Batch dialysis

Figure 2.5 Possible technological solutions to bioprocess problems a) Fed-batch culture b) Continuous product removal (eg dialysis, vacuum fermentation, solvent extraction, ion exchange etc) c) Two-phase system combined with extractive fermentation (liquid-impelled loop reactor) d) Continuous culture, internal multi-stage reactor e) Continuous culture, dual-stream multi-stage reactor f) Continuous culture with biomass feedback (cell recycling). (See text for further details).

Reversible inhibition ceases when the inhibiting molecule is removed from the system. The molecules can be eUminated from the feed in a flow system or from a batch reaction by a separation process such as dialysis. Two kinds of reversible inhibition are distinguished. Competitive inhibition occurs when an inhibitor molecule occupies a site before it is occupied by a substrate molecule. The assumed mechanism is... 

In the dialyzed batch start-up phase and the subsequent continuous operation a substantial increase in viable cell density and monoclonal antibody (MAb) titer was observed compared to a conventional suspension culture. The raw data, profiles of the viable cell density, viability and monoclonal antibody titer during the batch start-up and the continuous operation with a dialysis flow rate of 5 L/d are shown in Figures 17.6 and 17.7. The raw data are also available in tabular form in the corresponding input file for the FORTRAN program on data smoothing for short cut methods provided with the enclosed CD. 

Figure 17.6 Dialyzed Chemostat Monoclonal antibody concentration (raw and smoothed measurements) during initial batch start-up and subsequent dialyzed continuous operation with a dialysis flow rate of 5 L/d. [reprinted from the Journal of Biotechnology Bioengineering with permission from J. Wiley],...

The operation of the bioreactor was in the batch mode up to time t=2 2 h. The dialysis flow rate was kept at 2 Ud up to time t=91.5 h when a sharp drop in the viability was observed. In order to increase further the viable cell density, the dialysis flow rate was increased to 4 L/d and at 180 h it was further increased to 5 L/d and kept at this value for the rest of the experiment. 

Techniques of Purification. The purification techniques have included column and batch adsorption, extraction, dialysis, and column and paper chromatography. Liberal use was made of the fact that the gibberellate anion is not soluble in ethyl acetate, whereas the free acid is soluble, and that charcoal will adsorb GA3 from aqueous solutions but release it with acetone (26). As a rough rule, preliminary concentration by a factor of about 105 was necessary before significant use of paper chromatography could be made. For kudzu vine and pinto bean, a known amount of GA3 was added to an aliquot of the plant extract and taken through the same procedure as the initial extract. These controls are subsequently referred to as "spiked extracts, to differentiate them from the initial or "natural extract. 

Following its introduction in the early 1970s, the dialysis method has since developed into a useful method of preparing relatively large batches of liposomes for clinical use (Schwendener, 1986). 

Dissolve the mRNA pellet in 28 pL of Milli-Q water for batch, bilayer, or dialysis mode of translation. 

Either batch or continuous dialysis can be applied in the same manner [445],... 

Passive membrane dialysis is usually applied batch-wise, since its driving-force is the difference in gradient concentration between the two solutions separated by the membrane. In this case, the solute (reactants and products small molecules) from a hypertonic solution (the resulting solution of the catalytic reaction) permeates through the membrane to the hypotonic side (pure solvent) until equilibrium has been achieved, whereas the nanosized catalyst remains confined inside the membrane (similar to a tea-bag see Fig. 3A). 

Fig. 3 Schematic representation of batch-wise passive membrane dialysis (A) and continuous membrane filtration dead-end-filtration (B) and loop reactor (C)...

In addition to the vapor diffusion method described previously, other techniques such as the batch and micro-batch methods, bulk and micro dialysis, free interface diffusion, liquid bridge, and concentration dialysis have also been developed to produce crystals for x-ray diffraction analysis (see McPherson, 1982 and McPherson, 1999). 

On one side of the membrane is the solvent, and on the other is the solution to be separated. The particles will pass from the solution side to the solvent side, in the direction of decreasing solute concentration. In a batch dialysis process, the mass transfer of solute passing through the membrane at a given time is ... 

AC will decrease with time in the case of a batch dialysis cell, which is not at steady state. If the same type of membrane were run in a continuous process where the flow of the solution is countercurrent to the solvent flow direction, AC would be constant. In most applications, many of the cells are pressed together to make a stack, and all the cells are run in parallel. 

Ultrafiltration was used to speed up the concentration step and as a substitute for dialysis in washing the concentrated virus. In this modified centrifugation process, the total number of centrifuges was reduced almost in half and the processing time with each centrifuge lowered from 8 hours to k hours. Final concentration of a 30 liter batch was accomplished with 30 ft of 100,000 NMWCO ultrafiltration membrane. Inlet pressure was 20 psi and the outlet pressure was 10 psi (Table V). 

FIGURE 2.8 Release profiles of drags encapsulated in lipospheres. (A) Lipophilic compounds, such as retinyl acetate ( ) and progesterone ( ). (B) Hydrophilic compounds, such as sodium cromoglycate, encapsulated using gelatin (0) or poloxamer 407 ( ) as the stabilizer. As a reference, the release of free SCG is also reported ( ). The releases were determined by dialysis method. Data represent the average of five independent experiments on different microsphere batches. 

A significant leap forward was made in 1988 by Spirin et al. [4], who developed a continuous-flow apparatus for the continuous supplementation of reaction mixtures with substrates required for protein synthesis and continuous removal of reaction by-products. In this way, it was shown that the activity of a cell-free system could be sustained for many hours, compared with batch-mode reactions which became inactive after approximately 45 minutes. Since then, many reports have been made describing the use of simple dialysis systems [3, 5] that can maintain the high productivity of a reaction over many hours, without the use of a cumbersome apparatus. 

A push has been made more recently to develop productive systems that do not rely on flow apparatus or dialysis membranes, in order to make the methods compatible with robot-controlled formats. This has seen the emergence of fed-batch mode reactions where reactions are periodically supplemented with aliquots of reaction substrates [6]. Such a system has... 

PCR-generated DNA templates are excellent for rapid batch-mode screening of domain boundaries, product solubihty and expression levels, and for generating mutants. PCR conditions must be optimized to ensure that a single product is obtained, and high-fidelity enzymes should be used to minimize the introduction of random mutations. For scale-up to dialysis mode, hnear templates are less useful as they are prone to degradation by nucleases and plasmids are less expensive to amphfy. Successful PCR-generated constructs can be sub-cloned directly into an appropriate plasmid vector. 

The improved PEP/pyruvate system developed by Kim and Swartz [7] yielded 350 pg mL of chloramphenicol acetyl-transferase (CAT) in the first hour of an E. coli batch reaction. The same reaction system yielded 750 pg mL CAT in 3 hours when periodically fed with amino acids, PEP, and magnesium acetate [6]. This is remarkable efficiency for a system lacking a dialysis membrane, and is a feature that weU suits robotic handling and high-throughput screening (HTS) strategies. 

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