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Encyclopedia of Physical Science and Technology EN002G-67 May 25, 2001 20:8

Bioreactors 267

FIGURE 28 Total recycle of biomass by using a microfiltration device.

Medium design for viable culture needs to consider the os- actions involving cell-free enzymes. Essential aspects of
motic pressure of the broth as an important variable. kinetics for bioreactor design are discussed here.
Osmotic pressure of a solution is a measure of the con-
centration of dissolved molecules and ions present as indi-
vidual particles. All cells are susceptible to osmotic pres- 1. Cell Growth
sure of the surrounding medium, but cells without walls
Once a properly formulated and sterilized medium has
(e.g., animal cells, protoplasts) are especially susceptible
been inoculated with a seed culture or inoculum, the cells
to osmotic stress related damage. When the suspending
grow and multiply. In a batch culture, the cell or biomass
fluid has a higher osmotic pressure than the cells, water is
concentration increases with time as shown in Fig. 29,
drawn out of the cells and it may become so dehydrated
which is typical. A short lag phase of little or no growth is
that metabolism ceases. Incidentally, this is why high salt
followed by a period of exponential growth. The lag phase
and high sugar media often have a preservative action,
is an adaptation period in which the cells become acclima-
e.g. in pickling of vegetables. When the osmotic pressure
tized to a new environment. The length of the relatively
is low compared to that in the cell, the cell takes up water
unproductive lag phase may be shortened by increasing
and may burst. Osmotic shock produced by rapid dilu-
the inoculum size and ensuring that the growth environ-
tion is sometimes used to rupture cells, especially animal
ment (medium, pH, temperature, etc.) in the bioreactor is
cells. Osmotic pressure also influences plant cell suspen-
the same as the one in which the inoculum was grown.
sion culture.
Typically, the inoculum should be in the late exponen-
Media formulations do not generally specify the os-
tial phase of growth. The volume of a microbial inoculum
motic pressure; instead, the number of dissolved parti-
should between 5 and 10 percent of the volume of the
cles is given as osmolarity or osmolality. Osmolality is
medium being inoculated. Larger inocula are needed for
the number of moles of particles per kilogram of solution
slower growing cells such as animal and plant cells. In
whereas osmolarity is the number of moles per liter of so-
cultivation of animal cells, the inoculum size is selected
lution. One mole of particles is an osmole, abbreviated as 5
to provide an initial cell count of (2–4) × 10 cells/ml. Af-
Osm. Animal cell culture media have an osmolality of 280
to 320 mOsm kg −1 to conform to the osmolality of serum ter the lag period, the exponential growth persists usually
−1
(290 mOsm kg ). Osmolality is not easily calculated es- until an essential nutrient (the growth limiting nutrient
or substrate) runs low in the medium, or until some in-
pecially when a medium has many components and the
hibitory product of metabolism accumulates to a growth
degree of dissociation is not known. In such cases, osmo-
inhibitory concentration. The cells then enter a phase of
lality is estimated from measurements of freezing point
zero net growth, or the stationary phase (Fig. 29). This is
depression and other colligative properties (i.e., those de-
followed by a decline phase or death phase in which there
pendent on the concentration of dissolved particles). In
is a net loss of biomass as cells die and lyse. Usually an
cell culture media, sodium chloride is used for adjusting
attempt is made to achieve a maximum growth rate and
osmolality to the requisite value.
maintain it for as long as possible.
The biomass growth rate in a bioreactor, i.e., the change
F. Kinetics, Productivity, and Bioreactors
in biomass concentration with time, or dX/dt, depends
Design and performance analysis of a bioreactor are in- on the viable biomass concentration X present at any
separably linked with the kinetics of the bioreaction for time. In other words, growth is self catalyzing, or autocat-
which the reactor is intended. Some common bioreactions alytic. In exponential growth, the growth rate is expressed
include growth of microorganisms and other cells, and re- as follows: