CSIR-NET LIFE SCIENCES

Unit 12: Biochemical Engineering & Industrial Biotechnology

Complete Topic-Wise Syllabus

Bioprocess Engineering • Enzymes • Downstream Processing • Metabolic Engineering

A. Introductory Mathematics

📐 1. Calculus Review

• Functions and limits
• Differentiation
• Partial differentiation
• Integration
• Definite and indefinite integration
• Applications of derivatives
• Applications of integration

📌 Important Calculus Concepts

• Rate of change
• Maxima and minima
• Area under curves
• Optimization
• Derivative-based analysis of biological processes

📊 2. Ordinary Differential Equations

• First-order differential equations
• Initial value problems
• Separable equations
• Linear differential equations
• Applications to biological systems

📈 3. Second and Higher Order Differential Equations

• Second-order differential equations
• Higher-order equations
• Homogeneous equations
• Non-homogeneous equations
• Boundary conditions

🔢 4. Linear Algebra

• Matrices
• Determinants
• Vectors
• Systems of linear equations
• Eigenvalues
• Eigenvectors
• Matrix operations

💻 5. Numerical Methods

• Numerical solution of equations
• Root finding
• Numerical integration
• Numerical differentiation
• Approximation methods
• Computational solution of differential equations

dx/dt = f(x,t)

General idea: Initial condition → Mathematical model → Numerical solution

B. Engineering Principles

⚖️ 1. Material Balance

Material balance follows the conservation of mass principle and is fundamental for analyzing bioprocesses.

Accumulation = Input − Output + Generation − Consumption

• Overall material balance
• Component balance
• Batch material balance
• Continuous material balance
• Steady-state balance

🔥 2. Energy Balance

• Conservation of energy
• Energy input
• Energy output
• Heat generation
• Heat removal
• Energy requirements of bioprocesses

⚙️ 3. Steady-State Material and Energy Balance

At steady state, the accumulation term becomes zero.

At steady state:

Input − Output + Generation − Consumption = 0

🧪 4. Properties of Substances

• Density
• Viscosity
• Specific heat
• Thermal conductivity
• Diffusivity
• Pressure
• Temperature

🌊 5. Introduction to Transport Phenomena

• Momentum transfer
• Heat transfer
• Mass transfer

Transport Process	Major Quantity Transferred
Momentum Transfer	Momentum
Heat Transfer	Thermal energy
Mass Transfer	Chemical species

💨 6. Momentum Transfer

• Fluid flow
• Viscosity
• Shear stress
• Laminar flow
• Turbulent flow
• Fluid behaviour in bioreactors

🔥 7. Heat Transfer

• Conduction
• Convection
• Radiation
• Heat exchangers
• Temperature control in bioprocesses

💧 8. Mass Transfer

• Molecular diffusion
• Concentration gradients
• Mass-transfer coefficients
• Gas-liquid mass transfer
• Oxygen transfer in bioreactors

🏭 9. Mass Transfer Equipment

• Absorption equipment
• Extraction equipment
• Membrane systems
• Gas-liquid contactors
• Bioprocess separation equipment

C. Thermodynamics in Biological Systems

🔥 1. First Law of Thermodynamics

The first law represents the conservation of energy.

ΔU = Q − W

• Internal energy
• Heat transfer
• Work
• Energy conservation

🌡️ 2. Second Law of Thermodynamics

• Entropy
• Direction of spontaneous processes
• Irreversibility
• Energy degradation

ΔS ≥ 0

🧬 3. Biological Systems as Open Non-Equilibrium Systems

Living systems continuously exchange matter and energy with their surroundings and therefore operate away from thermodynamic equilibrium.

• Continuous energy input
• Continuous material exchange
• Metabolic flux
• Energy dissipation
• Maintenance of biological organization

🧬 Important Concept:

Cells maintain internal organization by continuously exchanging matter and energy with their environment.

⚠️ 4. Limitations of Classical Thermodynamics

• Classical thermodynamics mainly describes equilibrium states.
• Living systems are dynamic and non-equilibrium.
• Biological systems contain complex coupled processes.
• Energy and matter continuously flow through living systems.

↔️ 5. Thermodynamic Flux and Force

• Thermodynamic force drives a process.
• Flux represents the rate of a thermodynamic process.
• Flux-force relationships describe irreversible processes.

♨️ 6. Entropy Production

Entropy production is associated with irreversible processes occurring in biological systems.

• Heat dissipation
• Metabolic reactions
• Transport processes
• Chemical reactions

📐 7. Constitutive Equations

• Describe relationships between thermodynamic forces and fluxes.
• Used in non-equilibrium thermodynamics.
• Useful for describing biological transport and reaction processes.

🔗 8. Thermodynamics of Coupled Biochemical Reactions

Unfavourable reactions can be driven by coupling them with favourable reactions.

Example:

ATP hydrolysis can be coupled to energetically unfavourable biochemical reactions.

⚡ 9. Thermodynamic Analysis of Oxidative Phosphorylation

• Electron transport
• Proton gradient
• Proton motive force
• ATP synthesis
• Energy coupling

🔄 10. Glycolytic Oscillations

• Periodic changes in metabolic intermediates
• Dynamic regulation of glycolysis
• Nonlinear biochemical behaviour
• Feedback regulation

⏰ 11. Biological Clocks

• Circadian rhythms
• Oscillatory biochemical networks
• Feedback loops
• Periodic gene expression
• Metabolic rhythms

D. Bioprocess Engineering and Technology

🦠 1. Microbial Growth

• Microbial growth principles
• Growth phases
• Lag phase
• Exponential phase
• Stationary phase
• Death phase

📈 2. Factors Affecting Microbial Growth

• Temperature
• pH
• Nutrients
• Oxygen
• Osmotic pressure
• Inhibitory compounds

📊 3. Growth Kinetics

μ = (1/X)(dX/dt)

Monod Equation: μ = μmax S / (Ks + S)

• Specific growth rate
• Maximum specific growth rate
• Substrate concentration
• Saturation constant

🧪 4. Batch Culture

• Closed cultivation system
• No continuous nutrient feed
• Growth occurs through different phases
• Widely used for laboratory and industrial fermentation

💧 5. Fed-Batch Culture

• Nutrients are continuously or intermittently added.
• Useful for controlling substrate concentration.
• Can prevent substrate inhibition.
• Widely used for high-cell-density cultivation.

🔄 6. Continuous Culture

• Fresh medium continuously enters the reactor.
• Culture broth continuously leaves the reactor.
• Used for steady-state operation.
• Dilution rate is an important parameter.

📌 Major Culture Modes

System	Main Characteristic
Batch	No continuous feed or removal during cultivation
Fed-Batch	Feed added during cultivation
Continuous	Continuous feed and product removal

🏭 7. Introduction to Bioreactors

• Batch bioreactor
• Fed-batch bioreactor
• Continuous stirred-tank reactor
• Plug-flow reactor
• Enzyme reactor

🧫 8. Sterilization

• Medium sterilization
• Equipment sterilization
• Air sterilization
• Steam sterilization
• Filtration
• Prevention of contamination

⚖️ 9. Mass and Energy Balance in Microbial Processes

• Substrate consumption
• Biomass formation
• Product formation
• Oxygen consumption
• Carbon dioxide production
• Heat generation

🫧 10. Dissolved Oxygen

• Oxygen requirement of aerobic microorganisms
• Dissolved oxygen concentration
• Oxygen limitation
• Oxygen transfer rate

OTR = kLa (C* − C)

🌬️ 11. Oxygen Mass Transfer

• Gas-liquid oxygen transfer
• Volumetric mass-transfer coefficient (kLa)
• Oxygen saturation concentration
• Oxygen uptake rate

💨 12. Aeration and Agitation

• Air supply
• Mixing
• Gas dispersion
• Mass transfer enhancement
• Heat distribution

