B.Tech. IV (CH), Semester – VII L T P CES-1: Fundamentals of Biochemical Engineering 3 0 0 3(Elective Subject from Department)• INTRODUCTION (06 Hours)Scope and possibilities; Characteristics and classification of biological matter; Basics of microbial growth; • OVERVIEW OF BIOSEPARATIONS (02 Hours)An Overview of bioseparations; Cell disruptions; Genetically modified organism • SEPARATION METHODS (25 Hours)Filtration; Centrifugation; Adsorption; Extraction; Membrane separation processes; Concepts of precipitation, Chromatography – Basic concepts; Gel filtration Ion exchange chromatography; Hydrophobic chromatography; Affinity chromatography; Suitable examples; Electrokinetic methods of separations; Finishing operations and formulations• INDUSTRIAL APPLICATIONS (12 Hours)Biomass to Biofuels; Bioremediation; Biocatalysts; Biofouling; Microbial Polymer and plastics; Natural resources recovery

Total Contact Hours: 45)BOOKS RECOMMENDED:1.Shuichi Aiba, Arthur E. Humphrey, Nancy F. Millis, “Biochemical Engineering”, 2nd Ed., Academic Press, New York, 1973.2. James E. Bailey, David F. Ollis, “Biochemical Engineering Fundamentals”, 2nd Ed., McGraw hill, 1986.3. L. Weatherley, “Engineering Processes for Bioseparations”, Butterworth-Heinemann Ltd., Oxford, 1995.4. D.L. Pyle, “Separation for biotechnology”, Royal Society of Chemistry, Cambridge, 19945. A. Scragg, “Environmental Biotechnology”, 2nd Ed., Oxford University Press, 2005.

Fundamentals of Biochemical Engineering

Topics already Discussed• The importance of the need of the day of the subject• Similarity and difference between the Chemical and

Biochemical Engineering• Advantages of bioprocesses (ambient temperature,

high product specificity, relatively clean technology)• Application areas like Pharmaceuticals, drug

intermediates, food chemicals, beverages, organic fine chemicals and solvents, industrial enzymes, dairy products etc.

Characteristics of Bioproducts

Need for Downstream Processing

• Downstream processing (bioseparation)is essential part of bioprocess Technology

• The products are manufactured using a variety of equipment like Fermenters or bioreactors, other reactors such as airlift, membrane and immobilized cell reactors are also used.

• The products formed are usually in low concentrations and for separation unit operations are involved

Characteristics of Fermentation Broths

The characteristics of fermentation broths that influence the downstream processing of biomolecules include:

•The type of microorganisms and their morphological features (size and shape)

• Concentration of cells, products and byproducts

•Physical and rheological characteristics

Morphology of Cells

Figure 1: Morphology of Cell

•Cells size ranges from 1 micro m to 4000 micro m

•The cells and the cell agglomerates exhibit a variety of shapes.

•Bacterial and yeast cells occur mostly as homogeneously suspended particles in the fermentation broth

•Fungi form a network of hyphi thereby increasing the viscosity of the broth

•Cell size decreases capacity of the separation process decreases and cost increases

Concentration of cells, products and byproducts

The concentrations of the biomass and that of the products in the fermentation broth are important in deciding on the choice of the separation process

Physical and Rheological CharacteristicsThe density of the dry biomass would be about 1400 kg/m3; however the density of the fermentation broth is lower , around 1100 kg/m3 as the cells have a high water content of about 70-80%.

The rheological property of fermentation broth is of importance for downstream processing in the case of centrifugation and membrane separations.

Newtonian and Non-newtonian models can describe most of the fermentation broths.

The viscosity of the broth is strongly influenced by the cell concentration as well as cell shape and to a minor extent by the changes in the concentration of the nutrients and other metabolites.

The stages of bioseparations

The bioseparation stages are:1. Product release and pre-treatment: Involves cell disruption and release of intracellular products2. Removal of insoluble's or particulates that is solid- liquid separation: Filtration or centrifugation achieves the removal or separation of particulates 3. Product concentration and recovery: Involves adsorption, extraction, precipitation or membrane separation4.Purification: Involves high resolution techniques like chromatography and affinity separations5. Finishing operations and formulations: Includes drying, crystallization etc.Note: This is purely for academic purpose and there is no rigid stages for bioseparation

The separation Techniques

Typical bioprocesses

Typical bioprocesses

Cell wall of microorganisms

Cell wall•The murein layer is about 10-80 nm thick, made of petidoglycan exists in one form or other in almost all the species.• The space below the murien layer called periplasmic space, is about 8 nm thick and often contains enzymes•The inner layer called the plasma or cytoplasmic membrane is about 8 nm is a double layer made of phospholipids and some proteins and metal ions• The cell interior, called the cytoplasm, is an aqueous solution of salts, amino acids and biopolymers including proteins, enzymes, RNA and DNA.It is necessary to rupture the cell to release the protein in the cytoplasm

Cell Disruption

Cells for disruptions

Physical Methods

Thermoly

sis

For

products of stable of heat

shock

Osmotic shoc

k

Cells in pure water (double amt) , cells

swells

and

disrupts

Ultrasonication

Ultrasound waves of frequency .20kHz

ruptures the

cells

Chemical and Enzymatic methods

Alkali treatment

Detergent solubilization

Cell wall permeabilization

Lipid solubilization by organic solvents

Enzyme digestion

Lytic enz yme to the cell

suspension

Mechanical Disruptions

Bead Mill disruption

High prressure

homogenizer

Disruption of microbial cell

Bead mill disruption