GENIE ENZYMATIQUE

IMMOBILIZATION DES ENZYMES

ENZYME IMMOBILIZATION
• What is enzyme immobilization and Why immobilize enzymes ?
• Before giving answers to these questions, let’s first of all review what enzyme
are.
• Enzymes are protein molecules which serve to accelerate the chemical reactions of living
cells (often by several orders of magnitude).
• Without enzymes, most biochemical reactions would be too slow to even carry out life
processes. Enzymes display great specificity and are not permanently modified by their
participation in reactions.
A review of what enzyme are
• Since they are not changed during the reactions, it is cost-effective to
use them more than once. However, if the enzymes are in solution
with the reactants and/or products it is difficult to separate them.
• Therefore, if they can be attached to the reactor in some way, they
can be used again after the products have been removed.
• That is Why enzymes are immobilized.
What is an immobilized enzyme?
• The term "immobilized" means unable to move or stationary. And
that is exactly what an immobilized enzyme is: an enzyme that is
physically attached to a solid support over which a substrate is
passed and converted to product.
What are therefore the advantages of
immobilizing enzymes?
• There are a number of advantages to attaching
enzymes to a solid support and a few of the major
reasons are listed below:
• Multiple or repetitive use of a single batch of enzymes
• The ability to stop the reaction rapidly by removing the enzyme
from the reaction solution (or vice versa)
• Enzymes are usually stabilized by bounding
• Product is not contaminated with the enzyme (especially useful in
the food and pharmaceutical industries)
• Analytical purposes - long 1/2-life, predictable decay rates,
elimination of reagent preparation, etc
What are the factors which will affect the rate of
immobilized enzymes?
• It is important to understand the changes in physical and chemical
properties which an enzyme would be expected to undergo upon
immobilization (sometimes referred to as insolubilization). There are
a number of factors that affect the rate of the enzyme’s catalytic
activities.
• An enzymatic reaction is the conversion of one molecule into another;
a chemical reaction catalyzed at the reactive sites on the enzyme.
Considering the complex nature of the enzyme itself, it is not
unreasonable to expect that many parameters will affect the rate of this
catalytic activity. Enzyme activity can be influenced by:
Which are the factors which will affect the rate of
immobilized enzymes?
• Spacing (steric hindrance)
• pH
• Temperature
• Substrate Concentration (Michaelis-Menten Kinetics)
Spacing (Steric hinderance)
• Spacing
Any groups that separate the enzyme from the support (or
backbone) are referred to as spacing groups. For an enzyme only one
spacing group away, it would be very difficult for a substrate to find
the active site. The backbone interferes sterically. But with more than
one CH2 (or other spacing groups), the enzyme can whip around and
twist so that the active site is much more accessible. Usually, spacers
that provide as much distance as six CH 2 groups are enough.
Effect of pH Change
• Effect of pH Change
Since enzymes are proteins, they are very sensitive to
changes in pH. Each enzyme has its own optimum range for
pH where it will be most active. pH changes could be due to a
combination of factors:
(1) the binding of the enzyme to substrate,
(2) the catalytic activity of the enzyme,
(3) the ionization of the substrate, and
(4) the variation of protein structure.
The initial rates for many enzymatic reactions exhibit bell
shaped curves as a function of pH as shown in the example
below. (Note that this particular enzyme is most active at a pH
of zero, but this is not the case for all.)
Effect of pH Change
Effect of Temperature Change

• Effect of Temperature Change

As temperature increases, the rate of reaction also increases, as is
observed in many chemical reactions. However, the stability of the
protein also decreases due to thermal degradation. Holding the
enzyme at a high enough temperature for a long period of time may
cook the enzyme.
Effect of Substrate Concentration
• Enzymes are not passive surfaces on which reactions take place but
rather, are complex molecular machines that operate through a great
diversity of chemical mechanisms. According to Michaelis-Menten kinetics,
enzyme-substrate reactions are actually comprised of two elementary
reactions. The first is the when the substrate forms a complex with the
enzyme and then in the second, the complex decomposes to product and
enzyme.
• k1 k2
Enzyme + Substrate <----> Complex ----> Products + Enzyme
k-1

• According to this model, when the substrate concentration becomes

high enough to entirely convert all of the enzyme to the complex form, the
second step of the reaction becomes the rate-limiting step. Therefore, the
overall conversion to product becomes insensitive to further increases in
substrate concentration. The general expression for the rate of this
reaction (velocity) becomes:
• v = d[P]/dt = k2*[complex]
Stability of immobilized enzyme

• Stability
• The stability of the enzymes can be expected to
change upon immobilization. It will increase if the
carrier provides a micro environment capable of
stabilizing the enzyme and decrease if the carrier is
capable of denaturing the enzymatic protein. It has
been found that enzymes coupled with inorganic
carriers are generally more stable than those attached
to organic polymers.
• Stability with respect to denaturing agents (agents that
will affect the stability of proteins) may also be changed
upon immobilization.
Methods of Immobilization

• What guides in the choice of method for enzyme immobilization?

