CBB20203 - FERMENTATION

TECHNOLOGY

ASSIGNMENT 1:
FERMENTATION PROCESS FOR
DUMMIES

PREPARED BY:
MOHD NADLY AIZAT BIN MOHD NUDRI
55202113502

MUHAMMAD NOR AZHRE SHAH BIN MALEK RAZUAN

55202113502
TABLE OF CONTENTS.
CONTENT
PAGE
1) INTRODUCTION
2) PRODUCTS DERIVED FROM FERMENTATION
3) INDUSTRIAL SCALE FERMENTATION
4) TYPES (MODES OF PROCESS)
5) ADVANTAGES AND DISADVANTAGES
6) CONCLUSION
7) REFERENCES

1.0) INTRODUCTION
The definition of fermentation in general came from Latin word fervere, which means
to boil. In the 18th century, Louis Pasteur has given a meaning that close to fermentation
which is life without air. Nout, (2014) has defined fermentation in larger scale which called
fermentation technology as the use of microorganisms to achieve desirable properties in the
bioprocessed product, fermented food or beverages.
A general definition of fermentation is an energy-yielding anaerobic metabolic
process in which organisms convert nutrients (typically carbohydrates) to alcohols and acids
(lactic acid and acetic acid). Even though the word fermentation indicates anaerobic
metabolism, which is happening in the living cells, it is also used in a broader sense to
indicate all anaerobic and aerobic microbiological and biochemical modifications that result
in desirable quality modifications of bioproducts internal compositions.
The fermentation itself is a part of the manufacturing process, consisting of several
operations that affect the microbial activity. The diversity of the industrial fermentations not
only includes typical anaerobic events, such as the lactic acid fermentation, alcoholic
fermentation, but also aerobic processes, such as alkaline fermentation and fungal
fermentation.
During the fermentation process, several metabolites were secreted by the
microorganism while utilizing the nutrients supplied. These metabolites are useful for human
consumption for various applications which have been mentioned earlier.

2.0) MECHANISM AND EXAMPLES OF PRODUCTS PRODUCED FROM THE

FERMENTATION PROCESS.
Basically, fermentation happened through a steps of process that illustrated in figure
1.

Figure 1: The general description of fermentation process. A glucose molecule is changed

anaerobically to produce ethanol and carbon dioxide with the expense of Adenosine
triphosphate (ATP). Reproduced from Amir, (n.d.).
Pyruvate is an intermediate compound in these serial steps of fermentation where the
final product that produced through fermentation is depending solely on the type of
microorganism that being involved in the reaction. The examples of microorganism and the
products produced are summarised in figure 2.

Figure 2: Examples of microorganism and the products from the fermentation process
associated with each microorganism. Reproduced from Amir, (n.d.).
The applications of the products from fermentation process are outlined in table 1.
Product
Ethanol

Organism
Saccharomyces cerevisiae

Use
Industrial solvents, beverages

Glycerol

Saccharomyces cerevisiae

Production of explosives

Lactic acid

Lactobacillus bulgaricus

Food and pharmaceutical

Acetone and butanol

Clostridium acetobutylicum

Solvents

-amylase
Bacillus subtilis
Starch hydrolysis
Table 1: The use of product formed from fermentation process that involved microorganism
respectively.

3.0) INDUSTRIAL SCALE FERMENTATION

Basically, the production line that involved fermentation process in industrial sector is
divided into three sections, which are the upstream, the main fermentation process and finally
the downstream processing (Villadsen, 2002). The upstream processing stage is where the
medium is prepared and preparation for sufficient amount of inoculum are conducted. The
main fermentation process is where the main fermentation reaction taken place within certain
period of time and known environmental conditions. While for downstream processing, this
is the stage where a series of unit operations are involved to recover, isolate, purify and
finally to polish the desired products from the reaction.
In the main fermentation process, the most important equipment that being used is a
bioreactor or also known as a fermentor. It contains the media to carry out fermentation, and
creates environment for fermentation at large scale. The size of the fermentor usually depends
on few factors such as the amount of raw materials, production demands and many more.
These are crucial factors that need to be considered during equipment selection and sizing
(Designing a bioreactor). The diagram of a bioreactor is shown in figure 3.

Figure 3: The detailed diagram of a bioreactor. Reproduced from Amir, (n.d.).

