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A novel continuous two-phase dispersion process was developed to produce κ-carrageenan gel microspheres, using static mixers. It was shown that yeast-loaded carrageenan beads, with controlled diameter and tight size distribution, can be produced on a continuous basis, in a scalable mixer, at production rates appropriate to both pilot plant-scale and, potentially, industrial-scale operations. Immobilized yeast are intended to be used in continuous brewing operations. The effects of the static mixer diameter (D), the number of mixing elements (N e ), the fluid linear velocity (V), and the volumetric fraction (e) of κ-carrageenan, on the mean diameter and size distribution of the resulting gel microspheres, were studied. Image analysis showed that mean diameter was strongly influenced by the average linear fluid velocity through the mixer, and by the mixer diameter. The number of mixer elements and the mixer diameter governed bead size dispersion. A productivity of 10 L h -1 of beads was attained using a 1.27-cm-diameter static miter. Because the productivity is proportional to the mixer diameter squared, this process, although suited for the production of small-size beads (down to 50 μm), would be technically and economically feasible for a large industrial immobilization process. However, because the coefficient of variability increased with mixer diameter, and thus with scale-up, operational improvements are suggested, such as the use of smaller-diameter mixers operating in parallel, to reduce the size dispersion.

Microcapsules are produced by a dropwise addition of one solution into a solidifying bath. By applying an electrostatic potential between the droplet formation device and the collecting solution, it is possible to obtain smaller droplets which are desirable for many applications. Droplet formation may be divided into two phases. Under a certain critical value of the electric potential Uc, liquid exits the nozzle as droplets. The surface tension decreases with increasing electric potential resulting in a reduction of droplet diameter to approximately 200 μm. At higher electric potential, liquid exits the nozzle as a jet which subsequently breaks into droplets, which are smaller than 200 μm. In this case, droplet size is mainly determined by the jet instability (theory of Rayleigh).

Smooth and spherical alginate microspheres and nylon-membrane bound microcapsules were formed in an air-agitated, liquid-liquid mixer by emulsification/internal gelation and interfacial polymerization respectively. The mean diameter of the alginate microspheres ranged from 100 to 800 microm, and was controlled by process modifications. Increase in emulsifier concentration, gas flowrate, and emulsification time resulted in smaller microsphere size as did a decrease in liquid height. Increase in the dispersed phase viscosity resulted in a longer emulsification time required for approaching a minimum microsphere size. Microspheres could be formed with the proportion of dispersed phase approaching 30%. The yield of alginate microspheres was 70%, with losses attributed to incomplete recovery during washing and filtration operations. The yield of DNA encapsulation within the fraction of recovered microspheres, was 94%. The small loss was thought to occur by surface release during the washing of the microspheres. (c) 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 56: 464-470, 1997.

The use of natural pigments such as lobster carotenoids in fish feed formulations offers advantages over the use of the synthetic alternatives. Microencapsulation of the pigments, with or without the addition of antioxidants to the formulation, may be of benefit in terms of stabilizing pigment colour. In the present study, lobster carotenoids were extracted from lobster shell into petroleum ether and microencapsulated by phase separation and salt coacervation within (poly vinyl alcohol) and poly(vinyl alcohol)/poly(D,L-lactic acid) membranes. Spherical microcapsules, with smooth, thin and resilient membranes were obtained with mean diameters ranging from 50 to 150 microns, depending on the membrane material, and source of pigment. The microcapsules were pink-orange in colour, and colour stability was followed spectrophotometrically. Enhanced stability was observed in both membrane materials, in comparison to the non-encapsulated control. Rates of discoloration were determined under a variety of storage conditions, including the absence of light, reduced temperatures and under nitrogen atmosphere. The best stability of lobster carotenoids was observed under a nitrogen atmosphere within PVA/PLA membranes, representing an 11-fold enhancement of pigment stability in comparison to the controls. Under ambient conditions, the enhancement in pigment stability was approximately 6-fold. The optimum concentration of PVA during microencapsulation was 3-4%, and the microencapsulated pigments appeared most stable under acidic conditions. The rate of discoloration appeared independent of pigment concentration.

