Resistência mecânica de hidrogéis termo-sensíveis constituídos de Alginato-Ca2+ / PNIPAAm, tipo Semi-IPN

Thermosensitive hydrogels were synthesized using alginate-Ca2+ in association with a thermosensitive polymer, such as PNIPAAm. The mechanical properties of the hydrogels were determined measuring the maximum tension of deformation. With the increase of the temperature by 25 to 40 ºC above the LCST the chains of PNIPAAm collapsed, dragging the alginate net and diminishing the size of the pores. The decrease in the size of the pores of the hydrogel was followed by an increase in the mechanicals resistance of the material.

Resistência mecânica de hidrogéis termo-sensíveis constituídos de Alginato-Ca2+ / PNIPAAm, tipo Semi-IPN

Thermosensitive hydrogels were synthesized using alginate-Ca2+ in association with a thermosensitive polymer, such as PNIPAAm. The mechanical properties of the hydrogels were determined measuring the maximum tension of deformation. With the increase of the temperature by 25 to 40 ºC above the LCST the chains of PNIPAAm collapsed, dragging the alginate net and diminishing the size of the pores. The decrease in the size of the pores of the hydrogel was followed by an increase in the mechanicals resistance of the material.

Multipoint covalent immobilization of lipase on chitosan hybrid hydrogels: influence of the polyelectrolyte complex type and chemical modification on the catalytic properties of the biocatalysts

This work aimed at the production of stabilized derivatives of Thermomyces lanuginosus lipase (TLL) by multipoint covalent immobilization of the enzyme on chitosan-based matrices. The resulting biocatalysts were tested for synthesis of biodiesel by ethanolysis of palm oil. Different hydrogels were prepared: chitosan alone and in polyelectrolyte complexes (PEC) with kappa-carrageenan, gelatin, alginate, and polyvinyl alcohol (PVA). The obtained supports were chemically modified with 2,4,6-trinitrobenzene sulfonic acid (TNBS) to increase support hydrophobicity, followed by activation with different agents such as glycidol (GLY), epichlorohydrin (EPI), and glutaraldehyde (GLU). The chitosan-alginate hydrogel, chemically modified with TNBS, provided derivatives with higher apparent hydrolytic activity (HA(app)) and thermal stability, being up to 45-fold more stable than soluble lipase. The maximum load of immobilized enzyme was 17.5 mg g(-1) of gel for GLU, 7.76 mg g(-1) of gel for GLY, and 7.65 mg g(-1) of gel for EPI derivatives, the latter presenting the maximum apparent hydrolytic activity (364.8 IU g(-1) of gel). The three derivatives catalyzed conversion of palm oil to biodiesel, but chitosan-alginate-TNBS activated via GLY and EPI led to higher recovered activities of the enzyme. Thus, this is a more attractive option for both hydrolysis and transesterification of vegetable oils using immobilized TLL, although industrial application of this biocatalyst still demands further improvements in its half-life to make the enzymatic process economically attractive.

Nitric oxide photorelease from hydrogels and from skin containing a nitro-ruthenium complex

Nitric oxide (NO) is a gaseous molecule that has specific functions dictated by its localization and its kinetics of release. As NO-donors have a range of potential uses in the skin, much attention has been paid to the development of topical NO delivery systems. The aim of this work was to study the release rate and the skin penetration of the NO-donor cis[Ru(NO(2))(bpy)(2)(4-pic)](+) from different gel formulations and their potential as topical NO delivery systems under light stimuli. Among the formulations developed, the anionic gel retarded the nitro-ruthenium complex diffusion and also obstructed NO release after light irradiation. On the other hand, NO release before light irradiation was observed when the complex was dispersed in the cationic chitosan gel, possibly due to oxi-redox reactions between the amino groups of the polymer and the drug molecule. Finally, the non-ionic gel released the NO after light irradiation to the same extent as a drug aqueous solution at the same pH. The drug dispersed in this gel also penetrated into the stratum corneum skin layer, and the nitro-ruthenium complex present in the skin was able to release the NO after light stimuli, suggesting the potential use of this formulation as a topical NO delivery system. (C) 2010 Elsevier B.V. All rights reserved.

