The document discusses enzyme catalysis and immobilization. It begins by outlining applications of hydrolytic enzymes like proteases, carbohydrases, and esterases. These enzymes catalyze the hydrolysis of proteins, carbohydrates, fats, and polymers into simpler units. The document then examines methods of immobilizing enzymes, including carrier binding, cross-linking, and entrapment. Immobilized enzymes have benefits like reusability, easy product separation, and stability. The kinetics of immobilized enzymes are also discussed.

1. CHAPTER 4
ENZYME
CATALYSIS

2. ENZYME CATALYSIS
 Application of Hydrolytic Enzymes.
 Enzyme Immobilization Technology:
- Methods of enzyme immobilization.
- Immobilized enzyme kinetics.
SUBCHAPTER

3. INTRODUCTION

4. OBJECTIVE OF THIS CHAPTER:
 To study some of the applications
of enzymes.
 To examine immobilized enzyme
catalyst formulations (allow
sustained, continuous use of the
enzyme).

5. Some enzymes of industrial importance
Name Source Application Notes
Diastase Malt
Digestive aid,
supplement to bread,
syrup
α-amylase activity,
β-amylase activity
Takadiastase Aspergillus
oryzae
Digestive aid;
supplement to bread,
syrup
Contains many
other enzymes,
protease, R-Nase
Amylase Bacillus
subtilis
Desizing textiles,
syrup, alcohol
fermentation industry,
glucose production
Crude preparation
contains protease
Acid-resistant
amylase
Aspergillus
niger Digestive aid Optimum pH 4-5

6. Cont' ( starch-saccharifying amylase)
Name Source Application
Amyloglucosidase
Rhizopus niveus,
Aspergillus niger,
Endomycopsis
fibuliger
Glucose
production

7. Cont' ( animal and vegetable protease)
Name Source Application
Trypsin Animal pancreas Medical uses, meat tenderizers, beer
haze removal
Pepsin Animal stomach Digestive aid, meat tenderizer
α-
Chymotrypsin Animal stomach medical uses
Rennet Calf stomach cheese manufacture
Pancreas
protease Animal pancreas Digestive aid, cleaning, leatherbathing,
dehairing, feed improvement
Papain Papaya digestive aid, medical uses, beer
haze removal, meat tenderizer
Bromelain,
ficin PIneapple, fig digestive aid, medical uses, beer
haze removal, meat tenderizer

8. Cont' ( microbial protease)
Name Source Application Notes
Protease
Aspergillus oryzae Flavoring of sake, haze
removal in sake -
Aspergillus niger feed, digestive aid
acid resistant
protease, optimum
pH
Bacillus subtilis
detergents, removal of
gelatin from film
(recovery of silver), fish
solubles, meat tenderizer
optimum pH 7
Streptomyces
griseus
detergents, removal of
gelatin from film
(recovery of silver), fish
solubles, meat tenderizer
optimum pH 8
Varidase Streptococcus sp. medical use lederle
Streptokinase Streptococcus sp. medical use profibrinolysin

9. APPLICATION OF
HYDROLYTIC ENZYMES

10. Hydrolytic Enzymes ???
An enzymes that catalyzes hydrolysis.

11.  Protein
 Carbohydrate
 Fat molecules
 Polymers
Simplest
Units
Hydrolysis Process
Hydrolytic Enzymes ???
Hydrolytic enzymes

12. Hydrolytic Enzymes
Enzyme Substrate Hydrolysis product
Esterases:
Lipase Glycerides (fats) Glycerol + fatty acids
Phosphatases:
Lecithinase
Pectin esterase
.
Lecithin
Pectin methyl ester
.
Choline + H3PO4 + fat
methanol + polygalacturonic acid
Carbohydrases:
Fructosidase
α-glucosidase (maltase)
β-glucosidase (cellobiase)
β-galactosidase (lactase)
Amylases
Cellulase
Cytase
Polygalacturonase
.
Sucrose
Maltose
Cellobiose
Lactose
Starch
Cellulose
.
Polygalacturonic acid
.
fructose + glucose
glucose
glucose
galactose + glucose
maltose or glucose + maltooligosaccharides
cellobiose
simple sugars
galacturonic acid
Nitrogen-carryingcompounds:
Proteinases
Polypeptidases proteins
proteins
polypeptides
amino acids
Desaminases:
Urease
Asparaginase
Deaminase
urea
asparagine
amino acids
CO2 + NH3
Aspartic acid + NH3
NH3 + organic acids

