Chitosan (CS) is a man-made sugar based biopolymer derived from chitin, the second most abundant natural polymer after cellulose. Chitin is sourced from crustacean species such as shrimps and crabs. The chemical structure of chitin contains N-Acetyl D-glucosamine monomer units which forms CS upon deacetylation. In CS, ?-(1-4) linked D-glucosamine units are randomly distributed. Approximately 75% - 80% sugar units contains primary amine groups in commercially available low molecular weight CS. Biodegradability, low toxicity, mucoadhesive and transfecting properties of CS polymer are attractive for applications as oral and nasal drug delivery systems. Chitosan polymer is water insoluble at neutral pH. To solubilize CS, dilute mineral acid (such as hydrochloric acid and nitric acid) or organic acid (such as acetic acid) is often used. CS contains both hydroxyl and primary amine groups in its structure. In acidic solution, the amine functional groups become protonated (positively charged). Positively charged CS remains stable only in low pH condition due to electrostatic repulsion of charged polymer segments. Therefore, by using a suitable anionic (negatively charged) cross-linker, stable CS particles (such as nanoparticles and microspheres) can be prepared. This is popularly known as ionic gelation method. Extensive studies have been done on the synthesis of drug loaded CS particles where particle integrity is maintained by ionic gelation using tripolyphosphate (TPP, an anionic cross-linker). Drug encapsulated CS-TPP composite particles are shown to maintain biodegradability and biocompatibility. The CS-TPP composite particles exhibits very limited dispersibility at neutral pH conditions specifically in neutral buffered conditions. A number of biomedical applications (including systemic drug formulations) however demands buffer-stable CS composite particles for achieving optimal therapeutic outcome. To overcome the above dispersibility issues, CS polymer and CS particles units have been chemically modified using water soluble motifs (such as water soluble polymer or ligands). This approach is very cumbersome and usually involves multiple purification steps. Chemical modification of natural CS chain introduces risks of compromising biodegradability and biocompatibility. Therefore, there is a strong need for developing a straightforward method of making water soluble CS and CS particles. Chapter 1 of this dissertation presents an overview of the CS polymer, various applications of CS polymers, methods of making CS polymers and CS particles, current limitations of synthesis methods for preparing stable chitosan particles at neutral pH conditions and finally delineates the scope of the proposed research work. Chapter 2 describes development of chemo-hydrothermal synthesis method for producing water soluble CS polymer and water dispersible CS composite particles. In this method, a chemical (depolymerizing agent) is used to treat CS polymer in a hydrothermal (high temperature and high pressure) condition. Two types of depolymerizing agents have been used, an inorganic acid (e.g. hydrochloric acid, HCl) and a bicarboxylic organic acid (e.g. tartaric acid, TA). In both cases, 100% depolymerized CS polymer was obtained. Chemical characteristics of the depolymerized CS were comparable to acid solubilized CS. CS polymer exhibits weak fluorescence. Interestingly, hydrothermally depolymerized CS shows strong fluorescence properties irrespective of the nature of depolymerizing agent used. TA not only depolymerized CS but also formed CS-TA composite particulate structures in solution via self-assembly. The CS-TA composite particles are stable in a wide pH range from 5 to 11. Detailed spectroscopic and microscopic studies have been done to understand the basic mechanism of particle formation and increase in fluorescence properties (i.e. structure-property relationship). Usefulness of CS-TA in solubilizing water-insoluble cargos (such as fluorescein isothiocyanate, FITC) has been demonstrated. Chapter 3 is focused on hydrothermal synthesis of mixed-valence copper (Cu) oxide loaded CS-TA composite particles and their characterization. Crystalline Cu oxide nanoparticles were coated with the CS-TA layer. Water dispersibility of Cu oxide greatly improved upon coating with CS-TA material. To demonstrate catalytic activity of Cu-oxide loaded CS-TA film in sequestering carbon dioxide (CO2), an electrochemical setup was used. Electrochemical reduction of CO2 was successfully demonstrated. It was observed that CS-TA environment not only maintained catalytic properties of Cu oxide but also allowed solution processing of Cu-oxide film onto the electrode surface. Chapter 4 discusses a convenient method of making monodispersed water dispersible Cu loaded chitosan nanoparticles (Cu-CS) using HCl depolymerized CS polymer. The purpose of this study was to investigate if there was any improvement in antibacterial properties of Cu-CS nanoparticles prepared using hydrothermally treated CS polymer. Interestingly, it was observed that the antibacterial efficacy of Cu was not compromised in Cu-CS nanoparticles. Moreover, the materials exhibited improvement in antibacterial efficacy against both Gram-negative and Gram-positive bacteria species. A plausible mechanism has been proposed to explain antibacterial results. Chapter 5 summarizes major findings of this dissertation research and presents future research directions.

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Il fattore antisecretivo (AF) è una proteina secreta nel plasma e nei tessuti dei mammiferi, che ha dimostrato di essere un potente inibitore dell’ipersecrezione intestinale e dell'infiammazione. Dopo il bando degli AGP, diversi approcci nutrizionali finalizzati all’ottimizzazione della fase di transizione dello svezzamento e alla riduzione delle malattie intestinali sono stati proposti, ma con scarsi risultati rispetto agli antibiotici. Le diete in grado di indurre la secrezione endogena di AF potrebbero essere una valida alternativa agli AGP. Si ritiene che AF contenuto nel colostro e nel latte delle scrofe possa essere un fattore di protezione contro la diarrea nei suinetti. E’ stato dimostrato che è possibile incrementare il livello di AF nel plasma attraverso la dieta, sia nell’uomo che negli animali, con l’ingestione di cereali sottoposti ad un particolare processo idro-termico (HPC). In questa tesi è stato studiato l’effetto dell’aggiunta di HPC alla dieta di suinetti svezzati, sulle performance e sullo stato infiammatorio dell’epitelio intestinale. La supplementazione della dieta con HPC ha migliorato ADG e FCR durante tutto il periodo sperimentale. I valori I-FABP, considerato un parametro del danneggiamento della mucosa intestinale, nel plasma dei suinetti sono risultati bassi in tutti i gruppi e non influenzati dalla dieta. L'attività antisecretiva di AF-16, il dominio attivo della proteina, è stata valutata mediate su colture cellulari IPEC-J2 trattate con tossina colerica (CT) in Ussing chamber. AF-16 non ha inibito l’incremento di Isc indotto da CT. E’ stato osservato un ridotto incremento di Isc durante la somministrazione contemporanea di AF-16 e CT. L'attività anti-infiammatoria di AF-16 è stata studiata in colture di macrofagi RAW 264,7, trattati con LPS, e su macrofagi alveolari di suino stimolati con PMA. AF-16 ha ridotto la produzione di NO nei macrofagi stimolati con LPS in funzione del dosaggio, con maggiori effetti a dosaggi più elevati. L’effetto antiinfiammatorio di AF-16 non è stata confermata dai ROS test, neanche usando dosi elevate di peptide. I risultati confermano l’efficacia dei cereali HPC come promotori della crescita nei suinetti. Ulteriori studi in vitro sono necessari per capire il meccanismo d’azione di AF. / The antisecretory factor (AF) is a protein secreted in plasma and other tissue fluids in mammalians which was shown to be a potent inhibitor of intestinal fluid secretion and inflammation. After the AGP ban, several nutritional approaches aimed to the optimization of the weaning transition and reduction of gut diseases have been proposed, but the success rates are really low compared to antimicrobials. AF-inducing diets appear to be suitable alternatives to AGP. Indeed AF content in sows’ colostrum and milk appears to be a protection factor against diarrhoea in suckling piglets. Increased AF level in plasma by dietary means, such as feeding hydro-thermally processed cereals (HPC), has been demonstrated in human and animals. We tested the effect of two different HPC level of inclusion in a wheat-barley based diet on weaned piglets growth performance, reared in an experimental farm. The results confirmed the efficacy of HPC as growth promoters in piglet nutrition: HPC supplementation improved ADG and FCR. In vitro test were made to study the antisecretory and anti-inflammatory properties of the AF protein and its mechanism of action by using AF-16, the active region of the protein. The Ussing chamber experiments performed on polarized IPEC-J2 cells confirmed the neuronal involvement in the antisecretory activity of AF. In fact, AF-16 did not inhibit CT-induced Isc. A slower rate of Isc increase was observed during the simultaneous administration with AF-16 and CT. High dosages of AF-16 were found to reduce the LPS-stimulated NO production in RAW264.7 cell, thus supporting the hypothesis of an anti-inflammatory action. On the contrary, no significant results were obtained on PMA-stimulated ROS generation in pig alveolar macrophages.

Hydrothermally treated biomass could not only be used as a fuel or a fertilizer but it can also be refined into high-value products. Activated carbons are one of those. In the studies of this thesis, four different hydrothermally carbonized (HTC) biomasses, including horse manure, grass cuttings, beer waste and biosludge, have been successfully made into activated carbons. The activated carbon materials were in the forms of powdered activated carbons, powdered composites of activated carbon and iron oxide nano-crystals, and activated carbon discs. The HTC biomasses and the activated carbons were characterized and analyzed using several methods. The biomasses were carbonized to different extent during the hydrothermal treatment, which depended on the different natures of the biomasses. The HTC biomasses were activated into powdered activated carbons by both physical activation, using CO2, and by chemical activation, using H3PO4. Full factorial design matrices were applied to design experiments and study the influence of different parameters used during both physical and chemical activation. Activated carbons with embedded iron oxide nanoparticles were synthesized through hydrothermal carbonization followed by CO2 activation. These composites had high surface areas and showed a strong magnetism, and the powders could be separated from liquid phase by applying a magnetic field. Strong and dense activated carbon discs were also prepared from powdered HTC beer waste by pulsed current processing (PCP) and a subsequent CO2 activation procedure. The potential for carbon dioxide separation from nitrogen, and methylene blue adsorption in aqueous solution, were assessed for the powdered activated carbons produced from HTC biomasses. They showed good performance in both applications. / <p>At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 1: In press.</p>