Exploring the Factors Affecting the Solubility of Chitosan in Water

We explore the role of crystallinity and inter- or intramolecular forces in chitosan for its solubility in water and demonstrate the expansion of its solubility to a wider pH range. Due to its semicrystalline nature, derived mainly from inter- and intramolecular hydrogen bonds, chitosan is water-soluble only at pH < 6. In acidic conditions, its amino groups can be partially protonated resulting in repulsion between positively charged macrochains, thereby allowing diffusion of water molecules and subsequent solvation of macromolecules. We show that chemical disruption of chitosan crystallinity by partial re-acetylation or physical disruption caused by the addition of urea and guanidine hydrochloride broadens the pH-solubility range for this biopolymer.

Why is Chitosan Mucoadhesive?

Chitosan is a biocompatible and biodegradable amino polysaccharide, which is soluble in aqueous solutions at pH < 6.5. It has been widely used for developing drug delivery systems because of its excellent mucoadhesive properties. Although many studies report on chitosan being mucoadhesive, the nature of interactions between chitosan and mucin remains poorly defined. Here, we have examined the role of primary amino groups and the role of electrostatic attraction, hydrogen bonding, and hydrophobic effects on aggregation of gastric mucin in the presence of chitosan. Reducing the number of amino groups through their half acetylation results in expansion of chitosan’s pH-solubility window up to pH 7.4 but also reduces its capacity to aggregate mucin. We demonstrated that electrostatic attraction forces between chitosan and gastric mucin can be suppressed in the presence of 0.2 mol/L sodium chloride; however, this does not prevent the aggregation of mucin particles in the presence of this biopolymer. The presence of 8 mol/L urea or 10% v/v ethanol in solutions also affects mucin aggregation in the presence of chitosan, demonstrating the role of hydrogen bonding and hydrophobic effects, respectively, in mucoadhesion.

In vitro fermentation of chitosan derivatives by mixed cultures of human faecal bacteria

Stirred, pH controlled batch cultures were carried out with faecal inocula and various chitosans to investigate the fermentation of chitosan derivatives by the human gut flora. Changes in bacterial levels and short chain fatty acids were measured over time. Low, medium and high molecular weight chitosan caused a decrease in bacteroides, bifidobacteria, clostridia and lactobacilli. A similar pattern was seen with chitosan oligosaccharide (COS). Butyrate levels also decreased. A three-stage fermentation model of the human colon was used for investigation of the metabolism of COS. In a region representing the proximal colon, clostridia decreased while lactobacilli increased. In the region representing the transverse colon, bacteroides and clostridia increased. Distally a small increase in bacteroides occurred. Butyrate levels increased. Under the highly competitive conditions of the human colon, many members of the microflora, are unable to compete for chitosans of low, medium or high molecular weight. COS were more easily utilised and when added to an in vitro colonic model led to increased production of butyrate, but some populations of potentially detrimental bacteria also increased. (c) 2005 Elsevier Ltd. All rights reserved.

Effects of drying methods and conditions on antimicrobial activity of edible chitosan films enriched with galangal extract

The aim of this work was to study the effects of drying methods and conditions (i.e., ambient drying, hot air drying at 40 degrees C, vacuum drying and low-pressure superheated steam drying within the temperature range of 70-90 degrees C at an absolute pressure of 10 kPa) as well as the concentration of galangal extract on the antimicrobial activity of edible chitosan films against Staphylococcus aureus. Galangal extract was added to the film forming solution as a natural antimicrobial agent in the concentration range of 0.3-0.9 g/100 g. Fourier transform infrared (FTIR) spectra and swelling of the films were also evaluated to investigate interaction between chitosan and the galangal extract. The antimicrobial activity of the films was evaluated by the disc diffusion and viable cell count method, while the morphology of bacteria treated with the antimicrobial films was observed via transmission electron microscopy (TEM). The antimicrobial activity, swelling and functional group interaction of the antimicrobial films were found to be affected by the drying methods and conditions as well as the concentration of the galangal extract. The electron microscopic observations revealed that cell wall and cell membrane of S. aureus treated by the antimicrobial films were significantly damaged. (C) 2009 Elsevier Ltd. All rights reserved.

Miscibility studies of the blends of chitosan with some cellulose ethers

The polymeric films have been prepared based on blends of chitosan with two cellulose ethers-hydroxypropylmethylcellulose and methylcellulose by casting from acetic acid solutions. The films were transparent and brittle in a dry state but an immersion of the samples in deionized water for over 24 h leads to their disintegration or partial dissolution. The miscibility of the polymers in the blends has been assessed by infrared spectroscopy, wide-angle X-ray diffraction, scanning electron microscopy and thermal gravimetric analysis. It was shown that although weak hydrogen bonding exists between the polymer functional groups the blends are not fully miscible in a dry state. (c) 2005 Elsevier Ltd. All rights reserved.

