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Unit:1
FERMENTATION AND ITS TYPES
Industrial microbiology - Types of fermentation- Solid, Submerged - Batch, Continuous, Fed
batch - Component parts of fermentation process - Fermentation economics
The field of industrial microbiology has been undergoing rapid transformations in recent
years. First, what has been described as the cook book approach, has been discarded for
the rational manipulation of micro-organisms, on account of the increased knowledge of their
physiology. Second, influential new tools and technologies, particularly genetic engineering,
genomics, proteomics, bioinformatics and such like new areas promisethrilling horizons for
the continued exploitation of micro-organisms by human beings. Third, new approaches
have become available for the utilization of some traditional microbialproducts, such as
restrained enzymes and cells, site-directed mutation and metabolicengineering.
Simultaneously, microbiology has addressed itself to some currentproblems, such as, the
fight against cancer by the production of anti-tumor antibiotics; ithas changed the traditional
practice in a number of areas: for example, the deep sea hasnow joined the soil as the
medium for the search for new bioactive chemicals, such asantibiotics (Okafor, 2007).
Even the search for organisms producing new products has now been broadened to include
organisms, which are isolated primarily on genes secluded from the environment. Finally,
greater consciousness of the effect of fossil fuels on the environment has increased the use
of more environmental friendly and renewable sources of energy, has led to a search for
alternate fermentation substrates, exemplified in cellulose, and a return to fermentation
production of ethanol and other chemicals. Due to the increased knowledge and changed
approach, even the definitions of acquainted words, such as, antibiotic and species seem to
bechanging. Industrial microbiology may be defined as the study of large-scale and profit
motivated production of micro-organisms or their products for direct use, or as inputs in the
manufacturing of other goods. Thus, yeasts may be produced for direct consumption as food
for humans or as animal feed, or for use in bread-making; their product, ethanol, may also
be consumed in the form of alcoholic beverages, or used in the manufacture of perfumes,
pharmaceuticals, etc. Industrial microbiology is a branch of biotechnology and includes the
traditional and nucleic acid aspects (Okafor, 2007).What is Industrial Microbiology?Industrial
microbiology or microbial biotechnology is the application of scientific andengineering
principles to the processing of materials by micro-organisms, such as bacteria, fungi, algae,
protozoa and viruses or plant and animal cells to create worthwhile products or processes.
The micro-organisms utilized may be native isolates, laboratory-selected mutants or
microbes that have been genetically modified using recombinant DNA methods.
Meta-genomics, the study of all of the genetic material in an environmental sample, is being
used toscreen for microbes with potentially useful industrial properties. In some cases, the
organismshave been developed using biology, the design of new biological systems or the
re-design of existing systems. The terms “industrial microbiology” and “biotechnology” are
often considered to be one and the same (Careers in Industrial Microbiology and
Biotechnology, 2014). Areas of industrial microbiology include discovery of new organisms
and passageways, such as anti-microbial drugs. For instance, most antibiotics come from
microbial fermentations involving a group of organisms called actinomycetes. Other
organisms, such as yeasts are used in baking, in the production of alcoholic beverages, and
in biofuel production. Additional groups of micro-organisms form products that range from
organic acids to enzymes used to create various sugars, amino acids, detergents, and
consumer products and specialty ingredients. For example, the sweetener aspartame is
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derived from microbially produced amino acids. Industrial microbiologists may also be
responsible for the bioremediation of air, soil, and water contamination. Industrial
microbiologists may also deal with products associated with the food, dairy, and consumer
products industries, along with the prevention or deterioration of processed or manufactured
goods, and with waste disposal systems. Quality assurance for the food, pharmaceutical,
and chemical industries is a large area, along with the health of animals used in testing
products (Careers in Industrial Microbiology and Biotechnology, 2014). History of Industrial
MicrobiologyThe practise of industrial microbiology has its roots deep in ancient times. Long
before their discovery, micro-organisms were exploited to serve the needs and requirements
of humans, for example to preserve milk, fruits, and vegetables, and to improve the quality
oflife by producing beverages, cheeses, bread, pickled foods, and vinegar. The use of yeasts
dates back to the ancient days. The oldest fermentation know-how, the conversion of sugar
to alcohol by yeasts was used to make beer in Sumeria and Babylonia as early as 7000 B.C.
