Study of the Thermal Phase Transition of Poly(N,N-diethylacrylamide-co-N-ethylacrylamide) Random Copolymers in Aqueous Solution.

N-alkyl-substituted polyacrylamides exhibit a thermal coil-to-globule transition in aqueous solution driven by an increase in hydrophobic interactions with rising temperature. With the aim of understanding the role of N-alkyl substituents in the thermal transition, this study focuses on the molecular interactions underlying the phase transition of poly(N,N-diethylacrylamide-co-N-ethylacrylamide) random copolymers. Poly(N,N-diethylacrylamide) (PDEAm), poly(N-ethylacrylamide) (PNEAm), and their random copolymers were synthesized by free radical polymerization and their chemical structure characterized spectroscopically. It was found that the values of the cloud-point temperature increased with PNEAm content, and particle aggregation processes took place, increasing the negative charge density on their surface. The cloud-point temperature of each copolymer decreased with respect to the theoretical values calculated assuming an absence of interactions. It is attributed to the formation of intra- and interchain hydrogen bonding in aqueous solutions. These interactions favor the formation of more hydrophobic macromolecular segments, thereby promoting the cooperative nature of the transition. These results definitively reveal the dominant mechanism occurring during the phase transition in the aqueous solutions of these copolymers.

3D Printing of Cellulase-Laden Cellulose Nanofiber/Chitosan Hydrogel Composites: Towards Tissue Engineering Functional Biomaterials with Enzyme-Mediated Biodegradation.

The 3D printing of a multifunctional hydrogel biomaterial with bioactivity for tissue engineering, good mechanical properties and a biodegradability mediated by free and encapsulated cellulase was proposed. Bioinks of cellulase-laden and cellulose nanofiber filled chitosan viscous suspensions were used to 3D print enzymatic biodegradable and biocompatible cellulose nanofiber (CNF) reinforced chitosan (CHI) hydrogels. The study of the kinetics of CNF enzymatic degradation was studied in situ in fibroblast cell culture. To preserve enzyme stability as well as to guarantee its sustained release, the cellulase was preliminarily encapsulated in chitosan-caseinate nanoparticles, which were further incorporated in the CNF/CHI viscous suspension before the 3D printing of the ink. The incorporation of the enzyme within the CHI/CNF hydrogel contributed to control the decrease of the CNF mechanical reinforcement in the long term while keeping the cell growth-promoting property of chitosan. The hydrolysis kinetics of cellulose in the 3D printed scaffolds showed a slow but sustained degradation of the CNFs with enzyme, with approximately 65% and 55% relative activities still obtained after 14 days of incubation for the encapsulated and free enzyme, respectively. The 3D printed composite hydrogels showed excellent cytocompatibility supporting fibroblast cell attachment, proliferation and growth. Ultimately, the concomitant cell growth and biodegradation of CNFs within the 3D printed CHI/CNF scaffolds highlights the remarkable potential of CHI/CNF composites in the design of tissue models for the development of 3D constructs with tailored in vitro/in vivo degradability for biomedical applications.

Chitosan Hydrogels Based on the Diels-Alder Click Reaction: Rheological and Kinetic Study.

The Diels-Alder reaction is recognized to generate highly selective and regiospecific cycloadducts. In this study, we carried out a rheological and kinetic study of N-furfuryl chitosan hydrogels based on the Diels-Alder click reaction with different poly(ethylene)glycol-maleimide derivatives in dilute aqueous acidic solutions. It was possible to prepare clear and transparent hydrogels with excellent mechanical properties. Applying the Winter and Chambon criterion the gel times were estimated at different temperatures, and the activation energy was calculated. The higher the temperature of gelation, the higher the reaction rate. The crosslinking density and the elastic properties seem to be controlled by the diffusion of the polymer segments, rather than by the kinetics of the reaction. An increase in the concentration of any of the two functional groups is accompanied by a higher crosslinking density regardless maleimide:furan molar ratio. The hydrogel showed an improvement in their mechanical properties as the temperature increases up to 70 °C. Above that, there is a drop in G' values indicating that there is a process opposing to the Diels-Alder reaction, most likely the retro-Diels-Alder.

Polyphosphazene-Based Nanocarriers for the Release of Camptothecin and Epirubicin.

