2. CONTENTS
Introduction of fermentation
Historical background
Types of fermentation (Alcoholic, Lactic acid ,Propionic , Butanol fermentation)
Mechanism of fermentation
Products of fermentation
Fermentation as a food preservative
Advantages and disadvantages of fermentation
Importance and application of fermentation
Bioreactors
4. Fermentation is a metabolic process
that produces chemical changes in
organic substrate through the action
of enzymes. In biochemistry, it is
narrowly defined as the extraction of
energy from carbohydrates in the
absence of oxygen.
In the context of food production, it
may more broadly refer to any process
in which the activity
of microorganisms brings about a
desirable change to a foodstuff or
beverage.The science of fermentation
is known as zymology.
INTRODUCTION
5. In microorganisms, fermentation is the
primary means of producing adenosine
triphosphate (ATP) by the degradation
of organic
nutrients anaerobically.Humans have
used fermentation to produce foodstuffs
and beverages since the Neolithic age.
For example, fermentation is used for
preservation in a process that
produces lactic acid found in such
sour foods as pickled
cucumbers, kombucha, kimchi,
and yogurt, as well as for producing
alcoholic beverages such as
wine and beer..
Fermentation also occurs within the
gastrointestinal tracts of all animals,
including humans.
9. HISTORY CONT.
The use of fermentation, particularly
for beverage, has existed since
the Neolithic and has been documented
dating from 7000–6600 BCE
in Jiahu, China,5000 BCE in India,
Ayurveda mentions many Medicated
Wines, 6000 BCE in Georgia, 3150 BCE
in ancient Egypt, 3000 BCE
in Babyl,2000 BCE in pre-Hispanic
Mexico, and 1500 BC
in Sudan. Fermented foods have a
religious significance
in Judaism and Christianity. The Baltic
god Rugutis was worshiped as the agent
of fermentation.
10. In 1837, Charles Cagniard de la
Tour, Theodor Schwan and Friedrich
Traugott Kützing independently
published papers concluding, as a result
of microscopic investigations, that yeast
is a living organism that reproduces
by budding. Schwann boiled grape juice
to kill the yeast and found that no
fermentation would occur until new
yeast was added. However, a lot of
chemists, including Antoine Lavoisier,
continued to view fermentation as a
simple chemical reaction and rejected
the notion that living organisms could
be involved. This was seen as a reversion
to vitalism and was lampooned in an
anonymous publication by Justus von
Liebig and Friedrich Wöhler.
11. The turning point came when Louis
Pasteur(1822–1895), during the
1850s and 1860s, repeated
Schwann's experiments and
showed that fermentation is
initiated by living organisms in a
series of investigations. In 1857,
Pasteur showed that lactic acid
fermentation is caused by living
organisms.In 1860, he
demonstrated that bacteria
cause souring in milk, a process
formerly thought to be merely a
chemical change, and his work in
identifying the role of
microorganisms in food spoilage
led to the process of pasteurization
12. Many scientists, including Pasteur,
had unsuccessfully attempted to
extract the
fermentation enzyme from yeast. Su
-ccess came in 1897 when the
German chemist Eduard
Buechner ground up yeast,
extracted a juice from them, then
found to his amazement that this
"dead" liquid would ferment a sugar
solution, forming carbon dioxide
and alcohol much like living
yeasts. In 1907, Buechner won
the Nobel Prize in chemistry for his
work.
14. ALCOHOLIC FERMENTATION
Alcoholic fermentation generally means production of
ethanol (CH3CH2OH). Commonly yeasts, particularly
Saccharomyces cerevisiae, are used for production of
various alcoholic beverages, as well as industrial alcohol.
Yeasts are essentially aerobic organisms, but they can also
grow as facultative anaerobes.
The energy-yield under anaerobic conditions is much lower
and hence the growth is slower with much lower cell-yield.
When grown with aeration, the cell-yield increases
dramatically, but alcohol production falls. Thus, oxygen
inhibits fermentation. This is known as Pasteur-effect.
