PART 1 - Kinetics of Substrate Utilization, Product Formation and Biomass Production in Cell Cultures Flashcards
Batch Growth, CSTR or Chemostat, Balanced Growth
___________ are apparatus or structures in which a chemical, biological and/or
physical process are facilitated
Reactors
____________ are model systems for which the transport processes
and chemical reactions are exactly defined.
Ideal reactors
For a reactor to be ideal, it must be:
- controlled
- analog and prototype
- mathematically modeled
Types of ideal reactors in the context of enzymatic and culture processes:
- Batch operation of mixed enzyme reactor
- Continuous operation of mixed enzyme reactor
Batch process is an ____________________ operation where the __________,_________ changes with ______.
unsteady state
concentration of cell mass, substrate changes with TIME
Batch processes operate in ________________
closed systems
Explain what happens in batch processes
substrate is added at the beginning of the process and products removed only at the end.
What are classified as batch
Bioreactors with neither input nor output of liquid
or solid material
In Batch, the mass of substrate in the reactor, M, is equal to the ________________ multiplied by the ________________.
substrate concentration s
liquid volume V
Recite the initial equation of Batch Time from Mass Balance for Enzymatic Reaction
d(sV)/dt = -(VmaxS/Km+S)V
Since V is constant,
Take V outside the differential, and cancel from both sides
Enzymes are subject to _______________. Accordingly, the concentration of active enzyme in
the reactor, and therefore the value of __________, may change during reaction.
deactivation, vmax
Microorganisms have specific ___________ and _____ ranges at which they thrive
temperature, pH
Cell growth process got two different manifestations according to the morphology of cell
involved: EXPLAIN UNI ORGANISMS AND MOLDS
For unicellular organisms which divide as they grow, increases in biomass are accompanied by increases in the number of cells present.
Unlike unicellular organisms, molds do not necessarily undergo cell division as the primary means of growth. Instead, they grow by
elongation and branching of their filamentous structures.
Certain parameters/ phenomena to
consider in determining the cell population kinetics
What does it do to formulate a simple kinetic model for cellular activities
- characteristics of culture broth,
- nutrients and substrates used for growth and production of metabolites
- cell to cell heterogeneity
- microbial cells of different ages manifesting metabolic activities
- characterization of biochemical pathway
THEY ARE COMPLEX
They make it difficult to formulate a simple kinetic model
Two processes are associated with cell growth
- utilization of materials from the medium
by the cells - generation of metabolic end products in the medium
Define the RATE-LIMITING SUBSTRATE
First, it is assumed that
the concentration of all components present in the medium is sufficiently high and the rate of
reaction depends on the concentration of that component only.
The process by which cells generate energy, involves various metabolic pathways.
Cellular respiration
These are used to maintain optimal growth conditions
Incubators, Bioreactors, and Temperature-Controlled Environments
What can be indicative of alterations in cell structure,
function, or health
Changes in the
rheological properties of cells, including viscosity,
This refers to the study of the flow and deformation of matter, and it plays a role in understanding the mechanical properties of cells
Rheology
This may contribute to the diversity observed in different phases of cellular activities.
Stochastic events
This can contribute to genetic variability among cells within a population
Genetic drift, as
a random process
They classified the microbial systems according to the number of components used in the
cellular representation.
Arnold Fredrickson and Henry
Tsuchiya
Model classifications for mathematical representation of cell populations: ENUMERATE AND EXPLAIN EACH
This is one perspective in which microbial cells are considered as
multicomponent systems. These models are very complex and not used very often.
Structured model
One common type of structured model in cell growth is the
age-structured model
The structured model usually deals with the kinetics of the change in individual components present in the cells, such as
RNA, DNA, proteins, etc
When cell population is treated as one component system, it is referred to as
Unstructured
model
Model assumes balanced growth where cell components do not change with time. Much less complex and much more commonly used.
Unstructured Model
This consider individual cells, in recognition of the fact that cells in a
population – a pure culture – are different and are most often formulated as a population balance
model.
Segregated Model
These models form an important class and describe the biomass as
consisting of several variables (such as NADH, precursors, metabolites, ATP, biomass).
Unsegregated structured models
The real condition of the living system is
a structured, segregated one
In ideal conditions,
the cell growth kinetics is assumed to be in the
unsegregated, unstructured mode
This is typically prepared by using a seed culture in a liquid medium.
Inoculum
It is the period where the individual bacteria are maturing and not yet able to divide.
Lag phase
The log phase is sometimes called
the logarithmic phase or the exponential phase.
It is the period characterized by cell doubling and the growth rate is independent of nutrient and substrate concentration.
Log phase/ Exponential phase
No net growth of cell numbers or cell mass. Secondary metabolites like alkaloids and glycosides are produced.
Stationary Phase
This phase shows an exponential decrease in the number of living cells in the media while nutrients are depleted.
The rate of cell decline is ___________.
The death phase
first-order
Batch process is ________________ operation where the concentration of cell mass, substrate
changes with time.
an unsteady state
This is a type of chemical reactor that operates continuously, provides thorough mixing.
Continuous Stirred-Tank Reactor (CSTR)
This is used in a variety of industrial processes for chemical reactions where steady-state conditions are desired.
