Biotechnology Flashcards
What is biotechnology?
The industrial use of living organisms (or parts of living organisms) to produce food, drugs or other products
What are the main advantages of using microorganisms in biological processes?
- No ethical issues when growing
- Large variety of microorganisms
- Genetically modified to produce protein required
- Short life cycle & rapid growth rate - huge quantities can be produced in short periods of time
- Cheap
What microorganism is involved in baking?
- Yeast: mixed with sugar and water to respire aerobically
- CO2 produced makes bread rise
- Yeast cells are killed during cooking
- Takes a couple hours
What microorganism is involved in brewing?
- Yeast: respires anaerobically to produce ethanol
- Mixed with malted barley and hot water
- Fermentation continues for days in anaerobic conditions
What conditions are needed to make bread rise?
- Oxygen
- Heat
- Carbohydrates (flour)
What microorganisms are involved in cheese making?
- Bacteria: feed on lactose in milk, changing texture and taste, and inhibiting the growth of bacteria which makes the milk go off
What microorganisms are involved in yoghurt-making?
- Bacteria: to form ethanol and lactic acid
- Both produce extracellular polymers that give yoghurt its smooth, thick texture
Why is milk pasteurised before being used commercially to make cheese and yoghurt?
- To kill of most natural bacteria that would make it go bad rapidly
- This is done by heating it for a certain amount of time.
Why is milk homogenised before being used commercially to make cheese and yoghurt?
- To allow the fat droplets to evenly distribute through the milk
- This will make the final product smoother and with less fat content
Give 2 differences between the production processes of cheese and yoghurt
- The milk is heated in cheese making and cooled in yoghurt making
- Whole milk is used in cheese making and skimmed milk is used in yoghurt making
Give 4 advantages of using microorganisms to produce human food?
1- Microorganisms reproduce fast and produce protein faster than animals and plants
2- They have high protein content with little fat
3- No welfare issues when growing microorganisms
4- Can be made to taste like anything
Give 4 disadvantages of using microorganisms to produce human food
1- Need sterile conditions that are carefully controlled which adds to cost
2- Protein has to be purified to ensure it contains no toxin or contaminants
3- Many dislike the thought of eating microorganisms grown on waste
4- Has little natural flavour - needs additives
Producing penicillin
- Needs relatively high oxygen levels and a rich nutrient medium to grow well
- It is sensitive to pH and temperature
- A semi-continuous batch process is used
- First stage = fungus grows
- Second stage = produces penicillin
- Drug is extracted from medium and purified
Key points about producing penicillin
- Small fermenters (40-200dm3) as it’s difficult to maintain high levels of oxygenation in large ones
- mixture is continuously stirred to keep it oxygenated
- Rich nutrient medium
- Growth medium contains a buffer to maintain pH at around 6.5
- Bioreactors are maintained at about 25-27°C
What is bioremediation?
Where microorganisms are used to break down pollutants and contaminants in the soil or water
What are the different approaches to bioremediation?
- Using natural organisms: many microorganisms naturally break down organic materials producing CO2 and water
- GM organisms: scientists are trying to develop GM bacteria which can break down or accumulate contaminants which they would not naturally encounter
What are the requirements for optimal growth when culturing microorganisms in the lab?
- Optimum temperature
- Optimum pH
- Oxygen (unless anaerobic)
- Food (nutrient medium)
- Aseptic technique conditions
How do you grow a broth culture?
- Mix sterile nutrient medium with water to make broth
- Mix known mass of dried bacterial culture with water to make suspension
- Inoculate the sterile broth with a known volume of the bacterial suspension
- Stopper the flask, with cotton wool or a bing with air inlet, to prevent contamination from the air
- Incubate at suitable temperature, shaking regularly to aerate
How do you inoculate an agar plate?
- The inoculating loop must be sterilised by holding it in a Bunsen flame until it glows red hot. It must not touch any surfaces as it cools to avoid contamination
- Dip the sterilised loop in the bacterial suspension. Remove lid of Petri dish and spread liquid across surface
- Replace the lid of the Petri dish. It should be held down with tape but not sealed comply so O2 can get in, or even the growth of anaerobic bacteria
- Incubate at a suitable temperature
What type of plates are there?
- Spread (investigating antibiotics)
- Pour (serial dilutions)
- Streak (isolating a small number of bacteria)
Aseptic techniques?
- Heat loop to kill any microorganisms
- Don’t put the lid down on the bench to avoid getting microorganisms on lid
- The screw capped bottles are held at an angle so less SA is exposed to air = less contamination
- Lid of Petri dish opened as little as possible = less air contamination
- Mouth of second bottle is flamed = kill bacteria on neck
- Used wire loop must be heated again = kill remaining bacteria
What is the lag phase?
When bacteria are adapting to their new environment. They are growing, synthesising the enzymes they need and are not yet reproducing at max rate
- Rates are EQUAL
What is the exponential phase?
Birth rate is GREATER than death rate
What is the stationary phase?
Total growth rate is equal
Death rates are EQUAL to birth rates
What is the death stage?
Death rate is GREATER than birth rate
What are the limiting factors that prevent exponential growth in a culture?