• When immobilizing an enzyme to a surface, it is most important to;
• choose a method of attachment that will prevent loss of enzyme activity by not
changing the chemical nature or reactive groups in the binding site of the enzyme. In
other words, attach the enzyme but do as little damage as possible.
What guides in the choice of method for enzyme immobilization?

• The surface on which the enzyme is immobilized is

responsible for retaining the structure in the enzyme
through hydrogen bonding or the formation of electron
transition complexes. These links will prevent vibration of
the enzyme and thus increase thermal stability.
• The micro environment of surface and enzyme has a
charged nature that can cause a shift in the optimum pH
of the enzyme of up to 2 pH units. This may be
accompanied by a general broadening of the pH region in
which the enzyme can work effectively, allowing enzymes
that normally do not have similar pH regions to work
together.
Carrier-Binding Method of immobilization
• Carrier-Binding (the binding of enzymes to water-insoluble carriers )
The carrier-binding method is the oldest immobilization technique
for enzymes. In this method, the amount of enzyme bound to the
carrier and the activity after immobilization depend on the nature of
the carrier. The following picture shows how the enzyme is bound to
the carrier:
Carrier-Binding Method
On what depends the selection of the
Carrier?
• The selection of the carrier depends on:
• the nature of the enzyme itself, as well as the
• Particle size
• Surface area
• Molar ratio of hydrophilic to hydrophobic groups
• Chemical composition
Carriers
• In general,
• an increase in the ratio of hydrophilic groups and in the
concentration of bound enzymes, results in a higher activity
of the immobilized enzymes.
• Some of the most commonly used carriers for enzyme
immobilization are polysaccharide derivatives such as:
• cellulose,
• dextran,
• agarose, and
• polyacrylamide gel
Sub-classification of Carrier-binding method

• According to the binding mode of the enzyme, the carrier-binding

method can be further sub-classified into:
• Physical Adsorption
• Ionic Binding
• Covalent Binding
 Physical Adsorption

• This method for the immobilization of an enzyme is based on the

physical adsorption of enzyme protein on the surface of water-
insoluble carriers.
• Hence, the method causes little or no conformational change of the
enzyme or destruction of its active center.
• If a suitable carrier is found, this method can be both simple and
cheap.
• However, it has the disadvantage that the adsorbed enzyme may
leak from the carrier during use due to a weak binding force
between the enzyme and the carrier.
Physical Adsorption
• The earliest example of enzyme immobilization using
this method is the adsorption of beta-D-fructo-
furanosidase onto aluminum hydroxide.
• The processes available for physical adsorption of
enzymes are:
• Static Procedure
• Electro-deposition
• Reactor Loading Process
• Mixing or Shaking Bath Loading
Of the four techniques, the most frequently used in the lab is
Mixing-Bath Loading . For commercial purposes the
preferred method is Reactor Loading .
Physical Adsorption
• Advantages
• A major advantage of adsorption as a general
method of immobilizing enzymes is that usually no
reagents and only a minimum of activation steps
are required.
• Adsorption tends to be less disruptive to the
enzymatic protein than chemical means of
attachment because the binding is mainly by
hydrogen bonds, multiple salt linkages, and Van
der Waal's forces. In this respect, the method
bears the greatest similarity to the situation found
in natural biological membranes and has been
used to model such systems.
Physical Adsorption
• Disadvantages
• Because of the weak bonds involved, desorption of
the protein resulting from changes in temperature,
pH, ionic strength or even the mere presence of
substrate, is often observed.
• Another disadvantage is non-specific, further
adsorption of other proteins or other substances as
the immobilized enzyme is used. This may alter the
properties of the immobilized enzyme or, if the
substance adsorbed is a substrate for the enzyme, the
rate will probably decrease depending on the surface
mobility of enzyme and substrate.
 Ionic Binding