Mucoadhesive and elastic films based on blends of chitosan and hydroxyethylcellulose

Mucoadhesive polymeric films have been prepared based on blends of chitosan and hydroxyethylcellulose. The blends have been characterized by IR spectroscopy, DSC, WAXD, TGA, SEM, and mechanical testing. It is demonstrated that the mechanical properties of chitosan are improved significantly upon blending with hydroxyethylcellulose. An increase in hydroxyethylcellulose content in the blends makes the materials more elastic. The thermal treatment of the blends at 100 degrees C leads to partial cross-linking of the polymers and formation of water-insoluble but swellable materials. The adhesion of the films towards porcine buccal mucosa decreases with increasing hydroxyethylcellulose content in the blends.

Production and evaluation of dry alginate-chitosan microcapsules as an enteric delivery vehicle for probiotic bacteria

This study investigates the production of alginate microcapsules, which have been coated with the polysaccharide chitosan, and evaluates some of their properties with the intention of improving the gastrointestinal viability of a probiotic (Bifidobacterium breve) by encapsulation in this system. The microcapsules were dried by a variety of methods, and the most suitable was chosen. The work described in this Article is the first report detailing the effects of drying on the properties of these microcapsules and the viability of the bacteria within relative to wet microcapsules. The pH range over which chitosan and alginate form polyelectrolyte complexes was explored by spectrophotometry, and this extended into swelling studies on the microcapsules over a range of pHs associated with the gastrointestinal tract. It was shown that chitosan stabilizes the alginate microcapsules at pHs above 3, extending the stability of the capsules under these conditions. The effect of chitosan exposure time on the coating thickness was investigated for the first time by confocal laser scanning microscopy, and its penetration into the alginate matrix was shown to be particularly slow. Coating with chitosan was found to increase the survival of B. breve in simulated gastric fluid as well as prolong its release upon exposure to intestinal pH.

Chitosan-based mucoadhesive tablets for oral delivery of ibuprofen

Chitosan and its half-acetylated derivative have been compared as excipients in mucoadhesive tablets containing ibuprofen. Initially the powder formulations containing the polymers and the drug were prepared by either co-spray drying or physical co-grinding. Polymer–drug interactions and the degree of drug crystallinity in these formulations were assessed by infrared spectroscopy and differential scanning calorimetry. Tablets were prepared and their swelling and dissolution properties were studied in media of various pHs. Mucoadhesive properties of ibuprofen-loaded and drug-free tablets were evaluated by analysing their detachment from pig gastric mucosa over a range of pHs. Greater polymer–drug interactions were seen for spray-dried particles compared to co-ground samples and drug loading into chitosan-based microparticles (41%) was greater than the corresponding half-acetylated samples (32%). Swelling and drug release was greater with the half-acetylated chitosan tablets than tablets containing the parent polymer and both tablets were mucoadhesive, the extent of which was dependent on substrate pH. The results illustrate the potential sustained drug delivery benefits of both chitosan and its half-acetylated derivative as mucoadhesive tablet excipients.

Chitosan coated alginate beads for the survival of microencapsulated Lactobacillus plantarum in pomegranate juice

This work studied the effect of multi-layer coating of alginate beads on the survival of encapsulated Lactobacillus plantarum in simulated gastric solution and during storage in pomegranate juice at 4 °C. Uncoated, single and double chitosan coated beads were examined. The survival of the cells in simulated gastric solution (pH 1.5) was improved in the case of the chitosan coated beads by 0.5–2 logs compared to the uncoated beads. The cell concentration in pomegranate juice after six weeks of storage was higher than 5.5 log CFU/mL for single and double coated beads, whereas for free cells and uncoated beads the cells died after 4 weeks of storage. In simulated gastric solution, the size of the beads decreased and their hardness increased with time; however, the opposite trend was observed for pomegranate juice, indicating that there is no correlation between cell survival and the hardness of the beads.

