Filter sterilization

Bioprocess Sterilization

1. Filter Sterilization
Air and Liquid media

2. Filter Sterilisation
Filtration is used for the removal of
microbes from solutions that cannot
easily be treated in other fashions.
Typically heat-sensitive compounds
such as antibiotics and vitamins are
filtered before addition to sterile cool
media. In the food industry, filtration
finds utility in beer making to remove
yeast before final bottling.

3. Filter Sterilisation
Filtration physically removes microbes
because it employs membranes whose
precisely defined pores are too small to
allow their passage. It is obviously only useful
for liquids and gases. Filtration does not
effectively remove viruses from solution
because they are typically too small. A filter
system with sufficiently small pores to
remove viruses would have an extremely
slow flow rate due to the viscosity of water.
Because viruses are not removed, filtration
cannot technically be considered a form of
sterilization, although it is common to refer to
the process as “filter sterilization.”

4. Types of Filters
There are three major types of filters.
Depth filters, are the oldest type and
consist of overlapping layers of fibrous
sheets of paper, asbestos or glass
fibers. The random nature of the fibers
laying on one top of another creates
torturous paths through the filter that
trap many particles.

5. Depth Filters
Due to their nature they do not trap all
particles of a given size, but they find
utility as pre-filters before final filtration.
That is, their ability to remove the vast
majority of things means that this
removed material does not plug up
the more efficient filters noted below.
They are also useful for the filter
sterilization of gases. Glass and
asbestos depth filters can tolerate high
heat and can be conveniently
sterilized with steam.

6. Membrane Filters
The membrane filter is the most
common type of filtration system used
in modern microbiology laboratories.
These are made from high tensile
strength polymers of cellulose acetate,
cellulose nitrate, polycarbonate,
polyester, polypropylene or
polysulfone.

7. Membrane Filters
At high magnification the
polymers appear as a thick mesh
of interconnected strands with
rather precisely defined spaces
between the polymer. By adjusting
polymerization conditions, the size
of the spaces or pores can be
controlled to create membranes
with various maximal sizes.

8. Nucleopore membranes
Nucleopore Membranes are created
by exposing a very thin polycarbonate
film (10 µm) to nuclear radiation, which
creates areas of weakness in the
polymer. The membrane is then
treated with an etching solution that
degrades the weak areas creating
pores in the membrane.

9. Nucleopore Membranes
Changing the strength of the etching
solution or the time of exposure of the
membrane controls the size of the
pores created and results in
remarkably precise pore sizes.
Nucleopore membranes are
particularly useful for trapping bacteria
on the surface of a membrane for
subsequent microscopic examination.

10. Sterilization of Air
An aerobic process requires large amount
of air ( of the order .1-1 volume of air per
volume of liquid media). Since the cost
incurred for sterilizing such huge amount
of air would be enormous air filters are
used to reduce costs. These filters are
periodically changed when blocked dirty.

11. Air Sterilization

12. Filters come in many shapes and sizes
from small (13 mm) filters that are used for
the filtration of a few milliliters of liquid to
industrial filters with surface areas of
several square meters that can process
hundreds of liters a minute. The figure
below illustrates several different types of
filters and filtration apparatus.

13. Some Filter Types

14. Diagram of Air Filter ( membrane Type)

15. Critical oxygen concentration
Critical oxygen concentration is the
term used to indicate the value of
specific oxygen absorption rate which
permits the respiration without
hindrance

16. Dissolved Oxygen Concentration
QO
2
C
critical
Effect of dissolved O
2 concentration on the
QO
2
of a microorganism
Specific O
2
uptake increases with increase in
dissolved O
2
levels to a certain point C
crit.
QO2= Oxygen Consumption rated

17. Critical dissolved oxygen levels for a range
of microorganisms
Organism Temperature Critical dissolved
o
C Oxygen concentration
(mmoles dm
-3
)
Azotobacter sp. 30 0.018
E. coli 37 0.008
Saccharomyces sp. 30 0.004
Penicillium chrysogenum24 0.022

18. Critical dissolved oxygen levels
To maximise biomass production you must satisfy the
organisms specific oxygen demand by maintaining the
dissolved O
2
levels above C
crit
Cells become metabolically disturbed if the level drops
below C
crit
High dissolved O
2
levels also promote product
formation

Amino acid biosynthesis by Brevibacterium flavum
Cephalosporium synthesis by Cephalosporium sp.

19. FACTORS AFFECTING OXYGEN DEMAND
·Rate of cell respiration
·Type of respiration (aerobic vs anaerobic)
·Type of substrate (glucose vs methane)
·Type of environment (e.g pH, temp etc.)
·Surface area/ volume ratio
large vs small cells (bacteria v mammalian cells)
hyphae, clumps, flocs, pellets etc.
·Nature of surface area ( shape)

20. Size of sparger
gas bubble
Gas composition, volume &
velocity
Design of Impeller
size, no. of blades
rotational speed
Baffles
width, number
FACTORS INFLUENCING OXYGEN SUPPLY
Foam/antifoam
Temperature
Type of liquid
Height/width ratio
‘’Hold up’’
Process factors