The design and study of efficient polymer-based drug delivery systems for the controlled release of anticancer drugs is one of the pillars of nanomedicine. The fight against metastatic and invasive cancers demands therapeutic candidates with increased and selective toxicity towards malignant cells, long-term activity and reduced side effects. In this sense, polyphosphazene nanocarriers were synthesized for the sustained release of the anticancer drugs camptothecin (CPT) and epirubicin (EPI). Linear poly(dichloro)phosphazene was modified with lipophilic tocopherol or testosterone glycinate, with antioxidant and antitumor activity, and with hydrophilic Jeffamine M1000 to obtain different polyphosphazene nanocarriers. It allowed us to encapsulate the lipophilic CPT and the more hydrophilic EPI. The encapsulation process was carried out via solvent exchange/precipitation, attaining a 9.2-13.6 wt% of CPT and 0.3-2.4 wt% of EPI. CPT-loaded polyphosphazenes formed 140-200 nm aggregates in simulated body physiological conditions (PBS, pH 7.4), resulting in an 80-100-fold increase of CPT solubility. EPI-loaded polyphosphazenes formed 250 nm aggregates in an aqueous medium. CPT and EPI release (PBS, pH 7.4, 37 °C) was monitored for 202 h, being almost linear during the first 8 h. The slow release of testosterone and tocopherol was also sustained for 150 h in PBS (pH 7.4 and 6.0) at 37 °C. The co-delivery of testosterone or tocopherol and the anticancer drugs from the nanocarriers was expected. Cells of the human breast cancer cell line MCF-7 demonstrated good uptake of anticancer-drug-loaded nanocarriers after 6 h. Similarly, MCF-7 spheroids showed good uptake of the anticancer-drug-loaded aggregates after 72 h. Almost all anticancer-drug-loaded polyphosphazenes exhibited similar or superior toxicity against MCF-7 cells and spheroids when compared to raw anticancer drugs. Additionally, cell-cycle arrest in the G2/M phase was increased in response to the drug-loaded nanocarriers. Almost no toxicity of anticancer-drug-loaded aggregates against primary human lung fibroblasts was observed. Furthermore, the aggregates displayed no hemolytic activity, which is in contrast to the parent anticancer drugs. Consequently, synthesized polyphosphazene-based nanocarriers might be potential nanomedicines for chemotherapy.

Self-Assembled Silk Fibroin-Based Aggregates for Delivery of Camptothecin.

A water-soluble hydrolysate of silk fibroin (SF) (~30 kDa) was esterified with tocopherol, ergocalciferol, and testosterone to form SF aggregates for the controlled delivery of the anticancer drug camptothecin (CPT). Elemental analysis and 1H NMR spectroscopy showed a degree of substitution (DS) on SF of 0.4 to 3.8 mol %. Yields of 58 to 71% on vitamins- and testosterone-grafted SF conjugates were achieved. CPT was efficiently incorporated into the lipophilic core of SF aggregates using a dialysis-precipitation method, achieving drug contents of 6.3-8.5 wt %. FTIR spectra and DSC thermograms showed that tocopherol- and testosterone-grafted SF conjugates predominantly adopted a ß-sheet conformation. After the esterification of tyrosine residues on SF chains with the vitamin or testosterone, the hydrodynamic diameters almost doubled or tripled that of SF. The zeta potential values after esterification increased to about -30 mV, which favors the stability of aggregates in aqueous medium. Controlled and almost quantitative release of CPT was achieved after 6 days in PBS at 37 °C, with almost linear release during the first 8 h. MCF-7 cancer cells exhibited good uptake of CPT-loaded SF aggregates after 6 h, causing cell death and cell cycle arrest in the G2/M phase. Substantial uptake of the CPT-loaded aggregates into MCF-7 spheroids was shown after 3 days. Furthermore, all CPT-loaded SF aggregates demonstrated superior toxicity to MCF-7 spheroids compared with parent CPT. Blank SF aggregates induced no hemolysis at pH 6.2 and 7.4, while CPT-loaded SF aggregates provoked hemolysis at pH 6.2 but not at pH 7.4. In contrast, parent CPT caused hemolysis at both pH tested. Therefore, CPT-loaded SF aggregates are promising candidates for chemotherapy.

