Cell distribution

Biochemical Separation
Cell Disruption

Several different adsorption operations are used in bioprocessing, particularly for medical and pharmaceutical products. Ion-
exchange adsorption is established practice for recovery of amino acids, proteins, antibiotics and vitamins. Adsorption onto
activated charcoal is a method of long standing for purification of citric acid; adsorption of organic chemicals onto charcoal or
porous polymeric adsorbents is common in wastewater treatment.
Downstream processing of fermentation broths usually begins with
separation of cells by filtration or centrifugation.
The next step depends on location of the desired product.
The product secreted during the process of fermentation is either
intracellular, extracellular, or periplasmic.
If the product is produced extracellularly, the desired product can
be obtained from the liquid broth followed by further purification
steps.
On the other hand, if the product of interest is produced inside the
cell (either cytoplasm or periplasm), it is indispensable to disrupt or
disturb (in the case of periplasmic expression) the cell in order to
extract the intracellular products.

Cell disruption involves the pervasion or lysis of the cell that
enhances the release of intracellular products.
For substances such as ethanol, citric acid and antibiotics which are
excreted from cells, product is recovered from the cell-free broth
using unit operations.
For products such as enzymes and recombinant proteins which
remain in the biomass, cell disruption must be carried out to release
the desired material. A variety of methods is available to disrupt
cells.

Several different adsorption operations are used in bioprocessing, for medical and pharmaceutical products. Ion-exchange
adsorption is established practice for recovery of amino acids, proteins, antibiotics and vitamins. Adsorption onto activated
charcoal is a method of long standing for purification of citric acid; adsorption of organic chemicals onto charcoal or porous
polymeric adsorbents is common in wastewater treatment.
This requires an appropriate selection of the cell disruption
method, which in turn affects the purification steps in downstream
operations. Application of the disruption method depends highly
on the nature and type of the cell.
Other factors such as low cost, maximum product release, ease
extraction from the cell debris, and product stability govern the
selection of disruption techniques.
Cellular disruption methods are briefly categorized into mechanical
and non-mechanical methods. Figure shows the various existing
cell disruption methods for the release of biological products.

What is a cell ?
A cell is a mass of cytoplasm that is bound externally by a cell
membrane. Usually microscopic in size, cells are the smallest
structural units of living matter and compose all living things. Most
cells have one or more nuclei and other organelles that carry out a
variety of tasks. Some single cells are complete organisms, such as
a bacterium or yeast. Others are specialized building blocks
of multicellular organisms, such as plants and animals.

Centrifuge capacity cannot be increased by simply increasing the size of the equipment without limit; mechanical stress in
centrifuges increases in . proportion to (radius) 2 so that safe operating speeds are substantially lower in large equipment. The
need for continuous throughput of material in industrial applications also restricts practical operating speeds.To overcome
these difficulties, a
Liquid ExtractionMechanism

CellDisruptionTechniques
Certain measures
should be taken
before the cell
disruption
technique in
order to increase
the efficiency of
the disintegration
method.

relies on grinding and abrasion, while fluid shear relies on high pressure and velocity of fluid.
MechanicalMethods
Mechanical cell disruption methods use both solid and liquid shear, and the cells
are subjected to high pressure/agitation.
Chemical agents or external reagents are not added in mechanical cell disruption,
which is a chief advantage.
Solid shear mainly relies on grinding and abrasion, while fluid shear relies on high
pressure and velocity of fluid.
Mechanical methods of disruption have high efficiency and suits almost all types
of cells.
Most of the methods require cooling after the disruption processes. Heat
generation, product degradation, and high cost are some of the drawbacks of
mechanical lysis techniques.

Mechanical Methods
1. High Pressure Homogenizer
2. Bead Mill
3. Ultrasonication And Cavitation
4. French Press

HighPressureHomogenizer(HPH)
High-pressure homogenizer used
in industries.
Tanks 1 and 2 process the cell
suspension alternatively, and the
cells are disrupted at the valve
seats.

HighPressureHomogenizer(HPH)
HPH is the extensively used method for lysing cells mechanically.
Though HPHs are commonly used for large-scale purposes, they
are also available on smaller scales and can process 25 200 mL of
sample volume.
Cells present in the media after harvest are allowed to pass
through an orifice (0.1 0.2 mm) where they are compressed by
the application of high shear force.
High shear, impact, and cavitation are the principles used in HPH.
Two storage tanks are available in HPH that work alternatively
and process the homogenate. Compressed cells are collected
using a positive displacement pump.
The amount of protein released by HPH is given by Equation:

BeadMill
Bead mills are commonly used on a large scale, yet some are also employed in
laboratories for disrupting cells.
In this technique, cell suspension is mixed with glass, steel, or ceramic beads and
agitated at high speed.
High shear force is applied to the cells when they collide with the beads, which in
turn disintegrate the cell membrane.
The type, size, and weight of the beads to be employed largely depends on the
nature of the cells to be disrupted.
Glass beads with a diameter greater than 0.5
m are suitable for yeast cell disruption, whereas
those smaller than 0.5 m are suitable for
bacterial cell disruption.
The main governing parameters of cell disruption
are bead diameter, number of beads, and agitator speed.

UltrasonicationAndCavitation
The process of cavitation uses the principle of sonochemistry.
In this process, sound energy is generated electrically at a frequency ranging
between 20 and 50 Hz.
The sound energy travels through a probe that passes through the media solution
or water placed in an ultrasonic bath.
This process causes the formation of bubbles, which ultimately causes the cell
membrane to rupture.

UltrasonicationandCavitation
Alternate methods are nitrogen cavitation and hydrodynamic cavitation.The
physical stress is less in cavitation methods compared to ultrasonic method.
Cells are placed in a pressure vessel and nitrogen free of oxygen is passed into the
cells under high pressure (5500 KPa).
Nitrogen bubbles are created, which causes the rupture of cell walls. It is best
suited for fragile cell walls such as mammalian and plant cells and some bacterial
cells, but not for fungi and yeast cells.
Hydrodynamic cavitation is an efficient method for the extraction of lipids in
microalgae; it also causes less stress on the proteins and enzymes compared to
the ultrasonic method.
In hydrodynamic cavitation, the sample is passed through a small channel, which
increases the velocity, thus causing the membrane to rupture and releasing the
intracellular products.

FrenchPress
This is similar to HPH where application of high-pressure technique is employed.Yet
this technique is used only for small-scale purposes.
Initially, the cells are passed through a valve into a pump cylinder, after which they
are allowed to pass through an annular gap. At this region, the pressure applied is
1500 bar.
Then the cells are passed through a discharge valve where the pressure is close to
atmospheric pressure.
Cell disruption occurs at the discharge valve, where the pressure drops suddenly.
Complete disintegration requires more than one pass. However, the number of
passes can be reduced by increasing the pressure, which sometimes causes product
degradation.
Apart from cell concentration and pressure application, the cellular product
released depends on the valve and valve seats.
Heat generated during the process can be reduced by external cooling.
Compact mechanical devices are available that use the similar principles of shear,
friction, grinding, and abrasion (Table).

ComparisonofCompactMechanicalCellDisruptionTechniques
Table:

PhysicalMethods
Nonmechanical or physical methods of cell disruption do not involve any force to
disintegrate the cell.
Nonmechanical methods are usually preferred when there is a small sample size
and also if there is a need to disrupt any specific part of the cell without any
contaminants or a minimal amount of contaminants.
They do not cause much shear to the cell, unlike mechanical methods of cell
disruption.
Physical methods can be applied for cells that do not have a tough cell wall.
Sometimes they are combined with mechanical cell disruption methods to
achieve complete disintegration of cells.

TypesofPhysicalMethods
Freeze-thaw
Thermolysis
Osmotic shock

Freeze-Thaw
The freeze-thaw technique is used to disrupt mostly mammalian and bacterial
cells.
It involves submerging the sample cell solution in dry ice or ethanol for 2 min
followed by thawing the cells in a water bath at 37°C for about 8 min.
This causes the formation of ice on the cells, which leads to rupture of the cell
membrane and release of the intracellular components.
This method cannot be used for cells that are sensitive to temperature.
However, this method is most suitable for highly expressed proteins from E. coli
and also for isolation of recombinant proteins from cytoplasm.
Fifty percent of the recombinant proteins are found to be released in relatively pure
form using the freeze-thaw technique.
However, there are some disadvantages. The freeze-thaw method is time
consuming because more cycles of freezing and thawing are required for efficient
disruption of cell membrane and release of cellular components.
Also, for temperature-sensitive components, this affects the activity.

Thermolysis
Thermolysis is a simple technique that employs only a stirring tank where the
cell suspension is placed.
It is an economical method of cell disruption provided the cells are thermally
stable.
The principle of heat shock is applied, where the cells are heated to 50°C to
disintegrate the cell membrane, leaving the products intact.
For the extraction of cytoplasmic proteins, cells have to be heated to 90°C, at
which some of the protein molecules and enzymes are unstable and they are
degraded.
The solubility of the protein is also varied at higher temperatures, which is a
factor to consider.
These are the major drawbacks of this method.

There are two main mechanisms by which solids are retained by a filter (see Fig. 10.2):
OsmoticShock
Turgor pressure (ranging from 2 6 atm) is necessary for the cells to remain intact and is
directly related to the elasticity of cell membranes.
However, if the environmental conditions are altered, this pressure varies, which
elasticity and in turn the size of pores present in the cell membrane.
Variation in pore size causes the release of intracellular contents.
In this process, cells are first exposed to hypertonic solution (salt or sugar solution) for
them to shrink; then the cells are treated using hypotonic solution (cold water) for the
shrunk cells to swell.
The shrinking and swelling of cells during hypertonic and hypotonic treatments,
respectively, affects the intactness of the cell membrane, thus increasing the pore size.
Increase in pore size causes the release of periplasmic proteins, leaving the cytoplasmic
contents intact.This is the major advantage of this method.