Water diffusion in hydroxyethyl methacrylate (HEMA)-based hydrogels formed by gamma-radiolysis

Polymer hydrogels based upon methacrylates are used extensively in the pharmaceutical industry, particularly as controlled release drug delivery systems. These materials are generally prepared by chemically initiated polymerization, but this can lead to the presence of unwanted initiator fragments in the polymer matrix. In the present work, initiation of polymerization by gamma-irradiation of hydroxyethyl methacrylate, with and without added crosslinkers, has been investigated, and the diffusion coefficients for water in the resulting polymers have been measured through mass uptake by the polymers. The diffusion of water in poly(hydroxyethyl methacrylate) at 310 K was found to be Fickian, with a diffusion coefficient of 1.96 +/- 0.1 x 10(11) m(2) s(-1) and an equilibrium water content of 58%, NMR imaging analyses confirmed the adherance to a Fickian model of the diffusion of water into polymer cylinders. The incorporation of small amounts (0.2-0.5 wt%) of added ethyleneglycol-dimethacrylate-based crosslinkers was found to have only a small effect on the diffusion coefficient and the equilibrium water content for the copolymers. (C) 1999 Society of Chemical Industry.

Copolymer hydrogels of 2-hydroxyethyl methacrylate with n-butyl methacrylate and cyclohexyl methacrylate: synthesis, characterization and uptake of water

The bulk free radical copolymerizations of 2-hydroxyethyl methacrylate (HEMA) with n-butyl methacrylate (BMA) or cyclohexyl methacrylate (CHMA) were studied over the composition mole fraction interval of 0-1 for HEMA in the monomer feed. The C-13 NMR (125 MHz) spectra of the copolymers were analysed to determine the copolymer composition and the stereochemical configuration of the copolymers. The terminal model reactivity ratios of HEMA and BMA were found to be r(HEMA) = 1.73 and r(BMA) = 0.65 and for HEMA and CHMA, r(HEMA) = 1.26 and r(CHMA) = 0.31. The BMA and CHMA homopolymers were found to be predominantly syndiotactic with isotacticity parameters of theta(BB) = 0.18 and theta(CC) = 0.19, respectively. The copolymers were also found to be predominantly syndiotactic, indicating a strong tendency for racemic additions of the monomers in the formation of the copolymers. The diffusion of water into cylinders of poly(HEMA-co-BMA) and poly(HEMA-co-CHMA) was studied over a range of copolymer compositions and was found to be Fickian. The diffusion coefficients of water at 37 degrees C were determined from swelling measurements and were found to vary from 1.72 x 10(-11) m(2) s(-1) for polyHEMA to 0.97 x 10(-11) m(2) s(-1) for poly(HEMA-co-BMA) having a mole fraction F-HEMA = 0.80 and to 0.91 x 10(-11) m(2) s(-1) for a poly(HEMA-co-CHMA) also having F-HEMA = 0.80. The mass of water absorbed at equilibrium relative to the mass of dry polymer varied from 58.8 for polyHEMA to 27.2% for poly(HEMA-co-BMA) having F-HEMA = 0.85 and to 21.3% for poly(HEMA-co-CHMA) having F-HEMA = 0.80. (C) 1999 Elsevier Science Ltd. All rights reserved.

The states, diffusion, and concentration distribution of water in radiation-formed PVA/PVP hydrogels

Hydrogels with various compositions of polyvinyl alcohol (PVA) and poly(1-vinyl-2-pyrrolidinone) (PVP) were prepared by irradiating mixtures of PVA and PVP in aqueous solutions with gamma-rays from Co-60 sources at room temperature. The states of water in the hydrogels were characterized using DSC and NMR T-2 relaxation measurements and the kinetics of water diffusion in the hydrogels were studied by sorption experiments and NMR imaging. The DSC endothermic peaks in the temperature range -10 to +10 degrees C implied that there are at least two kinds of freezable water present in the matrix. The difference between the total water content and the freezable water content was refer-red to as bound water, which is not freezable. The weight fraction of water at which only nonfreezable water is present in a hydrogel with F-VP = 0.19 has been estimated to be g(H2O)/g(Polymer) = 0.375. From water sorption experiments, it was demonstrated that the early stage of the diffusion of water into the hydrogels was Fickian. A curve-fit of the early-stage experimental data to the Fickian model allowed determination of the water diffusion coefficient, which was found to lie between 1.5 x 10(-11) m(2) s(-1) and 4.5 x 10(-11) m(2) s(-1), depending on the polymer composition, the cross-link density, and the temperature. It was also found that the energy barrier for diffusion of water molecules into PVA/PVP hydrogels was approximate to 24 kJ mol(-1). Additionally, the diffusion coefficients determined from NMR imaging of the volumetric swelling of the gels agreed well with the results obtained by the mass sorption method.

