Group13 Fermentation Industries

8/2/2019 Group13 Fermentation Industries

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SUBMITTED BY: GROUP XIII

MD. HASSAN FARAZ 09CH1011

NITIN GAUTAM 09CH3018

ADARSH KUMAR 09CH1035

PRANITA PADALWAR 09CH1043

RAKESH KUMAR 09CH1032

A REPORT ON

FERMENTATION INDUSTRIES


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INTRODUCTION

Definition: Industrial fermentation is the use of fermentation by microorganisms such as bacteria and

fungi to make products useful to humans and have economic value. Used in food, pharmaceutical and

biotechnology industry. The term fermentation derives from the Latin wordfeveremeaning to

ferment.Nutrient Sources for industrial fermentation:

Growth media are required for industrial fermentation, since any microbe requires water, (oxygen), an

energy source, a carbon source, a nitrogen source and micronutrients for growth.

Carbon & energy source + nitrogen source + O2+ other requirements Biomass + Product + byproducts

+ CO2 + H2O + heat

Fermentation technique:

Techniques for large-scale production of microbial products must provide an optimum environment for

the microbial synthesis of the desired product and be economically feasible on a large scale. They can be

divided into two types majorly.

Surface (emersion): In the surface techniques, the microorganisms are cultivated on the surfaceof a liquid or solid substrate. These techniques are very complicated and rarely used in industry.

Submersion: In this technique microorganisms grow in a liquid medium which is held infermenters and stirred to obtain a homogeneous distribution of cells and medium. Most

processes are aerobic, and for these the medium must be vigorously aerated. All important

industrial processes (production of biomass and protein, antibiotics, enzymes and sewage

treatment) are carried out by submersion processes. It can be run in batch, fed batch or

continuous reactors.

Microbial Growth Kinetics: Describes how the microbe grows in the fermenter.

Stationary Phase: At some point the cell growth rate will level off and become constant. Themost likely cause of this leveling off is substrate limited inhibition which means that the

microbes do not have enough nutrients in the medium to continue multiplying.

Death Phase: The number of cells dying is greater than the number of cells multiplying. Thecause of the death phase is usually that the cells have consumed most of the nutrients in the

medium and there is not enough left for sustainability.

This information is important to determine

optimal batch times. The growth of

microbes in a fermenter can be broken

down into four stages:

Lag Phase: The cells have just been

injected into a new environment and

they need time to adjust accordingly.

Cell growth is minimal in this phase.

Exponential Phase: The cells have

adjusted to their environment and

rapid growth takes place. Cell

growth rate is highest in this phase


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Types of fermentation process:

Batch fermentation process: A tank of fermenter is filled with the prepared mash of raw materials to be

fermented. The temperature and pH for microbial fermentation is properly adjusted, and occasionally

nutritive supplements are added to the prepared mash. The mash is steam sterilized in a pure culture

process. The inoculum of a pure culture is added to the fermenter, from a separate pure culture vessel.

Fermentation proceeds, and after the proper time the contents of the fermenter, are taken out for

further processing. The fermenter is cleaned and the process is repeated. Thus each fermentation is a

discontinuous process divided into batches.

Batch Bioreactor: The bioreactor's environmental conditions like gas (i.e., air, oxygen, nitrogen, carbon

dioxide) flow rates, temperature, pH and dissolved oxygen levels and agitation speed/ circulation rate

need to be closely monitored and controlled. Biological fermentation is a major source of heat,

therefore in most cases bioreactors need refrigeration. They can be refrigerated with an external jacket

or, for very large vessels, with internal coils. In an aerobic process - Oxygen is poorly soluble in water

and is relatively scarce in air (20.95%). Oxygen transfer is usually helped by agitation, which is also

needed to mix nutrients and to keep the fermentation homogeneous but there are limits to the speed of

the agitation. In practice, bioreactors are often pressurized; this increases the solubility of oxygen inwater.