Cellulose Nanofiber-Reinforced Chitosan Hydrogel Composites for Intervertebral Disc Tissue Repair.

The development of non-cellularized composites of chitosan (CHI) hydrogels, filled with cellulose nanofibers (CNFs) of the type nanofibrillated cellulose, was proposed for the repair and regeneration of the intervertebral disc (IVD) annulus fibrosus (AF) tissue. With the achievement of CNF-filled CHI hydrogels, biomaterial-based implants were designed to restore damaged/degenerated discs. The structural, mechanical and biological properties of the developed hydrogel composites were investigated. The neutralization of weakly acidic aqueous CNF/CHI viscous suspensions in NaOH yielded composites of physical hydrogels in which the cellulose nanofibers reinforced the CHI matrix, as investigated by means of microtensile testing under controlled humidity. We assessed the suitability of the achieved biomaterials for intervertebral disc tissue engineering in ex vivo experiments using spine pig models. Cellulose nanofiber-filled chitosan hydrogels can be used as implants in AF tissue defects to restore IVD biomechanics and constitute contention patches against disc nucleus protrusion while serving as support for IVD regeneration.

Novel Brassinosteroid-Modified Polyethylene Glycol Micelles for Controlled Release of Agrochemicals.

Two synthetic analogues of brassinosteroids (DI31 and S7) exhibit good plant growth enhancer activity. However, their hydrophobicity and quick metabolism in plants have limited their application and benefits in agriculture. Our objective was to prepare novel brassinosteroid-modified polyethylene glycol (PEG) micelles to achieve controlled release with extended stability while retaining agrochemical activity. Spectroscopic studies confirmed quantitative disubstitution of studied PEGs with the brassinosteroids, while elemental analysis assessed purity of the synthesized conjugates. Conjugates were also characterized by X-ray diffraction and thermal analysis. Dynamic and static light scattering showed stable and homogeneous approximately spherical micelles with average hydrodynamic diameters of 22-120 nm and almost neutral ζ potential. Spherical 30-140 nm micelles were observed by electron microscopy. Sustained in vitro releases at pH 5.5 were extended up to 96 h. Prepared PEG micelles showed good agrochemical activity in the radish seed bioassay and no cytotoxicity to the human microvascular endothelial cell line in the MTS test.

Preparation and Characterization of Chitosan Obtained from Shells of Shrimp (Litopenaeus vannamei Boone).

The main source of commercial chitosan is the extensive deacetylation of its parent polymer chitin. It is present in green algae, the cell walls or fungi and in the exoskeleton of crustaceans. A novel procedure for preparing chitosan from shrimp shells was developed. The procedure involves two 10-minutes bleaching steps with ethanol after the usual demineralization and deproteinization processes. Before deacetylation, chitin was immersed in 12.5 M NaOH, cooled down and kept frozen for 24 h. The obtained chitosan was characterized using scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), UV, X-ray diffraction (XRD) and viscosimetry. Samples of white chitosan with acetylation degrees below 9 % were obtained, as determined by FTIR and UV-first derivative spectroscopy. The change in the morphology of samples was followed by SEM. The ash content of chitosan samples were all below 0.063 % . Chitosan was soluble in 1 % acetic acid with insoluble contents of 0.62 % or less. XRD patterns exhibited the characteristic peaks of chitosan centered at 10 and 20 degrees in 2 θ . The molecular weight of chitosan was between 2.3 and 2.8 × 10 5 g/mol. It is concluded that the procedure developed in the present work allowed obtaining chitosans with physical and chemical properties suitable for pharmaceutical applications.

Fine microstructure of processed chitosan nanofibril networks preserving directional packing and high molecular weight.

Crystalline chitosan nanofibril networks were prepared, preserving the native structural packing and the polymer high molecular weight. The fine microstructure of the nanomaterial, obtained by mild hydrolysis of chitosan (CHI), was characterized by using synchrotron small- and wide-angle X-ray scattering (SAXS and WAXS), transmission electron microscopy (TEM) and electron diffraction. Hydrolysis of chitosan yielded a network of crystalline nanofibrils, containing both allomorphs of chitosan: hydrated and anhydrous. The comparison of WAXS data in transmission and reflection mode revealed the preferential orientation of the CHI crystals when subjected to mechanical compression constrains. The results are in agreement with the existence of a network nanostructure containing fiber-like crystals with the principal axis parallel to the polymer chain axis. The evolution of the CHI allomorphic composition with temperature was studied to further elucidate the mechanism of structural transitions occurring during CHI nanofibril network processing.

