Ca-alginate Gel Beads Research Articles

Diffusion of (de)acylated antibiotic A40926 in alginate and carrageenan beads with or without cells and/or soybean meal.

Effective diffusion coefficients (D(e)) of antibiotic A40926 and its deacylated derivative were determined in Ca-alginate (2% wt/wt) and kappa-carrageenan (2.6% wt/wt) gel beads with or without immobilized Actinoplanes teichomyceticus cells and/or soybean meal (SBM). The method used was based on transient concentration changes in a well-stirred antibiotic solution in which gel beads, initially free of solute, were suspended. Unsteady-state diffusion in a sphere was applied and D(e) determined from the best fit of experimental data. A40926 showed markedly different diffusion characteristics than its deacylated derivative. Diffusivity of deacyl-A40926 in alginate or carrageenan gel beads was six to seven times that of A40926. Large differences in partition coefficients (Kp) were also found. In case of beads without additions, A40926, in contrast to deacyl-A40926, strongly partitioned to the liquid phase. Introduction of SBM and/or mycelium in the gel beads decreased the effective diffusivity of deacyl-A40926, but increased its partitioning to the solid phase. Our findings indicate that a relatively moderate structural change of a lipoglycopeptide molecule could lead to a major change in its diffusion/partition characteristics.

Volatile Compound Formation in White Wines Fermented Using Immobilized and Free Yeast

Using free yeast or yeast immobilized in double-layer Ca-alginate gel beads or strands, or single-layer beads or strands, white table wines were made from Koshu grape musts with initial pH between 2.80 and 3.70, containing 0 to 325 mg SO₂/L and 477 to 2178 mg total phenols gallic acid equivalent (GAE)/L, at fermentation temperatures ranging from 10 to 40°C. Yeast cells immobilized in beads could be added to and removed from fermenting vessels more easily than those in strands and markedly more easily than free yeast cells. Yeast immobilized in double-layer beads produced the most visibly clear wine, with the least leakage of viable cells from the gel matrices into the wine at the end of the fermentation process. Double-layer bead-immobilized yeast was also more tolerant than free yeast of high SO₂ and phenol concentrations, low pH, and high fermentation temperature. Fermentations with both immobilized and free yeast were negligibly influenced by differences in the initial pH. Lower concentrations of undesirable volatile compounds, such as methyl alcohol, ethyl acetate, and acetaldehyde, were observed in wines made using yeast immobilized in double-layer beads than in wines produced with free yeast. Concentrations of major higher alcohols (aroma compounds) varied with the use of either immobilized or free yeast, the SO₂ concentration in the must, and the fermentation temperature.

Cultures of Nicotiana tabacum Cells Immobilized in Calcium Alginate Gel Beads Coated with Cell-Free Gel Film.

Nicotiana tabacum cells were immobilized in the Ca-alginate gel beads coated with the cell-free gel film and then cultivated in the flask. The effects of the coating film thickness on intraparticle cell growth and cell leakage were investigated. The beads with the coating film of a desired thickness can be prepared by controlling the amount of free Ca2+ in the core beads. The optimum thickness for preventing cell leakage without inhibiting intraparticle cell growth was determined. The productivity of the phenolics, which was one of the secondary metabolites of tobacco cells, for the immobilization cultures was compared with that for the suspension culture. The phenolics production and release to the medium seemed to be promoted by the immobilization. The immobilization culture using the coated beads was very effective in promoting the extracellular productivity of the phenolics as well as preventing cell leakage. The immobilization method was applied to the culture system using an airlift bioreactor with a draft tube. Cell leakage was completely prevented throughout the 25-day cultivation without inhibiting cell growth.

Immobilisation of Solanum chrysotrichum plant cells withinCa-alginate gel beads to produce an antimycotic spirostanol saponin

Plant cell cultures of the Mexican species Solanum chrysotrichum can be established from friable calli. These cells produce an antifungal spirostanol saponin designated SC1. Plant cell immobilisation within Ca-alginate gel beads can lead to an enhancement in secondary metabolite production. Different matrices, obtained by varying calcium (0.1 to 0.8 mol·L –1) and alginate (1 to 1.5 % w/v) concentrations were used to study the influence of immobilisation parameters on the behaviour of S. chrysotrichum cells. Throughout the growth cycle and in the various immobilisation conditions, fresh and dry weight, SC1 yield and viability were measured. Important metabolic, morphological and physiological modulation seemed to be generated by the gel bead networks and by the cell-cell interactions inherent to the microenvironmental constraints. An accumulation of starch grains was observed through amyloplasts. Immobilised S. chrysotrichum cells accumulated 5 mg·g –1 dry weight of SC1. Although this was four times less than the free cells, the desired metabolite was excreted into the surrounding medium (around 40 % of the total production).

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.

Control of molecular weight cut-off for immunoisolation by multilayering glycol chitosan-alginate polyion complex on alginate-based microcapsules

Glycol chitosan is a positively charged polysaccharide which is water-soluble at pH 7.4, and is able to form a polyion complex (PIC) with anionic polymers, such as alginate. The authors attempt to develop a novel type of alginate-based microcapsule using this glycol chitosan for a islets-encapsulated bioartificial pancreas. The number of layers composed of glycol chitosan-alginate (GC-Alg) PIC were optimized, in order to cut off immunoglobulin transport and to protect encapsulated islets from the host immune reaction, and the transport characteristics were evaluated of glucose, bovine serum albumin (BSA) and gammaglobulin. To add mechanical stability to the microcapsule, calcium ions, which crosslinked the alginate polymers close to the interface between core Caalginate and multilayered membrane, were partially substituted with barium ions after the formation of multilayered Ca-alginate gel beads. The partition coefficients of BSA and gamma-globulin were decreased with the increasing number of layers. The immunoisolation was achieved againstgamma-globulin with four layers of the GC-Alg PIC membrane, while BSA could permate the membrane. The four-layered Ba-alginate gel bead had a good permeability for glucose, giving a diffusion coefficient corresponding to 80% of that in pure water. Insulin secretion from the islets in the four-layered Ba-alginate microcapsule was satisfactorily observed with the fractional stimulation ratio of 2.17. This result indicates that the encapsulated islets maintained their viability even after encapsulation. It was, thus, shown that the Ba-alginate microcapsule with four layers of the GC-Alg PIC membrane is promising as the microencapsulation material for a bioartificial pancreas.

Effect of protective solutes on leakage from and survival of immobilized lactobacillus subjected to drying, storage and rehydration

When lactic acid bacteria are used industrially as fermentation starters it is important to obtain stable and highly viable bacterial cultures. Six strains of Lactobacillus encapsulated in Ca-alginate gel beads were investigated to determine whether dehydration, storage and rehydration may inflict injury. A negative relationship between leakage of lactate dehydrogenase and survival rates was found. Mesophilic lactobacilli showed only negligible leakage compared with thermophilic strains when dehydrated at 30 degrees C to a level of 0.11 g H20 (g dry wt)-1. The choice of an appropriate suspending medium to be introduced before drying was therefore very important for thermophilic lactobacilli in order to increase the survival rates during dehydration, storage and rehydration. The osmoregulatory solutes tested were adonitol, betaine, glycerol and reconstituted non-fat milk solids (NFMS). Less injury was inflected during dehydration for Lactobacillus helveticus with adonitol, glycerol and NFMS. Survival rates for the strains subjected to immobilization, dehydration, storage and rehydration varied with the strain and the protective solute when fluidized-bed drying was used at 5 degrees C to a level as high as 0.34 g H20 (g dry wt)-1. Non-fat milk solids gave the best protection for thermophilic lactobacilli, while adonitol and NFMS were best for mesophilic lactobacilli.

Survival of Lactobacillus helveticus entrapped in Ca-alginate in relation to water content, storage and rehydration.

Lactobacillus helveticus CNRZ 303 entrapped in Ca-alginate gel beads was investigated for improved survival and stability during fluidized-bed drying, storage and rehydration. Addition of protective solutes was very important. Studies of the conditions showed that inactivation of entrapped L. helveticus started when the water content exceeded 0.3-0.4 g H2O (g dry wt)-1 for adonitol, glycerol and reconstituted non fat milk solids (NFSM). With Ringer's solution (control) and betaine, the fall in viability was evident above 1 g H2O (g dry wt)-1. Drying down to 0.2 g H2O (g dry wt)-1 required the removal of 98.5-98.9% of the water. The best survival rate with the least injured cells among survivors was experienced with adonitol and NFMS, respectively, 71% and 57% (compared to the initial) immediately after dehydration. Adonitol and NFMS were also best for survival during storage. The highest cell recovery was obtained by rehydrating the cells in cheese whey permeate between 20-30 degrees C done at pH 6.0-7.0, satisfying the demands for cell survival, repair and slow swelling (adaptions).

