BIOTECHNOLOGY

SEMINAR ON
CITRIC ACID
PRESENTED BY:
BHAVYA RAJ
DEEPAK C V
ARSH UNIYAL
CH01
HISTORY 17th-18th Century: Lemon juice
was recognized for its ability to
prevent scurvy among sailors. This
was due to its high vitamin C
Early Observations: Citric acid's
content, which is a form of citric
natural occurrence in citrus
acid.
fruits like lemons, oranges, and
limes was known for centuries.
Ancient civilizations, including 18th Century: Chemists began
Greeks and Romans, were aware isolating citric acid from lemon juice.
of the acidic properties of citrus The first recorded isolation of citric
fruits. acid was by the Swedish chemist Carl
Wilhelm Scheele in 1784. He extracted
the acid from lemon juice and named it
"acid of lemon."
 What Is Citric Acid?
● Citric Acid is a poor organic acid but a strong organic
compound. It is generally found in citrus fruits. This acid is
usually found in fruits and vegetables such as oranges,
tomatoes, lemons, pomelo, etc.
● Citric acid 2-hydroxy-propane-1,2,3-tricarboxylic acid
Industrial Production:
Industrial-scale citric acid production first began in 1890 based on
the Italian citrus fruit industry, where the juice was treated with
hydrated lime (calcium hydroxide) to precipitate calcium citrate,
which was isolated and converted back to the acid using diluted
sulfuric acid.
In 1893, C. Wehmer discovered Penicillium mold could produce
citric acid from sugar. However, microbial production of citric acid
did not become industrially important until World War I disrupted
Italian citrus exports.
In 1917, American food chemist James Currie discovered that
certain strains of the mold Aspergillus niger could be efficient citric
acid producers, and the pharmaceutical company Pfizer began
industrial-level production using this technique two years later.

● Post-World War II: Citric acid's usage expanded significantly after

World War II due to its diverse applications in food,
pharmaceuticals, and other industries.
Citric Acid Properties
Some properties of citric acid are:
1. It is a weak organic acid.
2. The molecular formula of citric acid is C₆H₈O₇.
3. It is a colourless, odourless, and tasteless crystalline compound.
4. The structure of citric acid crystals is monoclinic.
5. The molar mass of citric acid is 192.124g/mol.
6. Its density is 1.665g/cm3.
7. The boiling point of citric acid is 310°C, while its melting point is 153°C.
8. Other than water, it is soluble in ether, alcohol, and acetone.
9. At room temperature, it exists in the white crystalline powder form.
APPLICATIONS OF CITRIC ACID
Citric acid has a wide range of applications in the industrial world and in our
day to day lives.

● Due to its pleasant taste, low toxicity and excellent palatability, citric acid
is widely used in industry for the preparation of food and sugar
confectionery (21% of total production) and beverages (45%).
● pharmaceutical and detergent/cleaning industry
● stabilisation of oils and fats or ascorbic acid against metal ioncatalysed
oxidation.
● useful as a buffer over a broad range of pH values
● non-toxic plasticisers
● trisodium citrate may be used as a blood preservative
Biochemical pathways of citric acid accumulation
B
 Microbial strains used for citric acid
accumulation
Several microbial strains are used for the industrial production of citric acid through fermentation. Among them, the fungus
Aspergillus niger is the most widely employed due to its high citric acid yield and efficiency. Here are some examples of the
microbial strains commonly used for citric acid accumulation:

1. Aspergillus niger: 2. Yarrowia lipolytica:

3. Saccharomyces cerevisiae: 4. Candida species

REGULATIONS OF CITRIC ACID ACCUMULATION

Citric acid accumulation is observed under a set of strictly controlled nutrient conditions .

The conditions required for optimum yields vary with the type of fermentation

Accumulation of Citric acid depends on the following factors:-

● Sugar type and concentration

● Trace metal ions
● pH
● Dissolved O2 tension
● Effect of Nitrigen sources
 SUGAR TYPE AND CONCENTRATION

● sugars that are rapidly catabolised by the fungus -- such as sucrose, maltose or glucose.
● beet and cane molasses are most common carbon sources
● Concentration of carbon source used is very high (100 to over 200 g l−1)
● Very little citric acid is produced at sugar concentrations below 50 g l−1.
1. Trace metal ions
● trace metal ions such as Fe, Zn, Cu, Mn, Co are essential for A. niger growth
● Excess traces of metal ions also affects the production rate. Mn2+ conc even high as 2 µg
l−1 will decrease citric acid accumulation by about 20%
● Therefore purification of carbon sources by precipitation after complexation with
hexaferrocyanide or cation exchange treatment is required.

