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Entwicklung von Fertigungstechnologien zur Herstellung biomimetischer faserbasierter Scaffolds aus Kollagen für das Tissue Engineering und die regenerative MedizinTonndorf, Robert 14 June 2022 (has links)
Die enormen Fortschritte und Erkenntnisse der Medizin und der damit einhergehenden gestiegenen mittleren globalen Lebenserwartung von indes knapp 75 Jahren fußen auch auf den medizinischen Entwicklungen des 20. Jahrhunderts, da durch diese z. B. infektiöse und onkologische Erkrankungen, Diabetes, Bluthochdruck, Herzinsuffizienz, Magengeschwüre, Depressionen, Hämophilie und andere Krankheiten erfolgreich therapiert werden können. Die entwickelten Therapiemethoden beruhten im Wesentlichen auf chirurgischen und intensivmedizinischen Neuerungen, chemischen Wirkstoffen, belastungsfähigen Implantaten und extrakorporalen Systemen. Im 21. Jahrhundert hingegen sind medizinische Neuerungen im molekularbiologischen Bereich zu erwarten, wie beispielsweise in der Zellbiologie, DNA-Analyse und -Transfer oder in der regenerativen Medizin. In Letzterer werden autologe regenerative Mechanismen als therapeutisches Prinzip genutzt, um funktionsgestörte Zellen, Gewebe und Organe entweder durch den biologischen Ersatz oder durch die Anregung körpereigener Regenerations- und Reparaturprozesse zu erhalten bzw. wiederherzustellen.
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Immobilization of Beta-Glycosidase BglX from Escherichia coli on Chitosan Gel BeadsPickens, Tara L., L 30 August 2018 (has links)
No description available.
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Arsenic removal using biosorption with Chitosan : Evaluating the extraction and adsorption performance of Chitosan from shrimp shell wasteWestergren, Robin January 2006 (has links)
Nicaragua is a country in which the toxic metal contamination of freshwater resources has become an increasingly important problem in certain regions posing a threat to the environment as well as to human health. Among the metals found in the waters of Nicaragua, arsenic is one of the most problematic since its long time consumption is connected to serious health problems such as cancer and neurological disorders. The arsenic contamination of water recourses in Nicaragua is mostly attributable natural factors, even though anthropogenic activities including gold mining may be a contributing factor. In this work the biopolymer Chitosan was studied as a potential adsorption material for the removal of arsenic from aqueous solutions for water treatment design purposes. The Chitosan used in this study was extracted from shrimp shells with an overall yield of 40% and a deacetylation grade of 59%. The maximum adsorption capacity was determined to 20.9 mg As/g at a controlled pH of 5.5 using the Langmuir isotherm. The adsorption was found to be strongly pH dependant with a fourfold increase in adsorption capacity when pH was well under the pKa of Chitosan. The pH dependence indicates that ionic exchange was the most important mechanism. No difference in adsorption capacity with respect to the initial pH of the solution was detected in the pH range 3-7. This was attributed to the ability of Chitosan to act as a weak base in water solutions. The arsenic was desorbed from Chitosan using NaOH, (NH4) 2SO 4 and NaCl, with a 1M NaOH solution being the most efficient displaying a concentration ratio of 1.08. The NaOH and (NH4) 2SO 4 solutions displayed a steep desorption curvature with a large fraction of the arsenic being easily desorbed. The arsenic was, however, not completely desorbed from the Chitosan implying that the adsorption capacity would decrease for the coming cycles. Being a biopolymer the Chitosan is quite easily degraded in acid and alkali solutions, which might be a limiting step for the process applicability.
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Synthesis of In Situ Gelling Hydrogels Based on PolysaccharidesSträtz, Juliane 18 December 2023 (has links)
The synthesis of hydrogels still uses frequently starting materials based on petroleum. Since it is a non-renewable resource, the deposits will be exhausted sooner or later, besides the exploitation of fossil oil is environmentally harmful and contributes to climate change. For these reasons, alternative materials for hydrogels, which should be renewable and sustainable, have to be investigated. The focus of this thesis was to synthesize \textit{in situ} gelling hydrogels based on polysaccharides and to characterize their properties.
Cellulose and chitosan were chosen as starting materials, because they belong to the most abundant biopolymers on earth and therefore are readily available and also renewable. Additionally, cellulose sulfate (CS) is capable of attracting growth factors and chitosan has antimicrobial properties, all characteristics that are desirable for hydrogels in tissue engineering applications.
First of all, the sulfation of the cellulose was performed. Celluloses of two origins, sulfite pulp and cotton linters, were used to investigate the influence of the DP towards the DS_sulf. By applying direct sulfation and acetosulfation as sulfation methods a broader range of DS_sulf from 0.8 to 2.0 has been achieved. Values below 1.0 were obtained by acetosulfation for both celulloses without a remarkable influence of the DP. In direct sulfation, DS_sulf values of 1.1 to 2.0 could be achieved by adjusting the ratio of chlorosulfuric acid and anhydroglucose unit of cellulose between 3.0:1; 4.5:1 and 6.0:1. Starting from a ratio of 4.5:1 the effect of the DP is obvious, the longer chains of the cotton linters reach lower DS_sulf values compared to the sulfite pulp. Due to the longer cellulose chains the accessibility of the sulfating agent is limited, this is in particular noticeable at higher DS_sulf once the outer and easier accessible positions are sulfated.
