Design Features of Stirred Tank Bioreactor

Design Features of  Stirred Tank Bioreactor

Dr. Rosfarizan Mohamad Dept. of Bioprocess Technology, FBSB, UPM

Main function : • to provide a controlled environment for the growth of a microorganism,, or a defined microorganism mixture of microorganisms microorganisms,, to obtain a desired product product..

Consideration in designing and constructing a bioreactor 1) Microbiological and biochemical characteristics of  cell systems. 2) Hydrodynamic characteristics of the bioreactor. 3) Mass and heat characteristics of the bioreactor. 4) Kinetic of cell growth and product formation. 5) Genetic stability characteristics of the cell system.

•6) Aseptic equipment design. •7) Control of bioreactor environment (both macro and micro-environments). •8) Implication of bioreactor design on downstream product separation. •9) Capital and operating costs of the bioreactor. •10) Potential for bioreactor scale up.

Stirred tank bioreactor and its main component

• Most important bioreactor for industrial application (Low capital and operating cost). •However, no single system adequately meets the needs all biological systems can be constructed.

• Laboratory scale bioreactors liquid volume < 10 L constructed out of Pyrex glass.

• For larger bioreactors, stainless steel is used. Stainless steel = refers to various alloys of primarily iron, nickel and chromium.

•Different grade of SS = 302, 304, 316, 318. (higher the number, the greater the resilience of the steel.

•316 L – most widely used (L indicates the steel has low carbon content). •The stainless steel used in bioreactors are polished to a mirror finish (makes cleaning and sterilization easier). •Components joined in an oxygen-free environment to avoid corrosion, displaced by argon (TIG technique, Total Inert Gas)

STANDARD GEOMETRY OF A STIRRED TANK BIOREACTOR

• .

Geometry measurement of stirred tank bioreactor • A mechanically stirred tank bioreactor fitted with a sparger and a Rushton turbine impeller will typically have relative dimensions (Table 1)

Table 1: Geometry Dimension of Stirred Tank Bioreactor Typical values ~0.7-0.8

Remarks

Height of reactor to diameter H /D t t of tank

~1 - 2

European reactors tend to be taller than those designed in the USA

Diameter of impeller diameter in tank

1/3 - 1/2

Rushton Turbine reactors are generally 1/3 of the tank diameter. Axial flow impellers are larger.

 Height of liquid in reactor to HL /Ht height of reactor

to Da /Dt

Diameter of baffles to diameter Db /Dt of tank Impeller blade height diameter of impeller

to W/Da

~0.0.08 0.1 0.2

Impeller blade width diameter of impeller

to L/Da

0.25

Distance between middle of E/W impeller blade and impeller blade height

1

Depends on the level of foaming produced during the fermentation

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A tank's height:diameter ratio is often referred to as its aspect ratio. A stirred tank bioreactor is approximately cylindrical in shape. It has a total volume (Vt) of 100,000 litres. The geometry of the reactor is defined by the following ratios: Dt:Ht

0.50

Da:Dt

0.33

Db:Dt

0.10

•Ex: A cylindrical reactor has a liquid volume of 100,000 L. It has an aspect ratio of 1:1. The height of the liquid in the reactor will be approximately….?? (Ans:5.03m)

Head space volume •A bioreactor is divided in a working volume and a head space volume. •A working volume = fraction of the total volume taken by the medium, microbes and gas bubbles = 70- 80% of the total fermenter volume = but depending on the rate of foaming formation during fermentation. •The remaining volume is called the head space volume.

A modern mechanically agitated  bioreactor will contain: •

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An agitator system An oxygen delivery system A foam control system

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A temperature control system

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A pH control system

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Sampling ports

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A cleaning and sterilization system.

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A sump and dump line for emptying of the reactor.

Main components and parts of stirred tank bioreactor

 AGITATION SYSTEM SYSTEM (Agitator and and Baffles)

Function: •provide good mixing and thus increase mass transfer rates through the bulk the bulk liquid and  bubble boundary layers layers.. •provide the appropriate shear conditions required for the  breaking up of bubbles bubbles..

Stirrer's shaft seal - subject to high mechanical loads. - Important for good operation; to avoid risk of contamination; shaft jamming, possible leak. - All component with contact to the product must have smooth surfaces and may not have any recesses. - < 1 L bioreactor = plastic coated magnetic rods are used located on bearings on the bottom (associated with possible aeration problems) or suspended for rotational movement.

• Magnetically coupled agitator from ELECTROLUX for pilot scale.

•Stirred vessels > 10 L volume, mechanical drive coupling together with rotating mechanical seals. •Seal components: carbon and ceramic. •Laboratory scale: simple rotating mechanical seal. •Larger scale: double-action rotating mechanical seals; achieving longer operation times. •The rotating mechanical seal must be free from cracks and the cavities between the packing must be steamable.

Rotating mechanical seals

Drive Configuration (the drive for the agitator shaft) • Can be installed either above through the reactor cover or from below through the bottom flange. • Bottom drive; - leaves cover free for the installation of other components and connection, the agitator shaft can kept shorter. • Top drive; - easily protected against leakages; sterility is easier to maintain.

Agitator (has to fulfill the following tasks) -

Dispersion of culture air in the form of   bubbles and the creation of higher transfer rates at the gas/liquid interface for supplying oxygen to the microorganisms and extracting CO2.

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Creation of constant living conditions (substrate, pO2, pH, temperature, etc.) by homogenous distribution.

- Improvement of the heat transfer at the heat

transfer surfaces for dissipating the heat generated by the  biological reaction and mechanical work.