• The ionic binding method relies on the ionic binding of the enzyme
protein to water-insoluble carriers containing ion-exchange residues.
• Polysaccharides and synthetic polymers having ion-exchange
centers are usually used as carriers.
Ionic Binding
• Advantages
• The binding of an enzyme to the carrier is easily
carried out, and the conditions are much milder than
those needed for the covalent binding method.
• Hence, the ionic binding method causes little changes in
the conformation and the active site of the enzyme.
• Therefore, this method yields immobilized enzymes with
high activity in most cases.
 Ionic Binding

• Disadvantages
- Leakage of enzymes from the carrier may occur in substrate
solutions of high ionic strength or upon variation of pH.
This is because the binding forces between enzyme proteins and
carriers are weaker than those in covalent binding.
Ionic Binding
• What is the main difference between ionic and physical adsorption
binding modes?
• The main difference between ionic binding and physical adsorption is
that:
• the enzyme to carrier linkages are much stronger for ionic binding although
weaker than in covalent binding .
Covalent Binding
• The most intensely studied of the immobilization
techniques is the formation of covalent bonds
between the enzyme and the support matrix.
• When trying to select the type of reaction by
which a given protein should be immobilized,
the choice is limited by two characteristics:
(1) the binding reaction must be performed
under conditions that do not cause loss of
enzymatic activity, and
(2) the active site of the enzyme must be
unaffected by the reagents used.
Covalent Binding
• The covalent binding method is based on:
• the binding of enzymes and water-insoluble carriers by covalent
bonds.
The functional groups that may take part in this
binding are listed below:
• Amino group Carboxyl group Sulfhydryl group Hydroxyl
group Imidazole group Phenolic group
Thiol group Threonine group Indole group
Covalent Binding
• This method can be further classified into :
• diazo,
• peptide and
• alkylation methods according to the mode of linkage.
Disadvantage
The conditions for immobilization by covalent binding are
much more complicated and less mild than in the cases of
physical adsorption and ionic binding.
Therefore, covalent binding may alter the conformational
structure and active center of the enzyme, resulting in
major loss of activity and/or changes of the substrate.
Covalent Binding
• Advantage
• The binding force between enzyme and carrier is so strong that no leakage of
the enzymes occurs, even in the presence of substrate or solution of high ionic
strength.
Covalent Binding
• Protective methods of active sites:
• Covalent attachment to a support matrix must involve
only functional groups of the enzyme that are not
essential for catalytic action.
• Higher activities result from prevention of inactivation
reactions with amino acid residues of the active sites.
A number of protective methods have been devised:
• Covalent attachment of the enzyme in the presence of:
• a competitive inhibitor or substrate.
• A reversible, covalently linked enzyme-inhibitor
complex.
• A chemically modified soluble enzyme whose covalent
linkage to the matrix is achieved by newly incorporated
residues.
• A zymogen precursor.
Effective Covalent Binding
• Hence, covalent binding can be brought about by the following:

• Diazotization :
• Amide bond formation :
• Alkylation and Arylation:
• Schiff's base formation :
• Amidation reaction :
• Thiol-Disulfide interchange :
SUPPORT--N=N--ENZYME.
SUPPORT--CO-NH--ENZYME
SUPPORT--CH2-NH-ENZYME
SUPPORT--CH2-S--ENZYME
SUPPORT--CH=N--ENZYME
SUPPORT--CNH-NH--ENZYME
SUPPORT--S-S--ENZYME

• UGI reaction
• Mercury-Enzyme interchange
• Gamma-Irradiation induced coupling
• Carrier binding with bifunctional reagents :
• SUPPORT-O(CH2)2 N=CH(CH2)3 CH=N-ENZYME
• The active site of the enzyme must not be hindered. There must be ample space
Ugi reaction
• The Ugi reaction is a multi-component reaction in organic chemistry
involving a ketone or aldehyde, an amine, an isocyanide and a
carboxylic acid to form a bis-amide. The reaction is named after Ivar
Karl Ugi, who first published this reaction in 1962.
Increasing the yield of the immobilized enzyme
• It is possible in some cases to increase the number of
reactive residues of an enzyme in order to increase
the yield of the immobilized enzyme.
• This provides alternative reaction sites to those
essential for enzymatic activity.
• Similiarity between covalent bonding and
cross-linking
• provide stable, immobilized enzyme derivatives that do
not leach enzyme into the surrounding solution.
• The wide variety of binding reactions and insoluble
carriers (with functional groups capable of covalent
coupling or being activated to give such groups) makes this
a generally applicable method of immobilization. This is
true even if very little is known about the protein structure
or active site of the enzyme to be coupled.
Cross-Linking