OsmoticShock
It is more suitable for gram
negative bacteria and also
mammalian cells.
The
reduced, however, for cells
that have tougher cell walls.
Sometimes enzymatic
treatment methods are
initially applied followed by
osmotic shock process, which
increases the efficiency of the
disruption process.
The process of osmotic shock
is illustrated in Figure.

ChemicalandEnzymaticMethods
In addition to mechanical and physical methods, chemicals and enzymes are
also used for cell disruption.
Multiple processes are often combined to achieve efficient cell disruption.
The main disadvantage of the method is the need to remove the chemical or
enzyme after the disruption process to follow the downstream processing steps
more easily.
Detergents, solvents, and enzymes are the agents used in chemical and
enzymatic methods.

Detergents
Detergents are amphipathic molecules because they contain a polar head and a
nonpolar tail.
They are used to disrupt the cells or sometimes only the cell membranes for
protein extraction.
They operate by incorporating themselves into the cell wall or cell membrane,
thereby solubilizing lipids and proteins on the cell wall and creating pores on the
cell wall.
This mechanism results in the release of cellular components such as RNA,
DNA, and proteins.
A typical cell contains both hydrophilic and hydrophobic molecules.The unique
property of detergents is that they disturb the hydrophilic-hydrophobic bonding
in cellular components such as lipids, proteins, and polysaccharides.
This interaction of detergents with the cell components is based on the charges
carried by them. Accordingly, they are classified into anionic, cationic, and non-
ionic detergents.

Solvents
Cell disruption, or disturbing the cell membrane to extract proteins by using chemical
solvents, is one of the methods commonly used in labs.
Solvents such as toluene, alcohol, dimethyl sulfoxide, and methyl ethyl ketone are
employed in cell disruption.
Cell lysis and the extraction of the components is based directly on the polarity of the
solvents. Different solvents suit different types of cell.
The major drawback in utilizing solvents is that they damage the cellular components,
especially protein molecules. Hence, they have to be used in minimal quantities. Also,
some of the solvents cause serious environmental problems.
Scaling up is difficult, and the solvent is required in larger amounts when it is applied
to biomass.These reasons have restricted the use of solvents to only small-scale
processes.
Alkaline compounds are also used in the cell lysis process.The working principle is
-glycerol ester bonds in the cell wall,
thus increasing cell permeability for the release of intracellular components.

Enzymes
Different types of enzymes are used to digest cell walls depending
on the type of cell wall. Lysozyme is a well-known enzyme used
for cell disruption.
Enzymes are limited in availability and also very expensive.
Hence, their usage is restricted only to labs.
Recently, microfluidics platforms are being used extensively used
in the process of cell lysis, and they are classified into mechanical,
thermal, chemical, electrical, and acoustic lyses methods

Example
An industrial wastewater contains 10 mg/L chlorophenol, and
is going to be treated by carbon adsorption. 90% removal is
desired. The wastewater is discharged at a rate of 0.1 MGD.
Calculate the carbon requirement for
a) a single ,completely mixed contactor
b) two completely mixed contactors in series
c) a column contactor.
q = 6.74xC0.41
mg/g C mg/L
Freundlich isoherm

q = 6.74x10.41
=6.74
Organic Load = 10-1 mg/Lx 3.78x105
L/d =3.4x106
mg/day
Carbon requirement =
3.4x106
mg/day x
g C
x
1 kg
6.74 mg 1000 g
=505 kg/day
a) Single CMFR
Cinf = 10 mg/L
Ceff = 1 mg/L
mg/g C

b) Two CSTRs connected in series
We assume it. It is not given in the question. If you change it,
you will calculate a different value.
5 mg/L
10 mg/L
1 mg/L

Contactor 1
Organic Load = 10-5 mg/L x 3.78x105
L/d =1.89x106
mg/day
Cinf = 10 mg/L
Ceff = 5 mg/L
q = 6.74x50.41
=13.0 mg/g C
1.89x106
mg/day x
g C
x
1 kg
13.0 mg 1000 g
=145 kg / day

Contactor 2
Cinf = 5 mg/L
Ceff = 1 mg/L
q = 6.74x10.41
=6.74 mg/g C
L/d =1.51x106
mg/day
1.51x106
mg/day x
g C
x
1 kg
6.74 mg 1000 g
=224 kg/day

20
18
16
14
12
10
8
6
4
2
0
1
0 2 3 4 5 6 7 8 9
Ce
10 11 12
qe
Co
Ce of 1 CMFR
and 2nd Contactor
Ce of 1st Contactor

Total C requirement = 145+224=369 kg/day
a) a single , mixed contactor (CMFR)
b) two mixed (CMFR) contactors in
series
C requirement decreased because, in the 1st contactor, we
are able to put more on the surface of the carbon.

Column
Height
Flow
Direction 0 Co
Concentration (mg/L)
Here you start observing your breakthrough curve
when the last layer starts getting saturated.
Everything happens in the primary
adsorption zone (or mass transfer
zone, MTZ). This layer is in contact with
the solution
concentration
at its highest
level, Co. As time
passes, this layer will start saturating.
Whatever escapes this zone will than be
trapped in the next zones. As the
polluted feed water continues to flow
into the column, the top layers of carbon
become, practically, saturated with
solute and less effective for further
adsorption. Thus the primary adsorption
zone moves downward through the
column to regions of fresher adsorbent.

Flow
Direction
0 Co
Last primary adsorption zone. It is called
primary because the upper layers are
not doing any removal job. They are
saturated.
When breakthrough occurs there is some
amount of carbon in the column still not used.
Generally, this is accepted to be 10-15%.

Primary adsorption zone
Region where the solute is most effectively
and rapidly adsorbed.
This zone moves downward with a constant
velocity as the upper regions become
saturated.

Active zones at various times during adsorption and the
breakthrough curve.

Column Contactor
Cinf = 10 mg/L
Ceff = 1 mg/L
q = 6.74x100.41
=17.3 mg/g C
L/d =3.4x106
mg/day
3.4x106
mg/day x
g C
x
1 kg
17.3 mg 1000 g 90%
x
100%
=218.4 kg/day
Assume that the breakthrough occurs while 10%
of the carbon in the column is still not used.

Packed Column Design
It is not possible to design a column accurately without a
test column breakthrough curve for the liquid of interest
and the adsorbent solid to be used.
breakthrough curve

Theoretical Breakthrough Curve

i. Scale up procedure
and
ii. Kinetic approach
are available to design adsorption columns . In
both of the approaches a breakthrough curve
from a test column, either laboratory or pilot
scale, is required, and the column should be as
large as possible to minimize side wall effects.
Neither of the procedures requires the adsorption
to be represented by an isotherm such as the
Freundlich equation.

Scale up Procedure for Packed Columns
Use a pilot test column filled with the carbon to be
used in full scale application.
Apply a filtration rate and contact time (EBCT)
which will be the same for full scale application (to
obtain similar mass transfer characteristics).
Obtain the breakthrough curve.
Work on the curve for scale up.

An industrial wastewater having a TOC of 200 mg/L will be
treated by GAC for a flowrate of 150 m3/day. Allowable TOC
in the effluent is 10 mg/L.
Pilot Plant Data
Q = 50 L/hr
Column diameter = 9.5 cm
Column depth (packed bed) = 175 cm
Packed bed carbon density = 400 kg/m3
Vbreakthrough = 8400 L
Vexhaustion = 9500 L
Example

Breakthrough Curve of the Pilot Plant

a) Filtration rate of the pilot plant
1 1000cm3
x
d
2
Q
= 50
L
x
A hr 1L
2
cm3
= 705
hr.cm2
FR =
d = 9.5 cm
The same FR applies to Packed Column.

b) Area of the Packed Column
FR =
Q
A
Q
A =
FR
3 3
2
1 m
m3 1 h.cm2
1 d 106
cm3
A = 150
d
x x x
24 h
705 cm
. = 8865cm
d=
4 x 8865
= 106cm

c) Empty Bed Contact Time of the Pilot Plant
15 mins.is the EBCT of the Packed Column.
Q
=
d
2
9.5
2
= A x Height = x H = 3.14 x x175 = 12,404 cm3
= 12.4 L
2 2
=
12.4 L
= 0.248 hr = 14.88 15 min
50 L/hr

d) Height of the Packed Column
The same as the height of the Pilot Plant. Because height
is set by and Q/A , and these are the same for Pilot
Plant and the Packed Column.
cm3
2
Q 1 hr
x = 15 min x 705 x = 176cm
A hr.cm 60 min

e) Mass of Carbon required in the Packed Column
2
= 1.76 m x x 1.06
4
3
= 1.553 m
Packed bed carbon density is given by the supplier.
density = 400 kg/m3
m3
1.553 m3
x 400
kg
= 621kg

f) Determination of qe
Mass of carbon in the pilot column =
Volume of the pilot column = 12.4 L
x density= 0.0124 m3
m3
x 400
kg
= 4.96 5 kg
TOC removed by 5 kg of carbon =
200 mg/L x 9500 L = 1.9x106
mg
1.9x106
mg TOC mg
qe = = 380 C
g
5 kg C

g) Fraction of Capacity Left Unused (Pilot Plant)
Fraction of capacity left unused = f =
Total capacity =
mg
9500 L x 200
L
6
= 1.9x10 mg
TOC removed before breakthrough =
mg
8400 L x 200
L
6
= 1.68x10 mg
1.9-1.68 x106
1.9x106
x100 12%
This fraction of capacity left unused will apply to the
Packed Column also.

The same as the Packed Column :
6
30x10 x
mg 1
= 78.9 kg/d
380 mg/g C
d
Amount of carbon consumed =
Carbon consumption rate =
h) Breakthrough time of the Packed Column
3
d 1 m
mg m3
1000 L
Organic Loading = 200 6
L
x 150 x = 30x10 mg/d
621 kg x 1-0.12 = 546.5 kg
Breakthrough time =
546.5 kg
7 days
78.9 kg/d
50 L 24 hr
8400 L x
1 hr
x
1 d
= 7 days

i) Volume Treated Before Breakthrough
m3
treated = 150
d
3
x 7 days = 1050 m

Kinetic Approach
This method utilizes the following kinetic equation.
1
qoM - CoV
k
o
1 + eQ
C 1
C

where
C = effluent solute concentration
Co = influent solute concentration
k1 = rate constant
qo = maximum solid phase concentration of
the sorbed solute, e.g. g/g
M = mass of the adsorbent. For example, g
V = throughput volume. For example, liters
Q = flow rate. For example, liters per hour

Kinetic Approach
The principal experimental information required is
a breakthrough curve from a test column, either
laboratory or pilot scale.
One advantage of the kinetic approach is that the
breakthrough volume, V , may be selected in the
design of a column.

Assuming the left side equals the rigth side,
cross multiplying gives
Rearranging and taking the natural logarithms of
both sides yield the design equation
t
o o
k
q M - C V
1 + eQ =
Co
C

Rearranging and taking the natural logarithms of
both sides yield the design equation.
-
Q Q
k1qoM k1CoV
ln
Co
-1 =
C
y = b - mx

A phenolic wastewater having a TOC of 200 mg/L is to be
treated by a fixed bed granular carbon adsorption column
for a wastewater flow of 150 m3/d, and the allowable
effluent concentration, Ca, is 10 mg/L as TOC. A
breakthrough curve has been obtained from an
experimental pilot column operated at 1.67 BV/h. Other
data concerning the pilot column are as follows:
inside diameter = 9.5 cm , length = 1.04 m,
mass of carbon = 2.98 kg , liquid flowrate = 12.39 L/h ,
unit liquid flowrate = 0.486 L/s.m2 , and
the packed carbon density = 400 kg/m3 .
The design column is to have a unit liquid flowrate of 2.04
L/s.m2 , and the allowable breakthrough volume is 1060 m3.
Example

Example
Using the kinetic approach for design, determine :
The design reaction constant, k1 , L/s-kg.
The design maximum solid - phase concentration,
qo , kg/kg.
The carbon required for the design column, kg.
The diameter and height of the design column, m.
The kilograms of carbon required per cubic meter
of waste treated.

V (L) C(mg/L) C/Co Co/C Co/C-1 ln(Co/C-1)
0 0 0,000
378,0 9 0,045 22,222 21,222 3,06
984,0 11 0,055 18,182 17,182 2,84
1324,0 8 0,040 25,000 24,000 3,18
1930,0 9 0,045 22,222 21,222 3,06
2272,0 30 0,150 6,667 5,667 1,73
2520,0 100 0,500 2,000 1,000 0,00
2740,0 165 0,825 1,212 0,212 -1,55
2930,0 193 0,965 1,036 0,036 -3,32
3126,0 200 1,000 1,000 0,000

200
180
160
140
120
100
80
60
40
20
0
0 500 1000 1500 2000 2500 3000 3500
V, Liters
C,
mg/L

-9
16
11
6
1
-4
0 1000 3000 4000
ln(Co/C
-
1)
2000
V (L)
Plot of Complete Data Set

Take the linear range only!
y = -0,0064x + 15,787
-5
0
5
10
15
0 3000 4000
ln(Co/C
-
1)
1000 2000
Volume treated (L)

15.787= 0 1
Q
q k M
-1
0.0064L = 1 0
Q
k C
-4 L
h
a)k = mg
1 200 L
(0.0064L-1) (12.39L)
=3.96 10
mg h
3.96 10-4 L 1h 106mg=0.11 L
mg h 3600s 1kg kg s

h 3600s
12.39L 1h
q 0.11L 2.98kg
b)15.787= 0 kg s
q = h 3600s
0
15.787 12.39L 1h
0.11 L 2.98kg
kg s
q =0.166kg
0 kg

Q = 6250 L / h
V = 1050000 L
C0 = 200 mg / L
q =0.166 kg
0 kg
k =3.96 10 4 L
1 mg h
k1qoM k1CoV
-
Q Q
ln
Co
-1 =
C
Using
ln
200
-1 = -
M 3.96 10 4 L
10 6250 6250
kg
3.96 10 4 L
0.166
kg
mg h mg h L
200
mg
1050000L
L
h
L
h
M =1545009487 mg =1545 kg

Q = 6250 L/ h =1.736 L/ s
Unit liquid flowrate = 2.04 L/ s m2
(given)
(given)
400
kg
m3
M =1545 kg
Packet carbon density = 400 kg / m3
Then, design bed volume is;
V
1545 kg
3.86 m3
0.85m2
2.04 L/ s m2
1.736 L/ s
Cross section area =
3.86 m3
Column height =
0.85 m2
4.54 m
d =1.04 m
1050m3
TB =
150m3
/d
7d
Breakthrough time is;

Scale-up approach:
1. The design bed volume (BV) is found as;
150 m3
/ d kg 1000 L
Mt =
m3
8.954 kg/h
24 h 698 L
3
1.67 BV / h = = 6.25 m / h
150 m3
/ d
24 h / d
BV = 3.74 m3
2. The mass of carbon required is;
M = BV = 3.74 m3
400 kg / m3
=1500 kg
From the breakthrough curve the volume treated at the allowable
breakthrough (10 mg/L TOC) is 2080 L. So, the solution treated per
kilogram of carbon is 2080 L/2.98 kg or 698 L/kg (pilot scale). The same
applies to the design column; for a flow rate of 150 m3/d.
3. The weight of carbon exhausted per hour (Mt) is

200
180
160
140
120
100
80
60
40
20
0
0 500 1000 1500 2000
V, Liters
2500 3000 3500
C,
mg/L

B
V = Q T =150 m3
/ d 7 d =1050 m3
4. The breakthrough time is;
1500 kg
T = =168 h = 7 d
8.954 kg / h
5. The breakthrough volume of the design column is;
Comparing the results of two approaches:
Kinetic approach
M=1545kg
Scale-up approach
M =1500 kg
V =1050 m3
B
TB =7d
V =1050 m3
B
TB =7d
=3.86m3
Design
V Design
V =3.74m3

A phenolic wastewater that has phenol concentration of 400
mg/L as TOC is to be treated by a fixed bed granular
carbon adsorption column for a wastewater flow of 227100
L/d, and the allowable effluent concentration, Ca, is 35 mg/L
as TOC. A breakthrough curve has been obtained from an
experimental pilot column operated at 1.67 BV/h. Other
data concerning the pilot column are as follows:
inside diameter = 9.5 cm , length = 1.04 m,
mass of carbon = 2.98 kg , liquid flowrate = 17.42 L/h ,
unit liquid flowrate = 0.679 L/s.m2 , and
the packed carbon density = 401 kg/m3 .
The design column is to have a unit liquid flowrate of 2.38
L/s.m2 , and the allowable breakthrough volume is 850 m3.
HOME WORK
Problem statement

HOME WORK
Using the kinetic approach for design, determine :
The design reaction constant, k1 , L/s-kg.
The design maximum solid - phase
concentration, qo , kg/kg.
The carbon required for the design column, kg.
The diameter and height of the design column,
m.
The kilograms of carbon required per cubic
meter of waste treated.

V
(L)
C
(mg/L) C/Co Co/C Co/C - 1 ln(Co/C - 1)
15 12 0.030 33.333 32.333 3.476
69 16 0.040 25.000 24.000 3.178
159 24 0.060 16.667 15.667 2.752
273 16 0.040 25.000 24.000 3.178
379 16 0.040 25.000 24.000 3.178
681 20 0.050 20.000 19.000 2.944
965 28 0.070 14.286 13.286 2.587
1105 32 0.080 12.500 11.500 2.442
1215 103 0.258 3.883 2.883 1.059
1287 211 0.528 1.896 0.896 -0.110
1408 350 0.875 1.143 0.143 -1.946
1548 400 1.000 1.000 0.000

Extraction

Introduction
Liquid extraction is used to isolate many pharmaceutical products from animal
and plant sources. In liquid extraction of fermentation products, components
dissolved in liquid are recovered by transfer into an appropriate solvent.
The simplest equipment for liquid extraction is the separating funnel used for
laboratory-scale product recovery.
Liquids forming two distinct phases are shaken together in the separating
funnel; solute in dilute solution in one solvent transfers to the other solvent to
form a more concentrated solution.
The two phases are then allowed to separate and the heavy phase is
withdrawn from the bottom of the funnel.The phase containing the solute in
concentrated form is processed further to purify the product.

Liquid-Liquid Extraction
In liquid liquid extraction (solvent extraction), a liquid feed of two or more
components is contacted with a second liquid phase, called the solvent,
which is immiscible or only partly miscible with one or more feed
components and completely or partially miscible with one or more of the
other feed components.
The solvent is selected to partially dissolve certain species of the liquid
feed, effecting at least a partial separation of the feed components.
The solvent may be a pure compound or a mixture. If the feed is an
aqueous solution, an organic solvent is used; if the feed is organic, the
solvent is often water.