The determination of the sulfur content, to quantify the DS_sulf, in CS became a key challenge in this thesis. At the beginning the sulfur content was determined by elemental analysis, because it is the common method in the literature. It is fast, does not require any pretreatment and only a low amount of sample is needed. However, it was found that the carbon and sulfur content in a sample did not interact logically. Theoretically, the relative carbon content should decrease and the relative sulfur content should rise with increasing DS_sulf, because pure cellulose does not contain sulfur. In practice, the percentage for carbon and sulfur increased both or the decrease of the carbon content was lower than expected for the increase in sulfur, so that the resulting DS_sulf was not the same when calculated with sulfur or carbon content. Hence additional direct methods of measuring the DS_sulf, which do not need a calibration sample, had to be investigated to validate the DS_sulf. ICP-OES and precipitation as BaSO4 were chosen as further methods and showed consistent results, but differed considerably from the findings of the elemental analysis. In contrast to elemental analysis, the other methods involve the digestion of the sample. So it could be possible that by using elemental analysis the reaction is incomplete and therefore the result is non-reproducible, however, it is not very likely. Although elemental analysis is faster and straightforward, it is recommended to use ICP-OES or precipitation as BaSO4 when determining the DS_sulf to receive reliable results.
The introduction of aldehyde groups in CS was necessary to provide reactive groups for the later hydrogel formation. In cellulose chemistry the widely known Malaprade reaction, an oxidation using sodium periodate, was performed. This oxidation stops after forming the aldehyde and does not further oxidize through to the carboxylic acid, additionally the reaction is possible in water which is essential for CS, since it is only soluble in water and it was aimed for a homogeneous reaction. A requirement for the oxidation is the presence of vicinal diols. The carbon-carbon bond is cleaved and an aldehyde is formed at each hydroxyl group. The maximum DS_ald of 2, which is possible for pure cellulose, cannot be reached, because the prior introduced sulfate groups reduce the number of vicinal OH groups. Although acetosulfation only took place at C6-position and all vicinal diols were still available, the DS_ald reached not more than 0.35. It is possible that steric hindrance through the sulfate group is the reason for these low values. Overall the DS_ald of the oxidized cellulose sulfate (oCS) ranged from 0.09 to 0.35; with increasing DS_sulf the DS_ald decreased.
The oCS are intended to be used in medical applications, accordingly their toxicity had to be investigated. Indeed, oCS are just one component for the hydrogel synthesis, but in case of an incomplete gelation the single components have to be non-toxic as well. In general, all oCS were non-toxic at low concentrations (0.5 mg/ml) yet an increasing concentration or a DS_ald of 0.3 and higher resulted in toxic effects. Aside from that, a coherence between M_w and toxicity was ascertained. The toxicity increased when the M_w of the oCS was 70 kDa or less.
The second component which is required for the hydrogel formation is a chitosan. The amino group of the chitosan can react with the aldehyde of the oCS by forming an imine. That way both biopolymers are crosslinked and result in a hydrogel. The used chitosan needs to be soluble under physiological conditions, consequently pure chitosan is not suitable since it is only water-soluble under acidic conditions. Hence, three chitosan derivates -- chitosan acetate, chitosan lactate and carboxymethyl chitosan (CMCh) -- were chosen which fulfill the criterion of solubility. To examine their aptitude for hydrogel formation 10 mg of each chitosan derivative and 10 mg of oCS were solved separately in 0,5 ml phosphat buffered saline. Afterwards, a chitosan derivative and oCS were mixed together while stirring until a reaction took place. In case of chitosan acetate and chitosan lactate a white sediment was the result, whereas the use of CMCh led to a colorless hydrogel, on this account all further studies were performed with CMCh.
To establish a basis for a targeted hydrogel synthesis, the storage modulus G' was investigated regarding selected parameters of the hydrogel: DS_ald, M_w, mixing ratio and time for gelation. The cross-linking was conducted with four mixing ratios of oCS:CMCh (1:1; 1:3; 1:5; 1:10) and correlated to the amount of substance of the aldehyde of the oCS and the corresponding amount of required CMCh. Since the CMCh solution has a high viscosity, the range of mixing ratios was limited. The total amount of substance was the same for all gels to ensure the comparability of the different hydrogels and the mixed volumes had always a 1:1 ratio to guarantee a fine blending of the components. Like presumed, G' increased with increasing DS_ald. The DS_ald is the specifying magnitude for the cross-linkage of the hydrogel, because the frequency of the aldehyde group is much lower compared to the frequency of the amine group of CMCh. Aldehyde and amine interact by forming an imine bond, the more of these bonds are formed the stiffer the resulting hydrogel becomes and as a result of that, G' increases as well. Furthermore, the storage modulus rises with rising M_w, the reason for it is that with increasing chain length the possibility of polymer entanglement increases. This physical type of cross-linkage makes for a stiffer gel too and therewith a higher G' is the consequence. With respect to the mixing ratio, maximum values for G' are attained if the ratio is shifted towards the component which defines the number of cross-linkages. During the study of the time for gelation it appeared that the time is not an independent parameter, in fact the time depends on the DS_ald. Samples with the shortest time for gelation showed the highest values for G', additionally they had the highest value for DS_ald. Thus, the time for gelation can not be considered when adjusting G'.