Recubrimiento de microesferas de quitosana-ibuprofeno con un complejo interpolimérico pH dependiente / Coating of chitosan-Ibuprofen microspheres with a pH-depending interpolymer complex

Introducción: el efecto irritante sobre la mucosa gástrica que producen los antiinflamatorios no esteroideos es una de sus principales reacciones adversas. La encapsulación de estos en matrices poliméricas con propiedades entéricas constituye una alternativa tecnológica para solucionar dicho problema. Objetivo: obtener micropartículas de quitosana cargadas con ibuprofeno recubiertas con un complejo interpolimérico pH dependiente a base de poli(ácido acrílico)/poli(N-vinil-2-pirrolidona). Métodos: se prepararon micropartículas de quitosana cargadas con ibuprofeno mediante secado por aspersión y se determinó el rendimiento del proceso y la eficiencia de encapsulación. Las micropartículas se recubrieron con un complejo interpolimérico pH dependiente de poli(ácido acrílico)/poli(N-vinil-2-pirrolidona), empleando la técnica de emulsión/evaporación del disolvente. Mediante espectroscopia infrarroja de transformada de Fourier, se comprobó la formación del complejo, y la evaluación morfológica se realizó por microscopia electrónica de barrido. Los estudios de liberación se realizaron en fluido gástrico e intestinal simulados (FGS pH= 1,2; FIS pH= 6,8). Resultados: en el proceso de obtención de las micropartículas de quitosana y quitosana-ibuprofeno hubo un rendimiento de 69 ± 1 por ciento y 54,4 ± 0,8 por ciento respectivamente. La eficiencia de encapsulación resultó de 46,8 ± 0,7 por ciento. Las micropartículas recubiertas presentaron una superficie rugosa. La formación del complejo se confirmó a través de los cambios observados en la posición de las bandas de absorción de los grupos funcionales involucrados en la formación del enlace por puente de hidrógeno. La liberación de ibuprofeno en FGS resultó del 40 por ciento para las micropartículas sin recubrimiento, mientras que fue despreciable en el caso de las micropartículas recubiertas durante el intervalo de tiempo estudiado. Conclusiones: los resultados muestran las potencialidades del complejo interpolimérico...

Introduction: the irritating effect on the gastric mucosa caused by non-steroidal anti-inflammatory drugs is one of the main adverse reactions. Their encapsulation in polymer matrices with enteric properties is a technological alternative to solve the problem. Objective: to obtain ibuprofen-loaded chitosan microparticles coated with a pH dependent interpolymer complex based on poly(acrylic acid)/poly(N-vinyl-2-pyrrolidone). Methods: Ibuprofen-loaded chitosan microparticles were prepared through the spray drying technique; the yield and the efficiency of encapsulation were evaluated. Microparticles were coated with a pH-dependent interpolymer complex based on poly(acrylic acid)/poly(N-vinyl-2-pyrrolidone) using the emulsion/solvent evaporation technique. The complex formation was verified by Fourier transform infrared spectroscopy and the morphological evaluation was made with the electronic scanning microscopy. Release studies used simulated gastric (SGF, pH= 1.2) and intestinal (SIF, pH= 6.8) fluids. Results: in the process of obtaining the chitosan and chitosan-ibuprofen microparticles, the yield rates amounted to 69 ± 1 percent and 54.4 ± 0.8 percent respectively were obtained. The encapsulation efficiency was 46.8 ± 0.7 percent. The coated microparticles presented rough surface. Complex formation was confirmed by changes in the position of the absorption bands of the functional groups involved in hydrogen bonding. The release of ibuprofen from uncoated microparticles in simulated gastrointestinal fluid reached 40 percent whereas it was neglectable in the coated microparticles during the study interval. Conclusions: the results show the potential of poly(acrylic acid)/poly(N-vinyl-2-pyrrolidone) interpolymer complex as pH dependent cover for use as enteric coating to reduce the side effects on the gastric mucosa of medications such as non-steroidal anti-inflammatory drugs(AU)

