Researchers realized the benefits of developing large-scale chromatographic processes operating continuously and employing the countercurrent contact between a fluid or gas and an appropriate chromatographic packing. Two main schools of thought developed: the moving-bed and moving-column systems. This chapter focuses on the development and application of the latter systems, with particular emphasis on their application to the separation of carbohydrate mixtures and the continuous fractionation of macromolecules. The moving-column system, however, could be particularly promising in analytical applications with further mechanical development of the system. The simulated moving-column chromatographic systems have been found to overcome the foregoing problems and offer better promise in large-scale continuous applications. Although the moving-tube bundle systems described previously were successful in gas and liquid separations, their weaknesses are associated with the difficulties in manufacturing and maintaining uniformly flat metal surfaces and thus preventing any leakages.

This chapter focuses on developments made with the simulated moving bed, with particular emphasis on Aston University semicontinuous chromatographic refiner (SCCR) systems and their applications in liquid separations. The various SCCR systems used for dextran fractionation were packed with Spherosil XOB075 of 200- to 400-μm porous silica beads, employing the slurry packing technique. The SCCR7 Mkl system was used in the anion-exchange mode to investigate factors affecting the separation of synthetic equimolar glucose-fructose and hydrolyzed sucrose feedstocks. The great separation potential and versatility of chromatography have been widely accepted and has been established as a powerful analytical tool. In moving-bed systems the stationary phase flows under gravity countercurrent to a stream of mobile phase flowing upward. The moving-column continuous systems consist of a circular array of interconnected columns, which rotate as a whole past fixed inlet and outlet ports.

Describes the latest developments in the scaling-up and application of chromatographic operations and demonstrates that production-scale chromatography is a powerful and invaluable separation process. The book covers every important process design and reveals actual, immediately applicable techniques and is designed to appeal to design, chemical/biochemical, and research and development engineers, process development managers, bioprocess technologists, analytical and clinical chemists and biochemists, pharmacists, and upper-level undergraduate, graduate, and continuing-education students in these disciplines.

The hydrolysis of lactose using the enzyme lactase from Aspergillus oryzae and the simultaneous separation of the products galactose and glucose was examined in a Simulated Counter-Current Chromatographic Bioreactor-Separator (SCCR-S) system. The system was operated in continuous and batch modes and the results obtained used to compare the performance in both modes. Mathematical modelling and computer simulation of the system operating in the continuous mode are reported.

Combined bioreaction and separation has been successfully carried out in a Simulated Counter-Current Chromatographic Reactor-Separator (SCCR-S) system for the saccharification of modified starch to maltose and dextrin. The effects of the operating parameters (switch time, eluent flowrate, feed concentration and enzyme activity) on the performance of the SCCR-S system were investigated. By using an eluent of dilute enzyme solution, starch conversions of up to 60% were achieved using lower amounts of enzyme than the theoretical amount required by a conventional bioreactor to produce the same amount of maltose over the same time period. Comparing the SCCR-S system with a continuous rotating annular chromatograph (CRAC) for the saccharification of modified starch showed that the SCCR-S system required only 34.6-47.3% of the amount of enzyme required by the CRAC, system.

Combined biosynthesis and product separation has been successfully performed for the first time using a zonal centrifugal bioreactor-separator. The biosynthesis of dextran polymer from sucrose using the dextransucrase enzyme was investigated in order to evaluate bioreactor performance. The bioreactor consisted of a bowl, fitted with a Reorienting gradient rotor which facilitated substrate and enzyme loading and the reproducible unloading of solutions from the system. This allowed the distribution of substrate, enzyme and product materials in the bowl at the end of each trial to be accurately determined. Studies have indicated that combined bioreaction-separation is possible using this system. However, viscosity build-up in the bioreactor must be minimised in order to increase the yield of polymer product per unit time and improve product separation.

High activities of the enzyme dextransucrase were repeatedly produced using slowly agitated non-aerated fed-batch fermentations of Leuconostoc mesenteroides B-512(F). Activities in excess of 24.0 U cm-3 were obtained consistently in a 16 dm3 laboratory fermenter using a 6 dm3 initial work volume. Yeast extract type was identified to be one of the important factors influencing the enzyme yield.Studies on aerating the medium with different gases indicated that the presence of carbon dioxide in the medium favoured high enzyme production. Agitation rates did not appear to have significant effects on either cell growth or enzyme production. One type of antifoam (silicone antifoam) was observed to affect enzyme production but not the cell growth.Scale-up of the non-aerated process was carried out up to a 1000 dm3 scale with enzyme broths containing up to 21.0 U cm-3 being produced. Two batches of the enzyme that were produced at the large scale were used for the first time to synthesize dextran at a 50000 dm3 industrial scale. The dextran yields were up to 95.5 % of the conventional industrial yields and were achieved in much shorter reaction time intervals.