Further research on the hydrogels presented in this work needs to focus on investigating the toxicity and long-term behavior of the hydrogel, like stability and degradation, as well as its impact on tissue regeneration. The bioactivity and harmlessness of the hydrogel need to be ensured before it can be utilized in tissue engineering for example as cartilage tissue.
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Efeito da incorporação de quitosana e quitosana modificada na qualidade, funcionalidade tecnológica e vida de prateleira de salsichas com reduzido teor de gorduraAmaral, Deborah Silva do 11 August 2016 (has links)
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Previous issue date: 2016-08-11 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / The aim of this study was to evaluate the inclusion of chitosan and modified chitosan (chitosan-glucose) in appropriate concentration to meet the health claim in sausages with reduced fat, on the technological functionality and shelf life of these sausages. For this, an initial study was conducted, in which chitosan (2% w/w) was added to sausages prepared with pork and three levels of fat (5%, 12.5% and 20% w/w). The sausages were submitted to cold storage at 4 ± 1 °C for 15 days and evaluated in terms of microbiological, physic-chemical and sensory parameters. Subsequently, considering the absence of studies on incorporation of chitosan in meat goat products, carried out a second study, similar to the above to evaluate the effect of incorporation of chitosan (2% w/w) in sausage made with goat meat, and similar levels of fat in the microbiological and physico-chemical quality during cold storage (4 ± 1 °C) for 15 days. Finally, in the third study was evaluated the effect of the derivative of chitosan (2% w/w) in comparison with chitosan base (2% w/w) in sausage made with goat meat and reduced fat content (10% w/w). The sausages were stored at 4 ± 1 ° C for 21 days and evaluated by microbiological, physic-chemical and sensory parameters. In all three studies a formulation without the addition of polymers (chitosan and modified chitosan) was used as control. The results indicated that the use of 2% (w / w) chitosan and its derivative (corresponding to 1 g of chitosan per sausage of 50g each in a portion of 150g) to ensure ingestion of 3 g of chitosan per day is technologically feasible for use in the preparation of sausage with pork and goat meat. In addition to the functional potential, chitosan and chitosan derivative provided increased stability of the treated sausages since it caused the reduction of microbial growth when compared to the control sample. Additionally, they provided an improvement in red color, resulting in positive effects on the appearance, a more stable emulsion for the greater ability to bind water and fat, and a firmer texture to increase the compression forces, without negatively affecting the sensory properties of sausages. The chitosan and modified chitosan showed similar behavior, but the use of chitosan resulted in higher antioxidant activity than its derivative, considering further reduction of TBARS values and increased content of unsaturated fatty acids and poly-unsaturated. However, the chitosan derivative promoted a greater red color (a *) and lower hardness than chitosan. Therefore, the results of the use of chitosan in the manufacture of goat sausages, which was studied for the first time, indicated that the chitosan and modified chitosan showed potential interest to be used in the preparation of meat products, since they can provide higher quality and shelf life, adding value to the raw material of low commercial value, besides allow to obtain of a functional product with the health claim and fat reduction. / O objetivo deste estudo foi avaliar a inclusão da quitosana e quitosana modificada (quitosana-glicose), em concentração adequada para atender a alegação de saúde, em salsichas com reduzido teor de gordura, sobre a funcionalidade tecnológica e a vida de prateleira destas salsichas. Para isso, um primeiro estudo foi realizado, no qual a quitosana (2 % m/m) foi adicionada em salsichas elaboradas com carne suína e três níveis de gordura (5%, 12,5% e 20%, m/m). As salsichas foram submetidas ao armazenamento refrigerado a 4 ± 1 ºC por 15 dias e avaliadas em relação aos parâmetros microbiológicos, físico-químicos e sensoriais. Posteriormente, considerando a inexistência de estudos sobre a incorporação de quitosana em produtos cárneos caprino, realizou-se um segundo estudo, semelhante ao anterior, para avaliar o efeito da incorporação da quitosana (2 % m/m) em salsicha elaborada com carne caprina e similares níveis de gordura na qualidade microbiológica e físico-química durante o armazenamento refrigerado (4 ± 1 ºC) por 15 dias. Por fim, no terceiro estudo foi avaliado o efeito do derivado da quitosana (2% m/m), comparando com a quitosana base (2% m/m), em salsicha elaborada com carne caprina e reduzido teor de gordura (10% m/m). As salsichas foram armazenadas a 4 ± 1 ºC por 21 dias, e avaliadas nos parâmetros microbiológicos, físico-químicos e sensoriais. Nos três estudos uma formulação sem adição dos polímeros (quitosana e quitosana modificada) foi usada como controle. Os resultados indicaram que o uso de 2% (m / m) de quitosana e seu derivado (correspondendo a 1 g de quitosana por salsicha de 50g cada em uma porção de 150g) para garantir a ingestão de 3 g de quitosana por dia é tecnologicamente viável para uso na elaboração de salsicha com carne de porco e cabra. Além do potencial funcional, a quitosana e o derivado da quitosana proporcionaram aumento da estabilidade das salsichas tratadas, uma vez que provocaram a redução do crescimento microbiológico quando comparadas a amostra controle. Adicionalmente, proporcionaram uma melhoria na cor vermelha, resultando em efeitos positivos na aparência, uma emulsão mais estável, pela maior capacidade de ligar água e gordura, e uma textura mais firme por aumentar as forças de compressão, sem afetar negativamente as propriedades sensoriais das salsichas. A quitosana e a quitosana modificada mostraram comportamento semelhante, mas o uso da quitosana resultou em maior atividade antioxidante do que o seu derivado, considerando a maior redução dos valores de TBARS e maior teor de ácidos graxos insaturados e poli-insaturados. Entretanto, o derivado da quitosana promoveu maior cor vermelha (a*) e menor dureza que a quitosana. Portanto, os resultados da utilização da quitosana na fabricação de salsichas caprina, a qual foi estudada pela primeira vez, indicaram que a quitosana e a quitosana modificada apresentaram potencial interessante para serem utilizados na elaboração de produtos cárneos, uma vez que podem proporcionar maior qualidade e vida de prateleira, agregando valor à matéria-prima de baixo valor comercial, além de permitir a obtenção de um produto funcional com alegação de saúde e redução de gordura.