Continuous fermentation process: Growth of microorganisms during batch fermentation conforms to

the characteristic growth curve, with a lag phase followed by a logarithmic phase. This, in turn, is

terminated by progressive decrements I in the rate of growth until the stationary phase is reached. This

is because of limitation of one or more of the essential nutrients. In continuous fermentation, the

substrate is added to the fermenter continuously at a fixed rate. This maintains the organisms in the

logarithmic growth phase. The fermentation products are taken out continuously. The design and

arrangements for continuous fermentation are somewhat complex.

Chemostat: is a bioreactor to which fresh medium is continuously added, while culture liquid is

continuously removed to keep the culture volume constant. By changing the rate with which medium is

added to the bioreactor the growth rate of the microorganism can be easily controlled. One of the most

important features of chemostats is that micro-organisms can be grown in a physiological steady state.

In steady state, growth occurs at a constant rate and all culture parameters remain constant (culture

volume, dissolved oxygen concentration, nutrient and product concentrations, pH, cell density, etc.)

Aerobic Fermentation: A number of industrial processes, although called 'fermentations', are carried on

by microorganisms under aerobic conditions. In modern fermentation processes aerobic conditions are

maintained in a closed fermenter with submerged cultures. The contents of the fermenter are agitated

with an impeller and aerated by forcing sterilized air.

Anaerobic Fermentation: Basically a fermenter designed to operate under anaerobic conditions will be

the same as that designed to operate under aerobic conditions, except that arrangements for intense

agitation and aeration are unnecessary. Many anaerobic fermentation does, however, require mild

aeration for the initial growth phase, and sufficient agitation for mixing and maintenance of

temperature.

Application: Medicines i.e. pharmaceuticals, including antibiotics, steroids, human protein, vaccines

and vitamins. E.g. Penicillin; Alcohol production; Beverages Beer, wine; Organic Acid & solvent

production, Dairy Product, Yogurt, cheese, etc.; Sewage Disposal
http://en.wikipedia.org/wiki/Oxygenhttp://en.wikipedia.org/wiki/Nitrogenhttp://en.wikipedia.org/wiki/Carbon_dioxidehttp://en.wikipedia.org/wiki/Carbon_dioxidehttp://en.wikipedia.org/wiki/PHhttp://en.wikipedia.org/wiki/Agitator_(device)http://en.wikipedia.org/wiki/Refrigerationhttp://en.wikipedia.org/wiki/Oxygenhttp://en.wikipedia.org/wiki/Airhttp://en.wikipedia.org/wiki/Bioreactorhttp://en.wikipedia.org/wiki/Population_growth_ratehttp://en.wikipedia.org/wiki/Microorganismhttp://en.wikipedia.org/wiki/Microorganismhttp://en.wikipedia.org/wiki/Population_growth_ratehttp://en.wikipedia.org/wiki/Bioreactorhttp://en.wikipedia.org/wiki/Airhttp://en.wikipedia.org/wiki/Oxygenhttp://en.wikipedia.org/wiki/Refrigerationhttp://en.wikipedia.org/wiki/Agitator_(device)http://en.wikipedia.org/wiki/PHhttp://en.wikipedia.org/wiki/Carbon_dioxidehttp://en.wikipedia.org/wiki/Carbon_dioxidehttp://en.wikipedia.org/wiki/Nitrogenhttp://en.wikipedia.org/wiki/Oxygen


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Major Engineering Problems:

1. Scale-up of Submerged FermentationEscape velocity of air = (vV/A) D where v=volume of air per unit volume of liquid per time and

V is tank volume.

We see that the escape velocity increases with tank size. If small diameter tanks are used,

foaming will increase and become unmanageable. But an increase in diameter results in ill-mixing. Thus the design must be a compromise between tank size, shape, optimized mixing of

air supply and exit velocity.

2. Air and Media sterilization: Air needs to be appropriately sterilized in order to avoidcontamination of the microbes used in fermentation. A common method to achieve this is by

heat sterilization where air is adiabatically compressed to 200C and removing the dust and oil

by passage through fibrous filters. The substrate is similarly sterilized.