2. pH
● Significant amount of citric acid is accumulated when pH is below 2.5.
● This prevents the production of other acids like gluconic acid and oxalic acid

3. Dissolved O2 tension
● Accumulation of citric acid is dependent on strong aeration.
● Dissolved O2 tension higher than that required for vegetative growth of A. niger is required.

4. Effect of Nitrogen and Phosphate

● Ammonia salts, nitrates and urea are the nitrogen sources used.
● The concentration of the phosphate source is usually kept low in the media
 PRODUCTION PROCESSES FOR CITRIC ACID
There are three main production processes for citric acid.
● Surface process

Older and more labour-intensive version of citric acid fermentation

● Submerged process

This is the preferred method for industrial citric acid production on a large scale due
to ease of control and better growth of microbes.

● Solid state fermentation (koji process)

Carried out in the absence of free-flowing liquid using a non soluble material that
acts as both a solid support and a source of nutrients

There are also some small scale production processes employed regionally in some parts of the world
like Japan, China and some contries in South-East Asia.
 Surface Fermentation Process
● Liquid surface fermentation
● The process is carried out in the fermentation chamber which
provides uniform air circulation, maintains relative
temperature and humidity.
● It helps control humidity and temperature by evaporative cooling
since the chamber needs to be well ventilated. The below figure
shows the apparatus for surface fermentation.

First, the spores of A. niger is blown onto the

surface of the culture medium for about 5-6 days
and then dry air is passed.
The growth of white mycelium is observed on the
surface of the culture medium.
In the surface culture process, a small amount of
citric acid is produced as the primary metabolites
by the A. niger.
Submerged Fermentation Process
● Accounts to about 80% of the citric acid production
● Performed in stirred tanks or aerated tower fermenters made of stainless steel compiled
with proper aeration, cooling system, impellers
● Involves cultivating microorganisms, typically strains of the fungus Aspergillus niger, in a
liquid medium
● Inoculation is performed by adding a suspension of spores
● Mycelial development - Germinating spores aggregate to small (0.2--0.5 mm) pellets which
has a firm, smooth surface.
● The citric acid is recovered by the recovery process
● A final yield of 0.8--0.9 kg kg−1 is obtained after 7 to 10 days
Solid State Fermentation Process for Citric acid production
1. Substrate Selection: The process begins with the selection of a suitable substrate, such as sugarcane bagasse, rice bran, or
other agricultural residues, which will serve as the solid medium for the fermentation.

2. Inoculum Preparation: A suitable strain of Aspergillus niger, a fungus commonly used in citric acid production, is selected
and prepared for inoculation onto the solid substrate. The inoculum may be prepared by growing the fungus on a liquid medium and then
transferring it to the solid substrate.
3.
4. Fermentation Conditions: The solid-state fermentation is carried out under controlled conditions of temperature,
moisture, and aeration to promote the growth of the fungus and the production of citric acid. Typically, the process is carried out at
temperatures between 25-30°C and with moisture levels around 50-70%

5. Fermentation Period: The fermentation process typically lasts for several days to allow the fungus to grow and produce citric
acid. During this time, the solid substrate is periodically mixed to ensure uniform growth and distribution of the fungus.

6. Citric Acid Extraction: After the fermentation period, the solid substrate is processed to extract the citric acid. This may
involve methods such as solvent extraction, filtration, and purification to obtain the pure citric acid product.

7. Product Drying: The extracted citric acid is then dried to remove any remaining moisture and obtain the final product in a
powdered form.
RECOVERY PROCESS OF CITRIC ACID
● Filtration of culture broth and thorough washing of the mycelial cake
● Filtration of mycelium from submerged process often requires the use of filter aids
due to the formation of a slimy heteropolysaccharide protective layer
● Lime at pH 3 is added to the broth to precipitate any oxalic acid.
● Recovery of citric acid from the broth is then generally accomplished by three basic
procedures:
a. Precipitation
b. Extraction
c. Ion exchange adsorption
● Crystallisation of citric acid is finally performed in vacuum crystallisers.
Citric Acid Flow Chart
 CONCLUSION

In conclusion, citric acid stands as a remarkable compound with a rich history and
multifaceted significance in our lives and industries. From its early discovery in
citrus fruits to its industrial production using microbial fermentation, citric acid has
played a pivotal role in shaping our understanding of chemistry, nutrition, and
manufacturing processes

As we continue to innovate and explore new frontiers, citric acid remains a staple
compound with ever-evolving possibilities, offering a bright future of continued
discoveries and applications.
THANK YOU