Reactor configuration enzyme concentration

The discontinuous stirred tank reactor represents one of the most traditional reactor configurations for enzymatic reactions. It consists of a stirred tank where the enzyme, substrates, and cofactors are added at the beginning of the operation with no inlet and/or outlet stream during the reaction time. This type of reactor is usually considered to present an ideal hydrodynamic behavior therefore, the reactor is supposed to be completely mixed and the concentration of all... 

When macromolecular substrates are involved in the transformation under study, concentration polarization phenomena affect the EMR performance more severely. Diffusion limitations of macromolecular substrates hamper the use of immobilized enzymes in the hydrolysis of high-molecu-lar-weight substrates. By selecting membranes with an appropriate molecular weight cut-off, both enzyme and substrate are retained in an EMR in touch with each other, and hydrolysis products and/or inhibitors are continuously removed from the system. Soluble enzymes can then act directly on substrate macromolecules without diffusion limitations and steric hindrance imposed by enzyme fixation to a solid support. The stirring features of CST EMRs moreover assures that substrates and/or inhibitors within the reactor vessel are maintained at the lowest possible concentration level. Such reactor configuration is then extremely useful when substrate inhibited reaction patterns are involved, or when inhibiting species are assumed to exist in the feed stream. 

The operational conditions, that is, the concentration of substrate and enzyme, the temperature range, and the reactor configuration are summarized in Table 13.2. The activation energy of the reaction, E, was typically obtained for a batch reactor and compared with that calculated for a CSMR. The data obtained in the CSMR at steadystate enabled us, by using a semi-log plot of reaction rate versus time, to identify a first-order mechanism of enzyme deactivation and to determine both its first-order deactivation constant, kj, and the reaction rate at time zero, r, for each substrate and temperature. It was thus possible to compare the effect of the operational parameters on the activity and stability of these two enzymes. From the Arrhenius plot of these Tq, the E,-values were determined for each substrate, and were found to match the values obtained in the batch reactors. 

The stirred batch reactors are easy to operate and their configurations avoid temperature and concentration gradient (Table 5). These reactors are useful for hydrolysis reactions proceeding very slowly. After the end of the batch reaction, separation of the powdered enzyme support and the product from the reaction mixture can be accomplished by a simple centrifugation and/or filtration. Roffler et al. [114] investigated two-phase biocatalysis and described stirred-tank reactor coupled to a settler for extraction of product with direct solvent addition. This basic experimental setup can lead to a rather stable emulsion that needs a long settling time. 

When immobilized enzymes are employed in a continuous reactor, many of these limitations are avoided. Moreover, in this case the output signal is recorded at the reactor outlet, and this procedure therefore cannot affect the processes taking place in the reactor, and the signals obtained can be used in another system as actual concentrations without conversion. Yet, in this configuration the cofactor enters the reactor in the feed stream, which requires large amounts of cofactor, especially when a high flow rate is employed. 

Figure 27.18 Common configuration for postcolumn reactors with electrochemical analysis. (A) LC-chemical reaction-EC. Postcolumn addition of a chemical reagent (for example, Cu2+ or an enzyme). (B) LC-enzyme-LC. Electrochemical detection following postcolumn reaction with an immobilized enzyme or other catalyst (for example, dehydrogenase or choline esterase). (C) LC-EC-EC. Electrochemical generation of a derivatizing reagent. The response at the second electrode is proportional to analyte concentration (for example, production of Br2 for detection of thioethers). (D) LC-EC-EC. Electrochemical derivatization of an analyte. In this case a compound of a more favorable redox potential is produced and detected at the second electrode (for example, detection of reduced disulfides by the catalytic oxidation of Hg). (E) LC-hv-EC. Photochemical reaction of an analyte to produce a species that is electrochemically active (for example, detection of nitro compounds and phenylalanine). Various combinations of these five arrangements have also been used. [Reprinted with permission from Bioanalytical Systems, Inc.]...

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. 

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