ERT 313BIOSEPARATION ENGINEERING

INTRODUCTION

Prepared by:Pn. Hairul Nazirah Abdul Halim

Topic Outline• Introduction• What is separated in bioseparation?• Economic Important in Bioseparation• Nature of Bioseparation• Basis of Separation in Bioseparation Process• Bioseparation Techniques• The RIPP Scheme• Example of Bioseparation

BIOPROCESS ENGINEERING

UPSTREAM

DOWNSTREAM

Growth of biomolecules usually by bacterial or mammalian cell lines & harvest - BIOREACTOR

cell mass from the UPSTREAM are processed to meet purity and quality requirements

• What is Bioseparation Engineering???

- Systematic study of the scientific & engineering principles utilized for the large-scale purification of biological products.

Introduction

• What is Bioprocessing???- deals with manufacture of biochemicals- biopharmaceuticals- foods- nutraceuticals- agrochemicals

What is separated in Bioseparation?• Biological derived products can be classified based

on:a) chemical nature (Table 1.1)b) application (Table 1.2)

• Particle – liquid separation :1. Separation of cells from cell culture medium2. Separation of blood cells from plasma in the manufacture of plasma proteins3. Removal of bacteria and viruses from protein solutions

• Solute – solvent separation1. Removal of a solvent from a solute product (e.g. protein concentration enrichment)2. Removal of dissolved impurities from a liquid product

• Solute – solute separation1. Separation of serum albumin from other serum proteins

• Liquid – liquid separation:1. Separation of ethanol and acetone from an aqueous medium (in the manufacture of solvent)

Economic Important in Bioseparation

Nature of Bioseparation• Bioseparation is largely based on chemical separation processes.• The separations usually aim to achieve removal of specific

components, in order to increase the added value of the products, which may be the residue, the extracted components or both.

• Bioseparation is differ from chemical separation:1. biological products – very low concentrations in the starting material2. impurities have similar chemical/physical properties with the target products3. Denaturation & degradation4. Stringent quality requirement for products – therapeutic products should be free from endotoxins and pyrogens.

Basis of Separation in Bioseparation Processes

Factor Types of Separation

Size Filtration, Membrane Separation, Centrifugation

Density Centrifugation, Sedimentation, FloatationDiffusivity Membrane Separation

Shape Centrifugation, Filtration, SedimentationPolarity Extraction, Chromatography, Adsorption

Solubility Extraction, Precipitation, CrystallizationElectrostatic

ChargeAdsorption, Membrane Separation, Electrophoresis

Volatility Distillation, Membrane Distillation, Pervaporation

Types of Separation

1. Gas-liquid separation - Absorption2. Vapor-liquid separation - Distillation3. Liquid-liquid separation – Extraction 4. Fluid – solid separation – Adsorption, Leaching,

Chromatography, Crystallization5. Mechanical- physical separation – Precipitation,

Centrifugation, Filtration, Membrane Separation.

Bioseparation TechniquesLow-resolution + high-throughput

High-resolution + low-throughput

Cell DisruptionPrecipitation

CentrifugationLiquid-liquid Extraction

LeachingFiltration

Supercritical Fluid ExtractionMicrofiltrationUltrafiltration

AdsorptionCrystallization

UltracentrifugationChromatography

Affinity separationElectrophoresis

Common Stage in BioseparationRIPP Scheme• Recovery• Isolation (volume reduction)• Purification• Polishing

•RIPP SCHEME

• Recovery of product– Filtration, centrifugation, microfiltration

• Isolation of product (volume reduction)– Cell disruption, extraction, adsorption, ultrafiltration, precipitation

• Purification– Adsorption, elution chromatography, ultrafiltration, electrophoresis, precipitation, crystallization

• Polishing– Crystallization, drying, auxiliary process, solvent recovery

Example: Fractional Precipitation

• Ethanol fractionation of human plasma protein factors:

pH, temperature, ionic strength & protein concentration change

• cohns method 6: primary means of preparing albumin, γ-globulin, fibrinogen

Bioseparation of reagent grade monoclonal antibody from cell culture supernatant