Вы находитесь на странице: 1из 7

NMEICT-MHRD (Govt.

of India) Project on - Creation of e-Contents on Fermentation Technology

Module-24: Introduction to Product Recovery & Purification

 After the production of the desired fermentation product,the next most important step
includes product recovery and purification.
 The recovery and purification of fermentation products refers Downstream
Processing.
 Downstream processing includes many technologies for laboratory& industrial-scale
separation of biological products.
 The Downstream Processing of fermentation products may be difficult and costly.
 Main purpose of this is to obtain a high-quality product as fast as possible at an
effectiverecovery rate using least costs.
 The recovery costs of microbial products may vary from 15% to 70% of the total
costs.
 It is an essential step in the manufacture of many products such as,
1. antibiotics,
2. hormones,
3. industrial enzymes,
4. natural fragrance and
5. flavor compounds.
 At the time of harvesting, the product concentration may be low in an aqueous
solution, which may containmicro-organisms, cell debris, soluble and insoluble media
components and other metabolic products.
 The product may also be intracellular, heat labile and easily damagedbycontaminating
micro-organisms which increase the difficulties of product recovery.
 If product is heat labile,speed of operation may be the dominant factor in noble
product recovery.
 To ensure the proper processing of harvested broth within a reasonable time
limitprocessing equipment must be of the correct type andof correct size.
 The choice of recovery process is based on the following criteria:
1. The location of the product: Intracellular or Extracellular
2. The quantity of the product in the fermentation broth.
3. The proposed use of the product.

Project control No: RE-02091011297, Christ College, Rajkot, Gujarat, India


NMEICT-MHRD (Govt. of India) Project on - Creation of e-Contents on Fermentation Technology

4. The marginal acceptable standard of purity.


5. The physical and chemical nature of the desired product.
6. The magnitude of bio-hazard
7. The impurities in the fermented broth.
8. The market potential of the product.

Stages of downstream processing

Stage – 1 Removal of Insoluble

 The first stage for the recovery of an extracellular product aims to remove large solid
particles and microbial cells.
 It is possible by various measures like by centrifugation, filtration, sedimentation,
precipitation, flocculation, electro-precipitation, and gravity settling.

Stage – 2ProductIsolation

 In this stage, the broth is extracted into major fractions using various techniques
likeultrafiltration, reverse osmosis, adsorption/ion-exchange/ gel filtration or affinity
chromatography, liquid- liquid extraction, two phase aqueous extraction, precipitation
etc.
 It allows the removal of unnecessary components whose properties differdistinctly
from that of the desired product.

Stage – 3 Product Purification

 Once, the product-containing fraction is purified by fractional precipitation, further


more precise chromatographic techniques and crystallization is used to obtain a highly
concentrated product which is essentially free from impurities.
 Steps in this stage are expensive because itrequires sensitive and sophisticated
equipment which contributes a majorsection of the entire downstream processing
expenditure.

Project control No: RE-02091011297, Christ College, Rajkot, Gujarat, India


NMEICT-MHRD (Govt. of India) Project on - Creation of e-Contents on Fermentation Technology

Stage – 4 Product Polishing

 It is the last processing steps which end with packaging of the product.
 It includes Crystallization, desiccation and spray drying.
 Sometime it alsoincludes operationsto sterilize the product by removing contaminants
which otherwise affect product safety.
 In short the downstream processing involves the following major procedures:

i. Removal of microbial cells and other solid matter:Charged properties of


microbial cells to be exploited by electrophoresis and dielectrophoresis&
flocculation characteristics and magnetic separations to beimproved byultrasonic
treatment
ii. Foam separation:It exploits differences in surface activity of materials. The
material is selectively adsorbed to the surface of gas bubbles rising through a
liquid.Then it separated and finally removed.
iii. Precipitation:This is thechemical process in which solid gets formed in a
solution or inside another solid.Various agentscan be used in precipitation like
acids, bases, salts of sodium and ammonium, organic solvents, non-ionic
polymers (polyethylene glycol), protein binding dyes etc.
iv. Filtration: it is commonly used method which separate suspended particles from
liquid or gas. It uses a porous medium that retain the suspended particles by
allowing the liquid or gas to pass through.So it allows separation of solids by
interfering a medium through which only the fluid can pass.
v. Centrifugation:Centrifuge works using the sedimentation principle, where the
centripetal speed causes separation of denser substances along the radial
direction.Various types of centrifuges are used like the basket centrifuge, the
tubular-bowl centrifuge, the solid-bowl scroll centrifuge etc.
vi. Cell disruption:It is a method for releasing intracellular molecules. To release
the cellular contents from their extremely tough cell wall a number of methods
have been established. For examplephysico mechanical methods such as liquid
shear, solid shear, freeze thawing, agitation with abrasives, and ultra-sonication.
The chemical methods includeosmotic shock, usage of detergents, alkali
treatment, and enzyme treatment.