By 4000 B.C., the Egyptians had discovered that carbon dioxide was produced by the action
of brewer’s yeast could leaven bread. Ancient individuals are also known to have made
cheese with molds and bacteria. Another ancient product of fermentation, wine, was made in
Assyriain 3500 B.C. and reference to wine can be found in the book of Genesis, where it is
noted thatNoah consumed too much of the beverage (Demain, 2010).According to the
Talmud, ‘a man without salt and vinegar is a lost man.’ In the field of human health, vinegar
has a long history of use. The Assyrians used it to treat chronic middle ear diseases,
Hippocrates treated patients with it in 400 B.C., and, according to the New Testament,
vinegar was offered to Jesus on the cross. The use of micro-organisms in food also has a
long history. In 100 B.C., ancient Rome had over 250 bakeries, which were making leavened
bread. As a method of preservation, milk was fermented to lactic acid to make yogurt and
also converted into kefyr and koumiss, using kluyveromyces species in Asia. The use of
molds to saccharify rice in the koji process dates back to A.D. 700. By the fourteenth century
A.D., the distillation of alcoholic spirits from fermented grain, a practise thought to have
originated in China or the Middle East, was common in many parts of the world. Vinegar
manufacture began in Orleans, France, at the end of the fourteenth century and the surface
technique used is known as the Orleans method (Demain, 2010). In the seventeenth
century, Antonie van Leeuwenhoek, a Dutch merchant did not acquire any professional
educational qualifications. But he developed keen interest in the
construction of microscopes, made use of a simple lens in the examination of water,
decayingmatter, and scrapings from his teeth. There he reported on the presence of tiny
animalcules, moving organisms less than a thousandth the size of a grain of sand.
Leeuwenhoek’s lack of university connection might have caused his findings to go
unrecognized, had it not been for the Royal Society in England and its secretary. Henry
Oldenburg, who corresponded with European science individuals, who were not very skilled.
From 1673 to 1723, Leeuwenhoek ’s observations as a microscopist were communicated to
the society in a series of letters (Demain, 2010).Most scientists at the time thought that
microbes arose spontaneously from non-living matter. What followed was a 100 year debate
over spontaneous generation, appropriately called the ‘War of the Infusions’. Supporters had
previously claimed that maggots were naturally created from decaying meat. But this theory
was a generation, originally postulated by Aristotle, among others, have been condemned
with respect to higher forms of life, so the proponents concentrated their arguments on
bacteria. The theory did seem to provide explanation in terms of how a clear broth became
cloudy via growth of large numbers of spontaneously generated micro-organisms as the
broth aged (Demain, 2010).However, others possessed the viewpoint that micro-organisms
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only came from previously existing microbes and that their universal presence in air was the
reason that they would develop in organic infusions after gaining access to these rich liquids.
In the early nineteenth century, three independent investigators, Charles Cagniard de la Tour
of France, Theodor Schwann, and Friedrich Traugott Kutzing of Germany proposed that the
products of fermentation, mainly ethanol and carbon dioxide, were created by a microscopic
form of life. This concept was severely opposed by the leading chemists of the period, such
as Jons Jakob Berzelius, Justus von Liebig, and Friedrich Wohler. They possessed the
viewpoint that fermentation is a chemical reaction. They maintained that the yeast in the
fermentation broth was a lifeless decaying matter (Demain, 2010).Organic chemistry was
flourishing at the time, and these opponents of the theory of living microbial origins were
quite successful in putting forth their viewpoints. Attentiveness in the instruments of these
fermentations resulted in the later investigations by Louis Pasteur. These investigations not
only brought about advancements in microbiology as a distinct discipline, but also led to the
development of vaccines and concepts of hygiene which revolutionized the practise of
medicine. In 1850, Casimir Davaine detected rod-shaped objects in the blood of
anthrax-infected sheep and was able to produce the disease in healthy
sheep by inoculation of such blood. In the next 25 years, Pasteur of France and John Tyndall
of Britain finally demolished the concept of spontaneous generation and proved that
existingmicrobial life came from pre-existing life (Demain, 2010).The work of Pasteur
originated in his work as a chemist on chirality. In the1850s, Pasteur had detected two
optical types of amyl alcohol, that is, D and L, but he was not able to separate the two. For
this reason, he began to study living microbes carrying out fermentation which led to his
conclusion. In 1857, that fermentation was a living process of yeast. In 1861, Pasteur proved
the presence of microbes in the air, which discredited the theory of spontaneous generation
of microbes, and it was at this point that fermentation microbiology was originated.
