In recent years, the application of microparticles in different fields including cosmetics, agriculture, pharmaceutics and biomedicine has been widely investigated.In this project, we aimed to improve the preparative methods for chitosan–triphosphate microparticles (Cs/TPP) for perspective application in the fields of cell encapsulation and controlled drug delivery. Prior to the preparation of chitosan-based microparticles, in order to confirm good biocompatibility and reproducibility, protocols have been established for the purification and characterisation of chitosan including the assessment of average molecular weight, protein content and degree of deacetylation.This study then primarily focused on the use of β-glycerophosphate (βGP) as an excipient, which is known to solubilize chitosan at neutral pH, thus allowing the preparation of chitosan microparticles (microspheres and toroidal) via ionotropic gelation under physiological conditions. The preparation of Cs-βGP/TPP microparticles was optimized varying several key process variables (concentration, flow rate, and frequency) and these microparticles were produced with a narrow size distribution (400 – 500 μm, spherical shape) and compared to Cs/TPP controls. The main result was the possibility to perform this process at neutral pH, although we have also demonstrated an improved toughness and cross-linking density of these microspheres as a result of the presence of β-glycerophosphate. Further, we have investigated the application of this method to a toroidal geometry, which provides advantages in terms of better transfer of oxygen and nutrients to any encapsulated materials. Cs/TPP and Cs-βGP/TPP ‘micro-doughnuts’ were also prepared and characterised. This research highlighted the evidence of a higher cross-linking density of these microparticles in comparsion with the spherical ones. In order to optimise the physicochemical characteristics of these microparticles for future applications as biomaterials, the surface of Cs/TPP and Cs-βGP/TPP microparticles was modified through an additional polyelectrolyte complexation with poly (sodium 4- styrene sulphonate) (PSS). The improved toughness and cross-linking density was confirmed by measuring the mechanical properties and solid content which indicated the successful complexation of PSS onto the surface of these microparticles.

