C: Questions Flashcards
What is a chemostat?
An open cultivation system that permits a continual state of exponential growth. Fresh media is continually delivered to the vessel, and spent medium, cells, and product removed at the same rate. Growth yield is controlled by substrate concentration, and growth rate by the dilution rate.
What is meant by a continuous culture system?
Bioreactor system that is ‘open’: there is continual addition of fresh medium from a reservoir, and simultaneous removal of cells, product and expended medium. Steady state culture where dx/dt = 0 at equilibrium.
Two desirable properties of a microbe in an industrial fermentation?
Genetic ability, ease of genetic modification, microbial metabolism is controllable (inclusion bodies), purificcation aspects, rapid growth, able to grow on cheap culture media, inducible production, post-translational modifications, low levels of extracellular proteins, amenable to simple process monitoring, thermotolerance, safety.
What is meant by a batch culture system?
A ‘closed’ microbial cultivation system where nothing is added or removed after inoculation.
Difference in operation of a chemostat and a perfusion culture system. Which type of culture system would be suitable for the cultivation of mammalian cells?
A chemostat is an open cultivation system that permits a continual state of near-exponential growth. Fresh media is continually delivered to the vessel, and spent medium, cells and product are removed at the same rate. Growth yield is controlled by substrate concentration and growth rate by the dilution rate. Key difference is that in perfusion culture, there is a cell-retention mechanism to prevent cell wash-out. Perfusion is a variant of continuous culture used with mammalian cells.
Perfusion culture is often used for industrial scale in vitro mammalian cell production systems. Explain the main principle of perfusion culture
This system is a variant of continuous culture: fresh medium is continually pumped into the vessel with removal of equal volume of spent medium and product (this may either be completely removed and replenished or recirculated). Cells are retained within the bioreactor, but in some cases, a limited ‘bleed’ of cells may also occur to make room for growing cells.
Provide one advantage of perfusion culture?
high cell density and product; independent control of growth rate and nutrients through bleed-perfusion ratio; most nutrients go to product and only minimally to biomass formation (steady state); less down-time; in-line cell separation
Briefly explain the operation of fed-batch culture and provide one example of its usefulness in maximising product yield?
essential nutrient or precursor is fed to the culture on a specific rate basis (gradually) so that the period of production of the target molecule can be extended.
If substrate in inhibitory or adversely affects mass transfer rates (catabolite repression, overflow metabolism, amylase and starch); by extending length of stationary phase - support penicillin biosynthesis while minimizing by-products (antibiotic); feeding of precursor (penicillin); induction of metabolite production (rDNA approaches).
Example of perfusion culture
Recombinant Factor VIII (Kogenate) used in hemophilia treatment (159 Kg needed per annum) is manufactured using perfusion technology with suspension-cultivated Baby Hamster Kidney cells. It is harvested continuously over 6 month production runs.
Provide one disadvantage of perfusion culture?
(i) Even more complex (cell retention); (ii) Cell death and debris; (iii) Fouling of probes.
Advantages of continuous systems
- Well suited to growth-related products: a continuous flow process operated at a high dilution rate is the most economic production system for a biomass related product;
- Uses smaller bioreactors (lower capital cost) than batch to produce the same amount of product;
- Most downstream processing operations are most productive when operated in a continuous manner (smaller equipment needed). Using a continuous culture allows the fermentation to be in-tune with other operations in the plant. Thus, overall plant productivity is easier to optimize;
- It is not necessary to shut the continuous fermentor down as frequently as a batch fermenter (less down-time). Theoretically, a continuous fermentor could operate indefinitely without having to be shut down. In practice, however this is not possible;
- Low productivity lag times avoided;
Disadvantages of continuous culture
- Relatively complex operation;
- Contamination risk;
- Not all growth-related products are produced well in a continuous flow system. For full flavour development, some fermented foods and beverages require cellular products released from different phases of batch culture growth. Beer and soya sauce for example cannot be successfully produced by continuous culture;
- Potential loss of complete production run;
5. Maintenance of sterile conditions for long time periods can be challenging;
Industrial processes of continuous culture
SCP (Quorn), ethanol production, gluconic acid, wastewater treatment (activated sludge, trickle beds and anaerobic digesters).
advantage of fed-batch
(i) High cell densities and product; (ii) Less down-time compared to batch; (iii) good biomass growth; (iv) Less overflow metabolism due to slow feeding; (v) Because cells are not removed during the fermentation, fed-batch fermentors are well suited for the production of compounds produced during very slow or zero growth.
disadvantage of fed-batch
(i) More complex; (ii) No removal of waste products; (iii) Reactor working volume reduced due to need to accommodate nutrient feeding.
