INDUSTRIAL BIOTECHNOLOGY

Lecture 31: Citric acid fermentation

PROF. DEBABRATA DAS

DEPARTMENT OF BIOTECHNOLOGY
IIT KHARAGPUR

1
Citric acid is a tricarboxylic acid

Prof. Debabrata Das

Department of Biotechnology 2
Properties of citric acid: Transition temperature: 36.6 oC

C6H8O7 .H2O (CAM)

Prof. Debabrata Das

Department of Biotechnology 3
 Prof. Debabrata Das
Department of Biotechnology 4
 Uses of Citric acid
• Citric acid is a weak organic tricarboxylic acid having the chemical formula C6H8O7.

• 75% citric acid is used in food, confectionery and beverages; 10% use in
pharmaceuticals and 15% use in industries

• Food acidulant, pleasant taste and enhancing flavour

• Cleaning of power station boiler

• Ferric ammonium citrate tablet for anemia patient

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Department of Biotechnology 5
 History of citric acid production
• Scheele in 1784 first isolated citric acid from lemon juice and crystallized it as a
solid.
• Wehmer in 1893 described citric acid as a product of mold fermentation.
• In 1917, Currie published the results that citric acid can be produced from
Aspergillus niger.
• The pharmaceutical company, Pfizer began industrial-level production using this
technique two years later.
• John & E. Sturge Ltd., UK took the technology from Pfizer
and marketed throughout the world.
• In 2007, worldwide annual production stood at
approximately 1,600,000 tons https://en.wikipedia.org/wiki/Citric_acid

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Department of Biotechnology 6
 Organisms involved in citric acid production
• Aspergillus niger
• Aspergillus clavatus
• Penicillium luteum
• Penicillium citrinum
• Corynebacterium sp.
• Mucor piriformis
• Candida catenula

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Department of Biotechnology 7
 Aspergillus niger
Although various strains have been found to produce citric acid, only strains of A.niger are
of industrial importance.
It is an aerobic organism

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Department of Biotechnology 8
 Aspergillus niger
Advantages of using A.niger for industrial
production of citric acid:

•High yields

•Uniform biochemical characteristics

•Easy cultivation

•Production of negligible quantity of

undesirable end products.
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Department of Biotechnology 9
 Biochemistry of citric acid fermentation

• Citrate is an intermediate in the TCA cycle a central metabolic pathway

for animals, plants and bacteria.

• Glucose is the predominant carbon source for citric acid production

• The biosynthetic pathway for citric acid production involves glycolysis

wherein glucose is converted to two molecules of pyruvate.

• Pyruvate in turn forms acetyl CoA and oxaloacetate which condenses to

finally give citrate
.

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Department of Biotechnology 10
 Biochemistry
of citric acid
fermentation

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 Production of citric acid
Fermentation is the most economical way to produce citric acid

Industrial citric acid production

Surface culture Submerged fermentation

• Solid medium • Stirred reactors

•Liquid medium • Airlift reactors

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 Production of citric acid
Surface fermentation
• Solid nutrient media is used.
• Used for the production of complex flavour extracts by bacteria and moulds
(Aspergillus, Mucor)
• Limited water availability
• The media is placed in simple thermostatic boxes on baking-tray like plates to which
the microorganisms are inoculated.

Submerged fermentation
• Fluid nutrient media is used where microbes are propagated in bioreactors
• Sufficient water availability

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Department of Biotechnology 13
 Nutrient requirements for citric acid production

Carbohydrate source
• A high yield occurs if the sugars are rapidly metabolised are used
e.g. sucrose, glucose, maltose.
• Cane molasses and beet molasses are commonly used.

Trace metals
• Certain trace elements (Fe, Cu, Zn, Mn, Mg, Co) are essential for
the growth of A. niger.
• Manganese ions promote glycolysis and reduce respiration
• Iron is a cofactor for the enzyme aconitase (of TCA cycle).
• Fe concentration of 0.05-0.5 ppm is ideal for optimal citric acid
production.

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Department of Biotechnology 14
 Nutrient requirements for citric acid production
pH
• pH -2.5.
• At low pH, transport of citric acid is much higher.
• If the pH is above 4, gluconic acid accumulates at the expense of citric acid.
• And when the pH goes beyond 6, oxalic acid accumulates.
• Another advantage with low pH is that the risk of contamination is very minimal, since many
organisms cannot grow at this pH.

Dissolved O2
• The yield of citric acid production proportional to the dissolved O2. This can be achieved by strong
aeration or by sparging with pure O2.

Nitrogen source
• Ammonium salts, nitrates and urea are the nitrogen sources

Prof. Debabrata Das

Department of Biotechnology 15
 Prof. Debabrata Das
Department of Biotechnology 16
 INDUSTRIAL BIOTECHNOLOGY
Lecture 32: Citric acid fermentation (continued)

PROF. DEBABRATA DAS

DEPARTMENT OF BIOTECHNOLOGY
IIT KHARAGPUR

1
 Surface fermentation - Solid surface
fermentation
•Solid substrates are such as wheat bran or pulp from sweet
potato starch are used as culture media.
•pH - 4-5
•Inoculum in the form of spores of A. niger is spread as layers (3-6
cm thickness)
• Incubated at 28oC.
• Growth is accelerated by the addition of α-amylase.
• Solid-state fermentation time - 80 to 100 h
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Department of Biotechnology 2
 Liquid surface fermentation
• It is the oldest method for citric acid production.
Advantages
• Simple technology
• Low energy costs
• Higher reproducibility
• Interference of trace metals and dissolved O2 tension is minimal

About 20% of the citric acid in the world is produced by surface processes.

Prof. Debabrata Das

Department of Biotechnology 3
 Liquid surface fermentation

• Nutrient supply – beet molasses.

• Fermentation- carried out in aluminium trays filled with sterile nutrient

medium

• Inoculum - spores is sprayed

• Sterile air - for supplying O2 and cooling

• Temperature – 30oC

• As the spores germinate (that occurs within 24 h of inoculation), a layer

of mycelium is formed over the medium.

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Department of Biotechnology 4
 Liquid surface fermentation
• The pH of the medium drops to 2, as the mycelium grows in size and forms a
thick layer on the surface of the nutrient solution.

• Fermentation is stopped after 7-15 d.

• The mycelium and nutrient solution are separated.

• The mycelium is mechanically pressed and thoroughly washed to obtain

maximum amount of citric acid.

• The nutrient solution is subjected to processing for the recovery of citric acid.

• The final yield of citric acid is in the range of 0.7-0.9 g per g of sugar.

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Department of Biotechnology 5
 Submerged fermentation

• 80% of the world’s supply of citric acid is produced by submerged

processes.

Advantages

• High efficiency
• Easy automation

Disadvantages

• Adverse influence of trace metals and other impurities,

• Variations in O2 tension,
• Advanced control technology that requires highly trained personnel.

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Department of Biotechnology 6
 Submerged fermentation

•Two types of bioreactors are in use—

stirred tanks and aerated towers.

•Bioreactors – made of high-quality

stainless steel.

•An adequate supply of O2 (20-25% of

saturation value) is required for good
production of citric acid.

•Problem of foam formation

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Flow diagram of Citric acid fermentation
process

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Department of Biotechnology 8
Inoculum preparation:

Sporulation of A. niger is prepared in the laboratory under aseptic

condition.

Spores are prepared as suspension in saline water.

This is inoculated in the Inoculum vessel located in the

fermentation plant for the production of vegetative cells.

These vegetative cells are used as inoculum for the production

fermenter.

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Department of Biotechnology
 Submerged fermentation
• The success and yield of citric acid production mainly depend on the structure of mycelium.
• Mycelium with forked and bulbous hyphae which aggregate into pellets is ideal for citric acid
formation.
• If the mycelium is loose and filamentous with limited branches- no citric acid production
occurs.
• Mixing and shear stress is also a crucial parameter.

Different morphological characteristics of Aspergillus niger (volumetric power input P/V = 100 W m-3), a) biopellet at pH 5.5 after a
cultivation period of 32 h, b) free dispersed mycelia at pH 3 after 24 h, c) morphology after an initial growth at pH 3 for 8 h,
subsequent pH-shift to pH 5.5 and further cultivation for 24 h
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Downstream processing of citric acid
Crude raw material

Upstream
processing
Processed raw materials
Fermentation
process

Crude products
Downstream
processing

Citric acid

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Department of Biotechnology 11
Block flow diagram of citric acid purification process