Liquid-Liquid Extraction
Extraction with organic solvents is a major separation technique in
bioprocessing, particularly for recovery of antibiotics.
However, organic solvents are unsuitable for isolation of proteins and other
sensitive biopolymers.
Techniques are being developed for aqueous two-phase extraction of these
molecules.
Aqueous solvents which form two distinct phases provide favourable
conditions for separation of proteins, cell fragments and organelles with
protection of their biological activity.
Two-phase aqueous systems are produced when particular polymers or a
polymer and salt are dissolved together in water above certain
concentrations.

Fundamentals
The low-density, solvent-rich stream, called the extract,
exits from the top of the extractor with 99.8% of the
acetic acid in the feed.
The high-density, carrier-rich stream, called the raffinate,
exiting from the extractor bottom, contains only 0.05
wt% acetic acid.
The extract is sent to a distillation column, where glacial
acetic acid is the bottoms product.
The overhead vapor, which is rich in ethyl acetate but
also contains appreciable water vapor, splits into two
liquid phases when condensed.

Fundamentals
These are separated in the decanter by gravity. The
lighter ethyl acetate-rich phase is divided into reflux and
solvent recycle to the extractor.
The water-rich phase from the decanter is sent, together
with the raffinate from the extractor, to a second
distillation column, where wastewater is the bottoms
product, and the ethyl-acetate-rich overhead is recycled
to the decanter.
Makeup ethyl-acetate solvent is provided for solvent
losses to the glacial acetic acid and wastewater.

Fundamentals
In general, extraction is preferred over distillation for:
1. Dissolved or complexed inorganic substances in organic or aqueous
solutions.
2. Removal of a contaminant present in small concentrations, such as a color
former in tallow or hormones in animal oil.
3. A high-boiling component present in relatively small quantities in an aqueous
waste stream, as in the recovery of acetic acid from cellulose acetate.

Fundamentals
4. Recovery of temperature-sensitive materials, where extraction may be
less expensive than vacuum distillation.
5. Separation of mixtures according to chemical type rather than relative
volatility.
6. Separation of close-melting or close-boiling liquids, where solubility
differences can be exploited.
7. Separation of mixtures that form azeotropes.
The key to an effective extraction process is a suitable solvent.

Fundamentals
In addition to being stable, non-toxic, inexpensive, and easily recoverable, a
solvent should be relatively immiscible with feed components other than the
solute, and have a different density from the feed to facilitate phase
separation by gravity.
It must have a high affinity for the solute, from which it can be easily
separated by distillation, crystallization, or other means.
Ideally, the distribution (partition) coefficient for the solute between the
liquid phases should be greater than one, or a large solvent-to-feed ratio
will be required.

AqueousTwo-PhaseLiquidExtraction
Aqueous two-phase separations are of special interest for extraction of
enzymes and recombinant proteins from cell debris produced by cell
disruption.
After partitioning, product is removed from the extracting phase using other
unit operations such as precipitation or crystallisation.
The extent of differential partitioning between phases depends on the
equilibrium relationship for the system.The partition coefficient Kis defined
as:
where CAu is the equilibrium concentration of componentA in the upper phase
and CAl is the equilibrium concentration of A in the lower phase.

Even when the partition coefficient is low, good product recovery or yield can
be achieved by using a large volume of the phase preferred by the solute.
Yield of A in the upper phase,YU, is defined as:
Where
VU = volume of the upper phase,
V I = volume of the lower phase,
V 0 = original volume of solution containing the product and
CA0 = original product concentration in that liquid

In the lower phase, yieldYl is defined as:
Another parameter used to characterise two-phase partitioning is the
concentration factor or purification factor, ,defined as the ratio of product
concentration in the preferred phase to the initial product concentration:

Example
Aqueous two-phase extraction is used to recover a-amylase from solution. A
polyethylene glycol-dextran mixture is added and the solution separates into
two phases.The partition coefficient is 4.2. Calculate the maximum possible
enzyme recovery when:
(a) the volume ratio of upper to lower phases is 5.0; and
(b) the volume ratio of upper to lower phases is 0.5.

Solution:
As the partition coefficient is greater than 1, enzyme prefers the upper phase.
Yield at equilibrium is therefore calculated for the upper phase. Dividing both
numerator and denominator of Eq. byV l gives:

Liquid-LiquidMassTransfer
Liquid-liquid mass transfer between immiscible solvents is most often
encountered in the product-recovery stages of bioprocessing.
Organic solvents are used to isolate antibiotics, steroids and alkaloids from
fermentation broths; two-phase aqueous systems are used in protein
purification.
Liquid-liquid mass transfer is also important when hydrocarbons are used as
substrates in fermentation, e.g. in production of microbial biomass for single-
cell protein.

The situation at the interface between
two immiscible liquids is shown in
Figure.
ComponentA is present at bulk
concentration CAI in one liquid phase;
this concentration falls t0 CAI i at the
interface.
In the other liquid, the concentration of A
falls from CA2 i at the interface to CA2 in
the bulk.

The rate of mass transfer N A in each liquid phase can be obtained from Equation:
It can be assumed that there is negligible resistance to mass transfer at the actual
interface, i.e. within distances corresponding to molecular free paths on either
side of the phase boundary.

A typical equilibrium curve relating concentrations
of solute A in two immiscible liquid phases is
shown in Figure.
The points making up the curve are obtained
readily from experiments.
Equilibrium distribution of one solute between two
phases is conveniently described in terms of the
distribution law.
At equilibrium, the ratio of solute concentrations in
the two phases is given by the distribution
coefficient or partition coefficient, m.

If we now multiply Eq. by m:
and divide Eq. by m:

Equipment for Solvent Extraction
Equipment similar to that used for absorption, stripping, and
distillation is sometimes used for extraction, but such devices
are inefficient unless interfacial tension and liquid viscosities
are low and differences in phase density are high.
Generally, mechanically agitated or centrifugal devices are
preferred, especially if many equilibrium stages are required.
During passage through extraction equipment, one phase is
the dispersed phase (discontinuous phase) in the form of
droplets and, the other phase is the continuous phase.
In static extraction columns of the spray, packed, and sieve
tray type, it is preferred to disperse the phase of higher
entering volumetric flow rate, unless the other phase has a
high viscosity.

Mixer-Settlers
In mixer-settlers, the two liquid phases are first mixed in
a vessel by one of several types of impellers and then
separated in a second vessel by gravity-induced settling.
Any number of mixer-settler units may be connected
together to form a multistage countercurrent cascade.
However, floor space can be a major factor.
During mixing, one of the liquids is dispersed in the
form of small droplets in the other liquid. The dispersed
phase may be either the heavier or the lighter phase.

Spray Columns
The simplest and one of the oldest extraction devices is
the spray column. Either the heavy phase or the light
phase can be dispersed, as seen in Figure 8.6.
The droplets of the dispersed phase are generated at
the inlet, usually by spray nozzles. Because of the lack of
column internals, combined volumetric throughputs can
be large, depending upon phase-density difference and
phase viscosities.
As in gas absorption, axial dispersion (back-mixing) in
the continuous phase limits these devices to applications
where only one or two stages are required.

A

Packed Columns
Axial dispersion in a spray column can be reduced, but not eliminated,
by packing the column. This also improves mass transfer by breaking up
large drops to increase interfacial area and promote mixing in drops by
distorting droplet shape.
With the exception of Raschig rings, the packings used in distillation and
absorption are suitable for liquid liquid extraction; however, choice of
packing material is more critical.
For best performance, the packing should be preferentially wetted by
the continuous phase. Throughput, especially with newer packings, is
large. Because of back-mixing, the HETS is generally large, making
packed columns suitable only when few equilibrium stages are needed.

Sieve-Tray Columns
Sieve trays reduce axial mixing and promote a stagewise type of
contact. The dispersed phase, which is analogous to vapor bubbles in
distillation, flows up the column, with redispersion at each tray.
The heavy phase is continuous, flowing at each stage through a
downcomer, and then across the tray like a liquid in a distillation
tower. If the heavy phase is dispersed, upcomers are used for the
light phase.
Sieve-tray extractors are subject to the same limitations as distillation
columns: flooding, entrainment, and, to a lesser extent, weeping.
An additional problem is scum formation at phase interfaces due to
small amounts of impurities.

Columns with Mechanically Assisted Agitation
If:
1. Interfacial tension is high,
2. Density difference between liquid phases is low, and/or
3. Liquid viscosities are high, then gravitational forces are
inadequate for proper phase dispersal and turbulence
creation.

In that case, mechanical agitation is necessary to increase
interfacial area per unit volume, thus decreasing mass-
transfer resistance.
For packed and plate columns, agitation can be provided by
an oscillating pulse to the liquid, either by mechanical or
pneumatic means.
Pulsed, perforated-plate columns find considerable
application in the nuclear industry.
The most prevalent agitated columns are those that employ
rotating agitators driven by a shaft extending axially through
the column. The agitators create shear mixing zones, which
alternate with settling zones. Nine of the more popular
mechanically-agitated devices are shown in Figure 8.7a i.

Comparison of Industrial Extraction Columns
Maximum loadings and sizes for industrial extraction columns are listed in
Table 8.2.
As seen, the Lurgi tower, RDC, and Graesser extractors have been built in
very large sizes. Combined volumetric throughputs per unit cross-sectional
area are highest for the Karr extractor and lowest for the Graesser extractor.
Table 8.3 lists the advantages and disadvantages of the various types of
extractors, and Figure 8.8 shows a selection scheme for commercial
extractors.

