Immobilized Cells

269 Pages·2001·6.969 MB·English

by  René H. Wijffels (auth.), Dr. René H. Wijffels (eds.)

SPRINGER
LAB MANUALS
Springer-Verlag Berlin Heidelberg GmbH
Rene H. Wijffels
(Ed.)
Immobilized Cells
With 54 Figures and 8 Tables
Springer
Dr. RENE H. WIJFFELS
Wageningen Un iversity
Food and Bioprocess Engineering Group
P.O.B.8129
6700 EV Wageningen
The Netherlands
e-mall: [email protected]
Library of Congress Cataloging-in-Publication Data
Immobilized cells / Rene H. Wijffels (ed.).
p. cm. - (Springer lab manuals)
Include bibliographical references and index.
ISBN 978-3-540-67070-4 ISBN 978-3-642-56891-6 (eBook)
DOI 10.1007/978-3-642-56891-6
1. Immobilized cells - Laboratory manuals. 1. Wijffels, Rene H., 1960-II. Series
QH585.5.145 14662000
571.6-dc21
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Contents
Introduction
Chapter 1
Characterization of Immobilized Cells; Introduction
RENE H. WUFFELS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1
Immobilization
Chapter 2
Description of the Support Material
EMILY J.T.M. LEENEN ................................... 6
Chapter 3
Description of the Immobilization Procedures
DENIS PONCELET, CLAIRE DULlEU, and MURIEL JACQUOT. . . . . . 15
Subprotocol 1: Ionic Gelation (Alginate Beads). ............... 20
Subprotoco12: hermal Gelation (K-Carrageenan Beads) ......... 23
Subprotocol 3: Ionic Polymer Coating (Chitosane on Alginate Beads) 25
Subprotocol 4: Coating by Transacylation Reaction ............ 26
Subprotocol 5: Polyelectrolyte Complex Membrane
(Sulfoethylcellulose/Polydiallyldimethyl
Ammonium Chloride) . . . . . . . . . . . . . . . . . . . . . .. 27
Chapter 4
Measurement of Density, Particle Size and Shape of Support
ERIK VAN ZESSEN, JOHANNES TRAMPER, and ARJEN RINZEMA 31
VI Contents
Chapter 5
Mechanical Stability of the Support
EMILY J.T.M. LEENEN. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 36
Chapter 6
Diffusion Coefficients of Metabolites
EVELIEN E. BEULING . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 44
Subprotocol 1: Preparation of the Membrane ................. 48
Subprotoco12: Diffusion Experiments. . . . . . . . . . . . . . . . . . . . . .. 51
Subprotoco13: Step-Response Method Utilizing Micro-Electrodes. 54
Kinetics
Chapter 7
Quantity of Biomass Immobilized, Determination
of Biomass Concentration
ELLEN A. MEIJER and RENE H. WIJFFELS . . . . . . . . . . . . . . . . . . .. 65
Chapter 8
Kinetics of the Suspended Cells
RENE H. WIJFFELS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 74
Chapter 9
Diffusion Limitation
RENE H. WIJFFELS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 77
Chapter 10
Micro-Electrodes
DIRK DE BEER . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 85
Subprotocol 1: Manufacturing of O Microsensors . . . . . . . . . . . .. 87
2
Subprotocol 2: Measurements . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 92
Chapter 11
Biomass Gradients
RENE H. WIJFFELS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 101
Contents VII
Chapter 12
NMR and Immobilized Cells
JEAN-NoEL BARBOTIN, JEAN-CHARLES PORTAlS,
PAULA M. ALVES, and HELENA SANTOS ..................... 123
Engineering
Chapter 13
Immobilization at Large Scale by Dispersion
DENIS PONCELET, STEPHANE DESOBRY, ULRICH JAHNZ,
and K. VORLOP ........................................ 139
Subprotocol 1: Jet Breaking Methods ....................... 140
Subprotocol 2: Lentikats ................................. 142
Subprotocol 3: Rotating Devices ........................... 144
Subprotocol 4. Emulsification Using Static Mixer . . . . . . . . . . . . .. 146
Chapter 14
Immobilization at Large Scale with the Resonance Nozzle Technique
JAN H. HUNIK ......................................... 150
Chapter 15
External Mass Transfer
RENE H. WUFFELS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 162
Chapter 16
Liquid Fluidization of Gel-Bead Particles
ERIK VA N ZESSEN, JOHANNES TRAMPER, and ARJEN RINZEMA .. 175
Chapter 17
Gradients in Liquid, Gas or Solid Fractions
RENE H. WIJFFELS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 182
Chapter 18
Support Material Stability at the Process Conditions Used
EMILY J.T.M. LEENEN ................................... 191
VIII Contents
Cases
Chapter 19
Immobilized Cells in Food Technology:
Storage Stability and Sensitivity to Contamination
CLAUDE P. CHAMPAGNE. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 199
Chapter 20
Immobilized Cells in Bioremediation
BRONAGH M. HALL and AlDEN J. Mc LOUGHLIN 213
Subprotocoll: Alginate Encapsulation ...................... 214
Subprotocol2: Carrageenan Encapsulation. . . . . . . . . . . . . . . . . .. 217
Subprotocol3: Co-Immobilization with Adjuncts. . . . . . . . . . . . .. 219
Subprotocol 4: Immobilization with Synthetic Polymers. . . . . . . .. 221