Prof. Debabrata Das

Department of Biotechnology
 Prof. Debabrata Das
Department of Biotechnology 13
 INDUSTRIAL BIOTECHNOLOGY
Lecture 33: Citric acid fermentation (continued)

PROF. DEBABRATA DAS

DEPARTMENT OF BIOTECHNOLOGY
IIT KHARAGPUR

1
 Prof. Debabrata Das
Department of Biotechnology
 Prof. Debabrata Das
Department of Biotechnology
 Prof. Debabrata Das
Department of Biotechnology
 Prof. Debabrata Das
Department of Biotechnology
 Steps in the downstream processing
Step 1
• Filtration of the culture broth and
washing of mycelium (which may
contain about 10 %w/v of citric acid).
• Filtration is done rotary vacuum filter
• Oxalic acid is an unwanted byproduct
and it can be removed by precipitation
by adding lime at pH < 3.

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Prof. Debabrata Das 6

Department of Biotechnology
 Steps in the downstream processing
Step 2

• Ca(OH)2 is added to the filtrate until the pH becomes 6.8.

• Calcium hydroxide neutralizes the broth and forms the insoluble precipitate calcium
citrate.

• Calcium citrate contains about 74% citric acid.

• The calcium citrate is then washed, heated, and filtered to remove any number of the
contaminants
Prof. Debabrata Das
Department of Biotechnology
 Steps in the downstream processing
Step 3
• Pannevis filter is then used to separate Calcium citrate. The choice of filter plays an
important role in the downstream processing.

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8
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 Steps in the downstream processing
Step 4

• H2SO4 is added at 60oC.

• Calcium citrate then reacts with sulphuric acid to form calcium sulphate
(Gypsum)and citric acid
• Further, filtration is done to separate Calcium sulfate (gypsum, CaSO4.2H2O)
• Name of the filter is Gypsum filter

Prof. Debabrata Das

9
Department of Biotechnology
 Steps in the downstream processing
Step 5

• The final steps for citric acid recovery are — treatment with
activated charcoal and crystallization.

• The dilute citric acid is treated with activated charcoal to

decolorize the solution.

• Evaporation is done to produce crystals of citric acid.

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10
Department of Biotechnology
Decolorization by
activated charcoal

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11
Department of Biotechnology
 Steps in the downstream processing
Step 6
• Evaporation of the citric acid crystals is done
by continuous centrifugation

• They are sieved and packaged into crystals.

• The degree of purity of citric acid produced

depends on the purpose for which it is
required. For instance, pure forms of citric acid
are needed for use in food preparations, while
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for industrial use it can be crude form.
Prof. Debabrata Das
12
Department of Biotechnology
 Downstream processing of citric acid
Further Purification

Citric acid can be produced in two forms – monohydrate and anhydrous. These forms may

require additional purification steps to reach the desired purity.

Monohydrate
• contains one water molecule for every citric acid molecule
• Requires repeated crystallization until water content is approx. 7.5-8.8%

Anhydrous
• Processed to remove all water from end product
• Prepare by dehydrating the monohydrate citric acid product at a
temperature above 36.6⁰ C.

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13
Department of Biotechnology
 Prof. Debabrata Das
Department of Biotechnology
 Prof. Debabrata Das
Department of Biotechnology
 Prof. Debabrata Das
Department of Biotechnology
Problem:
100 m3 of citric acid fermentation broth has been harvested in harvesting tank,
the broth has 11% (w/v) of cell mass and 12% w/v of citric acid. Compute the
followings:
i) Total amount of mycelium produced;
ii) Lime required for calcium citrate precipitation process;
iii) Maximum of amount of gypsum produced (CaSO4 2H2O);
Amount of water is to be removed to increase the citric acid concentration from
22% to 60 %w/v.
Prof. Debabrata Das
Department of Biotechnology
 Prof. Debabrata Das
Department of Biotechnology
 Prof. Debabrata Das
Department of Biotechnology
 Prof. Debabrata Das
Department of Biotechnology
 Prof. Debabrata Das
Department of Biotechnology
 Prof. Debabrata Das
Department of Biotechnology 22
INDUSTRIAL BIOTECHNOLOGY
Lecture 34: Lactic acid production

