DOWNSTREAM PROCESSING

DOWNSTREAM PROCESSING

The separation and purification of desired fermentation product is downstream processing or bioseperation.

Steps involved in downstream processing are –

• Removal of insoluble's• Product Isolation • Product Purification

Removal of Insoluble’s

• It is the first and the crucial step in downstream processing

• It includes separation of cells, cell debris or other particulate matter from fermentation broth

•Separation by Filtration or Centrifugation

FILTRATIONFiltration separates particles by forcing the fluid through a filtering medium on which solids are deposited

FILTRATION•The two types of filters – filter press and rotary drum filters.

•A filter press is often employed for the small-scale separation of bacteria and fungi from broths.

•For large-scale filtration, rotary drum filters are usually used.

CENTRIFUGATIONIt uses the centrifugal force for the separation of mixtures

More-dense components migrate away from the axis of the centrifuge while less-dense components migrate towards the axis

CENTRIFUGATIONTwo basic types of centrifuges are – tubular and the disk centrifuge

CELL RUPTURE• To obtain intracellular products the cells first have to be disrupted to release these into a liquid medium before further separation can be carried out

• The cell rupture techniques have to be very powerful, but they must be mild enough so that desired components are not damaged

•Physical Methods include mechanical disruption by homogenization, or ultrasonication

CELL RUPTURE• Chemical methods include the treatment of cells with detergents, alkalis, organic solvents, or by osmotic shock

•Detergents : (SDS), Triton X-IOO,

•Alkali : cell walls- saponification of lipids

•Organic solvents: Toluene ruptures the cell wall by penetrating the cell wall lipids

•Biological methods include Enzymatic digestion of the cell wall

PRODUCT ISOLATION

• From the obtained Aqueous layer product is isolated or recovered

•The different techniques used are –

ExtractionPrecipitationAdsorption

Solvent Extraction•It is a method used to separate compounds based on their relative solubilities in two different immiscible liquids

•After the extraction, the solvent-rich phase is called the extract and the residual liquid from which solute has been removed is called the raffinate

Solvent Extraction•Ideal solvent should show – mutual insolubility of the two liquid, easy recoverability, anontoxicity

Precipitation•Formation of a solid in a solution during a chemical reaction

•Solid formed is called the precipitate and the liquid remaining above the solid is called the supernate

Adsorption•Adsorption is the binding of molecules or particles to the surface

•Conventional adsorption is a reversibleprocess as the result of intermolecular forces of attraction

•Ion exchange resins are polymers that are capable of exchanging particular ions within the polymer with ions in a solution that is passed through them

Adsorption•Affinity adsorption is based on the chemicalinteraction between a solute and a ligand which is attached to the surface of the carrier particle by covalent or ionic bonds

PRODUCT PURIFICATION•After a product is recovered or isolated, it may need to be purified further.

•The purification can be accomplished by numerous methods – precipitation, chromatography, electrophoresis,ultrafiltration

Chromatography•In this, separation of compound is based on liquid-liquid partitioning which takes place between static or stationary phase

•Column Chromatography and Gel filtration chromatography

Electrophoresis•It is the separation of charged species based on their specific migration rates in an electrical field

•When a mixture of solutes is placed in an electrical field, the positively charged species are attracted to the anode and the negatively charged ones to the cathode