Encapsulation of therapeutic peptides and proteins into polymeric micro and nanoparticulates has been proposed as a strategy to overcome limitations to oral protein administration. Particles having diameter less than 5 µm are able to be taken up by the M cells of Peyer's patches found in intestinal mucosa. Current formulation methodologies involve organic solvents and several time consuming steps. In this study, spray drying was investigated to produce protein loaded micro/nanoparticles, as it offers the potential for single step operation, producing dry active-loaded particles within the micro to nano-range. Spherical, smooth surfaced particles were produced from alginate/protein feed solutions. The effect of operational parameters on particle properties such as recovery, residual activity and particle size was studied using subtilisin as model protein. Particle recovery depended on the inlet temperature of the drying air, and mean particle size ranged from 2.2 to 4.5 µm, affected by the feed rate and the alginate concentration in the feed solution. Increase in alginate:protein ratio increased protein stability. Presence of 0.2 g trehalose/g particle increased the residual activity up to 90%. Glycol-chitosan-Ca(2+)alginate particles were produced in a single step operation, with resulting mean diameter of 3.5 μm. Particles showed fluorescein isothiocyanate labeled bovine serum albumin (BSA)-protein entrapment with increasing concentration toward the particle surface. Similar, limited release profiles of BSA, subtilisin and lysozyme were observed in gastric simulation, with ultimate full release of the proteins in gastrointestinal simulation.

Smooth and spherical alginate microspheres and nylon-membrane bound microcapsules were formed in an air-agitated, liquid-liquid mixer by emulsification/internal gelation and interfacial polymerization respectively. The mean diameter of the alginate microspheres ranged from 100 to 800 microm, and was controlled by process modifications. Increase in emulsifier concentration, gas flowrate, and emulsification time resulted in smaller microsphere size as did a decrease in liquid height. Increase in the dispersed phase viscosity resulted in a longer emulsification time required for approaching a minimum microsphere size. Microspheres could be formed with the proportion of dispersed phase approaching 30%. The yield of alginate microspheres was 70%, with losses attributed to incomplete recovery during washing and filtration operations. The yield of DNA encapsulation within the fraction of recovered microspheres, was 94%. The small loss was thought to occur by surface release during the washing of the microspheres. (c) 1997 John WileySons, Inc. Biotechnol Bioeng 56: 464-470, 1997.

Co-guanidine membranes were shown to form intact, ionically complexed membranes on alginate beads, serving as an alternative to the commonly used polymers, poly-L-lysine and chitosan. DNA was encapsulated and membrane thickness, the level of DNA protection from nuclease diffusion and the degree of DNA-complexation with co-guanidine membranes were all shown to be dependent on both polymer concentration and coating time. The highest level of DNAse exclusion was possible within beads coated with a polymer concentration of 5 mg/ml. Recovery of double-stranded DNA after nuclease exposure for 60 min reached 90% of that initially encapsulated. The molecular weight cut-off for these co-guanidine membranes was approximately 31 kDa, sufficient to exclude extracapsular nuclease. The level of DNA protection was found to be comparable to high molecular weight poly-L-lysine membranes (197.1 kDa). Intracapsular DNA was accessible to the carcinogen ethidium bromide, which showed a 4-fold increase in uptake in uncoated beads and 2-fold uptake in co-guanidine coated beads compared to beads lacking in DNA. Co-guanidine membranes coating alginate result in a molecular weight cut-off sufficient to retain DNA and exclude 31 kDa DNAse, while providing access to the low molecular weight carcinogen, ethidium bromide.

Alginate gels produced by an external or internal gelation technique were studied so as to determine the optimal bead matrix within which DNA can be immobilized for in vivo application. Alginates were characterized for guluronic/mannuronic acid (G/M) content and average molecular weight using 1H-NMR and LALLS analysis, respectively. Nonhomogeneous calcium, alginate, and DNA distributions were found within gels made by the external gelation method because of the external calcium source used. In contrast, the internal gelation method produces more uniform gels. Sodium was determined to exchange for calcium ions at a ratio of 2:1 and the levels of calcium complexation with alginate appears related to bead strength and integrity. The encapsulation yield of double-stranded DNA was over 97% and 80%, respectively, for beads formed using external and internal calcium gelation methods, regardless of the composition of alginate. Homogeneous gels formed by internal gelation absorbed half as much DNAse as compared with heterogeneous gels formed by external gelation. Testing of bead weight changes during formation, storage, and simulated gastrointestinal (GI) conditions (pH 1.2 and 7.0) showed that high alginate concentration, high G content, and homogeneous gels (internal gelation) result in the lowest bead shrinkage and alginate leakage. These characteristics appear best suited for stabilizing DNA during GI transit.