🌊 13. Fluid Rheology

• Viscosity
• Newtonian fluids
• Non-Newtonian fluids
• Shear stress
• Shear rate

💊 14. Industrial Fermentation Products

• Antibiotics
• Organic acids
• Alcohols
• Bioplastics
• Vitamins
• Industrial enzymes

🧪 15. Biotransformation of Steroids

• Microbial conversion of steroid compounds
• Selective chemical modification
• Use of microbial enzymes
• Industrial pharmaceutical applications

📋 16. Process Flow Sheet

Raw Materials → Medium Preparation → Sterilization → Inoculum Development → Fermentation → Harvest → Downstream Processing → Purification → Formulation → Final Product

💰 17. Process Economics

• Raw material cost
• Energy cost
• Equipment cost
• Labour cost
• Downstream processing cost
• Product yield
• Process productivity
• Overall profitability

E. Enzymes and Microbial Technology

🧪 1. Enzymes in Organic Solvents

• Enzyme activity in non-aqueous environments
• Organic solvent tolerance
• Altered enzyme specificity
• Industrial biocatalysis

⚗️ 2. Enzymes in Ionic Liquids

• Non-conventional reaction media
• Enzyme stability
• Solubility advantages
• Biocatalytic applications

🧬 3. Biocatalysts

Biocatalysts are biological molecules or organisms used to catalyze chemical reactions.

• Enzymes
• Whole cells
• Engineered enzymes
• Microbial catalysts

🧬 4. Enzyme Engineering

• Modification of enzyme properties
• Improved stability
• Improved catalytic activity
• Altered substrate specificity
• Improved solvent tolerance

🎲 5. Random Approach to Protein Engineering

• Random mutagenesis
• Generation of mutant libraries
• Screening of variants
• Selection of improved enzymes

🎯 6. Rational Protein Engineering

• Structure-guided mutation
• Knowledge-based modification
• Active-site engineering
• Specific amino-acid substitution

🧬 Protein Engineering:

Random Approach → Random Mutations → Large Library → Screening

Rational Approach → Structural/Functional Knowledge → Targeted Mutation → Screening

⚙️ 7. Biocatalysis

• Enzyme-mediated chemical conversion
• High specificity
• Mild reaction conditions
• High stereoselectivity
• Industrial synthesis

🔗 8. Enzyme Immobilization

• Physical adsorption
• Covalent attachment
• Entrapment
• Encapsulation
• Cross-linking

🦠 9. Whole-Cell Immobilization

• Immobilization of complete microbial cells
• Multiple enzyme systems retained within cells
• Useful for multistep biotransformations

🏭 10. Immobilized Enzyme Reactors

• Batch immobilized enzyme reactors
• Continuous reactors
• Packed-bed reactors
• Fluidized-bed reactors

📈 11. Kinetics of Immobilized Enzymes

• Effect of immobilization on enzyme activity
• Substrate accessibility
• Effective reaction rate
• Internal diffusion limitations

🧮 12. Diffusional Resistance

Diffusional resistance occurs when substrate or product movement through the immobilization matrix limits the overall reaction rate.

📐 13. Thiele Modulus

The Thiele modulus relates the intrinsic reaction rate to diffusion within an immobilized enzyme particle.

Thiele Modulus: φ ∝ √(Reaction Rate / Diffusion Rate)

📌 Exam Concept:

A high Thiele modulus generally indicates stronger internal diffusion limitations in an immobilized enzyme particle.

F. Downstream Processing in Biotechnology

🦠 1. Biomass Removal

• Filtration
• Centrifugation
• Sedimentation
• Membrane separation

💥 2. Cell Disruption

• Mechanical disruption
• High-pressure homogenization
• Ultrasonication
• Bead milling
• Enzymatic disruption
• Chemical disruption

🧂 3. Precipitation by Salts

• Protein precipitation
• Salt-induced precipitation
• Selective protein recovery
• Concentration of biomolecules

