Industrial Biotechnology is a branch of science which harneses the power of Microorganisms to produce Metabolites of interest. They can be classified as Primary Metabolities and Secondary Metabolites in technical terms. These metabolites are produced during the metabolism process of the microorganism for various growth related functions. Microorganisms produce a wide range of sustances essential for their growth such as nucleic acids, proteins, vitamins, carbohydrates, organic acids, antiobiotics and by-products of high energy yielding metabolism such as ethanol, acetone and butanol.
During its growth, the microorganism utilizes a wide range of nutrients in the form of Carbon sources, Nitrogen Source, Inducers, Growth factors etc. To assimilate these nutrients for its metabolism process, they must be broken down into simpler molecules. It is during this phase that the microorganism produces various Protein molecules which catalyse the hydrolysis of the compounds.
These biological catalysts are known as Enzymes.
Nature has a vast collection of a wide range of Microorganisms. Advances in modern microbiology have enabled us to isolate microorganisms thriving in a plethora of environments and understand the conditions of their growth..
For manufacture of products of Industrial importance, it is essential to screen a host of microorganisms under lab conditions to identify their commercial viability. For example, we may be looking to isolate a microorganism from diverse conditions which hold the potential to produce an Alkaline Protease stable for applications involving the enzyme to perform efficiently in a high Alkaline pH. The temperature at which the enzyme performs at its most optimum is also of importance for industrial applications since industries are increasingly looking to reduce energy costs by looking for solutions which demand less energy to produce the same results.
Hence it is essential to thoroughly screen every isolated microorganism for its capabilities as a Cell Factory which produces innovative products of Industrial importance. This would involve constant research in the field of discovering novel microrganims isolated from various environmental conditions.
We at Archimedes Enzymes understand this basic principle of Industrial Biotechnology and constantly work to strengthen our position in isolating and stocking of novel microorganisms from various sources.
When screening for a multitude of microorganisms in the laboratory, the ones which promise the best results for an industrial process are separated from the ones which do not meet expectations. Now comes the most difficult part of the whole process, where the exact optimum growth conditions of the microorganism need to be mimicked in the laboratory.
A microorganism requires a host of nutrients in its growth media to multiply quickly and produce the desired products, but designing an optimum growth media is no simple task. Our Scientists work hard to understand the metabolism process of the microorganism and its behaviour in various media sources. A media composition which works well for one strain may not work well for another. The growth may be limited hence the limited production of the required products. This hampers the economic feasibility of the production source.
The key is finding an optimum nutrient medium which allows the microorganism to achieve the highest growth and produce the product of interest in high volumes. Our team of Scientists and Biochemists work to identify this and develop a solution which guarantees the most valid results.
Finding an optimum growth medium is not the only part of turning the microorganism of interest into a powerful producer. A strain isolated from soil conditions is commonly refered to as Wild type, since it produces a host of other products along with the product of interest. This poses two problems: The yield of the product of interest is relatively low due to production of by-products and the growth medium contains by-products which are not of economic important or sometimes may even be restricted for the targeted end use.
Thus another objective of an efficient process development is preparing a microorganism which does not produce unwanted by-products while enhancing the production capability of the product of interest. This is referred to as Strain improvement and is achieved by Induced Mutations or by Recombinant Technology. Both these processes alter the genetic structure of the microorganism bringing changes to its behaviour. If properly directed, these processes bring efficient changes to the gene of the microorganism such as changed or lower medium nutritional requirements and increased productivity of the product of interest.
Industrial Fermentation refers to the processes involved for large scale culture of microorganisms under controlled parameters of Nutrient composition, Temperature, pH, Oxygen level and other such factors which enable the production microorganism to mass produce the product of interest. It is an important process which has to be thoroughly executed to achieve desired level of target products and needs to be optimized at every stage prior to the main large scale fermentation. But the transition from Discovery of Microorganisms, to their optimization and the actual fermentation process has various stages in between.
Microorganisms have a tendency to adapt slowly to changes around them, hence there are procedures which need to be followed which in technical terms is known as "Upscaling" or "Upstream Processing". Since the cell population post optimization processes, such as Induced Mutations or Genetic Engineering, is very low and not sufficient for large scale fermentation, they need to be multiplied to a large population of microorganisms gradually.
The preserved production microorganism is first transferred to agar slants to start the process. A nutrient medium is added to the agar slants and agitated so a microbial suspension is prepared. This is now transferred to a bottle which contains sterile medium. The bottle is incubated at suitable temperatures to allow the microorganism to start multiplying.
Post a suitable amount of time and growth, the cell suspension from the bottle is transferred to shake flasks containing a liquid nutrient medium. The medium is sterilized in autoclaves prior to addition of the production microorganism. These flasks are now placed in an incubator shaker, where they are incubated at a controlled temperature along with constant agitation. The agitation allows for aeration and the cells now start multiplying at a rapid pace. This serves as a Working Culture which is used to prepare the Inoculum for the large scale fermentation.
There is yet another step between the actual large scale fermentation process and the working culture stage, the one of Seed Fermentation. This refers to the small scale fermentation carried out in Laboratory Fermenters (or Seed Fermenters). This is a small scale version of a large fermenter, having complete functionalities to control the parameters of pH, Temperature, Aeration and Monitor of Dissolved Oxygen, Foam control etc. The microbial cell suspensions from the Shake flasks are used as Inoculum for the Seed Fermentation. The medium contains all the essential nutrients in a calculated composition to allow for effective growth of the microorganism and produce the metabolite of interest. This simulates the large scale fermentation process and allows monitoring of the process before scaling it for bulk production.