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Le 17B-Estradiol combiné à un biopolymère à base de chitosan accroît la biocompatibilité des cellules progénitrices dérivées de la moelle osseuseTardif, Kim 07 1900 (has links)
Les cellules dérivées de la moelle osseuse, principalement les cellules endothéliales progénitrices, sont réduites chez les patients souffrant de maladies cardiovasculaires. Leur mobilisation et leur incorporation aux sites de lésion vasculaire sont des évènements prépondérants dans l’accélération des processus de réendothélialisation. Dans un modèle murin, le 17β-estradiol favorise les processus de guérison vasculaire par la mobilisation et le recrutement des cellules endothéliales progénitrices dérivées de la moelle osseuse. Il existe présentement plusieurs stratégies afin d’augmenter la mobilisation des cellules progénitrices ainsi que leur incorporation à la paroi vasculaire. Cependant, peu d’études privilégient la livraison locale d’un nombre élevé de cellules progénitrices fonctionnelles par un véhicule biodégradable et leur maintien au site de lésion afin de favoriser la réendothélialisation ciblée. Un polymère d’intérêt pour cette application s’avère être le chitosan. Ce biopolymère non toxique et biodégradable est couramment utilisé dans l’ingénierie tissulaire et, depuis peu, est utilisé dans la guérison vasculaire. Le chitosan complexé à la phosphorylcholine voit sa solubilité s’accroître dans les solutions aqueuses ainsi que sa biocompatibilité cellulaire en condition physiologique.
Le projet de ce mémoire visait donc : 1) à étudier in vitro, la capacité d’un polymère de chitosan complexé à la phosphorylcholine à influencer l’adhésion, la survie, la différenciation et la fonctionnalité cellulaire dans un modèle murin de culture mixte de cellules dérivées de la moelle osseuse et 2) de déterminer l’impact de la présence du 17β-estradiol sur ces mêmes comportements cellulaires.
Nos travaux démontrent que la matrice de chitosan-phosphorylcholine s’avère compatible avec notre modèle de culture cellulaire. En effet, ce polymère est capable de promouvoir l’organisation et le développement des cellules dérivées de la moelle osseuse de façon comparable à la matrice normalement utilisée dans la croissance in vitro des cellules endothéliales progénitrices, la fibronectine. De plus, ce polymère n’a nullement compromis l’activité migratoire des cellules, laissant supposer qu’il pourrait éventuellement être un véhicule approprié pour effectuer une livraison cellulaire à un site de lésion.
Il s’avère que le 17β-estradiol, lorsqu’ajouté au milieu de culture ou complexé au polymère de chitosan phosphorylcholine, est capable de moduler le comportement cellulaire, et ce, de façon différente. Le 17β-estradiol complexé au polymère de chitosan-phosphorylcholine démontre, par rapport à sa forme soluble, une plus grande aptitude à accroître le nombre de cellules hématopoïétiques ainsi que des cellules endothéliales progénitrices dérivées de la moelle osseuse in vitro. De plus, le 17β-estradiol complexé au polymère de chitosan-phosphorylcholine permet une amplification marquée des cellules endothéliales progénitrices et leur offre un support adéquat afin de favoriser la guérison vasculaire.
L’ensemble de nos travaux suggère que le polymère de chitosan complexé à la phosphorylcholine en présence ou non de 17β-estradiol est une matrice compatible avec les cellules progénitrices dérivées de la moelle osseuse in vitro. Le 17β-estradiol complexé au polymère est toutefois plus efficace que sa forme soluble à promouvoir l’amplification du nombre de cellules progénitrices. Ce polymère représente un outil thérapeutique attrayant et une matrice de livraison d’agent bioactif prometteuse pour le recrutement cellulaire dans l’accélération de la guérison vasculaire. / Bone marrow derived cells, including endothelial progenitor cells, are reduced in numbers in patient with cardiovascular disease or risk factors. Mobilization and incorporation of these cells at the vascular lesion site are important events in the reendothelialization process. 17β-estradiol was shown in a mouse model of injury, to favour this healing process through mechanisms which involve the mobilization and incorporation of endothelial progenitor cells derived from the bone marrow. At the moment, there are many strategies to increase endothelial progenitor cells mobilization as well as recruitment into the vascular wall. However, few studies favour local delivery of a large number of functional progenitor cells on a biodegradable scaffold and to maintain them at the lesion site in order to promote reendothelialization. An interesting biopolymer for this application is chitosan. This non toxic and biodegradable biopolymer is commonly used in tissue engineering and was recently used in vascular healing. Phosphorylcholine modified chitosan can increase the water solubility and cell biocompatibility of the biopolymer in physiological condition.