Nevertheless, it took almost two decades, until 1876, to disprove the chemical hypothesis of
Berzelius, Liebig, and Wohler, i.e., that fermentation wasthe result of contact with decaying
matter (Demain, 2010).In 1876, the great German microbiologist Robert Koch proved that
bacteria from anthrax infections were capable of causing the disease. His contributions
involving the growth of microbes in pure culture led to the decline of the pleomorphism
theory, which postulated that one form of bacteria developed into another. It was primarily
Koch’s work that led to the acceptance of the idea that specific diseases were caused by
particular organisms, each of which had a specific form and function. In 1884, his students
Gaffky and Loeffler were able to confirm the etiologic role of infectious bacteria in typhoid
fever and diphtheria. Pasteur was called on by the distillers of Lille to find out, why the
contents of their fermentation vats were turning sour. Using his microscope, he noted that
the fermentation broth contained not only yeast cells, but also bacteria, and he already knew
that these could produce lactic acid. This observation led to his suggestion that such souring
couldbe prevented by a mild heat treatment, which later came to be known as pasteurization
(Demain, 2010).During World War I, the need for glycerol, used to manufacture ammunition,
resulted in the application of yeast to convert sugars into glycerol. This development led after
the war to a comprehensive study by Neuberg of the mechanisms, involved in these
reactions and those converting sugars to ethanol. This was followed by the studies of Dutch
scientists in Delft looking at oxidation, reduction reactions and the kinetics of enzyme catalyzed reactions. Also during World War I, Chaim Weizmann at the University of
Manchester, applied the butyric acid bacteria used for centuries for the retting of flax and
hemp, for production of acetone and butanol. His use of Clostridium to produce acetone and
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butanol was the first non-food fermentation developed for large scale production. With it
came the
problems of viral and microbial contamination that had to be solved. Although use of this
fermentation faded because it could not compete with chemical methods of solvent
production, and it did provide a base of experience for the development of large scale
cultivation of fungi for production of citric acid. Soon after World War I, an aerobic process
was developed in which Aspergillus niger was used. Not too many years later, the
discoveriesof penicillin and streptomycin and their commercial development indicated the
beginning of the antibiotic era (Demain, 2010).Multi-disciplinary Nature of Industrial
MicrobiologyUnlike many other areas of the discipline of microbiology, the microbiologist in
anindustrial establishment does not function by himself. He is usually only one of a number
of different functionaries with whom he has to interact constantly. In a modern industrial
microbiology organization, others may include chemical or production engineers,
biochemists, economists, lawyers, marketing experts, and other high-level
functionaries.They all co-operate to achieve the purpose of the firm, which is not
philanthropy, but the generation of profit or wealth (Okafor, 2007).Despite the necessity for
team-work emphasized above, the microbiologist has a central role in his organization.