• The Cross-linking method of enzyme immobilization

entails:
• The intermolecular cross-linking of enzymes by bi-
functional or multi-functional reagents.
Is the cross-linking of an enzyme to itself
of any use?
• NO!
• Cross-linking an enzyme to itself is both :
• expensive and insufficient, as some of the
protein material will inevitably be acting
mainly as a support.
• This will result in relatively low enzymatic
activity.
• Generally, cross-linking is best used in
conjunction with one of the other methods.
• It is used mostly as a means of stabilizing
adsorbed enzymes and also for preventing
leakage from polyacrylamide gels.
• Advantage of covalent-linking in cross-linkage
immobilization
• Since the enzyme is covalently linked to the support
matrix, very little desorption is likely using this
method.
The most common reagent used for cross-linking
is glutaraldehyde.

• Disadvantage of Cross-Linking
• Cross-linking reactions are carried out under
relatively severe conditions. These harsh conditions
can change the conformation of active center of the
enzyme; and so may lead to significant loss of
activity.
Enzyme Immobilization by Entrapment

• Entrapping Enzymes:
• incorporating enzymes into the lattices of a semi-permeable gel or enclosing
the enzymes in a semi-permeable polymer membrane.
• It is done in such a way as to retain protein while allowing penetration of
substrate. It can be classified into lattice and micro capsule types.
Enzyme Immobilization by Entrapment
Enzyme Immobilization by Entrapment

• How does it differ from the covalent binding and cross linking
methods of immobilization?
• This method differs from the covalent binding and cross linking in that the
enzyme itself does not bind to the gel matrix or membrane. This results in a
wide applicability.
• The conditions used in the chemical polymerization reaction are relatively
severe and result in the loss of enzyme activity. Therefore, careful selection of
the most suitable conditions for the immobilization of various enzymes is
required.
Lattice-Type entrapment
• Lattice-Type entrapment:
- involves entrapping enzymes within the interstitial spaces of
a cross-linked water-insoluble polymer.
- Some synthetic polymers such as polyarylamide,
polyvinylalcohol, etc... and natural polymer (starch) have been used
to immobilize enzymes using this technique.
Microcapsule-Type entrapping
• Microcapsule-Type entrapment:
• involves enclosing the enzymes within semi permeable polymer membranes.
• The preparation of enzyme micro capsules requires extremely well-controlled
conditions and the procedures for micro capsulation of enzymes can be
classified as:
• Interfacial Polymerization Method
• Liquid Drying
• Phase Separation
Microcapsule-Type entrapping
• Interfacial Polymerization Method
• In this procedure, enzymes are enclosed in semi permeable membranes of
polymers.
• An aqueous mixture of the enzyme and hydrophilic monomer are emulsified in a water-
immiscible organic solvent.
• Then the same hydrophilic monomer is added to the organic solvent by stirring.
Polymerization of the monomers then occurs at the interface between the aqueous and
organic solvent phases in the emulsion. The result is that the enzyme in the aqueous
phase is enclosed in a membrane of polymer.
Microcapsule-Type entrapping

• Liquid Drying:
• In this process, a polymer is dissolved in a water-immiscible organic solvent
which has a boiling point lower than that of water.
• An aqueous solution of enzyme is dispersed in the organic phase to form a
first emulsion of water-in-oil type.
• The first emulsion containing aqueous micro droplets is then dispersed in an
aqueous phase containing protective colloidal substances such as gelatin,
and surfactants, and a secondary emulsion is prepared.
• The organic solvent in then removed by warming in vacuum. A polymer
membrane is thus produced to give enzyme micro capsules.
Microcapsule-Type entrapping
• Phase Separation:
• One purification method for polymers involves dissolving the polymer in an
organic solvent and re-precipitating it. This is accomplished by adding
another organic solvent which is miscible with the first, but which does not
dissolve the polymer.
Form of an immobilized enzyme
• The form of an immobilized enzyme can be
classified into four types:
• particles,
• membranes,
• Tubes, and
• filters.
Most immobilized enzymes are in particle form
for ease of handling and ease of application.
Form of an immobilized enzyme
• Particles - The particle form is described in the above
section.
• Membranes - Enzyme membranes can be prepared by
attaching enzymes to membrane-type carriers, or by
molding into membrane form. The molding is done after
the enzymes have been enclosed within semi-permeate
membranes of polymer by entrapment.
• Tubes - Enzyme tubes are produced using Nylon and
polyacrylamide tubes as carriers. The polymer tube is first
treated in a series of chemical reactions and the enzyme is
bound by diazo coupling to give a tube in a final step.
• Fibers - Enzymes that have been immobilized by
entrapment in fibers to form enzyme fibers.
Solid Supports
• The solid supports used for enzyme immobilization can be:
• inorganic or
• organic .
Some organic supports include:
Polysaccharides, Proteins, Carbon,
Polystyrenes, Polyacrylates,
Maleic Anhydride based Copolymers,
Polypeptides, Vinyl and Allyl Polymers, and Polyamides.
Choice of Reactor for Enzyme immobilization

• Types of Reactors
• In an enzyme reactor, the highest specific enzyme activity
is desirable. It is considered an added bonus if the support
that is used also aides in separation.
• One approach is to use a molecular sieve as the support and pulse
the reactor bed with the alternating passage of substrate
solution and water. The result is that bands of unused substrate
and product progress down the column. It so happens that the
enzymes for which this technique would be useful are also those
which in some cases benefit in having the enzyme immobilized on
a porous support.
Case of an industrial reactor
• For an industrial reactor,
• it is preferable to use supports that are non-biodegradable
such as:
• glass,
• silica,
• Celite,
• Bentonite,
• alumina, or
• titanium oxide, if possible.
Even the linkages between enzyme and support can be non-
biodegradable, as they are in the case of titanium.
Case of an industrial reactor
• In some of the of the above supports,
• the physical nature of the surface becomes a major problem. Thus, some
supports that form excellent packed beds fail to do so when coated with
enzyme. Particles which ideally self-suspend in a fluid bed may form
aggregates during use which will require more power to pump through
substrate. Many problems were encountered using porous glass supports
until someone realized that the glass itself could dissolve. This problem has
been eliminated by treatment of the glass surface with zirconium.
Stirred Tank Reactors
A batch stirred tank reactor :
• is the simplest type of reactor. It is composed of a reactor
and a mixer such as a stirrer, a turbine wing or a propeller.
The batch stirred tank reactor is illustrated below:

•
Limits of a batch stirred tank reactor
• This reactor is useful for substrate solutions of high
viscosity and for immobilized enzymes with relatively
low activity.
• However, a problem that arises is that an
immobilized enzyme tends to decompose upon
physical stirring.
• The batch system is generally suitable for the
production of rather small amounts of chemicals.
A continuous stirred tank reactor
• A continuous stirred tank reactor is shown below. It is
more efficient than a batch stirred tank reactor but
the equipment is slightly more complicated.
Packed Bed Reactors

• Packed Bed Reactors

• Continuous packed bed reactors are the most
widely used reactors for immobilized enzymes and
immobilized microbial cells.
• In these systems, it is necessary to consider:
• the pressure drop across the packed bed or column, and
• the effect of the column dimensions on the reaction rate.
There are three substrate flow possibilities in a packed bed and they
are illustrated below:
• Downward flow method
• Upward flow method
• Recycling method
Packed Bed Reactors
Packed Bed Reactors
• The recycling method is advantageous when the
linear velocity of the substrate solution affects the
reaction flow rate. This is because the recycling
method allows the substrate solution to be passed
through the column at a desired velocity.
• For industrial applications, upward flow is
generally preferred over downward flow because it
does not compress the beds in enzyme columns as
downward flow does. When gas is produced during
an enzyme reaction, upward flow is preferred.
Advantages continuous packed bed reactor over a batch
packed bed reactor
• A continuous packed bed reactor has the
following advantages over a batch packed bed
reactor:
• Easy, automatic control and operation
• Reduction of labor costs
• Stabilization of operating conditions
• Easy quality control of products
Types of Continuous reactors
• Continuous reactors may include:
• Stirred Tank Reactor with Filtration Recovery
• Stirred Tank Reactor with Settling Tank Recovery
• Stirred Tank Reactor with Immobilized Enzyme Basket
Paddles
• Stirred Tank Reactor with Ultra filtration Recovery
 Combined CSTR/UF Reactor