Optimizing lipase activity, enantioselectivity, and stability with medium engineering and immobilization for beta-blocker synthesis.

Lipase from Pseudomonas cepacia showed poor activity and moderate enantioselectivity (E) in pure aqueous systems for hydrolysis of a racemic mixture (+/-)-1-chloro-2-acetoxy-3-(1-naphthyloxy)-propane, which is a potential intermediate for beta-blocker synthesis. However, addition of polar organic solvents to the reaction can change both the activity and the enantioselectivity for this chiral reaction significantly. It was observed, in general, that the activity increases and the enantioselectivity decreases with the increase in the polarity of the organic solvent added to the medium. Among the six solvents chosen (i.e., dimethylsulfoxide [DMSO], 1, 4-dioxane, dimethylformamide [DMF], acetone, 1-propanol, and tetrahydrofuran [THF]), maximum activity and minimum enantioselectivity was obtained with DMSO, whereas minimum activity and maximum enantioselectivity was obtained with THF as the cosolvents. In the subsequent studies, native or polyethylene glycol (PEG)-modified lipase was immobilized by entrapping in Caalginate gel beads. In a fixed-bed continuous reactor containing these catalyst beads, the enzyme was found to be at least three times more enantioselective than the native form in a batch reactor. This fixed-bed reactor with the beads could be operated with high concentration of acetone (33% v/v) for about 1 month without a significant loss of enzyme activity and enantioselectivity.

DEVELOPMENT, CHARACTERIZATION AND IN VIVO EVALUATION OF POLYELECTROLYTE COMPLEX MEMBRANE GEL MICROCAPSULES CONTAINING MELATONIN-RESIN COMPLEX FOR ORAL USE

A novel oral multiple-unit delivery system for melatonin, (Ca) alginate/polyethyleneimine (PEI) gel microcapsules containing melatonin-resin (Dowex 1-X8 Cl-) complex, which overcome many of the problems encountered with the conventional dosage forms, Ca-alginate gel beads was designed through an ionic crosslinking/interpolyelectrolyte complexation technique. The effect of different processing variables viz., gelation time, PEI concentration, melatonin/resin ratio, core/coat ratio, pH and ionic strength of the dissolution medium on the release characteristics of microcapsules in comparison with alginate beads was investigated. The results obtained indicated that a PEI concentration of 2% and gelation time of 24 h. were the optimal conditions for a marked release retardation. Of ultimate importance is the ionic character of the synthetic polyelectrolyte complex (PEC) membrane (alginate/ PEI) that produced microcapsules which did not disintegrate in simulated intestinal fluid (S.I.F., pH 7.4) over a period of 3 days of testing and allowed a pH-independent release rate of melatonin up to 2 h. in phosphate buffer (0.154 M Na+ ions) (K1 = 5.42x10-1, 5.995x10-1 and 5.78x10-1 hr-1 for pHs of 5.5, 7.4 and 9.0, respectively). The metatonin release rate of microcapsules (K1= 1.144z`0-1 hr-1) in S.I.F. (pH 7.4) was significantly reduced when compared to that of the original alginate beads (K1= 19.59x10-1 hr-1) which showed a dramatic disintegration and dissolution in S.I.F. (pH 7.4) within 1 h. Most importantly, successful attempts to prepared crosslinked protein or polyamide/PEI-graft co-polymer membranes-coated alginate microcapsules were achieved by making use of a modified interfacial polymerization process. The coated microcapsules offered a remarkable reduction of the release rate in simulated gastric fluid (S.G.F., pH 1.2) as compared to alginate beads. The modulatory influence of melatonin (biological half-life = 24 min) on liver detoxifying and antioxidant enzyme systems (glutathione-S-transferases (GSTs) and glutathione peroxidase (GSH-Px), as well as lipid peroxides (LPER) levels was studied in rats. The obtained results revealed that melatonin resonates-loaded (Ca) alginate/PEI gel microcapsules (T50% = 6.06 h) produced a significant marked elevation in hepatic cytosolic activities levels of GSTs and GSH-Px as well as glutathione (GSH) content with a reduction in (LPER) levels, when compared with the corresponding controls, suggesting that alginate/PEI gel microcapsules could be a useful reservoir system for controlled-release melatonin delivery.