Some of the functions include: (Okafor, 2007).1. The selection of the organism to be used in
the processes.2. The choice of the medium of growth of the organism.3. The determination
of the environmental conditions for the organism’s optimumproductivity i.e., pH, temperature,
aeration, etc.4. During the actual production, the microbiologist must monitor the process for
theabsence of contaminants, and participate in quality control to ensure uniformity ofquality
in the products.5. The proper custody of the organisms, usually in a culture collection, so
that theirdesirable properties are retained.6. The improvement of the performance of the
micro-organisms by genetic manipulation or by medium reconstitution.The Use of the Word
‘Fermentation’ in Industrial MicrobiologyThe word fermentation comes from the Latin verb
fevere, which means to boil. It originated from the fact that early at the start of wine
fermentation, gas bubbles are produced continuously to the surface, giving the impression of
boiling (Okafor, 2007). Fermentation is referred to any process involving the mass culture of
micro-organisms, aerobic or anaerobic,
any biological process that occurs in the absence of O2, food spoilage, the production of
alcoholic beverages, use of an organic substrate as the electron donor and acceptor, use of
an organic substrate as an electron donor, and of the same partially degraded organic
substrate asan electron acceptor and growth dependent on substrate level phosphorylation
(Niel, n.d.).It has three different meanings, which might be perplexing. The first meaning
relates to microbial physiology. In strict physiological terms, fermentation is defined in
microbiology as the type of metabolism of a carbon source in which energy is generated by
substrate level phosphorylation and in which organic molecules function as the final electron
acceptor or as acceptors of the reducing equivalents generated during the break-down of
carbon containing compounds or catabolism. As it is well-known, when the final acceptor is
an inorganic compound, the process is called respiration. Respiration is referred to as
aerobic,if the final acceptor is oxygen and anaerobic, when it is some other inorganic
compound outside oxygen, e.g. sulphate or nitrate (Okafor, 2007).The second usage of the
word is in industrial microbiology, where the term‘fermentation’ is any process in which
micro-organisms are grown on a large scale, even if the final electron acceptor is not an
organic compound, i.e. even if the growth is carried out under aerobic conditions. Thus, the
production of penicillin, and the growth of yeast cells, which are both aerobic to a major
extent, and the production of ethanol or alcoholic beverages, which are fermentations in the
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physiological sense, are all referred to as fermentations. The third usage concerns food. A
fermented food is one, in the processing of which micro-organisms play a major part.
Micro-organisms determine the nature of the food through producing the flavoured
components as well as making decisions regarding the general character of the food. But
micro-organisms form only a small portion of the finished product by weight. Foods such as
cheese, bread, and yoghurt are fermented foods (Okafor, 2007).Organizational Set-up in an
Industrial Microbiology EstablishmentIndustrial microbiology and biotechnology both involve
the utilization of micro-organisms to achieve specific goals. Whether, these are creating new
products with monetary value or making improvements in the environmental conditions.
Industrial microbiology, as ithas traditionally developed, put emphasis on the products, such
as, pharmaceutical and medical compounds, i.e. antibiotics, hormones, transformed
steroids, solvents, organic acids, chemical feed-stocks, amino acids and enzymes that have
direct economic value. The micro-organisms employed by the industry have been secluded
from nature and in many cases, were
modified, making use of classic mutation-selection procedures. The first task of the industrial
micro-biologist is to find a suitable micro-organism for use in the preferred processes. A
comprehensive range of approaches are available, ranging from isolating micro-organisms
from the environment to making use of refined molecular strategies to modify an existing
micro-organism. Micro-organisms are made use of in the industrial microbiology and
biotechnology to create a wide variety of products and to assist in maintaining and bringing
about improvements in the environmental conditions (Chapter 42, 2002).Microbiologists are
rendering a significant contribution in expanding the micro-organisms on a rapid scale, with
characteristics, which are desirable for use in industrial microbiology and biotechnology.
