INDUSTRIAL PRODUCTION OF ENZYMES: PART 1 ENZYMES IN FERMENTATION INDUSTRIES

The good thing about the use of enzymes in various industrial processes amenable to enzyme activity is that the reactions are simpler, faster, easier to control and you do not have to worry much about unwanted side products or unwanted products downstream.
However in various industrial fermentations, the use of enzymes are restricted to a few steps which more than often are prelude steps before the real microbial fermentation process. The restrictions in the use of enzymes for industrial processes are often restricted by the type and suitability of the enzymes needed for the various processes in the complete fermentation process.
In industrial fermentations, the use of enzymes is more in the hydrolysis of substrates prior to its fermentation. This stage is often called the liquefaction stage where large molecules are broken down to simpler monomers to be easily used by the fermenting microorganisms. Enzymes too are widely used in the clarification process of various fermentation juices
Using the enzymes therefore help in making the fermentation process more efficient both technically and economically.
THE NATURE OF ENZYMES
Enzymes are basically catalysts, whose main function is to speed up the specific reactions. In a way, the enzymes are no better than the inorganic catalysts used in many of the industrial chemical reactions.
The differences between enzymes and the inorganic catalysts are more in the sense organic enzymes carry out their reactions under a less demanding conditions. Their reactions are more under gentler physiological conditions found in the normal metabolism of the cells.
Organic enzymes are usually large protein structures which are easily denatured by conditions of extreme ph, temperature and salts and metals. The activity of these enzymes are not stable as they are easily denatured . Their active sites where the reactants interact are very sensitive to such conditions
Industrial catalysts operate under very extreme conditions of pressure, temperature, chemical toxicities which would not be suitable for the living microorganisms.
There is high demand today for enzymes to be used in various food and beverage industries among others. One of the biggest outlet for the production of enzymes is by the use of fermentation technology where the enzymes itself is the sought fermentation products.
Before discussing further the production of enzymes using fermentation technology let us acquaint ourselves with the type and location of enzymes in the microbial cells.
Many new students tend to fantasize the enzymes as equivalent to the magic elixir that can transform almost anything to anything. As we have said earlier the function of enzymes are nothing more than speeding up certain biochemical reactions. The living cell which contains cytoplasm within the membrane sac is nothing more than a complex mixture or soup of enzymes. Without enzymes it’s doubtful life will persist!
In reality the function of enzymes are very limited to carry out certain chemical reactions which involve breaking or making new bonds between the chemical groups. We classify enzymes by the nature of the chemical reactions they carry out!
We also classify enzymes by their location that is they intracellular or extracellular. Or we can classify enzymes whether they are constitutive or inductive, or whether they are soluble or attached
The nature of thee enzymes as stated above will determine the conditions for the production of the enzymes and its downstream extraction isolation and purification
MICROBIAL ENZYMES
Most industrially important enzymes produced by microorganisms are those which are induced and secreted out by the cells into the fermentation broth. It is therefore of great importance and cautions that only suitable microorganisms are used in the production of enzymes using fermentation technology. Common microorganisms used are Bacillus subtilis and Aspergillus oryzae
Much work and research are needed before these microbial enzymes can be used in large fermentors. What is most important is that at least the microorganism chosen must have the ability to produce the desired enzymes. The productivity of the microorganisms can further be enhanced by using various techniques of biotechnology such as gene manipulation
The enzymes produced by these microorganisms do not come in market ready forms! A lot of work needs to be done especially in the downstream activities.




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BIRD’S EYE VIEW OF FERMENTATION PROCESS

One of the first things the students taking fermentation technology will be the outline of the fermentation process flow. He will learn that the complete industrial fermentation process will be made up of three main stages:
1 Upstream activities
2 Midstream activities
3 Downstream activities
In any industrial fermentation process is seen as a pipeline of activities originating upstream with the fermentation products finally being obtained at the end of the pipe or at the downstream stage.
Thus in a simplification in the process flow we see masses being introduced into the process and as it flow down the pipe these masses are being transformed to the desired fermentation products by the fermentation microorganisms.
At each stage of the transformation optimal conditions are implemented not only to obtain the highest yield of the products but also in terms of energy and economic efficiency. This attempts can only be carried out if we assumed that the flow of substrates through the fermentation process will pass through various unit processes which could be identified and optimized.
In such situation, it is important that:
1 The flow of masses from the beginning stages to the final products will be smooth and not interrupted. To achieve this hydrodynamic flow restrictions imposed by various volumetric and hydrodynamic restrictions must be overcome by using different bioreactor or volumetric configurations
2 That each unit process is working under its optimal operating parameters such as temperature, mixing, ph, seeding etc
3 Elimination of physical and physiological bottlenecks in terms of pumping and even removing the effect of catabolite repressions or inhibitions
4 That the impact of any changes in upstream activities will affect the efficiency of the downstream activities. Any improvement or changes in upstream activities will affect downstream and not vice versa
5 That there must be allocation or allowances for changes even within this narrow range of optimized parameters



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PROBLEMS OF HIGH CELL DENSITY FERMENTATION

The microorganisms are important components in any microbial fermentation. The microorganisms in the fermentation process are either the agents of change which convert the substrate to valuable fermentation products or they themselves are the products sought in the fermentation process.
In either situation the objective of the fermentation process it selves is to obtain the highest number of microbes per unit volume. This trend is termed as achieving high cell density fermentation.
If we can achieve high cell density it is akin to saying that we have higher number of cells to carry out the transformation of substrate to products. More cells mean more “factory workers” to carry out the production. Thus more products will hopefully be produced over the period of fermentation process.
It should be noted although increasing the number of cells can increase the amount of fermentation products formed, that does not mean that the real efficiency of the fermentation process has increased in terms of fermentation yield by the unit cell. It is only the numbers of cells has increased leading to more conversions.
Only new strains or improved strains of producing microorganisms will increase the fermentation yield.
Logically, it is a good option to increase the fermentation product formation by increasing the number of microorganisms that will transform the fermentation substrates to fermentation products. However, this is not easily achieved in reality as there are many physical and physiological constraints that will try to prevent this objective.
The microbial cell is unique in its own right. During the process of fermentation not only it transforms the fermentation substrates to fermentation products, it will in the process of metabolism extract energy and carbon from the substrate to produce new biomass and undergo microbial growth.
In a way this would be ideal in achieving the high cell density fermentation, but such growth is not infinite and will ultimately slow down as in the typical sigmoid growth curve of the batch culture fermentation
Even before achieving this stationary phase the cells will it selves undergoes major changes in its metabolism or physiology will be of negative impact to the fermentation process and product formation
So how can we achieve the optimal high cell density fermentation? How shall we operate so that the fermentation of high cell density will be at optimum fermentation product formation in terms of economic and efficient control?
The whole idea of obtaining high cell density fermentation is ideal when:
1 You can have high density or number of microbial cells in the bioreactor
2 The high density protocol will not negatively affect the fermentation process or the products in terms of the quantity, quality and stability of products
3 That the fermentation process will be able to be carried out efficiently and economically in terms of substrate usage and efficient mass transfers
There are various strategies which could be applied in trying to obtain high density cell fermentation not only through feeding mode but also bioreactor configurations and physiological manipulations of the microbial populations





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