Different types of impeller

EKATO INTERPROP impeller

• Multi-blade disc impellers- produce radial flow and high energy dissipation density in the proximity of the agitator. • Propeller impellers- create an axial flow (for mammalian cell cultivation), low shear forces in connection with low oxygen requirements are applicable • INTERMIG impellers- create both radial and axial flows, for high viscous media or products

•Kaplan Turbine – normally for loop systems(mixing effect is achieved by circulating the liquid) •EKATO INTERPROP – improved heat and mass transfer, high dispersion of drops and bubbles, homogenous distribution of solid particles as well as low investment costs.

The functions of the agitation system include; • increasing mass transfer rates, especially KLa, through the bulk liquid • providing appropriate levels of shear • increasing heat transfer rates • reducing the size of boundary layers

KLa can be influenced in 3 ways by agitation; • a) The impeller can break up the air into smaller bubbles thus increasing the gas/liquid interfacial area • b) Agitation can delay the loss of air from the bioreactor. • c) Turbulence shear can reduce film thickness at the gas/liquid interface.

Effect of Impeller Spacing on Flow Patent and Power Absorption a) Spaced too closely – multiple impellers tend to  behave like a single large impeller.  b) Spaced too far apart – appearing regions in the liquid (Stagnant area). - Optimal spacing is about one impeller apart, with the lowest impeller about one impeller above the bottom of the vessel.

Impellers have 2 distinct function • I) To provide mixing by pumping liquid round the vessel. For this function, it requires a large diameter, low speed impeller with a small number of blades.

• II) To dispersed the injected gas steam as small  bubbles and re-disperse coalesced bubbles. It requires a high speed, small diameter impeller with a large number of blades

Agitators characteristics: Radial Flow - the liquid flow from the impeller is initially directed towards the wall of the reactor Axial Flow - The liquid flow from the impeller is directed downwards towards the base of the reactor

Radial flow impellers ( contain two or more impeller blades which are set at a vertical pitch)

Radial flow impellers - Shear characteristics

Radial flow impellers

Flow pattern using radial flow impellers.

Radial flow impellers - Rushton turbine

Generation of high shear conditions by radial flow impeller

Radial flow impellers - Rushton turbine

Six bladed Rushton turbine impeller

Function of Rushton turbine impeller. •Mixing is achieved with the use of baffles. •higher input energy •Mixing is not efficient as axial flow mixing

Axial flow impellers

Marine and hydrofoil impellers

Flow pattern of axial impeller •More energy efficient, effective at lifting solids from the base of the tank, low shear properties •Use for shear sensitive process such as crystallization, precipitation reactions and culture of animal cells. •Not suitable for bacterial and fungal aerobic fermentation (ineffective breaking up bubbles).

Axial flow impellers - Intermig Impeller •- for microbial fermentation •Bottom: has large axial flow section •The tips of impeller contain finger like extensions, create a turbulence wake for breaking bubbles.

Flow pattern created by INTERMIG impeller

•Overall shear conditions in the reactor are lower than would be generated by a radial flow impeller (Rushton turbine).

Top entry and bottom entry impellers Bottom driven impeller (need

Top driven impeller (more expensive to install)

higher maintenance due to damage of the seals by particulates in the medium and by medium components that crystallize in the seal when reactor is not in use)

Baffles (Aid in satisfactory mixing, heat and mass transfer) • Liquid mixing; a) Baffled b) Unbaffled

Unbaffled bioreactor. Note the presence of a large vortex. The liquid is circulating around the impeller.

Baffled bioreactor. Note the presence of small bubbles from gas entrainment and the absence of a large vortex.

Formation of eddies by baffles

•Baffles break the liquid flow lines causing the formation of turbulent eddies.

OXYGEN DELIVERY SYSTEM Consists of: • a compressor • inlet air sterilization system • an air sparger • exit air sterilization system

A compressor –  • forces the air into the reactor, need sufficient pressure to force the air; for large reactors, produce air at 250kPa.The air should be dry and oil free so as to not block the inlet air filter (not to use “instrument air”)

Air sterilisation system –  •to prevent contaminating organisms from entering the reactor as well as to prevent the microorganism in the reactor from contaminating the air. •Common method: filtration •Smaller bioreactor ( 1000L) – pleated membrane filters housed in polyproplylene cartridge. •By pleating the membrane, it is possible to create a compact filter with a very large surface area for air filtration. Large scale membrane filtration is very expensive process. •filtration is not possible; too expensive; Heat sterilisation (steam is use to sterilize the air)

•For small reactor, the exit air system, will include a condenser.

• Condenser –  • simple heat exchanger through which cool water is passed; minimize water evaporation and the loss of volatiles; Drying the air also prevents blocking the exit air filter with water

 AIR SPARGER

Ring Sparger

•The air sparger breaks the incoming air into small bubbles •The sparger must located below the agitator to facilitate bubble break up

• Formation of bubbles from ring sparger •During emptying of a fermenter, it is important that the air feed valve is closed. This will minimize contamination of the inlet air line.

Effect of agitation speed on bubbles distribution in liquid

•Slow impeller speed

• Fast impeller speed

•Slow impeller speed – bubbles will not be broken, rise directly to to the surface, accumulate and coalesce under impeller leading to from large bubbles and low oxygen transfer rates.

•Fast speed – smaller bubbles will be generated, move throughout the reactor increasing the gas hold up and bubble residence time.

 AIR FLOW RATES

•Volume per volume per minute, vvm. •The airflow rate and liquid volume must have the same basal unit. The air flow rate must be expressed in terms of volume per min.

CONDENSOR TEMPERATURE • A cold condenser temperature can help to control the foam. • The density of the foam increases when it moves from the warm headspace volume to the cold condenser region. • This causes the foam to collapse