CELL GROWTH

SUBSTRATE CONSUMPTION
PRODUCT FORMATION
CELL GROWTH: STATEMENT
Microbial growth is Autocatalytic Reaction
➢ The rate of Growth  Cell Concentration
CELL GROWTH: QUANTIFICATION
▪ Can Quantify Either as Cell Mass or Cell
Number
▪ Purpose:
❖ For Determination of Kinetics and Stoichiometry of microbial
growth
▪ Two Categories
❖ Direct – May be less precise if suspended
solids/interfering compounds present in the medium
❖ Indirect
QUANTIFICATION BY CELL MASS (DIRECT)
▪ Dry Cell Weight (DCW)
❖ Sample broth is centrifuged, washed, and dried
at 80oC and weighed until a constant reading
is achieved
▪ Wet Cell Weight (WCW)
❖ Off-line measurement, performed in process

▪ Optical Density (OD)

❖ Turbidity - based on absorption of light by
suspended cells in culture media
QUANTIFICATION BY CELL MASS (INDIRECT)
▪ When direct method is inapplicable
▪ E.g., culturing mold on solid state media
▪ Indirect method is therefore applied, based on measurement of
substrate consumed and/or product formed
▪ Cell mass can be determined by measurement of protein release, DNA
or ATP,
❖ E.g. assuming, 1 mg ATP/g dry cell weight (DCW)
❖ If 100 mg ATP/L measured,  cell = 100 g DCW/L
QUANTIFICATION BY CELL NUMBER
▪ Hemocytometer
❖ Direct microscopic count
❖ Count all cells (viable & non-viable)
❖ Immediate results

𝑻𝒐𝒕𝒂𝒍 𝑵𝒐 𝒐𝒇 𝑪𝒆𝒍𝒍𝒔
N (cells/mL) = 𝐱 10,000 x dilution Factor
𝑵𝒐 𝒐𝒇 𝑺𝒒𝒖𝒂𝒓𝒆 𝑩𝒐𝒙
QUANTIFICATION BY CELL NUMBER
▪ Agar Plates
❖ Count only live cells (viable)
❖ Delayed results
❖ Each viable cell = 1 colony
❖ Results in Colony Forming Unit (CFU)
QUANTIFICATION BY CELL NUMBER
▪ Coulter particle counter
❖ Count all cells present
❖ For discrete cells in particulate-free medium
❖ Can distinguish between cells of different sizes
CELL GROWTH: PROFILE

Cell density (Y-axis) can be X (biomass cell density)

N (cell number)
CELL GROWTH: MALTHUSIAN MODEL
General rate of cell growth, rx = dX 1
dt
Or as Malthus (1798) put it, rx = X 2

Where  termed as specific growth rate, unit = h-1

Thus 1 1 = 22, & rearrange, we get,  = 1 dX 3

X dt
ANOTHER WAY TO EXPLAIN:
▪ As highlighted previously, cell growth is autocatalytic reaction.
▪ During balanced growth, the growth mimics a first order chemical reaction –
the product of reaction will be a reactant as well, therefore influence the rate.
▪ Rate of cell growth  cell concentration
▪ dX/dt  X
▪ dX/dt = kX
*Note that μ (specific growth rate) is actually
the constant k.

Therefore, why =
1 2
REMEMBER
THIS?
3

dt

5
KINETIC PARAMETER: MAX SPECIFIC GROWTH RATE
In batch culture, the rate of change of biomass concentration is the same as
the rate of generation of biomass (mass balance). This is equivalent to:

ln X t = ln X 0 +   t 5

ln X = max t 6
X0
Or  max t
X = X0  e What is the use of this
expression?

Where µmax = Maximum specific growth rate of cells

X0 = Initial cell concentration at t = 0
KINETIC PARAMETER: MAX SPECIFIC GROWTH RATE
lag
Based on the linear equation 3 ln X t = ln X 0 +   t
phase Stationary
phase
▪ A plot of the natural logarithm
of biomass concentration against
time should yield a straight line,
ln biomass concentration

Log or
exponential the slope of which should equal
phase
to μ. (based on eq. 4)
▪ During the exponential phase, the
organism is rapidly multiplying
& growing at its Maximum
Specific Growth Rate, μmax.
Time
KINETIC PARAMETER: DOUBLING TIME
▪ Another important parameter in  Xt 
the exponential growth phase is ln  = t
the Doubling Time, td (time X0 
required to double the microbial If X t = 2X 0 and t = td ,
mass)
Therefore
▪ By considering X = 2Xo when t is ln 2 = t d
at td, the relationship between
0.693
specific growth rate and the td =
doubling time (td) of the biomass: 
MALTHUS MODEL LIMITATION
▪ Malthus model assumes that µmax is constant and not
dependent on substrate concentration
▪ It is based on the empirical observation during maximum
growth phase.
▪ Valid only for the exponential growth phase.
▪ Model application is limited as it provides no actual limit to
the cell growth.
▪ The obvious limit would be the exhaustion of substrate, which
is not built into the model.
CELL GROWTH: LOGISTIC MODEL
To provide mechanism limiting the growth, a
modification was made by Verhulst to Malthus
equation: *Note that Xm is included – maximum
concentration of cell. Because, in this model, it
 Xm − X 
 = max   assume that the growth rate reduces when Xm is
 Xm  reached as a result of environmental pressures.

In its linear form of equation, for a batch growth of constant

volume, cell balance subjected to growth rate given by:
 max t
This model has the ability to describe X0 e
Exponential Phase, Deceleration Phase, and
X =
Stationary Phase. 1−
X0
X
(
1− e  max t
)
SUBSTRATE DEPENDENT GROWTH MODEL (MONOD)
▪ During Growth and Decline Phases of batch culture,  is dependent
on the concentration of many nutrients in the medium.
▪ But, in many cases, a single substrate exerts a dominant influence
on ; this component is known Growth-limiting substrate. It
can be:
➢ Carbon or Nitrogen source
➢ Some Oxygen
➢ Oxidant such as Nitrate.

▪ During balanced growth,  is related to the concentration of

growth-limiting substrate by the Monod Equation
SUBSTRATE DEPENDENT GROWTH MODEL (MONOD)
rX max S
MONOD MODEL  = = Ks = Monod Saturation Constant (g/L)
X Ks + S
*Notice the state variable S is now being included!
max
When S → 0, µ → S
KS
It generally means when the S is too small /nearing 0 (but not exactly
equal to 0), only the S in the denominator is cancelled out.

When S → , µ → µmax
 max
When S → Ks, µ →
KS
LIMITATION TO MONOD MODEL
▪ Application Limited to Single Substrate
▪ Existence of Long Lag Phase &
Acceleration Phase of Batch Culture
SOLVING FOR MONOD?
-Graphical determination of constant. Linearising the model.
▪ Lineweaver- Burke Plot
▪ Eadie-Hoftee

*How do you obtain the linearised equations?

 Lineweaver-Burke Eadie-Hoftee

*Note that v = μ (specific growth rate), since the above example

is based on Michaelis-Menten.
KS VALUES FOR SEVERAL
MICROORGANISMS
RATE OF CELL DEATH
▪ During the course of fermentation a significant number of
cells might die or deactivated during the course of
fermentation. The rate of expression to describe this can be
written as:
rd = kdX

Where, kd = coefficient for cell death rate (h-1)