bioprocess control - universitas...
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Successful of bioprocessing depends on controlling key processvariables:
1. Nutrient metabolite concentrations,
2. Growth factor compositions, and
3. Physiological parameters (e.g., temperature, pH, and oxygen)
Nur Istianah-THP-FTP-UB-2014
Bioprocess Parameter
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They affect cell growth, viability, and differentiation.
A fed-batch strategy is often considered most suitable for
tuning and optimizing cell metabolism. It is more efficient,
which reduces metabolite accumulation in culture
supernatant.
Nutrient metabolite concentrations
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This play a crucial role in regulation of stem cell behavior.
So perfusion mode has been preferentially adopted for
most stem cell bioprocesses because it ensures continuous
renewal of nutrients and other factors as well as
continuous removal of metabolic by-products.
Growth factor compositions
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1. MSC differentiation is enhanced at lower temperatures (32 °C)than in 37 °C conditions (70), whereas high temperatures (39°C) enhanced megakaryopoiesis in CD34-enriched cord bloodcells (71).
2. High pH (7.60) enhanced differentiation and maturation ofmegakaryocyte progenitors (72), whereas lower values (7.1)increased their expansion capacity (73).
3. Oxygen is critical to hESC culture (18), and emerging evidencesuggests that reducing its concentration to low levels (74, 75)can be beneficial for in vitro maintenance of pluripotenthESCs, supporting their self-renewal and reducing spontaneousdifferentiation while maintaining karyotypic integrity (76, 77)compared with normoxia conditions (20%). So a robust strategyhas been developed for mass production of undifferentiatedhESCs using pO2 –controlled bioreactors (61).
Physiological parameters (case study)
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Optimization
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For optimized process yields, control system performance iscritical to managing and documentingperfusion, recirculation, and feeding of bioreactor cultures foran optimal growth environment and maximized cell viability.This is particularly important for the dynamic environmentscreated by differentiating stem cells.
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Monitoring and controling
Process
Measure all of variable
Decide which variable relevant
Compare the measured value with the calculated(desired) one
Decide the action of result
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Measurement of variable
2 Using sensor 2 Manual analysis
1. Temperature
2. pH
3. Pressure (PO2, PCO2)
4. CO2, CCO2 (dissolved)
5. Biomass
6. Optical density
7. Redox potential
8. Thermodynamics
1. Cell mass concentration
2. Cell number concentration
3. Substrate
4. Product
5. Intermediet
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Fig. 1. Common measurement and control of bioreactors as generally accepted as routine equipment
Bernhard Sonnleitner, 1999
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Measurement of variable
Taking sample Insitu
measurement
The methods of direct growthmeasurement are cell opticaldensity, total cellcounters, coulter counter, celldry weight, packed cell volumeand optical detectors.
The indirect measurements ofcell growth are based on cellularcomponents, measurements ofATP, bioluminescence, substrateconsumption and productformation, oxygen uptakerate, respiration quotient andheat evolution.
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Basic control fasilities
Temperaturecontrol
pHcontrol
Dissolvedoxygencontrol
Foamingcontrol
Level control
The vessel is jacketed for cooling and heating
Use base, acid or buffer
Rate of oxgygen supply
Antifoam,agitation
Rate of substrate (valve, flow meter)
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Temperaturecontrol Heat is generated in the fermenter by dissipation
of power, resulting in an agitated system; heat isalso generated by the exothermic biochemicalreactions (related to the rate of cell growth).
Measuring temperature: using thermometers/thermocouples/ thermistors/ platinum resistancethermometers/ miniature integrated circuit devices
Controlling: cooling by jacketed system with theprinciple of heat transfer; heat production is equalto the heat transfer by the jacketed system.
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pHcontrol
The pH has a major effect on cell growth and productformation by influencing the breakdown of substrates andtransport of both substrate and product through the cell wall
Measurement of pH is based on the absolute standard of theelectrochemical properties of the standard hydrogen electrode. Ag/AgClelectrode and KCl electrolyte saturated with AgCl2.
A constant potential ismaintained at the inner surfaceof the glass membrane by fillingthe tube of the electrode witha buffered solution ofaccurately determined andstable composition, and withconstant and accurate hydrogenion activity.
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Dissolvedoxygencontrol
Method of DO measurement:
(1)The tubing method;
an inert gas flows through a coil of permeable silicon
rubber tubing, oxygen diffuses from the broth then
measured by an oxygen gas analyser
(3) electrochemical detectors (the most common)
types: galvanic and polarographic detectors
It use membranes to separate electrochemical cell components
from the broth
(2) Use of mass spectrometer probes;priciple: the ability of the gas to diffuse across the surface membrane.
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Exemple of sensoring scheme (PO2)
O2 changes
O2 react with H2O to release electron (make a current)
Current was presented as “PO2”
by termistor
.P x= PO since
POhigher theis O More
2O2
22
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Foamingcontrol
The problems caused by foam are the loss of broth, clogging ofthe exhaust gas system and possible contamination, a problemthat is due to wetting of the gas filters.
Foam breaker: blades or disks operate on the centre of the shaft and generally mounted on the same agitator shaft.
Chemical anti-foams, based on silicon, prevent any foamingby reducing the interfacial tension of the broth. Use ofchemical anti-foam may complicate the microbialfermentation process, and some may act as an inhibitor.
Ultrasonic waves also can be used to destroy the foam.
Measuring foam use foam detector
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Level control
Rv = resistancce of effluent rate (related to the biomass growth)
Level control is needed for continuous bioreactor.This aimed to maintain its continuity and alsoprevent overflow that causes broth losses
When liquid reach the level detector (sensor), conductancechanges and controller sent information to the inlet andeffluent valve to adjust liquid level by consider this equation:
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