13. Hydrolysis of Starch & Cellulose
Amylases
can hydrolyze the glucosidic bonds in
starch and related glucose-containing
compounds

14. Saccharifying
enzyme?
Hydrolysis of Starch & Cellulose
α-amylase
β-amylase
Starch-liquefying
enzyme

15. Industry Use
Glucose &
syrup
total or partially hydrolysis of corn starch to give a large quantity of
sweeteners.
Brewing conversion of crushed grain starch to maltose
Breadmaking leavening: conversion of sufficient starch to fermentable saccharides
needed for CO2 generation
Fruit juice hydrolysis of starch causing turbidity due to insolubility
Papermaking alpha-amylase action to liquefy starch coating to a desired viscosity
for application to fibers
Textiles
Sizing: alpha-amylase activity to liquefy starch
Desizing: alpha,beta-amylse action to remove size from woven
material so that all threads will dye uniformly and fabric will have
desired texture.
Candy production of candy of desired softness
Common applications of amylse
preparations

16. Proteolytic enzyme
Synthesized in an inactive form suitable either
for STORAGE or for TRANSPORT from the site
of synthesis to the desired site of activity
HOW to activate the proteolytic
enzyme?
From a precursor
By presence of a metal ion

17. OHRCOOHROHCOORR 2121 
Esterase Applications
To synthesize of various ester bonds to
yield an acid and an alcohol.

18. Hydrolytic enzyme applications which
are expanding (in development)
Enzyme Process
Penicillin
acylase
Production of semysynthetic penicillin
core from nature penicillin G
Lactase Removal of lactose from whey, milk
Ribonuclease Production of 5'-nucleotides from RNA
Dextranase Removal of tooth plaque
Isoamylase Production of maltose from starch
Keratinase Modification of wool, hair, leather
Tannase Removal of tannic acid from foods

19. ENZYME IMMOBILIZATION
TECHNOLOGY
 Methods of
enzyme
immobilization
 Immobilized
enzyme kinetics

20. Immobilized Enzymes?
Introduction

21. Enzyme Immobilization????
To restrict enzyme
mobility in a fixed space.

22. Enzyme Immobilization????
Immbolized enzymes are
ATTACHED
to an insoluble suppport medium or
enclosed by the support medium
which is also known as a CARRIER
In some cases, the enzymes molecules are
cross-linked to each other so that their
movement is restricted but their catalytic
activities are retained.

24. Needed for Immobilizaton
ECONOMICAL
CONVENIENCE
STABILITY
continuous use of bio-
catalyst is possible
separation of bio-catalyst &
product is much easier than
conventional batch process
Immobilized enzymes
typically have greater thermal
& operational stability than
the soluble form of the
enzyme

25. Methods for Enzyme
Immobilization

27. Methods for immobilization enzyme

28. Carrier-binding Method
 The oldest immobilization
method.
 Carriers (water insoluble)
such as polysaccharide
derivatives, synthetic
polymers, and porous glass,
etc.

29. Cross-linking Method
 Based on the formation of chemical bonds, but
water-insoluble carriers are not used in this method.
 Reagents such as glutaldehyde, bisdiazobenzidine,
and hexamethylene diisocyanate, etc.

30. Entrapping Method
 Based on confining enzymes in the lattice of a polymer matrix or enclosing
enzymes in semipermeable membranes.
 A chemical polymerization reaction e.g. cllagen, gelatin, cellulose etc.

31. Preparation &
Characteristics
of immobilized
enzyme

32. Preparation & Characteristics of
immobilized enzyme

33. Benefits of immobilizing and ENZYME
 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.

34. Immobilized Enzyme Kinetics