PROF. DEBABRATA DAS

DEPARTMENT OF BIOTECHNOLOGY
IIT KHARAGPUR

1
 Lactic acid
• Lactic acid is an organic compound with the formula CH3CH(OH)COOH.
• In its solid state, it is white and water-soluble.
• In its liquid state, it is clear. It is produced both naturally and synthetically.
• Lactic acid is found primarily in sour milk products, such as yogurt and kefir,
and some cottage cheeses.
• The casein in fermented milk is coagulated due to lactic acid.
• Lactic acid is also responsible for the sour flavour of
sourdough bread.
• Also finds application in pharmaceutical industry
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 Forms of lactic acid

The lactic acid formed during fermentation is usually L (-) form (a racemic mixture).
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Department of Biotechnology 3
 History of lactic acid production
• It was isolated and identified by Scheele as the principal acid in sour milk in 1780.
• Lactic acid was first discovered as a fermented product by Blondeau in 1847.
• It was investigated by Pasteur as one of his first microbiological problem.
• Schultze (1868) demonstrated the presence of lactic acid bacteria in yeast
cultures of distilleries.
• During this same period Delbruck was endeavoring to determine the most
favorable temperature of lactic acid fermentation in distilleries.
• He concluded that relatively high temperature favored high yield of lactic acid.
• In 2006, global production of lactic acid reached 275,000 tonnes with an average
annual growth of 10%.

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Department of Biotechnology 4
 Lactic acid bacteria
• Lactic acid bacteria (LAB) are a diverse group of bacteria
capable of lactic acid production.
• Found in cheeses, yoghurts or decomposing plants.
• Gram-positive and non sporulating
• Ability to produce lactic acid as a major metabolic end
product of carbohydrate fermentation
• They are non-respiratory (anaerobic), but tolerate also
aerated environments and can survive high acid (pH 3-6)
and high ethanol concentrations.
• The Lactobacillales can be divided into different genera,
such as Lactobacillus, Leuconostoc, Pediococcus,
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functional-ingredients

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Department of Biotechnology 6
Lactic acid production
pathway

Homofermentative lactic acid

bacteria use the EMP pathway to
make pyruvate and then reduce it to
lactate using up their excess NADH in
the process subsequently.

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Department of Biotechnology 7
 Activity requirements
• Lactic acid bacteria requires complex medium for their growth.

• They fail to grow in the medium containing any lack of amino acid.

• They utilize the B-vitamins.

• They are now produced commercially from corn sugar, molasses and whey.

• Usually, yeast extract or malt extract is used which contains all the amino acids
required for their growth.
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Department of Biotechnology 8
 Desirable characteristics of lactic acid bacteria
• During fermentation, it should yield lactic acid as one of the major product. Lactic acid
bacteria may form components other than lactic acid e.g. 2, 3 butanediol.

• It should act as a preservative

• It should prevent putrefactive changes from taking place in sauerkraut and pickles and
souring of the milk.

• It should aid flavor development in the fermentation product. Characteristic flavor in

the dairy products such as yogurt and cheese have is due to the lactic acid bacteria.

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Department of Biotechnology 9
 Importance of LAB addition to yeast
Acid dough contains lactic acid bacteria and yeast. LAB addition is done to the following reasons:

• It acts as a preservative.

• Most yeast utilize lactic acid as a C-source.

• Most of the contamination to yeast is caused by the alkalinity of the medium. To avoid
undesirable bacteria grow like putrefying bacteria and to prevent contamination In the
fermentation process, lactic acid bacteria is used. So, aseptic fermentation is relaxed.

• In ancient age, large inoculums are used to prevent contamination. It is used to acidulate (worts in
the manufacture of beer) to adjust the pH of the brine in the manufacture of pickled green olives,
and to inhibit the development of butyric acid bacteria in the manufacture of yeast.

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Department of Biotechnology 10
 Classification of lactic acid bacteria
Lactic acid bacteria

Homofermentative Heterofermentative
bacteria bacteria
•They convert about 95% of fermentable hexoses to
lactic acid. It follows Embden-Meyerhof pathway. •They differ from the homo fermentative
species in the fact that lactic acid is only one
•C6H12O6 → 2 CH3 CHOH. COOH
•Small amounts of volatile acids and CO2 are also of several principal products formed from
produced sugar other compounds include ethyl alcohol,
•Disaccharides are fermented in a similar manner acetic and formic acids and carbon dioxide. It
e.g. L. casae, L. delbrueekii, L. bulgaricus. follows Pentose phosphate pathway.
e.g. L. acidophilus.

