Calcium Alginate Hydrogel Research Articles

Simple method to determine flow characteristics of injectable hydrogels

This article presents a simple inexpensive method to measure flow and viscoelastic properties of calcium alginate hydrogels or gel solutions and is particularly suitable for quality control in small laboratory settings. This method can be considered an extension of the Melt Flow Index (MFI) method, which is routinely used in the polymer industry to indirectly determine the molecular weight of polymers. Results of alginate gels at various crosslinking densities obtained with this new test are compared to complex viscosity data obtained from a standard cone and plate rheometer, and demonstrate very good correlation. A mathematical model describing the apparent viscosity of the alginate gels in this test has also been developed. © 2010 Wiley Periodicals, Inc. J Appl Polym Sci, 2010

Specific rebinding property of protein macromolecularly imprinted polymer microspheres based on calcium alginate hydrogel via gas jetting‐dropping method

AbstractAlginate hydrogel polymer microspheres are usually prepared by inverse suspending method. The main problem is the remained suspending medium and surfactant on the beads. Syringe dropping or extruding method avoids using of the organic solvents. However, it is difficult to control the particle dimension and produce in large quantity. We have developed a new method called nitrogen gas jetting‐dropping (GJD) method for the preparation of bovine serum albumin (BSA) imprinted polymer microspheres based on calcium alginate hydrogel. The microspheres can be produced continuously without using any hydrophobic medium or surfactant. The particle diameter is controlled by adjusting pressure and needle dimension. BSA‐imprinted alginate microspheres were prepared by the GJD method and the rebinding tests were investigated. The equilibrium rebinding capacity ranged from 0.01 to 0.014 μmol/g. The imprinting efficiency towards BSA was from 1.46 to 2.17 with the greatest separation factor of 1.523 against a contrastive template ovalbumin (OVA). © 2010 Wiley Periodicals, Inc. J Appl Polym Sci, 2010

Encapsulation of fibroblasts causes accelerated alginate hydrogel degradation

Calcium-alginate hydrogel has been widely studied as a material for cell encapsulation for tissue engineering. At present, the effect that cells have on the degradation of alginate hydrogel is largely unknown. We have shown that fibroblasts encapsulated at a density of 7.5 x 10(5) cells ml(-1) in both 2% and 5% w/v alginate remain viable for at least 60 days. Rheological analysis was used to study how the mechanical properties exhibited by alginate hydrogel changed during 28 days in vitro culture. Alginate degradation was shown to occur throughout the study but was greatest within the first 7 days of culture for all samples, which correlated with a sharp release of calcium ions from the construct. Fibroblasts were shown to increase the rate of degradation during the first 7 days when compared with acellular samples in both 2% and 5% w/v gels, but after 28 days both acellular and cell-encapsulating samples retained disc-shaped morphologies and gel-like spectra. The results demonstrate that although at an early stage cells influence the mechanical properties of encapsulating alginate, over a longer period of culture, the hydrogels retain sufficient mechanical integrity to exhibit gel-like properties. This allows sustained immobilization of the cells at the desired location in vivo where they can produce extracellular matrix and growth factors to expedite the healing process.

Inkjet fabrication and characterization of calcium alginate microcapsules

The present work describes a method for producing calcium alginate hydrogel microcapsules in the size range of 50–70 μm by means of a piezoelectric drop-on-demand inkjet device. Particles were prepared by emitting droplets of 0.5% and 1% (w/w) sodium alginate solutions into a magnetically stirred pool of CaCl 2 solution of variable viscosity ranging from 1 to 100 mPas. The effect of viscosity on the morphology of the resulting microcapsules was systematically investigated — lower viscosity of the receiving solution has lead to the formation of elongated particles, medium viscosities lead to spherical capsules, and for higher viscosities flattened particles were obtained. The applied voltage used for driving the piezoelectric inkjet print-head was found to be the most significant parameter for influencing the droplet size. The duration of the voltage pulse or the droplet viscosity had only minor effects. The release rates of two model substances (methylene blue dye and vitamin B12) from alginate beads were measured and their effective diffusion coefficients determined as function of alginate concentration.

Reactive inkjet printing of calcium alginate hydrogel porogens—a new strategy to open-pore structured matrices with controlled geometry

Taking advantage of inkjet's ability to dispense uniform droplets in the picolitre/nanolitre ranges of volumes, we have generated reversible hydrogel porogen beads using reactive printing, which we use as templates for creating networks of pores with monomodally distributed pore sizes.

Novel technique to control inner and outer diameter of calcium-alginate hydrogel hollow microfibers, and immobilization of mammalian cells

Calcium-alginate hydrogel hollow microfibers have a potential to be utilized in various fields such as tissue engineering. In this study, we developed a new technique for exquisite control of inner diameter (id) as well as outer diameter (od) of the microfibers using a coaxial triple cylinder. Aqueous solutions of dextran (core solution), sodium alginate (sample solution) and CaCl 2 (sheath solution) were extruded simultaneously from the inner, intermediate and outer cylinders, respectively. The ratio of id to od could be controlled only by the ratio of flow volume of core solution to total flow volume of core and sample solutions (core volume ratio). The od depended on core volume ratio, flow velocities of core and sample solution at the tip of the intermediate cylinder, tip id of intermediate cylinder and flow velocity of sheath solution. Two types of mammalian cells could be separately located in the hollow core and the gel part of the microfibers. We confirmed that the cell-enclosing process scarcely influenced the viability of the enclosed cells. Although we did not perform long-term culture of the cell-enclosing microfibers in this study, these results indicate that our system has a high potential to fabricate biomimetic scaffold for tissue-engineered tubular tissues.

Structural characterization of calcium alginate matrices by means of mechanical and release tests.

In this paper we have concentrated on the characterization of calcium alginate hydrogels loaded with a model drug (myoglobin) by means of a mechanical approach; in addition, release tests of myoglobin from alginate hydrogels were performed. At a fixed temperature, relaxation tests (mechanical study) were carried out on matrices constituted by different polymer concentrations. The interpretation of the relaxation behavior of the different matrices was conducted using the generalized Maxwell model; as a result of this investigation it was possible to conclude that for polymer concentrations greater than 0.5 g/ 100 mL the matrices behaved as solid materials. In addition, it was observed that the mechanical properties of the matrices increased with polymer concentration. With regard to the release tests, the diffusion coefficient of myoglobin in the matrix in relation to polymer concentrations was determined. The mechanical and release data where then analyzed by Flory’s theory and by a modified free-volume theory, respectively, to estimate the network mesh size ξ. The comparison between the mesh sizes obtained by the two approaches showed a satisfactory agreement for polymer concentrations greater than 0.5 g/100 mL. It should be noted that the approach proposed here to determine the polymeric network meshes is absolutely general and can be advantageously applied to the characterization of other similar polymeric systems.

Electroaddressing of Cell Populations by Co‐Deposition with Calcium Alginate Hydrogels

AbstractElectroaddressing of biological components at specific device addresses is attractive because it enlists the capabilities of electronics to provide spatiotemporally controlled electrical signals. Here, the electrodeposition of calcium alginate hydrogels at specific electrode addresses is reported. The method employs the low pH generated at the anode to locally solubilize calcium ions from insoluble calcium carbonate. The solubilized Ca2+ can then bind alginate to induce this polysaccharide to undergo a localized sol‐gel transition. Calcium alginate gel formation is shown to be spatially controlled in the normal and lateral dimensions. The deposition method is sufficiently benign that it can be used to entrap the bacteria E. coli. The entrapped cells are able to grow and respond to chemical inducers in their environment. Also, the entrapped cells can be liberated from the gel network by adding sodium citrate that can compete with alginate for Ca2+ binding. The capabilities of calcium alginate electrodeposition is illustrated by entrapping reporter cells that can recognize the quorum sensing autoinducer 2 (AI‐2) signaling molecule. These reporter cells were observed to recognize and respond to AI‐2 generated from an external bacterial population. Thus, calcium alginate electrodeposition provides a programmable method for the spatiotemporally controllable assembly of cell populations for cell‐based biosensing and for studying cell‐cell signaling.

Apparent Production of Enzymatically Synthesized Amylose in DMSO by Means of Calcium Alginate Hydrogel Beads/DMSO System

In this paper, we describe the apparent production of enzymatically synthesized amylose in DMSO by means of the calcium alginate hydrogel beads/DMSO system as the reaction field of the phosphorylase-catalyzed enzymatic polymerization. When the calcium alginate hydrogel beads including glucose-1-phosphate, maltoheptaose, and phosphorylase were suspended in DMSO and the system was slowly stirred at 40°C for 12 h, the reaction proceeded to produce amylose, which eluted to the DMSO solution. The obtained amylose was purified by the treatment with ion-exchange resins, and its structure was confirmed by the 1H NMR spectrum. The time-course experiment in the present system revealed that the phosphorylase-catalyzed enzymatic polymerization was carried out for 15 min on the inside of the calcium alginate hydrogel beads and the produced amylose was gradually eluted to the surrounding DMSO solution. The comparison of the present system with the general enzymatic polymerization in aqueous buffer solution suggested that the yield and the degree of polymerization of amylose in the present system were comparable to those in aqueous buffer solution.

Structure−Function Relationship of Calcium Alginate Hydrogels: A Novel Crystal-Forming Engineering

Hydrogels of calcium alginate are the cross-linked networks containing a large fraction of water, which were used as the precursors of calcium carbonate (CaCO3) mineralization for the first time. The well-defined geometry, the permeability, and the ion-exchange property of these pregels favored the facile fabrication of calcite superstructures through the slow inpouring of ammonia and carbon dioxide gases. When calcium alginate hydrogels sponged up a relatively high amount of liquid, the resulting products with the outside calcite sequences transcribed the spongelike pregel beads with the outside nucleation sites of carboxyl groups. The inner characteristic of these CaCO3 superstructures showed the endocentric growth trends of calcite, indicating the permeability of hydrogel beads and the diffusing directions of permeated gases. In the presence of low liquid content, the pregels favored the formation of calcite superstructures with relatively smooth surfaces, demonstrating the inside lamellar array of cal...

Preparation and characterization of porous biodegradable microspheres used for controlled protein delivery

In this study, a new approach for protein delivery is introduced, involving biodegradable porous poly( d, l-lactic-co-glycolic acid) (PLGA) microspheres blocked with calcium alginate hydrogel. Porous microspheres were prepared by solvent evaporation method using sodium oleate as an extractable porogen. The resultant microspheres show porous surface and three-dimensional network structure with interconnecting pores. Human serum albumin (HSA) used as model protein, was adsorbed into micropores of porous microspheres by a solution immersing method. Thus, the harsh preparation conditions, such as the aqueous/organic interface produced by a water-in-oil microemulsion and the acute shearing strength brought by a high-speed homogenizer or a sonicater, inducing protein denaturation and aggregation could be avoided. Then, calcium alginate hydrogel was fabricated to block micropores of protein-loaded porous microspheres. A sustained protein release in vitro could be achieved in this way. Furthermore, it is noteworthy that the protein maintained its structural integrity after this preparation. The porous polymeric microspheres may have great potential application for water-soluble drugs, such as protein, vaccine and gene delivery.

Injectable and in situ gelling hydrogels for modified protein release

A novel injectable polysaccharide system based on calcium Alginate (Ca-Alg) hydrogel and two Dextran methacrylate derivatives (DexMA) was recently developed. The resulting Interpenetrating Polymer Network showed a synergistic mechanical behavior that can be exploited to target the hydrogel properties towards specific biomedical needs. In the present paper, hydrogels composed of 3% (w/v) Ca-Alg and Dextran (Mw 40 x 10(4) and 500 x 10(4)), derivatized with methacrylic groups (derivatization degrees 5 and 30%) at concentrations 5% (w/v), were characterized. The data reported here evidenced that Mw and derivatization degree of Dex chains can deeply affect the mechanical as well a model protein (Horseradish peroxidase) delivery rate. The enzymatic activity of such model protein was never significantly altered by the adopted experimental conditions.

Preparation and characterization of an injectable composite

Hydrogels are increasingly used in medicine due to their potential to be delivered into the body in a minimally invasive manner and to be gelated at the site of introduction subsequently. The aim of this study was to develop a novel injectable and in situ-forming gel composite (GC) comprised of calcium alginate hydrogel and nano-hydroxyapatite/collagen (nHAC), assess its rheological, mechanical and in vitro degradable properties, and discuss the gelation mechanism. Injectable property test showed that the injectability of GC was tunable. Rheological results indicated that three phases of pre-gel, sol-gel phase transformation and post-gel could be found in the process of gelation. The compressive elastic modulus (E) and shear modulus (G) are in the range of 17.0-56.0 kPa and 24.7-55.0 kPa, respectively. During the in vitro degradation, the wet weight increased in the first week, then declined in the following 3 weeks, but the dry weight lost continuously during whole study. Meanwhile, the surface changed greatly after 2 weeks, but samples did not break down up to 28 days. These data indicate that GC exhibits controllable initial setting time and final setting time, tunable injectability, which provides a possible injectable material for bone repair and bone tissue engineering.

Treatment of soy milk with Debaryomyces hansenii cells immobilised in alginate

Whole cells of Debaryomyces hansenii UFV-1 were permeabilised with ethanol and immobilised in calcium alginate hydrogel. The optimum pH and temperature for α-galactosidase activities of permeabilised free (PFC) and permeabilised immobilised cells (PIC) were 4.5 and 60 °C; and 4.0 and 70 °C, respectively. PIC α-galactosidase was more stable than that of PFC. The incubation of PIC at 60 and 70 °C promoted an increase in α-galactosidase activity. The α-galactosidase activity was maintained when PIC was used in three repeated batches. The K m values for PIC and PFC α-galactosidases, with ρNPαGal, were 0.82 mM and 0.30 mM, respectively. Soy milk treatment with PIC for 6 h at 60 °C promoted 100% hydrolysis of raffinose oligosaccharides.

The benefit of human embryonic stem cell encapsulation for prolonged feeder-free maintenance

The majority of methodologies for maintaining human embryonic stem cell (hESC) pluripotency require the use of human or animal feeder cell layers, the most common being murine embryonic fibroblasts. In this study, we applied a protocol aimed at maintaining hESCs in culture without exposure to animal cells or proteins. hESCs were encapsulated in 1.1% (w/v) calcium alginate hydrogels and grown in basic maintenance medium for a period of up to 260 days. Investigation of the cell aggregates formed within the hydrogels yielded no evidence of the formation of any of the three germ layers, although the hESCs retained their pluripotency and could differentiate when they were subsequently cultured in a conditioned environment. Immunohistochemistry and RT-PCR showed that the hESC aggregates expressed protein and gene markers characteristic of pluripotency including Oct-4, Nanog, SSEA-4, TRA-1-60 and TRA-1-81. At the ultrastructural level, the cells were arranged in closely packed clusters and showed no cytoplasmic organelles, suggesting an undifferentiated state. These data show that it is possible to maintain hESCs in an undifferentiated state, without passaging or embryoid body formation, and without animal contamination.

Immobilized Manihot esculenta preparation as a novel biocatalyst in the enantioselective acetylation of racemic alcohols

The enzymatic preparation obtained from a discard of Manihot esculenta roots has been successfully immobilized on calcium alginate hydrogels. This preparation has been tested as a chiral biocatalyst in the enzymatic acylation of a set of racemic aromatic alcohols. Depending on the reaction conditions, excellent enantioselectivities can be achieved. Some parameters that can alter the biocatalytic properties of the enzyme, such as solvent, temperature, acyl donor and substrate structure have been studied exhaustively in order to establish a deeper knowledge of this novel biocatalyst.

Comparison of different technologies for alginate beads production

AbstractThis paper describes the results of the round robin experiment “Bead production technologies” carried out during the COST 840 action “Bioencapsulation Innovation and Technologies” within the 5th Framework Program of the European Community. In this round robin experiment, calcium alginate hydrogel beads with the diameter of (800 ± 100) μm were produced by the most common bead production technologies using 0.5–4 mass % sodium alginate solutions as starting material. Dynamic viscosity of the alginate solutions ranged from less than 50 mPa s up to more than 10000 mPa s. With the coaxial air-flow and electrostatic enhanced dropping technologies as well as with the JetCutter technology in the soft-landing mode, beads were produced from all alginate solutions, whereas the vibration technology was not capable to process the high-viscosity 3 % and 4 % alginate solutions. Spherical beads were generated by the electrostatic and the JetCutter technologies. Slightly deformed beads were obtained from high-viscosity alginate solutions using the coaxial airflow and from the 0.5 % and 2 % alginate solutions using the vibration technology. The rate of bead production using the JetCutter was about 10 times higher than with the vibration technology and more than 10000 times higher than with the coaxial air-flow and electrostatic technology.

Microfluidic scaffolds for tissue engineering

Most methods to culture cells in three dimensions depend on a cell-seedable biomaterial to define the global structure of the culture and the microenvironment of the cells. Efforts to tailor these scaffolds have focused on the chemical and mechanical properties of the biomaterial itself. Here, we present a strategy to control the distributions of soluble chemicals within the scaffold with convective mass transfer via microfluidic networks embedded directly within the cell-seeded biomaterial. Our presentation of this strategy includes: a lithographic technique to build functional microfluidic structures within a calcium alginate hydrogel seeded with cells; characterization of this process with respect to microstructural fidelity and cell viability; characterization of convective and diffusive mass transfer of small and large solutes within this microfluidic scaffold; and demonstration of temporal and spatial control of the distribution of non-reactive solutes and reactive solutes (that is, metabolites) within the bulk of the scaffold. This approach to control the chemical environment on a micrometre scale within a macroscopic scaffold could aid in engineering complex tissues.

Fabrication of Artificial Endothelialized Tubes with Predetermined Three‐Dimensional Configuration from Flexible Cell‐Enclosing Alginate Fibers

One possible strategy for creating three-dimensional (3D) tissue-engineered organs in vitro is to develop a vasculature for sufficient transport of oxygen and nutrients within these constructs. Here, we describe a novel technique to fabricate endothelialized tubes with predetermined 3D configuration, as a starting point for self-developing capillary-like networks in vitro. Calcium-alginate hydrogel fibers of ca. 250 and 500 mum in diameter, enclosing bovine carotid artery vascular endothelial cells (BECs), were used as templates for endothelialized tubes. Fibers were prepared by extruding a 2% (w/v) sodium alginate solution containing BECs into a 100 mM calcium chloride solution flowing in the same direction. Fibers were embedded in type I collagen gels and enzymatically degraded by alginate lyase, resulting in channels with predetermined 3D configuration filled with a BEC suspension. Cells attached to and covered the surfaces of the channels. Exposing the cells to medium containing basic fibroblast growth factor resulted in their migration into the ambient collagen gel and self-assembly into capillary-like structures. These results demonstrate that using artificial endothelialized tubes with predetermined 3D configuration, as a starting point for a self-developing capillary-like network, could be potentially useful for constructing 3D tissue-engineered organs.

Enhanced Overhauser contrast in proton–electron double-resonance imaging of the formation of an alginate hydrogel

Proton–electron double-resonance imaging (PEDRI) was recently employed to monitor the process of formation of a calcium alginate hydrogel at a field of 16 mT. Here, under the same experimental conditions, images obtained through this technique are compared to images obtained by conventional T 2-weighted method. The results confirm that the image contrast obtained using PEDRI, thanks to the Overhauser effect, exhibits an improved sensitivity with respect to changes in water mobility as previously suggested in the literature. Furthermore, by increasing the echo time interval for the T 2-weighted images, important features of the gelling dynamics obtained via PEDRI could not be reproduced.