Equipment for Solvent
Extraction
For example, if only a small
number of stages is required, a
set of mixer-settler units might
be selected. If more than five
theoretical stages, a high
throughput, and a large load
range (m3 2-h) are needed,
and floor space is limited, an
RDC or ARD contactor should
be considered.

General Design Considerations
Liquid liquid extraction involves more design variables than distillation. To
determine stages, one of the three cascade arrangements in Figure 8.9, or an
even more complex arrangement, must be selected.
Packed-column configurations are shown in Figure 8.9, but other extraction
equipment may be preferred. The single-section cascade of Figure 8.9a, which
is similar to that used for absorption and stripping, will transfer solute in the
feed to the solvent.
The two-section cascade of Figure 8.9b is similar to that used for distillation.
Solvent enters at one end and reflux, derived from the extract, enters at the
other end.

The feed enters in between. With two sections, depending on solubilities, it is
sometimes possible to achieve a separation between two feed components; if
not, a dual-solvent arrangement with two sections, as in Figure 8.9c, with or
without reflux at the ends, may be advantageous.
For configurations 8.9b and 8.9c, calculations should be made by a process
simulator. For the configuration of Figure 8.9a, it is useful and instructive to
make the graphical calculations.

A

Operative factors are:
1. Entering feed flow rate, composition, temperature, and pressure.
2. Type of stage configuration (one- or two-section).
3. Desired degree of recovery of one or more solutes for one-section cascades.
4. Degree of feed separation for two-section cascades.
5. Choice of solvent(s).
6. Operating temperature (often ambient).
7. Operating pressure (greater than the bubble point of both phases).
8. Minimum-solvent flow rate and actual-solvent flow rate as a multiple of the minimum rate for one-
section cascades or reflux rate and minimum reflux ratio for two-section cascades.

9. Number of equilibrium stages.
10. Emulsification and scum-formation tendency.
11. Interfacial tension.
12. Phase-density difference.
13. Maximum residence time to avoid degradation.
14. Type of extractor.
15. Extractor cost and horsepower requirement.

The ideal solvent has:
1. High selectivity for the solute relative to the carrier to minimize the need to
recover carrier from the solvent.
2. High capacity for dissolving the solute to minimize solvent-to-feed ratio.
3. Minimal solubility in the carrier.
4. A volatility sufficiently different from the solute that recovery of the solvent
can be achieved by distillation, but not so high that a high extractor pressure
is needed, or so low that a high temperature is needed if the solvent is
recovered by distillation.

5. Stability to maximize the solvent life and minimize the solvent makeup
requirement.
6. Inertness to permit use of common materials of construction.
7. Low viscosity to promote phase separation, minimize pressure drop, and
provide a high-solute mass-transfer rate.
8. Non-toxic and non-flammable characteristics to facilitate its safe use.
9. Availability at a relatively low cost.

10. Moderate interfacial tension to balance the ease of dispersion and the
promotion of phase separation.
11. Large difference in density relative to the carrier to achieve a high capacity in
the extractor.
12. Compatibility with the solute and carrier to avoid contamination.
13. Lack of tendency to form a stable rag or scum layer at the phase interface.
14. Desirable wetting characteristics with respect to extractor internals.

Chromatography

Chromatography is a separation
procedure for resolving mixtures
and isolating components.
The basis of chromatography is
differential migration, i.e. the
selective retardation of solute
molecules during passage
through a bed of resin particles.
A schematic description of
chromatography is shown in
Figure.

The pattern of solute peaks emerging from a chromatography
column is called a chromatogram.
The fluid carrying solutes through the column or used for elution is
known as the mobile phase.
The material which stays inside the column and effects the
separation is called the stationary phase.

ChromatographicTerms
The analyte is the component(s) to be separated from the mixture.
A bonded phase that is covalently linked to the support particles
present in the inside wall of the column is termed as stationary phase.
Analytical chromatography detects the presence and concentration of
an analyte(s) in a sample.
A chromatogram is the output displayed in the monitor of the
chromatograph. Each peak or pattern on the chromatogram
corresponds to a component(s) present in the mixture.
The analyte is carried by the mobile phase solvent, called eluent, and
eluate is the mobile phase that leaves the column. An eluotropic series
is the order of solvents based on their eluting power.

ChromatographicTerms
The mobile phase is the phase that flows in a defined direction. It may be a liquid
(LC), a gas (GC), or a supercritical fluid (supercritical-fluid chromatography [SFC]).
The sufficient quantities of a substance are purified using preparative
chromatography for further use.
The time taken by a particular analyte to pass through the entire system is termed
retention time.
The sample is the mixture to be separated in chromatography.
The solute is the single component or mixture of components present in the
sample.
The solvent has the capacity to solubilize another substance completely.
The detector refers to the instrument used to analyze the analytes qualitatively
and quantitatively after separation.

Chromatographic System

Gas chromatography (GC)
In gas chromatography, the mobile phase is a gas.
Gas chromatography is used widely as an analytical tool for
separating relatively volatile components such as alcohols,
ketones, aldehydes and many other organic and inorganic
compounds.
Liquid chromatography (LC)
However, of greater relevance to bioprocessing is liquid
chromatography, which can take a variety of forms.
Liquid chromatography finds application both as a laboratory
method for sample analysis and as a preparative technique for
large-scale purification of biomolecules.

Methods
Chromatography is a high-resolution technique and therefore suitable
for recovery of high-purity therapeutics and pharmaceuticals.
Chromatographic methods available for purification of proteins,
peptides, amino acids, nucleic acids, alkaloids, vitamins, steroids and
many other biological materials include:
Adsorption chromatography
Partition chromatography
Ion-exchange chromatography
Gel chromatography
Affinity chromatography

Adsorption Chromatography
Biological molecules have varying tendencies to adsorb onto polar
adsorbents such as silica gel, alumina, diatomaceous earth and charcoal.
Performance of the adsorbent relies strongly on the chemical composition
of the surface, i.e. the types and concentrations of exposed atoms or
groups.
The order of elution of sample components depends primarily on molecule
polarity. Because the mobile phase is in competition with solute for
adsorption sites, solvent properties are also important.

Partition Chromatography
Partition chromatography relies on the unequal distribution of solute between
two immiscible solvents.
This is achieved by fixing one solvent (the stationary phase) to a support and
passing the other solvent containing solute over it.
The solvents make intimate contact allowing multiple extractions of solute to
occur.
Several methods are available to chemically bond the stationary solvent to
supports such as silica.
When the stationary phase is more polar than the mobile phase, the technique is
called normal-phase chromatography.
When non-polar compounds are being separated it is usual to use a stationary
phase which is less polar than the mobile phase; this is called reverse-phase
chromatography.

relies on grinding and abrasion, while fluid shear relies on high pressure and velocity of fluid.
Ion exchange Chromatography
The basis of separation in this procedure is electrostatic attraction
between the solute and dense clusters of charged groups on the column
packing.
Ion-exchange chromatography can give high resolution of
macromolecules and is used commercially for fractionation of antibiotics
and proteins.
Solutes are eluted by changing the pH or ionic strength of the liquid
phase; salt gradients are the most common way of eluting proteins from
ion exchangers.

Gel Chromatography
This technique is also known as molecular-sieve chromatography, exclusion
chromatography, gel filtration and gel-permeation chromatography.
Molecules in solution are separated in a column packed with gel particles of
defined porosity.
Gels most often used are cross-linked dextrans, agaroses and polyacrylamide gels.
The speed with which components travel through the column depends on their
effective molecular size.
Large molecules are completely excluded from the gel matrix and move rapidly
through the column to appear first in the chromatogram.
Small molecules are able to penetrate the pores of the packing, traverse the
column very slowly, and appear last in the chromatogram.
Molecules of intermediate size enter the pores but spend less time there than the
small solutes.

AffinityChromatography
This separation technique exploits the binding specificity of biomolecules.
Column packing is prepared by linking a binding molecule called a ligand to an
insoluble support; when sample is passed through the column, only solutes with
appreciable affinity for the ligand are retained.
The ligand must be attached to the support in such a way that its binding
properties are not seriously affected; molecules called spacer arms are often
used to set the ligand away from the support and make it more accessible to the
solute.
Many ready-made support-ligand preparations are available commercially and
are suitable for a wide range of proteins.
Conditions for elution depend on the specific binding complex formed: elution
usually involves a change in pH, ionic strength or buffer composition.
Affinity chromatography using antibody ligands is called immuno-affinity
chromatography.

Principles of Liquid Chromatography
Basic Requirements:
For high capacity, the solid support must be porous with high internal surface
area; it must also be insoluble and chemically stable during operation and
cleaning.
Ideally, the particles should exhibit high mechanical strength and show little or
no non-specific binding.
The low rigidity of many porous gels was initially a problem in industrial-scale
chromatography; the weight of the packing material in large columns and the
pressures developed during flow tended to compress the packing and impede
operation.
However, many macroporous gels and composite materials of high rigidity are
now available for industrial use.

Chromatographic Separations
Two methods for carrying out chromatographic separations are:
High-performance liquid chromatography (HPLC) and
Fast protein liquid chromatography (FPLC)
In principle, any of the types of chromatography described above can be
executed using HPLC and FPLC techniques.

Chromatographic separations traditionally performed under atmospheric
pressure in vertical columns with manual sample feed and gravity elution are
carried out faster and with better resolution using densely-packed columns and
high flow rates in HPLC and FPLC systems.
The differences between HPLC and FPLC lie in the flow rates and pressures used,
the size of the packing material, and the resolution accomplished.
In general, HPLC instruments are designed for small-scale, high-resolution
analytical applications; FPLC is tailored for large-scale purification.