Prof. Debabrata Das

Department of Biotechnology
 Spoilage reactions caused by lactic acid bacteria

• It acts as spoilage in beer, malt beverages, wine and distillery industries.

They produce turbidity, off flavor and precipitates.
• In acetone butanol fermentation, the lactic acid bacteria is undesirable. It
uses sugar for their growth.
• In dairy industry it can also cause souring in milk.
• In orange juice concentrates, lactic acid bacteria causes spoilage because
of off flavor, souring.
• In sugar refinery industry, Leuconostoc meseteroids can grow which forms
high viscous sugar which may interfere in crystallization of sugar.
• In meat products, it can form turbidity and surface growth which is
undesirable.
Prof. Debabrata Das
Department of Biotechnology 12
Steps in the biological and
chemical manufacture of
lactic acid

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Department of Biotechnology 13
 Raw materials
Carbohydrates suitable for utilization
• A large number of carbohydrates may be utilized for lactic acid production.
• The acid is generally produced from glucose, sucrose, or lactose.
• Starches, corn and potato especially, may be hydrolyzed by enzymes, or by acids
(preferably sulfuric acid) to maltose and glucose.
• Molasses and whey are generally low priced sources of sugars for the fermentation.
• The sugar in mashes is normally adjusted to a concentration of 5 to 20 %

Oxygen relationship
• The bacteria used to produce L.A. industrially are usually microaerophilic or anaerobic in
nature.
• Small out of O2 is required for the conversion of lactic acid also.

Prof. Debabrata Das

Department of Biotechnology 14
 Fermentation
pH
• The fermentation proceeds best when the pH is on the acid side of neutrality.
• Owing to the addition of calcium carbonate, calcium hydroxide, or some other neutralizing
agent to the fermentation mash, the pH tend to approach neutrality (pH 5.5-6.5).

Growth factors for lactic acid bacteria

• Certain growth factors appear to be essential for certain lactic acid bacteria.
• Riboflavin acts as an activator
• Nicotinic acid stimulated growth and acid production
• Some strains require biotin, panthothenic acid and nicotinic acid for their normal growth.

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Department of Biotechnology 15
 Accessory nutrients
• The yield and the fermentation time depend on the kind, quantity and combination of the
accessory nutrients.

• They also supply organic nitrogen and stimulatory substances.

• Good fermentation yields and short fermentation time result from the addition of accessory
nutrients (when black strap molasses is used as substrate) such as

Malt sprouts (unheated malt sprouts accelerated the fermentation due to the addition of a
heat-labile growth factor contained in sprouts), Steep water etc.

Prof. Debabrata Das

Department of Biotechnology 16
 Fermentation
Temperature
• The L.A. fermentation is carried out at comparatively high temperature.
• Fermentation temperature depends upon the organism used
• L. delbrueekii temperature of 45°C, or higher, may be maintained.
• L. bulgaricus - 45 to 50°C.
• L. pentosus, L. casei, Streptocuccus lactic - 30°C.

Fermentation time
1-6 d

Yields obtained
85 to 90 %
end

17
 Prof. Debabrata Das
Department of Biotechnology 18
 INDUSTRIAL BIOTECHNOLOGY
Lecture 35: Lactic acid production (continued)

PROF. DEBABRATA DAS

DEPARTMENT OF BIOTECHNOLOGY
IIT KHARAGPUR

1
 Lactic acid production from whey
 Whey is a cheap raw material.

 It contains 4.6 %w/v lactose

 The cream is removed from the milk and casein is precipitated from the
use of hydrochloric acid.

 The whey, which is the product left after the separation of the cream and
casein from the milk, contains albumin (protein) approximately 4.6
%w/v lactose and Vitamin G, mineral salts and water.

 This medium serves as the nutrient substrate in the manufacture of lactic

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acid.

Prof. Debabrata Das

2
Department of Biotechnology
 Steps involved in lactic acid production

• Preparation of the starter

• Fermentation
• Lactic acid recovery Filtration
Evaporation
Purification
Slant Shake Culture in
Culture flask whey in Lactic acid
culture the
fermenter
Production
fermenter

Prof. Debabrata Das

3
Department of Biotechnology
 Step 1: Preparation of the starter
 Sterilized skim milk is inoculated with a culture of L. bulgaricus, containing yeast that
causes the fermentation to become more vigorous.

