Whole cell bioprocesses

Several of the problems associated with whole cell bioprocesses are related to the highly effective metabolic control of microbial cells. Because cells are so well regulated, substrate or product inhibition often limits the concentration of desired product that can be achieved. This problem is often difficult to solve because of a poor understanding of the kinetic characteristics of the metabolic pathway leading to the desired product. 

In whole cell bioprocesses, extracellular products are preferable because this removes the requirement for cell disruption and tins reduces the level of impurities in the product solution. Nevertheless, product isolation and purification can be prohibitively expensive particularly for low concentration product streams, which is a feature of many bioprocesses. 

Keywords. In situ product removal (ISPR), Integrated bioprocessing. Whole cell bioprocesses... 

A significant number of general reviews on ISPR techniques in whole cell bioprocessing have been published [1,3,14-21]. Furthermore,more specialized reviews exist that cover either a certain product category such as ethanol [4] or butanol [22,23] or the use of certain specialized ISPR techniques such as aqueous two-phase system [24,25], organic-aqueous two-phase systems [26-30],or solid adsorbents [31]. 

The alternative to batch mode operation is continuous operation. In the continuous mode there is a continuous flow of medium into the fermentor and of product stream out of the fermentor. Continuous bioprocesses often use homogenously mixed whole cell suspensions. However, immobilised cell or enzyme processes generally operate in continuous plug flow reactors, without mixing (see Figure 2.1, packed-bed reactors). 

When compared to traditional chemical synthesis, processes based on biocatalysts are generally less reliable. This is due, in part, to the fact that biological systems are inherently complex. In bioprocesses involving whole cells, it is essential to use the same strain from the same culture collection to minimise problems of reproducibility. If cell free enzymes are used the reliability can depend on the purity of the enzyme preparation, for example iso-enzyme composition or the presence of other proteins. It is, therefore, important to consider the commercial source of the enzyme and the precise specifications of the biocatalyst employed. 

The application of whole-cells or enzyme-based catalysts was protected in two different bioprocess patents ([56] and [57], respectively). The patent specifies the process [57] involving a sulfur-specific reactant with membrane fragments, an enzyme, or a composition of enzymes having the ability to selectively react with sulfur by cleavage of organic C—S bonds, derived from R. rhodochrous strain ATCC No. 53968 or B. sphaericus strain ATCC No. 53969. 

The book is divided into three sections Enzyme mediated bioprocess, whole cell mediated bioprocess and the engineering principle in bioprocess... 

Protein patterns of whole cells by simple SDS-PAG-electrophoresis [68] have been used for a long time in strain identification. Such an analysis resolves something like 20 to 50 bands of protein groups in the order of their (apparent) molecular mass. Detailed analysis of a high number of cellular proteins is usually performed by two-dimensional electrophoresis (see Sect. 7.2 on stress response markers) e.g. for E. coli, this was done by Pedersen et al. [69]. As mentioned above, the two-dimensional technique is not suitable for routine bioprocess monitoring. 

In a bioprocess the desired end product may be present as whole cells or intracellular or extracellular material at the end of a fermentation. Therefore in this first bioseparation stage, it may be necessary to recover either the solid or aqueous phase, with as much of the unwanted phase removed as possible, and with minimal loss of the desired material to maximize product yield. 

There has been much work conducted recently in the area of plant cell culture, or phytoproduction, especially where the product is a plant-unique mixture of Individual flavor substances such as vanilla extract of which vanillin is the major component. As well, the possibility of genetically engineering improved varieties of plants for high yield and consistent quality products is of considerable interest especially for more complex plant-unique flavors. Many flavor compounds are secondary metabolites for which a detailed understanding of their production is not well understood. Presently more knowledge exists In microbial metabolism relative to plant biosynthetic pathways and therefore has resulted in more successful development of microbial-based flavor bioprocessing. As well, scale-up of microbial cultures and isolated enzymes has become relatively common practice while the translation of plant cell culture to large commercial scales is not yet well established. This review will focus on the microbial whole cell and isolated enzyme systems for flavor production. 

There are numerous ways to exploit the calorimetric detection principle in combination with biological materials, not only in metabolite assays, but also for analyses of proteins and other macromolecules as well as for whole cells. These include metabolite determination, bioprocess monitoring, measurements of enzymic activities in separation procedures, determinations in organic solvents, and miniaturized thermal biosensors. 

Enzyme technology is the application of free enzymes and whole-cell biocatalysts in the production of foods and services. A more narrow definition limits enzyme technology to the technological concepts that allow the use of enzymes in competitive large-scale bioprocesses. Enzyme technology is an interdisciplinary field, recognized by the Organization for Economic Cooperation and Development (OECD) as an important component of sustainable industrial development. 

First applications of membrane reactors can be foimd in the field of bioprocess engineering using whole cells in fermentations or enzymatic bioconversions [6, 7]. Most of these processes use polymeric membranes, as temperatures seldomly exceed 60 °C. The development of inorganic membrane materials (zeolites, ceramics and metals) has broadened the application potential of membrane reactors towards the (petro) chemical industry [8]. Many of these materials can be applied at elevated temperatures (up to 1000°C), allowing their application in catalytic processes. 

---