This master project was thus designed to :1) evaluate, in vitro, the capacity of phosphorylcholine modified chitosan to influence cell adhesion, survival, differentiation and functionality in a mouse model of bone marrow mixed culture and 2) determine the impact of 17β-estradiol on these cell behaviours.
Our results suggest an adequate biocompatibility of phosphorylcholine modified chitosan with our cell culture system. Indeed, this polymer was able to promote cell organization and development of bone marrow derived cells in the same way that fibronectin, the most commonly matrix used in the progenitor cells in vitro culture. Moreover, cell migratory activity was not compromised by the chitosan polymer.
It appears that 17β-estradiol, when added to cell culture media or attached on phosphorylcholine modified chitosan is able to modulate differently cell behaviour. Our data suggest that 17β-estradiol coupled to the chitosan polymer was superior to increase the number of haematopoietic and endothelial progenitor cells derived from bone marrow in vitro compared to the soluble form. 17β-estradiol coupled to the polymer of phosphorylcholine modified chitosan allowed an increased amplification of progenitor cell number and provided adequate scaffold to favour vascular healing.
We propose that phosphorylcholine modified chitosan in presence or not of 17β-estradiol is a compatible matrix with bone marrow derived progenitor cells in vitro. 17β-estradiol enhances the amplification of progenitor cell in vitro when associated to the polymer compared to its soluble form. This biopolymer may be an attractive matrix and a promising vehicle in a drug delivery therapeutic system for progenitor cells recruitment and to promote vascular healing.
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Enhancement of the intestinal epithelial permeability of peripherally acting opioid analgesics by chitosanRubelt, Miriam 13 December 2013 (has links)
Die schmerzstillende Wirkung von Opiaten wird über Opioidrezeptoren im zentralen und peripheren Nervensystem vermittelt. Die Schmerzlinderung kann jedoch mit sehr starken Nebenwirkungen einhergehen, die das Patientenwohlbefinden beeinträchtigen. Dies legt die Bedeutung von neuen Opioidanalgetika nahe, die ihre schmerzstillende Wirkung ausschließlich über Opioidrezeptoren im PNS entfalten, ohne unerwünschte zentrale Nebenwirkungen zu induzieren. Die orale Gabe von Medikamenten minimiert Unannehmlichkeiten für den Patienten, jedoch müssen die Substanzen die intestinale Barriere passieren können, um in die Blutzirkulation eintreten zu können. Die intestinale Permeabilität von zwei peripher wirksamen Opiaten (AS006 und Loperamid) wurde in Ussing-Kammer Experimenten untersucht. Um die Darmepithelpermeabilität für beide Opiate zu erhöhen, wurde der Absorptionsverstärker Chitosan verwendet. Chitosan bewirkte nach 30 Minuten bei HT29/B6 und Caco-2 Zelllinien eine Abnahme des epithelialen Widerstands in vitro. Die Permeabilität für AS006 war bei beiden Zelllinien erhöht, für Loperamid nur bei HT29/B6, jedoch nicht bei Caco-2 Zellmonolayern. Verhaltensexperimente zur Messung des antinozizeptiven Effektes von oral appliziertem Loperamid auf Entzündungsschmerz wurden an Ratten durchgeführt. Die orale Gabe von Loperamid induzierte eine Dosis-abhängige antinozizeptive Wirkung in der entzündeten Hinterpfote. Bei oraler Gabe von Loperamid in Kombination mit Chitosan wurde keine signifikante Verstärkung des maximalen antinozizeptiven Effekts von Loperamid beobachtet. Zusammenfassend ist Chitosan ein geeigneter Absorptionsverstärker für intestinale Permeabilitätsstudien von peripher wirksamen Opioidanalgetika in vitro. Die in vitro Ergebnisse haben gezeigt, dass der Effekt von Chitosan auf Loperamid möglicherweise schwächer ist als auf AS006. Dementsprechend fiel die Wirkung des Absorptionsverstärkers auf Loperamid-induzierte Analgesie im Verhaltensversuch eher gering aus. / Analgesic effects of opioids are mediated by opioid receptors that are widely distributed in the central and peripheral nervous systems (CNS and PNS, respectively). Although opioids are the most powerful analgesics, severe side effects restrict their use and affect patient convalescence. This suggests an advantage of new analgesic opioids which selectively bind to opioid receptors in the PNS. After oral administration however, peripherally restricted opioids first have to cross the intestinal epithelial barrier before absorption into the circulation and distribution to opioid receptors in peripheral tissues. Here, the transport across intestinal epithelia of two opioid ligands (AS006 and loperamide) that selectively activate peripheral opioid receptors without entering the CNS were investigated. To increase the intestinal passage of these drugs, the absorption enhancer chitosan was used. Chitosan significantly decreased the transepithelial resistance of HT29/B6 and Caco-2 cell monolayers after 30 min in vitro. The permeability values for AS006 increased from < 0.3 × 10-6 cm/s up to 10 × 10-6 cm/s in the presence of chitosan. In contrast, HT29/B6 monolayers showed moderate loperamide permeability in the presence of chitosan, and chitosan had no effect on the permeability of loperamide using Caco-2 monolayers. Oral administration of loperamide induced a dose-depended elevation of paw pressure thresholds in inflamed paws that lasted for 60 min. Oral administration of loperamide combined with chitosan slightly but nonsignificantly enhanced the antinociceptive effect of loperamide. In conclusion, chitosan is a suitable absorption enhancer for in vitro intestinal permeability studies. Future in vivo experiments might investigate different formulations and application schedules, and further address the effects of chitosan on the antinociceptive efficacy of hydrophilic opioids.