Recubrimiento de microesferas de quitosana-ibuprofeno con un complejo interpolimérico pH dependiente / Coating of chitosan-Ibuprofen microspheres with a pH-depending interpolymer complex

INTRODUCCIÓN: el efecto irritante sobre la mucosa gástrica que producen los antiinflamatorios no esteroideos es una de sus principales reacciones adversas. La encapsulación de estos en matrices poliméricas con propiedades entéricas constituye una alternativa tecnológica para solucionar dicho problema. OBJETIVO: obtener micropartículas de quitosana cargadas con ibuprofeno recubiertas con un complejo interpolimérico pH dependiente a base de poli(ácido acrílico)/poli(N-vinil-2-pirrolidona) MÉTODOS: se prepararon micropartículas de quitosana cargadas con ibuprofeno mediante secado por aspersión y se determinó el rendimiento del proceso y la eficiencia de encapsulación. Las micropartículas se recubrieron con un complejo interpolimérico pH dependiente de poli(ácido acrílico)/poli(N-vinil-2-pirrolidona), empleando la técnica de emulsión/evaporación del disolvente. Mediante espectroscopia infrarroja de transformada de Fourier, se comprobó la formación del complejo, y la evaluación morfológica se realizó por microscopia electrónica de barrido. Los estudios de liberación se realizaron en fluido gástrico e intestinal simulados (FGS pH= 1,2; FIS pH= 6,8). RESULTADOS: en el proceso de obtención de las micropartículas de quitosana y quitosana-ibuprofeno hubo un rendimiento de 69 ± 1 por ciento y 54,4 ± 0,8 por ciento respectivamente. La eficiencia de encapsulación resultó de 46,8 ± 0,7 por ciento. Las micropartículas recubiertas presentaron una superficie rugosa. La formación del complejo se confirmó a través de los cambios observados en la posición de las bandas de absorción de los grupos funcionales involucrados en la formación del enlace por puente de hidrógeno. La liberación de ibuprofeno en FGS resultó del 40 por ciento para las micropartículas sin recubrimiento, mientras que fue despreciable en el caso de las micropartículas recubiertas durante el intervalo de tiempo estudiado. CONCLUSIONES: los resultados muestran las potencialidades del complejo interpolimérico poli(ácido acrílico)/poli(N-vinil-2-pirrolidona) como cubierta pH dependiente, con vistas a obtener un recubrimiento de tipo entérico que reduzca los efectos adversos sobre la mucosa gástrica de fármacos como los antiinflamatorios no esteroideos(AU)

INTRODUCTION: the irritating effect on the gastric mucosa caused by non-steroidal anti-inflammatory drugs is one of the main adverse reactions. Their encapsulation in polymer matrices with enteric properties is a technological alternative to solve the problem. OBJECTIVE: to obtain ibuprofen-loaded chitosan microparticles coated with a pH dependent interpolymer complex based on poly(acrylic acid)/poly(N-vinyl-2-pyrrolidone). METHODS: Ibuprofen-loaded chitosan microparticles were prepared through the spray drying technique; the yield and the efficiency of encapsulation were evaluated. Microparticles were coated with a pH-dependent interpolymer complex based on poly(acrylic acid)/poly(N-vinyl-2-pyrrolidone) using the emulsion/solvent evaporation technique. The complex formation was verified by Fourier transform infrared spectroscopy and the morphological evaluation was made with the electronic scanning microscopy. Release studies used simulated gastric (SGF, pH= 1.2) and intestinal (SIF, pH= 6.8) fluids. RESULTS: in the process of obtaining the chitosan and chitosan-ibuprofen microparticles, the yield rates amounted to 69 ± 1 percent and 54.4 ± 0.8 percent respectively were obtained. The encapsulation efficiency was 46.8 ± 0.7 percent he coated microparticles presented rough surface. Complex formation was confirmed by changes in the position of the absorption bands of the functional groups involved in hydrogen bonding. The release of ibuprofen from uncoated microparticles in simulated gastrointestinal fluid reached 40 percent whereas it was neglectable in the coated microparticles during the study interval. CONCLUSIONS: the results show the potential of poly(acrylic acid)/poly(N-vinyl-2-pyrrolidone) interpolymer complex as pH dependent cover for use as enteric coating to reduce the side effects on the gastric mucosa of medications such as non-steroidal anti-inflammatory drugs(AU).

