COMMERCIAL APPLICATION OF BIOTECHNOLOGY

Compare and contrast the processes of continuous & batch culture Explain the importance of manipulating the growing conditions in the fermentation vessel in order to maximise the yield of product required Explain the importance of asepsis in the manipulation of micro-organisms

Commercially, fermenters are large tanks used to grow.. micro organisms such as bacteria and fungi. The products of these organisms include
Single cell protein

Antibiotics, e.g. penicillin Alcohol

On an enormous scale

Can be up to 200,000 litres of liquid culture Whole process is highly automated probes monitor conditions; computers react to changes to keep conditions optimum

The diagram shows an industrial fermenter

Sparger Cooling water out Harvest Cooling jacket Impeller

Culture broth

E F

B G C H I

D J

STARTER CULTURE

Pure culture of microorganisms grown in sterile nutrient broth & added at the beginning to act as the starter population

The conditions within fermenters need to be .. 1. Aseptic (Asepsis - Sterile) to avoid competition and production of unwanted byproducts (contaminants)
2.

Optimum to allow for maximum yield

Sterile conditions
Nutrient medium would support growth of many unwanted micro-organisms
This would: a) reduce yield of product

b) result in competition for nutrients and space

c) results in product being contaminated/spoiled d) may produce toxic chemicals e) may destroy the culture organisms and/or their products f) May result in food/medical product being unsafe - thus discarded

ASEPTIC TECHNIQUE
Required to ensure asepsis from initial culture, through scaling up, fermentation and product harvesting For details, see table on p 163

TYPE AND TIMING OF NUTRIENT ADDITION

Nutrients Micro-organisms require a source of energy and minerals to grow and respire. These may need to be very specific. e.g. Glucose as a source of carbon Ammonium ions/amino acids as a source of nitrogen Essential vitamins and minerals Timing for primary metabolite early

For secondary metabolite towards end of main growth period

OXYGEN CONCENTRATION
Most use aerobic conditions so Oxygen needed Sterile air is pumped in at the bottom and it bubbles up (sparger) helps mix culture with nutrients Stirrers help distribute air Lack of oxygen unwanted products of anaerobic respiration & less growth N.B. facultative vs obligate

TEMPERATURE
Micro-organisms are very sensitive to changes in temperature.

Optimum temp.used; why?

Reactions release heat Temperature is monitored and controlled by a water jacket - often used to heat to get reaction started and then cool (remove respiratory heat)

pH

Changes in pH would affect metabolic rate/enzyme action of micro-organisms

Buffer or Acid or alkali added as needed

FOAMING

Add anti-foaming agent

AGITATION/STIRRING

Culture medium is stirred using paddles (impellers) or air bubbles to prevent micro organisms settling and maintaining an even concentration of nutrients & even temperature.

PRODUCT HARVEST
Removed via a tap at the bottom May be:

Purified Packaged Marketed

BATCH VERSUS CONTINUOUS CULTURE

Batch (fed-batch) Culture Growth for fixed period of time Growth with fixed quantity of nutrients (no extra added) Products removed and fermentation tank emptied
Useful for processes involving secondary metabolite production as product made in stationary phase Little glucose added at regular intervals keeps culture alive; how?
e.g. Penicillin from Penicillium e.g. Humulin (human insulin from GM E. coli)

Continuous Culture Growth for long period of time Nutrients added at regular intervals or continuously Products removed at regular intervals or continuously keep volume constant
Useful for processes involving primary metabolite production as pop. growth kept in exponential phase

BATCH VERSUS CONTINUOUS CULTURE (ADVANTAGES AND DISADVANTAGES)

Batch (fed-batch) Culture
Growth level slower nutrient levels decline with time Easy to set up and maintain

Continuous Culture
Higher growth rate, nutrients always at high levels More difficult to set up and maintenance of growing conditions (e.g. temp./pH) can be difficult to achieve Contamination could affect huge volumes of product depends on when it is detected More efficient, fermenter operating continuously

Contamination would only affect one batch Less efficient, fermenter not in operation all of the time; need cleaning between batches