Title: The growth curveHomework: complete learning package 12
3 May 2023
Learning Outcomes
• Describe, with the aid of diagrams, and explain the standard growth curve of a microorganism in a closed culture.
Standard Growth curve
Growth curve in a closed culture
• Lag phase– Bacteria adjusting to new
conditions– Takes a while for enzyme
production• Log phase
– Number of bacteria increase rapidly
• Stationary Phase– Rate of growth is equal
to rate of death• Decline Phase
– Death rate is greater than “birth rate”
• The first three stages represent a sigmoid growth curve
Primary and secondary metabolites
• Primary metabolites– Substances produced by on
organism as part of its growth– Amino acids, proteins,
enzymes, nucleic acids, ethanol and lactate
– Production matches growth of population of organism
• Secondary metabolites– Substances produced not as
part of normal growth– Antibiotic chemicals are
mostly secondary metabolites– Begins after main growth
period and does match population growth.
Learning Outcomes
• Explain the importance of manipulating the growing conditions in a fermentation vessel in order to maximise the yield of product required.
Large-Scale production
• Microorganisms are cultured in large containers called fermenters
• The growing conditions within the fermenter are manipulated and controlled– Precise growing conditions• Temperature• Type and time of the addition of the nutrient• Oxygen concentration• pH
A batch fermenter
Large scale production
• Three examples are– The production of penicillin– The production of protease enzymes– The production of mycoprotein
Learning Outcomes
• Compare and contrast the processes of continuous culture and batch culture.
• Describe the differences between primary and secondary metabolites.
Metabolism and metabolites
• Metabolism (process)– Sum total of all the chemical reactions– Processes produce• New cell and cell components• Chemicals• Waste products
• Metabolites (products)– A substance produced during cell processes
Primary and secondary metabolites
• Primary metabolite– Substance produced by
organism as part of it’s normal growth
– E.g. amino acids, proteins, enzymes
– Production of primary metabolites matches the growth in population
• Secondary metabolite– A substance only produced
at a particular growth phase– No direct involvement in
fundamental metabolite processes
– Production usually begins after the main growth phase of the micro organisms
Batch culture
• Starter population is mixed with a specific quantity of nutrient solution
• Allowed to grow for a fixed period• Products removed• Fermentation tank emptied• Examples
• Penicillin production• Enzyme production
Continuous Culture
• Nutrients are added and products are removed from the fermentation tank at regular intervals
• Examples• Insulin production from genetically modified E.Coli• Production of mycoprotein
Learning outcomes
• Explain the importance of asepsis in the manipulation of microorganisms
Asepsis
• Asepsis – absence of unwanted microorganisms
• Aseptic techniques– Any measure taken during a biotechnological
process to prevent contamination by unwanted microorganisms
The importance of asepsis
• Unwanted microorganisms– Compete with the culture microorganisms– Reduce the yield of useful products– Cause spoilage of the product– Produce toxic chemicals– Destroy the culture microorganism or its products.
Methods to maintain asepsis• Ensure all fermenters and attachments are sterile
– Cleaning with pasteurised steam– Chemical sterilisation
• Sterilise all liquids, solids and gases that enter the reaction vessel
• Maintain a pressure difference between the air in the room where fermentation is taking place and outside– Maintains a steady airflow out of the room
• Ensure culture of microorganisms is pure• Ensure the workers do not introduce unwanted
microorganisms from their skin.
Learning Outcomes
• Describe how enzymes can be immobilised.• Explain why immobilised enzymes are used in
large-scale production.
Immobilising enzymes
• Enzymes act as catalysts in metabolic reactions• Enzymes are useful in industrial processes– Specificity– Temperature of enzyme action
• Enzymes in solution need to be separated from the products.
• Immobilised enzymes can be re-used many times and leaves the product enzyme free.
Methods for immobilising enzymes
• Gel entrapment– Example – immobilising lactase in alginate– Stages• Enzyme solution is mixed with sodium alginate solution• Droplets of this solution are added to a solution of
calcium chloride• The droplet turns into a bead which contains the
enzyme
Immobilising lactase in alginate
Immobilising lactase in alginate
• The beads can be tightly packed into a column
• The liquid substrate can be trickled over the beads
• The product trickles out of the bottom of the column
• The product is collected and purified.
Methods of immobilising enzymes
• Adsorption / carrier bound– Enzyme molecules are
mixed with immobilising support e.g. glass beads or clay
• Covalent Bonding / cross-linked– Enzyme molecules
covalently bonded to a support
Methods of immobilising enzymes
• Entrapment / inclusion– Enzymes trapped in their natural
state in a gel bead– Reaction rate can be reduced as
substrate needs to get through the trapping barrier
• Membrane separation– Substrate separated from the
mixture by a partially permeable membrane.
Advantages of immobilised enzymes
• The advantages of using immobilised enzymes over enzymes in solution are– Immobilised enzymes can be reused– Product is enzyme free– Immobilised enzymes are more tolerant to pH and
temperature changes