Batch fermentation
Batch fermentation
Batch culture is defined as an operational technique in biotechnological
processes where all the required nutrients (substrates) are supplied to the
bioreactor during cultivation and in which the product(s) remain in the
bioreactor until the end of the run.
Batch cultivation is closed system where there is no interaction between
the system and the surrounding during the process.
Except air during the aerobic cultivation.
In Batch cultivation
we prepare medium,
sterilize it and
inoculate the culture into the bioreactor.
Allow the cells to grow and produce the product.
Once the product formation reaches maximum harvest the fermentation
broth.
Batch fermentation is carried out in batches.
• Substrate and microorganism are loaded into the fermenter batchwise.
• Seed culture is needed for each new batch.
Fermentation media is filled up to 80% space by fermentor, and the
remaining space is used as head space.
Head space plays an important role as some area of a fermentor is
required for collection of air, gases, and foam which is produced during
the fermentation process.
After inoculation of media the fermentor is steam sterilized
After sterilization, the nutrient media is cooled and inoculated with desired
volume of inoculums under aseptic condition.
Fermentation process is carried out under optimum growth condition.
It is stopped after specific period of time and the fermented media or broth
are removed from the fermentor and the desired product is obtained.
The product obtained is passed through recovery and purification process.
• Later, the fermentor is cleaned and reused for the next batch.
• In this process, as the fermentation proceeds, the quantity of nutrients
from the media gets depleted, and microbes and products increase.
• In batch fermentation, growth of micro-organism is slower down due to
decrease of nutrients.
This is the most popular and simple method for ethanol production.
A higher initial sugar concentration is required in order to achieve more
efficient ethanol production;
A high sugar concentration will inhibit the growth and function of
fermenting microorganisms due to excessive osmosis to result in a low
fermentation yield with a prolonged fermentation period.
The composition of the culture medium, the biomass concentration, and the
metabolite concentration generally change constantly as a result of the
metabolism of the cells.
After the inoculation of a sterile nutrient solution with microorganisms and
cultivation under physiological conditions, four typical phases of growth
are observed
• Growth Phases
• Lag phase
• Physicochemical equilibration between microorganism and the
environment.
• Log phase
• Growth of the cell mass can now be described quantitatively as a
doubling of cell number per unit time for bacteria.
• Stationary phase
• As soon as the substrate is metabolized or toxic substances have been
formed, growth slows down or is completely stopped.
• Death phase
• In this phase the energy reserves of the cells are exhausted.
Advantages of batch fermentation
low investment costs,
simple control and operations, and
easy-to-maintain complete sterilization.
It requires less space,
there are fewer chances of contamination and this process is easy to
handle.
Reduced risk of contamination or cell mutation as the growth period
is short.
Lower capital investment when compared to continuous processes for
the same bioreactor volume.
More flexibility with varying product/biological systems.
Higher raw material conversion levels, resulting from a controlled
growth period.
• The disadvantages include:
The disadvantage is that it is a time consuming process
it requires more time for cleaning, sterilization, cooling.
The yield of the product is also low.
Lower productivity levels due to time for filling, heating, sterilization,
cooling, emptying and cleaning the reactor.
Increased focus on instrumentation due to frequent sterilization.
Greater expense incurred in preparing several subsultures for
inoculation.
Higher costs for labour and/or process control for this non-stationary
procedure.
Larger industrial hygiene risks due to potential contact with
pathogenic microorganisms or toxins.
• How cells grow during Batch cultivation
• After inoculating the medium and start measuring the biomass at
different time intervals, you may find six different phases.
• They are
[Link] phase
[Link] growth phase
[Link] growth phase
[Link] growth phase
E. Stationary phase
F. Death phase
Lag phase
• Lag Phase is an initial period of cultivation during which the change of cell
number is zero or negligible.
• Short period immediately after inoculation
• Organisms are synthesising the enzymes needed to exploit the new
medium.
• Eventhough the cell number doesnot increase(usually), the cells may grow
in size during this period.
• The Lag phase results from several factors.
• When cells are placed in the fresh medium,
• the intracellular concentrations of cofactors, amino acids, ions will
decrease and
• these have to be synthesized / transported first before cell division to
occur.
• When the cells are inoculated into medium containing different carbon
source then the enzymes for its metabolism have to be transported.
• (If organisms have been transferred from an identical medium, at the same
temperature, the lag phase may be very short)
• When cell are placed in medium containing several carbon sources then
several lag phases may result. This is known as diauxic growth.
• When glucose and lactose are present then glucose will be utilized first
then lactose.
• Presence of glucose will have catabolite repression on galactosidase
enzyme which is required for lactose utilization
• How to avoid lag phase
• Lag phase is non productive period in the industrial fermentations. Hence
minimizing it is essential.
• The stage of culture from where the inoculum is drawn is important.
• Exponentially growing cells will have adequate concentrations of
intermediates and intracellular pool of compounds.
• Hence if the inoculum is drawn from this stage they will not suffer
dilution effect.
• Size of the inoculum
– If the size of the inoculum is large then the lag phase can be
minimized
• Generally 10% are used for yeast and mold and 5% for bacteria.
• Medium of inoculum should be same of that production medium.
• In certain cases such as recombinant [Link] cultivation to minimize
plasmid loss higher percentage of inoculum will be used.
• Accelerated growth phase
• At the end of Lag phase, when growth begins the division rate
increase gradually and reaches a maximum value.
• The sp growth rate increases to maximum during this phase.
Exponential phase (log phase)
• Organisms have started to reproduce
• Abundant food,
• absence of competition and of inhibiting factors such as toxic
waste products
• allows cell division to occur at a high rate.
• The population doubles at regular intervals.
• Growth of the cell mass can now be described quantitatively as a
doubling of cell number per unit time (yeast and bacteria)
or
• a doubling of biomass per unit time (filamentous organisms such as
streptomycetes and fungi).
• By plotting the number of cells or biomass against time on a
semilogarithmic graph, a straight line results, hence the term “log
phase”.
• Although the cells alter the medium through uptake of substrates and
excertion of metabolic products, the growth rate remains constant during
the log phase.
• Growth rate is independent of substrates concentration as long as excess
substrate is present.
Decelerated growth phase
• The end of the exponential phase occur when any of the essential
nutrients is depleted or toxic metabolite accumulated in the system.
• During this phase the growth rate declines.
• Stationary phase will follow this phase.
• Stationary phase
• In a closed culture food soon runs out or the organisms become poisoned
by their own accumulated wastes
• As a results growth slows down and stops.
• If the population is being measured by a total cell count, or by turbidity
readings, there may be no apparent further change.
• The length of stationary phase may vary with cell type, previous
growth conditions etc.,
• In certain cases the product formation will occur during this phase
• Death phase:
• Following this is the death phase where the cells will start to lyse and
the cell density decreases.
• Environmental conditions affecting cell growth
• Substrate concentration
• Temperature
• pH
• Dissolved oxygen
• Others
• Redox potential
• Ionic concentrations
• Dissolved Carbondioxide concentration