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Bionanocompósitos de derivados de quitosana/montmorilonita/nanopartículas de prata preparadas via fotoquímica / Bionanocomposites of chitosan derivatives/montmorillonite/silver nanoparticles prepared by PhotochemistryGabriel, Juliana dos Santos 28 July 2017 (has links)
O presente trabalho apresenta a síntese e a caracterização de derivados de quitosana, bem como o preparo e caracterização de filmes de nanocompósitos à base de quitosana comercial (ou seus derivados), argila (MMT) e nanopartículas de prata (NPs-Ag), obtidas via Fotoquímica. Para tanto, foram preparados, a partir da quitosana comercial (QC), os derivados: quitosana desacetilada (Q30des), quitosana purificada (QP), quitosanas parcialmente despolimerizas (QD30, QD21 e QD5), quitosanas hidrofílicas (QD21-40DEAE e QD5-49DEAE) e quitosanas anfifílicas (QD21-40DEAE-6DD, QD21-40DEAE-18DD, QD5-49DEAE-6DD e QD5-49DEAE-17DD). O grau médio de desacetilação das QC, QP e Q30des e de substituição por grupos DEAE e dodecila foram determinados por Espectroscopia de Ressonância Magnética Nuclear de Hidrogênio (RMN de 1H). Ademais, os biopolímeros foram caracterizados por Espectroscopia no Infravermelho (FTIR-ATR), Viscosimetria, Análise Termogravimétrica e Microscopia Eletrônica de Varredura (MEV). Em seguida, foi estudada a síntese de nanopartículas de prata, sob radiação UV, em filmes de nanocompósitos de quitosana comercial ou seus derivados e argila. Em um primeiro momento, estudou-se a formação das NPs-Ag em filmes de QC com diferentes formulações e posteriormente em filmes de derivados de quitosana contendo 10% de MMT (m/m). A técnica de Difração de Raios-X (DRX) foi utilizada para a determinação do espaçamento interlamelar da argila montmorilonita pura e nos compósitos preparados. A síntese das NPs-Ag foi acompanhada por Espectrofotometria de Absorção Molecular no UV-vis, e monitorada após um ano de sua formação, sendo suas características morfológicas, bem como a dispersão da argila nos nanocompósitos examinados por Microscopia Eletrônica de Transmissão (MET). Por fim, a atividade antimicrobiana dos filmes de nanocompósitos foi avaliada pelo método de Disco de Difusão contra as bactérias Escherichia coli e Bacillus subtilis. / The present work presents the synthesis and characterization of chitosan derivatives, as well as the preparation and characterization of nanocomposite films based on commercial chitosan (or its derivatives), clay (MMT) and silver nanoparticles (NPs-Ag) obtained by photochemical method. Therefore, were prepared from commercial chitosan (QC): deacetylated chitosan (Q30des); purified chitosan (QP); partially depolymerized chitosans (QD30, QD21 and QD5); hydrophilic chitosans (QD21-40DEAE and QD5-QD5) and amphiphilic chitosans (QD21-40DEAE-6DD, QD21-40DEAE-18DD, QD5-49DEAE-6DD and QD5-49DEAE-17DD). The deacetylation degrees of QC, QP and Q30des were determined by Nuclear Magnetic Resonance Spectroscopy (1H-NMR). This technique also used to determine the degrees of substitution by DEAE and dodecyl groups. In addition, the biopolymers were characterized by Infrared Spectroscopy (FTIR-ATR), Viscosimetry, Thermogravimetry and Scanning Electron Microscopy. Moreover, the NPs-Ag synthesis under UV radiation was studied on nanocomposite films of commercial chitosan or its derivatives and clay. At first, the Ag-NPs formation was studied on QC films with different formulations and secondarily, on films of chitosan derivatives containing 10 wt % of MMT. The X-Ray Diffraction (XRD) was used to determine the interlamellar spacing of pure montmorillonite clay and in the nanocomposites prepared. The synthesis of the NP-Ag was accompanied by UV-vis Spectroscopy. Its morphological characteristics, as well as the clay dispersion in the nanocomposites were examined by Electron Transmission Electron Microscopy (TEM). Finally, the antimicrobial activities of materials were investigated by the disk diffusion method against the bacteria Escherichia coli e Bacillus subtilis.