Subprotocol 5: Microencapsulation . . . . . . . . . . . . . . . . . . . . . . . .. 225
Subprotocol6: Monitoring Microbial Inoculum in the Environment 230
Chapter 21
Plasmid Stability in Immobilized Cells
JEAN-NoEL BARBOTIN .................................. 235
Chapter 22
Immobilization for High-Throughput Screening
NICOLE M. NASBY, TODD C. PETERSON, and CHRISTOPHER J. SILVA 247
Subject Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 259
Chapter 1 OVERVIEW
Characterization of Immobilized Cells; Introduction
RENE H. WIJFFELS
Introduction
In processes with immobilized cells, the cells are attached to or entrapped
in an inert support. In a continuously operated bioreactor, medium con
taining the substrate will be supplied. Substrate will be converted into a
product by the immobilized cells and product and remaining substrate
disappear with the outflowing medium. The immobilized cells are retained
easily in the bioreactor and as such utilized continuously. In this way the
capacity of the process is independent of the growth rate of the micro
organisms involved. In particular in cases where cells grow slowly, immo
bilized-cell process are more advantageous over processes with suspended
cells. Also for very specific situations where the presence of biomass in the
product should be prevented (e.g. champagne), drugs should be added gra
dually to the medium (e.g. islets of Langerhans), phage infections in starter
cultures should be prevented (e.g. lactic-acid bacteria for cheese produc
tion) and plasmids should be stabilized (for application of genetically en
gineered cells in a continuous mode), immobilized cells perform better
than suspended cells.
Immobilized cells have been studied widely during the last decades. An
enormous quantity of papers have been published. For many processes the
complex physiology in a heterogeneous environment is now becoming
clear. In addition it is shown that in many processes it is more efficient
to use immobilized cells than suspended cells.
In 1995 the symposium "Immobilized Cells: Basics and Applications"
was organized under auspices of the Working Party of Applied Catalysis of
the European Federation of Biotechnology (Wijffels et al. 1996). The sym-
Rene H. Wijffels, WageningenUniversity, Food and Bioprocess Engineering Group,
P.O. Box 8129, Wageningen, 6700 EV, The Netherlands (phone [phone];
fax [phone]; e-mail [email protected])
2 RENE H. WIJFFELS
posium covered the path from basic physiological research to applications,
and scientists were brought together from different disciplines from aca
demia, industry and research institutes. For applications, physiology needs
to be integrated with engineering. The goal of the symposium was to relate
basic research to applications. Another aim was to extract guidelines for
characterization of immobilized cells in view of process design and appli
cation from the contributions.
Laboratory methods for scientific research and manufacturers
It is important to recognize that research on immobilized metabolizing
cells should lead, whenever possible, to eventual successful implementa
tion of such catalysts in industrial processes. One of the aims of the pro
posed guidelines was to ensure that research done is effective in achieving
this objective. It is difficult otherwise to justify the research.
Of course there are differences between scientific research and applica
tions; applications should be fed by the scientific efforts. The scientific
guidelines could be a tool to bridge the gap. As a result of following
the guidelines it should not be strictly necessary for manufacturers to re
peat the tests that should have been done in the scientific research phase.
The guidelines if implemented should ensure a certain minimal quality of
research data. Development of an application can thus focus on more spe
cific research by manufacturers, e.g. the scale-up aspects.
The goal of research, however, is not always application oriented. Dif
ferent goals of the research can be:
• development of methodology
• scientific question
• application
In the different phases of the development different parts of the guidelines
should be used. If research is done in order to apply the process, the scale
up and applications should be considered at a very early stage, so that ex
periments done, for instance, on physiology are relevant. In case of the
development of methodologies and scientific questions this is not always
necessary.
Guidelines should not be too complicated and be a help for research
rather than a number of restrictive procedures. Guidelines should cover the
whole procedure of the development of processes with immobilized cells.
This means that within a single research paper not all guidelines need to be