Slide 1: Welcome….
Slide 2: NATIONAL COLLEGE OF PHARMACYDEPT. OF PHARMACOGNOSYSHIMOGA -577201
An evaluation Seminar on
Subject : Medicinal Plant Biotechnology
Topic: Enzyme Immobilization: Techniques and Application
Presented By: Neelanjan Chatterjee (I M.Pharm)Under the guidence of:Dr. G. Narayana Murthy
Department of Pharmacognosy NCP, Shimoga.2010
Slide 3: Enzymes: Enzymes are defined as soluble, colloidal, organic catalysts which are produced by living cells but are capable of acting independently of the cells.
In 1873 Berzelius recognized ferments catalyses chemical reactions
Later it was coined Enzymes (in side the yeast)
1897 Edword Buchner extracted enzymes from yeast that catalyzes sugars into alcohol
Slide 4: Immobilized Enzyme Systems:
Enzymes physically confined or localized in a certain defined region of space with retention of their catalytic activities and which can be used repeatedly and continuously
Retrospection:
In 1916 Nelson and Griffin discovered that Invertase enzyme showes same activity when absorbed on a solid media ( charcoal, aluminium hydroxide) at the bottom of the reaction vessel
Oreste .J. Lantero invented a method for the immobilization of an enzyme producing microorganism by introducing an aqueous solution containing bacterium cells into polyethyleneimine and adding gluteraldehyde and chitosan(US Patent-4,760,024, 1988)
Slide 5: Tosa et al invented a method of immobilization of enzymes in microbial cells using k-carrageenan as matrix. Gelatin in k-Carrageenan is a new polymer for the immobilization
Pietro crimonesi has invented a method of enzyme immobilization by adding a suspension of polysaccharide in aqueous media containing vinyl monomer and an enzyme, then adding ferric salt as catalyst and irradiating the mixture with UV light that lead to polymerization to form a copolymer of polysaccharide having enzymatic activity(USPatent-4,338,401. 1982)
Ephraim et al has reported a new carrier Eupergit© for the immobilization of enzymes of industrial potential. It is a carrier consisting nanopore beads of immobilized enzymes
Slide 6: Types of enzyme immobilization:
1. On surface immobilization:
Covalent coupling with polymers
Functional group containing polymers
Gluteraldehyde containing protein coupling
Polysaccharide as solid support
Complexation and chelation
2.With support immobilization
Inclusion in gels
Inclusion in fibers
Inclusion in microcapsules
Slide 7: Covalent coupling:
The protein functional groups can be used for the covalent coupling
NH2- lysine
COOH- α and β Aspertic acid, Glutamic acid
OH- Phenol ring on tyrosine
SH- Cysteines
Polymeric support s which are widely used:
Hydroxyl groups of polysaccharide, PVA, Poly methyl acetate
Amino ethyl coated polysaccharides, silica gels
Aldehyde and acetal groups of polymers
Amide groups of polypeptides
Slide 8: Hydroxyl group containing polymers:
Slide 9: Gluteraldehyde based protein coupling :
Bi functionality of gluteraldehyde can be used for the formation of covalent bond
Slide 10: Polysaccharides as solid support:
Naturally occurring polysaccharides like cellulose containing 1,4 linked β-D glucose, agarose containing 1,3 linked β-D glucose are derivatized and used as solid support for enzyme immobilization.
Slide 11: Adsorption method:
Various sold supports like aluminium, amberlite, bentonite, CMC, silica gels are available to adsorb the enzyme molecule and immobilize depending on various forces like vanDer Walls force and hydrogen boding
Slide 12: Complexation and chelation methods:
In this process chelation and complexation of transition metals are used to immobilize enzymes on solid support. Titanium or zirconium is widely preferred for their non toxic nature.
Slide 13: With support immobilization:
Here enzymes are physically entrapped in a polymer matrix, gels or capsules (micro encapsulation)
The support should have very small size pores which facilitates the movement of substrate inside the compartment
Inclusion in gels: Poly acrylamide gels
Poly vinyl alcohol gels
Inclusion in fibers: Cellulose and Poly acrylamide gels
Inclusion in micro capsules: polyamine, Polybasic acid chloride monomers
Slide 14: Inclusion in gels and fibers Inclusion in Microencapsulation
Slide 15: Examples of Immobilized enzymes by entrapment:
Slide 16: Applications of immobilized enzymes:
Production of antibiotics- immobilized penicillin amidase used for production of Penicillin G, Amoxicillin and ampicillin
Production of steroids- immobilized cells of Cornybacterium simplex is used to convert hydrocortisone and prednisolone from cortesolone
Production of amino acids- β tyrosinase used for the production of l-dopa
Production of organic compounds-
Propiniobacterium produce vit B 12
Catharanthus roseus produce ajmalcine
Digitalis lanata produce digitoxin
Slide 17: Clinical applications- Glucokinase as glucose sensor
Cholesterol oxidase as lipid sensor
Industrial applications-
Slide 18: References:
Pharmaceutical Biotechnology: Vyaas SP and Dixit VK (page-13-159)
A Textbook of Biotechnology: Dubey RC (page 238-240)
Molecular Biology and Biotechnology: Kumar HD(page-394-404)
Elements of Biotechnology: Gupta PK (Page-417-433)
Concepts of Biotechnology: Balasubramaniyan D et al (page-114-134)
http//:www.cheric.org/ippage/ipdata/2004/05/file/e200405-1101.pdf
http//:www.scribd.com/doc/14657530/applications-0f-enzyme-immobilization.pdf
http://www.youtube.com/watch?v=XYmioTsy4Cc
http://www.youtube.com/watch?v=gV_oIuQEmGE
Slide 19: Thank you…