15. Conversion of pyruvic acid to ethanol proceeds in
two steps: pyruvic acid to acetaldehyde and
acetaldehyde to ethanol. The first step is catalysed
by pyruvic acid decarboxylase which requires TPP
as coenzyme, and the second step by alcohol
dehydrogenase which requires NADH2 as
coenzyme.
NADH2 is thereby oxidized to NAD which can
be reused for reduction of GAP to DPGA in the
EMP:
16. For example, for production of baker’s yeast used in bread
industry, strongly aerated cultures favour large cell-yield
with little or no alcohol. Extract of malted (partly
germinated) barley serves as substrate for beer production.
The starting material contains large amount of maltose (a
dissacharide of two glucose units) produced by hydrolysis
of starch present in barley seeds. Maltose is split into
glucose and serves as substrate for alcohol fermentation
under anaerobic conditions.
Similarly, for production of wine, grape juice is the
substrate of choice. Specific selected strains are employed
to impart characteristic flavour and taste of different
alcoholic beverages. For manufacture of industrial alcohol,
generally molasses is used as the starting material. Also
sulfite liquor, which is a waste product of paper industry, is
used as a cheap substrate for industrial alcohol production.
17. LACTIC ACID FERMENTATION
Lactic acid bacteria are both morphologically and
physiologically diverse. The lactic cocci, previously included in
the genus Streptococcus, have been transferred to the genus
Lactococcus. The rod-shaped lactic acid bacteria are distributed
in several genera, though majority are placed in the genus
Lactobacillus. Some representative species of homo-fermentative
lactic acid bacteria are Lactococcus lactis, L. cremoris, L.
diacetilactis, L. thermophilus, Lactobacillus lactis, L. bulgaricus,
L. acidophilus etc. Representatives of heterofermentative type
include Lenconostoc mesenteroides, Lactobacillus brevis,
Bifidobacterium bifidum etc. There is also a spore-forming lactic
bacterium, Sporolactobacillus. The lactic acid bacteria prefer
anaerobic conditions for optimal growth as they do not have
cytochromes or catalase, though they can also grow in
microaerophilic environment.
18. Homo-fermentative
In the first type, lactic acid is produced as the sole product by
reduction of pyruvic acid with the help of the enzyme lactic acid
dehydrogenase. The reaction regenerates NAD from
NADH2 which is reused for oxidation of GAP to DPGA in the
glycolytic pathway.
As one molecule of lactic acid is formed from one molecule of
pyruvic acid, two molecules of lactic acid are produced from
each molecule of glucose, when it is dissimilated through EMP
Homolactic fermentation is the simplest of all fermentations,
involving only a single step in which pyruvic acid is reduced to
lactic acid. Lactic acid is formed also in muscles by a similar
reaction.
19. Hetero-fermentative
In heterofermentative type, the products are lactic acid and ethanol or acetic
acid and CO2. The heterofermentative lactic acid bacteria dissimilate glucose
via PPC. They produce lactic acid from one-half of the glucose molecule, and
ethanol or acetic acid and CO2 from the other half.
The heterofermentative lactic acid bacteria lack two vital enzymes of the
glycolytic pathway — aldolase and triose phosphate isomerase. Hence, they are
unable to use EMP. As an alternative, they employ the pentose phosphate
pathway. An intermediate of this pathway is xylulose 5-phosphate.
The heterofermentative bacteria cleave xylulose 5-phosphate by a TPP-linked
pentose phosphate ketolase into glycerin aldehyde phosphate (GAP) and acetyl
phosphate. GAP is then converted to pyruvic acid by the usual EMP enzymes,
while acetyl phosphate is reduced either to acetic acid or to ethanol. From
pyruvic acid, lactic acid is formed by the lactate dehydrogenase activity.
Leuconostoc mesenteroides produces from one molecule of glucose, one
molecule of lactic acid, one molecule of ethanol and one molecule of CO2. On
the other hand, Lactobacillus brevis produces acetic acid in place of ethanol.