Chromatography is essentially a batch operation; however industrial
chromatography systems can be monitored and controlled for easy automation.
Cleaning the column in place is generally difficult.
Depending on the nature of the impurities contained in the samples, rather harsh
treatments with concentrated salt or dilute alkali solutions are required; these
may affect swelling of the gel beads and, therefore, liquid flow in the column.
Regeneration in place is necessary as repacking of large columns can be laborious
and time-consuming.
Repeated use of chromatographic columns is essential because of their high cost.

Differential Migration
Differential
migration
provides the
basis for
chromatographic
separation and is
explained
diagrammatically
in Figure.

Several parameters are used to characterise differential migration.
An important variable is the volumeVe of eluting solvent required to
carry the solute through the column until it emerges at its maximum
concentration.
Each component separated by chromatography has a different elution
volume. Another parameter commonly used to characterise elution is
the capacity factor, k:
V0 is the void volume
For two solutes, the ratio of their capacity factors kI and k2 is called the
selectivity or relative retention,

In gel chromatography where separation is a function of effective
molecular size, the elution volume is easily related to certain physical
properties of the gel column.The total volume of a gel column is:
where
VT is total volume,
Vo is void volume outside the particles,
Vi is internal volume of liquid in the pores of the particles, and
Vs is volume of the gel itself

Solutes which are only partly excluded from the stationary phase elute
with a volume described by the following equation:
where Kp is the gel partition coefficient, defined as the fraction of
internal volume available to the solute. For large molecules which do
not penetrate the solid, Kp = 0. From above equation:

Experimental determination of Kp depends on knowledge ofVi, which is
difficult to measure accurately.Vi is usually calculated using the equation:
where a is mass of dry gel andWr is the water regain value, defined as the
volume of water taken up per mass of dry gel.The value forWr is generally
specified by the gel manufacturer.
If, as is often the case, the gel is supplied already wet and swollen, the value of
a is unknown andVi is determined using the following equation:

Example
A pilot-scale gel-chromatography column packed with Sephacryl resin is used
to separate two hormones A and B. The column is 5 cm in diameter and 0.3m
high; the void volume is 1.9 x 10 -4 m 3. The water regain value of the gel is 3
x 10 -3 m 3 kg-1 dry Sephacryl; the density of wet gel is 1.25 x 103 kg m-3. The
partition coefficient for hormone A is 0.38; the partition coefficient for hormone
B is 0.15. If the eluant flow rate is 0.7 L h-1, what is the retention time for each
hormone?

Solution
The total column volume is:

Zone Spreading
Zone spreading is not so important when migration rates vary widely because
there is little chance that solute peaks will overlap.
However if the molecules to be separated have similar structure, migration rates
will also be similar and zone spreading must be carefully controlled.
Spreading of the solute peak is caused by several factors represented
schematically in Figure .
Axial diffusion. As solute is carried through the column, molecular diffusion of
solute will occur from regions of high concentration to regions of low
concentration.
Eddy diffusion. In columns packed with solid particles, actual flow paths of liquid
through the bed can be highly variable.
Local non-equilibrium effects. In most columns, lack of equilibrium is the most
important factor affecting zone spreading, although perhaps the most difficult to
understand.

Zone Spreading
Zone spreading in a Chromatography

TheoreticalPlatesinChromatography
The chromatography column is considered to be made up of a number of
segments or plates of height H, the magnitude of H is of the same order as the
diameter of the resin particles. Within each segment equilibrium is supposed to
exist.
Nevertheless the idea of theoretical plates is applied extensively, mainly
because it provides a parameter, the plate height H, which can be used to
characterise zone spreading.
Use of the plate height, which is also known as the height equivalent to a
theoretical plate (HETP), is acceptable practice in chromatography design even
though it is based on a poor model of column operation.
HETP is a measure of zone broadening; in general, the lower the HETP value the
narrower is the solute peak.

HETP depends on various processes which occur during elution of a
chromatography sample.
A popular and simple expression for HETP takes the form:
where H is plate height, u is linear liquid velocity, and A, B and C are
experimentally-determined kinetic constants.
A, B and C include the effects of liquid-solid mass transfer, forward and
backward axial dispersion, and non-ideal distribution of liquid around the
packing.

HETP for a particular component is related to the elution volume and width of
the solute peak as it appears on the chromatogram.
If, as shown in Figure, the pulse has the standard symmetrical form of a normal
distribution around a mean value x, the number of theoretical plates can be
calculated as follows:
where N is number of theoretical plates, Ve is the distance on the chromatogram
corresponding to the elution volume of the solute, and w is the base line width of
the peak between lines drawn tangent to the inflection points of the curve.
Above equation applies if the sample is introduced into the column as a narrow
pulse. Number of theoretical plates is related to HETP as follows:
where L is the length of the column.

Resolution
Resolution is a measure of zone overlap in chromatography and an indicator
of column efficiency. For separation of two components, resolution is given by
the following equation:
where
RN is resolution,
Vel andVe2 are distances on the chromatogram corresponding to elution
volumes for components 1 and 2, and
wI and w2 the baseline widths of the chromatogram peaks as shown in Figure.

Resolution
Column resolution can be expressed in terms of HETP. Assuming wI and w2 are
approximately equal,
Equation becomes:
Substituting for w2 from Equation :
The term can be expressed in terms of k2 and using the equations
So above Equation becomes:
Using the expression for N ( ),
The equation for column resolution is:

ScalingUpChromatography
The solution to scale-up is to keep the same column length, linear flow velocity
and particle size as in the small column, but increase the column diameter.
The larger capacity of the column is therefore due solely to its greater cross-
sectional area.
Sample volume and volumetric flow rate are increased in proportion to column
volume.
In this way, all the important parameters affecting the packing matrix, liquid flow,
mass transfer and equilibrium conditions are kept constant; similar column
performance can therefore be expected.
Because liquid distribution in large-diameter packed columns tends to be poor,
care must be taken to ensure liquid is fed evenly over the entire column cross-
section.

Membrane Separation Technologies

Theoretical Background
In a membrane-separation process, a mixture of two or
more components is partially separated by means of a
semipermeable barrier (the membrane) through which
some species travel faster than others.
The most general membrane process is shown in Figure,
where the feed mixture is separated into a retentate
(that part of the feed that does not pass through the
membrane) and a permeate (that passes through the
membrane).
The feed, retentate, and permeate are usually liquid or
gas.

The barrier is most often a thin, nonporous, polymeric
film, but may also be porous polymer, ceramic, or metal
material, or even a liquid, gel, or gas.
To maintain selectivity, the barrier must not dissolve,
deform, disintegrate, or break.
The optional sweep, is a liquid or gas used to facilitate
removal of the permeate.

In membrane separations:
1. The two products are usually miscible,
2. The separating agent is a semipermeable barrier, and
3. A sharp separation is often difficult to achieve.
Membrane separations differ in some respects from the
more common separation operations of absorption,
distillation, and liquid liquid extraction.

Membrane Materials
Originally, membranes were made from processed natural polymers such as
cellulose and rubber; however, since 1930, many are custom-made synthetically.
Polymers are amorphous or crystalline. The former refers to a polymer that is glass-
like in appearance and lacks crystalline structure, whereas the latter refers to a
polymer that is usually opaque and has a crystalline structure.
If the temperature of a glassy polymer is increased to the glass-transition
temperature, Tg, the polymer becomes rubbery. In rubbery polymers, portions of the
chain can move and the backbone can rotate, resulting in high diffusion rates of the
permeate.
In glassy polymers, thermal motion of the polymer is largely curtailed, resulting in
low permeate diffusion rates.

Membrane Materials
The linear-chain polymers soften with an increase in temperature, are
soluble in organic solvents, and are referred to as thermoplastic polymers.
At the other extreme, highly cross-linked polymers decompose at high
temperature, are not soluble in organic solvents, and are referred to as
thermosetting polymers.
For polymeric membranes, a classification based on the arrangement or
conformation of the polymer molecules is useful.

Membrane Materials
To separate a binary chemical mixture, a polymer membrane must
possess high permeance and a high permeance ratio for the two
components being separated.
The permeance for a given species diffusing through a membrane of
given thickness is analogous to a mass-transfer coefficient, i.e., the flow
rate of that species per unit cross-sectional area of membrane per unit
driving force (concentration, partial pressure, etc.) across the membrane
thickness.
The molar transmembrane flux (flow rate per unit cross-sectional area of
membrane) of species i is:

Membrane Materials
The molar transmembrane flux (flow rate per unit cross-sectional area of
membrane) of species i is:

Membrane Materials
Polymer membranes can be characterized as:
Dense, amorphous membranes
Pores of microscopic dimensions may be present, but they are generally less than a few Å
(angstroms) in diameter.
Microporous membranes
Contain interconnected pores and are categorized by their use in microfiltration (MF),
ultrafiltration (UF), and nanofiltration (NF).

Membrane Materials
The pores are formed by a variety of proprietary techniques, some of
which are described by Baker [5]. Such techniques are valuable for
producing isotropic (symmetric), microporous, crystalline membranes.
Permeability for microporous membranes is high, but selectivity is low
for small molecules, due in part to pore-size distributions that can be
variable and broad.
However mixtures of molecules smaller and larger than the pore size
may be separated almost perfectly by size.

Membrane Materials
Polymer membrane applications are usually limited to temperatures
below 200 C and to mixtures that are chemically inert.
Operation at high temperatures and with chemically active mixtures
requires membranes made of inorganic materials.
These include microporous ceramics, metals, and carbon; and dense
metals, such as palladium, that allow the selective diffusion of small
molecules such as hydrogen and helium.