Mathematical description of ethanol fermentation by immobilised Saccharomyces cerevisiae

Fermentation characteristics of Saccharomyces cerevisiae ATCC 9763 immobilised in Ca-alginate gel beads have been investigated in a stirred batch system at 2, 4, 8, 10% (w/v) initial glucose concentrations. The experimental results were tested using eleven different kinetic models relating biomass and ethanol production and glucose utilisation in different forms. The models proposed by Monod and Hinshelwood were found to be more appropriate for describing the batch growth and ethanol production of immobilised S. cerevisiae at low (2–4%) and high (8–10%) initial glucose concentrations, respectively. The validation of the models chosen was done using data obtained from experiments in an inclined reactor with nutrient recirculation.

Enhanced production of scopolin by Solanum aviculare cells immobilised within Ca-alginate gel beads

Enhanced production of scopolin by Solanum aviculare cells immobilised within Ca-alginate gel beads

Enhanced production of scopolin bySolanum aviculare cells immobilised within Ca-alginate gel beads.

Different matrices, obtained by varying calcium (0.1 to 1.5M) and alginate (1 to 1.5%) concentrations, were used to study the influence of immobilisation parameters on the behaviour ofS. aviculare. A significant modulation of cell growth, cell release, and scopolin production and excretion has been observed. Physiological and morphological characteristics ofSolanum aviculare cells immobilised within Ca-alginate beads were notably different from those of suspended cells. ImmobilisedS. aviculare have accumulated scopolin (up to 120 μg·g-1 FWB) within beads and excreted it into the culture medium (up to 8 μg·g-1 FWB). Contrary to suspended cells which have accumulated only traces of this metabolite within intracellular compartments (1 μg·g-1 FWB), no scopolin has been found into the culture medium.

The effective diffusion coefficient and the distribution constant for small molecules in calcium‐alginate gel beads

The effective diffusion coefficient, D(e), and the distribution constant, K(i), for selected mono- and disaccharides and organic acids were determined in homogeneous calcium-alginate gel with and without entrapped bacteria. Results were obtained from transient concentration changes in well-stirred solutions of limited volume, in which the gel beads were suspended. The effective diffusioncoefficients and the distribution constants were estimated by fitting mathematical model predictions to the experimental data using a nonlinear model fitting program (MODFIT). Both single solute diffusion and multiple solute diffusion were performed. A small positive effect was obtained onthe values of D(e) for the system of multiple solute diffusion; however, the values of K(i) were not significantly influenced. For the nine solutes tested, D(e) for 2% Ca-alginate gel beads was found to be approximately 85% of the diffusivity measured in water. The effects on D(e) and K(i), for lactose and lactic acid were determined for variations of alginate concentration, pH, temperature, and biomass content in the beads. D(e) decreased linearly for both lactose and lactic acid with increasing cell concentration in the Ca-alginate gel. K(i), was constant for both lactose and lactic acid with increasing cell concentration. D(e) was significantly lower at pH 4.5 than at pH 5.5 and 6.5 for both lactose and lactic acid. Furthermore, D(e) seemed to decrease with increased alginate concentration in the range of 1% to 4%. The diffusion rate increased with increasing temperature, and the activation energy for the diffusion process for both lactose and lactic acid was constant in the temperature range tested. (c) 1995 John Wiley & Sons Inc.

Effects of alginate composition and gelling conditions on diffusional and mechanical properties of calcium-alginate gel beads.

Dependencies of effective diffusivity of glucose in Ca-alginate gel beads and mechanical compression strength of the gel on alginate composition, viscosity, concentration, temperature and pH of the alginate solution were systematically investigated. Diffusional and mechanical properties of Ca-alginate gel beads depended on alginate composition and the gelling conditions. The results were discussed in relation to get structure and conformation of alginate molecule. Differences in reported diffusivities of glucose could be explained by the differences in alginate composition and the gelling conditions such as alginate concentration, temperature and pH. Furthermore, selection guidelines for preparation of Ca-alginate gel having low diffusional obstruction and large compression strength were presented.

Efficient production of celery embryos and plantlets released in culture of immobilized gel beads

Celery embryos and plantlets were found to be selectively released in a culture of immobilized Ca-alginate gel beads in which celery callus was entrapped under regeneration conditions. We studied the feasibility of use of this process for celery embryogenesis in an artificial seed system. The cells released from the gel beads were larger than those obtained in suspension culture. The optimal concentration of alginate gel for embryo and plantlet production was 2% for the immobilized cell culture. Considering the maintenance of the gel bead structure and detrimental effect of CaCl 2 on plantlet development, 5 mM CaCl 2 supplementation gave the best result in terms of the number of heart and torpedo embryos and plantlets. The ratio of the number of heart embryos, torpedo embryos and plantlets to total number of cells in the immobilized cell culture was higher than that in the suspension culture. Repeated batch culture with 5 mM CaCl 2 provided long-term (more than 154 d) embryo and plantlet production without gel beads disruption. Productivity of plantlets in the immobilized cell culture with 5 mM CaCl 2 was 2.2-fold as high as that in the suspension culture.