Project control No: RE-02091011297, Christ College, Rajkot, Gujarat, India


NMEICT-MHRD (Govt. of India) Project on - Creation of e-Contents on Fermentation Technology

vii. Liquid- liquid extraction:It is a method to separate compounds on the basis of


their relative solubilities in two different immiscible liquids.So, an extraction of a
substance from one to another liquid phase in which the desired product is
preferentially soluble is known as liquid-liquid extraction. It is also known as
solvent extraction and partitioning.
viii. Solvent recovery:Distillation is the best method for solvent recovery
whichincludes three stages evaporation, vapour-liquid separation in a column and
condensation.
ix. Chromatography: In chromatography separation is based on differential
partitioning between two phases that is mobile and stationary. Different types of
chromatography techniques are used like adsorption chromatography, ion-
exchange chromatography, gel permeation chromatography, affinity
chromatography, reverse-phase chromatography and high-performance liquid
chromatography.
x. Drying:To allow minimum loss in viability, activity and nutritional value drying
is necessary.It removes water from a heat-sensitive material assuringleast
damage.
xi. Crystallization:It is used for final purification of a diverse range of compounds
including the recovery of organic acids and amino acids.

 It must be remembered that the upstream and downstream processing are integral
parts of an overall process.
 As they are interconnected, neither stage should be developed independently, as this
might result in problems and unnecessary expenditure.
 So, by taking care of some steps in the upstream process productrecovery may be
made easier. i.e.
1. Selection of test strain: By selecting test microorganisms that do not produce any
pigment and/or undesirable metabolites.
2. Environmental setup: By adjusting the production environments to allow least
production of undesirable secondarymetabolites.
 Beside this some process parameters should be checked and maintained like:
1. Time of harvesting
2. pHmaintenance during fermentation and harvesting

Project control No: RE-02091011297, Christ College, Rajkot, Gujarat, India


NMEICT-MHRD (Govt. of India) Project on - Creation of e-Contents on Fermentation Technology

3. Temperature maintenance
4. Use of suitable chemicals for flocculation and separation
 The recovery and purification of many compounds may be achieved by a number of
alternative ways.
 The decision to follow a particular way involves study of the following factors:
1. Capital expenses
2. Processing expenses
3. Probable yield
4. Product quality
5. Availability oftechnical skill
6. Requirements of waste disposal
7. Type of Processing: Continuous or batch
8. Automation
9. Health and safety
 There are so many problems associated with the product recovery program.
 For example, the recovery of extra cellular enzymes might be difficult as it needs
immediate processing.
 One another problem is pigment production by test organisms because some time the
pigment binds to the same resin as the enzyme.
 At the time of foam separation, selection of antifoam should be proper otherwise it
affect ultrafiltration or ion exchange resins used in recovery stages.
 In short it should always be remembered that good recovery starts in the fermentation
by the selection of proper upstream processing like the correct media and proper time
of harvesting.
 The major problem currently faced in product recovery is transfer of small-scale
preparative methods to the production scale without disturbingyield of the process,
quality of the product and purity levelof the product.