Layer-by-layer electrostatic entrapment of protein molecules on superparamagnetic nanoparticle: new strategy to enhance adsorption capacity and maintain biological activity

There is a recent interest to use inorganic-based magnetic nanoparticles as a vehicle to carry biomolecules for various biophysical applications, but direct attachment of the molecules is known to alter their conformation leading to attenuation in activity. In addition, surface immobilization has been limited to monolayer coverage. It is shown that alternate depositions of negatively charged protein molecules, typically bovine serum albumin (BSA) with a positively charged aminocarbohydrate template such as glycol chitosan (GC) on magnetic iron oxide nanoparticle surface as a colloid, are carried out under pH 7.4. Circular dichroism (CD) clearly reveals that the secondary structure of the entrapped BSA sequential depositions in this manner remains totally unaltered which is in sharp contrast to previous attempts. Probing the binding properties of the entrapped BSA using small molecules (Site I and Site II drug compounds) confirms for the first time the full retention of its biological activity as compared with native BSA, which also implies the ready accessibility of the entrapped protein molecules through the porous overlayers. This work clearly suggests a new method to immobilize and store protein molecules beyond monolayer adsorption on a magnetic nanoparticle surface without much structural alteration. This may find applications in magnetic recoverable enzymes or protein delivery.

Isolation of lactoperoxidase using different cation exchange resins by batch and column procedures

Lactoperoxidase (LP) was isolated from whey protein by cation-exchange using Carboxymethyl resin (CM-25C) and Sulphopropyl Toyopearl resin (SP-650C). Both batch and column procedures were employed and the adsorption capacities and extraction efficiencies were compared. The resin bed volume to whey volume ratios were 0.96:1.0 for CM-25C and ≤ 0.64:1.0 for SP-650 indicating higher adsorption capacity of SP-650 compared to CM-25C. The effluent LP activity depended on both the enzyme activity in the whey and the amount of whey loaded on the column within the saturation limits of the resin. The percentage recovery was high below the saturation point and fell off rapidly with over-saturation. While effective recovery was achieved with column extraction procedures, the recovery was poor in batch procedures. The whey-resin contact time had little impact on the enzyme adsorption. SDS PAGE and HPLC analyses were also carried out, the purity was examined and the proteins characterised in terms of molecular weights. Reversed phase HPLC provided clear distinction of the LP and lactoferrin (LF) peaks. The enzyme purity was higher in column effluents compared to batch effluents, judged on the basis of the clarity of the gel bands and the resolved peaks in HPLC chromatograms.

Layer-by-layer deposition of praseodymium oxide on tin-doped indium oxide (ITO) surface

Praseodymium oxide as a thin film of controllable layer is known to display many unique physiochemical properties, which can be useful to ceramic, semiconductive and sensor industries. Here in this short paper, we describe a new chemical method of depositing praseodymium oxide on tin-doped indium oxide (ITO) surface using a layer-by-layer approach. The process is carried out by dipping the ITO in solutions of adsorbable polycationic chitosan and alkaline praseodymium hydroxide Pr(OH)(3) alternatively in order to build up the well-defined multi-layers. XRD suggests that the predominant form of the oxide is Pr6O11, obtained after heat treatment of the deposited ITO in static air at 500 degrees C. Microscopic studies including AFM, TEM and SEM indicate that the deposited oxide particles are uniform in size and shape (cylindrical), mesoporous and the thickness of the film can be controlled. AC impedance measurements of the deposited materials also reveal that the oxide layers display a high electrical conductivity hence suitable for sensor uses. (c) 2006 Elsevier B.V. All rights reserved.

Site specificity of glycation and carboxymethylation of bovine serum albumin by fructose

We report an investigation of the site specificity, extent and nature of modification of bovine serum albumin (BSA) incubated with fructose or glucose at physiological temperature and pH. Sites of early glycation (Heyns rearrangement products (HRP) from fructose; fructoselysine (FL) from glucose) as well as advanced glycation (N-epsilon-(carboxymethyl)lysine; CML) wereanalyzed by liquid chromatography-mass spectrometry. The major site of modification by fructose, like glucose, is Lysine-524 and this results in, respectively, 31 and 76% loss of the corresponding unmodified tryptic peptide, Gln525-Lys533. In addition, total lysine, HRP, FL, CML and N-epsilon-(carboxyethyl)lysine in the incubations, was quantified. Almost all of the loss of lysine in the fructose-modified BSA was attributed to the formation of CML, with the yield of CML being up to 17-fold higher than glucose-modified BSA. A mechanism for the formation of CML from the HRP is proposed.

Mass spectrometric analysis of glucose-modified ribonuclease

RNase A (1 mM) was incubated with glucose (0.4 M) at 37°C for up to 14 days in phosphate buffer (0.2 M, pH 7.4), digested with trypsin and analysed by LC-MS. The major sites of fructoselysine formation were Lys1, Lys7, Lys37 and Lys41. Three of these sites (Lys7, Lys37 and Lys41) were also the major sites of Ne-(carboxymethyl)lysine formation.