3. Continuous Vs Batch Distillation:Continuous Batch

Easier control, uniformity of product, reduced

labor cost

Long residence time, reduced hazards of

contamination and mutation of microbes

4. Mixing limitations:Rate=KG (Ygas - Yliq), where = total pressure.

Y= equivalent mole fraction of O2 based on gas phase.

Kg= overall mass transfer coefficient.

Kg varies as 1/3rd power of Pv (mixing power per unit volume)

Initially the rate increases as impeller

power is increased as it ensures bettermixing and turbulence. But at high Pv

there is a fall in the rate due to the

damage caused to the cell walls of the

microorganisms under high shear stresses

developed in mixing.

Economics of Fermentation Industry:

One of the hallmarks of business of manufacturing is to produce the products at the lowest cost while

still making profits. Lowering the price will make it attractive to consumers and reduce or eliminate

competitors of the same products. This can only be achieved if the volume of production is higher thus

the cost of producing the unit price will be lowered. In fermentation this would mean involving bigger

fermentation capacity which is not normally achieved in small cottage industries.

Prerequisitesfor a successful fermentation processes are:


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Microorganisms must be specific and produce good yield of desired end-product Rapid fermentation rates must be achieved Use economic raw materials with dependable supply Products easily recovered and purified

Cheap substrates are difficult to find even if they are by-products from other processes. One interesting

development in the world-wide fermentation picture is the use of cheap petroleum hydrocarbons to

produce food for human consumption. Japan is actively engaged in this research.

Examples in India of typical fermentation industries presently economic are Ethyl alcohol and Penicillin.

ETHYL ALCOHOL

Physical Properties:

Mol. Wt. 46.07 MP 112C

Density 0.791 @ 20C BP 78.3C

Flash Point 21C Ignition Temp 372CExplosive limits Lower= 3.5% by vol.

Upper= 19% by vol.

Toxicity Limit 1000ppm

India is the fourth largest producer of ethanol after Brazil, the United States and China.

Raw Materials:

Corn is used majorly on industrial scale. Corn is largely composed of starch which can be hydrolyzed

(broken down) into sugars and fermented to ethanol using yeast.

But in India sugarcane is the key raw material for producing ethanol. It is produced by fermentation of

molasses which is a bi-product of sugar.

Typically 88lbs of molasses are produced per ton of cane from which 10lt of ethanol can be made. If the

sugarcane is used completely for molasses then 70lt/ton can be produced.

0%5%

10%15%20%

25%30%35%40%

End Use Pattern

percentage


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Type of yeast preferred: Saccharomyces cerevisiae. Enzyme furnished Invertase and zymase

Raw Materials processing:

i. Preparation of the corn grain involves cleaning and conditioning steps, as well as generating anaqueous solution high in simple sugars. Firstly broken corn kernels and foreign materials (metal

dirt etc.) are removed by blowers and screens. It is then ground in hammer mills fitted with

suitable screens. Grinders serve the purpose of breaking the tough outer coating of the corn

kernel and increase the surface area of exposed starch.

ii. Liquefaction involves combining the ground corn with process water to form slurry which isapproximately 30% solids by weight. Ammonia and lime are added for pH control and as a

nutrient to the yeast. Some thermostable enzymes (alpha-amylase) is added to convert starch

into simple sugar.

iii. Following liquefaction sulfuric acid is added to lower the pH to 4.5 as required. Nutrients such asammonium and magnesium sulfate or phosphate is added at various steps if lacking in the feed.

iv. Air Sterilization: Contamination of pure cultures used in fermentators is avoided by heatsterilization of the air. This is mostly accomplished by adiabatic compression to 200C or a steam

coil can also be used. This reduces the bacteria level which inhibit fermentation process by yeastcells.

Chemical Reactions:

a. Main Reactions:C12H22O11 + H2O = 2C6H12O11 Enzyme responsible: Invertase

C6H12O11 = 2C2H5OH + 2CO2 Enzyme responsible: Zymase H = -31.2Kcal.

b. Side Reactions:2C6H12O11 + H2O = ROH + RCHO (Higher mol. Wt. alcohols.)

Process Description:

i.