Membrane Materials
Examples of inorganic membranes are:
1. Asymmetric, microporous -alumina tubes with 40 100 Å pores at the
inside surface and 100,000 Å pores at the outside;
2. Microporous glass tubes, whose pores may be filled with other oxides
or the polymerization pyrolysis product of trichloromethylsilane;
3. Silica hollow fibers with 3 5 Å pores;
4. Porous ceramic, glass, or polymer materials coated with a thin, dense,
palladium metal film just a few m thick;
5. Sintered metal;
6. Pyrolyzed carbon; and
7. Zirconia on sintered carbon.
Extremely fine pores (<10 Å) are necessary to separate gas mixtures. Larger
pores (>50 Å) are satisfactory for the separation of large molecules or solid
particles from solutions containing small molecules.

Membrane Modules
Membrane Separation Technologies

Mass Transfer in Membranes
Membranes can be macro-porous, microporous, or dense (nonporous). Only
microporous or dense membranes are permselective.
Macro-porous membranes are used to support thin microporous and dense
membranes when significant pressure differences across the membrane are
necessary to achieve high flux.
The theoretical basis for mass transfer through microporous membranes is
more highly developed than that for dense membranes, so porous-
membrane transport is discussed first, with respect to bulk flow, liquid
diffusion, and then gas diffusion.

This is followed by nonporous (dense) membrane solution-diffusion
transport, for liquid and gas mixtures. External mass-transfer resistances in
the fluid films on either side of the membrane are treated where appropriate.
It is important to note that, because of the range of pore sizes in
membranes, the distinction between porous and nonporous membranes is
not always obvious.
The distinction can be made based only on the relative permeabilities for
diffusion through pores and diffusion through the solid, amorphous regions
of the membrane, respectively.

Mass Transfer Through Porous
Membranes
Mechanisms for transport of liquid
and gas molecules through a porous
membrane are depicted in Figures,
where flow is downward.

Transport Through Nonporous Membranes
Mass transfer through nonporous (dense) solid
membranes is the predominant mechanism in membrane
separators for reverse osmosis, gas permeation, and
pervaporation (liquid and vapor).
As indicated in Figure, gas or liquid species absorb at the
feed-side of the membrane, diffuse through the
membrane, and desorb at the permeate-side.
Liquid diffusivities are several orders of magnitude less
than gas diffusivities, and diffusivities of solutes in solids are
a few orders of magnitude less than diffusivities in liquids.

Concentration and partial pressure
profiles for solute transport through
membranes.
Liquid mixture with (a) a porous
and (b) a nonporous membrane
Gas mixture with (c) a porous and
(d) a nonporous membrane

Module Flow Patterns
The flow pattern can significantly affect the degree of separation and the
membrane area.
(a) Perfect mixing (b) Countercurrent flow (c) Cocurrent flow (d) Cross flow

Cascades
A single membrane module or a number of such modules arranged
in parallel or in series constitutes a single-stage membrane-
separation process.
The extent to which a feed mixture can be separated in a single stage
This factor depends, in turn, on module flow patterns; permeability
ratio (ideal separation factor); cut,
and the driving force for membrane mass transfer.

For example, the composition of retentate leaving stage
1 and entering stage 2 would be identical to the
composition of permeate flowing from stage 3 to stage
2.
This corresponds to the least amount of entropy
production for the cascade and, thus, the highest
second-law efficiency. Such a cascade is referred to as
Calculation methods for cascades, as discussed by
Hwang and Kammermeyer [30], utilize single-stage
methods that depend upon the module flow pattern, as
described in the previous section.

The calculations are best carried out on a computer but results for a
binary mixture can be conveniently displayed on a McCabe Thiele-
type diagram of the type used for distillation.
For a membrane cascade, the component mole fraction in the
permeate leaving each stage, yi, is plotted against the mole fraction
in the retentate leaving each stage, xi.
For a membrane cascade, the equilibrium curve becomes the
S.

In Figure 14.11, it is assumed that pressure drop on the feed or upstream
side of the membrane is negligible. Thus, only the permeate must be
pumped to the next stage if a liquid or compressed if a gas. In the case
of gas, compression costs are high.
Thus, membrane cascades for gas permeation are often limited to just
two or three stages, with the most common configurations shown in
Figures 14.11b, c, and d.

Compared to one stage, the two-stage stripping cascade is designed to
obtain a purer retentate, whereas a purer permeate is the goal of the two-
stage enriching cascade.
Addition of a pre-membrane stage, as shown in Figure 14.11d, may be
attractive when feed concentration is low in the component to be passed
preferentially through the membrane, desired permeate purity is high,
separation factor is low, and/or a high recovery of the more permeable
component is desired.

Concentration Polarization and Fouling
Gases produced during electrolysis accumulate on and around the
electrodes of the electrolytic cell, reducing the flow of electric current.
This is referred to as polarization. A similar phenomenon, concentration
polarization, occurs in membrane separators when the membrane is
permeable to A, but relatively impermeable to B.
Thus, molecules of B are carried by bulk flow to the upstream surface of
the membrane, where they accumulate, causing their concentration at
the surface of the membrane to increase in a polarization layer.

The equilibrium concentration of B in this layer is reached when its back-diffusion
to the bulk fluid on the feed-retentate side equals its bulk flow toward the
membrane.
Concentration polarization is most common in pressure driven membrane
separations involving liquid mixtures, such as reverse osmosis and ultrafiltration,
where it reduces the flux of A.
The polarization effect can be serious if the concentration of B reaches its solubility
limit on the membrane surface.
Then, a precipitate of gel may form, causing fouling on the membrane surface or
within membrane pores, resulting in a further reduction in the flux of A.
Concentration polarization and fouling are most severe at high fluxes of A.

Membrane Separation Processes

Membrane Process
The heart of the membrane process is the membrane itself. A membrane is an
ultra-thin semipermeable barrier separating two fluids that permits the transport
of certain species through the barrier from one fluid to the other.
The membrane is typically made from various polymers, but ceramic and metallic
membranes are also used in some applications.
The membrane is selective since it permits the transport of certain species while
rejecting others.The term semipermeable is frequently used to describe this
selective action.

Membrane Processes
Reverse osmosis uses non-porous membranes and can separate down to the
ionic level as with the example of seawater in the rejection of dissolved salt.
Nanofiltration performs separations at the nanometer range.
Ultrafiltration uses porous membranes and separates components of
molecular weight ranging from the low thousand to several hundred thousand
molecules; an example includes components of biochemical processing.
Microfiltration uses much more porous membranes and is typically used in the
macromolecular range to remove particulate or larger biological matter from
a feed stream.

Membrane Processes
Dialysis membrane processes use a concentration driving force for separation of
liquid feeds across a semipermeable membrane with the major application in
the medical field of hemodialysis.
Electrodialysis separates a liquid feed solution through ion selective membranes
by means of an electrical driving force and is widely used in water purification
and industrial processing.
Gas separation processes can be divided into two categories gas permeation
through non-porous membranes and gas diffusion through porous membranes.
Both of these processes utilize a concentration driving force.

Reverse Osmosis
Reverse osmosis is an advanced separation technique that may be used
whenever low molecular weight solutes such as inorganic salts or small organic
molecules (e.g., glucose) are to be separated from a solvent (usually water).
Reverse osmosis is widely utilized today by a host of industries for a surprising
number of operations.
Reverse osmosis processes are classified into the following two basic categories:
1. Purification of a solvent such as in desalination where the permeate or purified
water is the product.
2. Concentration of the solute such as in concentration of fruit juices where the
retentate is the product.

Reverse Osmosis
Reverse osmosis membranes may be configured into certain geometries for system
operation:
In the plate and frame configuration, flat sheets of membrane are placed between
spacers with heights of approximately 0.5 1.0 mm.These are, in turn, stacked in parallel
groups.
Tubular units are also used for RO.This is a simpler design in which the feed flows inside
of a tube whose walls contain the membrane.
There is also the hollow fine fiber (HFF) arrangement.This geometry is used in 70% of
worldwide desalination applications.
A spiral wound cartridge is occasionally employed. In this configuration, the solvent is
forced inward towards the product tube while the concentrate remains in the spacing
between the membranes.
A flat film membrane is made into a

Seawater desalination by RO
A simple material balance can be written
on the overall process flows and for that of
the solute:
Subscripts f, r, and p refer the feed,
retentate, and permeate, respectively.
Verify that the quantities provided in Figure
satisfy both a componential and overall
material balance.