H-1 NMR study of the states of water in equilibrium Poly(HEMA-co-THFMA) hydrogels

The spin-spin relaxation times, T-2, of hydrated samples of poly(hydroxymethyl methacrylate), PHEMA, poly(tetrahydrofurfuryl methacrylate),PTHFMA, and the,corresponding HEMA-THFMA copolymers have been examined to probe the states of,the imbibed water in these polymers. The decay in the transverse magnetization of water. in fully hydrated samples of PHEMA, PTHFMA, and copolymers of HEMA and THFMA was described by a multiexponential function. The short component of T-2 was interpreted as water molecules that were strongly interacting with the polymer chains. The intermediate component of T-2 was assigned to water residing in the porous structure of the samples. The long component of T-2 was believed to arise from water residing in the remnants of cracks formed in the polymer network during water sorption.

PFG-NMR measurements of the self-diffusion coefficients of water in equilibrium poly(HEMA-co-THFMA) hydrogels

The self-diffusion coefficients for water in a series of copolymers of 2-hydroxyethyl methacrylate, HEMA, and tetrahydrofurfuryl methacrylate, THFMA, swollen with water to their equilibrium states have been studied at 310 K using PFG-NMR. The self-diffusion coefficients calculated from the Stejskal-Tanner equation, D-obs, for all of the hydrated polymers were found to be dependent on the NMR storage time, as a result of spin exchange between the proton reservoirs of the water and the polymers, reaching an equilibrium plateau value at long storage times. The true values of the diffusion coefficients were calculated from the values of D-obs, in the plateau regions by applying a correction for the fraction of water protons present, obtained from the equilibrium water contents of the gels. The true self-diffusion coefficient for water in polyHEMA obtained at 310 K by this method was 5.5 x 10(-10) m(2) s(-1). For the copolymers containing 20% HEMA or more a single value of the self-diffusion coefficient was found, which was somewhat larger than the corresponding values obtained for the macroscopic diffusion coefficient from sorption measurements. For polyTHFMA and copolymers containing less than 20% HEMA, the PFG-NMR stimulated echo attenuation decay curves and the log-attenuation plots were characteristic of the presence of two diffusing water species. The self-diffusion coefficients of water in the equilibrium-hydrated copolymers were found to be dependent on the copolymer composition, decreasing with increasing THFMA content.

Development of 2-oxazoline-based hydrogels and porous polyester microparticles in scCO2

Thesis submitted to Faculdade de Ciências e Tecnologia from Universidade Nova de Lisboa in partial fulfillment of the requirements for the obtention of the degree of Master of Science in Biotechnology

Production and characterization of electrospun composite fibers: confinement of thermosensitive microgels

Materials engineering focuses on the assembly of materials´ properties to design new products with the best performance. By using sub-micrometer size materials in the production of composites, it is possible to obtain objects with properties that none of their compounds show individually. Once three-dimensional materials can be easily customized to obtain desired properties, much interest has been paid to nanostructured poly-mers in order to build biocompatible devices. Over the past years, the thermosensitive microgels have become more common in the framework of bio-materials with potential applicability in therapy and/or diagnostics. In addition, high aspect ratio biopolymers fibers have been produced using the cost-effective method called electrospinning. Taking advantage of both microgels and electrospun fibers, surfaces with enhanced functionalities can be obtained and, therefore employed in a wide range of applications. This dissertation reports on the confinement of stimuli-responsive microgels through the colloidal electro-spinning process. The process mainly depends on the composition, properties and patterning of the precur-sor materials within the polymer jet. Microgels as well as the electrospun non-woven mats were investigated to correlate the starting materials with the final morphology of the composite fibers. PNIPAAm and PNIPAAm/Chitosan thermosensitive microgels with different compositions were obtained via surfactant free emulsion polymerization (SFEP) and characterized in terms of chemical structure, morphology, thermal sta-bility, swelling properties and thermosensitivity. Finally, the colloidal electrospinning method was carried out from spinning solutions composed of the stable microgel dispersions (up to a concentration of about 35 wt. % microgels) and a polymer solution of PEO/water/ethanol mixture acting as fiber template solution. The confinement of microgels was confirmed by Scanning Electron Microscopy (SEM). The electrospinning process was statistically analysed providing the optimum set of parameters aimed to minimize the fiber diameter, which give rise to electrospun nanofibers of PNIPAAm microgels/PEO with a mean fiber diameter of 63 ± 25 nm.