- Nutrients available: initially there’s plenty of food, but as numbers increase it’s used up and won’t support further growth
- O2 levels: as population rises, so does the demand for O2
- Temperature: enzyme-controlled reactions within microorganisms are affected by temperature of culture medium
- Build-up of waste: as bacterial numbers rise, the build up of toxic material may inhibit further growth and can even poison/ kill the culture
- Change in pH: as CO2 produced through respiration increases, the pH of the culture falls until a point where the low pH affects enzyme activity and inhibits population growth
What are serial dilutions used for?
To dilute a bacterial culture, so that when you spread a sample onto an agar plate containing nutrients, only 30 to 300 bacteria will grow and produce colonies
What are the two main ways of growing microorganisms?
- Batch fermentation
- Continuous fermentation
Outline the steps in batch fermentation
- Microorganisms are inoculated into a fixed volume of medium
- As growth takes place, nutrients are used up and both new biomass and waste products build up
- As the culture reaches the stationary phase, overall growth ceases but during this phase the microorganisms often carry out biochemical changes to form the desired end products
- The process is stopped before the death phase and the products are harvested. The whole system is then cleaned and sterilised and a new batch culture started up
Outline the steps in continuous fermentation
- Microorganisms are inoculated into sterile nutrient medium and they start to grow
- Sterile nutrient medium is added continually to the culture once it reaches the exponential point of growth
- Culture broth is continually removed - the medium, waste products, microorganisms and products - keeping the culture volume constant
Why and how are bioreactors controlled?
- Temperature: if too low they won’t grow quickly enough, if too high the enzymes start to denature and microorganisms are inhibited/destroyed
- Nutrients and oxygen: O2/nutrient medium can be added in controlled amounts to the broth when probes or sample tests indicate that levels are low
- Mixing things up: inside a bioreactor there are large volumes of liquid, which may be quite thick/viscous due to the growth of microorganisms. Simple diffusion is not enough to ensure all microorganisms receive enough food/oxygen. Most have a mixing mechanism and many are stirred continuously
- Asepsis: if a bioprocess is contaminated by microorganisms from the air or workers, it can affect the yield. To solve this problem, most are sealed, aseptic units
What are the advantages of using isolated enzymes?
- Less wasteful: whole microorganisms use up substrate when growing, producing biomass rather than product
- More efficient: work at higher concentrations
- More specific: no unwanted enzymes present
- Less downstream processing: cheaper
Why are most of the isolated enzymes extracellular?
- They are secreted so are easier to isolate and use
- Each microorganism produces few extracellular enzymes, making it easy to identify and isolate the required enzymes
- More robust than intracellular. Conditions outside a cell are less tightly controlled so can cope with greater variations of temp and pH
What are immobilised enzymes?
Enzymes attached to an insoluble material
What are the advantages of using immobilised enzymes?
- Can be reused (cheaper)
- Easily separated from reactants and products = reduced downstream processing (cheaper)
- More reliable as there’s a high degree of control
- Greater temp tolerance, less easily denatured
- Ease of manipulation, catalytic properties altered to fit a particular process
What are the disadvantages of using immobilised enzymes?
- Reduced efficiency. The process of immobilising may reduce activity rate
- Higher initial cost of materials. It’s more expensive than free enzymes or microorganisms
- Higher initial costs of bioreactor. System is different from traditional fermenters
- More technical issues. Reactors are more complex so more things can go wrong
How are enzymes immobilised?
- Absorption: surface immobilisation
- Covalent bonding: surface immobilisation
- Entrapment: in matrix
- Encapsulation: membrane entrapment in micro capsules behind a semi permeable membrane
What are the advantages and disadvantages of absorption surface immobilisation?
+ Simple and cheap to do
+ Can be used with many different processes
+ Enzymes very accessible to substrate and their activity is virtually unchanged
- Enzymes can be lost from matrix relatively easily
What are the advantages and disadvantages of covalent bonding immobilisation?
+ Cost varies
+ Enzymes strongly bound so unlikely to be lost
+ Enzymes very accessible to substrate
+ pH and substrate concentration often have little effect of enzyme activity
- Cost varies
- Active site of the enzyme may be modified in process, making it less effective
What are the advantages and disadvantages of entrapment?
+ Widely applicable to different processes
- May be expensive
- Can be difficult to entrap
- Diffusion of the substrate to and and product from the active site can be slow and hold up the reaction
- Effect of entrapment on enzyme activity very variable, depending on matrix
What are the advantages and disadvantages of encapsulation?
+ Relatively simple to do
+ Relatively small effect on enzyme activity
+ Widely applicable to different processes
- Relatively expensive
- Diffusion of the substrate to and product from the active site can be slow and hold up the reaction
What are secondary metabolites?
- Substances that are only produced by an organism in the stationary phase
- E.g. antibiotics (penicillin)
What are primary metabolites?
- Substances produced as an essential part of the normal functioning of the organism
- E.g. alcohol produced by yeast as a product of anaerobic respiration
What is the hormone auxin used for?
Stimulate growth of root/root hairs
What is the hormone cytokinins used for?
Stimulate me growth of shoot/stem