There are few factors that are crucial in order to conduct a fermentation process with
high yield. The factors are elaborated as follows:
a) Pure culture: The inoculum with sufficient amount and in a favourable physiological
state. Research under genetic engineering are conducted to improve the performance and
efficiency of the microorganism so that the yield of production could be improved.
b) Sterilized medium: Medium that contains enough nutrients to support the growth of
microorganism. Medium need to be sterilized so that no foreign microbes or germs interfere
with the fermentation process.
c) Seed fermenter: Seed fermenter is a flask shaken or to a certain extent, a small scale
bioreactor to provide enough amount of inoculum before transferred into the main bioreactor.
The amount of inoculum produced here should be around 5 10% of the main bioreactor.
d) Controlled parameters: The microenvironment of the fermentation process need to be
monitored based on the parameters such as temperature, pH, foams formed and since it is an
anaerobic process, the amount of Oxygen need to be kept to a very minimum level.

According to Chisti, (2014) there are two kind of fermentation techniques which are:
a) Surface, or also known as Solid State Fermentation (SSF).
In SSF, the microorganism are cultivated on the surface of a liquid or solid substrate. The
process itself is somehow complicated and rarely used in industry. Some of the products
produced by SSF are foods such as tempeh, bread and cocoa.

b) Submersion techniques.
For submerged fermentation, microorganism grows in a liquid medium. The process is much
simpler and widely used in the industry, ranging from foods and beverages,
biopharmaceuticals, and also wastewater treatment. Biomass, proteins, antibiotics and sewage
treatment are carried out through this process.

4.0) TYPES (MODES OF PROCESS)

4.1 Batch culture
Batch fermentation refers to a partially closed system in which most of the
materials required are loaded onto the fermenter, decontaminated before the process
starts and then, removed at the end. The only material added and removed during the
course of a batch fermentation is the gas exchange and pH control solutions. In this
mode of operation, conditions are continuously changing with time, and the fermenter
is an unsteady-state system, although in a well-mixed reactor, conditions are supposed
to be uniform throughout the reactor at any instant time. The principal disadvantage of
batch processing is the high proportion of unproductive time (down-time) between
batches, comprising the charge and discharge of the fermenter vessel, the cleaning,
sterilization and re-start process.

4.2 Continuous culture

Continuous culture is a technique involving feeding the microorganism used
for the fermentation with fresh nutrients and, at the same time, removing spent
medium plus cells from the system. An unique feature of the continuous culture is that
a time-independent steady-state can be attained which enables one to determine the
relations between microbial behavior (genetic and phenotypic expression) and the
environmental conditions.
4.3 Fed-Batch processes
The fed-batch technique was originally devised by yeast producers in the early
1900s to regulate the growth in batch culture of Saccharomyces cerevisiae. Yeast
producers observed that in the presence of high concentrations of malt, a by-product ethanol - was produced, while in low concentrations of malt, the yeast growth was
restricted. The problem was then solved by a controlled feeding regime, so that yeast
growth remained substrate limited. The concept was then extended to the production
of other products, such as some enzymes, antibiotics, growth hormones, microbial
cells, vitamins, amino acids and other organic acids. Basically, cells are grown under a
batch regime for some time, usually until close to the end of the exponential growth
phase. At this point, the reactor is fed with a solution of substrates, without the
removal of culture fluid. This feed should be balanced enough to keep the growth of
the microorganisms at a desired specific growth rate and reducing simultaneously the
production of by-products (that can be growth or product production inhibitory and
make the system not as effective).
These by-products may also affect the culture environment in such a way that
might lead to early cell death even though sufficient nutrients are available or are still
being provided. A fed-batch is useful in achieving high concentration of products as a
result of high concentration of cells for a relative large span of time. Two cases can be
considered: the production of a growth associated product and the production of a
non-growth associated product. In the first case, it is desirable to extend the growth
phase as much as possible, minimizing the changes in the fermentor as far as specific
growth rate, production of the product of interest and avoiding the production of byproducts.

For non-growth associated products, the fed-batch would be having two

phases: a growth phase in which the cells are grown to the required concentration and
then a production phase in which carbon source and other requirements for production
are fed to the fermenter. This case is also of particular interest for recombinant
inducible systems: the cells are grown to high concentrations and then induced to
express the recombinant product.
Also, considering that plasmid stability is very often guaranteed by the
presence of an antibiotic marker gene and that the lifetime of this antibiotic in a
fermenter can be limited, it might be of interest to use the fed-batch concept to feed
this same antibiotic continuously so that the presence of the plasmid in the cells is
more of a reliable fact.
Fed-batch fermentations can be the best option for some systems in which the
nutrients or any other substrates are only sparingly soluble or are too toxic to add the
whole requirement for a batch process at the start.
Finally, in fermentations such as mycelial culture the increase of viscosity with
time can be compensated by the addition of relatively small quantity of water during
the fermentation time, although the efficacy of this protocol is controversial among
researchers. Two basic approaches to the fed-batch fermentation can be used: the
constant volume fed-batch culture - Fixed Volume Fed-Batch - and the Variable
Volume Fed-Batch.