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Bionanocompósitos de derivados de quitosana/montmorilonita/nanopartículas de prata preparadas via fotoquímica / Bionanocomposites of chitosan derivatives/montmorillonite/silver nanoparticles prepared by PhotochemistryJuliana dos Santos Gabriel 28 July 2017 (has links)
O presente trabalho apresenta a síntese e a caracterização de derivados de quitosana, bem como o preparo e caracterização de filmes de nanocompósitos à base de quitosana comercial (ou seus derivados), argila (MMT) e nanopartículas de prata (NPs-Ag), obtidas via Fotoquímica. Para tanto, foram preparados, a partir da quitosana comercial (QC), os derivados: quitosana desacetilada (Q30des), quitosana purificada (QP), quitosanas parcialmente despolimerizas (QD30, QD21 e QD5), quitosanas hidrofílicas (QD21-40DEAE e QD5-49DEAE) e quitosanas anfifílicas (QD21-40DEAE-6DD, QD21-40DEAE-18DD, QD5-49DEAE-6DD e QD5-49DEAE-17DD). O grau médio de desacetilação das QC, QP e Q30des e de substituição por grupos DEAE e dodecila foram determinados por Espectroscopia de Ressonância Magnética Nuclear de Hidrogênio (RMN de 1H). Ademais, os biopolímeros foram caracterizados por Espectroscopia no Infravermelho (FTIR-ATR), Viscosimetria, Análise Termogravimétrica e Microscopia Eletrônica de Varredura (MEV). Em seguida, foi estudada a síntese de nanopartículas de prata, sob radiação UV, em filmes de nanocompósitos de quitosana comercial ou seus derivados e argila. Em um primeiro momento, estudou-se a formação das NPs-Ag em filmes de QC com diferentes formulações e posteriormente em filmes de derivados de quitosana contendo 10% de MMT (m/m). A técnica de Difração de Raios-X (DRX) foi utilizada para a determinação do espaçamento interlamelar da argila montmorilonita pura e nos compósitos preparados. A síntese das NPs-Ag foi acompanhada por Espectrofotometria de Absorção Molecular no UV-vis, e monitorada após um ano de sua formação, sendo suas características morfológicas, bem como a dispersão da argila nos nanocompósitos examinados por Microscopia Eletrônica de Transmissão (MET). Por fim, a atividade antimicrobiana dos filmes de nanocompósitos foi avaliada pelo método de Disco de Difusão contra as bactérias Escherichia coli e Bacillus subtilis. / The present work presents the synthesis and characterization of chitosan derivatives, as well as the preparation and characterization of nanocomposite films based on commercial chitosan (or its derivatives), clay (MMT) and silver nanoparticles (NPs-Ag) obtained by photochemical method. Therefore, were prepared from commercial chitosan (QC): deacetylated chitosan (Q30des); purified chitosan (QP); partially depolymerized chitosans (QD30, QD21 and QD5); hydrophilic chitosans (QD21-40DEAE and QD5-QD5) and amphiphilic chitosans (QD21-40DEAE-6DD, QD21-40DEAE-18DD, QD5-49DEAE-6DD and QD5-49DEAE-17DD). The deacetylation degrees of QC, QP and Q30des were determined by Nuclear Magnetic Resonance Spectroscopy (1H-NMR). This technique also used to determine the degrees of substitution by DEAE and dodecyl groups. In addition, the biopolymers were characterized by Infrared Spectroscopy (FTIR-ATR), Viscosimetry, Thermogravimetry and Scanning Electron Microscopy. Moreover, the NPs-Ag synthesis under UV radiation was studied on nanocomposite films of commercial chitosan or its derivatives and clay. At first, the Ag-NPs formation was studied on QC films with different formulations and secondarily, on films of chitosan derivatives containing 10 wt % of MMT. The X-Ray Diffraction (XRD) was used to determine the interlamellar spacing of pure montmorillonite clay and in the nanocomposites prepared. The synthesis of the NP-Ag was accompanied by UV-vis Spectroscopy. Its morphological characteristics, as well as the clay dispersion in the nanocomposites were examined by Electron Transmission Electron Microscopy (TEM). Finally, the antimicrobial activities of materials were investigated by the disk diffusion method against the bacteria Escherichia coli e Bacillus subtilis.
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Le 17B-Estradiol combiné à un biopolymère à base de chitosan accroît la biocompatibilité des cellules progénitrices dérivées de la moelle osseuseTardif, Kim 07 1900 (has links)
Les cellules dérivées de la moelle osseuse, principalement les cellules endothéliales progénitrices, sont réduites chez les patients souffrant de maladies cardiovasculaires. Leur mobilisation et leur incorporation aux sites de lésion vasculaire sont des évènements prépondérants dans l’accélération des processus de réendothélialisation. Dans un modèle murin, le 17β-estradiol favorise les processus de guérison vasculaire par la mobilisation et le recrutement des cellules endothéliales progénitrices dérivées de la moelle osseuse. Il existe présentement plusieurs stratégies afin d’augmenter la mobilisation des cellules progénitrices ainsi que leur incorporation à la paroi vasculaire. Cependant, peu d’études privilégient la livraison locale d’un nombre élevé de cellules progénitrices fonctionnelles par un véhicule biodégradable et leur maintien au site de lésion afin de favoriser la réendothélialisation ciblée. Un polymère d’intérêt pour cette application s’avère être le chitosan. Ce biopolymère non toxique et biodégradable est couramment utilisé dans l’ingénierie tissulaire et, depuis peu, est utilisé dans la guérison vasculaire. Le chitosan complexé à la phosphorylcholine voit sa solubilité s’accroître dans les solutions aqueuses ainsi que sa biocompatibilité cellulaire en condition physiologique.
