Biotechnology

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Biotechnology basics

 Biotechnology- the industrial use of living organisms to produce food, drugs or other products

Food: Cheese and yoghurt- Lactoballicus, Mycoprotein- Fusarium

Drugs: Penicillin- PenicilliumInsulin-E.coli

Why do we use micro-organisms?

  • grow rapidly in favourable conditions- double every 30mins 
  • produce proteins or chemicals that can be harvested 
  • genetically engineered to produce specific products 
  • can grow at low temps, most chem. processes at 400degrees, so 40 is low 
  • Not dependent on climate-can grow anywhere in the world 
  • Can be grown using waste nutrient products, e.g. waste paper 
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The growth curve

Culture- growth of microorganisms, could be a single species (pure) or a mixture of species (mixed). Can be cultured in a liquid e.g. agar.

Standard growth culture models the pop. size of microorganisms in a closed culture. "Closed"=conditions are fixed, where no new materials are added or waste products removed.

PHASE 1: LAG PHASE=

  • microorganisms adjusting to conditions
  • releasing enzymes for extracellular digestion-break down surrounding nutrient agar,
  • cells active but not reproudcing yet-pop=constant.

PHASE 2: Exponential/log phase=

  • Cells have broken down and absorbed nutrients
  • Lots of nutrients available 
  • Favourable conditions,  little intraspecific competition=pop size doubles each generation-every 20-30mins
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The growth curve

PHASE 3- STATIONARY PHASE=

  • Nutrients levels decrease
  • CO2 builds up
  • Some individuals die-death rate=growth rate
  • in an open system, this would be called carrying capacity

PHASE 4- DEATH PHASE=

  • Nutrients very low
  • Waste products=HIGH
  • death rate increases
  • death rate > growth rate, pop. size decreases
  • in closed system, all will eventually die
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Phases of Microbial Growth

(http://web.deu.edu.tr/atiksu/toprak/curve.gif)

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Metabolites

Metabolite= product of metabolism (all the chemical reactions occuring in an organism)

These include: New cells, waste products, hormones/enzymes

Primary metabolites- 

  • produced by an organism as part of its normal growth=(amino acids, enzymes, nucleic acids)
  • produced in good conditions 
  • ALL MICROORGANISMS PRODUCE THESE

Secondary metabolies-

  • Not produced as part of normal growth
  • begins after the main growth period, therefore doesnt match the pop.
  • produced in less favourable conditions
  • produces mols not essential for growth, but still useful, e.g. penicillium produces penicillin to help it compete w bacteria 
  • NOT ALL MICROORGANISMS CAN PRODUCE SECONDARY METABOLITES
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Fermenters

Fermenters= huge tanks where internal conditions can be controlled 

Sensorsdetect current conditions and feed this back to output devices to bring conditions back to norm

Temp- too hot=denaturing of enzymes, too cold=rate of growth too slow 

Nutrient supply-
timing of adding nutrients determined by whether u are producing secondary or primary metabolites
Must be high in carbon, hydrogen, nitrogen and oxygen

O2 concentration= growth=aerobic resp. bc no oxygen leads to anaerobic conditions=waste products+slow growth rate

pH=affects activity of enzymes, alters 3D shape and breaks hydrogen bonds

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Asepsis

ABSENCE OF UNWANTED MICROORGANISMS=ASEPTIC TECHNIQUES

Unwanted microorganisms=contaminants

Contaminants must be kept out so they don't- 

  • Compete w the microorganisms for space+nutrients
  • Reduce the yield of the product
  • Spoil the product
  • Produce toxic chemicals as metabolites
  • Make food/medicine unsafe
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Batch and Continuous Culture

Batch= 

  • starter pop. mixed w a fixed amount of nutrient and allowed to grow for a fixed period w no further nutrient added  
  • Products collected at the end 
  • Growth Rate slower-nutrients run out
  • Not operating all the time
  • Easy to set up and maintain
  • Useful for producing secondary metabolites (allows unfavourable conditions to develop)
  • Penicillin produced this way 

Continuous=

  • Nutrients added and waste products removed regularly 
  • Set up more difficult
  • Maintaining conditions difficult
  • More efficient as operates continuously 
  • good for primary metabolites
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Immobilised Enzymes

Used because: 
Highly specific-catalyse reactions between specific chemicalsFewer by-products and less purification necessary 
Lowers temp- enzymes function well at low temps which are cheaper 

However you have to separate the enzyme from the product, and downstream (where enzymes are extracted from the product) is time-consuming and expensive, therefore immobilse them!

Enzymes not present w the product =low downstream costs, enzymes immediately available for reuse=increases continuity, more stable and less likely to have pH or temp damage 

Takes more time and equipment=high costs, less active as they don't mix freely w substrate, enzyme leakage can still happen.

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Immobilised Enzymes- 4 methods to set up

FIRST METHOD: ADSORPTION

  • enzyme molecules mixed with immobilising support and bind to it through hydrophobic reactions and ionic links. Adsorbing agents=glass beads, clay
  • Not strong bond-enzyme leakage can happen, but active site is exposed 

SECOND METHOD: COVALENT BONDING

  • enzymes covalently bonded to insoluble material, e.g. clay, using cross linking agent like sepharose.
  • Little leakage and exposed active site--but doesnt immobilise the whole enzyme

THIRD METHOD: ENTRAPMENT

  • enzymes trapped within a substance, eg gel bead. Slow rate of reaction as molecules have to get through barrier-active site less accessible

FOURTH METHOD: MEMBRANE SEPARATION

  • enzyme separated from substrate using membrane-enzyme too big to pass through it. 
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