Orientador: Ângela Maria Moraes / Dissertação (mestrado-) - Universidade Estadual de Campinas, Faculdade de Engenharia Química / Made available in DSpace on 2018-08-22T01:56:28Z (GMT). No. of bitstreams: 1 Pires_AnaLuizaResende_M.pdf: 3161193 bytes, checksum: c48592744224fd8b24f5563cdc1a3415 (MD5) Previous issue date: 2013 / Resumo: Polímeros biodegradáveis estão sendo amplamente aplicados na constituição de curativos para o tratamento de lesões de pele de diferentes origens, estando a quitosana e o alginato dentre os mais estudados. A associação de alginato e quitosana em curativos pode atuar de maneira positiva na absorção de fluidos das lesões, além de propiciar a incorporação de fármacos. Entretanto, a flexibilidade do dispositivo é baixa e o processo de obtenção é lento em função de etapas como a correção do pH para a neutralidade, secagem e reticulação. Neste trabalho teve-se por objetivo melhorar as propriedades mecânicas de membranas de quitosana e alginato pela incorporação de um gel de silicone comercial (Silpuran® 2130 A/B), avaliar os efeitos da correção do pH em etapa única, da temperatura de secagem e também da eliminação das etapas de reticulação nas características de membranas de quitosana e alginato na presença e ausência de AlphaSan®RC2000, um agente microbicida à base de prata. As membranas foram caracterizadas quanto à espessura, à absorção de fluidos, à perda de massa quando expostas a diferentes fluidos fisiológicos e à resistência mecânica. Análises complementares de FTIR e EDS também foram realizadas. Os resultados obtidos mostraram que as membranas secas a 60 °C apresentaram superfície mais lisa que as secas a 37 °C. A espessura diminuiu com a correção do pH para 7,0 e aumentou na presença de AlphaSan®RC2000. Para as formulações livres do antimicrobiano, a elevação do pH aumentou a absorção de soro fetal bovino e solução salina e aumentou também a perda de massa nesta última solução. Já as amostras obtidas na presença de AlphaSan® RC2000 apresentaram aumento na absorção de água. A elevação da temperatura reduziu somente a perda de massa após o contato com a água. A reticulação, após a secagem, com CaCl2 a 2% mostrou-se indispensável para a estabilização das membranas, resultando em materiais com aspecto mais rugoso e maior espessura. A absorção de fluidos, a estabilidade em diferentes fluidos e a resistência mecânica diminuíram tanto na presença quanto na ausência de prata. Análises por EDS e FTIR mostraram que tanto o AlphaSan® RC2000 quanto o Silpuran® 2130 A/B, não são removidos após a etapa de lavagem das membranas. As formulações contendo o composto siliconado apresentaram-se mais homogêneas, flexíveis e com maior caráter adesivo, assim como menor espessura. A absorção de soluções por estas formulações foi menor e a estabilidade em solução foi maior. A resistência mecânica aumentou significativamente com a incorporação de silicone, atingindo-se uma tensão de ruptura máxima de 63 MPa. Dessa forma, é viável a diminuição do tempo de processamento das membranas pelo ajuste do pH em etapa única, pelo uso de temperaturas mais elevadas de secagem e pela eliminação da etapa de reticulação primária. A incorporação de Silpuran® 2130 A/B mostrou-se uma boa alternativa para a melhora das propriedades mecânicas dos curativos de quitosana e alginato / Abstract: Biodegradable polymers are widely applied in the constitution of dressings for treating various types of skin lesions, being chitosan and alginate two of the most studied raw materials. The association between alginate and chitosan in wound dressings can act positively in the absorption of fluids from lesions, allowing also the incorporation of drugs. However, the device flexibility is frequently low and its production is time-consuming due to steps such as adjusting the pH to neutrality, sample drying and crosslinking. This study aimed to improve the mechanical properties of chitosan-alginate wound dressings by incorporating a commercial silicone gel (Silpuran® 2130 A/B) and to evaluate the effects of pH correction in one step, of the drying temperature and also of the elimination of the crosslinking steps in the characteristics of chitosan-alginate membranes obtained in the presence and absence of AlphaSan® RC2000, an antimicrobial agent containing silver in its composition. The membranes were characterized regarding thickness, absorption of fluids, mass loss in different fluids and mechanical resistance. Complementary analyzes of FTIR and EDS were also performed. The results showed that membranes dried at 60 °C presented smoother surfaces than when dried at 37 °C. Membrane thickness decreased with the correction of the pH to 7,0 and increased in the presence of AlphaSan® RC2000. For formulations free of the antimicrobial agent, the pH adjustment increased bovine fetal serum and saline solution absorption, increasing also sample mass loss in the latter solution. Devices containing AlphaSan® RC2000, on the other hand, had increased water absorption. The increase of drying temperature, however, reduced mass loss due to prolonged contact with water. The crosslinking with CaCl2 2% after a drying step was shown to be essential for membrane stabilization, conducting to materials with less smooth surfaces and with greater thickness. The absorption of fluids decreased for samples prepared both in the presence and absence of silver, as did membrane stability and mechanical properties. EDS and FTIR analyzes showed that both AlphaSan® RC2000 and Silpuran® 2130 A/B were not removed from the membranes after the washing step. The formulations incorporating the silicone compound showed more homogeneous surfaces, greater flexibility and adhesivity, as well as lower thickness. The solution absorption was lower and membrane stability was higher for those devices. The mechanical strength increased with the incorporation of silicone, reaching a maximum of 63 MPa. Thus, it is viable to decrease the processing time of the membranes by pH adjustment in a single step, through the use of higher temperatures during drying and by the eliminating of the primary crosslinking step. The incorporation of Silpuran® 2130 A/B proved to be a good alternative for the improvement of the mechanical properties of chitosan and alginate dressings / Mestrado / Desenvolvimento de Processos Biotecnologicos / Mestra em Engenharia Química

