生化工程,第三章固定化酶反应动力学

§3.1 Advantages of Immobilized Enzymes
Advantages:
1. Enzyme reutilization: catalyst reuse;
2. Elimination of enzyme recovery and purification processes: product purity , while effluent handling problems ; 3. Provide a better environment for enzyme activity;
Problems:
1. Leakage of enzymes into solution: Reducing the MW cutoff of membranes or the pore size of solid matrices; 2. Considerable diffusional resistance emerges: Reducing the particle size of the matrices and/or capsules; 3. Reduction of enzyme activity and stability: Alter the unfavorable microenvironmental conditions; 4. Lack of control of the microenvironment:
Disadvantages: Desorption is quit common because the binding forces are so week, especially in the presence of strong hydrodynamic forces. Advantages: Normally, active site is not affected, nearly all activity is retained after immobilization.

Cross-Linking The enzyme molecules can also cross-link with each other. But this may cause significant changes in the active site of enzymes, and also severe diffusion limitations may result.
Surface of immobilized enzyme Substrate diffusion into permeable catalyst
CONCENTRATION PROFILE PARAMETERS
[P ]max
Bulk solution
[ Sb ]
[S ]min
Effective diffusion coefficient (DES, DEP)
生化工程
第三章 固定化酶反应动力学
ImБайду номын сангаасobilized Enzyme Systems
Definition
The restriction of enzyme mobility in a fixed space is known as enzyme immobilization. In general, immobilized enzymes are enzymes that are attached to, or entrapped within, a macroscopic support matrix so that the resulting catalyst can be reused. Immobilized enzyme Two opposite concept Free enzyme
Among 2000 types of enzymes found, only 50 ~ 60 have been applied in industry, most of which are extracellular enzymes.
§3.3 Methods of Immobilization
SUMMARY
The most suitable support material and immobilization method vary depending on the enzyme and particular application.
Then how to select support materials ?
A little difficult to handle.
Bound:

Adsorption A physical attachment of enzyme on the surfaces of support particles by weak physical forces: van der Waals. Support materials: Inorganic materials: alumina, silica, porous glass, ceramics, diatomaceous earth; Organic materials: cellulose (CMC, DEAE-cellulose), starch, activated carbon, ionexchange resin (Amberlite, Sephadex, Dowex).
Solid matrices: activated carbon, porous ceramic, diatomaceous earth.
When immobilized in a polymer matrix, enzyme solution is mixed with polymer solution before polymerization takes place.
How to select
method of immobilization ?
Table: Comparison of the Characteristics of Different Methods of Enzyme Immobilization
Carrier Binding Methods Characteristic Physical adsorption Easy Low Unchangeable Weak Possible Low Low Ionic Binding Easy High Unchangeable Moderate Possible Moderate Low Covalent Binding Difficult High Changeable Strong Impossible Moderate High Crosslinking Method Difficult Moderate Changeable Strong Impossible Low Moderate Entrapping Method Difficult High Unchangeable Strong Impossible High Low
Preparation Enzyme Activity Substrate Specificity Binding Force Regeneration General Application Cost of Immobilization
§3.4 Immobilized Enzyme Kinetics
[ Pb ]
Intrinsic kinetics (本征反应动力学) Bioreactor kinetics
Mass transfer of substrate and product
PHENOMENA
Bulk Fluid Stagnant Film or Boundary Layer Immobilized Enzyme Slab
MicroEncapsulated
Entrapment:
The physical enclosure of enzymes in a small space.

Matrix entrapment Matrices used for enzyme immobilization are usually: Ploymeric materials: Ca-alginate, agar, ĸ-carrageenin, polyacrylamide, collagen;
Disadvantages:
Increase the diffusion resistance, so decreases the reaction rate.
§3.2 Applications of Immobilized Enzymes
Immobilized enzyme are employed in many fields.

Membrane entrapment Membrane: nylon, cellulose, polysulfone, polyacrylate. In all cases, a semi-permeable membrane is used to retain high-MW compounds (enzyme) while allowing small-MW compounds (substrates or products) access to the enzyme. Microencapsulation: a special form of membrane entrapment. In this technique, microscopic hollow spheres are formed. The spheres contain the enzyme solution, while the sphere is enclosed with a porous membrane.
Immobilized Soluble Enzymes Entrapped MatrixEntrapped MembraneEntrapped Adsorbed (Physical or Ionic) Bound CovalentlyBound
To Support
To Enzyme
Between Macroscopic Membranes

Covalent Binding It is the retention of enzymes on support surfaces by covalent bond formation, via certain functional groups, such as amino, carboxyl, hydroxyl, and –SH group. The functional groups must not be the active sites.
a. Binding capacity, which is a function of charge density, functional groups, porosity, and hydrophobicity;
b. Stability and retention of the enzyme activity, which is a function of functional groups on support material and microenvironmental conditions.
Analytical devices:
For example: in immobilized enzyme electrodes for the detection of various chemical substances.
As catalyst in industrial processes: