Alginate-chitosan Microcapsules Research Articles

Investigation of mechanical properties of soft hydrogel microcapsules in relation to protein delivery using a MEMS force sensor

This article reports the investigation of mechanical properties of alginate-chitosan microcapsules and the relation to protein delivery. For microscale compression testing, a system based on a microelectromechanical systems (MEMS) capacitive force sensor was developed. The bulk microfabricated capacitive force sensors are capable of resolving forces up to 110 microN with a resolution of 33.2 nN along two independent axes. The monolithic force sensors were directly applied to characterizing mechanical properties of soft hydrogel microparticles without assembling additional end-effectors. Protein-loaded alginate-chitosan microcapsules of approximately 20 mum in diameter were prepared by an emulsion-internal gelation-polyelectrolyte coating method. Young's modulus values of individual microcapsules with 1, 2, and 3% chitosan coating were determined through microscale compression testing in both distilled deionized (DDI) water and pH 7.4 phosphate-buffered saline (PBS). Protein release rates were also determined in DDI water and PBS. Finally, protein release rates were correlated with mechanical properties of the microcapsules.

Genipin Cross-Linked Polymeric Alginate-Chitosan Microcapsules for Oral Delivery: In-Vitro Analysis

We have previously reported the preparation of the genipin cross-linked alginate-chitosan (GCAC) microcapsules composed of an alginate core with a genipin cross-linked chitosan membrane. This paper is the further investigation on their structural and physical characteristics. Results showed that the GCAC microcapsules had a smooth and dense surface and a networked interior. Cross-linking by genipin substantially reduced swelling and physical disintegration of microcapsules induced by nongelling ions and calcium sequestrants. Strong resistance to mechanical shear forces and enzymatic degradation was observed. Furthermore, the GCAC membranes were permeable to bovine serum albumin and maintained a molecular weight cutoff at 70 KD, analogous to the widely studied alginate-chitosan, and alginate-poly-L-lysine-alginate microcapsules. The release features and the tolerance of the GCAC microcapsules in the stimulated gastrointestinal environment were also investigated. This GCAC microcapsule formulation offers significant potential as a delivery vehicle for many biomedical applications.

Evaluation of Two Techniques for the Production of Alginate-Based Microcapsules for Oral Delivery of Ibuprofen

A widely used approach for the microencapsulation of drugs especially for controlled drug delivery is emulsion polymerization. The purpose of this study was to comparatively evaluate chitosan-alginate microcapsules for oral delivery of ibuprofen using two techniques. Alginate-based microcapsules were prepared using the emulsion/internal gelation and electrostatic droplet generator/counter-ion coacervation methods. The effect of production parameters on the size of the microcapsules and in vitro ibuprofen release at pH 1.2 and 7.4 was determined. For emulsion/internal gelation method, the results showed that increase in stirring rate, volume of dispersion medium and viscosity of the coating materials did not significantly (p > 0.05) affect the size of the microcapsules. Microcapsules produced by the electrostatic droplets generator method had a narrower size distribution compared to the emulsion method. Drug release at pH 7.4 was considerably more than at pH 1.2, especially for uncoated microcapsules, microcapsules coated with high-viscosity (undigested) chitosan, and microcapsules prepared by the emulsion method. The encapsulation efficiencies of the two methods were similar as over 90% of ibuprofen was entrapped in each case. The two microencapsulation techniques can be applied in the microencapsulation of ibuprofen for oral delivery.

Chitosan–alginate microcapsules for oral delivery of egg yolk immunoglobulin (IgY): In vivo evaluation in a pig model of enteric colibacillosis