They are also identifying the micro-organisms involved in mutualistic and protocol-operative
relationships with other micro-organisms and with plants and animals. There is a continuing
interest in bio-prospecting in all areas of the world and major companies have been
organised in continuing and exploring microbial diversity and identification of
micro-organisms with new capabilities (Chapter 42, 2002). In industrial microbiology, the
individuals are engaged in preparation of products, which may also include, foods and
beverages. In order to carry out their job duties well, they need to possess efficient
knowledge in terms of the methods and procedures. Investigation is regarded as an
important aspect that lead to progression of the field of microbiology (Industrial Microbiology,
n.d.).The organization of a fermentation industrial establishment will differ from one firm to
another and will depend on the production processes. The culture usually comes from the
firm’s culture collection, but may have been sourced initially from a public culture collection
and connect to a patent. On the other hand, it may have been secluded by the firm from soil,
the air, the sea, or some other natural body. The nutrients, which go into the medium are
compounded from various raw materials, sometimes after proper preparation or
modification,including saccharification as in the case of complex carbohydrates, such as
starch or cellulose. An inoculum is first prepared regularly from a lyophilized vial, whose
purity must be checked on an agar plate. The organism is then grown in shake flasks of
increasing volumes until about 10 percent of the volume of the pilot fermentor is attained. It
is then introduced into pilot fermenters, before final transfer into the production fermentors
(Okafor, 2007).The extraction of the material depends on what the end product is. There are
differences between the methods depending on whether the organism itself, or its metabolic
product is the desired commodity. If the product is the required material, the procedure will
be dictated by its chemical nature. Quality control must be carried out on a regular basis to
ensure that the right material is being produced. Sterility is important in industrial
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microbiology processes and is maintained by various means, including the use of steam
filtration or by chemicals. Air, water, steam and other services must be supplied and
appropriately treated before use. The wastes generated in the industrial processes must also
be disposed off. Packaging and sales are to be carried out at the end, but are by no means
the least important. Indeed they are about the most important, as they are the points of
contact with the consumer, for whose satisfaction, all the hard work was carried out in the
first occurrence (Okafor, 2007).Job Duties of Industrial MicrobiologistsJob duties of industrial
microbiologists have been highlighted in the factors, which have been stated as follows:
(Careers in Industrial Microbiology and Biotechnology, 2014). Antibiotics/Antimicrobials Both natural and chemically improved microbial productscan be used to control the diseases
caused in humans, animals, and plants. Using traditional genetics or recombinant DNA
techniques, the micro-organisms can be reformed to improve the yield or action of antibiotics
and other antimicrobial agents. New research guidelines are aimed at determining microbial
metabolites with pharmacological activities, suitable in the treatment of diseases such as,
hypertension, obesity, coronary heart disease, cancer, and inflammation.Vaccines - Vaccines
are crucial to protect humans and animals from microbial diseases. Recombinant DNA
technology has permitted for the production of vaccines that offer protection without risk of
infection, e.g., hepatitis B vaccine. Industrial microbiologists are enthusiastically involved in
the development of these new vaccines.Health-Care Products - The development and
production of diagnostic assays that utilize monoclonal antibody or DNA probe technology
are crucial in the manufacturing of health-care products, such as rapid tests for strep throat,
pregnancy, and AIDS. Micro-organisms are also used to produce human or animal biological
such as, insulin, growth hormone, antibodies, and components for cosmetics. The industrial
microbiologist orbiotechnologist may screen new microbial sources, e.g., marine or
cave-dwelling micro-organisms for their ability to produce new pharmaceuticals or develop
new diagnostic assays.Food and Beverages Produced by Microbial Activity – Food items
such as, yogurt, cheese, chocolate, butter, pickles, sauerkraut, soy sauce, food
supplements, such as vitamins and amino acids, food thickeners, produced from microbial
polysaccharides, alcohol (beer,
whiskeys, and wines), sausages, and silage from animals are a small sample of products of
microbial activity. Industrial microbiologists and biotechnologists may be involved in
producing concentrated microbial inocula for fermentations or the maintenance of
fermentation systems utilized in the production services. They may also take part in
identifying the organisms involved in and maintaining proprietary culture
collections.Agriculture - Conventional, recombinant DNA and monoclonal antibody
techniques are used to make improvements in microbial inoculants. These serve as fertilizer
supplementsby fixing atmospheric nitrogen to improve plant yields and to serve as plant pest
controls. All of these require a microbiologist to insure product usefulness and
worth.Enzymes - Industrial applications of enzymes include the production of cheese, the
interpretation of apple juice, the development of more efficient laundry detergents, pulp and
paper production, and the treatment of sewage. These processes have been vividly
enhanced by the use of recombinant DNA techniques to design enzymes and increase
action, constancy,and specificity.Carbohydrates - Some molecular sieves for purification and
separation processes e.g.,dextran and thickening agents e.g., xanthan used in salad
dressings, which are steady at high temperatures, are examples of microbial carbohydrates.
