Bioreactors immobilized enzymes/cells

Moo-Young, M., Bioreactor immobilized enzymes and cells fundamentals and appli-... 

Engineering Aspegts Trevan, M. D., Immobilized Enzymes An Introduction and Applications in Biotechnology, Wiley, 1980. Moo-Young, M., Bioreactors Immobilized Enzymes and Cells Fundamentals and Applications, Elsevier, London, 1988. 

Moo-Young, M. Bioreactor Immobilized Enzymes and Cells Eundamentals and Applications, Elsevier Science Publishing, Inc. New York, 1998. 

E. J. Vandamme, Immobilized Biocatalysts and Antibiotic Production Biochemical, Ge-netical and Biotechnical Aspects, in Bioreactors, Immobilized Enzymes and Cells (Ed. M. Moo-Young), Elsevier, 1988, pp. 

Itianes A, Zuniga ME, Chamy R et al. (1988) Immobilization of lactase and invertase on crossUnked chitin. In Moo-Young M (ed). Bioreactor immobilized enzymes and cells. Elsevier, London, pp 233-249... 

As most biochemical reactions occur in the liquid phase, bioreactors usually handle liquids. Processes in bioreactors often also involve a gas phase, as in cases of aerobic fermentors. Some bioreactors must handle particles, such as immobilized enzymes or cells, either suspended or fixed in a liquid phase. With regard to mass transfer, microbial or biological cells may be regarded as minute particles. 

In the design and operation of various bioreactors, a practical knowledge of physical transfer processes - that is, mass and heat transfer, as described in the relevant previous chapters - are often also required in addition to knowledge of the kinetics of biochemical reactions and of cell kinetics. Some basic concepts on the effects of diffusion inside the particles of catalysts, or of immobilized enzymes or cells, is provided in the following section. 

Bubble columns in which gas is bubbled through suspensions of solid particles in liquids are known as slurry bubble columns . These are widely used as reactors for a variety of chemical reactions, and also as bioreactors with suspensions of microbial cells or particles of immobilized enzymes. 

A bioreactor is a vessel in which biochemical transformation of reactants occurs by the action of biological agents such as organisms or in vitro cellular components such as enzymes. This type of reactor is widely used in food and fermentation industries, in waste treatment, and in many biomedical facilities. There are two broad categories of bioreactors fermentation and enzyme (cell-free) reactors. Depending on the process requirements (aerobic, anaerobic, solid state, immobilized), numerous subdivisions of this classification are possible (Moo-Young, 1986). 

In fermentation reactors, cell growth is promoted or maintained to produce metabolite, biomass, transformed substrate, or purified solvent. Systems based on macro-organism cultures are usually referred as tissue cultures. Those based on dispersed non-tissue forming cultures of micro-organisms are loosely referred as microbial reactors. In enzyme reactors, substrate transformation is promoted without the life-support system of whole cells. Frequently, these reactors employ immobilized enzymes, where an enzyme is supported on inert solids so that it can be reused in the process. Virtually all bioreactors of technological importance deal with a heterogeneous system involving more than two phases. 

Fumarase. The development and use of this immobilized enzyme by Tanabe Seiyaku for production of L-malic acid is very similar to that of aspartase ( 3). Lysed Brevibacterium ammoniagenes or B. flavin cells are treated with bile acid to destroy enzymatic activity which converts fumarate to succinate. As with aspartase, the cells can be immobilized in polyacrylamide or k-carrageenan gels. Using a substrate stream of 1 M sodium fumarate at pH 7.0 and 37°C, L-malic acid of high purity has been produced since 1974 by a continuous, automated process (3,39) for example, using a 1000-L fixed-bed bioreactor, 42.2 kg L-malic acid per hour was produced continuously for 6 months. 

Enzymes catalyze the reversible enantioselective addition of ammonia to a./ -unsaturated carboxylic acids to give L-a-amino acids. Immobilized enzymes and whole-cell bioreactor technology competes well with chemical methods31,32. For practical purposes, immobilized whole cells are preferred over immobilized enzyme because of the added cost of enzyme isolation. 