Hydrogel wound dressing preparation at the laboratory scale by using electron beam and gamma radiation / Preparación de apósitos de membranas de hidrogeles a escala de laboratorio mediante haz de electrones y radiación gamma

The present work describes the preparation of hydrogel based on cross-linked networks of poly (N-vinylpirrolidone), PVP, with polyethyleneglicol and agar with 90% water and PVP nancomposites with a synthetic nanoclay, Laponite XLG, for use as burn dressings. These systems were obtained in two ways: using gamma Co-60 and electron beam radiation. The gelation obtained dose was Dg= 1.72 kGy. The elastic modulus of hydrogel was independent of the method of irradiation. It was 0.39 MPa for the hydrogel irradiated with gamma Co-60 and 0.38 MPa for electron beam irradiation. The elastic modulus of the nanocomposite membrane was 1.25 MPa, three times higher. These results indicate that the PVP/Laponite XLG nanocomposite hydrogel membrane is the best choice for wound dressing applications due to its high water sorption capacity and its superior mechanical properties.

En el presente trabajo se describe la preparación de hidrogeles basados en redes entrecruzadas de poli (N-vinilpirrolidona), PVP con polietilenglicol, agar y un 90% de agua, y nanocomposites de PVP con una nanoarcilla sintética, la Laponita XLG para su empleo como apósito para quemaduras. Estos sistemas se obtuvieron por dos vías: radiación gamma de Co-60 y haz de electrones. La dosis de gelificación obtenida fue de Dg= 1.72 kGy. El módulo elástico de los hidrogeles resultó independiente del método de irradiación, siendo igual a 0.39 MPa para el irradiado con Co-60 y 0.38 MPa para el irradiado con haz de electrones. El módulo elástico de la membrana de nanocomposite fue 3 veces superior, 1.25 MPa. Estos resultados muestran que los hidrogeles de nanocomposites de PVP/Laponita XLG resultan superiores para su aplicación en el tratamiento de quemaduras, por su alta capacidad de sorción de agua y sus mejores propiedades mecánicas.

Kinetics study of the solid-state acid hydrolysis of chitosan: evolution of the crystallinity and macromolecular structure.

The heterogeneous hydrolysis of fully deacetylated chitosan solid samples was carried out with concentrated HCl. The hydrolysis kinetics was studied at different temperatures and HCl concentrations. From 5 to 50 degrees C in the hydrolysis time range up to 50 h, a monomodal distribution of molecular weights was observed connected to the only degradation of amorphous domains. Between 70 and 90 degrees C and for the hydrolysis longest times, a multimodal distribution appeared with the additional hydrolysis of the crystalline phase. The crystallinity index increased from 57 to 73% with the elimination and partial recrystallization of amorphous regions. X-ray diffraction patterns revealed the presence of the anhydrous polymorph, absent in the starting materials only containing the hydrated polymorph. The apparent crystallite width (from the Scherrer equation) of both the anhydrous and hydrated allomorphs did not vary significantly with time despite the increase in the fraction of anhydrous allomorph. Therefore, the hydrolysis in the solid state was complex, revealing several regimes. The activation energy parameters were deduced, and the mechanisms were discussed.

Ferrocene branched chitosan for the construction of a reagentless amperometric hydrogen peroxide biosensor.

Chitosan was chemically branched with ferrocene moieties and further used as a support for the immobilization of horseradish peroxidase on a glassy carbon electrode. The reagentless biosensor device showed a linear amperometric response toward hydrogen peroxide concentrations between 35 x 10(-6) M and 2.0 x 10(-3) M. The biosensor reached 95% of the steady-state current in about 20 s and its sensitivity was 28.4 x 10(-3) microA x M(-1). The enzyme electrode retained 94% of its initial activity after 2 weeks of storage at 4 degrees C in 50 x 10(-3) M sodium phosphate buffer at pH 7.0.