In our previous study, the applicability of chitosan–alginate microcapsules for oral delivery of egg yolk immunoglobulin (IgY) was established in a simulated gastrointestinal tract environment. The objective of the present study was to evaluate the protective efficacy of microencapsulated IgY against K88+ ETEC (enterotoxigenic Escherichia coli)-induced diarrhea in 40-day-old pigs. Groups of pigs orally challenged with 10 11 cfu/mL of K88+ ETEC were fed with non-encapsulated IgY, microencapsulated IgY and aureomycin-treated feed respectively. The clinical response of each group was monitored and evaluated in terms of lethargy, inappetence, occurrence of diarrhea, fecal consistency score, weight loss and recovery rate. The results showed that treatment of infected pigs with microencapsulated IgY significantly ( P < 0.05) reduced the K88+ ETEC-induced diarrhea at 24 h post-infection. In contrast, the diarrhea-reducing effect of non-encapsulated IgY was delayed (only evident after 72 h) while normal saline-treated pigs (controls) continued to suffer from diarrhea and dehydration. Similarly, weight gain in microencapsulated IgY-treated pigs was better and significantly different ( P < 0.05) than in non-encapsulated IgY and saline-treated controls. Collectively, these results support previous in vitro observations showing that chitosan–alginate microcapsules can be an effective method of protecting IgY from gastric inactivation, enabling its use for the widespread prevention and control of enteric diseases.

Optimization and characterization of chitosan-alginate microcapsules containing egg yolk immunoglobulin (IgY)

Optimization and characterization of chitosan-alginate microcapsules containing egg yolk immunoglobulin (IgY)

Biocompatibility and membrane strength of C3H10T1/2 cell-loaded alginate-based microcapsules

C3H10T1/2 cells, from a mouse embryonic fibroblast cell line, were used to investigate the improvement of alginate-based microencapsulated cells for cellular therapy. Purified sodium alginate (PSA) and non-purified sodium alginate (SA) were used to prepare alginate-based microcapsules, and their biocompatibility and membrane strength were then compared for the purposes of analyzing the advantages of purifying SA. In addition, poly-l-lysine (PLL) was replaced by chitosan for alginate-chitosan microcapsule preparation. The process of optimization and chemical modification of alginate-chitosan microcapsules using polyethylene glycol was also reviewed. The results showed improved biocompatibility and membrane strength of PSA-based microcapsules. Under optimal conditions, mesenchymal stromal cell (MSC)-loaded alginate-chitosan microcapsules with good morphology could be obtained using PSA and chitosans of medium molecular weight (1.0-2.5 x 10(5)). A chitosan solution of 0.1% (w/v) and a reaction time of 7 min between alginate and chitosan were determined as optimal preparation parameters. It could be concluded that the chemical modification of alginate-based microcapsules can improve their biocompatibility.

Ionic gelation controlled drug delivery systems for gastric-mucoadhesive microcapsules of captopril

A new oral drug delivery system was developed utilizing both the concepts of controlled release and mucoadhesiveness, in order to obtain a unique drug delivery system which could remain in stomach and control the drug release for longer period of time. Captopril microcapsules were prepared with a coat consisting of alginate and a mucoadhesive polymer such as hydroxy propyl methyl cellulose, carbopol 934p, chitosan and cellulose acetate phthalate using emulsification ionic gelation process. The resulting microcapsules were discrete, large, spherical and free flowing. Microencapsulation efficiency was 41.7-89.7% and high percentage efficiency was observed with (9:1) alginate-chitosan microcapsules. All alginate-carbopol 934p microcapsules exhibited good mucoadhesive property in the in vitro wash off test. Drug release pattern for all formulation in 0.1 N HCl (pH 1.2) was diffusion controlled, gradually over 8 h and followed zero order kinetics.

Effect of Solvent Type and Drying Method on Protein Retention in Chitosan-Alginate Microcapsules

Purpose: The effect of solvent used in dissolving chitosan (membrane material) and the microcapsule drying method used, on protein retention in chitosan-alginate microcapsules were studied since these factors affect the physicochemical characteristics of the coating membrane. Method: The microcapsules were prepared by extruding a solution containing alginate and BSA into chitosan/calcium chloride solution prepared with different acid solvents – acetic acid, formic acid, tartaric acid and hydrochloric acid. A portion of the microcapsules was air-dried at ambient temperature while the remaining portion was freeze-dried. The elution of protein from the microcapsules in simulated gastric fluid was monitored spectrophotometrically at &#955;max 280 nm. Results: Tartaric acid effected the highest mean protein retention (54%) after 9 h followed by acetic acid (35%), hydrochloric acid (31%) and formic acid, (30%). There appears to be a link between the pKa of the acids and the degree of chitosan–solvent interaction on the one hand, and protein retention on the other hand. Increase in elution pH from 1.2 to 5.0 did not significantly (P>0.05) affect protein retention. Furthermore, there was no significant difference (p>0.05) between the protein retention capacities of air-dried and freeze-dried microcapsules as both types showed protein retention of 50% after 5 h. Conclusion: Tartaric acid was the most suitable solvent for enhancing protein retention in chitosan-alginate microcapsules in simulated gastric fluid . Keywords: Tartaric acid, chitosan, solvent type, microcapsules, air-drying, freeze-drying.> Tropical Journal of Pharmaceutical Research Vol. 5 (2) 2006: pp. 583-588

Chitosan−Alginate Microcapsules for Oral Delivery of Egg Yolk Immunoglobulin (IgY)

Chitosan-alginate microcapsules were evaluated as a method of oral delivery of IgY antibodies. Physical characteristics, encapsulation efficiency (EE%), the loading capacity for IgY (IgY loading percentage, %, w/w of microcapsules), gastro-resistance, and release characteristics of these microcapsules in vitro under varying pH were investigated. Optimum physical factors were established for preparation of homogeneous, spherical, and smooth microcapsules. IgY loading% was not significantly altered by pH of the encapsulation medium. Encapsulation efficiency was highest (73.93%) at a pH of 3.5, above which EE% decreased significantly (p < 0.05). IgY was released from microcapsules upon exposure to simulated intestinal fluid (SIF, pH 6.8), and decreasing pH increased significantly IgY release (p < 0.05). The stability of IgY in simulated gastric fluid (SGF, pH 1.2) was greatly improved by encapsulation in chitosan-alginate microcapsules, and the residual activity was not affected by pH of the encapsulation medium. Moreover, microencapsulated IgY was significantly resistant to pepsin hydrolysis. This approach may enable intact IgY to reach target microorganisms within the lower digestive tract.

Preparation and characterization of novel polymeric microcapsules for live cell encapsulation and therapy

This article describes the preparation and in vitro characterization of novel genipin cross-linked alginate-chitosan (GCAC) microcapsules that have potential for live cell therapy applications. This microcapsule system, consisting of an alginate core with a covalently cross-linked chitosan membrane, was formed via ionotropic gelation between calcium ions and alginate, followed by chitosan coating by polyelectrolyte complexation and covalent cross-linking of chitosan by naturally derived genipin. Results showed that, using this design concept and the three-step procedure, spherical GCAC microcapsules with improved membrane strength, suppressed capsular swelling, and suitable permeability can be prepared. The suitability of this novel membrane formulation for live cell encapsulation was evaluated, using bacterial Lactobacillus plantarum 80 (pCBH1) (LP80) and mammalian HepG2 as model cells. Results showed that capsular integrity and bacterial cell viability were sustained 6 mo postencapsulation, suggesting the feasibility of using this microcapsule formulation for live bacterial cell encapsulation. The metabolic activity of the encapsulated HepG2 was also investigated. Results suggested the potential capacity of this GCAC microcapsule in cell therapy and the control of cell signaling; however, further research is required.

Immobilization of Laccase by Alginate–Chitosan Microcapsules and its Use in Dye Decolorization

Laccase is a ligninolytic enzyme that is widespread in white-rot fungi. Alginate–chitosan microcapsules prepared by an emulsification–internal gelation technique were used to immobilize laccase. Parameters of the immobilization process were optimized. Under the optimal immobilization conditions (2% sodium alginate, 2% CaCl2, 0.3% chitosan and 1:8 ratio by volume of enzyme to alginate), the loading efficiency and immobilized yield of immobilized laccase were 88.12% and 46.93%, respectively. Laccase stability was increased after immobilization. Both the free and immobilized laccase alone showed a very low decolorization efficiency when Alizarin Red was selected for dye decolorization test. When 0.1 mM 2,2′-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) was added into the decolorization system, the decolorization efficiency increased significantly. Immobilized laccase retained 35.73% activity after three reaction cycles. The result demonstrated that immobilized laccase has potential application in dyestuff treatment.

Genipin Cross-Linked Alginate-Chitosan Microcapsules: Membrane Characterization and Optimization of Cross-Linking Reaction

The genipin cross-linked alginate-chitosan (GCAC) microcapsule, composed of an alginate core and a genipin cross-linked chitosan membrane, was recently proposed for live cell encapsulation and other delivery applications. This article for the first time describes the details of the microcapsule membrane characterization using a noninvasive and in situ method without any physical or chemical modifications on the samples. Results showed that the cross-linking reaction generated the fluorescent chitosan-genipin conjugates. The cross-linked chitosan membrane was clearly visualized by confocal laser scanning microscopy (CLSM). A straightforward assessment on the membrane thickness and relative intensity was successfully achieved. CLSM studies showed that the shell-like cross-linked chitosan membranes of approximately 37 microm in thickness were formed surrounding the microcapsule. The reaction variables, including cross-linking temperature and time significantly affected the fluorescence intensity of the membranes. Elevating the cross-linking temperature from 4 to 37 degrees C drastically intensified the membrane fluorescence, suggesting the attainment of a high degree of cross-linking on the chitosan membrane. Extended cross-linking time altered the cross-linked membranes in modulation. Although genipin concentration and cross-linking time had little effects on the membrane thickness, cross-linking at higher temperatures tended to form relatively thinner membranes.

In vitro study of alginate?chitosan microcapsules: an alternative to liver cell transplants for the treatment of liver failure

The application of alginate-chitosan (AC) microcapsules to liver cell transplantation has not been previously investigated. In the current in vitro study, we have investigated the potential of AC microcapsules for the encapsulation of liver cells and show that the AC membrane supports the survival, proliferation and protein secretion by entrapped hepatocytes. The AC membrane provides cell immuno-isolation and has the potential for cell cryopreservation. The AC microcapsule has several advantages compared to more widely used alginate-poly-L-lysine (APA) microcapsules for the application of cell therapy.

A protein delivery system: biodegradable alginate–chitosan–poly(lactic-co-glycolic acid) composite microspheres

In the present study we developed alginate–chitosan–poly(lactic-co-glycolic acid) (PLGA) composite microspheres to elevate protein entrapment efficiency and decrease its burst release. Bovine serum albumin (BSA), which used as the model protein, was entrapped into the alginate microcapsules by a modified emulsification method in an isopropyl alcohol-washed way. The rapid drug releases were sustained by chitosan coating. To obtain the desired release properties, the alginate–chitosan microcapsules were further incorporated in the PLGA to form the composite microspheres. The average diameter of the composite microcapsules was 31 ± 9 μm and the encapsulation efficiency was 81–87%, while that of conventional PLGA microspheres was just 61–65%. Furthermore, the burst releases at 1 h of BSA entrapped in composite microspheres which containing PLGA (50:50) and PLGA (70:30) decreased to 24% and 8% in PBS, and further decreased to 5% and 2% in saline. On the contrary, the burst releases of conventional PLGA microspheres were 48% and 52% in PBS, respectively. Moreover, the release profiles could be manipulated by regulating the ratios of poly(lactic acid) to poly(glycolic acid) in the composite microspheres.

INVESTIGATION OF A NOVEL MICROCAPSULE MEMBRANE INTEGRATING POLYETHYLENE GLYCOL TO ALGINATE, POLY-L-LYSINE AND CHITOSAN MICROCAPSULES FOR THE APPLICATION OF LIVER CELL TRANSPLANTATION