This study evaluated the potential toxicity, genotoxicity, and cellular uptake of various sizes and concentrations of chitosan (CS) nanoparticles cultured with normal human dental pulp cells. Normal human dental pulp cells (hDPCs) were derived from human dental pulp tissues and cultured with (50–67) nm and (318–350) nm CS-nanoparticles in concentrations of 0.1 mg/mL, 0.5 mg/mL, 1 mg/mL, 2 mg/mL, and 4 mg/mL as study groups and 0 mg/mL as a control group for time intervals of 16 hours, 24 hours, 3 days, 7 days and 14 days. Attachment efficiency and proliferation rate were assessed by measuring the optical density of crystal violet-stained cells. Cell viability was determined by the activity of mitochondrial dehydrogenase enzymes. Genotoxicity was assessed using the cytokinesis-block micronucleus method and by measuring the fluorescent intensity of phosphorylated H2AX nuclear foci. Cellular uptake was determined by tagging chitosan nanoparticles with FITC stain and then measuring the fluorescence intensity of FITC-tagged chitosan nanoparticles using a spectrophotometer. Statistical analysis was performed using chi-square, one-way ANOVA, and post-hoc Tukey tests. All concentrations of the (50–67) nm group significantly reduced attachment efficiency in comparison with control (P< 0.01) and with (318–350) nm group (p<0.01). Proliferation rate and cell viability were significantly reduced in cells exposed to various concentrations of (50-67) nm chitosan when compared to (318-350) nm group (P<0.05) and control group (P<0.05). For both size groups, higher concentrations significantly showed lower proliferation rate and cell viability when compared to lower concentration (P< 0.01). CS-nanoparticles were able to internalize hDPCs and significantly induced micronuclei, nuclear buds, and pH2AX at concentrations of 0.5 mg/mL and 2 mg/mL as compared to 0.1 mg/mL (P<0.01) and control groups (P< 0.01). At both the 0.5 mg/mL and 2 mg/mL concentrations, (50–67) nm chitosan significantly induced higher proportions of micronuclei (P= 0.001), nuclear buds (P= 0.009), and pH2AX nuclear foci (P= 0.00004) compared to (318–350) nm chitosan. In conclusion, CS-nanoparticles at sizes (50–67) nm and (318–350) nm at a concentration of (0.5–4) mg/mL internalized hDPCs and exhibited cytotoxic and genotoxic effects in dose-dependent and size-associated manners.

Articular cartilage provides an almost frictionless surface for the articulating ends of the bone. Cartilage functions to lubricate and transmit compressive forces resulting from joint loading and impact. If the cartilage is damaged, through traumatic injury or disease, it lacks the ability of self-repairing as the tissue lacks vascular system. If the injuries to articular cartilage are left untreated, they may progress to Osteoarthritis. Osteoarthritis, a degenerative disease, is one of the leading disabilities in the United States. Tissue engineering has the potential to regenerate healthy hyaline cartilage, which can alleviate pain and restore the functions of normal tissue. This study explores the production of engineered cartilage on top of composite calcium phosphate scaffold. The current research is related to a biphasic approach to cartilage tissue engineering — in which one layer supports to form subchondral bone (osteogenesis) and another supports cartilage formation (chondrogenesis). Chondrocyte and bone marrow-derived stem cell attachment to chitosan will be investigated for producing a bilayered construct for osteochondral repair. The main objectives of my research include the following: attachment and proliferation of human mesenchymal stem cells on chitosan calcium phosphate scaffolds, techniques to create a biphasic construct, the effect of coating chitosan calcium phosphate scaffolds with type I collagen and determining the ideal bead size for making chitosan calcium phosphate scaffolds.

Graphene Oxide (GO) has emerged within the last decade as a next generation material for water treatment. Fabrication of graphene oxide membranes has been limited in scale and application due to repulsive hydration forces causing GO layers to electrostatically separate. In this study, chitosan is utilized to increase GO stability in the wet state through interactions with the negatively charged GO sheets (CSGO). This simple aqueous self-assembly allows scalable fabrication and enhanced stability for membrane applications in crosslow. The CSGO membrane’s performance was tested in a crosslow reactor and challenged with methylene blue at concentrations ranging from 1 to 100 ppm at 345 kPa with fluxes ranging from 1 to 4.5 L/(m2 hr) with 100% removal by physical rejection. This work demonstrates that the CSGO composite matrix is a potential alternative to traditional polymeric membranes for water treatment using a renewable biopolymer and minimal chemical input.