Molasses is diluted 10-15% concentration and pH is adjusted to support yeast growth whichfurnishes invertase and zymase catalytic enzymes. This liquid mixture called mash is sent to

steel/wooden fermentation tanks.

ii. The sterile mash is inoculated to grow pure yeast culture and then sent to the fermentationtank. The temperature is maintained at 20-30C over a 30-70 hours period. Since the reactions is

exothermic cooling coils ared provided to remove heat.

iii. We get 8-10% alcohol in the fermented liquor which is concentrated by a series of distillations.iv. In the first step, the beer is processed through a beer column where steam is used to strip off

almost all of the ethanol along with some water and some undesirable volatiles such as

aldehydes from the slurry. The ethanol (50-60%) leaves the beer column from the top and the

whole stillage (containing less than 0.1% ethanol by weight) exits from the bottom.

v. The overhead vapor flows to an aldehyde still to remove the volatiles from the top and ethanolis pulled off as a side stream split to the rectifying coloumn. Here azeotropic mixture of

alcohol(95%)and water is taken off as top side stream condensed and run to storage.Liquid

exiting the bottom has less than 0.1% ethanol by weight, and is recycled as process water for

dilution of molasses.


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Figure: Process Flow Diagram of Ethyl Alcohol Production by Fermentation

Product Despatch:

The product in the storage is split into three streams:

Direct sale as potable government controlled alcohol. Denatured by small addition of toxic ingredients to make it unsuitable to drink and sold for

industrial uses.

Made anhydrous by ternary azeotropic distillation using benzene or extractive distillation usingethylene glycol.

Co-Product Recovery:

The stillage exiting from the bottom can be evaporated and used as livestock fodder. Whole stillage is

sent through a centrifuge to separate the coarse solids from the liquid. The liquid is referred to asDistillers Solubles, or thin stillage. Thin stillage is concentrated to about 30% solids by evaporation

resulting in Condensed Distillers Solubles (CDS), or syrup. The coarse solids from the centrifuge and CDS

are combined to form Wet Distillers Grains with Solubles (WDGS) and can be dried to form DDGS, a high

quality nutritious livestock feed.

The CO2 released during fermentation is captured and sold for use in carbonating soft drinks and

beverages and the manufacture of dry ice.


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Figure: Flow chart for typical coproducts processing

Major Engineering Problems:

Collection and storage of molasses. Maintainance of sterile and specific yeast culture Batch versus continuous operation: Continuous molasses dilution and distillation are

incorporated to save space equipment and operating costs. Waste Disposal Problem: If uneconomic to concentrtate for cattle feed, must use trickling filters,

activated sludge or anaerobic digestion to lower the BOD before discharging to water runn off.

Development of methods to produce anhydrous alcohol from the 95% alcohol azeotrope.Safety Factors:

R11: Highly Flammable

S7 : Keep containers tightly closed

S9 : Keep container in a well-ventilated place

S16: Keep away from source of ignition - No Smoking

S33: Take precautionary measures against static discharges

Economics of Alcohol Industry:

Alcohol by fermentation in India is limited to sucrose substrate because insufficient supplies ofstarch and paper mill waste.

This type of process furnished the bulk of alcohol requirements till 1960s when alcohol frompetroleum processing became available in India. After that alcohol produced from petro-

chemical complexes started dominating due to their large scale nature and lower cost of


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production. Furthermore it is impossible to supply the entire future demand for alcohol with

industries based on agricultural raw materials.

Improvement in yields can be made by controlling yeast culture and conditions in the tank,recovering ethanol vapours from CO2 produced using activated carbon.

Present day scenario is that the alcohol for drinking purposes still produced by fermentationwhile industrial alcohol demand is met by petrochemical-complexes majorly.

Presently India has a capacity of 2900ML of annually by fermentation process itself. In addition to existing 10 facilities, Indian sugar producers were to build 30 new facilities by

2007.

