Fermentation Products Recovery

Numerous endeavors have been implemented in fermentation process development in order to alleviate the solvent-induced inhibitory effects and thus improve the fermentation productivity. Usually, the selective removal of fermentation products was integrated simultaneously into the fermentation process in order to maintain a low solvent concentration in the fermentation broth. Many online butanol removal techniques have been reported with various advantages and efficiencies, including liquid-liquid extraction, perstraction, gas stripping, pervaporation, adsorption, reverse osmosis, etc. 

However, liquid-liquid extraction can form emulsions, and the extractant is usually expensive and sometimes toxic to the cell culture (Diirre, 1998). To solve these problems, the perstraction technique was developed based on the liquid-liquid extraction concept. In the perstraction separation process, the fermentation broth and the extractant are separated by a membrane, where two immiscible phases can exchange butanol. In this process, butanol can diffuse preferentially across the membrane, while other components (substrates, cell culture, and other fermentation products) are retained in the fermentation broth. This strategy effectively avoids the potential problems in the liquid-liquid extraction system, but it needs to point out that 

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Chlor-alkali production Electrochemical synthesis Water-organic liquid separation Organic liquid mixture separaion Fermentation products recovery and purification Cell harvesting, virus and antibody concentration Protein desalting, concentration and fractionation Blood processing, including artificial kidney Isolation, concentration, and identification of solutes and particulates... 

Process developers should check existing patents and patent applications in order to avoid patent infringement with any use or process patent. On the other hand, there may be possibilities to patent the particular process newly developed or critical elements of it. Economic aspects such as yields from cell culture or fermentation, product recovery after purification and cost of materials and equipment also need constant attention. Production cost forecasts should be calculated and updated regularly. 

Miranda EA, Berglund KA (1993) Evaluation of column flotation in the downstream processing of fermentation products recovery of a genetically engineered -amylase. Biotechnol Prog 9 411-420... 

The processes and unit operations that are coimnonly used in the pharmaceutical industry include chemical synthesis, fermentation, product recovery and purificatiMi, extraction, formulation, filtration, crystallization, and drying. Drying is appUed to pharmaceutical products to remove excess solvents, as these heat-sensitive materials are more stable in dry than in wet form. 

Biotechnology/medicine Fermentation products recovery and purification... 

Recovery nd Purifica.tion. The production of EH Lilly s human insulin requires 31 principal processing steps of which 27 are associated with product recovery and purification (13). The production process for human insulin, based on a fermentation which yields proinsulin, provides an instmctive case study on the range of unit operations which must be considered in the recovery and purification of a recombinant product from a bacterial fermentation. Whereas the exact sequence has not been pubUshed, the principle steps in the purification scheme are outlined in Figure la. 

Some of the economic hurdles and process cost centers of this conventional carbohydrate fermentation process, schematically shown in Eigure 1, are in the complex separation steps which are needed to recover and purify the product from the cmde fermentation broths. Eurthermore, approximately a ton of gypsum, CaSO, by-product is produced and needs to be disposed of for every ton of lactic acid produced by the conventional fermentation and recovery process (30). These factors have made large-scale production by this conventional route economically and ecologically unattractive. 

Polar organic solvents readily precipitate exopolysaccharides from solution. The solvents commonly used are acetone, methanol, ethanol and propan-2-ol. Cation concentration of the fermentation liquor influences the amount of solvent required for efficient product recovery. In the case of propan-2-ol, increasing the cation concentration can lead to a four-fold reduction in die volume of solvent required to precipitate xanthan gum. Salts such as calcium nitrate and potassium chloride are added to fermentation broths for this purpose. 

Ultrafiltration has the advantage that there is removal of low molecular weight fermentation products and medium components during concentration of the exopolysaccharide. In addition, biological degradation is minimised because fluid is held only for a short time during the filtration process. Other advantages lie in file fact that there is no requirement for solvent recovery and the process is carried out at ambient (not elevated) temperature. 

In this chapter we consider amino acid production by fermentation and by chemo-enzymatic methods. We first consider the stereochemistry of amino adds and the importance of chirality in chemical synthesis. General approaches to amino add fermentation and recovery of amino adds from fermentation broths are then dealt with, followed by a detailed consideration of the production of L-phenylalanine by direct fermentation. Later in this chapter, chemo-enzymatic methods of amino acid... 

The ultimate goal of process development is to achieve feasibility where it is possible to produce amino adds on a large scale at a production cost per kg of amino add comparable to, or cheaper than, the processes currently used by other companies. If we presume that the technical performance (fermentation and recovery) are sorted out on a laboratory scale and scaling up looks promising, then it is time to find out whether it is possible to operate economically on a large scale. 