P668 Aims: Liver transplantation is the main form of therapy for most liver diseases. A major drawback to transplantation is the lack of donors and continuous requirement of immunosuppressants. Microencapsulation is an emerging technology which can be used to entrap isolated hepatocytes for cell transplanation as an alternative treatment to several liver diseases. One of the limiting factors in the progress of such therapy is attaining a biocompatible polymer enabling the long-term entrapment and growth of the hepatocytes without causing adverse host immune responses. Presently the most commonly studied membranes are the alginate-poly-l-lysine-alginate (APA) and alginate-chitosan (AC) microcapsules, however there remain limitations associated with these membranes. In the current study, improvements to the biocompatibility and stability of these membranes are investigated by the addition of a polyethylene glycol (PEG) coating and the potential of a novel membrane combining alginate, poly-l-lysine, chitosan and PEG is studied with the objective of proposing a membrane most suitable for cell entrapment. Methods: Five different microcapsules were prepared including APA, APA with PEG, AC, AC with PEG and the novel alginate-chitosan-PEG-PLL-alginate (ACPPA) microcapsule. Mechanical strength of the capsules were assessed using an osmotic pressure test and a rotational stress test. A fluorescence reader (FLx800) was used to detect permeability of dextran through the membranes. Morphological studies on capsule integrity in serum was observed microscopically. Cytotoxicity tests were perfomed by encapsulating Human HepG2 cells and performing an MTT calorimetric assay for monitroing metabolic activity. Further studies on immuno protection using macrophages and cryopreservation potential are also to be investigated. Results: Microcapsules of approximately 400+/-30μm were prepared. Stability tests, using osmotic pressure techniques, reveal the addition of PEG resulted in an increase in mechanical stability of both APA and AC capsules by over 50%. The rotational stress test indicate the novel membrane formulation to exhibit mechanical strength similar to APA membranes and greater than the AC, ACPEG and APPEG microcapsules. The ACPPA membrane was found to retain integrity in FBS. Cytotoxicity tests using human HepG2 cells indicate low viability of AC membranes however, positive MTT was observed for the remaining 4 membranes studied which implies that the addition of PEG can support cellular growth. Conclusions: Results support previous studies which indicate PEG may improve biocompatibility of polymers. The integration of PEG to microcapsules enhances mechanical strength and supports the proliferation of liver cells. This study confirms that the novel membrane combining PLL, chitosan and PEG shows similar mechanical properties as APA capsules. As previous studies indicate that APA encapsulated cells can result in cell growth and protein adhesion to the membrane surface, the new membrane may be an alternative biomaterial for microencapsulation cell therapy.

Swelling behaviour of alginate–chitosan microcapsules prepared by external gelation or internal gelation technology

Swelling behaviour is one of the important properties for microcapsules made by hydrogels, which always affects the diffusion and release of drugs when the microcapsules are applied in drug delivery systems. In this paper, alginate–chitosan microcapsules were prepared by different technologies called external or internal gelation process respectively. With the volume swelling degree ( S w) as an index, the effect of properties of chitosan on the swelling behaviour of both microcapsules was investigated. It was demonstrated that the microcapsules with low molecular weight and high concentration of chitosan gave rise to low S w. Considering the need of maintaining drug activity and drug loading, neutral pH and short gelation time were favorable. It was also noticed that S w of internal gelation microcapsules was lower than that of external gelation microcapsules, which was interpreted by the structure analysis of internal or external gelation Ca–alginate beads with the aid of confocal laser scanning microscope.

Mechanical characterization of biocompatible microspheres and microcapsules by direct compression.

The mechanical properties of biocompatible microparticles including alginate microspheres and alginate-chitosan microcapsules with different wall thickness were determined using a micromanipulation technique. Single microparticles with diameters of 20-60 microm were compressed to a given deformation and held, and compressed to rupture at different speeds. The corresponding force imposed on them was measured simultaneously by a force transducer. Results showed that the force imposed on these particles increased when they were compressed, but relaxed significantly when they were held. For alginate microspheres, the faster the compression speed was, the greater the force being imposed on them at a given deformation. Alginate-chitosan microcapsules showed less force relaxation when they were held, compared with alginate microspheres. The thicker their wall was, the less significant force relaxation the microcapsules exhibited. The mean rupture force of alginate microspheres increased with the compression speed, but this effect in general became less for alginate-chitosan microcapsules, which depended on their wall thickness. However, the deformation at rupture for all three samples was independent of the compression speed. On average, the alginate-chitosan microcapsules were bigger than alginate microspheres and had a greater rupture force.