The extracellular enzyme dextransucrase was produced from Leuconostoc mesenteriodes NRRL B512F and purified by ultracentrifugation and cross-flow ultrafiltration for use in the biosynthesis of the macromolecule dextran by ion exchange chromatographic reaction-separation techniques. The two-stage purification process yielded over 90% pure dextransucrase with overall enzyme recovery of over 60%. A second stage of centrifugation was required to achieve complete cell removal. The purified enzyme contained 1-2 g l-1 of solute ions, which affected the operation of the chromatographic system. Gel filtration removed over 93% of the remaining ions but resulted in high activity losses. Two-phase separation with polyethylene glycol (PEG) and purification by ion exchange chromatography were less successful in desalting the enzyme. PEG precipitation was successful in concentrating the enzyme, but the ions remained predominantly with the enzyme portion of the two phases. The purified enzyme was found to be unstable during storage. Use of the enzyme in chromatographic reactor-separators for the production of dextran resulted in over 33% more high molecular weight dextran (the desired product) and a useful pure fructose byproduct being obtained than for a conventional reactor. Sodium and potassium ions in the enzyme hampered continuous operation by displacing calcium ions from the resin and thus reducing the separation efficiency of the system. Partial regeneration of the resin with calcium nitrate rather than complete enzyme desalting, which was very expensive and resulted in high activity losses, helped overcome this effect.

Efficient unit operations for the production and purification of the enzyme dextransucrase and the biosynthesis and fractionation of dextrans have been developed and tested. An alternative process for the production of clinical-grade dextran has been proposed by integrating these processes to achieve improved dextran product yield and the recovery of fructose as a valuable by-product. The enzyme dextransucrase, which synthesizes dextran from sucrose, has been produced either by non-aerated fed-batch or continuous fermentations, purified by continuous ultracentrifugation and ultrafiltration and used in a bioreactor-separator stage for the biosynthesis of dextran. High molecular weight dextrans have been biosynthesized on continuous counter-current chromatographic bioreactor-separator systems. Membrane filtration and size exclusion continuous chromatography were used for the fractionation of the native dextran and production of clinical-grade dextran. The advantages of the proposed process over the conventional process are related to cost reduction due to the elimination of ethanol usage and recovery, improved product quality control and the production and recovery of fructose as a by-product.

Simultaneous biochemical reaction and separation has been carried out successfully for the first time in a continuous rotating annular chromatograph (CRAC) by inverting sucrose to glucose and fructose using the enzyme invertase. The chromatograph was packed with 14.5 dm3 Dowex 50W-X4 calcium form ion exchange resin. Results from the initial experiments indicated that complete conversion could be achieved for feed concentrations of up to 50% w/v sucrose and at feed throughputs of up to 15 kg sucrose per m3 resin/h. Numerical simulation for the combined biochemical and separation on a CRAC has also been carried out. The model was solved using a finite difference method and the results indicate a good agreement between the experimental and the predicted elution concentration profile.

The application of a 12 column preparative scale semi-continuous counter-current chromatographic bioreactor-separator (SCCR-S) as a combined bioreactor-separator is described. The SCCR-S system has been used successfully in continuous sucrose inversion and simultaneous product separation. This paper focusses on continuous dextran biosynthesis from sucrose and simultaneous product separation also the continuous saccharification of modified starch to produce maltose using the enzyme maltogenase.

Simultaneous biochemical reaction and separation has been successfully carried out in a continuous rotating annular chromatograph (CRAC) by saccharifying liquefied starch to maltose using the enzyme maltogenase. Maltogenase is a thermostable, exo-acting alpha-amylase. A simple optimisation procedure involving both volume overload and concentration overload was used. For soluble potato starch, maltose conversions of up to 79% were achieved at feed flow rates of up to 400 cm 3 h -1 and 15.5% w/v concentrations. The resolution between maltose and dextrin was 0.92.

A four stage diafiltration cascade was used to enhance the fractionation of dextran. This experimental work was complemented by the development of a mathematical model. By using the model and experimental equipment a detailed insight into the important parameters effecting the efficiency and operation of the cascade was achieved. The results suggested that fractionation efficiency could be improved by as much as 18% and the amount of diafiltrate reduced by up to 33%.

A chromatographic process may operate continuously if there is relative motion between the stationary (absorbent) phase and the point where the feed is introduced into that phase. Under these circumstances, component column residence time differences are transformed into physical displacements so that each feed component may be withdrawn continuously at a fixed and characteristic distance away from the feed point. The objective of this work is to model the continuous separation on the rotating annular chromatograph and to compare model and experimental data. A simulation model of the rotating annulus separation of a binary mixture has been developed. The steady-state chromatographic cycle is considered as the successive application of the saturation and elution processes. The equations developed are complicated by considering the effects of interaction between the two solutions. Experimental data are obtained from the separation of the isomers, glucose and fructose.