Modulation of the secretome of hBMSCs by tailoring the macromolecular gradient in hydrogels to generate tissue-to-tissue interfaces

Tissue-to-tissue interfaces are commonly present in all tissues exhibiting structural, biological and chemical gradients serving a wide range of physiological functions. These interfaces are responsible for mediation of load transfer between two adjacent tissues. They are also important structures in sustaining the cellular communications to retain tissueâ s functional integration and homeostasis. [1] All cells have the capacity to sense and respond to physical and chemical stimulus and when cultured in three-dimensional (3D) environments they tend to perform their function better than in two-dimensional (2D) environments. Spatial and temporal 3D gradient hydrogels better resemble the natural environment of cells in mimicking their extracellular matrix. [2] In this study we hypothesize that differential functional properties can be engineered by modulation of macromolecule gradients in a cell seeded threedimensional hydrogel system. Specifically, differential paracrine secretory profiles can be engineered using human Bone Marrow Stem Cells (hBMSCâ s). Hence, the specific objectives of this study are to: assemble the macromolecular gradient hydrogels to evaluate the suitablity for hBMSCâ s encapsulation by cellular viability and biofunctionality by assessing the paracrine secretion of hBMSCâ s over time. The gradient hydrogels solutions were prepared by blend of macromolecules in one solution such as hyaluronic (HA) acid and collagen (Col) at different ratios. The gradient hydrogels were fabricated into cylindrical silicon moulds with higher ratio solutions assembled at the bottom of the mould and adding the two solutions consecutively on top of each other. The labelling of the macromolecules was performed to confirm the gradient through fluorescence microscopy. Additionally, AFM was conducted to assess the gradient hydrogels stiffness. Gradient hydrogels characterization was performed by HA and Col degradation assay, degree of crosslinking and stability. hBMSCâ s at P3 were encapsulated into each batch solution at 106 cells/ml solution and gradient hydrogels were produced as previously described. The hBMSCâ s were observed under confocal microscopy to assess viability by Live/Dead® staining. Cellular behaviour concerning proliferation and matrix deposition was also performed. Secretory cytokine measurement for pro-inflammatory and angiogenesis factors was carried out using ELISA. At genomic level, qPCR was carried out. The 3D gradient hydrogels platform made of different macromolecules showed to be a suitable environment for hBMSCâ s. The hBMSCâ s gradient hydrogels supported high cell survival and exhibited biofunctionality. Besides, the 3D gradient hydrogels demonstrated differentially secretion of pro-inflammatory and angiogenic factors by the encapsulated hBMSCâ s. References: 1. Mikos, AG. et al., Engineering complex tissues. Tissue Engineering 12,3307, 2006 2. Phillips, JE. et al., Proc Natl Acad Sci USA, 26:12170-5, 2008

Epidermis recreation in spongy-like hydrogels: New opportunities to explore epidermis-like analogues

[Excerpt] On the road to successfully achieving skin regeneration, 3D matrices/scaffolds that provide the adequate physico-chemical and biological cues to recreate the ideal healing environment are believed to be a key element [1], [2] and [3]. Numerous polymeric matrices derived from both natural [4] and [5] and synthetic [6], [7] and [8] sources have been used as cellular supports; nowadays, fewer matrices are simple carriers, and more and more are ECM analogues that can actively participate in the healing process. Therefore, the attractive characteristics of hydrogels, such as high water content, tunable elasticity and facilitated mass transportation, have made them excellent materials to mimic cells’ native environment [9]. Moreover, their hygroscopic nature [10] and possibility of attaining soft tissues-like mechanical properties mean they have potential for exploitation as wound healing promoters [11], [12], [13] and [14]. Nonetheless, hydrogels lack natural cell adhesion sites [15], which limits the maximization of their potential in the recreation of the cell niche. This issue has been tackled through the use of a range of sophisticated approaches to decorate the hydrogels with adhesion sequences such as arginine-glycine-aspartic acid (RGD) derived from fibronectin [16], [17] and [18], and tyrosine-isoleucine-glycine-serine-arginine (YIGSR) derived from laminin [18] and [19], which not only aim to modulate cell adhesion, but also influencing cell fate and survival [18]. Nonetheless, its widespread use is still limited by significant costs associated with the use of recombinant bioactive molecules.