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1 INDUSTRIAL MICROBIOLOGY

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INDUSTRIAL MICROBIOLOGY
Industrial Microbiology
• Use of microbes to obtain a product or service of economic
value constitutes industrial microbiology
• Any process mediated by or involving microorganisms in
which a product of economic value is obtained is called
fermentation
– The terms industrial microbiology and fermentation are
virtually synonymous in their scope, objectives and
activities.
Microorganisms: importance in industry
Microorganisms: importance in industry
• Negative or detrimental effects:
– Causative agents of diseases
– Causing food poisoning and spoilage
– Affecting water quality
– Deterioration of materials and products
• Positive effects: produce valuable products or useful services
– Producing therapeutic compounds, foods and beverages,
additives
– Pollution control
– Alternative fuels
– Agriculture
• Microorganisms can be used 3 ways industrially
– Products of microbial metabolism
• metabolic end product, pathway intermediates,
enzymes, vitamins
– Bioconversion
• microorganisms as biocatalysts, e.g. bioremediation
– Microbial cells or cellular components
• foods or food supplements
PROPERTIES OF AN INDUSTRIAL MICROORGANISM
• Genetically stable
• Suitable for scale-up
• Easily grown
• Rapid growth rate
• Grown on relatively cheap media or waste materials
– molasses, corn steep liquor, whey, cassava, potatoes,
grapes, etc
Factors affecting growth of industrial microorganisms
• Availability of carbon and energy sources
• Ability to replicate DNA
• Size and mass of cells
• Physical and chemical environment
– media (balance between cost and maximum growth and
product formation), conditions
• Maintenance energy requirements
– energy needed to keep the cell alive but not for growth
• Culture type
– batch cultures (closed culture systems, nutrients are not
renewed)
– continuous cultures (continuous supply of substrates gives
better control over variables)
• Growth rates (increase in cells per unit time)
– cell population goes through a number of phases when
inoculated into fresh medium
PRODUCT FORMATION
i. Primary metabolites
• Product formed during the primary growth phase, e.g.
ethanol
• Growth dependent
ii. Secondary metabolites
• Products formed near the end of the growth (or stationary)
phase, e.g. antibiotics
• Not essential for growth and reproduction
• Highly dependent on growth conditions, repression frequently
occurs
• Usually produced from primary or intermediate metabolites,
not the substrate
METHODS OF FERMENTATION
• The goal of biotechnology is to obtain useful metabolic
products from biological material.
• Biotechnology encompasses two distinct phases:
– Fermentation and Product recovery
• Fermentation procedures must be developed for the
cultivation of microorganisms under optimal conditions and
for the production of desired metabolites or enzymes by the
organisms.
• Product recovery involves the extraction and purification of
biological products
PRINCIPLES OF MICROBIAL GROWTH
• The growth of organisms may be seen
– as the increase of cell material expressed in terms of mass
or cell number.
• Growth is dependent on
– the availability and transport of necessary nutrients to the
cell and subsequent uptake and
– environmental parameters such as temperature, pH and
aeration being optimally maintained.
• Cells grow in four typical phases
– lag phase, log phase, stationary phase and death phase
LAG PHASE
• No increase in the number of cells
• Time for adaptation to the new environment arising from:
– Change in the pH value
– Increase in the supply of nutrients
– Decrease in growth inhibitors
• New transport system must be induced within the cell
• Enzymes of primary metabolism must be adjusted to the new
conditions
• Physical condition of the inoculum is essential to the length of
the lag phase
LOG PHASE
• By the of end of lag phase, the cells have adapted to the new
conditions of the growth
• Microbial growth proceeds at the maximum possible rate for
that organism, with nutrients in excess, ideal environmental
parameters and growth inhibitors absent
• Growth can be described as:
– Doubling of the cell number per unit time (yeasts &
bacteria)
– Doubling of biomass per unit time (filamentous organisms
such as fungi & Actinomycetes)
• Cells alter the medium through uptake of substrates and
excretion of metabolic products
STATIONARY PHASE
• As soon as the substrate is metabolized or toxic substances
have been formed, growth slows down or is completely
stopped.
• The biomass increases only gradually or remains constant
during this phase
• Due to lysis, new substrates are released which may the serve
as energy for the slow growth of the survivors
• The various metabolites formed in this phase are of great
biotechnological interest
DEATH PHASE
• In this phase the energy reserves of the cells are exhausted
• The cells die at an exponential rate
• In biotechnological processes microorganisms in a bioreactor
can be grown in
– batch or continuous fermentation
• Within the bioreactor reactions can occur with
– static or agitated cultures
– in the presence or absence of oxygen
– in liquid or low moisture conditions (e.g. on solid
substrates)
1. BATCH FERMENTATION
– Is considered to be a closed system
– A fixed volume of sterilized nutrient solution in the
fermenter is inoculated with microorganism
• incubation is allowed to proceed under optimal
conditions
– Catalysts and medium are mixed by a rotating central shaft
that carries several impellers.