A New Kind of Immobilized Lipase in Organic Solvent and Its Structure Model

In this paper, we used Ca-alginate gel beads coated with polyetheneimine and glutaraldehyde to adsorb Expansum penicillium lipase. The immobilized lipase catalyzed esterification of 1-dodecanol with dodecanoic acid in benzene. The results show that when the concentration of Ca-alginate, polyetheneimine (PEI) and glutaraldehyde is 1%, 6% and 1%, respectively, the activity of the immobilized lipase and the amount of adsorbed protein are the highest. The immobilized lipase is better than the SDS-immobilized lipase. The activity of the immobilized lipase connected by glutaraldehyde is higher than the activity of that without glutaraldehyde. The initial rate of the immobilized lipase and lyophilized lipase powder is 5.9 × 102 nmol/min.mgpr and 2.8 × 101 nmol/min.mgpr, respectively. After the immobilized lipase catalyzed the esterification reaction at 37°C for about 12 hours, 93.3% of 1-dodecanol was converted to ester, but for lyophilized lipase powder, only 17.5% converted. Based on all above results, we have presumed and explained the structure of this kind of immobilized lipase.

Imipramine Release from Ca-Alginate Gel Beads.

Ca-alginate gel beads loaded with a model cationic drug imipramine were prepared, and their drug release characteristics were examiend. By soaking plain Ca-alginate gel beads in an imipramine solution, the slightly translucent beads were transformed into white beads, seemingly due to the precipitation of a drug-alginate complex. The amount of drug uptake in the beads increased linearly as the initial drug concentration increased (1-10mg/ml). However, no significant difference in uptake efficiency was observed between the beads prepared over various periods of curing time. The drug release rate from the beads was in the following order : JP XII disintegration test solution No. 1 (pH 1.2)>0.9% (w/v) NaCl≒test solution No. 2 (pH 6.8)>distilled water. The release rates were also measured in HCl solutions, NaCl solutions, and acetate buffers with varying pHs. It was suggested that imipramine ions interacting with the acidic residues of alginates were replaced by cations in the release medium and diffused out of the beads. Therefore, Ca-alginate gel could be a useful vehicle for the controlled release of water-soluble cationic drugs.

Immobilization and culture of coffee (Coffea arabica L.) cells on novel culture systems using a basket-shaped unit, “EGSTAR”

A simple and new basket-shaped unit for agitation made of stainless steel (EGSTAR), in which immobilized coffee cells in Ca-alginate gel beads were packed, was placed in a jar fermentor (System-1). This system allowed the plant cells to grow submersed in the unit even at high agitation speed (650 rpm). Only a small amount of cells existed out of the EGSTAR. Most of the purine alkaloids produced were released into the medium. Suspended coffee cells in the jar fermentor were also possibly immobilized onto a polyurethane foam sheet fixed inside the net of the EGSTAR (System-2). The total cells in System-2 biotransformed theobromine to caffeine (77.9%). Other plant cell suspensions were also immobilized as efficiently as were the coffee cells in this system. Thus, System-2 is a simple and convenient system for immobilization of plant cells to produce secondary metabolites.

Cream fermentation by a mixed culture of lactococci entrapped in two-layer calcium alginate gel beads

This investigation was directed towards the development of a process which produces a fermented cream of greatly reduced cell number.Lactococcus lactis subsp.Lactis andLactococcus lactis subsp.lactis biovardiacetylactis were entrapped separately in normal or two-layer Ca-alginate gel beads. Pasteurized cream (31% fat content) was inoculated with free-cells and with normal or two-layer beads. When 8% of the total volume was occupied by the gel, there was 300–800 times more inoculum in this system and the fermentation time was considerably reduced (5h against 18h). When pH 5.0 was reached, the residual free-cell count was 150 and 1800 times less than for a classical inoculation method with free-cells for normal and two-layer beads respectively. This result was reproducible for several consecutive runs. Also, the problems linked to storage acidification (souring, wheying-off) appeared later and living lactic acid bacteria were maintained in the product.