20. Uses of lactic acid fermentation
Lactic acid bacteria are widely used for production of various
fermented food throughout the world. The bacteria ferment the milk
sugar (lactose) to produce lactic acid which curdles milk protein.
Various species are used to yield products of variable consistency, taste
and aroma. In different countries the products are variously known as
yogurt in Europe and America, dadhi or dahi in India, Kefir in Russia,
Kumiss, butter milk, acidophilus milk etc.
Lactic acid bacteria are also employed in producing fermented
vegetable products, like sauerkraut (fermented cabbage), cucumber
pickles and fermented olive. These bacteria are also used for
production of sausages from beef and pork.
Both heterofermentative and homo-fermentative lactic acid bacteria
are used as ‘starter’ for production of fermented food. Some of these
bacteria are Lactococcus cremoris, L. lactis, L. thermophilus,
Lactobacillus bulgaricus, L. plantarum, L. brevis, Leuconostoc
mesenteroides, Pediococcus cerevisiae etc.
21. The propionic acid bacteria dissimilate glucose via
EMP and produce pyruvic acid. By a biotin- linked
carboxylation reaction pyruvic acid is converted to
oxalacetic acid which is then reduced in two steps to
succinic acid through reversal of TCA cycle reactions.
Succinic acid is then converted to succinyl-CoA, also by a
reverse step of the TCA cycle. Next, succinyl-CoA produces
methyl malonyl- CoA by the action of a vitamin B12-linked
enzyme methyl malonyl mutase which catalyses an intra-
molecular rearrangement. Methyl malonyl-CoA is then
decarboxylated to propionyl-CoA.
In the final step, propionyl-CoA yields propionic acid, and
CoA is transferred to succinic acid by an enzyme, CoA-
transferase.Together with lactic acid bacteria, the propionic
acid bacteria are used for commercial production of Swiss
cheese. Propionic acid contributes to the special flavour of
this cheese.
22.
23. BUTYRIC ACID FERMENTATION
The bacteria carrying out butyric acid-butanol fermentation are
all obligately anaerobic spore- forming bacteria belonging to the
genus Clostridium. Besides butyric acid and n-butanol, several
other products of this fermentation are acetic acid, ethanol,
isopropanol and acetone depending on species.
For example, C. butyricum, C. lactoacetophilum, C.
pasteurianum etc. produce butyric acid together with acetic acid,
while C. butylicum and C. acetobutylicum produce butyric acid,
acetic acid and isopropanol or acetone. Also, as a fermentation
product, CO2 is always present.
Clostridia dissimilate glucose by the EMP to form pyruvic acid
which by decarboxylation produces acetyl-CoA.
The latter acts as the key intermediate in the butyric-
butylic fermentation and gives rise to all the products by
different pathways
24.
25. MECHANISM OF FERMENTATION
Fermentation takes place when the electron transport chain is
unusable (often due to lack of a final electron receptor, such as
oxygen), and becomes the cell’s primary means of ATP(energy)
production. It turns NADH and pyruvate produced in glycolysis
into NAD+ and an organic molecule(which varies depending on
the type of fermentation). In the presence of O2, NADH and
pyruvate are used to generate ATP in respiration. This is called
Oxidative phosphorylation.
26. Oxidative Phosphorylation generates much more energy that is
ATP .For that reason, cells generally benefit from avoiding
fermentation when oxygen is available, the exception being
obligate anaerobes which cannot tolerate oxygen.
The first step which is gylcolysis is common to the
fermentation process and pathways as well.
Therefore the Glycolysis is the cause of fermentation
C6H12O6 + 2 NAD+ + 2 ADP + 2 Pi → 2 CH3COCOO− + 2NADH
+ 2 ATP + 2 H2O + 2H+
The overall reaction of fermentation is mentioned above.
27. Pyruvate is CH3COCOO−. Pi is inorganic phosphate. Two ADP
molecules and two Pi are converted to two ATP and two water
molecules via substrate-level phosphorylation. Two molecules of
NAD+ are also reduced to NADH.