Le projet de ce mémoire visait donc : 1) à étudier in vitro, la capacité d’un polymère de chitosan complexé à la phosphorylcholine à influencer l’adhésion, la survie, la différenciation et la fonctionnalité cellulaire dans un modèle murin de culture mixte de cellules dérivées de la moelle osseuse et 2) de déterminer l’impact de la présence du 17β-estradiol sur ces mêmes comportements cellulaires.
Nos travaux démontrent que la matrice de chitosan-phosphorylcholine s’avère compatible avec notre modèle de culture cellulaire. En effet, ce polymère est capable de promouvoir l’organisation et le développement des cellules dérivées de la moelle osseuse de façon comparable à la matrice normalement utilisée dans la croissance in vitro des cellules endothéliales progénitrices, la fibronectine. De plus, ce polymère n’a nullement compromis l’activité migratoire des cellules, laissant supposer qu’il pourrait éventuellement être un véhicule approprié pour effectuer une livraison cellulaire à un site de lésion.
Il s’avère que le 17β-estradiol, lorsqu’ajouté au milieu de culture ou complexé au polymère de chitosan phosphorylcholine, est capable de moduler le comportement cellulaire, et ce, de façon différente. Le 17β-estradiol complexé au polymère de chitosan-phosphorylcholine démontre, par rapport à sa forme soluble, une plus grande aptitude à accroître le nombre de cellules hématopoïétiques ainsi que des cellules endothéliales progénitrices dérivées de la moelle osseuse in vitro. De plus, le 17β-estradiol complexé au polymère de chitosan-phosphorylcholine permet une amplification marquée des cellules endothéliales progénitrices et leur offre un support adéquat afin de favoriser la guérison vasculaire.
L’ensemble de nos travaux suggère que le polymère de chitosan complexé à la phosphorylcholine en présence ou non de 17β-estradiol est une matrice compatible avec les cellules progénitrices dérivées de la moelle osseuse in vitro. Le 17β-estradiol complexé au polymère est toutefois plus efficace que sa forme soluble à promouvoir l’amplification du nombre de cellules progénitrices. Ce polymère représente un outil thérapeutique attrayant et une matrice de livraison d’agent bioactif prometteuse pour le recrutement cellulaire dans l’accélération de la guérison vasculaire. / Bone marrow derived cells, including endothelial progenitor cells, are reduced in numbers in patient with cardiovascular disease or risk factors. Mobilization and incorporation of these cells at the vascular lesion site are important events in the reendothelialization process. 17β-estradiol was shown in a mouse model of injury, to favour this healing process through mechanisms which involve the mobilization and incorporation of endothelial progenitor cells derived from the bone marrow. At the moment, there are many strategies to increase endothelial progenitor cells mobilization as well as recruitment into the vascular wall. However, few studies favour local delivery of a large number of functional progenitor cells on a biodegradable scaffold and to maintain them at the lesion site in order to promote reendothelialization. An interesting biopolymer for this application is chitosan. This non toxic and biodegradable biopolymer is commonly used in tissue engineering and was recently used in vascular healing. Phosphorylcholine modified chitosan can increase the water solubility and cell biocompatibility of the biopolymer in physiological condition.
This master project was thus designed to :1) evaluate, in vitro, the capacity of phosphorylcholine modified chitosan to influence cell adhesion, survival, differentiation and functionality in a mouse model of bone marrow mixed culture and 2) determine the impact of 17β-estradiol on these cell behaviours.
Our results suggest an adequate biocompatibility of phosphorylcholine modified chitosan with our cell culture system. Indeed, this polymer was able to promote cell organization and development of bone marrow derived cells in the same way that fibronectin, the most commonly matrix used in the progenitor cells in vitro culture. Moreover, cell migratory activity was not compromised by the chitosan polymer.
It appears that 17β-estradiol, when added to cell culture media or attached on phosphorylcholine modified chitosan is able to modulate differently cell behaviour. Our data suggest that 17β-estradiol coupled to the chitosan polymer was superior to increase the number of haematopoietic and endothelial progenitor cells derived from bone marrow in vitro compared to the soluble form. 17β-estradiol coupled to the polymer of phosphorylcholine modified chitosan allowed an increased amplification of progenitor cell number and provided adequate scaffold to favour vascular healing.
We propose that phosphorylcholine modified chitosan in presence or not of 17β-estradiol is a compatible matrix with bone marrow derived progenitor cells in vitro. 17β-estradiol enhances the amplification of progenitor cell in vitro when associated to the polymer compared to its soluble form. This biopolymer may be an attractive matrix and a promising vehicle in a drug delivery therapeutic system for progenitor cells recruitment and to promote vascular healing.
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