Number of facilities Capacity(ML) State

11 75 Uttar Pradesh

7 62.5 Tamil Nadu

8 66.5 Karnatka

4 40 Andhra Pradesh

PENICILLIN

Introduction:

A class of antibiotics which comes from mold, penicillium fungi. Discovered by accident in 1928 by

Alexander Fleming observed that Penicillium notatum, a common mold, had destroyed Staphylococcus

aureus bacteria in culture, is the first antibiotic. Penicillin antibiotics include ampicillin, phenoxymethyl

penicillin, and amoxicillin.

How Penicillin Works: Penicillin resembles proteins required for production of cell wall. Penicillin binds

to cell wall of bacteria, prevents peptide chains from linking, and lyses it.

Properties of Penicillin:

I. Structure :

II. Types of penicillin:Name of derivative Designation

British

Designation

U.S.

R in side chain units/mg of Na salt

2-Pentenyl- I F CH3CH2CH=CHCH2- 1600

n-Amyl I F n-C5H11- 1500

Benzyl II G C6H5CH2- 1667


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p-hydroxy-benzyl III X HOC6H4CH2- 900

n-Heptyl IV K n-C7H15 2300

Phenoxy-methyl V Vee C6H5-OCH2- 1550

Changes of M in the main ring will impart solubility and ingestion rate control.

Direct rapid action injection : M= Na or K Oral tablets : M= K,Ca,Al Delayed action oil base-base injection: M= procaine or other derivative to impart limited water

solubility.

Penicillin II or V are considered most clinically desirable.Industrial production:

Broadly divided into 2 processes:

UPSTREAM PROCESSING: Upstream processing encompasses any technology that leads to thesynthesis of product. Upstream process includes the exploration, development and production

and inoculum preparation.

DOWNSTREAM PROCESSING: The extraction and purification of a biotechnological product fromfermentation is referred to as downstream processing.

Raw materials:

Carbon sources: Lactose acts as a very satisfactory carbon compound, provided that is used ina concentration of 6%. Others such as glucose & sucrose may be used.

Nitrogen sources: Corn steep liquor (CSL), Ammonium sulphate and ammonium acetate can beused as nitrogenous sources.

Mineral sources: Elements namely potassium, phosphorus, magnesium, sulphur, zinc andcopper are essential for penicillin production. Some of these are applied by corn steep liquor. Calcium: can be added in the form of chalk to counter the natural acidity of CSL. PAA:- precursor

Quantitative requirements:

BASIS: 1 kg of Penicillin II (1.67109 units)

Broth (dry basis) : 135 Kg

Lactose : 100 kg

Calcium carbonate : 35 Kg

Process water : 3 ton

Air : 7000 Nm3

Plant capacity: 10-50 Kg/day


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Process Description:

i ) Mass Production of Penicillin is carried out by aerobic batch fermentation process.

Production of Maize/corn steep liquor or corn steep liquor:

It is a by-product of corn wet milling which is used in the culturing of Penicillium. Wet -milling isa process in which feed material is steeped in water, with or without sulphur dioxide, to soften

the seed kernel in order to help separate the kernels various components.

Then steeping is done in which Saturation in a liquid solvent to extract a soluble ingredient,where the solvent is the desired product.

It is good source of organic nitrogen.ii) Maize liquor is diluted with water to 4-5% solids and fortified with

Lactose solids (carbon source) Nutrients (Mg, Zn, Na2(SO4) ,Ammonium Acetate and potassium dihydrogen sulphate in amount

less than 0.2%.

CaCO3 is added to adjust pH 5-6 (important for culture development).Types and amount of alkali and nutrients depends upon the starting liquor.

iii) Addition of Organic Precursor chemicals to control type and yields of penicillin.

Eg. : For TYPE II Penicillin phenyl acetic acid precursor is required.

Figure: Process Flow Diagram of Penicillin Production by Fermentation


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iv) Sterilization: Batch fermentation medium is sterilized with steam 1150C for half an hr.

Fermentator is then cooled down to 250C and inoculated with a specific mold culture.

For eg Penicillium chrysogenum for TYPE II. Sterile air is blown through the tank for 4-5

days under temperature control of +/- 1/20C. Penicillin yield is maximized by periodic

assay

v) Separation: use of solvent extraction for penicillin from fermented broth.