Fermentation broths are complex, aqueous mixtures of cells, comprising soluble extracellular, intracellular products and any unconverted substrate or unconvertible components. Recovery and extraction of product is important in bioprocess engineering. In particular separation is a useful technique it depends on product, its solubility, size of the process, and product value. Purification of high-value pharmaceutical products using chromatography such as hormones, antibody and enzymes is expensive and difficult to scale up.1 Tire necessary steps to follow a specific process depend on the nature of the product and the characteristics of the fermentation broth. There are a few steps for product recovery the following processes are discussed, which are considered as an alternative for product recovery from fermentation broth. 

Initially fermentation broth has to be characterised on the viscosity of the fluid. If the presence of the biomass or cells causes trouble, they have to be removed. Tire product is stored inside the cells, the cells must be ruptured and the product must be freed. Intracellular protein can easily be precipitated, settled or filtered. In fact the product in diluted broth may not be economical enough for efficient recovery. Enrichment of the product from the bioreactor effluents for increasing product concentration may reduce the cost of product recovery. There are several economical methods for pure product recovery, such as crystallisation of the product from the concentrated broth or liquid phase. Even small amounts of cellular proteins can be lyophilised or dried from crude solution of biological products such as hormone or enzymes.2,3... 

Stages of the extraction process using fermentation broth for product recovery are listed below ... 

The bacterial culture converts a portion of the supplied nutrient into vegetative cells, spores, crystalline protein toxin, soluble toxins, exoenzymes, and metabolic excretion products by the time of complete sporulation of the population. Although synchronous growth is not necessary, nearly simultaneous sporulation of the entire population is desired in order to obtain a uniform product. Depending on the manner of recovery of active material for the product, it will contain the insolubles including bacterial spores, crystals, cellular debris, and residual medium ingredients plus any soluble materials which may be carried with the fluid constituents. Diluents, vehicles, stickers, and chemical protectants, as the individual formulation procedure may dictate, are then added to the harvested fermentation products. The materials are used experimentally and commercially as dusts, wettable powders, and sprayable liquid formulations. Thus, a... 

After fermentation, subsequent midstream to downstream processes such as cell disruption, centrifugation, extraction and drying will be carried on for product recovery. Fig. 9 shows a white sheet of PHB obtained from fermentation of sweet sorghum juice (SSJ) by Bacillus aryahhattai. 

The product is extracted from the culture fluid by adsorption onto caibon or resins rather than by solvent. This illustrates an important general point that antibiotic manufacturing processes differ from one another much more in their product recovery stages than in their fermentation stages. Figure 7.4 illustrates a typical production ronte from inoculum to bulk antibiotic. 

Downstream Processing Microfiltration plays a significant role in downstream processing of fermentation products in the pharmaceutical and bioprocessing industry. Examples are clarification of fermentation broths, sterile filtration, cell recycle in continuous fermentation, harvesting mammahan cells, cell washing, mycelia recovery, lysate recovery, enzyme purification, vaccines, and so forth. 

Fine and specialty chemicals can be obtained from renewable resonrces via multi-step catalytic conversion from platform molecules obtained by fermentation. An alternative method decreasing the processing cost is to carry out one-pot catalytic conversion to final product without intermediate product recovery. This latter option is illustrated by an iimovative oxidation method developed in our laboratory to oxidize native polysaccharides to obtain valuable hydrophilic end-products useful for various technical applications. 

Several areas are receiving much of the research attention. Approaches that integrate product recovery with the fermentation in a three-phase fluidized bed bioreactor reflect general research trends in biochemical engineering (Yabannavar and Wang, 1991 Davison and Thompson, 1992). The successful use of three-phase biofluidization has also been demonstrated for recombinant protein systems, where it may have some benefit in improving plasmid stability (Shu and Yang, 1996). 

Apart from new catalytic methods, cascade conversions require new process technologies, such as in situ product recovery, reactor design, and compartmental-ization. In the long term, part of the present-day stoichiometric chemistry as well as bio- and chemocatalytic conversions in multi-step syntheses will gradually be replaced by cascade catalysis in concert, and full fermentations by cell factory design, or combinations thereof (Fig. 13.17). 

In order to develop a host strain for production of a variety of subtilisin enzymes, it was first necessary to delete the endogenous alkaline and neutral proteases (72,76). The strain was then constmcted to contain the optimal combination of subtilisin regulatory genes which were compatible with the proposed fermentation and recovery process. Once this strain had been produced, the recombinant enzyme... 

By developing a series of generic host production organisms, fermentation development time can be minimized when these host systems are used for production of multiple products. The investment of time and resources in developing the initial fermentation and recovery system can be recovered in subsequent programs in the form of more rapid commercial development timelines. 

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