>Magister Scientiae - MSc / Different nanoparticles have the ability to improve plant tolerance to drought stress. In the study we report for the first time novel morpholin-4-ium 4-methoxyphenyl (morpholino) phosphinodithioate capped- gold nanoparticles (GYY4137-capped AuNPs). The GYY4137 is a slow releasing hydrogen sulfide (H2S) donor. The GYY4137 AuNPs compared to preliminary experiments of L-serine and L-threonine gold nanoparticles. The nanoparticles were prepared using a simple reflux reduction method in a rolling boil flask at 80 oC. The uncapped GYY4137-AuNPs were relatively stable and had surface plasmon resonance at 562 nm compared to 524 nm and 560 nm of serine-AuNPs and threonine-AuNPs. The nanoparticles were capped with different concentrations (0.1-5 %) of water-soluble poly (ethylene) glycol (PEG) (Mw300) and 0.2% chitosan. The PEG did not fully encapsulate the gold nanoparticles, while the chitosan successfully produced positively charged gold nanoparticles. The formation of chitosan capped GYY4137-AuNPs were verified with UV-Visible spectroscopy (UV-Vis), High Resolution Transmission electron microscopy (HRTEM), Dynamic Light scattering (DLS) and the Zetasizer. The UV-Vis, HRTEM and STEM verified chitosan capped nanoparticles had a surface plasmon resonance peak at 560 nm, with icosahedral, tetrahedron and spherical shaped nanoparticles as the serine-AuNPs that absorb at 560 nm. The agglomerated threonine-AuNPs had a maximum absorbance peak at 524 nm. The chitosan GYY4137-AuNPs had hydrodynamic size of 347.9 nm and zeta potential of + 47 mV, while serine-AuNPs and threonine-AuNPs had hydrodynamic size of 110 nm, zeta potential of -2.9 mV and -230 mV respectively. The polydispersity index (PDI) of the chitosan capped gold nanoparticles was 0.357 compared to 0.406 of both the amino acid gold nanoparticles. The polydispersity index (PDI) showed that the nanoparticles were polydispersed nanoparticles with broad size range as confirmed by the HRTEM and STEM results/ of chitosan capped GYY4137-AuNPs. The sizes of the nanoparticles were 100 nm and 60 nm for GYY4137-AuNPs while the size serine-AuNPs were 60 nm. The gold nanoparticles structural compositions were further confirmed by energy-dispersive X-ray spectrometry (EDX) and Attenuated total reflection infrared spectroscopy (ATR-IR). EDX results proved successful gold nanoparticles synthesis by presence of the element Au in all three nanoparticles and the chitosan GYY4137-AuNPs had 48. 56 wt. % of gold. The FTIR-ATR new bands formation shows that new chemical bonds are formed between the reducing agents, the precursor gold salt solution and capping agents. The shifts showed successful encapsulation with chitosan in GYY4137-AuNPs. The chitosan encapsulation improved surface charge and reactivity of the gold nanoparticles to improve delivery of the hydrogen sulfide donor GYY4137 for later applications to plants.

No description available.

Introduction: The primary reason for endodontic treatment failure is microbial infection inside the root canal system by different microbes including Enterococcus faecalis. Endodontic sealers with Chitosan (CS), a compound with antimicrobial effects, can decrease the growth of microorganisms and potentially prevent failure. Purpose: The aim of this study was to assess the antimicrobial activity ex-vivo and flowability of calcium silicate root canal sealers with added CS fillers. Materials and methods: EndoSequence (BC) sealer was mixed with CS fillers in two different concentrations (5% and 10%wt). Flowability was analyzed according to ISO-6876. The antibacterial assessment was conducted using an ex-vivo model. Extracted single-rooted teeth were infected with E. faecalis for 7 days. Follow the root canal treatment, the sealers (BC, BC+CS-5%, BC+CS-10%, N=6) were used for obturation and re-incubated in the bioreactor for 24 hours. Untreated teeth were used as positive control. Cell viability (CFU/mL) and fluorescence microscopy were used to determine the number of bacteria at the bonded interface. ANOVA was used for statistical analysis. Results: Addition of CS fillers into BC resulted in a significant decrease in flowability (BC:22mm, BC+CS-5%:19mm, BC+CS-10%:16mm). All evaluated sealers showed antibacterial activity. The addition of CS significantly reduced the viable cells (log(CFU/mL)) (CS-5%:5.3, CS-10%:4.6) compared with the BC (5.9) and positive control (7.2). Fluorescence analysis showed that CS-Sealers group had an increased number of dead cells (CS-10%:77.3%, CS-5%:64.4%) compared with the BC-Sealer (37.9%) and positive control (23.3%). Conclusion: Endodontic sealers with added CS enhanced antimicrobial effects of the sealer. However, adding more than 5% of CS affects the flow properties of the sealer. / Oral Biology