 After incubation for 24 h at a temp of 43 °C, the contents of the bottle are placed in
container with pasteurized skim milk.

 The contents of the jug are transferred, after incubation at a temp. of 43 °C for 24 h into
a glass-lined steel tank containing 500 gal of pasteurized whey.

 Following incubation at 43 °C for 24 h, this starter is added to the main fermentation

tank.
Prof. Debabrata Das
Department of Biotechnology
 Step 2: Fermentation
Material of construction
Lactic acid is highly corrosive and therefore it has problems in relation to its production, processing and
handling.
The non-corrosion material is used for the construction of fermenter.
Two of the materials of construction which are most resistant to lactic acid are silver and tantalum.
Group of materials which are fairly resistant to lactic acid are nickel, Inconel (nickel-chromium-based
super alloys) and low iron alloys.
Group of materials which are poor resistant to lactic acid are iron, copper, chrome steel etc.
Usually fermenter is constructed by heavy wood.
Modern fermenter is constructed by stainless steel.
Fermenter is provided with agitation.

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Department of Biotechnology
 Step 2: Fermentation
• Inoculated whey is maintained at a temperature of 43 ⁰C.

• Fermentation time – 42 h

• Calcium hydroxide in the form of slurry is added to the fermenting

mash every 6 h to keep the acidity of the mash below 0.1 %.

• Neutralizing the lactic acid with lime has several advantages such as
– shorter fermentation time, higher yield and prevention of
bacterial inhibition due to the production of acid.

Prof. Debabrata Das

6
Department of Biotechnology
Process flow diagram of lactic acid production

Li et al., Appl. Biochem.

BioTechnol., 147, 1-9, 2008

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Department of Biotechnology
Polylactic acid formation

Polylactic acid

Prof. Debabrata Das

Department of Biotechnology
 Uses of polylactic acid

Polylactic Acid (PLA) is biodegradable. It is mostly used in food handling

and medical implants that biodegrade within the body over time. Like most

plastics, it has the potential to be toxic if inhaled and/or absorbed into the

skin or eyes as a vapor or liquid (i.e. during manufacturing processes).

Prof. Debabrata Das

Department of Biotechnology
 Lactic acid product recovery
 Ca(OH)2 is added to broth to lower the concentration of acid below 0.1%.

 Then heated at 80-95°C which help for the coagulation of proteins and filtered. The filtrate then
allowed to evaporate to dryness to get Ca-lactate.

 For the preparation of pure Ca-lactate, it is dissolved in water and treated with decoloring carbon
(norit) and mixed thoroughly and stand for 15 min and filtered.

 To prepare lactic acid from Ca-lactate, it is treated with dil. H2S04 to get dilute solution of lactic acid.
CaSO4 gets precipitate out.

 If the dilute solution is heated to high temperature then some chemical change may occur e.g. it
forms lactate anhydride.

Prof. Debabrata Das

Department of Biotechnology
Crude lactic acid recovery

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Containers used in distribution

• Aqueous solutions of lactic acid are sold and distributed in

wooden or paraffin-lined barrels of oak, cypress, or pitch pine
or in lined car tanks.

• The higher grades of acid are sold in glass-lined cars or

carboys.

Prof. Debabrata Das

12
Department of Biotechnology
 Different grades of lactic acid

There are four principal grades of lactic acid.

• Crude or technical grade → In contents 20-80% L.A strength

• Edible → 50-80% L.A.

• Plastic → 50-80% L.A.

• United States Pharmacopeia (U.S.P.) (medicinal) → 85% L.A.

Prof. Debabrata Das

Department of Biotechnology
 Different grades of lactic acid

Crude grade
• The colour differs from yellow to brown
• It is used in
• (1) Deliming of hides i.e. Limes are removed from the animal skim.
(2) Dying of silk and other textile

Edible
• Straw - yellow colour.
• It is used in jam, jelly, brine fermentation, casein precipitation, for fat extraction.

Prof. Debabrata Das

Department of Biotechnology
 Different grades of lactic acid
Plastic:
• It is colourless.
• Generally used for the manufacture of plastic.

U.S.P:
• It is a pharmaceutical grade lactic acid
• Used in pharmaceutical formulations

Prof. Debabrata Das

Department of Biotechnology
 Prof. Debabrata Das
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Department of Biotechnology