• A combined CSTR/UF reactor is a combination of a continuously

stirred tank reactor and an Ultra Filtration Unit.
• This type of reactor begins as a typical CSTR. However, the product
passes through an Ultra Filtration Unit where the enzyme is removed
and recycled back into the reactor.
• An example of what this combination rector can look like is shown
below:
A combined CSTR/UF reactor
A combined CSTR/UF
• In a combined CSTR/UF reactor :
• the enzymes are immobilized in that they can not leave the reactor because of the filtration
unit.
• This allows continuous processing with free enzymes in the CSTR.
• The Ultra Filtration Unit contains a membrane which provides a semipermeable
barrier which allows products and unreacted substrate, if there is any, to pass
through while holding back the enzyme.
• There are other possibilities for similar reactors, such as :
• combined reactor-separators. However, the combined CSTR/UF reactor has
proven useful for several types of reactions where a typical immobilized enzyme
would not be as effective. One example of this is for the conversion of
benzylpenicillin to 6-aminopenicillanic acid by penicillin amidase.
Recycle Reactor

• A recycle reactor:
• is a reactor that is not seen very often, but is very
important to consider when studying immobilized enzymes.
It is very important to chemical engineering because it
allows some substrates to be processed, which could not
be processed using other reactor types.
• An example of a recycle reactor can be seen below:
Recycle Reactor
Recycle Reactor
• In a recycle reactor,
• a portion of the product stream is recycled and mixed with the inlet flow to
the reactor. If the entire product stream is recycled back to the inlet stream,
then it is called a total recycle reactor.
• This can obviously only be used in a batch process, because if the entire
product stream is recycled back into the reactor in a continuous reactor, the
volume of the reactor would increase to infinity. Therefore, we will only
consider partial recycle streams in a continuous reactor on this page.
Recycle Reactor
• This type of rector
• is used when you have a substrate that cannot be
completely processed on a single pass, such as with an
insoluble substrate.
• These reactors continue to move the same substrate
through the reactor so that the effective contact time is
high enough to allow the substrate to be processed.
Recycle reactors also allow the reactor to operate at high
fluid velocities.
• This is important because it minimizes the bulk mass
transfer resistance to the transport of the substrate.
• It is important to remember that a recycle reactor is
simply a reactor, such as a CSTR or fluidized-bed reactor,
with a recycle stream.
Fluidized-Bed Reactor
• The fluidized bed reactor is:
• most suitable when a high viscosity substrate solution and a gaseous
substrate or product are used in a continuous reaction system. The figure
below illustrates a continuous fluidized bed reactor:
Fluidized-Bed Reactor
Fluidized-Bed Reactor
• In this system, care must be taken to avoid the destruction and
decomposition of immobilized enzymes. The particle size of
immobilized enzymes is an important factor for the formation of a
smooth fluidized bed.
Fluidized-bed reactor
• A fluidized-bed reactor is a combination of the two most common,
packed-bed and stirred tank, continuous flow reactors. It is very
important to chemical engineering because of its excellent heat and
mass transfer characteristics. The fluidized-bed reactor can be seen
below:
Fluidized-bed reactor
Fluidized-bed reactor
• In a fluidized-bed reactor,
• The substrate is passed upward through the immobilized
enzyme bed at a high enough velocity to lift the particles.
However, the velocity must not be so high that the
enzymes are swept away from the reactor entirely.
• This causes some mixing, more than the piston-flow model
in the packed-bed reactor, but complete mixing as in the
CSTR model. This type of reactor is ideal for highly
exothermic reactions because it eliminates local hot-spots,
due to its mass and heat transfer characteristics
mentioned before.
• It is most often applied in immobilized-enzyme catalysis
where viscous, particulate substrates are to be handled.
Membrane Reactor using hollow fibers
• Ultrafiltration Membrane Devices
• A continuous ultrafiltration membrane device is shown
below:
Ultrafiltration Membrane Devices

• This device is suitable for a substrate of high

molecular weight and a product of low molecular
weight. Since the enzyme used here is soluble, no
improvement in the stability of the enzyme can be
expected.
• A hollow fiber device can also be used and its
characteristics are essentially the same as those of an
ultrafiltration membrane.
Fluidised-bed reactors are
used in waste treatment
with sand or similar
material supporting mixed
microbial populations.
They are also used with
flocculating organisms in
brewing and for production
of vinegar.