity tests (MTT test, AChE inhibition and LDH release tests) and fate and rate of aerobic biodegradation in soil and water; iii) and on the demonstration of their effectiveness as reaction media in organo-catalysis and as extractive solvents in the recovery of vegetable oil from terrestrial and aquatic biomass. The results about eco-toxicity tests with Daphnia magna, Vibrio fischeri and algae, and toxicity assay using cultured cell lines, clearly indicate that the difference in toxicity between alkyl and oxygenated cations relies in differences of polarity, according to the general trend of decreasing toxicity by decreasing the lipophilicity. Independently by the biological approach in fact, all the results are in agreement, showing a lower toxicity for compounds with oxygenated lateral chains than for those having purely alkyl lateral chains. These findings indicate that an appropriate choice of cation and anion structures is important not only to design the IL with improved and suitable chemico-physical properties but also to obtain safer and eco-friendly ILs. Moreover there is a clear indication that the composition of the abiotic environment has to be taken into account when the toxicity of ILs in various biological test systems is analysed, because, for example, the data reported in the Thesis indicate a significant influence of salinity variations on algal toxicity. Aerobic biodegradation of four imidazolium ionic liquids, two alkylated and two oxygenated, in soil was evaluated for the first time. Alkyl ionic liquids were shown to be biodegradable over the 6 months test period, and in contrast no significant mineralisation was observed with oxygenated derivatives. A different result was observed in the aerobic biodegradation of alkylated and oxygenated pyridinium ionic liquids in water because all the ionic liquids were almost completely degraded after 10 days, independently by the number of oxygen in the lateral chain of the cation. The synthesis of new ionic liquids by using renewable feedstock as starting materials, has been developed through the synthesis of furan-based ion pairs from furfural. The new ammonium salts were synthesised in very good yields, good purity of the products and wide versatility, combining low melting points with high decomposition temperatures and reduced viscosities. Regarding the possible applications as surfactants and biocides, furan-based salts could be a valuable alternative to benzyltributylammonium salts and benzalkonium chloride that are produced from non-renewable resources. A new procedure for the allylation of ketones and aldehydes with tetraallyltin in ionic liquids was developed. The reaction afforded high yields both in sulfonate-containing ILs and in ILs without sulfonate upon addition of a small amount of sulfonic acid. The checked reaction resulted in peculiar chemoselectivity favouring aliphatic substrates towards aromatic ketones and good stereoselectivity in the allylation of levoglucosenone. Finally ILs-based systems could be easily and successfully recycled, making the described procedure environmentally benign. The potential role of switchable polarity solvents as a green technology for the extraction of vegetable oil from terrestrial and aquatic biomass has been investigated. The extraction efficiency of terrestrial biomass rich in triacylglycerols, as soy bean flakes and sunflower seeds, was comparable to those of traditional organic solvents, being the yield of vegetable oils recovery very similar. Switchable polarity solvents as been also exploited for the first time in the extraction of hydrocarbons from the microalga Botryococcus braunii, demonstrating the efficiency of the process for the extraction of both dried microalgal biomass and directly of the aqueous growth medium. The switchable polarity solvents exhibited better extraction efficiency than conventional solvents, both with dried and liquid samples. This is an important issue considering that the harvest and the dewatering of algal biomass have a large impact on overall costs and energy balance.

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.

Synthesis of Modified Amino Acids and Insertion in Peptides and Mimetics. Structural Aspects and Impact on Biological Activity.

I studied the effects exerted by the modifications on structures and biological activities of the compounds so obtained. I prepared peptide analogues containing unusual amino acids such as halogenated, alkylated (S)- or (R)-tryptophans, useful for the synthesis of mimetics of the endogenous opioid peptide endomorphin-1, or 2-oxo-1,3-oxazolidine-4-carboxylic acids, utilized as pseudo-prolines having a clear all-trans configuration of the preceding peptide bond. The latter gave access to a series of constrained peptidomimetics with potential interest in medicinal chemistry and in the field of the foldamers. In particular, I have dedicated much efforts to the preparation of cyclopentapeptides containing D-configured, alfa-, or beta-aminoacids, and also of cyclotetrapeptides including the retro-inverso modification. The conformational analyses confirmed that these cyclic compounds can be utilized as rigid scaffolds mimicking gamma- or beta-turns, allowing to generate new molecular and 3D diversity. Much work has been dedicated to the structural analysis in solution and in the receptor-bound state, fundamental for giving a rationale to the experimentally determined bioactivity, as well as for predicting the activity of virtual compounds (in silico pre-screen). The conformational analyses in solution has been done mostly by NMR (2D gCosy, Roesy, VT, molecular dynamics, etc.). A special section is dedicated to the prediction of plausible poses of the ligands when bound to the receptors by Molecular Docking. This computational method proved to be a powerful tool for the investigation of ligand-receptor interactions, and for the design of selective agonists and antagonists. Another practical use of cyclic peptidomimetics was the synthesis and biological evaluation of cyclic analogues of endomorphin-1 lacking in a protonable amino group. The studies revealed that a inverse type II beta-turn on D-Trp-Phe constituted the bioactive conformation.

Electrostatic supramolecular assembly of charged dendritic polymers and their biological application

The aim of this study was the development of functional multilayer films through electrostatic layer by layer (LbL) assembly of dendritic macromolecules, the investigation of the fundamental properties of these multilalyered films and the study of their biological applications. rnThe synthesis of the anionic hyperbranched polyglycerols (hbPG) and the preparation of multilayers made of hbPG/phosphorus dendrimer as well as the influences of deposition conditions on multilayers were reported. The thicknesses of multilayer films increase with a decrease of molecular weight of anionic hbPGs. The multilayer films fabricated by low molecular weight hbPGs grow less regularly due to the less charged carboxylic acid groups providing the relative weaker electrostatic forces for the deposition. The thicknesses of multilayer films are reduced with increasing pH values and decreasing the concentration of NaCl. The observed changes of multilayer thickness and surface morphology could be interpreted with the aid of theories regarding the charge density and conformation of the anionic hbPG chains in solution. rnBesides the study of fundamental properties of hbPG/phosphorus multilayer films, antifouling thin films derived from hbPG layers were developed. The antifouling properties of hbPG layers were found to correlate with factors of the molecular weight of anionic hbPG and the film thickness. It was demonstrated that anionic hbPG single layer with highest molecular weight can reduce non specific protein adsorption more efficiently than single layer with lower molecular weight and all the hbPG bilayers possessed excellent property of antifouling. rnPhosphorus dendrimer multilayers were successfully prepared as the platforms to detect DNA immobilization and hybridization. The effect of NaCl concentration on the multilayer film thickness was evaluated to obtain the optimized film thickness. Making use of the multilayer deposited at the optimized condition as a substrate, a high loading of DNA probes was achieved through covalent coupling of probe DNA with the as-formed multilayer films. The hybridization of target DNA with immobilized probe DNA was then carried out and studied by SPFS. The limit of detection upon hybridization was estimated on various dendrimer multilayer platforms. The minimum detection concentration for DNA hybridization is in the same order of magnitude compared with other neutral phosphorus dendrimer systems. Furthermore, the LbL deposition of phosphorus dendrimer multilayers provided a mild and simple way to prepare platforms as DNA microarrays. rnBased on the phosphorus dendrimer multilayer systems, dendritic star polymers were employed which have more reactive groups than that phosphorus dendrimers. The as-assembled dendritic star polymer multilayer films exhibited such distinct morphology characteristics that they underwent extensive structural reorganization upon post-treatment under different pH conditions. Kinetic binding of probe DNA molecules on the outermost negatively charged dendritic surface was studied by SPR as well. The binding capacities of probe DNA on the multilayer surfaces fabricated from the first-generation and the second-generation of dendritic star polymers were compared. The improved binding capacity was achieved from the second-generation of dendritic star polymer multilayer films due to their more reactive groups. DNA hybridization reaction on dendritic multilayer films was investigated by SPFS. The similar hybridization behaviors were found on both multilayer surfaces. Meanwhile, the hybridization kinetic affinities were compared with that of phosphorus dendrimer multilayer surfaces and showed improved detection sensitivity than phosphorus dendrimer multilayer films.rn

Elemetary processes of radiation damage in organic molecules of biological interest

It was observed in the ‘80s that the radiation damage on biological systems strongly depends on processes occurring at the microscopic level, involving the elementary constituents of biological cells. Since then, lot of attention has been paid to study elementary processes of photo- and ion-chemistry of isolated organic molecule of biological interest. This work fits in this framework and aims to study the radiation damage mechanisms induced by different types of radiations on simple halogenated biomolecules used as radiosensitizers in radiotherapy. The research is focused on the photofragmentation of halogenated pyrimidine molecules (5Br-pyrimidine, 2Br-pyrimidine and 2Cl-pyrimidine) in the VUV range and on the 12C4+ ion-impact fragmentation of the 5Br-uracil and its homogeneous and hydrated clusters. Although halogen substituted pyrimidines have similar structure to the pyrimidine molecule, their photodissociation dynamics is quite different. These targets have been chosen with the purpose of investigating the effect of the specific halogen atom and site of halogenation on the fragmentation dynamics. Theoretical and experimental studies have highlighted that the site of halogenation and the type of halogen atom, lead either to the preferential breaking of the pyrimidinic ring or to the release of halogen/hydrogen radicals. The two processes can subsequently trigger different mechanisms of biological damage. To understand the effect of the environment on the fragmentation dynamic of the single molecule, the ion-induced fragmentation of homogenous and hydrated clusters of 5Br-uracil have been studied and compared to similar studies on the isolated molecule. The results show that the “protective effect” of the environment on the single molecule hold in the homogeneous clusters, but not in the hydrated clusters, where several hydrated fragments have been observed. This indicates that the presence of water molecules can inhibit some fragmentation channels and promote the keto-enol tautomerization, which is very important in the mutagenesis of the DNA.

Boron determination in biological samples - Intercomparison of three analytical methods to assist development of a treatment protocol for neoplastic diseases of the liver with Boron Neutron Capture Therapy

Die Bor-Neuroneneinfang-Therapie (engl.: Boron Neutron Capture Therapy, BNCT) ist eine indirekte Strahlentherapie, welche durch die gezielte Freisetzung von dicht ionisierender Strahlung Tumorzellen zerstört. Die freigesetzten Ionen sind Spaltfragmente einer Kernreaktion, bei welcher das Isotop 10B ein niederenergetisches (thermisches) Neutron einfängt. Das 10B wird durch ein spezielles Borpräparat in den Tumorzellen angereichert, welches selbst nicht radioaktiv ist. rnAn der Johannes Gutenberg-Universität Mainz wurde die Forschung für die Anwendung eines klinischen Behandlungsprotokolls durch zwei Heilversuche bei Patienten mit kolorektalen Lebermetastasen an der Universität Pavia, Italien, angeregt, bei denen die Leber außerhalb des Körpers in einem Forschungsreaktor bestrahlt wurde. Als erster Schritt wurde in Kooperation verschiedener universitärer Institute eine klinische Studie zur Bestimmung klinisch relevanter Parameter wie der Borverteilung in verschiedenen Geweben und dem pharmakokinetischen Aufnahmeverhalten des Borpräparates initiiert.rnDie Borkonzentration in den Gewebeproben wurde hinsichtlich ihrer räumlichen Verteilung in verschiedenen Zellarealen bestimmt, um mehr über das Aufnahmeverhalten der Zellen für das BPA im Hinblick auf ihre biologischen Charakteristika zu erfahren. Die Borbestimung wurde per Quantitative Neutron Capture Radiography, Prompt Gamma Activation Analysis und Inductively Coupled Plasma Mass Spectroscopy parallel zur histologischen Analyse des Gewebes durchgeführt. Es war möglich zu zeigen, dass in Proben aus Tumorgewebe und aus tumorfreiem Gewebe mit unterschiedlichen morphologischen Eigenschaften eine sehr heterogene Borverteilung vorliegt. Die Ergebnisse der Blutproben werden für die Erstellung eines pharmakokinetischen Modells verwendet und sind in Übereinstimmung mit existierenden pharmakokinetische Modellen. Zusätzlich wurden die Methoden zur Borbestimmung über speziell hergestellte Referenzstandards untereinander verglichen. Dabei wurde eine gute Übereinstimmung der Ergebnisse festgestellt, ferner wurde für alle biologischen Proben Standardanalyseprotokolle erstellt.rnDie bisher erhaltenen Ergebnisse der klinischen Studie sind vielversprechend, lassen aber noch keine endgültigen Schlussfolgerungen hinsichtlich der Wirksamkeit von BNCT für maligne Lebererkrankungen zu. rn

Transport of nanoparticles into polymersomes : a minimal model system of particles passage through biological membranes

This thesis focuses on the design and characterization of a novel, artificial minimal model membrane system with chosen physical parameters to mimic a nanoparticle uptake process driven exclusively by adhesion and softness of the bilayer. The realization is based on polymersomes composed of poly(dimethylsiloxane)-b-poly(2-methyloxazoline) (PMDS-b-PMOXA) and nanoscopic colloidal particles (polystyrene, silica), and the utilization of powerful characterization techniques. rnPDMS-b-PMOXA polymersomes with a radius, Rh ~100 nm, a size polydispersity, PD = 1.1 and a membrane thickness, h = 16 nm, were prepared using the film rehydratation method. Due to the suitable mechanical properties (Young’s modulus of ~17 MPa and a bending modulus of ~7⋅10-8 J) along with the long-term stability and the modifiability, these kind of polymersomes can be used as model membranes to study physical and physicochemical aspects of transmembrane transport of nanoparticles. A combination of photon (PCS) and fluorescence (FCS) correlation spectroscopies optimizes species selectivity, necessary for a unique internalization study encompassing two main efforts. rnFor the proof of concepts, the first effort focused on the interaction of nanoparticles (Rh NP SiO2 = 14 nm, Rh NP PS = 16 nm; cNP = 0.1 gL-1) and polymersomes (Rh P = 112 nm; cP = 0.045 gL-1) with fixed size and concentration. Identification of a modified form factor of the polymersome entities, selectively seen in the PCS experiment, enabled a precise monitor and quantitative description of the incorporation process. Combining PCS and FCS led to the estimation of the incorporated particles per polymersome (about 8 in the examined system) and the development of an appropriate methodology for the kinetics and dynamics of the internalization process. rnThe second effort aimed at the establishment of the necessary phenomenology to facilitate comparison with theories. The size and concentration of the nanoparticles were chosen as the most important system variables (Rh NP = 14 - 57 nm; cNP = 0.05 - 0.2 gL-1). It was revealed that the incorporation process could be controlled to a significant extent by changing the nanoparticles size and concentration. Average number of 7 up to 11 NPs with Rh NP = 14 nm and 3 up to 6 NPs with Rh NP = 25 nm can be internalized into the present polymersomes by changing initial nanoparticles concentration in the range 0.1- 0.2 gL-1. Rapid internalization of the particles by polymersomes is observed only above a critical threshold particles concentration, dependent on the nanoparticle size. rnWith regard possible pathways for the particle uptake, cryogenic transmission electron microscopy (cryo-TEM) has revealed two different incorporation mechanisms depending on the size of the involved nanoparticles: cooperative incorporation of nanoparticles groups or single nanoparticles incorporation. Conditions for nanoparticle uptake and controlled filling of polymersomes were presented. rnIn the framework of this thesis, the experimental observation of transmembrane transport of spherical PS and SiO2 NPs into polymersomes via an internalization process was reported and examined quantitatively for the first time. rnIn a summary the work performed in frames of this thesis might have significant impact on cell model systems’ development and thus improved understanding of transmembrane transport processes. The present experimental findings help create the missing phenomenology necessary for a detailed understanding of a phenomenon with great relevance in transmembrane transport. The fact that transmembrane transport of nanoparticles can be performed by artificial model system without any additional stimuli has a fundamental impact on the understanding, not only of the nanoparticle invagination process but also of the interaction of nanoparticles with biological as well as polymeric membranes. rn

Photoactive nanodevices for potential biological applications

Biological systems are complex and highly organized architectures governed by noncovalent interactions, which are responsible for molecular recognition, self-assembly, self-organization, adaptation and evolution processes. These systems provided the inspiration for the development of supramolecular chemistry, that aimed at the design of artificial multicomponent molecular assemblies, namely supramolecular systems, properly designed to perform different operations: each constituting unit performs a single act, whereas the entire supramolecular system is able to execute a more complex function, resulting from the cooperation of the constituting components. Supramolecular chemistry deals with the development of molecular systems able to mimic naturally occurring events, for example complexation and self-assembly through the establishment of noncovalent interactions. Moreover, the application of external stimuli, such as light, allows to perform these operations in a time- and space-controlled manner. These systems can interact with biological systems and, thus, can be applied for bioimaging, therapeutic and drug delivery purposes. In this work the study of biocompatible supramolecular species able to interact with light is presented. The first part deals with the photophysical, photochemical and electrochemical characterization of water-soluble blue emitting triazoloquinolinium and triazolopyridinium salts. Moreover, their interaction with DNA has been explored, in the perspective of developing water-soluble systems for bioimaging applications. In the second part, the effect exerted by the presence of azobenzene-bearing supramolecular species in liposomes, inserted both in the phospholipid bilayer and in the in the aqueous core of vesicles has been studied, in order to develop systems able to deliver small molecules and ions in a photocontrolled manner. Moreover, the versatility of azobenzene and its broad range of applications have been highlighted, since conjugated oligoazobenzene derivatives proved not to be adequate to be inserted in the phospholipid bilayer of liposomes, but their electrochemical properties made them interesting candidates as electron acceptor materials for photovoltaic applications.