Biotechnology

Biotechnology - technology that involves the exploitation of living organisms or biological processes

why are microorganisms used?

• have rapid life cycles so large populations can be built up quick
• prokaryotes reproduce asexually so populations are genetically identical and carry out the same metabolic processes
• have simple needs for growth - can be grown in fermenters under controlled conditions with little attention
• can be grown using waste materials
• no one really minds what happens to microorganisms so expoliting them doesnt raise many ethical issues
• easy to genetically modify

1. Lag Phase - organisms are adjusting to surroundings - cells are active but not reproducing - population remians fairly constant

2. Log (exponential phase) - cells are growing and dividing at maximum rate for the conditions 

3. Stationary Phase - Nutrient levels decrease and waste products begin to build up - rate to reproduction slows down - cell reproduction and cell death are at the same rate

4. Death Phase - Nutrients become exhausted and toxic waste products lead to death rate exceeding that of the reproduction rate - all cells will eventually die in a closed system

Batch - single fermentation whereby nothing is put on or taken out while fermentation takes place, except waste gases

Advantages

• easy to set up and maintain
• smaller vessels can be used
• contamination means only one batch is lost and not large quantities of product
• good for production of secondary metabolites

Disadvantages

• growth rate is less because nutrient levels decline as fermentation progresses
• fermenter is not in operation all the time so is less efficient

- ongoing fermentation whereby substrates are added and products removed

advantages

• Growth rate is higher as nutrients are continuosuly added to the fermenter
• fermenter operates continuously so more efficient - no 'down time'
• good for processes involving the production of primary metabolites

disadvantages

• if contamination occurs huge amounts of prodcut could be lost
• set up is more difficult and growing conditions must be maintained which can also be difficult

Metabolite - is a substance that is made by a cell in the course of its metabolism

Primary metabolite - substance produced by an organism as part of its normal growth - matches the growth in population of an organism

Secondary metabolite - substances produced only by some cells or at certain growth stages - no fundamental involvement with fundamental metabolic processes - usally produced after main growth stage - penicillin is an example of a secondary metabolite

Penicillin is created using the fed batch culture whereby a carbohydrate source is added every 30 mintues. This increases the length of fermentation and produces more penicillin that standard batch culture

• Temperature - if temperature is too hot then essential enzymes will be denatured, if too cold then growth rate will fall
• Oxygen Concenration - a lack of oxygen can lead to unwanted anaerobic by products 
• pH - the correct pH is needed to maximise the activity of the ezymes and to prevent denaturing

- the abscence of unwantedorganisms 

Reasons

• unwanted organisms may compete with culture organisms for nutrients and space
• may reduce the yeild of useful products
• cause spoilage with could be hazardous to health
• may produce toxic byproducts that destroy culture organisms or product

Aseptic Techniques - measures taken to ensure unwanted microorganisms do not contaminate the culture

• ensure fermenters are steralised before use eg steam cleaning, chemical sterilistation 
• steralising liquids, solids and gases before they enter the reaction vessell
• wearing lab coats and other protective clothing to prevent organisms enterting via the skin
• filters on  inlet and outlet tubes to prevent microorganisms entering
• stainless steel to prevent microbes sticking

- measures taken to seperate enzymes from the reaction mixture but ensuring they still function

advantages - purification costs are lowered because enzyms arnt present in the final product, enzymes are able to be reused, immolised enzymes are more stable than normal enzymes

disadvantages - leakage can occur which may contaminate the product, immoblisation takes time and money, enzymes are less active because they dont mix with the culture

Techniques

Adsorption - enzymes are mixed with an immoblising support and bind to it via hydrophobic interactions and ionic links - enzymes may still leak but if not then adsorption can give high reaction rates

Covalent Bonding - enzymes are bound to a support then covalently bonded to each other - not a large amount of ezymes used but bonds are strong to prevent leakage

Entrapment - enzymes are trapped inside a alginate bead - reaction rates are reduced because substrates must pass through trapping barrier, meaning the active site is less readily available

Membrane Seperation - Enzymes can be physically seperate from mixxture by a partially permeable membrane 

• Occurs in two stages - microorganisms that produces enzyme are grown and then ezyme is extracted and purified
• usually thermophillic organisms are used as they produce enzymes that can withstand high temperatures
• bacteria are provided with carbon source and a nitrogen source - carbon source usually left over waste products - nitrogen can be ureas
• batch culture is used 
• enzyme usually remains in the cell
• when fermentation is finished the culture is heated to kill enzymes - cells are then broken up to remove enzymes which dissolve in culture
• this can be concentrated and filtered to remove enzymes

• Mycoprotein means fungus protein
• Culture medium contains glucose obtain from starch - provides fungus with respiratory substrate
• ammonium phosphate is added as a nitrogen source for fungus to make proteins and nucleic acids
• temperature, pH and oxygen are kept constant to maintain optimum growing conditions
• stirrirs arnt used because this would tangle the fungle hyphae
• continuous culture method is used
• liquid culture containing the fungus is run off the bottom and then centrifuged to seperate hyphae from liquid
• filteration and steam treatment can then be used to complete the process, leaving the mycoprotein