Filtration: removal of suspended particles at 1-20C. Clear filtrate or beer is adjusted to pH of 2.5 with dilute phosphoric acid. Penicillin salt is extracted with amyl acetate. The raffinate or water phase is sent to a solvent

recovery still.

The extract is buffered with sodium phosphate solution to a pH of 7.5 in a continuous mixer andthe crude penicillin returns to the aqueous phase.

The mixture is centrifuged and the solvent is recirculated.vi) Purification:

Second acidic extraction Reversion to a pyrogen-free distilled water solution containing the alkaline salt of the desired

element i.e. Na, K , Ca or Al. Purified aqueous concentrate is separated from solvent in a super-centrifuge Pressurised through Biological filters to remove final traces of bacteria and pyrogens. Solution is concentrated by freeze drying or vacuum spray drying.

vii) Crystallization:

Crystalline penicillin salts can be obtained by salting out a saturated solution with a neutral saltcontaining the cations desired in the final penicillin salt.

It is further purified by recrystallization from organic solvents.viii) The oil soluble procaine penicillin is made by reacting a penicillin concentrate (20-30%)

with a 50% aqueous solution of procaine hydrochloride. Procaine penicillin crystallizesfrom this mixture.

Major Engineering problems:

Similar to all fermentation processes, sterile operations are required. Recovery of penicillin: Solvent extraction method has replaced the original activated carbon

adsorption-elution procedure.

It is difficult to design large fermenters (100-200 m3) that provide a uniform and efficient air-liquid contact. Tanks are pressurized to 1 atm gauge pressure and air is added just underneath a

high speed turbine agitator. Foaming is avoided by addition of antifoaming agents like

octadecanol

Safety Factors:

R42/43 : May cause sensitization by inhalation and skin contact.

H317 : May cause an allergic skin reaction.

H334 : May cause allergy or asthma symptoms or breathing difficulties if inhaled.

P302+P352 : IF ON SKIN: Wash with plenty of soap and water.

P304+P341 : IF INHALED: If breathing is difficult, remove victim to fresh air and

keep at rest in a position comfortable for breathing.


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Economics of Penicillin Industry:

Penicillin continues to be a popular reliable antibiotic despite the appearance of at least a dozencompetitive products.

The most desirable fermentation based liquor is derived from maize and soybean both of whichare imported to a large extent. So emphasis is put on much larger production in the agricultural

sector.

Though Penicillin G and V are still important pharmaceutical drugs for the treatment of variousinfectious diseases, their use as drugs is decreasing, since the newer semi synthetic penicillin

and cephalosporin are being used more and more. Therefore, the future demand for Penicillin

G/V will be mainly as raw materials for the manufacture of 6-APA, 7-ADCA and 7-ACA, the

demand for which is steadily increasing.

There are four Indian manufacturers for penicillin-G and they are:i) Hindustan Antibiotics Ltd., Public Sector (H.A.L.) Pimpri, Pune (Maharashtra)(UNICEF)

ii) Indian Drugs & Pharmaceuticals Ltd., Public Sector (I.D.P.L.) Virbhadra, Rishikesh (U.P.)(USSR)

iii) Alembic Chemical Works Co. Ltd., Private Sector Baroda (Gujarat)(Meiji Seiko Kaisha Ltd.,

Japan)

iv) Standard Pharmaceuticals Ltd., Private Sector Serampore (West Bengal)(Synbiotics

Ltd., Baroda (a company of the Sarabhai Group)

Imported Penicillin is cheaper than the indigenous penicillin thats why heavy import duty isapplied on it to reduce the dependence on imported products.

REFRENCES:

i. Outlines of chemical technology by Charles E. Dryden.ii. Shreves Chemical Process Industriesiii. http://www.slideshare.net/ravivishnu/ethanol-production-indiaiv. http://chooseethanol.comv. http://bioweb.sungrant.orgvi. http://www.thecattlesite.com

Group13 Fermentation Industries

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