Characterization of calcium alginate and chitosan-treated calcium alginate gel beads entrapping allyl isothiocyanate

Calcium alginate (CA), chitosan-coated calcium alginate (CCA-I), and chitosan–calcium alginate complex (CCA-II) gel beads, in which an oil-in-water emulsion containing allyl isothiocyanate (AITC) was entrapped, were prepared and characterized for efficient oral delivery of AITC. The AITC entrapment efficiency was 81% for CA gel beads, whereas about 30% lower values were determined for the chitosan-treated gel beads. Swelling studies showed that all the gel beads suddenly shrunk in simulated gastric fluid (pH 1.2). In simulated intestinal fluid (pH 7.4), CA and CCA-I gel beads rapidly disintegrated, whereas CCA-II gel beads highly swelled without degradation probably due to the strong chitosan–alginate complexation. Release studies revealed that most entrapped AITC was released during the shrinkage, degradation, or swelling of the gel beads, and the chitosan treatments, especially the chitosan–alginate complexation, were effective in suppressing the release. CCA-II gel beads showed the highest bead stability and AITC retention under simulated gastrointestinal pH conditions.     

Use of polyelectrolyte complex-stabilized calcium alginate gel for entrapment of beta-amylase

Calcium alginate gel stabilized with a polyelectrolyte complex (PEC) consisting of potassium poly(vinyl alcohol) sulfate (KPVS) and trimethylammonium glycol chitosan iodide (TGCI) was used for the immobilization of beta-amylase. The immobilization was made by gelling aqueous droplets of enzyme solution including both sodium alginate and KPVS in a CaCl(2) solution containing TGCI. The activity of the enzyme entrapped into the stabilized gel beads was evaluated by studying the batch reaction kinetics of enzyme-catalyzed hydrolysis of maltotetraose. Repeated kinetic measurements, totaling 18, were carried out at fixed time intervals. After each measurement the beads were stirred for 1 day in a freshly prepared 10 mM NaCl solution at 3 degrees C. It was found that the immobilized system remained stable without leading to a serious loss of the activity or to a large leakage of the enzyme from the support. This was explained as being due to a PEC-crosslinked contracted network structure of the stabilized gel matrix.     

Glucose oxidase release from calcium alginate gel capsules

Diffusion of glucose oxidase within calcium alginate gel capsules has been assayed and the experimental data fitted to a simple semi-empirical power equation, which is used to analyse the solute release from polymeric devices. It was found that an increase in the concentration of sodium alginate and calcium chloride gives rise to a reduction in the enzyme leakage. This was verified when glucose oxidase (GOD) diffusion percentages were compared in capsules with thicknesses of the same order of magnitude but obtained under different experimental conditions. So, the use of sodium alginate and calcium chloride solutions of concentrations 0.5% w/v and 2.6% w/v, respectively, lead to a diffusion percentage of 25 +/- 2. This percentage was reduced to 8 +/- 3 when sodium alginate and calcium chloride concentrations were fixed at 1% w/v and 4% w/v, respectively, even though the thicknesses of the capsules were of the same order of magnitude.     

Effective diffusion coefficient of sucrose in calcium alginate gel.

The effective diffusion coefficient of sucrose in 5% calcium alginate gel containing 41.6 g.d.c. l-1. Saccharomyces cerevisiae was investigated. Both free and immobilized S. cerevisiae in 0.175 cm and 0.3 cm diameter particles were used and the reactions were achieved in a medium containing 100 g l-1 sucrose and 0.05 M CaCl2. With the assumption that the microorganisms did not grow or die in this medium, the results were analyzed according to Michaelis-Menten kinetics and the values of the parameters were determined as: Vm = 0.256 g ml-1 gel h-1, Km0 = 0.097 g ml-1, Km1 = 0.125 g ml-1, and Km2 = 0.165 g ml-1. Using these values, effectiveness factors were calculated as eta 1 = 0.89 and eta 2 = 0.76, and effective diffusion coefficients for sucrose in calcium alginate gel were determined as De1 = 4.1 X 10(-6) cm2 s-1 and De2 = 4.0 X 10(-6) cm2 s-1, for the particle size involved.     

Diffusivity of Cu2+ in calcium alginate gel beads

The diffusivity of Cu(2+) in calcium alginate beads calculated by the shrinking core model (SCM) was reevaluated in this work. The results obtained in this work were significantly different than those by the original authors. There were excellent agreements between the results obtained by the SCM in this work and those by the more rigorous linear absorption model (LAM) by the original authors. (c) 1994 John Wiley & Sons, Inc.     