The latter are also used for secondaryoil recovery in oil fields and as lubricants in drilling oil
wells, gelling agents in foods, and thickeners in both paints and food products.Organic
Chemicals - Compounds such as, acetone, methanol, butanol, and ethanol have numerous
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applications in industrial settings. They are frequently used as raw materials for industrial
manufacturing and production processes. The microbiologist is involved in research on
improvements in the production and detection of new metabolic passageways. Microbes will
progressively be used to supplant or replace those processes, which rely on petroleum and
natural gas for the production of these compounds.Contamination Control - The industrial
microbiologist develops assays to detect microbial contaminants in foods and carry out vital
functions. These include, development of preservatives, evaluation of natural or synthetic
agents for the prevention of diseases, deterioration, or spoilage and determination of minute
quantities of vitamins or amino acids in food samples. Microbiologists are also involved in
the development of procedures for the control of deterioration in cosmetics, steel, rubber,
textiles, paint, and petroleum products.Waste and Wastewater Management - The
production of clean water and the destruction of waste material are important for preserving
the environment and providing
clean drinking water. The industrial microbiologist is directly involved in developing microbial
strains to detoxify wastes of industrial, agricultural, or human origin.Oil Recovery/Mining - Oil
recovery may be facilitated by the development of distinctive bacteria, which produce a
surfactant that forces trapped oil out of the rocks. Extraction of minerals from low-grade ores
is augmented by some bacteria, i.e. microbial leaching. In addition, selective binding of
metals by bio-hydrometallurgical processes is important in recycling of metals, such as silver
and uranium. Research and developments in these areas also make provision of career
paths for industrial microbiologists and biotechnologists.Environmental Microbiology Examination of microbes living in unusual environments, e.g., high temperatures, salt, low
pH or temperature, high radiation may lead tofinding of microbes with new capabilities to
damage or convert pollutants and make improvements in the environment. Industrial
microbiologists and biotechnologists are involved in engineering microbes to provide
solutions to the contamination and recycling problems, and evaluating the environmental
protection of new and exciting products.ConclusionIndustrial microbiology includes the use
of micro-organisms in the production of foodand other products. Within the industrial setting,
once the medium is established, the physicalenvironment for the microbial functioning in the
mass culture system must be defined. This involves the control of the specific system of
agitation, temperature and oxygenation. This is particularly true, when a dense microbial
culture is growing. Micro-organisms can be grown in culture tubes, shake flasks, stirred
fermenters or other mass culture systems. Computers arenormally made use of to monitor
outputs from probes that determine the microbial biomass, levels of critical metabolic
products or other parameters. This information is needed for process and product control.
Changes can be brought about in the environmental conditions or they can be held constant
overtime, depending on the goals of the particular process. Research has indicated that
there are number of individuals, who have made their careers in the field of industrial
microbiology. The job duties of industrial microbiologists have been identified in terms of
areas, which are, antibiotics/antimicrobials, vaccines, health-care products, food and
beverages produced by microbial activity, agriculture, enzymes, carbohydrates, organic
chemicals, contamination control, waste and wastewater management,oil recovery/mining
and environmental microbiology. Industrial microbiology has provided products that have
influenced the lives of the individuals to a major extent. The influences
have been direct and are not appreciated in number of ways. The individuals are making use
of industrial products in their daily lives. These include, agricultural products, food products,
medical products and so forth. Finally, it can be stated that in the past few years,
non-antibiotic compounds that have been used in medical and health care have made major
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contributions in bringing about well-being of human beings and other organisms, such as
plants and animals.