The use of natural pigments such as lobster carotenoids in fish feed formulations offers advantages over the use of the synthetic alternatives. Microencapsulation of the pigments, with or without the addition of antioxidants to the formulation, may be of benefit in terms of stabilizing pigment colour. In the present study, lobster carotenoids were extracted from lobster shell into petroleum ether and microencapsulated by phase separation and salt coacervation within (poly vinyl alcohol) and poly(vinyl alcohol)/poly(D,L-lactic acid) membranes. Spherical microcapsules, with smooth, thin and resilient membranes were obtained with mean diameters ranging from 50 to 150 microns, depending on the membrane material, and source of pigment. The microcapsules were pink-orange in colour, and colour stability was followed spectrophotometrically. Enhanced stability was observed in both membrane materials, in comparison to the non-encapsulated control. Rates of discoloration were determined under a variety of storage conditions, including the absence of light, reduced temperatures and under nitrogen atmosphere. The best stability of lobster carotenoids was observed under a nitrogen atmosphere within PVA/PLA membranes, representing an 11-fold enhancement of pigment stability in comparison to the controls. Under ambient conditions, the enhancement in pigment stability was approximately 6-fold. The optimum concentration of PVA during microencapsulation was 3-4%, and the microencapsulated pigments appeared most stable under acidic conditions. The rate of discoloration appeared independent of pigment concentration.

The factors which influence the steady state performance of a CSTR operation with microencapsulated urease for the regeneration of a dialysated solution have been studied at various enzyme activities. The theoretical model considered the effect of microcapsule diameter, pH-dependent kinetics, and product inhibition and substrate depletion, in relation to urea conversion and the capsule effectiveness factor. The limiting effects of pH, product inhibition and substrate depletion were also studied individually and in combination under eight case studies. The base case which included these three limiting factors in the reaction process, predicted the lowest urea conversion values at the enzyme activities considered. However, the effectiveness factor for each case at a fixed microcapsule diameter depended on the enzyme activity. This behaviour was studied for two different microcapsule diameters, 5 microns and 500 microns. At enzyme activities lower than 1 mM/s, a model considering Michaelis-Menten kinetics alone, predicted the highest effectiveness factors. On the other hand, beyond 1 mM/s enzyme activity, the lowest effectiveness factors were predicted, although higher conversions than that of the base case were achieved. This might be due to the rapid depletion of substrate at high activities when Michaelis-Menten kinetics were considered, leads to residual substrate concentrations (Sr) lower than the Michaelis-Menten constant Km,o; a condition for a dramatic drop in the intraparticle reaction rate. The limiting factors in the base case held Sr at a relatively higher value than Km,o.

A 91.5% mass yield of urease and hemoglobin (Hb), co-encapsulated within polyamide membranes, was determined spectrophoto-metrically. The specific activity yield of microencapsulation was 84%, twofold higher than values previously reported, as a result of optimization of encapsulation conditions. The kinetic parameters and pH activity profiles of intracapsular urease were determined to be similar to those corresponding to the free enzyme. Similar activities were also observed for intact and microcapsule homogenate, indicating minimal mass transfer and diffusional limitation. The active configuration of the enzyme appears to remain intact upon microencapsulation. The application of a kinetic model for encapsulated urease further indicated that the kinetics were reaction-controlled with minimal mass transfer restrictions.

Rhizopus arrhizus biomass was found to absorb a variety of different metal cations and anions but did not absorb alkali metal ions. The amount of uptake of the cations was directly related to ionic radii of La 3+ , Mn 2+ , Cu 2+ , Zn 2+ , Cd 2+ , Ba 2+ , Hg 2+ , Pb 2+ , UO 2 2+ , and Ag + . The uptake of all the cations is consistent with absorption of the metals by sites in the biomass containing phosphate, carboxylate, and other functional groups. The uptake of the molybdate and vanadate anions was strongly pH dependent, and it is proposed that the uptake mechanism involves electrostatic attraction to positively charged functional groups.