Project control No: RE-02091011297, Christ College, Rajkot, Gujarat, India


NMEICT-MHRD (Govt. of India) Project on - Creation of e-Contents on Fermentation Technology

References

 Principles of Fermentation Technology: (2nd edition, by Peter F. Stanbury, Allan Whitaker and
Stephen J. Hall, Butterworth-Heinemann, An imprint of Elsevier Science.)
 Industrial Microbiology: (By Casida L. E.New Age international (P) ltd publications)
 A Text Book of Industrial Microbiology: (2nd edition By WulfCrueger&AnnelieseCrueger)
 Biotechnology: Food Fermentation Microbiology, Biochemistry & Technology Vol. 1 & 2:(By V.K.
Joshi & Ashok Pandey)
 Manual of Industrial Microbiology and Biotechnology: (2nd Edition by Arnold L. Demain and Julian
E. Davies, Ronald M. Atlas, Gerald Cohen, Charles L. Hershberger, Wei-Shou Hu, David H. Sherman,
Richard C. Willson and J. H. David Wu)
 Industrial Microbiology-An introduction: By Michael J. Waites, Neil L. Morgan, John S. Rockey and
Gary Higton)
 Comprehensive Biotechnology-The Principles, Applications and Rugulations of Biotechnology in
Industry, Agriculture and Medicine: (By Mrray Moo Young)
 Fermentation Technology : Up Stream Fermentation Technology- Vol-I: (By H. A. Modi-Pointer
Publications)
 Fermentation Technology : Down Stream Fermentation Technology- Vol-II: (By H. A. Modi-Pointer
Publications)
 Industrial Microbiology by Prescott and Dunn's: (4th edition, edited by Gerald Reed, CBR
publications)
 Fermentation Technology: (By M.L. Srivastava, NAROSA publications)
 Industrial Microbiology: (By A.H. Patel)
 International student edition: Microbiology- A laboratory Manual: (4th edition. By James G.
Chappuccino& Natalie Sherman)
 Bacteriological Techniques: (By F.J. Baker)
 Introduction to Microbial Techniques: (By Gunasekaran)
 Mannual of Industrial Microbiology and Biotechnology: (2nd Edition by Arnold L. Demain and Julian
E. Davies, Ronald M. Atlas, Gerald Cohen, Charles L. Hershberger, Wei-Shou Hu, David H. Sherman,
Richard C. Willson and J. H. David Wu)

Web references

 http://www.homebrew.net/ferment/
 http://www.soyinfocenter.com/HSS/fermentation.php
 http://www.ensymm.com/pdf/ensymm_fermentation_abstract.pdf
 http://scialert.net/fulltext/?doi=jm.2007.201.208
 http://aem.asm.org/content/7/1/57.full.pdf
 http://www.slideshare.net/yongkangbirdnest/lecture-4-sterilization
 http://www.ars.usda.gov/research/publications/publications.htm?seq_no_115=140721
 http://www.scribd.com/doc/30706834/Fermentation-Design
 http://www.wiley-vch.de/books/sample/3527318194_c01.pdf
 http://www.engineersirelandcork.ie/downloads/Biopharmaceuticals%2020Jan09%20-%202%20-
%20Ian%20Marison%20DCU.pdf
 www.yobrew.co.uk/fermentation.php
 http://bioscipub.com/journals/bbb/pdf/19-24.pdf
 http://gertrude-old.case.edu/276/materials/web/immobilizedenzymereview.pdf
 http://download.bioon.com.cn/upload/month_0902/20090223_b809d1c59ba2a6e2abfdJtWiJOiFDm02.att
ach.pdf
 http://bioprocess-maulik.blogspot.in/2007/07/design-of-industrial-fermentation.html

Project control No: RE-02091011297, Christ College, Rajkot, Gujarat, India


NMEICT-MHRD (Govt. of India) Project on - Creation of e-Contents on Fermentation Technology

 http://hsc.csu.edu.au/biology/options/biotechnology/3051/biotechnologyPart3.html
 http://www.rsc.org/ebooks/archive/free/BK9780854046065/BK9780854046065-00001.pdf
 http://www.biotech.upm.edu.my/academics/On%20Line%20Note/Bioprocess/BTK%205301/Lect6%28I
noculum%20Preparation%20and%20Development%29.pdf
 http://www.biotechresources.com/services-strain.shtml
 http://www.idosi.org/wjc/4%281%2909/14.pdf
 http://cheserver.ent.ohiou.edu/Paper-gu/DualFeed.pdf

Project control No: RE-02091011297, Christ College, Rajkot, Gujarat, India