Viability of ectomycorrhizal fungus mycelium entrapped in calcium alginate gel

 We studied the viability of fragmented mycelium of Pisolithus tinctorius and Paxillus involutus entrapped in calcium alginate gel to determine the efficacy of this method of producing ectomycorrhizal fungus inoculum. Fungi were grown in MMN solution at 28  °C before being fragmented in a blender and subsequently entrapped in calcium alginate. We tested different ratios of alginate and mycelium suspension to 0.7 M CaCl₂. The ratio 8 : 10 resulted in well-formed beads of the highest viability for Paxillus involutus (99%) and for Pisolithus tinctorius (75%). Paxillus involutus mycelium was more than 90% viable when entrapped mycelium was 10 to 50 days old, and Pisolithus tinctorius attained its highest viability (55%) for 20- to 40-day-old mycelium. Gel entrapped Paxillus involutus mycelium grew well at all temperatures after 30 days of storage, but viability significantly decreased after 60 days storage at 6  °C on dry filter paper. For gel-entrapped Pisolithus tinctorius mycelium, viability was highest when stored at 25  °C in 0.7 M CaCl₂. Entrapment of Paxillus involutus fragmented mycelium in calcium alginate beads under the conditions that we propose can be used successfully to produce inoculum. Accepted: 11 October 1998     

Diffusivity of Cu(2+) in calcium alginate gel beads

A linear absorption model (LAM) is used to describe the process of metal binding to spherically shaped biopolymers particles. The LAM was solved using a numerical algorithm which calculates diffusivities of metal ion in biopolymer gels. It assumes attainment of rapid metal-biopolymer binding equilibrium accompanied by rate limiting diffusion of the metal ions through the gel. The model was tested using batch experiments in which copper (Cu(2+)) binding with calcium alginate beads was investigated. Biopolymer density in the beads was varied between 2% and 5%. The diffusion coefficient of Cu(2+) calculated from the LAM ranged from 1.19 x 10(-9) to 1.48 x 10(-9) m(2) s(-1) (average 1.31 +/- 0.21 x 10(-9) m(2) s(-1)), independent of biopolymer density. The LAM has theoretical advantages over the shrinking core model (shell progressive model). The latter calculated an unreasonable exponential increase in the diffusion coefficient as density of alginate polymer in the bead increased. (c) 1993 John Wiley & Sons, Inc.     

A study of cryogenic tissue-engineered liver slices in calcium alginate gel for drug testing

To address issues such as transportation and the time-consuming nature of tissue-engineered liver for use as an effective drug metabolism and toxicity testing model, “ready-to-use” cryogenic tissue-engineered liver needs to be studied. The research developed a cryogenic tissue-engineered liver slice (TELS), which comprised of HepG2 cells and calcium alginate gel. Cell viability and liver-specific functions were examined after different cryopreservation and recovery culture times. Then, cryogenic TELSs were used as a drug-testing model and treated with Gefitinib. Cryogenic TELSs were stored at −80 °C to ensure high cell viability. During recovery in culture, the cells in the cryogenic TELS were evenly distributed, massively proliferated, and then formed spheroid-like aggregates from day 1 to day 13. The liver-specific functions in the cryogenic TELS were closely related to cryopreservation time and cell proliferation. As a reproducible drug-testing model, the cryogenic TELS showed an obvious drug reaction after treatment with the Gefitinib. The present study shows that the cryopreservation techniques can be used in drug-testing models.     

Growth and hydrocarbon production ofBotryococcus braunii immobilized in calcium alginate gel

Summary The colonial microalgaB. braunii, immobilized in calcium alginate beads, shows active photoautotrophic growth. Nevertheless, the rates of increase in cell number and, to a lesser extent, in biomass are substantially lower when compared to free cultures. Such features are related to steric contraints which occasion also the formation of large spherical colonies in the gel, showing an unsual mulberry organization. Some cracks due to the development of underlying colonies appear at the surface of the beads. Alga release remains low, however, during the cultures. EntrappedB. braunii retain the ability to produce extracellular hydrocarbons; the structure of the latter is not affected by immobilization but their relative abundances can undergo some variations. Entrapment leads to marked improvements in hydrocarbon production; decrease in growth rates is therefore associated, in alginate gel, with a still more pronounced diversion ofB. braunii metabolic activity towards hydrocarbon generation. It appears also that the improvements in hydrocarbon production, due to strain selection and to culture condition adjustment, obtained in free cultures, can be directly applied toB. braunii immobilized in alginate beads.     

Diffusivity of Cu2+ in calcium alginate gel beads: recalculation

Calculations of the diffusivity of Cu(2+) in calcium alginate gel beads using the shrinking core model were checked by us. Corrected results are reported here. Diffusivity was still found to increase with increasing alginate concentration, but at a lower rate than reported in the cited paper. The diffusivity increased by a factor of 2 over the range of alginate concentrations studied rather than 10. The original data is included with sample calculations. (c) 1994 John Wiley & Sons, Inc.     

