Cell Growth Measurement

Growth – orderly increased of all chemical components

Balaced Growth – growth during which a doubling of the biomass is accompanied by a doubling of all
measurable properties of the population such as protein, DNA, RNA and intracellular water.

Cell growth can be determined by:

- Cell number
- Cell mass
- Cell activity

MEASUREMENT OF CELL NUMBER

Microscopic Counts - the number of cells in a population can be measured under a microscope by
counting cells placed in special counting chambers.

Two types of Chambers:

1. HEMOCYTOMETER – blood cell counting chamber for use with microorganisms of 3 µm in

diameter or larger
2. PETROFF-HAUSSER COUNTING CHAMBER – for used primarily with bacteria

Direct-Counting Method

ADVANTAGES
1. Minimal equipment is required
2. Results are obtained rapidly
3. Morphological characteristics
organisms can be observed
DISADVANTAGES
1. Dead cells cannot usually be distinguished
from live cells
of the 2. The method is not suitable for cell
suspensions of low density
3. Small cells are difficult to see under the
microscope and can be missed when
counting
4. The actual counting procedure is tiresome
and may cause considerable eyestrain
5. It is not suitable for highly flocculating cells
such as mycelium

Viable Plate Count

Viable cell – one that is able to divide and form a colony

Two ways of performing plate count

1. Spread Plate Method – a volume of no larger than 0.1 ml is spread over the agar surface
2. Pour Plate Method – sample is mixed with melted agar and poured into a sterile plate

Coulter Counter

- To avoid tedium of direct microscopic counting

 - Using this, cell size can also be measured
- Cannot ditinguish between cells and any impure particles
- Difficult to use with microorganisms in chains and is useless with mycelial organisms

MEASUREMENT OF CELL MASS

Cell Dry Weight

- can be measured directly by taking an aliquot of cell suspension and centrifuging it

- most direct approach for the quantitative measurement of cell mass and is probably the
most reliable and reproducible
- can only be used with dense cell suspensions

Turbidity

- based on the fact that small particles scatter light proportionally, within certain limits, to
their concentration
- when a beam of light is passed through a suspension of organisms, the reduction in the
amount of light transmitted as a consequence of scattering is thus a measure of cell density.
Such measurements are usually made in spectrophotometer, which reads in Absorbency (A)
units.

Absorbency – logarithm of the ratio of intensity of light striking the suspension (I o) to that transmitted by
the suspension (I)

Io
A = log
I
Indirect Methods

The indirect methods for measuring cell mass are based on the overallstoichiometry for growth
and product formation, which may be written in the general form:

The change of the cell mass can be monitored indirectly by measuring the following:1

1. Nutrient Consumption
2. Product Formation
3. Cell Components
4. Heat Evolution
5. Viscosity

Effect of Environmental Conditions on Microbial Growth

Temperature - important factor affecting the performance of cells

Groups of Organisms

1. Psychrophiles- (Topt < 20°C)

2. Mesophiles - (Topt = from 20° to 50°C)
3. Thermophiles - (Topt > 50° C)

As the temperature is increased toward optimal growth temperature, the growth rate
approximately doubles for every 10°C increase in temperature. Above the optimal temperature range,
the growth rate decreases and thermal death may occur.

Temperature also affects product formation. However, the temperature optimum for growth and
product formation may be different. The yield coefficient is also affected by temperature. In some cases,
such as single-cell protein production, temperature optimization to maximize the yield coefficient is
critical. When temperature is increased above the optimum temperature, the maintenance
requirements of cells increase resulting in a decrease in the yield coefficient.

Temperature also may affect the rate-limiting step in a fermentation process. At high
temperatures, the rate of bioreaction might become higher than the diffusion rate, and diffusion would
then become the rate-limiting step.

Hydrogen-ion concentration (pH) affects the activity of enzymes and therefore the microbial growth
rate. The optimal pH for growth may be different from that for product formation. Generally, the
acceptable pH range varies about the optimum by ±1 to 2 pH units. Different organisms have different
pH optima: the pH optimum for many bacteria ranges from pH = 3 to 8; for yeast, pH = 3 to 6; for molds,
pH = 3 to 7; for plant cells, pH = 5 to 6; and for animal cells, pH = 6.5 to 7.5. Many organisms have
mechanisms to maintain intracellular pH at a relatively constant level in the presence of fluctuations in
environmental pH. When pH differs from the optimal value, the maintenance-energy requirements
increase. One consequence of different pH optima is that the pH of the medium can be used to select
one organism over another.

Dissolved oxygen (DO)- is an important substrate in aerobic fermentations and may be a limiting
substrate, since oxygen gas is sparingly soluble in water. At high cell concentrations, the rate of oxygen
consumption may exceed the rate of oxygen supply, leading to oxygen limitations. When oxygen is the
rate-limiting factor, specific growth rate varies with dissolved-oxygen concentration according to
saturation kinetics; below a critical concentration, growth or respiration approaches a first-order rate
dependence on the dissolved-oxygen concentration.

Above a critical oxygen concentration, the growth rate becomes independent of the dissolved-
oxygen concentration. Oxygen is a growth-rate-limiting factor when the DO level is below the critical DO
concentration.

Dissolved carbon dioxide (DCO2) concentration may have a profound effect on performance of
organisms. Very high DCO2 concentrations may be toxic to some cells. On the other hand, cells require a
certain DCO2 level for proper metabolic functions. The dissolved carbon dioxide concentration can be
controlled by changing the CO2 content of the air supply and the agitation speed.
 High substrate concentrations that are significantly above stoichiometric requirements are
inhibitory to cellular functions. Inhibitory levels of substrates vary depending on the type of cells and
substrate.