– Metabolically active microorganisms evolve heat
• Jackets that surround it provide cooling to maintain an
optimum operating temperature.
– Equipments to monitor and control the temperature and
the pH of the medium are common
– If the process is aerobic, oxygen must be supplied
continuously.
• The cheapest means of sterilization of air is
– to pass it through a sterile filter composed of glass,
wool, carbon particles, or some finely divided
material that will trap microorganism present in the
air.
– Aeration and agitation of a liquid medium can cause the
production of foam.
• medium containing high levels of proteins or peptides.
– Foam must be controlled if fermentation is to be carried
out in a proper manner.
• The usual procedure for controlling foam is to add an
antifoam agent.
– Samples of mixture can be removed for analysis by means
of other tubes and pipes.
• In the course of the fermentation, nothing is added except:
– Oxygen (in the form of air)
– An antifoam agent
– Acid or base to control the pH
• As a result of metabolism of microbial cells, the following
change constantly:
– The composition of the culture medium
– The biomass concentration
– The metabolite concentration
• In most batch processes, the biocatalyst is discarded with the
spent medium, which is potentially disadvantageous.
2. CONTINUOUS FERMENTATION
• An open system
• Are fermentations that are run continuously
– without empting the fermenter at each harvest of
microbial cells or biosynthetic products.
• Fresh nutrient solution is added to the bioreactor
continuously and
– an equivalent amount of converted nutrient solution with
microorganisms is simultaneously taken out of the system.
• Continuous methods of cultivation
– will permit organisms to grow under steady state
(unchanging) conditions
• growth occurs at a constant rate and in a constant
environment.
• Factors such as pH and the concentrations of nutrients and
metabolic products
– held near constant in continuous cultivations.
• Potential benefit of continuous methods is
– reducing losses of catalyst. In most batch processes, the
catalyst is discarded with the spent medium.
Approaches to control microbial activity in continuous culture
Turbidostat:
– the total cell population of microbial cells is held constant
• by employing a device that measures the culture
turbidity
– regulate both the nutrient feed rate to the fermenter and
the culture withdrawal rate from the fermenter.
– If the population numbers rise above a predetermined
level
• a greater amount of fresh medium is added to the
fermenter
Chemostat:
– In contrast to the turbidostat, a chemostat maintains the
nutrient feed and harvest culture withdrawal rates at
constant values.
– The growth rate is controlled by
• supplying only a limiting amount of a critical growth
nutrient in the feed solution.
• Thus, cell multiplication cannot proceed at a rate
greater than that allowed by the availability of this
critical nutrient.
Advantage of continuous fermentation
• The size of the bioreactor and other equipment used in
continuous fermentation are
– relatively smaller compared to batch fermentation for the production
of the same quality of product
• They yield of the product is more consistent since the
physiological state of the cells is uniform
• The time gap between two successive fermentations for
cleaning and preparing the bioreactor for reuse is avoided in
continuous fermentation
• Potential benefit in reducing losses of catalyst
Disadvantages of continuous fermentation
• Maintaining sterile conditions on an industrial scale over a
longer period of time is difficult
• It is not to maintain the same quality of the culture medium
for all the additions.
– Nutrient variations will alter the growth and physiology of the cells
and consequently the product yield
• While using high-yielding strains, reverse mutants arise, which
can overgrow the production strains in continuous culture
SOLID SUBSTRATE FERMENTATION
• Is a fermentation processes that do not involve liquid
medium.
– growth of the microorganisms is carried out on solid
substrates in the absence or near absence of free water
• The presence of moisture (about 15%) is necessary for solid
state (solid substrate) fermentation
• The most regularly used solid substrates are
– cereal grains, legume seed, wheat grain, lignocellulose
materials such as straws, sawdust or wood shavings
• Most of this compounds are cheap, easily obtainable and
represents a concentrated source of nutrients for microbial
growth
• There is a need to pre-treat the substrate raw material to
facilitate the availability of the bound nutrients.
Advantages of solid substrate fermentation
– Simple cheaper substrate
– Low technology, low energy expenses
– Low moisture content of materials gives economy
of bioreactor space
– No need for sterilization
– Easier downstream processing
– Yield of product can be high
• Disadvantages of solid substrate fermentation
– Process limited mainly to moulds that tolerate low
moisture levels
– Slower growth rate of microorganisms
– Process monitoring, e.g. moisture levels, oxygen
and carbon dioxide levels is difficult to achieve
accurately
FERMENTATION EQUIPMENT AND ITS
USE
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