🧪 4. Precipitation by Solvents

• Organic solvent precipitation
• Protein recovery
• Selective separation

🧫 5. Membrane-Based Purification

• Microfiltration
• Ultrafiltration
• Nanofiltration
• Reverse osmosis
• Diafiltration

Technique	Major Application
Microfiltration	Cells and large particles
Ultrafiltration	Macromolecules and proteins
Nanofiltration	Small solutes and ions
Reverse Osmosis	Water removal and concentration

🧲 6. Adsorption

• Binding of molecules to solid surfaces
• Activated carbon
• Ion-exchange materials
• Selective product recovery

🧪 7. Chromatography

• Ion-exchange chromatography
• Size-exclusion chromatography
• Affinity chromatography
• Hydrophobic interaction chromatography

💧 8. Solvent Extraction

• Partitioning between immiscible phases
• Recovery of small molecules
• Recovery of organic products

💧 9. Aqueous Two-Phase Extraction

• Two aqueous phases
• Selective partitioning of biomolecules
• Protein purification
• Cell and enzyme separation

🌫️ 10. Supercritical Fluid Extraction

• Uses supercritical fluids
• High mass-transfer properties
• Useful for extraction of bioactive compounds

🔥 11. Drying

• Removal of water
• Improved product stability
• Storage of biological products
• Spray drying
• Freeze drying

🔬 Downstream Processing Sequence:

Harvest → Biomass Removal → Cell Disruption → Clarification → Concentration → Purification → Polishing → Drying → Final Product

G. Bioprocess Plant Design

🏭 1. General Design Information

• Process requirements
• Raw material requirements
• Production capacity
• Equipment requirements
• Utilities
• Safety requirements
• Waste management

📋 2. Process Flow Sheet

A process flow sheet represents the sequence of major operations and material flows in an industrial process.

🏭 Typical Bioprocess Flow:

Raw Material → Preparation → Sterilization → Inoculum → Bioreactor → Harvest → Separation → Purification → Formulation → Packaging

📈 3. Scale-Up

Scale-up involves transferring a process from laboratory scale to pilot or industrial scale while maintaining desired performance.

• Laboratory scale
• Pilot scale
• Industrial scale
• Mixing
• Oxygen transfer
• Heat transfer
• Power input

🔽 4. Scale-Down

Scale-down models reproduce important industrial conditions at smaller scales for process development and testing.

⚠️ 5. Scale-Up Problems

• Reduced oxygen transfer
• Mixing limitations
• Heat removal problems
• pH gradients
• Substrate gradients
• Increased shear effects
• Changes in rheology

🧪 6. Scale-Up of Downstream Processes

• Large-volume clarification
• Membrane filtration
• Chromatography
• Extraction
• Drying
• Product recovery

⚙️ 7. Selection of Bioprocess Equipment

• Bioreactor selection
• Agitator selection
• Aeration system
• Heat exchanger
• Filtration system
• Centrifuge
• Chromatography system
• Drying equipment

📐 8. Equipment Specifications

• Capacity
• Material of construction
• Operating pressure
• Operating temperature
• Mixing requirements
• Sterility requirements
• Cleaning requirements

🎯 Important Scale-Up Principle:

A successful scale-up must maintain critical process parameters and critical quality attributes while achieving the required productivity and product quality.

H. Metabolic Engineering and Synthetic Biology

🧬 1. Metabolic Engineering

Metabolic engineering involves deliberate modification of cellular metabolic pathways to improve production of desired compounds.

• Pathway modification
• Gene overexpression
• Gene deletion
• Promoter engineering
• Flux redistribution
• Improvement of product yield
• Improvement of productivity

🔄 2. Metabolic Flux

Metabolic flux represents the rate at which metabolites move through biochemical pathways.

Substrate ↓ Pathway A → Desired Product ↓ By-products

🎯 3. Strategies of Metabolic Engineering

• Overexpression of rate-limiting enzymes
• Deletion of competing pathways
• Enhancement of precursor supply
• Reduction of by-product formation
• Modification of cofactors
• Optimization of pathway balance

🧬 4. Synthetic Biology

Synthetic biology combines biological principles with engineering concepts to design, construct and modify biological systems with predictable functions.