Konovalova VV, Dmytrenko GM, Nigmatullin RR, Bryk MT, and Gvozdyak PL Chromium(Vl) reduction in a membrane bioreactor with immobilized Pseudomonas cells. Enzyme Microbial Technol, 2003 33(7) 899-907. 

One can also envision applications involving the use of encapsulated or entrapped enzymes in bioreactors for therapeutic applications involving detoxification of deleterious substances or correction of metabolic deficiencies. In these applications, the enzymes could be contained within artificial cells [e.g., modified red blood cells (erythrocytes) or liposomes]. Liang, Li, and Yang have reviewed biomedical applications of immobilized enzyme bioreactors. 

The packed-bed reactor is a cylindrical, usually vertical, reaction vessel into which particles containing the catalyst or enzyme are packed. The reaction proceeds while the fluid containing reactants is passed through the packed bed. In the case of a packed-bed bioreactor, a liquid containing the substrate is passed through a bed of particles of immobilized enzyme or cells. 

For biotechnological applications, synthetic membranes entrapping enzymes, bacteria, or animal cells are used in membrane bioreactors disclosing new important developments mainly due to the increased stability of immobilized enzymes, the possibility of their continuous reuse and the absence of pollution of the products. Membrane bioreactors are of great interest as well for the possibility of continuously removing metabolites whose presence in the reaction environment could reduce the productivity of the reactor. 

Bioreactors with immobilized enzymes or whole cells are utilized in a variety of areas. EMR have found widespread applications in the hydrolysis of macromolecules - proteins, poly- and oligosaccharides, lipids, etc. - for food and pharmaceutical applications (Prazeres and Cabral, 1994). More recently they have also begun to find application for the removal of various pollutants from wastewaters. An overview of the literature in these fields is described below. 

The performance of immobilized reactors in continuous operation can be negatively influenced by several incidents such as enzyme/cell leakage, thermal denaturation of the enzyme, disintegration of the support, or microbial contamination. These parameters can be evaluated experimentally, and approaches can thus be designed in order to counter their negative effect on bioreactor performance. 

In airlift bioreactors the fluid volume of the vessel is divided into two interconnected zones by means of a baffle or draft-tube (Fig. 5). Only one of these zones is sparged with air or other gas. The sparged zone is known as the riser the zone that receives no gas is the downcomer (Fig. 5a-c). The bulk density of the gas-liquid dispersion in the gas-sparged riser tends to be less than the bulk density in the downcomer consequently, the dispersion flows up in the riser zone and downflow occurs in the downcomer. Sometimes the riser and the downcomer are two separate vertical pipes that are interconnected at the top and the bottom to form an external circulation loop (Fig. 5c). External-loop airlift reactors are less common in commercial processes compared to the internal-loop designs (Fig. 5a, b). The internal-loop configuration may be either a concentric draft-tube device or an split-cylinder (Fig. 5a, b). Airlift reactors have been successfully employed in nearly every kind of bioprocess—bacterial and yeast culture, fermentations of mycelial fungi, animal and plant cell culture, immobilized enzyme and cell biocatalysis, culture of microalgae, and wastewater treatment. 

Immobilized enzyme and cell particles may be used in packed bed bioreactors, or the particles may be suspended in stirred tanks, bubble columns, airlift bioreactors, and fluidized beds, as discussed in an earlier section of this... 

Other bioreactor configurations have been developed specifically for immobilized enzymes and cells. Enzymes immobilized within polymeric membranes are used in hollow fiber (Fig. 16) and spiral membrane bioreactors (Fig. 17). In the hollow fiber device, many fibers are held in a shell-and-tube configuration (Fig. 16) and the reactant solution (or feed) flows inside the hollow fibers. The permeate that has passed through the porous walls of the fibers is collected on the shell side and contains the product of the enzymatic reaction. Also, instead of being immobilized in the fiber wall, enzymes bound to a soluble inert polymer may be held in solution that flows inside the hollow fiber. The soluble product of the reaction then passes through the fiber wall and is collected on the shell side the enzyme molecule, sometimes linked to a soluble polymer, is too large to pass through the fiber wall. 

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