Immobilization of Nicotiana tabacum plant cell suspensions within calcium alginate gel beads for the production of enhanced amounts of scopolin

Scopolin-producing cells of Nicotiana tabacum were immobilized within Ca-alginate gel beads. Free cell suspensions accumulated scopolin within cytoplasmic compartments and cell disruption was necessary to recover scopolin. On the contrary, immobilized plant cells excreted considerable amounts of scopolin. Scopolin diffused throughout the gel matrix and reached the culture media. A large fraction of produced scopolin could then be recovered from the culture medium without disrupting cells. Immobilized N. tabacum cells produced more scopolin than free cell suspensions did (3.8 mg/g fresh weight biomass [into the culture media] versus 0.2 mg/g fresh weight biomass [intracellular]). Variation of the immobilization conditions revealed a marked influence on the behavior of N. tabacum plant cells: production of scopolin and enhanced excretion, cell growth, and morphological aspect of plant cell colonies. This excretion phenomenon could be used advantageously at an industrial production level.     

Diffusion of Cu2+ in calcium alginate gel beads: further analyses

The diffusivity of Cu(2+), as determined by previous authors from analysis of experimental data in terms of the shrinking core (SCM) and linear absorption (LAM) models, is examined in light of the ability of the models to curve fit all the data. It is concluded from this further analysis that previous conclusions depicting the LAM to have an advantage over the SCM for predictive value are not justified. It is also shown that equally good curve fits can be obtained with a recent absorption/desorption model of diffusion which considers directly, through distribution theory, the effect of heterogeneity of material properties on the rate of diffusion. (c) 1995 John Wiley & Sons, Inc.     

Conjugal transfer of plasmid DNA between streptococci immobilized in calcium alginate gel beads

A rapid and simple technique was developed for conjugation between group N and group D streptococci by using cells entrapped within calcium alginate gel beads. With this method, the frequencies of transfer of lactose metabolism from Streptococcus lactis ME2 to S. lactis LM2302 were comparable to those achieved with agar surface matings. Conjugal transfer of the chloramphenicol and erythromycin resistance plasmid pVA797::Tn917 from S. faecalis V1229 to S. faecalis V1102 in alginate beads occurred at frequencies comparable to those achieved with filter matings. The results demonstrated efficient conjugal transfer of plasmid DNA among alginate-immobilized streptococcal cells and suggested that this method could be used as an alternative to conventional solid-surface and filter matings with these organisms.     

Conditions for immobilizing Streptomyces erythreus in a calcium alginate gel and the apparatus setup for the process

A laboratory unit for production of calcium alginate gel granules with immobilized microorganisms is described. It provides sterile production of particles from tens micrometers to 2 mm in diameter. Expediency of using biocatalysts in the form of fine granules is exemplified with a number of immobilized microorganisms. Conditions for immobilizing the erythromycin producing organism by its incorporation into the calcium alginate gel were studied. Viability of the actinomycete in the gel was shown by consumption of the nutrients and biosynthesis of the antibiotic.     

Determination of the radial distribution of Saccharomyces cerevisiae immobilised in calcium alginate gel beads

Summary The dissolution of alginate gel beads in 20 g sodium citrate /l produces a linear decrease in bead diameter. The rate of dissolution is dependent on the concentration of CaCl₂ within the gel beads. This method allows the controlled release of Saccharomyces cerevisiae from alginate gel beads and permits the simple and rapid determination of the radial distribution of cell concentration.     

Cytological and physiological behaviour of Euglena gracillis cells entrapped in a calcium alginate gel

The immobilisation of Euglena gracilis Z cells in a calcium alginate matrix maintained respiratory and photosynthetic activities and ultrastructural integrity. Moreover, immobilization did not prevent Euglena cells from greening inside the gel beads. Electron microscopy demonstrated that the immobilized cells were fixed in the same cellular state as they were when the immobilization occurred. This can he explained by simultaneous reaction of both Ca2⁺ and the alginate with the cells. Some hypotheses about the role of C2⁺ are discussed. In addition, long term storage (2 years) in calcium alginate has been performed permitting applications in algal storage.     

Conjugal transfer of plasmid DNA between streptococci immobilized in calcium alginate gel beads.

A rapid and simple technique was developed for conjugation between group N and group D streptococci by using cells entrapped within calcium alginate gel beads. With this method, the frequencies of transfer of lactose metabolism from Streptococcus lactis ME2 to S. lactis LM2302 were comparable to those achieved with agar surface matings. Conjugal transfer of the chloramphenicol and erythromycin resistance plasmid pVA797::Tn917 from S. faecalis V1229 to S. faecalis V1102 in alginate beads occurred at frequencies comparable to those achieved with filter matings. The results demonstrated efficient conjugal transfer of plasmid DNA among alginate-immobilized streptococcal cells and suggested that this method could be used as an alternative to conventional solid-surface and filter matings with these organisms.     

Decolorization of a kraft mill effluent with fungal mycelium immobilized in calcium alginate gel

A white-rot fungus Coriolus versicolor was immobilized by entrapment in calcium alginate beads. Treatment of a kraft mill effluent with the immobilized fungus in the presence of sucrose resulted in 80% loss of color of the effluent within 3 days. The minimal concentration of sucrose required for the decolorization was 10 mM. Other carbon sources (xylose, glucose, glycerol, and ethanol) could also be used.