• Design of genetic circuits
• DNA assembly
• Promoter engineering
• Regulatory elements
• Genetic switches
• Engineered microorganisms

🧩 5. Biological Parts

• Promoters
• Ribosome-binding sites
• Coding sequences
• Terminators
• Regulatory sequences
• Genetic switches

🔧 6. Design-Build-Test-Learn Cycle

🧬 Synthetic Biology Engineering Cycle:

Design → Build → Test → Learn → Redesign

🧪 7. Applications

• Biofuel production
• Pharmaceutical production
• Industrial enzymes
• Bioplastics
• Specialty chemicals
• Vitamins
• Food ingredients
• Biomaterials

🤖 8. Computational Metabolic Engineering

• Genome-scale metabolic models
• Flux balance analysis
• Pathway prediction
• Metabolic network analysis
• Strain optimization
• Computer-aided pathway design

🚀 Metabolic Engineering + Synthetic Biology:

Genome → Pathway Analysis → Target Identification → Genetic Modification → Metabolic Flux Optimization → Strain Construction → Product Formation

📚 Unit 12 – Quick Revision Map

A → Introductory Mathematics

B → Engineering Principles

C → Thermodynamics

D → Bioprocess Engineering

E → Enzymes & Microbial Technology

F → Downstream Processing

G → Bioprocess Plant Design

H → Metabolic Engineering & Synthetic Biology

🎯 High-Yield CSIR-NET Topics

• Calculus and differential equations
• Material and energy balances
• Transport phenomena
• Momentum, heat and mass transfer
• First and second laws of thermodynamics
• Non-equilibrium thermodynamics
• Entropy production
• Coupled biochemical reactions
• Oxidative phosphorylation thermodynamics
• Glycolytic oscillations
• Biological clocks
• Microbial growth kinetics
• Monod equation
• Batch, fed-batch and continuous culture
• Bioreactor types
• Sterilization
• Dissolved oxygen
• Oxygen transfer rate
• kLa
• Aeration and agitation
• Fluid rheology
• Fermentation technology
• Process economics
• Enzyme engineering
• Random vs rational protein engineering
• Biocatalysis
• Enzyme immobilization
• Immobilized enzyme reactors
• Diffusional resistance
• Thiele modulus
• Biomass removal
• Cell disruption
• Protein precipitation
• Membrane separation
• Adsorption
• Chromatography
• Solvent extraction
• Aqueous two-phase extraction
• Supercritical extraction
• Drying
• Bioprocess plant design
• Scale-up and scale-down
• Equipment selection
• Metabolic engineering
• Metabolic flux
• Synthetic biology
• Genetic circuits
• Design-Build-Test-Learn cycle
• Genome-scale metabolic models
• Flux balance analysis

🧮 Important Formula Revision

Material Balance: Accumulation = Input − Output + Generation − Consumption

First Law: ΔU = Q − W

Specific Growth Rate: μ = (1/X)(dX/dt)

Monod Equation: μ = μmax S / (Ks + S)

Oxygen Transfer Rate: OTR = kLa(C* − C)

🏭 Complete Bioprocess Workflow

Raw Material → Medium Preparation → Sterilization → Inoculum Development → Bioreactor → Fermentation → Harvest → Downstream Processing → Purification → Drying/Formulation → Final Product

🧪 Downstream Processing Memory Trick

Remove → Break → Separate → Concentrate → Purify → Dry

Biomass Removal → Cell Disruption → Clarification → Concentration → Chromatography → Polishing → Drying

🧠 One-Line Unit 12 Revision:

Biochemical engineering applies engineering, mathematical and thermodynamic principles to biological systems, while industrial biotechnology uses microorganisms, enzymes and engineered cells for large-scale production of valuable biological and chemical products.

🧬 CSIR-NET Life Sciences – Unit 12

Biochemical Engineering • Bioprocess Technology • Downstream Processing

Understand → Calculate → Apply → Practice → Revise