Reverse Osmosis
Before osmosis
equilibrium
Osmosis of
solvent
Osmotic pressure
Reverse osmosis

Reverse Osmosis
Process Flow Diagram

Reverse Osmosis
Osmoticpressureisrelatedtoboththesoluteconcentrationandthetemperature
ofthesolutionasdescribedinthe Hoffequationbelow:

Reverse Osmosis
Thechangeinosmoticpressureacrossthemembraneinthisoperation
mustbeovercomeinordertoachieveRO.Thisisshownby
Thischangeinosmoticpressurecanalsobefoundusingtheconcentrations
ofboththefeedandthepermeate,aswellasacoefficientdenotedas
Thisformulaisshownby

Reverse Osmosis
ThepermeatefluxisanimportantcharacteristicoftheROoperation.Itis
relatedtothepermeateflowaswellastheareaofthemembrane.Thiscan
beseeninthefollowingequation

Reverse Osmosis
Thefluxcanbedeterminedbymeasuringeachincrementalvolumeofpermeate,
collectedintimeperiod anddividingbythesurfaceareaofthemembrane.Inwater-
basedprocessessuchasdesalination,thepermeateconsistsofmostlywater.Therefore,
thepermeatefluxcanbeconsideredtobeequaltothewaterflux.Equationdefinesthe
waterflux:

Reverse Osmosis
Another important factor is the solute flux.This can be determined through the utilization of:
The solute flux can also be related to the solute concentration by utilizing the solute permeability
factor.This relationship is provided by

Reverse Osmosis
The selectivity of a membrane to filter out a solute can be expressed as the percent rejection (%R).
ability to selectively allow certain species to permeate and others to be rejected. This is an
important characteristic when selecting a membrane for a separation process.The percent
rejection represents the percentage of solute that was not allowed to pass into the permeate
stream, and is given

Reverse Osmosis
Finally, the solvent recovery,Y, is a measure of how much solvent is allowed to pass through the
membrane.This is defined as the quotient of the permeate flow divided by the feed flow, as shown
by

Reverse Osmosis
Note the following two basic membrane transport equations.
The driving force can be a pressure, concentration, or electric field.The flux, J, may also be written
as

Example:
With reference to figure for sea water desalination,
a. Calculate the solvent flux and membrane selectivity.
b. If the applied pressure gradient across the membrane is 500 psi and the
membrane thickness is 10 mm, determine the permeability of the
membrane in m2 /s . Pa.

Solution:
a. For the flux
b. Employing equation

Example
A new membrane material is to be evaluated for its solute and solvent
permeability. A small test cell is utilized with a 5.0 cm diameter circular
membrane. The test solution is 6000 mg/L of NaCl in water at 250C.
Assume that the following relationship holds for the osmotic pressure of
NaCl in water (0.0114 psi/mg/L).
At an operating pressure gradient of 750 psi, the permeate flow rate is
0.0152 cm3 /s and the permeate solute concentration is 150 mg/L.
Assuming there is no concentration polarization and that the operating
conditions remain constant.
a. Determine the water flux in g/cm2 . s and the solute flux in g/cm2 . s.
b. Calculate the percent solute rejection.

Solution:
a.
The membrane surface area is
b.

Ultrafiltration
Ultrafiltration (UF) is a membrane separation process that can be used to concentrate
single solutes or mixtures of solutes.
Trans-membrane pressure is the main driving force in UF operations and separation is
Ultrafiltration may be thought of as a membrane separation technique where a solution
is introduced on one side of a membrane barrier while water, salts, and/or other low
molecular weight materials pass through the unit under an applied pressure.
Membrane separation processes can be used to concentrate single solutes or mixtures
of solutes.
The variety in the different membrane materials makes a wide temperature/pH
processing range possible.
The main economic advantage of UF is a reduction in design complexity and energy
usage.

Ultrafiltration
The general design factors for any membrane system (including UF) are reported as:
1. Thin active layer of membrane
2. High permeability for species A and low permeability for species B
3. Stable membrane with long service life
4. Mechanical strength
5. Large surface area of membrane in a small volume
6. Concentration polarization elimination or control
7. Ease in cleaning, if necessary
8. Inexpensive to build
9. Low operating costs

Ultrafiltration
System performance is usually defined in terms of permeate flux, Jp, with dimensions
of volume/area . time.The typical units are L/m2 . h. As with RO, Jp can be obtained
experimentally by measuring the incremental volume of the permeate, collected
in a time period, Thus, the permeate flux describing equation is

Ultrafiltration
Trans-membrane pressure is the main driving force in UF operations, and separation
is achieved through the aforementioned sieving mechanism. Since UF is a pressure-
driven separation process, it is appropriate to examine the effects of pressure on flux.
Equation shows how the flux varies with pressure.

Ultrafiltration
When a solute such as milk solids dissolved in water flows through a typical UF
process, some of the solute usually passes through the membrane since real
membranes are partially permeable.The apparent rejection on a fraction basis is then
once again calculated as follows:

Ultrafiltration
Concentration polarization occurs in many separations, and for large solutes where
osmotic pressure can be neglected, concentration polarization without gelling is
predicted to have no effect on the flux.Therefore, if a flux decline is observed, it can
be attributed to the formation of a gel layer with a concentration Cg.The gel layer,
once formed, usually controls mass transfer.When this happens, Equation can be
used to determine the solvent flux:

Ultrafiltration
When the gel layer controls mass transfer and Cp = 0 or the apparent rejection is
unity, the solvent flux can be expressed in terms of a mass transfer coefficient, k, as
follows:

Ultrafiltration
Empirical equations can be employed in the determination of the mass transfer
coefficient, k. Fully developed turbulent flow in UF devices appears to occur at Re
greater and equal to 2000.The Reynolds number can be calculated using an
equivalent diameter as follows:

Ultrafiltration
The following equation can be used to determine the mass transfer coefficient for
turbulent flow through a channel:
For laminar flow through a channel, the average mass transfer coefficient can be
estimated using the following equation:

Example
During a 12 psi UF run with pure water, the incremental volume of water collected for a
580 s time interval was 50 cm3 . If the effective surface area is 40 cm2 , calculate the
permeate flux.
Solution:

Example
The average concentration of the retentate for a UF run is 1.117 g/cm3 . If a 19 cm3
permeate sample is placed on a 1.0534 g tray and dried to a weight of 1.1454 g,
calculate the apparent rejection.

Calculate the mass transfer coefficient.The length of the channel is 41.4 cm.
Example
The mass transfer coefficient can be estimated from empirical equations. The
empirical equation used depends on whether the flow is laminar or turbulent. The
Reynolds number needs to be calculated to determine the type of flow. Consider the
following UF bench scale system and operating conditions. The height and width of
the channel available for flow are 0.038 and 0.95 cm, respectively. The average
been estimated to be 1.013 g/cm3 and 0.020 g/cm . s, respectively.
a. Calculate the Reynolds number
b. Calculate the mass transfer coefficient. The length of the channel is 41.4 cm.

b. Solution
The flow is laminar. For a laminar flow through a channel, the mass transfer
coefficient can be calculated using Equation

Microfiltration
Microfiltration (MF) is employed in modern biochemical and biological separation
processes.
For example, in cell harvesting, microfiltration can be used instead of centrifugation or pre-
coat rotary vacuum filtration to remove yeast, bacteria, or mycelial organisms from
fermentation broth.
Both MF and UF are used for cell harvesting. Microfiltration is used to retain cells and
colloids, while allowing passage of macromolecules into the permeate stream.
Ultrafiltration is used to concentrate macromolecules, cells, and colloidal material, while
allowing small organic molecules and inorganic salts to pass into the permeate stream.
Pore sizes in microfiltration are around 0.02 10 mm in diameter as compared with 0.001
0.02 mm (300 300,000 Daltons) for ultrafiltration (ranges vary slightly depending on the
source).
Ideal membranes possess high porosity, a narrow pore size distribution, and a low binding
capacity. When separating microorganisms and cell debris from fermentation broth, a
biological cake is formed.

Microfiltration
Factors affecting system performance
OperatingTemperature
AverageTransmembrane Pressure (ATP)
Yeast Concentration in the Feed
pH
Feed Preparation

Microfiltration
Example
The data corresponding to a pressure run of 1.5 psi in a MF resulted
in a flux for the pure water system of 196.7 mL/m2 . s.
a. Calculate the membrane hydrodynamic resistance (Rm).
b. Calculate the cake resistance if fouling is neglected. Assume the
steady-state (or limiting) flux value to be approximately half the
value calculated.

Microfiltration
Solution:
a. When

Microfiltration
Solution:
b. The cake resistance is calculated using the following formula:

Microfiltration
Example
Data for a yeast run in a MF system yielded the following
concentration volume data over a 5 min sampling period:
Cr = final concentration of yeast cells in retentate = 0.5 g/L
Co = initial cell concentration in feed = 1.2 g/L
Vr = final retentate volume = 290 L
Vo = initial feed volume = 150 L
a. Calculate the recovery.
b. Calculate the solute rejection, Ro , if the concentration of the yeast
cells in the permeate is 0.10 g/L.

Microfiltration
Solution:
a. The recovery of the system is calculated using the following formula:
b. The solute rejection is calculated using the following formula:

Gas Permeation
Several different types of membrane separation processes are used in the chemical
process industries, including systems for gas separation.
These processes are generally considered as new and emerging technologies because
they are not included in the traditional chemical engineering curriculum.
Gas permeation systems have and continue to gain popularity in both traditional and
emerging engineering areas.
These systems were originally developed for hydrogen recovery.There are presently
numerous applications of gas permeation in industry and other potential uses of this
technology are in various stages of development.
Applications include gas recovery from waste gas streams, landfill gases, and
ammonia and petrochemical products. Gas permeation membrane systems are also
employed in gas generation and purification, including the production of nitrogen and
enriched oxygen gases.

Mass Transfer Through Porous Membranes
Mechanisms for transport of liquid and gas
molecules through a porous membrane are
depicted in Figures, where flow is downward.
Transport Through Nonporous Membranes

Protein Separation

ProteinSeparation
Protein bioseparation which refers to the recovery and purification of protein products
from various biological feed streams is an important unit operation in the food,
pharmaceutical and biotechnological industry.
For the purpose of simplicity, these industries will be collectively referred to as
bioprocess industries.
Protein bioseparation is at the present moment more important in the bioprocess
industry than at any time before.This is largely due to the phenomenal developments in
recent years in the field of modern biotechnology.
More and more protein products have to be purified in larger quantities. A further boost
to protein bioseparation is likely to come from the developing science of proteomics.

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