Fixed volume fed-batch

In this type of fed-batch, the limiting substrate is fed without diluting the
culture. The culture volume can also be maintained practically constant by feeding the
growth limiting substrate in undiluted form, for example, as a very concentrated liquid
or gas (ex. oxygen). Alternatively, the substrate can be added by dialysis or, in a
photosynthetic culture, radiation can be the growth limiting factor without affecting
the culture volume.

A certain type of extended fed-batch - the cyclic fed-batch culture for fixed
volume systems - refers to a periodic withdrawal of a portion of the culture and use of
the residual culture as the starting point for a further fed-batch process. Basically,
once the fermentation reaches a certain stage, (for example, when aerobic conditions
cannot be maintained anymore) the culture is removed and the biomass is diluted to
the original volume with sterile water or medium containing the feed substrate. The
dilution decreases the biomass concentration and result in an increase in the specific
growth rate. Subsequently, as feeding continues, the growth rate will decline gradually
as biomass increases and approaches the maximum sustainable in the vessel once
more, at which point the culture may be diluted again.
Variable volume fed-batch
As the name implies, a variable volume fed-batch is one in which the volume changes
with the fermentation time due to the substrate feed. The way this volume changes it
is dependent on the requirements, limitations and objectives of the operator.
The feed can be provided according to one of the following options:
(i) the same medium used in the batch mode is added;
(ii) a solution of the limiting substrate at the same concentration as that in the initial
medium is added; and
(iii) a very concentrated solution of the limiting substrate is added at a rate less than
(i), (ii) and (iii) .
This type of fed-batch can still be further classified as repeated fed-batch process or
cyclic fed-batch culture, and single fed-batch process.
The former means that once the fermentation reached a certain stage after
which is not effective anymore, a quantity of culture is removed from the vessel and
replaced by fresh nutrient medium. The decrease in volume results in a increase in the
specific growth rate, followed by a gradual decrease as the quasi-steady state is
established. The latter type refers to a type of fed-batch in which supplementary
growth medium is added during the fermentation, but no culture is removed until the
end of the batch. This system presents a disadvantage over the fixed volume fed-batch

and the repeated fed-batch process: much of the fermenter volume is not utilized until
the end of the batch and consequently, the duration of the batch is limited by the
fermenter volume.

5.0) ADVANTAGES AND DISADVANTAGES

Advantages
1.
2.

Production of high cell densities due to the extension of working of working time.
Controlled conditions in the provision of substrates during the fermentation,
particularly regarding the concentration of specific substrates as for example the carbon
source.
3. Control over the production of byproducts or catabolite repression effects due to
limited provision of substrates solely required for product formation.
4. Allows the replacement of water loss by evaporation.
5. Increase of antibiotic marked plasmid stability by providing the correspondent
antibiotic during the time span of the fermentation.
Disadvantages:
1.

It requires previous analysis of the microorganism, its requirements and the

understanding of its physiology with the productivity.
2. In a cyclic fed batch culture, care should be taken in the design of the process to
ensure that toxins do not accumulate to inhibitory levels and that nutrients other than
those incorporated into the feed medium become limiting. Also if many cycles are run,
the accumulation of non-producing or low producing variants may result.

6.0 CONCLUSION
Fermentation process is widely known and the technology is a very vibrant and fast
growing area of biotechnology, absorbing an ever increasing process and product
development. With a longer history than any area of biological sciences, fermentation
technology process has a longer and brighter future, in the service mankind, covering such
important areas such as food, medicine and supplement.

REFERENCES
Amir, H. (n.d.). Fermentation Technology. Retrieved October 4, 2015, from
http://www.slideshare.net/zeal_eagle/fermentation-technology?qid=b112605a-e9474b54-9432-85a7a6252e55&v=qf1&b=&from_search=2
Chisti, Y. (2014). Encyclopedia of Food Microbiology. Encyclopedia of Food Microbiology.
Elsevier. doi:10.1016/B978-0-12-384730-0.00106-3
Nout, M. J. R. (2014). Encyclopedia of Food Safety. Encyclopedia of Food Safety. Elsevier.
doi:10.1016/B978-0-12-378612-8.00270-5
Villadsen, J. (2002). Bioprocess engineering. Chemical Engineering Science, 57(7), 1235
1236. doi:10.1016/S0009-2509(02)00006-4