In oxidative phosphorylation the energy for ATP formation is
derived from an electrochemical proton gradient generated across
the inner mitochondrial membrane (or, in the case of bacteria, the
plasma membrane) via the electron transport chain. Glycolysis has
substrate-level phosphorylation (ATP generated directly at the
point of reaction).Humans have used fermentation to produce
food and beverages since the Neolithic age.
28. For example: fermentation is used for preservation in a
process that produces lactic acid as found in such sour foods
as pickled cucumbers, kimchi and as well as for producing
alcoholic beverages such as wine (see fermentation in
winemaking) and beer. Fermentation can even occur within
the stomachs of animals, such as humans.
31. Fermentation as a food
Preservation
technique:-
Introduction:-
The deliberate fermentation by man Predates
written history and is possibly the oldest method for preserving perishable
foods.
Evidence suggests that fermented food were consumed About 7000 years
ago in Babylon.
Why is fermentation used to preserve food?
Preserving by fermentation not only makes foods available for future use, but
more flavorful.
The nutritional value produced by fermenting is another benefit of fermention.
Increased enzyme content helps us absorb nutrients reducing the need of vitamins
and supplements.
Fermentation leads to predigestion of the food product thus making it easily
digestible by us.
32. How fermentation
preserves food?
Due to fermentation, bacteria causes less
deterioration of the food as it inhibits their
growth.
Some fermentation process lower the pH
of the food material making it undesirable for micro-
organisms to thrive in.
Controlled fermentation encourage the growth of good
bacteria that outgrows the harmful microbes.
33. Examples of
fermented food and
their benefits:-
Fermentation enhances the flavour in case of
black tea, wine, beer, aged cheese, cocoa beans etc.
The sprouting of grain, seeds and nuts multiplies
amino acid, vitamins, mineral content and
antioxidant qualities of starting material.
Due to change in chemical properties due to
Fermentation, a poisonous plant, cassava can be
made edible.
Fermented dairy products like yoghurt, kifir,
cheese aid digestion and also helpful in case of
lactose intolerance and autism.
Poridge made from grain after fermentation is rich
in nuritional value and reduces risk of disease in
children.
Vinegar is used to leach out flavour and certain
compound from food material to make it tasty and
healthy.
Probotic supplements ( bacterial culture to
maintain microbial balance in body) is know to
help in curing diseases like cancer.
Table containing list of fermented
products,their raw material and
microbes responsible :-
34. Method of Preserving using
Fermentation:-
The Fermentation begins with 'starter culture’
with preferred bacteria introduced in the food
to be fermented. This can be done by adding a
small sample from the previous batch of
already fermented food or with commercially
distributed mixture.
Naturally Fermented:- Materials cured in
brine solution (salt, sugar or spices) are said to
be naturally fermented or brined. Brine
protects the vegetables from aerobic bacteria.
Not naturally Fermented:- Vinegar is used
to preserve in this type. It Imparts flavour of
herbs and spices to the food and its acidity
preserve it. It doesn’t ferment food but it is a
product of Fermentation.
35. Advantages of
Fermentation:-
It increases shelf life of the food.
Enhances texture, flavour and odour of food.
New energy sources and high vitamin content.
Makes food easily digestible and nutrituous.
Decrease toxicity and ailments
Restores proper bacterial balance in intestine with the help of
Probotic supplements.
Protects liver from diseases like hepatitis C and liver tissue
damage.
Improves immune system.
Simple technology and easy to perform.
This forms a natural environment for microbes to grow hence
productivity is high.
36. Disadvantages of
Fermentation:-
Fermentation on large scale is expensive and difficult.
Chances of contamination is high.
Botulism contamination is common in home preserved
food.
Controlling parameters like pH, temperature etc. is
essential and difficult.
Heat build up is a common problem.
Higher impurity product is formed hence purification cost
is higher.
Varying quality.
Not portable.
38. Biological role: Anaerobic
respiration/fermentation is the only process
that is common in all prokaryote and
eukaryotes. For prokaryote, it is the only
means of producing energy while for
eukaryotes, fermentation is useful when
oxygen supply is less. Thus even under low
oxygen supply, fermentation can help in
production of energy even if it’s low and
Fermentation utilises end product of
glycolysis and hence keeps the cycle running.
