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A bioreactor is a vessel in which raw materials under controlled conditions are converted into products by activity of living cells (microorganisms, mammalian, plant and stem cells, tissues and algae) or by cellular components such as enzymes.
Bioreactor and fermenter are similar terms, but with a distinct difference. The term bioreactor often relates to the cultivation of mammalian, plant and stem cells.
If the application is the cultivation of a bacteria, yeast or fungi, then the term fermenter is used. It would not be a mistake to use the term bioreactor in such cases as well, but in the case of cell cultivation only the term bioreactor is used. The name fermenter is associated with reactors in which fermentations are carried out, i.e. metabolic processes that produce chemical transformations in organic substrates through the action of enzymes. The main difference between the bioreactors used for cell and microorganism cultivation is in the mixing and aeration requirements, as well as height and diameter ratio H/D. The mixing environment for microorganism cultivation is usually intensive, with effective dispergation of gas bubbles, and the aeration rate is between 0.5- 3 vvm (volume cultivation media/volume air flow per minute). However, cell cultures require gentle mixing and aeration rate is between 0.01-0.1 vvm. In turn, the optimal H/D ratio of the vessel for microorganism cultivation is 3:1, but in the case of cell cultures it is 2:1.
Generally, bioreactors can be characterized in two ways:
The cultivation principle;
The operation mode.
Cultivation principles of microorganisms are further subdivided into: submerged, immobilisation and solid state.
Cultivation principles of microorganisms are further subdivided into: submerged, immobilisation and solid state.
Inoculation of the microbial culture into the liquid medium for generation of the desired product is known as submerged cultivation or fermentation. Aerobic and anaerobic fermentation processes are the two separate fermentation processes.
In submerged fermentation (or cultivation) the cells of the producer (microorganism) are supplied with nutrient medium and oxygen (in the case of aerobic process) in the entire working volume of bioreactor by mixing and aeration. This makes the process highly economical. The bioreactor creates favorable conditions to accumulate a large amount of actively functioning producer biomass and in turn the target product.
As an example, relating to the history of biotechnology, it can be pointed out that replacing surface fermentation (in flasks and bottles) with submerged fermentation made it possible to increase the production of penicillin in a short time, especially urgently needed during the Second World War.
The submerged fermentation can be aerobic or anaerobic. For example, antibiotics and enzymes are produced through aerobic fermentation, which involves the incorporation of oxygen into the liquid medium, while butanol production proceeds under conditions of anaerobic fermentation, wherein the oxygen influence will have a inhibitory effect. Certain fermentation processes, such as ethanol production, use facultative anaerobic organisms like Saccharomyces cerevisiae, which may grow in the presence of oxygen and produce cell biomass before switching to anaerobic mode during the ethanol fermentation phase. Enzymes (amylases and proteases, amylases, and so on) are often made through aerobic submerged fermentation.
Submerged fermentation processes can be differentiated depending on the target product. The target product can be biomass, ferments or low-molecular compounds (for example, ethanol, methanol, acetates, oxalic and formic acids). Metabolites can be primary or secondary. A primary metabolite is a type of metabolite that plays a direct role in normal development, growth, and reproduction. Some common examples of primary metabolites are lactic acid, and certain amino acids. Secondary metabolites are generated toward or at the conclusion of the stationary phase of growth and do not play a role in growth, development, or reproduction. Atropine and antibiotics like erythromycin and bacitracin are examples of secondary metabolites.
Fermentation processes can finally be differentiated in terms of technology and the type of target product. The target product can be biomass, an individual high-molecular substance (for example, an enzyme – constitutive or inducible) or low molecular weight metabolite. Metabolite, in turn, can be primary or secondary. Thus, the need for inductors and precursors, as well as the time of their introduction into the medium, depends on the target product. The biosynthesis of secondary metabolites is characteristic of certain stages in the development of the producer’s culture and is stimulated in stressful situations. In this regard, the introduction of inducers and precursors is mandatory in the case when the target of the fermentation process is a secondary metabolite.
The biomass accumulation curve usually correlates with the accumulation curve of primary metabolites and does not coincide with the accumulation curve of secondary metabolites.
Submerged fermentation is best suited for microorganisms such as bacteria and yeasts that require high moisture. Another advantage of this method is that product purification is simplified. Submerged fermentation is most commonly employed to extract secondary metabolites that must be used in liquid form.
To carry out submerged fermentations are used as most typical stirred tank, bubble column, airlift type bioreactors, as well photobioreactors and membrane bioreactors in special applications.
Solid state fermentation (SSF) is carried out using solid phase substrate. The microorganisms are growing on a solid substrate in absence or near absence of free water.
The substrate must generate enough moisture to support growth and metabolism of the microorganism. SSF are applied for the production of fermented food products, for example, bread, meat cheese, pickles and yogurt. Using SSF can be recycled agro-industrial residues to obtain, for example enzymes, organic acids, food aroma compounds, biopesticides, mushrooms, pigments, xanthan gum and vegetable hormones. SSF requires less instrumentation and design of bioreactors are relative simple. However, scale-up is bothered, because it is difficult to ensure precise monitoring and control, and can not be controlled environmental conditions of the microorganisms. SSF are long, because the growth rate of microorganisms on solid substrate is slow. There are the processes, which successfully can realized only by SSF. For example, the sporulation of some fungi can attained only by SSF since these fungi do not sporulate in liquid media.
For SSF are used horizontal drum, tray-type, packed-bed and bench scale bioreactors.
Immobilization means the binding of an enzyme to an insoluble carrier while maintaining the functionality, i.e. the catalytic activity of the enzyme. The need of immobilization is determined by the fact that in many applications the end product must be completely free from enzyme residues in order to avoid immune reactions.
The immobilization of enzymes not only significantly increases their stability, but allows the long-term use of one batch or series of industrial biocatalysts. The concept of “immobilization of a biological object” means the physical separation of a biocatalyst and a solvent, in which molecules of the substrate and reaction products can freely penetrate from a liquid to a solid medium, and vice versa. In other words, the substrate in the flow of the solvent is supplied to the bio-object associated with an insoluble carrier, and the reaction product in the flow of the solvent is removed from the bio-object and is used as the target product.
Enzymes, as well as entire cells, can be immobilized. The immobilization, i.e., the fixation of cells onto a carrier, for example, in ethanol production has several advantages. The cells could be reused and have extended lifespan. Some of the traditional purification procedures are not required because the desired final product is essentially free of biological substances and organisms. Inclusion of cells in gels, in which a cell solution is combined with gel-forming chemicals, is one of the most frequent immobilization procedures. Small molecules such as glucose can pass through the gel pores to reach the cells while their metabolic products (alcohol and carbon dioxide) can exit the beads. The living yeast cells so remain intact.
Various immobilization techniques such as the attaching of cells in stable porous gels (e.g., alginate, collagen, chitosan, agarose, cellulose, K-carrageenan, or gel-matrix polymers such as polyacrylamide-hydrazide) or hydrogels or immobilization in solid macroporous carriers has been established and is used on both laboratory and industrial sizes for a variety of applications, including the food, dairy, and beverage sector, medication production, wastewater treatment, agriculture, and biodiesel generation. In the case of the production of pharmaceutical preparations, the target substance will not contain components of the culture liquid (mycelium, products of partial lysis of cells, components of a complex nutrient medium, etc.), which greatly facilitates the task of isolating and purifying the target product, guarantees the absence of proteins and other harmful impurities.
The economic advantages of using immobilized biological objects in production conditions are undeniable. The use of immobilized systems makes it possible to make the conditions of biosynthesis more standard, and the entire production more compact. The resulting biological object works for a long time. At the same time, less raw materials are consumed per unit of production.
The application problem can be that the cells may contain numerous catalytically active enzymes, which can cause unwanted side reactions and the cell membrane itself may serve as a diffusion barrier, thus reducing productivity. It is difficult in immobilized cell bioreactors to control the physiological state of microorganisms, and due to the process variability and flexibility can not be ensured.
Immobilized cell bioreactors divide into stirred tank, fixed bed, fluidized bed, moving bed, packed bed and membrane reactors.