Types of Culture Fermentation - carried out as batch, continuous and fed-batch Dictated largely by type of product being produced
Batch Culture Closed culture system – initially contains limited amount of nutrient No growth – lag phase – time of adaptation Growth rate increases – grow constantly at maximum rate – Log or exponential phase
Batch Culture- Exponential phase/Trophophase Exponential phase equation: dx/dt = µx where x is the concentration of microbial biomass, t is time in hours and µ is the specific growth rate in hours Nutrients are in excess and organism is growing at its maximum specific growth rate = µmax
Batch Culture – Deceleration phase Cessation of growth – due to depletion of some essential nutrient in medium (substrate limitation) Accumulation of some autotoxic product of organism in medium (toxin limitation) Or combination of both
Batch Culture – Stationary phase/Idiophase Point where growth rate has declined to zero. Population is metabolically active – producing secondary metabolites Maximum population phase
What is Yield Factor (Y)? It is a measure of efficiency of conversion of any one substrate into biomass. It is not constant – varies according to growth rate, pH, temperature, the limiting substrate and concentration of substrates in excess.
How can deceleration phase be tested? Zone A to B – increase in initial substrate concentration – proportionally increases biomass produced at stationary phase x=Y(Sr-s) where x is concentration of biomass produced Y is yield factor, Sr is initial substrate con. and s is residual conc.
Growth kinetics of Metabolites Growth-linked products/Primary metabolites Formation of growth linked product is described by equation -dp/dt = qpx where p is the concentration of the product -qp is specific rate of product formation (mg product/g biomass/h) -Product formation is related to biomass production by equation dp/dx = Yp/x -Where Yp/x is the yield of product in terms of biomass (g product/g biomass)
Batch fermentation – used for? Producing biomass – fastest growth rate and maximum cell population Primary metabolites – extend exponential phase Secondary metabolites – decreased growth in log phase
Continuous Culture (Chemostat) Addition of fresh medium to the vessel – exponential growth Medium is substrate limited Overflow device – added medium displaces equal volume of culture – continuous production of cells - Formation of new biomass balanced by loss of cells from vessel
Chemostat culture Chemostat culture – Cells and spent medium are continuously removed - State of culture is dependent upon flow rate of fresh medium
Continuous Culture (Chemostat) Dilution rate - The flow of medium into the vessel is related to volume of the vessel D = F/V - F is flow rate (dm3/h) - V is volume (dm3) -D is dilution rate in per hour -Net change in cell concentration over a time period = dx/dt = growth (µx) – output (Dx)
Problems - Chemostat Problems – Imperfect mixing and wall growth -Imperfect mixing – increase in degree of heterogeneity in fermenter -Wall growth – Organism adheres to inner surfaces of reactor – increases heterogeneity -Limited by coating inner surfaces of vessel with Teflon
Feedback systems - Chemostat Internal feedback – Limiting exit of biomass External feedback – Subjecting effluent stream to biomass separation
Perfusion/Turbidostat Perfusion culture - Medium is pumped continuously -Cells are retained -Becoming popular for large-scale production -Attains high cell density -Cell separator
Continuous culture Biomass production Microbial biomass produced for human or animal consumption – Single Cell Protein (SCP) Practice 1960s
What is Fed-batch culture? In open system/Fed-batch culture – involves controlled nutrient feeding Partial media changes at regular intervals
Fed-batch culture Initial batch cultures – fed continuously or sequentially with medium No removal of culture fluid Three types – Variable volume, Fixed volume and Cyclic fed-batch
Fed-batch culture Batch culture is fed in following ways -Same medium + conc. used to establish batch culture is added –Variable volume - Conc. solution of limiting substrate is added at a rate less than initial -Very conc. Solution of limiting substrate at lesser rate than initial – Fixed volume
Cyclic fed-batch culture Life of variable fed-batch fermentation may be extended -Withdrawing a portion of culture/residual culture -Increase in dilution rate and specific growth rate
Fed-batch culture Used for biomass Primary metabolite Secondary metabolite
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