Enzyme immobilization in novel alginate–chitosan core-shell microcapsules

Alginate–chitosan core-shell microcapsules were prepared in order to develop a biocompatible matrix for enzyme immobilization, where the protein is retained either in a liquid or solid core and the shell allows permeability control over substrates and products. The permeability coefficients of different molecular weight compounds (vitamin B2, vitamin B12, and myoglobin) were determined through sodium tripolyphosphate (Na-TPP)-crosslinked chitosan membrane. The microcapsule core was formed by crosslinking sodium alginate with either calcium or barium ions. The crosslinked alginate core was uniformly coated with a chitosan layer and crosslinked with Na-TPP. In the case of calcium alginate, the phosphate ions of Na-TPP were able to extract the calcium ions from alginate and liquefy the core. A model enzyme, β-galactosidase, was immobilized in the alginate core and the catalytic activity was measured with o-nitrophenyl- β- d-galactopyranoside (ONPG). Change in the activity of free and immobilized enzyme was determined at three different temperatures. Na-TPP crosslinked chitosan membranes were found to be permeable to solutes of up to 17,000 Da molecular weight. The enzyme loading efficiency was higher in the barium alginate core (100%) as compared to the calcium alginate core (60%). The rate of ONPG conversion to o-nitrophenol was faster in the case of calcium alginate–chitosan microcapsules as compared to barium alginate–chitosan microcapsules. Barium alginate–chitosan microcapsules, however, did improve the stability of the enzyme at 37°C relative to calcium alginate–chitosan microcapsules or free enzyme. This study illustrates a new method of enzyme immobilization for biotechnology applications using liquid or solid core and shell microcapsule technology.

Studies on alginate–chitosan microcapsules and renal arterial embolization in rabbits

Spherical and well-dispersed alginate-chitosan microcapsules, with a mean diameter of 77.28±0.93 μm ( n=3), were prepared by the emulsification–gelation method. Adriamycin hydrochloride (ADM) was used as a model drug to investigate the drug loading capacity and release characteristics of the microcapsules. The drug/carrier ratio and chitosan concentration influenced the encapsulation efficiency of adriamycin. The adriamycin release from microcapsules was obviously different in 0.1 mol/l HCl from that in phosphate-buffered saline (PBS, pH 7.4). The drug was completely and rapidly released in 0.1 mol/l HCl, while it showed a sustained release after a burst release in PBS. The increase in chitosan concentration had no effect on adriamycin release in PBS. Using sulforhodamin B (SRB)-staining survival assay, the inhibition of adriamycin alginate-chitosan microcapsules (ADM-ACM) to different cancer cell lines (human BGC-823 cells, Bel-7402 cells and Hela cells) in vitro was determined. The inhibitory rate of ADM-ACM suspension to the three cell lines significantly outran that of ADM solution, no matter at high or low concentration. The effects of blank alginate-chitosan microcapsules (BACM) on renal arterial embolization were examined with transcatheter arterial embolization in rabbits. The angiogram and histopathological results indicated the blank microcapsules had excellent short- and long-term effects on renal arterial embolization.

Evaluation of protein release from chitosan-aginate microcapsules produced using external or internal gelation

A series of experiments was undertaken to evaluate the diffusion of a model protein, i.e. bovine serum albumin (BSA), from chitosan-alginate microcapsules produced using either internal or external gelation. Diffusion of BSA was quantified during the microcapsule manufacture processes (gelation, washing, rinsing) and during incubation in conditions simulating the pH encountered during the gastric and intestinal phases of digestion. Encapsulation of an acid phosphmonoesterase permitted in situ protein localization, providing evidence to explain results obtained with BSA. There was significantly greater protein loss from internally versus externally-gelled chitosan-alginate microcapsules during the manufacture process (37.6% versus 4.7%, respectively). Similar trends were observed during 24 h incubation in 0.1 N hydrochloric acid. Increasing alginate concentration from 2-4% (w:v) did not significantly reduce losses from internally-gelled microcapsules. Addition of 0.25 m NaCl to the gelling medium significantly increased protein diffusing during microcapsule manufacture and acid incubation from externally gelled microcapsules. In situ protein localization revealed a higher level of protein near the surface of the microcapsules of externally gelled microcapsules versus internal gelation. The above data indicate that externally-gelled microcapsules are inhomogeneous with a higher concentration of alginate near the microcapsule surface, thus reducing the porosity of the resulting microcapsules. These results suggest that the porous nature of internally-gelled chitosan-alginate microcapsules may result in low encapsulation efficiency, depending on the nature of the product being encapsulated.