The main objectives of this study were to investigate the applicability of chitosan as an effective wood preservative against subterranean termites, conduct metagenomic analysis of the bacterial hindgut community of Reticulitermes flavipes exposed to chitosan-treated wood, and perform chitosanase activity assay of metagenomics suggested bacterial species potentially responsible for chitosan breakdown. Chitosan showed termiticidal effects on subterranean termites at varying retention levels. Termite mortality increased when exposed to samples treated with higher chitosan concentration solutions. Approximately 40 - 100% of chitosan retained in treated-wood was leached depending on the initial retention. Post-leaching results indicate chitosan is not suitable for protection against both subterranean termites in outdoor conditions, but should be effective in non-leaching/indoor applications. For metagenomic analysis of the bacterial hindgut community of Reticulitermes flavipes, two methods were used for sequence data interpretation. The Illumina BaseSpace program identified twenty-six bacteria phyla with significant differences in abundance between the chitosan-treated and control groups. The second method, mothur, identified fifteen bacterial phyla also with significant differences in abundance between both treatment groups. Similar bacterial taxa were uniquely assigned to samples from termites fed on chitosan-treated wood using both methods. These results suggest a treatment driven effect on the hindgut bacteria diversity. While majority of the bacterial taxa were common to both methods, inconsistencies detected using the BaseSpace program suggests that the Greengenes database in its present state is not reliable for 16S rRNA gene sequence analysis. As for chitosanase activity of bacterial species with significance abundance from chitosan-treated wood exposed termites, three bacteria species, Lactococcus raffinolactis, Lactococcus lactis, and Dysgonomonas gadei, were examined. After culturing on chitosan media plates and broth, no conclusive activity could be detected from all three species. Further studies need to be conducted to understand the mechanism of chitosan toxicity to termites and insects in general and to prevent chitosan leaching from treated wood. A comparative metatransciptomic study needs to be implemented to supplement the metagenomic study performed herein, so as to elucidate the exact bacteria species involved in chitosan breakdown and the enzymes produced. Also, other bacterial species suggested by the metagenomic data to possess chitosanase activity should be investigated.

Cellulose nanofibers (CNFs) have established widespread attention in various industries with their potential applications. Production of CNFs from agricultural post harvesting wastes has several cost-effective and eco-friendly benefits. The objective of this research was to prepare four different types of CNFs from cotton stalks by different chemical treatments followed by ultra-sonication. CNFs via untreated bleached pulp, sulfuric acid hydrolysis, and TEMPO [(2,2,6,6-tetramethylpiperidin-1-yl) oxy radical]-mediated oxidation process were produced. Physical and chemical properties of these CNFs were investigated by morphological (FE-SEM, AFM), structural (FTIR), and thermal gravimetric analysis (TGA). Developed TEMPO-oxidized cellulose nanofibers (TEMPO-CNFs) were brighter and higher in yields (>90%). It was the first time uniform and very small sized (3-15 nm diameter and 10-100 nm length) nanofibers were produced. In application purpose, TEMPO-CNFs were introduced into chitosan matrix (prepared from shrimp exoskeletons) for the development of bionanocomposite food packaging films and into chitin for hydrogels preparation in order to eliminate heavy metals from water bodies. Development of bionanocomposite films (chitosan/TEMPO-CNFs compositions) was an effective and complete green approach with enhanced mechanical and barrier properties. Also use of TEMPO-CNFs in this method makes it possible to produce flexible, transparent, and low cost food packaging films with good antimicrobial activity against Salmonella enterica, E. coli O157:H7, and Listeria monocytogenes. In a similar way, generation of recyclable biobased adsorbents (chitin/TEMPO-CNFs compositions) with superior adsorption capacity and high surface area were effectively used in lead (Pb2+) removal from aqueous solutions, thus provide new opportunities as economical and environmentally friendly green remediation.