The mannoprotein which is a major component of the cell wall of Saccharomyces cerevisiae is an effective bioemulsifier. Mannoprotein emulsifier was extracted in a high yield from whole cells of fresh bakers' yeast by two methods, by autoclaving in neutral citrate buffer and by digestion with Zymolase (Miles Laboratories; Toronto, Ontario, Canada), a beta-1,3-glucanase. Heat-extracted emulsifier was purified by ultrafiltration and contained approximately 44% carbohydrate (mannose) and 17% protein. Treatment of the emulsifier with protease eliminated emulsification. Kerosene-in-water emulsions were stabilized over a broad range of conditions, from pH 2 to 11, with up to 5% sodium chloride or up to 50% ethanol in the aqueous phase. In the presence of a low concentration of various solutes, emulsions were stable to three cycles of freezing and thawing. An emulsifying agent was extracted from each species or strain of yeast tested, including 13 species of genera other than Saccharomyces. Spent yeast from the manufacture of beer and wine was demonstrated to be a possible source for the large-scale production of this bioemulsifier.

Calculation of a “theoretical yield” of microbial process is often a subject of controversy. Theoretical yield values of the solvents (butanol-acetone-ethanol) produced in the cultivation of Clostridium acetobutylicum on glucose have been calculated for 30 different culture conditions. Two different approaches were taken based on expressing the stoichiometric relationship between the substrate and the products of the process. The maximum theoretical yield under acceptable conditions was established ranging from 38.6% to 39.9%. It was considered for an ideal biosynthetic situation when no intermediate acids were left over in the system and no carbon was utilized in the production of biomass. The values of the solvent yield are dependent on the ratio between the solvent products. The coefficients of the process stoichiometric relationship and the ratios between hydrogen gas and butanol are presented for each set of process conditions. A three-dimensional plot of the yield versus the weight fractions of butanol and ethanol in the system has been developed reflecting the continuous variations of this parameter with the solvent ratio.

Bacillus licheniformis produced a water‐insoluble levan which has potential application as a selective plugging agent in microbial enhanced oil recovery (MEOR). The microorganism grew on sucrose, glucose, and fructose but produced levan only on sucrose. Plugging may thus be selectively controlled in the reservoir by substrate manipulation. B. licheniformis and a crude preparation of its extracellular enzymes were evaluated for their ability to produce levan under reservoir conditions. Oil reservoirs which have a temperature of less than 55°C, a pH between 6 and 9, a pressure less than 500 atm, and a salt concentration of 4% or less are potentially suitable. Examples of such reservoir conditions are found in Lloydminster on the Alberta‐Saskatchewan border, one of the largest Canadian oil reserves.

Acinetobacter calcoaceticus was grown in 11-liter batch fermentations with hexadecane or sodium citrate as the sole source of carbon. Surface and interfacial tension measurements of the microbial broth indicated that surface-active compounds were being produced only during growth on the hydrocarbon substrate. Contact angle measurements of an aqueous drop on a smooth lawn of cells in a hexadecane bath indicated a highly hydrophobic surface of the cells in the initial stages of the hydrocarbon fermentation (120° contact angle). At this stage, the entire cell population was bound to the hydrocarbon-aqueous interface. The contact angle dropped rapidly to approximately 45° after 14 h into the fermentation. This coincided with a shift of the cell population to the aqueous phase. Thus, the cells demonstrated more hydrophilic characteristics in the later stages of the fermentation. Contact angles on cells grown on sodium citrate ranged from 18 to 24° throughout the fermentation. The cells appear to be highly hydrophilic during growth on a soluble substrate. From the contact angle and aqueous-hydrocarbon interfacial tension, the surface free energy of the cells was calculated along with the cell-aqueous and cell-hydrocarbon interfacial tension. The results of these measurements were useful in quantitatively evaluating the hydrophobic nature of the cell surface during growth on hydrocarbons and comparing it with the hydrophilic nature of the cell surface during growth on a soluble substrate.

La biosorption est la propriete que possedent certaines biomasses microbiennes de capter les metaux lourds et les actinides. Le genre Rhizopus capte l'uranium et le thorium a 20% environ de son poids sec ce qui en fait un biosorbant interessant