39. Food products: Fermentation is carried
out to produce a number of food product
that are consumed daily such as:
1. Bread: Saccharomyces Cervidae/Yeast, a
fungus is used to carry out fermentation
to produce bread, the Yeast is hence also
know as baker’s yeast. Yeast breaks down
the sugar present in the dough mixture
and releases carbon dioide thereby
making the dough rise.
2. Preservation of food products: Lactic
acid fermentation is carried out to
preserve food products for a long time as
in yoghurt, kimchi, sauerkraut, pickles
etc.
40. 3. Vinegar: Acetic acid fermentation is
carried out to convert starch/sugar present
in fruits(apple) or grains(rice) to produce
apple cider vinegar or sour vinegar.
4. Fermentation is widely used for the
production of alcoholic beverages, for
instance, wine from fruit juices and
beer from grains. Potatoes, rich in
starch, can also be fermented and
distilled to make gin and vodka.
41. 5. Cheese: Cheese is manufactured by fermentation of milk,
which leads to coagulation of milk thereby retaining protein
content of milk. Cheese production is a major industry. Few
of them are listed below:
S.No Type of cheese Organism used
1 Brick cheese Lactobacillus lactis
2 Swiss cheese Streptococcus
thermophilus
3 Blue cheese Brevibacterium
linens
4 Roquefort cheese Penicillium
roqueforti
5 Gorgonzola cheese Penicillium glaucum
6 Monterey cheese Streptococcus
cremoris
Table 1. Types of cheese and microorganism used
42. Table 2. Few food products, their raw
material and microorganism used
43. Industrial application:
1. Production of organic acids: Organic acids such as
Acetic acid, Butyric acid etc are produced by
fermentation using bacteria and fungi
S. No Organic acid Microorganism used
1 Acetic acid Aetobacter aceti
2 Butyric acid Lactobacillus butyricum
3 Citric acid Aspergillus niger
4 Fumaric acid Rhizopus nigericans
5. Gibberellic acid Fusarium monoliformae
Table 3. Organic acid and organism
2. Production of vitamins and enzymes: Many of the
vitamins such as thiamine(vitamin B1), riboflavin(Vitamin
B1), Cobamaline (vitamin B1), Acetic acid(Vitamin C) etc are
produced industrially by fermentation in microbe.
44. Seventy percent of enzymes are made from Bacillus
bacteria via fermentation.Most commercial microbial
enzymes are hydrolases, which break down different
organic molecules such as proteins and lipids. The enzyme
glucose isomerase is important in the production of fructose
syrups from corn and is widely used in the food industry.
45. 3. Chemicals:Numerous chemicals, such as amino acids,
polymers, organic acids (citric, acetic, and lactic), and
bioinsecticides are produced by industrial fermentation.
Amino acids are used as a food and animal feed, as well as in
the pharmaceuticals cosmetic, and chemical industries.
Bacteria such as Micrococcus luteus and Corynebacterium
glutamicum are used for industrial fermentation to produce
chemicals. Bacterial toxins are effective against different
insects. Since the 1960’s, preparations of the bacteria
Bacillus thuringiensis have been produced by fermentation as
a biological insecticide
4. Biofuel: Ethanol, one of the major byproducts of
fermentation, is widely used as fuel to drive machines as well as
automobiles in many countries such as United States.
Biogas produced by Anaerobic fermentation in bacteria such as
Methanococcus us another source of biofuel and is widely used
in rural parts of India
46. Bioreactor
Bioreactor are specially designed
large vessels in which fermentation
is carried out. Main purpose of
fermentor is to provide suitable
controlled environment for the
growth of microorganisms. It is
specialised in such a way that pH,
temperature, etc can be monitored
in the system.
Fermentor are designed in such a
way that there is scope of proper
aeration(oxygen supply) and
stirring(mixing of contents)
Fig. Diagrammatic
representation of a
bioreactor