CSTR
CSTR is also called
Chemostat
Purpose of mixing
to maintain homogeneity in the fermentation broth
In the steady- state condition, the concentration of any
component in the vessel is ___________ of time.
independent
Ideal Chemostat Models
–
Monod Chemostat Models
–
Sterile feed means
X0 = 0
Cell mass productivity
Explain the plot of XSS, SSS, DXSS versus the dilution rate D.
At Dwashout, ________ will remain present in the reactor.
no cell
At Dwashout, _______, where washout of cells will take place
X→0
It should be noted that ______ < 𝐷𝑤𝑎𝑠ℎ𝑜𝑢𝑡 ≤ ______
𝐷𝑚𝑎𝑥
𝜇𝑚𝑎𝑥
If a mixed microbial culture contains both slow-growing and fast-growing microorganisms, then by adjusting the dilution rate_______ than the __________ of slow-growing but ______ than that of fast-growing, it is possible to separate fast-growing organisms from the slow-growing organisms.
higher
Dwashout
less
Advantages of Chemostat
- Growth rate can be controlled and maintained indefinitely. So one can operate the log phase
of growth for the maximum cell mass production for the infinite period of time. - Effect of growth-limiting substrate can be easily monitored.
- The chemostat can be used to study the period of unbalanced growth, which occurs during
the transition period between steady states at different growth rates. The formation of some
plant metabolite is found to increase during the transition of phases. - Results obtained are reliable and reproducible.
Disadvantages of Chemostat
- The major problem of chemostat is cell washout. It is difficult to operate at Dmax because
it is very close to Dwashout - Cell growth over long periods can cause mutation or contamination
Bioreactors are operated continuously in a few bioprocess industries such as
- brewing,
- production of bakers’ yeast
- waste treatment
In CSTRs, if the vessel is well mixed, the product stream has ____________ composition as the liquid in the reactor.
the same
Therefore, when continuous reactors are used with ___________________, the catalyst is _________________ from the vessel in the product stream.
freely suspended cells or enzymes
continuously withdrawn
Characteristic operating parameters for continuous reactors are
- dilution rate D defined
in - average residence time τ.
T = V/F
D = F/V
The different phases of growth are more readily distinguished when the ____________________ is plotted against _______; alternatively, a _________ plot can be used.
logarithm of viable cell concentration
time
semi-log
During the ___________ immediately after inoculation of the culture, the rate of growth is essentially _______.
lag phase
zero
This also refers to a period of time when the change of cell number is zero
Lag phase
Cells use this phase to adapt to their new environment.
lag
In this phase, new enzymes or structural components may be synthesized but the concentration of cells _______________.
Lag phase
does not increase
The length of this lag period depends on many factors such as
- the type and age of the microorganisms,
- the size of the inoculum,
- culture conditions
This phase is where growth starts for the inoculated cell culture
Acceleration phase
The cell number starts to increase, and the division rate increases to reach a ____________
Acceleration phase
maximum
During this period, growth achieves its maximum rate.
Growth phase or exponential growth phase
If growth is exponential (which is the most often case), the growth phase appears as a
_______________ on a semi-log plot.
straight line
Growth slows down due to _______________ or build-up of ____________________
Decline phase
nutrient exhaustion
inhibitory products.
During this period, cell growth ceases. No further cell growth can be observed
Stationary phase
The transition between the exponential phase and the stationary phase involves a period
of _____________________ during which the various cellular components are synthesized at
___________________.
unbalanced growth
unequal rates
Consequently, cells in the stationary phase have a chemical composition ___________ from that of cells in the exponential phase.
different
At this phase, cells lose _________ or are destroyed by ______
Death phase
viability
lysis
Death occurs either because of the depletion of the _________________, or the accumulation of ______________.
cellular reserves of energy
toxic products
Cell growth is considered to be a
first-order autocatalytic reaction
time required for the cell population to double
Doubling time
Starting with a cell concentration of 𝑁0, the concentration at 𝑡 = 𝑡𝑑 is
N=2𝑁0.
or
X=2X0
Doubling time is a valid representation of the growth rate only when
μ is constant
This represents an approximation where the average cellular synthesis activities are not affected by the growing cell population as the coordination is relatively perfect.
Balanced growth kinetics
In an environment favorable for growth, cells regulate their metabolism and adjust the rates of various internal reactions so that a condition of _______________ occurs.
balanced growth
Under this type of kinetics, the composition of the biomass remains constant.
Balanced growth
A balanced growth signifies that the cell is able to modulate the effects of _____________________ and keep the biomass composition steady despite changes in environmental conditions.
external perturbations
Balanced Growth
A cell can grow in size or mass in numbers. Hence, unstructured models can be used for the
bio-phase characterization
For the biomass composition to remain constant during growth, the _________________________ in the culture must be equal to the cell ______________________
specific rate of production
of each component or N
specific growth rate, μ
Balanced growth cannot be achieved if __________________affect the ________________. In most cultures, balanced growth occurs at the same time as _________________.
environmental changes
rate of growth
exponential growth.
In ______________ organisms, the progressive doubling of cell number results in a ____________________in the population.
unicellular
continually increasing rate of growth
A bacterial culture undergoing balanced growth mimics a ______________________________
first-order autocatalytic chemical
reaction.
Balanced growth
Hence, the rate of the cell population increase at any particular time is proportional to the
___________________ of bacteria present at that time.
number density (CN)
