Cell culture batch reactors

The difference of this operation mode compared with batch is that one or more nutrients are fed during cultivation, to replace those consumed by the cells. Fed-batch cultures begin with a working volume lower than the maximum working volume, so that nutrients can be added during the culture period, either from fresh culture medium or from a concentrated nutrient solution. Figure 9.13A shows typical profiles for cell and product concentrations, as well as the volume of medium in the reactor resulting from pulse additions of nutrients. 

Ozturk SS, Palsson BO (1991), Growth, metabolic, and antibody production kinetics of hybridoma cell culture 2. Effects of serum concentration, dissolved oxygen concentration, and medium pH in a batch reactor, Biotechnol. Prog. 7 481-494. 

Another favorable aspect of stirred batch reactors is the fact that they are compatible with most forms of a biocatalyst. The biocatalyst may be soluble, immobilized, or a whole-cell preparation in the latter case a bioconversion might be performed in the same vessel used to culture the organism. Recovery of the biocatalyst is sometimes possible, typically when the enzyme is immobilized or confined within a semi-permeable membrane. The latter configuration is often referred to as a membrane reactor. An example is the hollow fiber reactor where enzymes or whole cells are partitioned within permeable fibers that allow the passage of substrates and products but retain the catalyst. A hollow-fiber reactor can be operated in conjunction with the stirred tank and operated in batch or... 

Typical commercial cell culture systems include batch or fed-batch suspension reactors and perfused immobilized-cell reactors. However, the transient nature of batch culture causes difficulties in studying the effects of external stimuli on growth, metabolism and product formation. Due to metabolite concentration gradients, and the difficulty of obtaining representative cell samples, immobilized-cell reactors are also poorly suited for the analysis of cell growth and metabolism. As a result it is desirable to use well-defined model systems. Continuous-flow suspension reactors allow metabolic parameters to be measured at steady state, after cells have adapted to new (and possibly inhibitory) conditions. Perfusion reactors (with cells immobilized on suspended or stationary supports) extend these benefits to anchorage-dependent cells, and provide model systems for cell responses in vivo. However, while it is instructive to study the behaviour of cells under well-defined conditions, the results obtained must be verified in the culture system selected for commercial production. 

In this case F = F = F=Q. Reactor samples (i.e. F 0) do not affect the analysis because they reduce the culture volume without altering any of the concentrations. However, acid or base additions do affect concentrations. If the volume added is significant the reactor effectively becomes a fed-batch reactor. In this case Equations 4.2.11 and 4.2.12 (below) would apply. For negligible cell lysis the expressions for fx pp, x and for batch culture can be obtained from Equations 4.2.7-4.2.9 by setting D = 0 ... 

Because free cells are removed with the outlet medium, rip is normally much less than n. However, the second term in the RHS of Equation 4.2.26 may be important because the increase in cell concentration often slows as cells reach confluence on the beads. Parameter p. can be calculated from p. pp if p,p is known, and p.p can be determined by allowing the microcarriers to settle, growing the remaining suspended cells in batch culture and analysing the data as described above. Because rip < n, it should be possible to obtain an accurate estimate for p,p at the conditions of interest before the nutrient supply in the medium removed from the reactor is depleted. For many cell types p,p is very small for single cells and /p is of the order 0.5. In such cases, the second term in Equation 4.2.25 can be neglected, especially since rip < n. 

Once Kitt is known, the cellular oxygen consumption rate, qX, can be measured under culture conditions. The mass balance for oxygen in a batch reactor with cells is given by ... 

A fed-batch reactor (Fig. 2) is one in which either nutrients only are fed to the reactor or products only are withdrawn. The feed rate or withdrawal rate may be discontinuous as well. One common example of a fed batch reactor is when cells are grown batch-wise until the late exponential phase, then a small amount of feed is added to the reactor continuously to provide just enough nutrients to allow for the production of secondary metabolites. These secondary products are then removed at given time intervals and purified. Other examples of fed-batch reactors include the discontinuous removal of an inhibitory end product such as ethanol from a fermentor, or the timed addition of an inducer to turn on production of a growth associated product after a cell culture has entered midexponential growth. 

Compared to microbial cultures, cell cultures stiU operate at low ceU densities. While medium development and feed strategies during fed-batch cell culture increases cell densities and productivities, the fact that unused media components and products accumulate in the reactor represents a natural limitation. 

Mammalian growth usually shows respiratory quotients close to 1, that is, carbon dioxide formation (CER) equals oxygen uptake rate (OUR). With respect to the relatively poor mass transfer conditions installed in cell culture reactors (compared to microbial conditions), dissolved CO2 (dCOj) levels can accumulate during fed-batch processes. While optimum dC02 concentrations appear to be cell-specific, growth inhibiting partial pressures of approximately 100 mbar had been identified for, for example, hybridoma cells [81]. 

The first production of poly(3HB-co-3HV), P(3HB-co-3HV), from ohve oils by Aeromonas caviae was described by Doi et al. (1995). Here, the polyester content in the cells was stiU rather low (6-12%). The feasibihty of using olive oil mill effluents as a substrate in biodegradable polymer production was studied by Dionisi et al. (2005), where ohve oil mill effluents were anaerobicaUy fermented at various concentrations combined with different pretreatments and without pretreatment to obtain volatile fatty acids (VFAs) such as acetate, propionate, butyrate, isobutyrate and valerate, which were used as substrates for PHA production. Olive oil miU effluents were also tested for PHA production by using a mixed culture from an aerobic sequencing batch reactor where olive oil miU effluents were centrifuged and tested with or without fermentation. The best results with regard to PHA production were obtained with... 

Consider again a closed volume with uniform composition and temperature — in practice a batch reactor, in which only changes with time occur. From its initial composition the medium evolves after seeding by an inoculum, which is a small amount of the living cell culture with optimized composition, through respectively a lag phase, an exponential growth of the number of cells, a stationary phase during which the increase in number of cells is compensated for by their destruction and a death phase with an (exponential) decrease in the number of cells. 

Several bioreactor designs are used to produce bioproducts, and include, but are not limited to batch reactors, fed-batch reactors, continuous cultivation reactors, plug flow reactors, recycle bioreactor systems, immobilized cell reactors, biofilm reactors, packed bed reactors, fluidized-bed reactors, and dialysis cultivation reactors (Williams 2002). These reactor types can contain either mixed or pure cultures, and can stimulate heterotrophic and/or phototrophic cellular functions depending on the specific reactor design. Additionally, these reactor schemes can be used to produce products directly, or to harvest biomass or other products for downstream processes. Due to the complex nature of bioreactors, particularly anaerobic digesters, the use of metagenomics is helpful to understand the physiology of such systems. 

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