The Seed fermentation process is carried out for 1-2 days or more depending upon the production microorganism and this is now used as the microbial inoculum for the bulk fermentation process. The large scale process is achieved using our proprietary Solid Substrate Fermentation technology.
Solid Substrate Fermentation is the culture of microbial cells in a solid medium which serves as the nutrient and the anchor for cell growth. The metabolite production occurs near the surface of the solid medium which translates for fewer processing steps while downstream processing or recovery of the products. Also recent works in the field of Solid Substrate Fermentation have shown relatively higher yields as compared to Submerged Fermentation.
Since Fermentation is an important part of the manufacturing process, we are constantly looking at new approaches to optimization. Solid Substrate Fermentation holds the potential for future innovation and increased productivity in enzyme manufacturing, and we are actively involved in pursuing this production technology
The fermentation broth recovered after the Fermentation process is composed of a mix of substances such as unutilized nutrients, microorganisms, production proteins, fermentation additives and water. The end use of the protein of interest requires it to be at a specific purity depending upon the application such as Food Processing, Animal Feed etc. Hence an optimum Downstream Processing technique is essential to recover the products of interest from the large fermentation broth.
Though this technique may seem to be pure process oriented, the reality is more than it meets the eye. Downstream process is among the major factors which are directly related to the cost of the finished product. An efficient process allows for maximum extraction of the target protein from the fermentation broth while maintaining its activity specifications. Moreover perfecting a process in the laboratory and actually performing it in large scale gives quite different results. It is imperative to maintain the same efficiency standards during bulk processing as previously conducted in laboratory scales; else the final results may vary. Another factor towards achieving efficiency is developing a process which works constantly once initiated, saving valuable overhead costs and energy resources.
In Industrial Biotechnology, especially in the manufacture of Industrial Enzymes, the processing steps for recovery of enzymes from the fermentation broth are applied in a systematic and continuous order. They slightly differ depending upon the Fermentation type (Solid Substrate or Submerged) but basically have the same principles. The steps involve removal of insoluble by pre-filtration, cell disruption if the product is intra-cellular, cell separation and purification & concentration of the target protein prior to formulation. There is one additional step in Solid Substrate fermentation, i.e. Extraction of the Enzymes from the growth substrate or biomass.
Established industrial techniques such as Centrifugation, Homogenization & Ultrasonification, Precipitation, Membrane filtration such as Microfiltration & Ultrafiltration etc are employed for the steps individually.
Designing an efficient downstream processing technique and executing it to perfection is the ardent task which our bioprocess engineers undertake at Archimedes Enzymes.
An important aspect of enzyme manufacturing is that of formulation. An enzyme concentrate recovered after an exhaustive series of downstream processing techniques is to be made available to the end users in a form which best suits their application. For example, in the Textile Industry consumers prefer using liquid enzyme solutions which are easy to dose using automated dosing systems or even manually. This minimizes the wastage caused during handling of the product and results in cost savings. Likewise in Detergent powders, robust enzyme granulates are now preferred which protect the enzyme component from harsh alkaline chemicals and moisture. They also solve the issue of enzyme dust formation during handling. In the Baking segment, clients prefer using an enzyme powder which allows easy dispersion and blending of the enzyme during flour milling or dough making. The end product should meet such expectations and thus the delivery of each product has to be designed in a way which addresses these issues.
The formulation of an enzyme is not limited to the delivery mode but has a far important principle which deals with the very quality and performance of the enzyme. Enzymes being Protein have a three dimensional structure and are prone to denaturing. Thus an enzyme concentrate is required to be blended along with additives which stabilize its structure, thus preserving its biological activity. Additions of these Stabilizers are custom tailored keeping in mind the best stability and performance of the enzyme. Another important component of the formulation is Preservatives which exhibit anti-microbial action.
Formulation also allows incorporation of the enzyme concentrate at specific percentages, thus enabling for a wide spectrum of products depending upon the application and end use. For example, a process may require the enzyme component to be at dilute levels. Hence an optimized formulation can be worked out for the end user depending upon their exact requirements allowing for robust performance along with economic feasibility.
At Archimedes Enzymes, our Biochemists understand the principle of formulation and we work to meet the expectations of our clients while delivering quality products which provide robust performance
The performance of a product in a commercial application, distinguishes it from its competition. Products are designed to work in a specific way, solving a set of problems in an industrial process and their performance in addressing those problems and meeting expectations is what turns it into a commercial reality.
With a host of companies out there in the market, claiming to deliver such solutions what would cause a client to prefer one product over the other? How does a company manufacturing the product understand what sets its solutions apart from its competitors?
The key understands how an enzyme works under diverse industrially important parameters and the factors affecting them. It is about understanding what makes a good product perform efficiently and working to develop better alternatives.
At Archimedes Enzymes, a finished formulation is subjected to various enzymatic activity tests in our Quality control facility. We perform comparative assays of the enzymatic activity of interest and prepare comparative charts mapping the performance of an enzyme product under various parameters. A principle of backward integration is followed where the Marketing team works in close collaboration with the Quality Control and R&D divisions feeding market expectations and demands to them. After a thorough analysis, the product is forwarded to the marketing division for pre-launch trials to determine the actual Industrial Performance of a product