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Microbial High Cell Density Fermentations in a Stirred Single-Use Bioreactor

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Disposable Bioreactors II

Part of the book series: Advances in Biochemical Engineering/Biotechnology ((ABE,volume 138))

Abstract

Microbial fermentations are of major importance in the field of biotechnology. The range of applications is rather extensive, for example, the production of vaccines, recombinant proteins, and plasmids. During the past decades single-use bioreactors have become widely accepted in the biopharmaceutical industry. This acceptance is due to the several advantages these bioreactors offer, such as reduced operational and investment costs. Although this technology is attractive for microbial applications, its usage is rarely found. The main limitations are a relatively low oxygen transfer rate and cooling capacity. The aim of this study was to examine a stirred single-use bioreactor for its microbial suitability. Therefore, the important process engineering parameters volumetric mass transfer coefficient (k L a), mixing time, and the heat transfer coefficient were determined. Based on the k L a characteristics a mathematical model was established that was used with the other process engineering parameters to create a control space. For a further verification of the control space for microbial suitability, Escherichia coli and Pichia pastoris high cell density fermentations were carried out. The achieved cell density for the E. coli fermentation was OD600 = 175 (DCW = 60.8 g/L). For the P. pastoris cultivation a wet cell weight of 381 g/L was reached. The achieved cell densities were comparable to fermentations in stainless steel bioreactors. Furthermore, the expression of recombinant proteins with titers up to 9 g/L was guaranteed.

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Abbreviations

\( a \) :

Empirical value for mathematical k L a description

\( A \) :

Heat transfer area

AOX:

Alcohol oxidase

\( b \) :

Empirical value for mathematical k L a description

\( \Updelta c \) :

Concentration difference

\( \Updelta c_{0} \) :

Initial concentration difference

\( C(t) \) :

Dissolved oxygen concentration

\( C^{*} \) :

Oxygen saturation concentration

CQA:

Critical quality attributes

\( d_{1} \) :

Vessel diameter

\( d_{2} \) :

Impeller diameter

DCU:

Digital control unit

DCW:

Dry cell weight

E. coli :

Escherichia coli

\( F(t) \) :

Feed flow rate

F G :

Gas flow rate

HCDF:

High cell density fermentation

IPTG:

Isopropyl-β-D-thiogalactopyranoside

\( k \) :

Heat transfer coefficient

\( k_{L} a \) :

Volumetric mass transfer coefficient

KPP:

Key process parameters

\( M \) :

Goodness of mixture

\( n \) :

Stirrer speed

\( Ne \) :

Newton number

OD600 :

Optical density measured at 600 nm

\( OTR \) :

Oxygen transfer rate

\( OUR \) :

Oxygen uptake rate

\( P/V \) :

Power input per volume

P. pastoris :

Pichia pastoris

pO2 :

Oxygen partial pressure

\( q_{{O_{2} }} \) :

Specific oxygen uptake rate

\( Q \) :

Heat flow

\( Q_{prod} \) :

Released heat flow

QbD:

Quality by design

RM:

Rocking motion

RO:

Reverse osmoses

\( s \) :

Thickness of the reactor wall

S :

Correlation factor for the heat generation

\( S_{Feed} \) :

Concentration of the feed solution

SDS-PAGE:

Sodium dodecyl sulfate polyacrylamide gel electrophoresis

STR:

Stirred tank reactor

t :

Time

\( \Updelta T \) :

Temperature difference

u :

Tip speed

\( V \) :

Filling volume of the bioreactor

\( V_{0} \) :

Initial volume

WCW:

Wet cell weight

\( X \) :

Cell density (dry cell weight)

\( X_{0} \) :

Initial cell density

\( Y_{{X/O_{2} }} \) :

Oxygen yield coefficient

\( Y_{X/S} \) :

Yield coefficient

\( Z \) :

Empirical value for mathematical k L a description

\( \alpha_{1} \) :

Convective heat transfer of the cooling liquid inside the double wall

\( \alpha_{2} \) :

Convective heat transfer of the reactor wall into the medium

\( \lambda \) :

Wall thermal conductivity

\( \mu \) :

Specific growth rate

\( \mu_{set} \) :

Selected specific growth rate for the fed batch

ρ :

Density

θ:

Mixing time

\( \nu \) :

Superficial air velocity

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Authors and Affiliations

  1. Sartorius Stedim Biotech GmbH, August-Spindler-Str.11, 37079, Goettingen, Germany

    Thomas Dreher, Ute Husemann, Franziska Klingenberg, Christian Zahnow, Thorsten Adams & Gerhard Greller

  2. Ablynx nv, Technologiepark 21, 9052, Zwijnaarde, Belgium

    Bart Walcarius & Peter Casteels

  3. Sartorius Stedim Biotech, Leuvensesteenweg 248 b, 1800, Vilvoorde, Belgium

    Davy de Wilde

Authors
  1. Thomas Dreher
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  2. Bart Walcarius
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  3. Ute Husemann
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  4. Franziska Klingenberg
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  5. Christian Zahnow
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  6. Thorsten Adams
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  7. Davy de Wilde
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  9. Gerhard Greller
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Correspondence to Thomas Dreher .

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Editors and Affiliations

  1. IBT Institut für Biotechnologie Bioverfahrenstechnik, Zürcher Hochschule für Angew.Wissensch Life Sciences und Facility Management, Wädenswil, Switzerland

    Dieter Eibl

  2. Hochschule Wädenswil, Wädenswil, Switzerland

    Regine Eibl

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Dreher, T. et al. (2013). Microbial High Cell Density Fermentations in a Stirred Single-Use Bioreactor. In: Eibl, D., Eibl, R. (eds) Disposable Bioreactors II. Advances in Biochemical Engineering/Biotechnology, vol 138. Springer, Berlin, Heidelberg. https://doi.org/10.1007/10_2013_189

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